Editorial
1.
Introduction
2.
The pros and cons of
Dendritic cell vaccination against ovarian cancer -- tipping the Treg/TH17 balance to therapeutic advantage?
TH17-based immunotherapy 3.
Can dendritic cells be educated to drive TH17 responses against ovarian cancer?
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
4.
Expert opinion
Martin J Cannon†, Hannah Goyne, Pamela J B Stone & Maurizio Chiriva-Internati †
University of Arkansas for Medical Sciences, Department of Microbiology and Immunology, Little Rock, Arkansas, USA
The pathology of ovarian cancer is characterized by profound immunosuppression in the tumor microenvironment. Mechanisms that contribute to the immunosuppressed state include tumor infiltration by regulatory T cells (Treg), expression of B7-H1 (PDL-1), which can promote T cell anergy and apoptosis through engagement of PD-1 expressed by effector T cells, and expression of indoleamine 2,3-dioxygenase (IDO), which can also contribute to effector T cell anergy. Expression of both B7-H1 and IDO has been associated with differentiation and recruitment of Treg, and clinical studies have shown that each of these mechanisms correlates independently with increased morbidity and mortality in patients with ovarian cancer. In a remarkable counterpoint to these observations, ovarian tumor infiltration with TH17 cells correlates with markedly improved clinical outcomes. In this Future Perspectives review, we argue that dendritic cell (DC) vaccination designed to drive tumor-antigen-specific TH17 T cell responses, combined with adjuvant treatments that abrogate immunosuppressive mechanisms operative in the tumor microenvironment, offers the potential for clinical benefit in the treatment of ovarian cancer. We also discuss pharmacological approaches to modulation of MAP kinase signaling for manipulation of the functional plasticity of DC, such that they may be directed to promote TH17 responses following DC vaccination. Keywords: dendritic cells, ovarian cancer, p38 MAPK, regulatory T cells, TH17 T cells Expert Opin. Biol. Ther. (2011) 11(4):441-445
1.
Introduction
In recent years, it has become increasingly apparent that ovarian tumors avail themselves of multiple mechanisms of immune evasion, the most prominent of which is recruitment and infiltration of regulatory T cells that suppress anti-tumor immunity. Landmark studies from Curiel and colleagues showed that regulatory T cells (Treg) are recruited to ovarian tumors by the chemokine CCL22 (predominantly expressed by ovarian tumors), and that the presence of Treg confers immune privilege and is associated with a poor prognosis and increased mortality [1]. Other investigators have corroborated these observations, showing that high expression of the forkhead box transcription factor foxp3, which is preferentially expressed by CD4+ Treg, is an independent prognostic factor for reduced overall survival in ovarian cancer [2], and that a high CD8+ T cell:Treg ratio is associated with a more favorable prognosis for this disease [3]. These observations support the idea
10.1517/14712598.2011.554812 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
441
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Dendritic cell vaccination against ovarian cancer -- tipping the Treg/TH17 balance to therapeutic advantage?
that depletion of tumor-associated Treg, or inhibition of Treg function, may be beneficial, particularly in conjunction with active tumor-specific immunotherapy. In contrast with the strong evidence that Treg infiltration is associated with poor outcomes in ovarian cancer (and other malignancies), the recent observation that TH17 T cell infiltration in ovarian cancer correlates with markedly more favorable clinical outcomes provides a striking counterpoint [4]. Tumor-infiltrating TH17 cells were positively associated with effector cells and negatively associated with Treg infiltration, with the latter relationship arguably being founded on the known reciprocal regulation of Treg and TH17 differentiation [5,6]. Tumor-associated macrophages were shown to be efficient inducers of T cell IL-17 production, through an IL-1b-dependent mechanism [4], an observation that is consistent with evidence pointing to a critical role for IL-1b in the induction of human TH17 responses [7-9]. Furthermore, Kryczek and colleagues found a positive correlation between ascites IL-17 and the TH1-associated chemokines CXCL9 and CXCL10, and provided evidence that TH17 T cell production of IL-17 and IFN-g-induced expression of CXCL10. In turn, the levels of CXCL9 and CXCL10 in tumor ascites positively correlated with tumor-infiltrating CD8+ T cells [4]. 2. The pros and cons of TH17-based immunotherapy
These observations have inevitably led to the question of whether TH17 cells could be therapeutically induced or expanded, either by tumor vaccines or adoptive immunotherapy [10]. Although the current evidence in ovarian cancer appears to present a strong case in favor of TH17-based antitumor immunotherapy, this is a controversial issue, since a number of studies have indicated a role for IL-17 in promoting tumor growth and invasion [11-16]. On the other hand, several recent reports have supported the view that TH17 responses may have therapeutic benefit in promoting anti-tumor immunity and survival. In the B16 mouse model of melanoma, adoptive T cell therapy with tumor-specific TH17 cells prompted strong activation of tumor-specific CD8+ T cells (which were required for the antitumor effect), thus indicating that TH17-driven inflammation can play a pivotal role in antitumor immunity [17]. Induction of TH17 responses in a mouse model of pancreatic cancer has also been shown to delay tumor growth and improve survival [18]. In similar vein, tumor growth and pulmonary metastasis was enhanced following injection of the MC38 colon cancer cell line in IL-17-deficient mice [19], again suggesting a protective role for IL-17-expressing T cells. Most notably, the pretreatment frequency of CD4+ TH17 cells in prostate cancer patients was found to correlate with the clinical response to a whole-cell vaccine [20], suggesting that the association of TH17 cells with improved survival may not be unique to ovarian cancer. 442
Furthermore, and in marked contrast with the prevailing opinion that CD4+ TH1 T cell responses and CD8+ CTL responses represent an optimal line of attack for antitumor immunotherapy, recent evidence has suggested that TH17based cellular immunotherapy may offer the potential for greater therapeutic efficacy. Groundbreaking studies from the National Cancer Institute have clearly shown that adoptively transferred CD4+ TH17 cells were markedly more effective than CD4+ TH1 cells in eradication of advanced B16 melanoma in a mouse model [21]. These investigators further showed that, compared with TH1 cells, TH17 cells enjoy a survival advantage in vivo, suggesting that their improved persistence may be a key reason for their greater ability to control disease.
Can dendritic cells be educated to drive TH17 responses against ovarian cancer?
3.
This section is based on the premise that active immunotherapy, and particularly dendritic cell (DC) vaccination, designed to drive a tumor-antigen-specific TH17 T cell response holds the potential to be of clinical benefit for patients with ovarian cancer. Various studies have shown that TH17 T cell differentiation in vitro can readily be driven by cytokines, notably IL-1b (see above), suggesting that tumor-antigen-specific TH17-based adoptive T cell immunotherapy may be a viable approach for treatment of ovarian cancer. However, such procedures are cumbersome and complex, and are not readily translated to clinical practice. A more practical and efficient alternative may be found with DC vaccination. DC are remarkable for their plasticity in directing T cell differentiation and effector function, and thus the key to success may reside in our ability to educate DC to drive ovarian tumor-antigenspecific TH17 responses. How could this be achieved? Several recent studies have indicated that regulation of the p38 and extracellular-signal-regulated kinase(ERK)--MAP kinase (MAPK) signal transduction pathways in DC plays a central role in direction of T cell differentiation. Inhibition of MEK 1/2 and ERK MAPK signaling promotes IL-12 production and TH1 T cell responses, whereas inhibition of p38 MAPK increases signal transduction through ERK 1/2 and blocks IL-12 production [22]. At face value, these observations suggest that inhibition of p38 MAPK signaling would be disadvantageous for DC-driven antitumor T cell responses, since this would abrogate TH1 responses, which are widely held to be important for effective anti-tumor immunity. However, p38 inhibition promotes differentiation and survival of monocyte-derived DC [23], and p38 inhibition or MEK/ERK MAPK activation restores deficiencies in DC function in myeloma patients [24], suggesting that treatment of DC with pharmacological inhibitors of p38 signaling may confer some benefit. Furthermore, p38 MAPK signaling in DC is associated with increased expression of IL-10 and the induction of
Expert Opin. Biol. Ther. (2011) 11(4)
Cannon, Goyne, Stone & Chiriva-Internati
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
tolerance in a mouse model of melanoma, thus contributing to the suppression of anti-tumor T cell responses [25]. Inhibition of p38 signaling in DC from tumor-bearing mice markedly suppressed expression of IL-10 and restored the capacity of DC to stimulate T cells. It is of particular significance that blockade of the p38 pathway can attenuate Treg induction by DC [26], whereas blockade of the ERK pathway suppresses DC-driven TH17 responses [27], suggesting that p38 blockade (which enhances ERK phosphorylation) may favor a switch from Treg induction to TH17 differentiation and expansion. These observations could have major implications for the rational design of DC vaccines against ovarian cancer. 4.
Expert opinion
The proposal that tumor-antigen-specific CD4+ TH17 immune responses may benefit cancer patients is a challenging position to adopt. Based on experimental evidence, there is little doubt that TH17 responses can drive tumor progression, invasion and angiogenesis. On the other hand, it is equally evident from experimental models and clinical studies that TH17 responses can support robust anti-tumor immunity and favor patient survival. How can these apparently opposing observations be reconciled? First, it is probable that TH17 responses are not homogeneous, and that differing effector functions under that broad umbrella are likely to have different outcomes. The ultimate challenge for tumor immunologists will be to dissect the nuances of TH17 function, and to determine how to drive a response that favors anti-tumor immunity rather than disease progression [16]. In the case of ovarian cancer, clinical evidence presents a strong rationale for basing active immunotherapy on strategies that drive a TH17 response [4]. We propose that manipulation of DC function to drive ovarian tumor-antigen-specific TH17 responses may afford the best opportunity for immunological treatment of ovarian cancer through DC vaccination. We have also discussed experimental evidence that inhibition of the p38 MAPK signaling pathway in DC may be an appropriate line of investigation to achieve this goal. Assuming that such a strategy is viable, there remain numerous barriers to successful DC vaccination for ovarian cancer. Immunosuppressive mechanisms operative in the
ovarian tumor microenvironment include infiltrating Treg (discussed above), and expression of B7-H1 (programmed death ligand 1 (PDL-1)) by tumor cells and infiltrating macrophages, resulting in apoptosis and anergy [28,29]. Of particular clinical interest, a retrospective analysis of human ovarian cancers revealed that patients with higher B7-H1 expression had a significantly poorer prognosis than those for whom the tumors had lower B7-H1 expression [30]. Expression of indoleamine 2,3-dioxygenase (IDO), which can contribute to recruitment of Tregs [31,32], has also been associated with poor clinical outcomes in ovarian cancer [33,34]. Tumor expression of endothelin-1, which can inhibit effector T cell migration across vascular endothelium into the tumor microenvironment, may also reduce the efficacy of immunotherapy or vaccination [35]. The optimal strategy for DC vaccination may thus combine adjuvant treatments designed to abrogate immunosuppression in the tumor microenvironment. B7-H1 may be blocked with specific antibodies, and IDO function can be blocked with 1-methyl-tryptophan, a competitive inhibitor of enzyme function that is currently being tested in clinical trials. Small-molecule antagonists of endothelin receptors are also undergoing clinical tests [36]. Last, but not least, various strategies can be applied to abrogation of tumor-associated Treg activity, notably treatment with denileukin diftitox (ONTAK) or low-dose cyclophosphamide [37]. Paclitaxel, which is commonly used for treatment of ovarian cancer, may also have activity against Treg [38]. Given the current weight of evidence, we would advocate further studies on the potential for treatment of ovarian cancer with DC vaccination formulated to drive TH17 responses, in combination with adjuvant treatments designed to blockade immunosuppressive mechanisms that prevail in the ovarian tumor microenvironment.
Declaration of interest The authors are sponsored by an NIH grant UL1RR029884-01, Arkansas Center for Clinical Translational Research. MJ Cannon is founder of DCV Technologies Inc, a biotechnology company dedicated to the clinical developement of dendritic cell vaccines for the treatment of cancer. M Chiriva-Internati is founder of Kiromic Inc, a biotechnology company that seeks to develop therapeutic cancer vaccines. The other authors declare no conflict of interest.
Expert Opin. Biol. Ther. (2011) 11(4)
443
Dendritic cell vaccination against ovarian cancer -- tipping the Treg/TH17 balance to therapeutic advantage?
Bibliography Papers of special note have been highlighted as either of interest () or of considerable interest () to readers. 1.
..
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
2.
.
3.
..
4.
..
5.
6.
444
Curiel TJ, Coukos G, Zou L, et al. Specific recruitment of regulatory T cells in ovarian carcinoma fosters immune privilege and predicts reduced survival. Nat Med 2004;10:942-9 A landmark study showing that Treg infiltration of ovarian tumors correlates with increased morbidity and mortality. Wolf D, Wolf AM, Rumpold H, et al. The expression of the regulatory T cell-specific forkhead box transcription factor FoxP3 is associated with poor prognosis in ovarian cancer. Clin Cancer Res 2005;11:8326-31 A clinical study that furnishes further evidence that Treg are associated with a poor prognosis in ovarian cancer. Sato E, Olson SH, Ahn J, et al. Intraepithelial CD8+ tumor-infiltrating lymphocytes and a high CD8+/regulatory T cell ratio are associated with favorable prognosis in ovarian cancer. Proc Natl Acad Sci USA 2005;102:18538-43 An important study showing that increased infiltration of CD8+ effector T cells versus immunosuppressive Treg favors improved clinical outcomes in ovarian cancer patients. Kryczek I, Banerjee M, Cheng P, et al. Phenotype, distribution, generation, and functional and clinical relevance of Th17 cells in the human tumor environments. Blood 2009;114:1141-9 A high-impact report showing that IL-17 expression in ovarian cancer correlates with markedly improved overall survival, leading to the hypothesis that TH17-based immunotherapy may be of clinical benefit for these patients. Kryczek I, Wei S, Zou L, et al. Th17 and regulatory T cell dynamics and the regulation by IL-2 in the tumor microenvironment. J Immunol 2007;178:6730-3 Bettelli E, Oukka M, Kuchroo VK. TH-17 cells in the circle of immunity
in tumor immunity. Immunity 2009;31:787-98
and autoimmunity. Nat Immunol 2007;8:345-50 7.
8.
Acosta-Rodriguez EV, Napolitani G, Lanzavecchia A, Sallusto F. Interleukins 1beta and 6 but not transforming growth factor-beta are essential for the differentiation of interleukin 17-producing human T helper cells. Nature Immunol 2007;8:942-9 Wilson NJ, Boniface K, Chan JR, et al. Development, cytokine profile and function of human interleukin 17-producing helper T cells. Nature Immunol 2007;8:950-7
9.
Liu H, Rohowsky-Kochan C. Regulation of IL-17 in human CCR6+ effector memory T cells. J Immunol 2008;180:7948-57
10.
Munn DH. Th17 cells in ovarian cancer. Blood 2009;114:1134-5
11.
Tartour E, Fossiez F, Joyeux I, et al. Interleukin 17, a T-cell-derived cytokine, promotes tumorigenicity of human cervical tumors in nude mice. Cancer Res 1999;59:3698-704
12.
13.
14.
18.
Gnerlich JL, Mitchem JB, Weir JS, et al. Induction of Th17 cells in the tumor microenvironment improves overall survival in a murine model of pancreatic cancer. J Immunol 2010;185:4063-71
19.
Kryczek I, Wei S, Szeliga W, et al. Endogenous IL-17 contributes to reduced tumor growth and metastasis. Blood 2009;114:357-9
20.
Derhovanessian E, Adams V, Hahnel K, et al. Pretreatment frequency of circulating IL-17+ CD4+ T-cells, but not Tregs, correlates with preclinical response to whole-cell vaccination in prostate cancer patients. Int J Cancer 2009;125:1372-9 Clinical evidence suggesting that TH17 immune responses may contribute to the efficacy of active anti-tumor immunotherapy in prostate cancer is presented.
.
21.
Muranski P, Boni A, Antony PA, et al. Tumor-specific Th17-polarized cells eradicate large established melanoma. Blood 2008;112:362-73 An important experimental study showing that adoptive transfer of TH17 T cells is highly effective against established melanoma in mice.
Numasaki M, Watanabe M, Suzuki T, et al. IL-17 enhances the net angiogenic activity and in vivo growth of human non-small cell lung cancer in SCID mice through promoting CXCR-2-dependent angiogenesis. J Immunol 2005;175:6177-89
22.
Nam J-S, Terabe M, Kang M-J, et al. Transforming growth factor beta subverts the immune system into directly promoting tumor growth through interleukin-17. Cancer Res 2008;68:3915-23
Jackson AM, Mulcahy LA, Zhu XW, et al. Tumour-mediated disruption of dendritic cell function: inhibiting the MEK1/2-p44/42 axis restores IL-12 production and Th1-generation. Int J Cancer 2008;123:623-32
23.
Zhu X, Mulcahy LA, Mohammed RAA, et al. Il-17 expression by breast-cancer-associated macrophages: IL-17 promotes invasiveness of breast cancer cell lines. Breast Cancer Res 2008;10:R95
Xie J, Qian J, Yang J, et al. Critical roles of Raf/MEK/ERK and PI3K/AKT signaling and inactivation of p38 MAP kinase in the differentiation and survival of monocyte-derived immature dendritic cells. Exp Hematol 2005;33:564-72
24.
Wang S, Hong S, Yang J, et al. Optimizing immunotherapy in multiple myeloma: restoring the function of patients’ monocyte-derived dendritic cells by inhibiting p38 or activating MEK/ERK MAPK and neutralizing interleukin-6 in progenitor cells. Blood 2006;108:4071-7
25.
Zhao F, Falk C, Osen W, et al. Activation of p38 mitogen-activated protein kinase drives dendritic cells to become tolerogenic in Ret transgenic
15.
Wang L, Yi T, Kortylewski M, et al. IL-17 can promote tumor growth through an IL-6-Stat3 signaling pathway. J Exp Med 2009;206:1457-64
16.
Murugaiyan G, Saha B. Protumor vs antitumor functions of IL-17. J Immunol 2009;183:4169-75
17.
Martin-Orozco N, Muranski P, Chung Y, et al. T helper 17 cells promote cytotoxic T cell activation
Expert Opin. Biol. Ther. (2011) 11(4)
.
Cannon, Goyne, Stone & Chiriva-Internati
.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
26.
..
27.
.
28.
29.
mice spontaneously developing melanoma. Clin Cancer Res 2009;15:4382-90 This study shows that p38 MAPK signal transduction contributes to dendritic cell induction of tolerance, and that inhibition of p38 MAPK can restore the ability of dendritic cells to activate anti-tumor effector T cell responses. Jarnicki AG, Conroy H, Brereton CF, et al. Attenuating regulatory T cell induction by TLR agonists through inhibition of p38 MAPK signaling in dendritic cells enhances their efficacy as vaccine adjuvants and cancer immunotherapeutics. J Immunol 2008;180:3797-806 This paper provides strong evidence that inhibition of p38 MAPK signaling in dendritic cells can inhibit Treg induction and enhance the efficacy of anti-tumor dendritic cell vaccination. Brereton CF, Sutton CE, Lalor SJ, et al. Inhibition of ERK MAPK suppresses IL-23- and IL-1-driven IL-17 production and attenuates autoimmune disease. J Immunol 2009;183:1715-23 Experimental evidence that ERK MAPK signal transduction (which is negatively regulated by p38 MAPK) favors dendritic-cell-driven TH17 responses. Dong H, Strome SE, Salomao DR, et al. Tumor-associated B7-H1 promotes T-cell apoptosis: a potential mechanism of immune evasion. Nature Med 2002;8:793-800 Curiel TJ, Wei S, Dong H, et al. Blockade of B7-H1 improves myeloid dendritic cell-mediated antitumor immunity. Nature Med 2003;9:562-7
30.
..
31.
32.
33.
..
34.
..
35.
Hamanishi J, Mandai M, Iwasaki M, et al. Programmed cell death 1 ligand 1 and tumor-infiltrating CD8+ T lymphocytes are prognostic factors of human ovarian cancer. Proc Natl Acad Sci USA 2007;104:3360-5 This paper shows that B7-H1 (PDL-1) expression in ovarian cancer is a negative predictor of patient survival, whereas CD8+ T cell infiltration is a positive prognostic indicator of outcomes. Sharma MD, Hou D-Y, Liu Y, et al. Indoleamine 2,3 dioxygenase controls conversion of Foxp3+ Tregs to TH17-like cells in tumor-draining lymph nodes. Blood 2009;113:6102-11 Chung DJ, Rossi M, Romano E, et al. Indoleamine 2,3-dioxygenase-expressing mature human monocyte-derived dendritic cells expand potent autologous regulatory T cells. Blood 2009;11:555-63 Okamoto A, Nikaido T, Ochiai K, et al. Indoleamine 2,3-dioxygenase serves as a marker of poor prognosis in gene expression profiles of serous ovarian cancer cells. Clin Cancer Res 2005;11:6030-9 An important study showing that IDO expression in ovarian cancer is a marker for poor prognosis. Inaba T, Ino K, Kajiyama H, et al. Role of the immunosuppressive enzyme indoleamine 2,3-dioxygenase in the progression of ovarian carcinoma. Gynecol Oncol 2009;115:185-92 Further evidence that IDO expression correlates with poor outcomes in ovarian cancer patients. Buckanovich RJ, Facciabene A, Kim S, et al. Endothelin B receptor mediates the endothelial barrier to T cell homing to
Expert Opin. Biol. Ther. (2011) 11(4)
..
tumors and disables immune therapy. Nat Med 2008;14:28-36 An innovative study that reveals a critical role for tumor endothelin expression and inhibition of effector T cell infiltration across vascular enothelium as a barrier for tumor immunotherapy.
36.
Kandalaft LE, Facciabene A, Buckanovich RJ, et al. Endothelin B receptor, a new target in cancer immune therapy. Clin Cancer Res 2009;15:4521-8
37.
Kandalaft LE, Singh N, Liao JB, et al. The emergence of immunomodulation: combinatorial immunochemotherapy opportunities for the next decade. Gynecol Oncol 2010;116:222-33
38.
Zhang L, Dermawan K, Jin M, et al. Differential impairment of regulatory T cells rather than effector T cells by paclitaxel-based chemotherapy. Clin Immunol 2008;129:219-29
Affiliation
Martin J Cannon†1,2 PhD, Hannah Goyne1 MD, Pamela J B Stone2 MD & Maurizio Chiriva-Internati3 PhD † Author for correspondence 1 University of Arkansas for Medical Sciences, Department of Microbiology and Immunology, Little Rock, Arkansas, USA Tel: +1 501 296 1254; Fax: +1 501 686 5359; E-mail:
[email protected] 2 University of Arkansas for Medical Sciences, Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Little Rock, Arkansas, USA 3 University Health Sciences Center, Division of Hematology and Oncology, Department of Internal Medicine, Texas Tech Lubbock, Texas, USA
445
Review
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Stem cells and stroke: opportunities, challenges and strategies 1.
Introduction
Terry C Burns & Gary K Steinberg†
2.
Cell transplantation for stroke
Stanford University School of Medicine, Department of Neurosurgery, Stanford, CA, USA
3.
Stroke-induced neurogenesis
4.
Conclusion
5.
Expert opinion
Introduction: Stroke remains the leading cause of disability in the Western world. Despite decades of work, no clinically effective therapies exist to facilitate recovery from stroke. Stem cells may have the potential to minimize injury and promote recovery after stroke. Areas covered: Transplanted stem cells have been shown in animal models to migrate to the injured region, secrete neurotrophic compounds, promote revascularization, enhance plasticity and regulate the inflammatory response, thereby minimizing injury. Endogenous neural stem cells also have a remarkable propensity to respond to injury. Under select conditions, subventricular zone progenitors may be mobilized to replace lost neurons. In response to focal infarcts, neuroblasts play important trophic roles to minimize neural injury. Importantly, these endogenous repair mechanisms may be experimentally augmented, leading to robust improvements in function. Ongoing clinical studies are now assessing the safety and feasibility of cell-based therapies for stroke. Expert opinion: We outline the unique challenges and potential pitfalls in the clinical translation of stem cell research for stroke. We then detail what we believe to be the specific basic science and clinical strategies needed to overcome these challenges, fill remaining gaps in knowledge and facilitate development of clinically viable stem cell-based therapies for stroke. Keywords: clinical trial, differentiation, ischemic brain injury, migration, neural progenitor cell, neuroblast, neurogenesis, neuroprotection, neuroregeneration, plasticity, stem cell, stroke, subventricular zone, translational research Expert Opin. Biol. Ther. (2011) 11(4):447-461
1.
Introduction
With an incidence of almost 800,000 new victims per year in the USA alone, stroke persists as the leading cause of disability and the third leading cause of mortality in the Western world. Stroke leads to rapid destruction of brain tissue over several hours, with an estimated 1.9 million neurons, representing approximately 14 billion synapses, dying each minute [1]. It is important to recognize that each lost neuron was born at a specified time and location during development as a result of complex sequences of physical and chemical signals as well as intrinsic timing mechanisms guiding progenitor cell fate. After birth, the immature neurons were precisely guided into appropriate locations, from which they extended projections along intersecting gradients of diffusible, membrane and extracellular matrix-bound molecules. They then competed successfully for neurotrophic signals and established thousands of activity- and experience-dependant synaptic connections. In the wake of stroke, these intricate networks are swiftly reduced to an expanding necrotic milieu of dead and dying cells. Adjacent neurons teeter on the edge of viability with marginal blood supply, where mounting inflammatory responses may mediate additional cell death. In addition to neuronal cell loss, even greater numbers of glia with probably under-appreciated location-defining and regulatory as well as supportive roles are also destroyed. 10.1517/14712598.2011.552883 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
447
Stem cells and stroke: opportunities, challenges and strategies
Article highlights. .
.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
.
.
. .
. .
.
To date, demonstrated mechanisms of stem cell benefit have included direct inhibition of cell death, enhanced regeneration of vasculature, immunomodulation, induction of neuronal plasticity, and promotion of endogenous neurogenesis. Both bone marrow mononuclear cells (BMMCs) and mesenchymal stem cells (MSCs) seem to have quite limited survival in the brain after either local or systemic delivery. Thus it is likely that benefits are mediated predominantly via trophic signals of variable duration. Recent meta-analysis of preclinical studies employing intravenous cell delivery indicated that neural stem cells (NSCs) yielded the greatest behavioral improvements when compared with bone-marrow-derived or other cell types. The recent development of techniques to generate induced pluripotent stem (iPS) cells, opens the potential for autologous neural cell therapy, thereby averting the need for immunosuppression. The optimal timing for cell delivery is unclear, but may depend upon the predominant. mechanism of action. Therapies aiming for neuroprotection will require earlier delivery than those targeting neuroplasticity. Exogenous cell therapy may act, in part, by augmenting the endogenous neurogenic response to stroke. Increased endogenous progenitor cell proliferation and neuroblast recruitment may persist for at least several months after ischemic injury. Multiple cellular and molecular tools now exist to enhance endogenous responses to stroke. The failure of hundreds of neuroprotective compounds in clinical trials illustrates the sobering challenge ahead of translational therapy for stroke. In the treatment of stroke, it remains true that ‘time is brain’. Cell therapies should be developed in conjunction with optimized recanalization technologies to target residual areas of ischemia, combat reperfusion injury and provide trophic support in areas of hemorrhagic conversion.
This box summarizes key points contained in the article.
Restoration of blood flow within the first three to four hours of stroke onset enables measurable improvements in outcome. However, only a small minority of patients arrive early enough to receive effective therapy. Despite decades of work and promising animal data, neuroprotective strategies aiming to limit further exacerbation of cell loss within and beyond this timeframe have uniformly failed in human trials [2-5]. Stem cells have the potential to generate nearly unlimited numbers of neural cells. Given the complex fidelity of neuronal development and integration, however, true cell replacement has proven an elusive goal. During the past decade, dozens of cell types have been tested via multiple routes of delivery in numerous animal models of stroke; in many cases, markedly decreased lesion size and improved functional outcomes have been achieved. Though some have claimed ‘replacement’ of neurons by transplanted cells, others have encountered poor survival despite functional benefits, suggesting indirect 448
mechanisms of recovery. Others have sought to stimulate the brain’s own stem cells toward regeneration with promising preliminary results [6]. Here we evaluate the preclinical and clinical progress of stem cell therapy to date. We discuss current evidence regarding mechanisms of action, and outline pertinent opportunities, challenges and strategies for safe and effective translation of stem cell therapy into clinical practice. 2.
Cell transplantation for stroke
Exogenous stem cells Preclinical studies of cell transplantation have identified a surprising variety of cells that promote functional recovery after stroke. Work to optimize delivery parameters such as route, timing, cell dose and immunosuppression is ongoing. To date, demonstrated mechanisms of benefit have included direct inhibition of cell death, enhanced regeneration of vasculature, immunomodulation, induction of neuronal plasticity and promotion of endogenous neurogenesis. 2.1
Human fetal brain cells Pioneering cell transplantation work focused initially on replacement of dopaminergic neurons for Parkinson’s disease (PD). Studies employing fetal midbrain demonstrated behavioral benefits from ‘cell replacement’, prompting several clinical trials with variable outcomes. In the mid 1980s, Polezhaev and Alexandrova performed transplantations of fetal brain tissue into rat brains after ischemic injury. Robust engraftment was observed with evidence of synaptic integration. Grafts also decreased cell death and promoted the restoration of ‘dysfunctional’ neurons to their normal state [7]. Grafts, which seemed to survive best in the penumbra [8] improved local neurotransmitter levels and facilitated cognitive recovery [9]. Human fetal brain tissue is a limited and ethically challenging resource. As such, no clinical trials of fetal cells have been pursued for stroke and significant efforts have sought to develop alternate cell types that may be more readily amenable to widespread clinical application. 2.1.1
Human teratocarcinoma cells A teratocarcinoma cell line, NT2, was shown in 1984 to generate pure populations of post-mitotic neural-like cells upon exposure to retinoic acid [10]. In 2000, based on preclinical evidence for functional improvements in animal models of stroke [11], these became the first cells reported in a Phase I clinical trial of a cell-based therapy for stroke (Table 1) [12]. Cells were grafted stereotactically into patients with stable deficits after a basal ganglia infarct; immunosuppression was continued for 2 months. Overall, no adverse effects were noted, and surviving cells were observed post-mortem with no evidence of neoplasm at 27 months [13]. In 2005, the report of a Phase II randomized controlled trial involving 14 treatment and 4 control patients revealed functional improvements in some patients. Given the very small group sizes, improvements based on a primary outcome measure of European stroke scale at 6 months did not 2.1.2
Expert Opin. Biol. Ther. (2011) 11(4)
Cell number/
Yonsei University, Souel, S Korea
Bang et al., 2005 [29]*
Expert Opin. Biol. Ther. (2011) 11(4)
Infarct
125 -- 500 million/ 24 -- 65 NIHSS autologous 4 -- 17 BMMCs/IA 50 million 30 -- 75 NIHSS > 6 2/Autologous MSCs/IV
41 -- 64 NIHSS 10.6 ± 0.92
14 -- 55 million/ Autologous BMMCs/IC
16 + I/II 36 controls
5 -- 7 weeks
I
I
6
5
8 -- 12 weeks
1 -- 10 years
I
II
I
No
Safety
Neurologic deficits
Safety, tolerance
Safety
Safety
Yes
No
No
Yes
No
6 months
4 years
Safe. European Stroke Score improved at 6 months (p = 0.046) Feasible; primary outcome measure not met
Observer 5 years only
Safe, feasible; mRS improved (p = 0.046), best if SVZ intact
FDA terminated trial due to possible side effects Observer 12 months Safe, feasible; only Barthel index higher at 3 and 6 months only No 12 months Safe; neuropsychiatric improvements in some patients No 120 days No neurologic worsening
No
Observer 6 months only
No
Randomized? Blinding Follow up Results
European Yes Stroke Score
Safety, feasibility
outcome
Phase Primary
5+ I/II 25 controls
30 -- 75 NIHSS > 6
50 million 2/Autologous MSCs/IV
32 -- 61 days
25 -- 52 NIHSS 5 -- 11 1.5 -- 10 years 5
Up to 50 million/ fetal porcine cells/IC
14 + 4 controls
n
40 -- 70 Basal ganglia 1 -- 5 years infarct, ESS 10 -- 45
window
Treatment
12
severity/ type 44 -- 75 Basal ganglia 6 months -infarct 6 years
Age
*[30] Includes patients previously reported in [29]; mRS not significantly different between groups in [29]; Barthel index not reported in Lee [30]. BMMC: Bone marrow mononuclear cell; hNT: Human teratocarcinoma-derived neural cell line; IA: Intra-arterial; IC: Intracerebral; IV: Intravenous; MSC: Mesenchymal stem cell; na: Not available; NIHSS: National Institutes of Health Stroke Scale; SPECT: Single-photon emission-computed tomography.
Centro Internacional Suarezde Restauracion Monteagudo et al., 2009 [19] Neurologica, Habana, Cuba Universidad Federal, Barbosa da Rio de Janeiro, Brazil Fonseca et al., 2010 [18] Yonsei University, Lee et al., Souel, S Korea 2010 [30]*
Harvard MA and Cornell, NY, USA
Savitz et al., 2005 [16]
5 or 10 million/ hNT cells/IC
University of Kondiolka et al., 2005 [14] Pittsburgh, PA & Stanford University, CA, USA
type/route
2 or 6 million/ hNT cells/IC
Sponsor/location
Kondziolka University of et al., 2000 [12] Pittsburgh, PA, USA
Ref.
Table 1. Published trials of cell therapy for stroke.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Epileptic-like activity on EEG at 6 and 12 months; no clinical seizures Cells not seen in brain by SPECT beyond 24 h Enrollment suspended due to concern regarding animal products in culture media
Adverse events in two patients, possibly unrelated. Some measures improved Improvements in two patients remained stable for 4 years Decreased exvacuo ventricular dilitation at 12 months in cell group, p = 0.019
Adverse events in two patients, possibly unrelated
Comments
Burns & Steinberg
449
Stem cells and stroke: opportunities, challenges and strategies
reach statistical significance. Reported adverse outcomes included one seizure and one subdural hematoma requiring evacuation [14]. Porcine fetal neural cells A major challenge of adult neural cell therapy is the relatively inhibitory environment presented by the adult brain to neurite outgrowth. It has been suggested that molecular differences between species may permit better engraftment of xenograft than allograft neurons [15]. In 2005, Savitz et al. published results from a trial employing stereotactic delivery of up to 50 million anti-MHC1 antibody-treated fetal porcine cells for stable basal ganglia stroke [16]. Of five patients enrolled, one showed temporary worsening of symptoms and another had a seizure. Both had questionably concerning findings on MRI, prompting the FDA to terminate enrollment. Two of the five patients experienced improvements in symptoms over several months that persisted at four years [16].
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
2.1.3
Bone marrow mononuclear cells (BMMCs) Endogenous bone-marrow-derived cells are swiftly recruited to regions of ischemic injury. Administration of supplemental bone marrow mononuclear cells has been under investigation in animal models of stroke since 2000 [17], with benefits attributed to various trophic mechanisms, in spite of largely poor long term survival. To date, BMMCs have appeared well tolerated in multiple small clinical trials, mostly involving intravascular delivery [18]. However, Suarez-Monteafudo et al. recently reported long-term, asymptomatic EEG abnormalities after intraparenchymal BMMC administration [19]. Interestingly, meta-analysis of BMMCs in clinical trials for acute myocardial infarction indicated a 4.77% improvement in left ventricular ejection fraction after three months [20]. G-CSF, which also has direct neurotrophic effects, may in part replicate the action of BBMCs by promoting mobilization of bone marrow cells. G-CSF is now under clinical investigation for stroke [21,22], having previously enabled a 3% increase in ejection fraction in meta-analysis of clinical trials for acute myocardial infarction [23]. 2.1.4
Mesenchymal stem cells (MSCs) By selectively culturing bone-marrow-derived cells that adhere to a culture dish in serum-containing media, cells variably termed marrow stromal cells or mesenchymal stem cells (MSCs) are generated that have shown benefit in animal models of stroke [24]. Recent meta-analysis of intravenouslydelivered cells in preclinical studies for stroke showed the beneficial effect of MSCs on behavioral outcome to be roughly twice that of BMMCs, consistent with the same study’s finding that cell lines and cultured or genetically modified cells are significantly more efficacious than primary cells [25]. Much literature has focused on conditions that may promote the neuronal differentiation of such cells either in vitro or in vivo after transplantation. However, few if any such claims withstand current standards of scrutiny [26,27]. 2.1.5
450
MSCs offer a somewhat more homogeneous and well characterized cell population for cell transplantation. These cultured cells are also amenable to genetic manipulation, allowing targeted delivery of specific therapeutic compounds. Both BMMCs and MSCs seem to have quite limited survival in the brain after either local or systemic delivery. Thus it is likely that benefits are mediated predominantly via trophic signals of variable duration. Though widely regarded as safe, some reports of MSC-derived sarcomas have appeared, suggesting that limits on passage numbers and stringent standards of cytogenetic quality control will be required for clinical applications [28]. Lee and colleagues recently published five year followup data from a previously reported [29] randomized open label trial of intravenous administration of two doses of 50 million autologous MSCs. Five year outcomes suggested significantly improved modified Rankin Scale scores, as assessed by blinded observers (p = 0.046). It is of interest that levels of stromal cell-derived factor-1 (SDF-1), which have been associated with MSC, as well as neural stem cell (NSC) homing, were found to correlate positively with clinical outcomes [30]. MSCs have been suggested to stimulate endogenous neurogenesis after stroke [31]. Thus, it is worth noting that patients in whom the subventricular zone (SVZ) was spared from infarct (n = 5) uniformly improved with MSC therapy, though outcomes in MSCtreated patients with infarct extending to the SVZ (n = 11) were more variable. Although no adverse effects were observed in MSC-treated patients within five years, recruitment was suspended due to the publication of concerns regarding use of xenogenic bovine calf serum in culture media for grafted cells [30]. Neural stem cells Techniques for the in vitro culture of neural stem cells were first described in the early 1990s by Reynolds and Weiss [32]. With inherent neurogenic potential, demonstrated trophic benefit and minimal risk of tumorgenicity, NSCs represent an excellent cell therapy choice and have been widely employed in pre-clinical stroke studies during the past 10 years with encouraging results [25,33]. Recent meta-analysis of preclinical studies employing intravenous cell delivery indicated that NSCs yielded the greatest behavioral improvements when compared with bone-marrow-derived or other cell types [25]. Due in part to regulatory delays, very few clinical trials have been initiated employing neural stem cells. NSClike olfactory ensheathing cells from the olfactory mucosa have been employed in a clinical trial for spinal cord injury with early establishment of safety and feasibility [34]. Neural stem cells from Stem Cells, Inc. were employed in a recently completed Phase I trial of six patients for neuronal ceroid lipofuscinosis, also known as Batten’s disease [35]. The results of this study remain to be published. An open label trial of the neural stem cell line CTX0E03, from ReNeuron, began in June 2010 and plans to enroll 12 patients for 2.1.6
Expert Opin. Biol. Ther. (2011) 11(4)
Burns & Steinberg
intraparenchymal delivery of 2 -- 20 million cells, 6 -- 12 months following subcortical stroke (Table 2). Embryonic stem cells (ESCs) and induced pluripotent stem (iPS) cells
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
2.1.7
ESCs possess the defining capacity to generate all cell types of a developing embryo under appropriate conditions. ESCs have received particular attention as a source of cells for which no reliable tissue-specific progenitor is available, such as cardiomyocytes, as well as certain neuronal lineages not readily obtainable from NSCs, including dopaminergic neurons for PD and motor neurons for amyotrophic lateral sclerosis (ALS). The recent development of techniques to generate ES-like cells via epigenetic reprogramming, termed induced pluripotent stem cells, or ‘iPS cells’, opens the potential for autologous neural cell therapy, thereby averting the need for immunosuppression. Safety concerns regarding the viral constructs used to reprogram iPS cells are being mitigated by the development of transient transfection techniques that leave cells genetically unaltered after reprogramming [36]. By definition, however, ESCs do generate teratomas. As such, the development of differentiation and culture techniques that eliminate any residual undifferentiated ESCs continues to be a high priority. Neural stem cell lines derived from hESCs have been generated that promote functional recovery in animal models of stroke without tumor formation, and are under development for future clinical applications [37,38]. Researchers at Wernig’s lab recently demonstrated that selective genetic reprogramming may enable direct transdifferentiation of fibroblasts to functional neuronal cells without the need for an intermediate ES or iPS cell stage [39]. In 2009, Geron received FDA approval to initiate the first ever clinical trial employing hESC-derived cells. This trial, for treatment of spinal cord injury, is based on the observation that pure cultures of hESC-derived oligodendrocyte precursors cells (OPCs) promote functional recovery by remyelinating axons in spinal cord-contused rats. A clinical hold imposed shortly after initial approval by the FDA for further safety evaluations was lifted on 30 July 2010, allowing the trial to proceed. Delivery variables for exogenous cell therapy For any given cell type, a number of options are available regarding when, where and how to implant, and what adjunctive treatments should additionally be administered (Table 3). The variety of protocols in use suggests that ‘right’ answers to these questions are not easily determined; optimal parameters may vary depending on the model, cell type, extent of injury and outcome measure being assessed. 2.2
Administration route Though not without risks, stereotactic delivery allows precise targeting of defined numbers of cells to desired sites, with best survival seen in the peri-infarct region. The first clinical trials of NT2 stem cells for chronic basal ganglia stroke 2.2.1
involved 25 cell deposits along 5 stereotactic tracts throughout the infarct area [40]. A recent protocol involving up to 88 deposits targeted selectively to the peri-lesion area was recently described for administration of MSCs [19]. Most studies of bone-marrow-derived cells to date have employed intravenous delivery. Meta-analysis of preclinical results suggests robust benefit, in spite of limited evidence for significant numbers of cells reaching the infarct site [25]. Intra-arterial or intra-carotid therapy has been advocated by several groups to facilitate delivery to the ischemic region and minimize cell sequestration in systemic tissues such as liver, lung and spleen [41]. With appropriate protocols to regulate cell density and allow continued blood flow during injection, risk of microembolic infarcts resulting from adherent cell clusters or vessel occlusion can be minimized [42]. Brain penetration of NSCs after intra-arterial delivery appears to be dependent upon upregulation of vascular cell adhesion molecule-1 (VCAM-1) following stroke, which binds the cell surface integrin CD49 that is expressed on the NSCs [43]. Future studies may assess whether or not genetic manipulation of receptor expression enhances targeting to the ischemic region. By comparison with stereotactic implantation, intravascular approaches have the advantage of readily allowing repeated administrations of cells. Combinations of intraparenchymal and intravascular therapies may also be feasible. Cell dosage Recent meta-analysis of intravenously-delivered cells in animal studies showed a robust effect of cell dosage on lesion size, behavioral outcome and molecular measures of outcome such as apoptosis, neurogenesis and angiogenesis [25]. Darsalia et al., recently demonstrated that the percentage of surviving cells is decreased with intraparenchymal delivery of larger numbers of cells. However, the total number of surviving cells trended upwards with incremental increases in cell dose [44]. The potential benefits of higher cell dose must be weighed against potential risks, including potential mass effects, theoretical risks of increased tumorgenicity in certain cells and potential for embolic events with intra-arterial delivery. As such, potential toxicity must be evaluated via appropriate dose--response analyses in preclinical studies [2]. 2.2.2
Immunosuppression The use of immunosuppression for cell therapy in CNS disorders is controversial [45]. Erlandsson et al., recently demonstrated that immunosuppression promotes endogenous progenitor migration and tissue regeneration with enhanced accumulation of SVZ-derived cells at the site of cortical injury [46]. By contrast, in meta-analysis of preclinical studies of IV-delivered stem cells, immunosuppression had no significant impact on behavioral outcomes, though a trend was noted towards more favorable outcomes without immunosuppression [25]. It should be noted that preclinical studies of human cell lines in animal models will almost always require 2.2.3
Expert Opin. Biol. Ther. (2011) 11(4)
451
452
Expert Opin. Biol. Ther. (2011) 11(4)
500 million/ autologous BMMC/IA versus IV Imperial College na/autologous London, UK CD34+ cells from bone marrow/IA Duke University, 5 million NC, USA cells/kg/ autologous cord blood/IV National 25 or 50 cc bone marrow/ Cardiovascular Center, Osaka, autologous Japan BMMCs/IV Hospital Universitario na/autologous Central de Asturias, CD34+ cells from Spain bone marrow/IA The University na/autologous of Texas Health BMMCs/IV Science Center, Houston, USA China Medical 2 -- 8 million/ University Hospital, autologous Taichung, Taiwan CD34+ cells from peripheral blood/IC Hospital Universitario, 8 -- 10 cc bone Nuevo Leon, Mexico marrow/CD34+ cells post G-CSF/IT Indian Council of 30-500 million/ Medical Research, BMMCs/IV New Delhi (multi-institution) Stempeutics Research 2 million cells/kg/ Pvt Ltd, Malaysia adult allogenic MSCs IV
Federal University of Rio de Janeiro, Brazil
type/route
Cell number/
Infarct
R: NIHSS 6 -- 15; L NIHSS 6 -- 18
NIHSS 9 -- 20
18 -- 80
35 -- 70
10
20
12
12
10
15
n
mRS < 5 with motor weakness
20 -- 80
< 10 days
NIHSS > 7; 7 -- 30 d Anterior circulation infarct
1 month -18 years
I
II
I
I/II
I/IIA
I
I/II
I
78
I/II
No
No
No
No
No
No
Yes
Adverse events, Yes NIHSS
Modified Barthel Yes Index, Safety
Battelle Devel. Inventory
NIHSS
March 2012
January 2011
January 2009
July 2009
June 2009
January 2009
December 2011
June 2011
August 2010
December 2010
January 2014
September March 2008 2010
May 2008
January 2008
September June 2010 2007
December June 2011 2005
Start date End date
Double blind May 2010
Open label
Open label
Open label
Open label
Single-blind (assessor)
Open label
Open label
Open label
Open label
Randomized? Blinding
Safety, feasibility No
Adverse events
NIHSS
Adverse events
Toxicity
Neurologic deficits
outcome
Phase Primary
120 II
10
6 -- 60 months 30
24 -- 72 h
18 -- 70
1 month -- Cerebral palsy 18 years
NIHSS > 7; MCA
18 -- 80
5 -- 9 days
7 -- 10 days
NIHSS > 9
20 -- 75
< 7 days
< 14 days
Total anterior circulation syndrome
30 -- 80
3 -- 90 days
window
Treatment
< 14 days Neonatal hypoxic/ ischemic injury
NIHSS 4 -- 20
severity/type 18 -- 75
Age
*Transient Notch 1 transfection. BMMC: Bone marrow mononuclear cell; ESS: European Stroke Scale; IA: Intra-arterial; IC: Intracerebral; IV: Intravenous; MCA: Middle cerebral artery; MSC: Mesenchymal stem cell; na: Not available; NIHSS: National Institutes of Health Stroke Scale.
NCT01091701 [118]
CTRI/2008/091/ 000046 [117]
NCT01019733 [116]
NCT00950521 [115]
NCT00859014 [114]
NCT00761982 [113]
NCT01028794 [112]
NCT00593242 [111]
NCT00535197 [110]
NCT00473057 [109]
Clinical trials ID [Ref.] Sponsor/location
Table 2. Ongoing and pending trials of cell therapy for stroke.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Stem cells and stroke: opportunities, challenges and strategies
na
NCT00908856 [121]
*Transient Notch 1 transfection. BMMC: Bone marrow mononuclear cell; ESS: European Stroke Scale; IA: Intra-arterial; IC: Intracerebral; IV: Intravenous; MCA: Middle cerebral artery; MSC: Mesenchymal stem cell; na: Not available; NIHSS: National Institutes of Health Stroke Scale.
na Open label Adverse effects, No acute and long-term safety 6 -- 12 months 18 2.5, 5.0, 10 million/ 18 -- 75 SB623 cells -modified* BMMCs/IC
mRS 3 -- 4; ESS 40 -- 50; Subcortical MCA or striatum +/cortex
I/II
Yes II 33 2 -- 21 days NIHSS 7 -- 24; supratentorial
30
January 2011
December 2013 December 2012 September 2010 Double blind January 2011 Open label Yes II < 14 days
University Hospital, Grenoble, France University of California, Irvine, CA, USA SanBio, Inc., Mountain View, CA, USA NCT00875654 [120]
n Age
Feasibility, tolerance Mortality
Adverse effects, No MRI, NIIHSS, antibodies I 6 -- 24 months 12
NHSS > 5; Subcortical white matter or basal ganglia NIHSS > 2
2, 5, 10, 60 -- 85 20 million/ CTX0E03 neural stem cells/IC na/autologous 18 -- 65 MSCs/IV 30 cc bone marrow/ 18 -- 85 autologous BMMCs/IV NCT01151124 [119]
ReNeuron, Ltd, Glasgow, UK
outcome window severity/type
Phase Primary Treatment Infarct
type/route
Cell number/ Clinical trials ID [Ref.] Sponsor/location
Table 2. Ongoing and pending trials of cell therapy for stroke (continued).
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Randomized? Blinding
Open label
June 2010
na
Start date End date
Burns & Steinberg
host immunosuppression, which, along with nature of the xenograft model itself, may significantly influence results. To date, autologous cells have been limited to bone marrow, and clinical trials of allogeneic cells for stroke have employed temporary immunosuppression [13]. Given the difficulty of accurately extrapolating immunosuppression results from animal studies, optimal protocols for allogenic cell grafts may be best derived in the setting of appropriately controlled clinical trials. Further development of iPS or nuclear reprogramming technologies may ultimately enable autologous human cells to be differentiated toward neural or neural stem cell lineages prior to grafting without need for immunosuppression. Timing The optimal timing for cell delivery is unclear, but may depend upon the predominant mechanism of action. Therapies aiming for neuroprotection will require earlier delivery than those targeting neuroplasticity. Some studies suggest optimal survival of transplanted cells at early time points (e.g., 48 h) before inflammatory responses are maximal [44], though studies have demonstrated robust benefit even when delivered over one month after stroke [47]. Meta-analysis of animal studies employing IV cell delivery found that the degree of inhibition of apoptosis was the strongest predictor of functional outcome [25]. This same study demonstrated a non-significant trend towards improved benefit with earlier cell delivery [25]. Stem cells have yet to be evaluated as adjuncts to thrombolysis or thrombectomy. However, pre-banked allogeneic cells may be feasibly delivered at quite early time points in this setting. Autologous therapies and intraparenchymal delivery will predictably be associated with greater delays. 2.2.4
Adjuncts Cellular grafts may be supplemented by extracellular matrix products to improve survival and neurite outgrowth [48,49] Cells may be genetically modified to secrete selected growth factors, with significantly enhanced therapeutic outcomes [25]. Alternatively, cells may be grafted after specialized pre-treatment protocols ranging from relative hypoxia to cytokine pretreatment or cellular co-culture. Delivery of cocktails containing multiple cell types, or adjunctive viral constructs is also feasible. Finally, cells grafted in clinical trials of stroke have predominantly been identified based on histopathological evaluation at autopsy. The modification of cells with appropriate transgenic or other cellular labels may markedly improve the capacity to track cells in vivo after transplantation, via MRI and/or positron-emission tomography (PET) [50]. 2.2.5
2.3
Proposed therapeutic mechanisms Integration
2.3.1
Graft-derived neurons can survive and mature, forming synaptic connections to host brain circuitry after transplantation of fetal [51], ESC-derived [52,53], and NSC-derived [54] cells into stroke-lesioned rodents. However, what role such integration, plays, if any, in functional recovery is unclear.
Expert Opin. Biol. Ther. (2011) 11(4)
453
Stem cells and stroke: opportunities, challenges and strategies
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Table 3. Overview of exogenous cell types. Exogenous cell type
Tumor risk
Cost
Survival
Autologous
Comments
Progress of clincal trials
Human fetal brain cells
-
$$$
+
-
None planned
Teratocarcinoma cells
+
$$
+
-
Porcine fetal neural cells
-
$$
+
-
Bone marrow mononuclear cells MSCs
-
$
+/-
+
+
$$
+/-
+/-
Neural stem cells
+
$$$
++
-
Embyonic-stem-derived cells
++
$$$$
++
+ (iPS only)
Limited availablility; ethical challenges First cell type in trials for stroke Complex immune considerations Most readily available cell type Trophic effects despite poor survival Robust efficacy data in animal studies iPS cells now available. May be used to generate NSCs
Early trials completed. No more planned Prior trial aborted by FDA. None planned Trials completed and in progress Trials completed and in progress Trials now starting Still in pre-clinical phase
+: Applicable; ++: Highly applicable; -: Not applicable; +/-: Variable; iPS: Induced pluripotent stem cell; MSC: Mesenchymal stem cell; NSC: Neural stem cell.
Benefits are frequently seen at early time points, well before grafted cells could mature and form synaptic connections. Benefits may also be seen in the presence of few, if any surviving cells, and after grafting of cells devoid of neurogenic potential. As such, popular consensus now favors trophic mechanisms as the predominant basis for functional gains after cell transplantation [55]. Selective ablation of grafted cells, for example via administration of diphtheria toxin in rodents after human cell grafting, would be needed to critically assess the requirement for ongoing graft survival for maintenance of functional gains [56]. No such studies in stroke-lesioned animals have yet been reported. In lesions involving cell death of select neuronal subpopulations with maintenance of surrounding cytoarchitecture, integration of grafted cells may be more feasible than after focal infarct and cavitation [57]. Use of supportive extracellular matricies may further maximize integration potential [58]. Neuroprotection Bone-marrow-derived and neural stem cells produce an impressive array of neuroprotective compounds. In a metaanalysis of 60 preclinical studies of intravenously-delivered cells, Janowski et al. demonstrated that outcomes were most strongly correlated with inhibition of apoptosis [25]. Less significant correlations were also found with neurogenesis and angiogenesis. Endogenous NSC-derived neuroblasts intrinsically migrate to the injured region following stroke and promote neuroprotection. This endogenous neuroprotection may be significantly bolstered by supplementation with exogenous cells. Careful attention to cell source may be important. Takahashi et al. found that NSCs derived from embryos were more effective than those derived from adults in mitigating ischemic damage [59]. Neuroprotection may be direct via secretion of neuroprotective compounds or indirect, 2.3.2
454
via immunomodulation, angiogenesis or amplifying the endogenous NSC response. Immunomodulation After acute ischemic injury, secondary injury may occur as a result of inflammatory mediators. Microglia are among the predominant regulators of the local inflammatory environment and may be modulated by grafted cells [60]. It should be noted that meta-analysis of preclinical studies employing intravenous cell delivery failed to find a significant correlation between immunomodulation and outcome [25]. However, the interactions between inflammatory signals and stem cells are notoriously complex and conflicting literature abounds. As a general rule, although SVZ and hippocampal neurogenesis increases after stroke, inflammatory signals following stroke impair neurogenesis [61]. Anti-inflammatory treatments, such as indomethacin, can increase neurogenesis following focal stroke [62]. It should be noted, however, that ablation of activated microglia exacerbates infarct size [63], consistent with an additional role of inflammation in the reparative process [64]. The immunosuppressive and anti-inflammatory effects of multiple stem cell populations are well documented [65]. Exactly when and how these effects impact outcomes following stroke requires further study. Unlike simple antiinflammatory treatments, it is conceivable that stem cells respond dynamically to changing inflammatory signals over time and may adjust their regulatory activities accordingly. Ideally, future studies should be undertaken in humanized mouse models to maximize insights relevant to human clinical therapies [66]. 2.3.3
Vascular repair The integral relationship between endothelial cells and neural progenitors is well established [67,68]. Neural precursors 2.3.4
Expert Opin. Biol. Ther. (2011) 11(4)
Burns & Steinberg
promote endothelial proliferation in the peri-infarct region. Conversely, such proliferation appears to enhance the recruitment of SVZ-derived neuroblasts [68]. Bone-marrow-derived stem cells similarly secrete multiple pro-angiogenic compounds including VEGF, EGF and IGF-1 in response to signals from ischemic brain [69], promoting endothelial proliferation in the peri-infarct region [68]. Blockade of angiogenesis in BMMC-treated cells markedly impedes recovery [70]. Plasticity The adult brain possesses much greater plasticity than previously appreciated. The spontaneous development of new host projections after stroke has been significantly correlated with behavioral recovery [71]. Moreover, mice devoid of thrombospondin 1 and 2, important for synapse formation and plasticity, demonstrate poor functional recovery after stroke [72]. Recent studies demonstrate that plasticity, as evidenced by increased synapse formation and new neuronal projections, is significantly enhanced by treatment with bone marrow-derived or neural stem cells following stroke [60,73].
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
2.3.5
Recruitment of endogenous neural progenitors
2.3.6
Neural stem cells proliferate and give rise to neuroblasts that migrate toward the injured region following stroke. These neuroblasts exert neuroprotective and pro-angiogenic effects upon arrival in the peri-infarct region. Bone-marrow-derived stem cells are known to secrete a variety of compounds that promote the proliferation and migration of endogenous neural progenitor cells, suggesting that exogenous cell therapy may act, in part, by augmenting the endogenous neurogenic response to stroke [30,74]. 3.
Stroke-induced neurogenesis
In the adult brain, neural stem cells are located in the hippocampal dentate gyrus [75] and SVZ [76] that give rise to new functional neurons throughout life. Hippocampal NSCs modulate learning, memory and spatial navigation as well as psychiatric states [77]. In the SVZ, slowly dividing stem cells generate transit amplifying cells, which in turn generate neuroblasts [78]. Unlike hippocampal NSC progeny that remain in the dentate gyrus, SVZ neuroblasts migrate along the rostral migratory stream (RMS) to generate functional olfactory bulb neurons [79], though they can be redirected towards areas of injury. Subventricular zone response to focal infarcts After ischemic stroke, hypoxia-induced signals promote the proliferation of neural stem and progenitor cells. SDF-1 and angiopoietin redirect neuroblasts from the SVZ and RMS along blood vessels toward the infarct region [80]. Rare new neurons are generated, though most recruited cells die or remain undifferentiated in association with blood vessels near the infarct boundary zone [80,81]. Increased progenitor 3.1
cell proliferation and neuroblast recruitment may persist for at least several months after ischemic injury [82]. Given the very low numbers of new neurons generated, the relevance of stroke-induced neurogenesis to functional recovery has been controversial, and has been examined via several experiments in the past decade employing irradiation or chemotherapeutic agents to impede neurogenesis. These manipulations worsened stroke outcomes, but conclusions were tentative, given possible toxicity from the experimental treatment. In 2006, Won et al. demonstrated that reelin mice lacking doublecortin (dcx), a protein required for neuroblast migration, had larger infarcts and worsened behavioral outcomes following stroke [83]. However, baseline behavioral deficits in these mice somewhat hampered interpretation. In 2009, Jim et al. selectively ablated migrating dcx+ cells and similarly observed increased infarct size as well as substantially worsened behavioral scores within days following stroke [84]. These studies collectively imply that immature endogenous neuroblasts act locally at the peri-infarct region to promote neuronal survival, well before any new mature neurons could possibly be generated. The exact mechanisms via which endogenous neuroblasts enhance outcomes are incompletely defined. However, NSCs are known to produce neurotrophic factors such as nerve growth factor (NGF) and glial-cell-derived neurotrophic factor (GDNF), to regulate the inflammatory environment, and to produce pro-angiogenic complexes including netrin-4, laminin and integrins [85]. Notably, NSCs constitutively secrete factors implicated in synaptic plasticity, including those that are considered anti-angiogenic, such as thrombospondins [72]. It is likely that the factors generated by NSC progeny vary dynamically as the infarct injury evolves. The specific factors generated by endogenous recruited neuroblasts and their temporal patterns of expression after ischemia remain to be evaluated. Such analysis may provide fundamental insights regarding the endogenous response to ischemic injury. A plethora of signaling molecules including VEGF, brain derived neurotrophic factor (BDNF), erythropoietin (EPO), fibroblast growth factor 2 (FGF2), noggin, notch, IGF-1, TGF-alpha, stem cell factor (SCF), nitric oxide (NO), EGF, angiopoietin, microphage-interacting protein-1-alpha (MIP 1-alpha), stromal cell-derived factor 1 (SDF-1), cell surface molecules, including CXCR4, vascular cell adhesion molecule (VCAM), integrins and extracellular matrix molecules, are now known to regulate NSC proliferation, migration and differentiation [86,87]. Experimental manipulation of these pathways in rodents can substantially bolster the neurogenic response with subsequently decreased lesion size and improved functional outcomes [88]. The clinical implications of these findings may be substantial. Manipulations that increase progenitor proliferation and migration with improvements in functional outcomes may be observed without significant increases in the number of new post-mitotic neurons generated. However, increased numbers of number of new neurons may be observed with overexpression of basic fibroblast growth
Expert Opin. Biol. Ther. (2011) 11(4)
455
Stem cells and stroke: opportunities, challenges and strategies
factor (bFGF) [82]. Moreover, ‘filling’ of the infarct cavity with new neurons may be attained with sequential administration of EPO and EGF [89]. While robust functional improvements are observed in these cases, the capacity of the new neurons to form functional connections with surrounding circuitry or contribute to functional recovery remains to be assessed. Selective neuronal replacement by endogenous progenitors
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
3.2
In animal models of selective neuronal loss, endogenous neural progenitors appear more inclined to replace the lost cell type. CA1 neurons in the hippocampus are particularly susceptible to ischemic injury [90]. Spontaneous regeneration of CA1 cells was observed after hypoxic injury, with cells arising from the periventricular region. This regenerative response could be significantly augmented by intraventricular delivery of EGF and bFGF. Functional recovery occurred in a delayed manner, which correlated with the appearance, maturation and electrophysiological integration of new neurons. In another example, Macklis’ lab developed a selective photoablation technique to delete subsets of cortical projection neurons without hypoxia, ischemia or other injury. Remarkably, SVZ-derived neurons appeared to migrate along the corpus callosum to the affected region, and then replace ablated neurons with subsequent generation of long distance corticospinal and corticothalamic projections [91,92]. The signals responsible for recruitment and directed differentiation of the new cortical projection neurons in this case remain unclear, though would be of fundamental importance to any attempts to promote regeneration after cortical injury. New neurons from cortical progenitors? Increasingly frequent reports of cortical progenitor cells with neurogenic potential have appeared in recent years. Ohira et al. recently found that the rat layer 1 cortex contains Ki67+/67 kDa glutamic acid decarboxylase (GAD67)+ cells that bear few of the usual NSC markers, and generate no new neurons under baseline conditions. Impressively, in response to hypoxic injury, these cells generated new inhibitory neurons throughout the cortex that appeared to integrate into local circuitry and survive for at least eight weeks [93]. These cells probably correspond to the mitotically active glial fibrillary acidic protein (GFAP)+ cells in cortical layer 1 described recently by Xui et al. [94]. These cells expressed vimentin and nestin in response to a cortical insult, migrating into deeper cortical layers over subsequent days and assuming an immature neuron-like morphology. The tendency of these cells to generate GABAergic neurons suggests they may represent residual undifferentiated progeny from the medial ganglionic eminence where cortical interneurons originate during development. The factors regulating the behavior of this newly identified population of cells and their response to focal ischemic injury remain to be evaluated. Certain astroglial cells from the cortex or subcortical white matter may also de-differentiate into neural stem cells after 3.3
456
in vitro culture [95,96]. Heinrich et al. recently demonstrated that reactive adult astrocytes isolated after cortical injury can be differentiated into glutamatergic or GABAergic neurons after transduction with Neurogenein2, or distal-less homeobox 2 (Dlx2), respectively. These neurons formed mature synapses with electrophysiological properties of mature neurons in vitro. Whether or not a similar strategy may be employed to redirect glial cells toward neurogenic fates in vivo after cortical injury remains to be determined [97]. Some have argued that small cortical infarcts that spare the striatum provide a poor stimulus for SVZ neuroblast migration and that neural progenitors surrounding such infarcts may be locally derived [98]. Lineage tracing studies will be needed to fully characterize the identity, behavior and function of naturally occurring cortical progenitor cells. Human implications Though distinct in structure from those of rodents, the human subventricular zone [99] and rostral migratory stream [100] have been characterized. Moreover, evidence for neuroblasts has been found in multiple studies of patients after ischemic and hemorrhagic stroke [101-103]. As such, strategies to augment the human neurogenic response may yield improved outcomes. As always, rigorous preclinical safety studies will be needed to ensure factors employed to mobilize endogenous neural stem cells are safe. For example, EGF and BDNF have both been implicated in the development of glioma-like growths from SVZ progenitors [104,105], BDNF may induce spontaneous seizure activity [106], bFGF infusion has resulted in demyelination [107] and all-causes mortality was elevated in a large clinical trial of EPO for treatment of acute ischemic stroke [108]. Nevertheless, appropriate dosing in animal studies has enabled marked benefits with several cytokines without clinically untoward effects. 3.4
4.
Conclusion
Early recanalization remains the most effective treatment for acute stroke by minimizing infarct size. Neural and bonemarrow-derived stem cells appear to function via multiple synergistic mechanisms to augment natural recovery mechanisms. In the acute setting, endogenous endothelial and neural progenitor cells work together to minimize neuronal cell death and both may be bolstered by signals from exogenous stem cells. Both neural and bone marrow-derived stem cells directly secrete pro-survival factors in addition to modulating the endogenous response to stroke, thereby maximizing the amount of original neural circuitry that survives the ischemic insult. Thereafter, stem cells and their progeny function to promote synaptic plasticity, optimizing functional recovery. Multiple cellular and molecular tools now exist to enhance endogenous responses to stroke. While much work lies ahead, ample proof of principle suggests that substantial benefits may reward an ongoing investment in the science and translation of stem cell therapies for stroke.
Expert Opin. Biol. Ther. (2011) 11(4)
Burns & Steinberg
5.
Expert opinion
Unique challenges of stem cell therapy for stroke The failure of hundreds of neuroprotective compounds in clinical trials illustrates the sobering challenge ahead of translational therapy for stroke. Given the inherently heterogeneous nature of stroke, clinical trials will be prone to inadequate power, being based on preclinical data from models that imperfectly reflect human disease. They may also suffer from being based on a literature that is skewed towards the publication of only positive results. Collaborative strategies will be needed to ensure not only that scientifically wellfounded studies are initiated, but also that such potentially beneficial therapies are not aborted prematurely due to Type II errors. Considerations of statistical practicality and therapeutic potential may be in conflict when therapeutic windows are selected for trials. Should cells be given early when there is potentially more to gain, or later, when the gains can be most accurately measured from a stable baseline? As a novel technology, stem cells offer new risks, not only to patients, but also to the field as a whole, should early complications undermine public support. Will randomized doubleblind studies be acceptable for therapies that may be invasive and risky, including possible stereotactic intracerebral cell delivery and immunosuppression? Ongoing collaborative discussions between experts on all sides of the negotiations will be essential. While stem cell therapies have attracted significant media hype and venture capital, transparency regarding realistic expectations is critical. Regulatory bodies, stake holders and the public at large should all be prepared for a long term investment that is more likely to be marked by small steps, than home runs. Refinement of protocols related to cell preparation, delivery and detection after the onset of human studies will require diligence and patience.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
5.1
Pre-clinical directions Several preclinical questions remain that are pertinent to translational efforts. How long must cells survive for therapeutic benefit? Simple timed ablation studies, for example, using diphtheria toxin, are needed and may guide decisions regarding immunosuppression and cell labeling in clinical trials. Selective ablation of specific graft-derived cell types using cre--lox technology may help to dissect mechanisms underlying long-term functional benefits. Knockdown of putative therapeutic genes in transplanted cells may further illuminate molecular mechanisms of benefit. The endogenous neurogenic response remains poorly characterized. Lineage tracing labels based on selective markers such as dcx should be used for isolation and gene profiling of endogenous neuroblasts at various times post-infarct. Subgroups of 5.2
newly born neurons should also be ablated after augmentation studies to assess their functional contributions to recovery. Studies employing humanized mice may yield clinically relevant insights regarding immunomodulatory effects of grafted and mobilized cells, while guiding decisions regarding immunosuppression. Clinical directions In the treatment of stroke, it remains true that ‘time is brain’. Cell therapies should be developed in conjunction with optimized recanalization technologies to target residual areas of ischemia, combat reperfusion injury and provide trophic support in areas of hemorrhagic conversion. Defined ex vivo genetic modifications of grafted cell lines should be considered to introduce transgenic MRI-detectable labels, as well as inducible suicide genes as insurance against undesired proliferation or neoplastic transformation. Transgenes may also permit delivery of complementary therapeutic genes. Inducible viral constructs should also be prepared for in vivo or cell-based delivery of cytokines for mobilization of endogenous NSCs. Use of bicistronic constructs with MRI labels may facilitate monitoring for effects upon transduced endogenous cells, while regulatory elements may improve safety in case of untoward side effects. Therapeutic candidates may be expanded to include intracerebral hemorrhage, with stereotactic cell delivery following stereotactic clot evacuation. Additionally, global ischemic injury after myocardial infarction may be amenable to cytokine augmentation of endogenous cell replacement. Upon establishment of safety, stem cell pretreatment may be indicated for high-risk patients, such as in subarachnoid hemorrhage patients at risk for vasospasm, and patients undergoing embolizations, complex tumor resections or cerebral revascularization procedures. In sum, although challenges abound, and while vigilance regarding safety and monitoring will be paramount, maximal efforts are indicated to ensure timely translation of the most promising therapies for the treatment of stroke. 5.3
Acknowledgements The authors wish to thank C Samos for editorial assistance.
Declaration of interest The authors are supported in part by funding from the William Randolph Hearst Foundation, and Bernard and Ronni Lacroute. GK Steinberg has received grants from the National Institute of Neurological Disorders and Stroke (NINDS) and the California Institute of Regenerative Medicine. TC Burns declares no conflict of interest.
Expert Opin. Biol. Ther. (2011) 11(4)
457
Stem cells and stroke: opportunities, challenges and strategies
Bibliography 1.
Saver JL. Time is brain -- quantified. Stroke 2006;37:263-6
2.
Stem Cell Therapies as an Emerging Paradigm in Stroke Participants. Stem Cell Therapies as an Emerging Paradigm in Stroke (STEPS): bridging basic and clinical science for cellular and neurogenic factor therapy in treating stroke. Stroke 2009;40:510-15
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
3.
4.
Bliss TM, Andres RH, Steinberg GK. Optimizing the success of cell transplantation therapy for stroke. Neurobiol Dis 2010;37:275-83 Bersano A, Ballabio E, Lanfranconi S, et al. Clinical studies in stem cells transplantation for stroke: a review. Curr Vasc Pharmacol 2010;8:29-34
5.
Locatelli F, Bersano A, Ballabio E, et al. Stem cell therapy in stroke. Cell Mol Life Sci 2009;66:757-72
6.
Burns TC, Verfaillie CM, Low WC. Stem cells for ischemic brain injury: a critical review. J Comp Neurol 2009;515:125-44
7.
8.
9.
Polezhaev LV, Alexandrova MA, Girman SV. Normalization of dystrophic brain cortex neurons after hypoxia and transplantation of embryonic nervous tissue in rats. J Hirnforschung 1986;27:501-13 Hadani M, Freeman T, Munsiff A, et al. Fetal cortical cells survive in focal cerebral infarct after permanent occlusion of the middle cerebral artery in adult rats. J Neurotrauma 1992;9:107-12 Aihara N, Mizukawa K, Koide K, et al. Striatal grafts in infarct striatopallidum increase GABA release, reorganize GABAA receptor and improve water-maze learning in the rat. Brain Res Bull 1994;33:483-8
10.
Andrews PW. Retinoic acid induces neuronal differentiation of a cloned human embryonal carcinoma cell line in vitro. Dev Biol 1984;103:285-93
11.
Borlongan CV, Tajima Y, Trojanowski JQ, et al. Transplantation of cryopreserved human embryonal carcinoma-derived neurons (NT2N cells) promotes functional recovery in ischemic rats. Exp Neurol 1998;149:310-21
12.
Kondziolka D, Wechsler L, Goldstein S, et al. Transplantation of cultured human neuronal cells for patients with stroke. Neurology 2000;55:565-9
458
13.
Nelson PT, Kondziolka D, Wechsler L, et al. Clonal human (hNT) neuron grafts for stroke therapy: neuropathology in a patient 27 months after implantation. Am J Pathol 2002;160:1201-6
acute ischemic stroke: a randomized controlled trial. CMAJ 2006;174:927-33 23.
Kang S, Yang Y, Li CJ, Gao R. Effectiveness and tolerability of administration of granulocyte colony-stimulating factor on left ventricular function in patients with myocardial infarction: a meta-analysis of randomized controlled trials. Clin Ther 2007;29:2406-18
14.
Kondziolka D, Steinberg GK, Wechsler L, et al. Neurotransplantation for patients with subcortical motor stroke: a phase 2 randomized trial. J Neurosurg 2005;103:38-45
15.
Deacon T, Schumacher J, Dinsmore J, et al. Histological evidence of fetal pig neural cell survival after transplantation into a patient with Parkinson’s disease. Nat Med 1997;3:350-3
24.
Chen J, Li Y, Wang L, et al. Therapeutic benefit of intracerebral transplantation of bone marrow stromal cells after cerebral ischemia in rats. J Neurol Sci 2001;189:49-57
16.
Savitz SI, Dinsmore J, Wu J, et al. Neurotransplantation of fetal porcine cells in patients with basal ganglia infarcts: a preliminary safety and feasibility study. Cerebrovasc Dis 2005;20:101-7
25.
Janowski M, Walczak P, Date I. Intravenous route of cell delivery for treatment of neurological disorders: a meta-analysis of preclinical results. Stem Cells Dev 2010;19:5-16
26.
17.
Chen J, Li Y, Chopp M. Intracerebral transplantation of bone marrow with BDNF after MCAo in rat. Neuropharmacology 2000;39:711-16
18.
Barbosa da Fonseca LM, Gutfilen B, Rosado de Castro PH, et al. Migration and homing of bone-marrow mononuclear cells in chronic ischemic stroke after intra-arterial injection. Exp Neurol 2010;221:122-8
Burns TC, Ortiz-Gonzalez XR, Gutierrez-Perez M, et al. Thymidine analogs are transferred from prelabeled donor to host cells in the central nervous system after transplantation: a word of caution. Stem Cells 2006;24:1121-7
27.
Breunig JJ, Arellano JI, Macklis JD, Rakic P. Everything that glitters isn’t gold: a critical review of postnatal neural precursor analyses. Cell Stem Cell 2007;1:612-27
28.
Tolar J, Nauta AJ, Osborn MJ, et al. Sarcoma derived from cultured mesenchymal stem cells. Stem Cells 2007;25:371-9
29.
Bang OY, Lee JS, Lee PH, Lee G. Autologous mesenchymal stem cell transplantation in stroke patients. Ann Neurol 2005;57:874-82
30.
Lee JS, Hong JM, Moon GJ, et al. A long-term follow-up study of intravenous autologous mesenchymal stem cell transplantation in patients with ischemic stroke. Stem Cells 2010;28:1099-106
31.
Chen J, Li Y, Katakowski M, et al. Intravenous bone marrow stromal cell therapy reduces apoptosis and promotes endogenous cell proliferation after stroke in female rat. J Neurosci Res 2003;73:778-86
32.
Reynolds BA, Weiss S. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 1992;255:1707-10
19.
20.
21.
22.
Suarez-Monteagudo C, Hernandez-Ramirez P, Alvarez-Gonzalez L, et al. Autologous bone marrow stem cell neurotransplantation in stroke patients. An open study. Restor Neurol Neurosci 2009;27:151-61 Zhang SN, Sun AJ, Ge JB, et al. Intracoronary autologous bone marrow stem cells transfer for patients with acute myocardial infarction: a meta-analysis of randomised controlled trials. Int J Cardiol 2009;136:178-85 Sprigg N, Bath PM, Zhao L, et al. Granulocyte-colony-stimulating factor mobilizes bone marrow stem cells in patients with subacute ischemic stroke: the Stem cell Trial of recovery EnhanceMent after Stroke (STEMS) pilot randomized, controlled trial (ISRCTN 16784092). Stroke 2006;37:2979-83 Shyu WC, Lin SZ, Lee CC, et al. Granulocyte colony-stimulating factor for Expert Opin. Biol. Ther. (2011) 11(4)
Burns & Steinberg
33.
34.
35.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
36.
37.
38.
Park KI. Transplantation of neural stem cells: cellular & gene therapy for hypoxic-ischemic brain injury. Yonsei Med J 2000;41:825-35
44.
Darsalia V, Allison SJ, Cusulin C, et al. Cell number and timing of transplantation determine survival of human neural stem cell grafts in stroke-damaged rat brain. J Cereb Blood Flow Metab 2011;31:235-42
Mackay-Sim A, Feron F, Cochrane J, et al. Autologous olfactory ensheathing cell transplantation in human paraplegia: a 3-year clinical trial. Brain 2008;131:2376-86
45.
Selden NR, Guillaume DJ, Steiner RD, Huhn SL. Cellular therapy for childhood neurodegenerative disease. Part II: clinical trial design and implementation. Neurosurg Focus 2008;24(3-4):E23
Piccini P, Pavese N, Hagell P, et al. Factors affecting the clinical outcome after neural transplantation in Parkinson’s disease. Brain 2005;128:2977-86
46.
Erlandsson A, Lin A, Yu F, Morshead CM. Immunosuppression promotes endogenous neural stem and progenitor cell migration and tissue regeneration after ischemic injury. Exp Neurol 2010. [Epub ahead of print]
Gonzalez F, Barragan Monasterio M, Tiscornia G, et al. Generation of mouse-induced pluripotent stem cells by transient expression of a single nonviral polycistronic vector. Proc Natl Acad Sci USA 2009;106:8918-22 Daadi MM, Steinberg GK. Manufacturing neurons from human embryonic stem cells: biological and regulatory aspects to develop a safe cellular product for stroke cell therapy. Regen Med 2009;4:251-63 Daadi MM, Maag AL, Steinberg GK. Adherent self-renewable human embryonic stem cell-derived neural stem cell line: functional engraftment in experimental stroke model. PLoS One 2008;3(2):e1644
47.
48.
49.
Shen LH, Li Y, Chen J, et al. Therapeutic benefit of bone marrow stromal cells administered 1 month after stroke. J Cereb Blood Flow Metab 200;27:6-13 Zhong J, Chan A, Morad L, et al. Hydrogel matrix to support stem cell survival after brain transplantation in stroke. Neurorehabil Neural Repair 2010;24:636-44 Park KI, Teng YD, Snyder EY. The injured brain interacts reciprocally with neural stem cells supported by scaffolds to reconstitute lost tissue. Nat Biotechnol 2002;20:1111-7
39.
Vierbuchen T, Ostermeier A, Pang ZP, et al. Direct conversion of fibroblasts to functional neurons by defined factors. Nature 2010;463:1035-41
50.
Gera A, Steinberg GK, Guzman R. In vivo neural stem cell imaging: current modalities and future directions. Regen Med 2010;5:73-86
40.
Kondziolka D, Steinberg GK, Cullen SB, McGrogan M. Evaluation of surgical techniques for neuronal cell transplantation used in patients with stroke. Cell Transplant 2004;13:749-54
51.
Daadi MM, Lee SH, Arac A, et al. Functional engraftment of the medial ganglionic eminence cells in experimental stroke model. Cell Transplant 2009;18:815-26
41.
Pendharkar AV, Chua JY, Andres RH, et al. Biodistribution of neural stem cells after intravascular therapy for hypoxic-ischemia. Stroke 2010;41:2064-70
52.
Buhnemann C, Scholz A, Bernreuther C, et al. Neuronal differentiation of transplanted embryonic stem cell-derived precursors in stroke lesions of adult rats. Brain 2006;129:3238-48
42.
Chua JY, Gaeta X, Wang N, et al. Intraarterial neural stem cell delivery does not reduce cerebral blood flow. J Cereb Blood Flow Metab 2009;29:S561
53.
43.
Guzman R, De Los Angeles A, Cheshier S, et al. Intracarotid injection of fluorescence activated cell-sorted CD49d-positive neural stem cells improves targeted cell delivery and behavior after stroke in a mouse stroke model. Stroke 2008;39:1300-6
Daadi MM, Li Z, Arac A, et al. Molecular and magnetic resonance imaging of human embryonic stem cell-derived neural stem cell grafts in ischemic rat brain. Mol Ther 2009;17:1282-91
54.
Jin K, Mao X, Xie L, et al. Transplantation of human neural precursor cells in Matrigel scaffolding improves outcome from focal cerebral ischemia after delayed postischemic
Expert Opin. Biol. Ther. (2011) 11(4)
treatment in rats. J Cereb Blood Flow Metab 2010;30:534-44 55.
Guzman R. Cellular stroke therapy: from cell replacement to trophic support. Expert Rev Cardiovasc Ther 2009;7:1187-90
56.
Hooshmand MJ, Sontag CJ, Uchida N, et al. Analysis of host-mediated repair mechanisms after human CNS-stem cell transplantation for spinal cord injury: correlation of engraftment with recovery. PLoS One 2009;4(6):e5871
57.
Mudrick LA, Baimbridge KG, Peet MJ. Hippocampal neurons transplanted into ischemically lesioned hippocampus: electroresponsiveness and reestablishment of circuitries. Exp Brain Res 1989;76:333-42
58.
Ourednik J, Ourednik V, Lynch WP, et al. Neural stem cells display an inherent mechanism for rescuing dysfunctional neurons. Nat Biotechnol 2002;20:1103-10
59.
Takahashi K, Yasuhara T, Shingo T, et al. Embryonic neural stem cells transplanted in middle cerebral artery occlusion model of rats demonstrated potent therapeutic effects, compared to adult neural stem cells. Brain Res 2008;1234:172-82
60.
Daadi MM, Davis AS, Arac A, et al. Human neural stem cell grafts modify microglial response and enhance axonal sprouting in neonatal hypoxic-ischemic brain injury. Stroke 2010;41:516-23
61.
Iosif RE, Ahlenius H, Ekdahl CT, et al. Suppression of stroke-induced progenitor proliferation in adult subventricular zone by tumor necrosis factor receptor 1. J Cereb Blood Flow Metab 2008;28:1574-87
62.
Hoehn BD, Palmer TD, Steinberg GK. Neurogenesis in rats after focal cerebral ischemia is enhanced by indomethacin. Stroke 2005;36:2718-24
63.
Lalancette-Hebert M, Gowing G, Simard A, et al. Selective ablation of proliferating microglial cells exacerbates ischemic injury in the brain. J Neurosci 2007;27:2596-605
64.
Lo EH. A new penumbra: transitioning from injury into repair after stroke. Nat Med 2008;14:497-500
65.
Djouad F, Plence P, Bony C, et al. Immunosuppressive effect of mesenchymal stem cells favors tumor
459
Stem cells and stroke: opportunities, challenges and strategies
adult mammalian brain. J Neurosci 1997;17:5046-61
growth in allogeneic animals. Blood 2003;102:3837-44 Shultz LD, Ishikawa F, Greiner DL. Humanized mice in translational biomedical research. Nat Rev Immunol 2007;7:118-30
77.
67.
Shen Q, Goderie SK, Jin L, et al. Endothelial cells stimulate self-renewal and expand neurogenesis of neural stem cells. Science 2004;304:1338-40
78.
Lois C, Alvarez-Buylla A. Long-distance neuronal migration in the adult mammalian brain. Science 1994;264:1145-8
68.
Nakano-Doi A, Nakagomi T, Fujikawa M, et al. Bone marrow mononuclear cells promote proliferation of endogenous neural stem cells through vascular niches after cerebral infarction. Stem Cells 2010;28:1292-302
79.
Rochefort C, Gheusi G, Vincent JD, Lledo PM. Enriched odor exposure increases the number of newborn neurons in the adult olfactory bulb and improves odor memory. J Neurosci 2002;22:2679-89
Qu R, Li Y, Gao Q, et al. Neurotrophic and growth factor gene expression profiling of mouse bone marrow stromal cells induced by ischemic brain extracts. Neuropathology 2007;27:355-63
80.
Taguchi A, Soma T, Tanaka H, et al. Administration of CD34+ cells after stroke enhances neurogenesis via angiogenesis in a mouse model. J Clin Invest 2004;114:330-8
81.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
66.
69.
70.
71.
72.
73.
74.
75.
76.
460
Liu Z, Zhang RL, Li Y, et al. Remodeling of the corticospinal innervation and spontaneous behavioral recovery after ischemic stroke in adult mice. Stroke 2009;40:2546-51
82.
Shors TJ, Miesegaes G, Beylin A, et al. Neurogenesis in the adult is involved in the formation of trace memories. Nature 2001;410:372-6
Zhang RL, Chopp M, Gregg SR, et al. Patterns and dynamics of subventricular zone neuroblast migration in the ischemic striatum of the adult mouse. J Cereb Blood Flow Metab 2009;29:1240-50 Arvidsson A, Collin T, Kirik D, et al. Neuronal replacement from endogenous precursors in the adult brain after stroke. Nat Med 2002;8:963-70 Leker RR, Soldner F, Velasco I, et al. Long-lasting regeneration after ischemia in the cerebral cortex. Stroke 2007;38:153-61
83.
Liauw J, Hoang S, Choi M, et al. Thrombospondins 1 and 2 are necessary for synaptic plasticity and functional recovery after stroke. J Cereb Blood Flow Metab 2008;28:1722-32
Won SJ, Kim SH, Xie L, et al. Reelin-deficient mice show impaired neurogenesis and increased stroke size. Exp Neurol 2006;198:250-9
84.
Liu Z, Li Y, Zhang ZG, et al. Bone marrow stromal cells enhance inter- and intracortical axonal connections after ischemic stroke in adult rats. J Cereb Blood Flow Metab 2010;30:1288-95
Jin K, Wang X, Xie L, et al. Transgenic ablation of doublecortin-expressing cells suppresses adult neurogenesis and worsens stroke outcome in mice. Proc Natl Acad Sci USA 2010;107:7993-8
85.
Yoo SW, Kim SS, Lee SY, et al. Mesenchymal stem cells promote proliferation of endogenous neural stem cells and survival of newborn cells in a rat stroke model. Exp Mol Med 2008;40:387-97
86.
van Praag H, Schinder AF, Christie BR, et al. Functional neurogenesis in the adult hippocampus. Nature 2002;415:1030-4 Doetsch F, Garcia-Verdugo JM, Alvarez-Buylla A. Cellular composition and three-dimensional organization of the subventricular germinal zone in the
87.
Staquicini FI, Dias-Neto E, Li J, et al. Discovery of a functional protein complex of netrin-4, laminin gamma1 chain, and integrin alpha6beta1 in mouse neural stem cells. Proc Natl Acad Sci USA 2009;106:2903-8 Snapyan M, Lemasson M, Brill MS, et al. Vasculature guides migrating neuronal precursors in the adult mammalian forebrain via brain-derived neurotrophic factor signaling. J Neurosci 2009;29:4172-88 Abrous DN, Koehl M, Le Moal M. Adult neurogenesis: from precursors to network and physiology. Physiol Rev 2005;85:523-69
Expert Opin. Biol. Ther. (2011) 11(4)
88.
Leker RR. Fate and manipulations of endogenous neural stem cells following brain ischemia. Expert Opin Biol Ther 2009;9:1117-25
89.
Kolb B, Morshead C, Gonzalez C, et al. Growth factor-stimulated generation of new cortical tissue and functional recovery after stroke damage to the motor cortex of rats. J Cereb Blood Flow Metab 2007;27:983-97
90.
Nakatomi H, Kuriu T, Okabe S, et al. Regeneration of hippocampal pyramidal neurons after ischemic brain injury by recruitment of endogenous neural progenitors. Cell 2002;110:429-41
91.
Chen J, Magavi SS, Macklis JD. Neurogenesis of corticospinal motor neurons extending spinal projections in adult mice. Proc Natl Acad Sci USA 2004;101:16357-62
92.
Magavi SS, Leavitt BR, Macklis JD. Induction of neurogenesis in the neocortex of adult mice. Nature 2000;405:951-5
93.
Ohira K, Furuta T, Hioki H, et al. Ischemia-induced neurogenesis of neocortical layer 1 progenitor cells. Nat Neurosci 2010;13:173-9
94.
Xue JH, Yanamoto H, Nakajo Y, et al. Induced spreading depression evokes cell division of astrocytes in the subpial zone, generating neural precursor-like cells and new immature neurons in the adult cerebral cortex. Stroke 2009;40:e606-13
95.
Buffo A, Vosko MR, Erturk D, et al. Expression pattern of the transcription factor Olig2 in response to brain injuries: implications for neuronal repair. Proc Natl Acad Sci USA 2005;102:18183-8
96.
Nunes MC, Roy NS, Keyoung HM, et al. Identification and isolation of multipotential neural progenitor cells from the subcortical white matter of the adult human brain. Nat Med 2003;9:439-47
97.
Heinrich C, Blum R, Gascon S, et al. Directing astroglia from the cerebral cortex into subtype specific functional neurons. PLoS Biol 2010;8(5):e1000373
98.
Shimada IS, Peterson BM, Spees JL. Isolation of locally derived stem/ progenitor cells from the peri-infarct area that do not migrate from the lateral ventricle after cortical stroke. Stroke 2010;41:e552-60
Burns & Steinberg
99.
Quinones-Hinojosa A, Sanai N, Soriano-Navarro M, et al. Cellular composition and cytoarchitecture of the adult human subventricular zone: a niche of neural stem cells. J Comp Neurol 2006;494:415-34
100.
Kam M, Curtis MA, McGlashan SR, et al. The cellular composition and morphological organization of the rostral migratory stream in the adult human brain. J Chem Neuroanat 2009;37:196-205
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
101.
102.
Marti-Fabregas J, Romaguera-Ros M, Gomez-Pinedo U, et al. Proliferation in the human ipsilateral subventricular zone after ischemic stroke. Neurology 2010;74:357-65
Shen J, Xie L, Mao X, et al. Neurogenesis after primary intracerebral hemorrhage in adult human brain. J Cereb Blood Flow Metab 2008;28:1460-8
104.
Doetsch F, Petreanu L, Caille I, et al. EGF converts transit-amplifying neurogenic precursors in the adult brain into multipotent stem cells. Neuron 2002;36:1021-34
110.
Autologous bone marrow stem cells in ischemic stroke. Bethesda, MD: Clinical trials.gov, 2007. Available from: http://clinicaltrials.gov/ct2/show/ NCT00535197? term=NCT00535197&rank=1 [Last accessed 15 January 2011]
111.
Cord blood for neonatal hypoxic-ischemic encephalopathy. Bethesda, MD: Clinical trials.gov, 2008. Available from: http://clinicaltrials. gov/ct2/show/NCT00593242? term=NCT00593242&rank=1 [Last accessed 15 January 2011]
112.
Autologous bone marrow mononuclear cell transplantation for stroke patients. Bethesda, MD: Clinical trials.gov, 2009. Available from: http://clinicaltrials. gov/ct2/show/NCT01028794? term=NCT01028794&rank=1 [Last accessed 15 January 2011]
113.
Autologous bone marrow stem cells in middle cerebral artery acute stroke treatment. Bethesda, MD: Clinical trials. gov, 2008. Available from: http:// clinicaltrials.gov/ct2/show/ NCT00761982?term= NCT00761982&rank=1 [Last accessed 15 January 2011]
Assanah M, Lochhead R, Ogden A, et al. Glial progenitors in adult white matter are driven to form malignant gliomas by platelet-derived growth factor-expressing retroviruses. J Neurosci 2006;26:6781-90
106.
Scharfman HE, Goodman JH, Sollas AL, Croll SD. Spontaneous limbic seizures after intrahippocampal infusion of brain-derived neurotrophic factor. Exp Neurol 2002;174:201-14
107.
Butt AM, Dinsdale J. Fibroblast growth factor 2 induces loss of adult oligodendrocytes and myelin in vivo. Exp Neurol 2005;192:125-33
108.
Safety study of autologous stem cell transplantation for patients with ischemic stroke. Bethesda, MD: Clinical trials.gov, 2007. Available from: http://clinicaltrials. gov/ct2/show/NCT00473057? term=NCT00473057&rank=1 [Last accessed 15 January 2011]
Jin K, Wang X, Xie L, et al. Evidence for stroke-induced neurogenesis in the human brain. Proc Natl Acad Sci USA 2006;103:13198-202
103.
105.
109.
Ehrenreich H, Weissenborn K, Prange H, et al. Recombinant human erythropoietin in the treatment of acute ischemic stroke. Stroke 2009;40:e647-56
114.
115.
Safety/feasibility of autologous mononuclear bone marrow cells in stroke patients. Bethesda, MD: Clinical trials. gov, 2008. Available from: http:// clinicaltrials.gov/ct2/show/ NCT00859014?term= NCT00859014&rank=1 [Last accessed 15 January 2011] Efficacy study of CD34 stem cell in chronic stroke patients. Bethesda, MD: Clinical trials.gov, 2009. Available from: http://clinicaltrials.gov/ct2/show/ NCT00950521?term= NCT00950521&rank=1 [Last accessed 15 January 2011]
Expert Opin. Biol. Ther. (2011) 11(4)
116. Intrathecal Stem Cells in brain injury (ISC). Bethesda, MD: Clinical trials.gov, 2009. Available from: http://clinicaltrials. gov/ct2/show/NCT01019733? term=NCT01019733&rank=1 [Last accessed 15 January 2011] 117. Stem cells therapy for patients with acute ischemic stroke. New Delhi, India: Clinical Trials Registry - India (CTRI), 2010. Available from: http://www.ctri. nic.in/Clinicaltrials/ViewTrial.jsp? trialno=121 [Last accessed 24 January 2011] 118. Ex vivo cultured adult allogenic MSCs in ischemic cerebral stroke. Bethesda, MD: Clinical trials.gov, 2010. Available from: http://clinicaltrials.gov/ct2/show/ NCT01091701?term= NCT01091701&rank=1 [Last accessed 15 January 2011] 119. Pilot Investigation of Stem Cells in Stroke (PISCES). Bethesda, MD: Clinical trials.gov, 2010. Available from: http://clinicaltrials.gov/ct2/show/ NCT01151124?term= NCT01151124&rank=1 [Last accessed 15 January 2011] 120. Intravenous Stem Cells After Ischemic Stroke (ISIS). Bethesda, MD: Clinical trials.gov, 2009. Available from: http:// clinicaltrials.gov/ct2/show/ NCT00875654?term= NCT00875654&rank=1 [Last accessed 15 January 2011] 121. Autologous cell therapy after stroke. Bethesda, MD: Clinical trials.gov, 2009. Available from: http://clinicaltrials. gov/ct2/show/NCT00908856? term=NCT00908856&rank=1 [Last accessed 15 January 2011]
Affiliation Terry C Burns MD PhD & Gary K Steinberg† MD PhD † Author for correspondence Stanford University School of Medicine, Department of Neurosurgery, 300 Pasteur Drive, R281, Stanford, CA 94305-5487, USA Tel: +1 650 725 5562; Fax: +1 650 723 2815; E-mail:
[email protected]
461
Review
Update on stem cell therapy for cerebral palsy James E Carroll† & Robert W Mays †
Medical College of Georgia, Neurology, Augusta, GA, USA
1.
Introduction
2.
Stem cell trials for
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
cerebral palsy 3.
Potential cell sources
4.
Experimental models
5.
Possible mechanisms of action
6.
Risks of treatment
7.
What’s needed next
8.
Conclusion
9.
Expert opinion
Introduction: Due to the publicity about stem cell transplantation for the treatment of cerebral palsy, many families seek information on treatment, and many travel overseas for cell transplantation. Even so, there is little scientific confirmation of benefit, and therefore existing knowledge in the field must be summarized. Areas covered: This paper addresses the clinical protocols examining the problem, types of stem cells available for transplant, experimental models used to test the benefit of the cells, possible mechanisms of action, potential complications of cell treatment and what is needed in the field to help accelerate cell-based therapies. Expert opinion: While stem cells may be beneficial in acute injuries of the CNS the biology of stem cells is not well enough understood in chronic injuries or disorders such as cerebral palsy. More work is required at the basic level of stem cell biology, in the development of animal models, and finally in well-conceived clinical trials. Keywords: animal models, cerebral palsy, embryonic stem cells, induced pluripotent stem cells, mesenchymal cells, multipotent adult progenitor cells, stem cells, transplantation Expert Opin. Biol. Ther. (2011) 11(4):463-471
1.
Introduction
Cerebral palsy is a heterogeneous group of conditions, defined as nonprogressive motor disability due to an abnormality of the cerebral hemispheres. While a small proportion of patients with cerebral palsy have as their cause a perinatal hypoxic-ischemic insult, most have acquired cerebral palsy due to the presence of one of a wide variety of other illnesses, such as developmental brain abnormalities, genetic conditions, traumatic or infectious disorders. Furthermore, insults may occur at different times during gestation, resulting in even more variation in pattern and causation. This heterogeneity in cause makes the assessment of any treatment fraught with considerable difficulty. Parents, on the other hand, focus on the condition of cerebral palsy and seek treatment based on that terminology. Patoine, in a recent editorial [1], described the pressures of a supposed ‘miracle cure’ supplied by stem cells influencing the behavior of parents of children with cerebral palsy. The United Cerebral Palsy Foundation states that there are 800,000 children and adults in the USA with cerebral palsy. The Centers for Disease Control estimates that about 10,000 babies are born each year with cerebral palsy. Improvements in the care of neonates have done little to alter the percentage of children with cerebral palsy. In fact, the increased survival of very low birth weight infants has contributed to sustaining the present occurrence rate [2]. Thus, the issue of stem cells as a potential treatment for cerebral palsy has assumed a disproportionately elevated position among parents of children with cerebral palsy. Seven years ago we presented in this journal the state of stem cell research in cerebral palsy [3]. While there has been definite progress in the scientific study of multiple types of stem cells, particularly the discovery of induced pluripotent stem cells (iPS cells), 10.1517/14712598.2011.557060 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
463
Update on stem cell therapy for cerebral palsy
Article highlights. . . . . . .
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
Treatment with stem cells is a serious consideration for cerebral palsy parents. Several clinical trials are in progress. There are numerous types of stem cells that could be used. While there are many animal models of brain injury, none are completely satisfactory for cerebral palsy. The potential mechanism of action of stem cells is potentially multifaceted. Risks of stem cell transplantation are real and probably understated. What is needed includes more knowledge of stem cell biology, a better chronic injury model and, later on, well-conceived clinical trials.
This box summarizes key points contained in the article.
relevant animal models for cerebral palsy are still lacking in critical factors. Consequently, progress with the initiation of cell based clinical trials for treatment of cerebral palsy has been limited. An additional problem is the timing of treatment. In order to be effective for most patients with cerebral palsy, the treatment will need to address an established or longstanding brain abnormality. But as we accumulate more information about the potential mechanisms of action of stem cells in brain injury, we are led to the conclusion that stem cells are much more likely to be effective in the acute situation rather than long into the course of a chronic disability. However, it is possible that stem cells could act favorably in a chronic injury by replacing nerve cells, with even a small replacement being significant, by making existing connections more effective, or by promoting blood vessel regeneration. The purpose of this article is to present the current state of stem cell transplantation for cerebral palsy patients. We review the current efforts with patients, the types of cells that might be used, the experimental basis for the treatment, animal models for cerebral palsy, the possible mechanisms for therapeutic success, the need for additional work, and the potential for harm. 2.
Stem cell trials for cerebral palsy
There are two ongoing US trials (Duke University and the Medical College of Georgia) listed in ClinicalTrials.gov [4] testing the safety and efficacy of autologous umbilical cord blood for cerebral palsy. These trials are obviously dependent upon the fact that some parents chose to preserve their child’s umbilical cord blood at the time of birth. The fact that the cells are autologous gives a significant safety margin to the trials, which otherwise might not have been allowed to proceed. Given that the parents have a strong commitment to stem cell therapy and enter the trials only because they know their children will receive the cells, both these trials 464
are double-blinded with a crossover treatment protocol. The crossover allows the children to receive their cells at some point in the study. The trials attempt to pare down the long list of causes for cerebral palsy by having extensive exclusion criteria, such as athetoid cerebral palsy, autism, hypsarrthymia, intractable epilepsy, progressive neurological disorder, HIV infection, extreme microcephaly, known genetic disorder, obstructive hydrocephalus, significant defect of brain development, chromosomal disorder, presence of major congenital anomaly or severe intrauterine growth retardation. One of the main justifications for these trials is the need to investigate the efficacy of this treatment in the face of ongoing clinical usage of the treatment. Currently there are no US trials for cerebral palsy dealing with allogeneic cell therapies. While hypoxic--ischemic injury is a clear cut and easily definable cause of cerebral palsy and possibly the most potentially open to treatment, this cohort of patients is in the minority. The current US trials attempt to focus on this group. Perhaps fewer than 100,000 of the 800,000 individuals with cerebral palsy have hypoxic--ischemic injury as their cause. A third trial listed in ClinicalTrails.gov [4] is being conducted by the Sung Kwang Medical Foundation in the Republic of Korea. This study is double-blinded, randomized with placebo control using allogeneic umbilical cord blood in combination with erythropoietin. The three arms of the study are: i) umbilical cord blood, erythropoietin, and rehabilitation, ii) erythropoietin and rehabilitation, and iii) rehabilitation only. This study employs immunosuppression in order to allow for the use of allogeneic cells. A fourth trial listed in ClinicalTrials.gov [4] is active but not recruiting (Hospital Universitario, Monterrey, Mexico). In this trial the patients are given G-CSF in order to stimulate their bone marrow to produce stem cells, bone marrow is harvested, and CD 34+ cells are purified and delivered via the intrathecal route. Outside the USA, there are a number of facilities that offer treatment with various types of stem cell preparations for cerebral palsy. These facilities are not conducting formal clinical trials. Stem cells offered from these companies or institutions are usually autologous adult stem cells prepared from the patient’s own tissue, usually bone marrow. The specific details of the preparation methods are generally not available. The cells are delivered either intravenously or into the cerebrospinal fluid. Often multiple administrations are recommended. 3.
Potential cell sources
There are many potential cell sources that have been used for experimental treatment protocols in animal models. The studies employ either direct implantation into brain parenchyma or, more commonly, intravenous injection. We recently reviewed the various cell sources [5].
Expert Opin. Biol. Ther. (2011) 11(4)
Carroll & Mays
Mesenchymal stem cells (MSCs) Mesenchymal stem cells (MSCs) are bone marrow stromal cells, comprised of a mixture of cell types, capable of supporting hematopoiesis along with the capability to differentiate into multiple cell types. While bone marrow is considered the primary source of MSCs, they are also found in human umbilical cord blood and to a lesser degree in other tissues. MSCs are generally isolated based on their preferential attachment to tissue culture plastic. The cells are fibroblast-like and possess the ability for self renewal. Most of the adult stem cells currently studied share some similarities with MSCs. In all pre-clinical cerebral palsy studies to date testing MSCs, the cells have been administered in the short term [6-9], with the longest period being one month after injury [10]. The benefit is noted both with intravenous and intracerebral transplantation. The mechanism of cell action is unknown, but does not appear to be neuronal cell replacement. However, the treatment appears to lead to sparing of intrinsic cells. In a primate model, Li et al. [11] reported that the cell transplantation resulted in upregulation of IL-10 expression. In association they found a decrease in neuronal apoptosis and astroglial activity in the periischemic area. The number of proliferating cells in the subventricular zone was also increased.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
3.1
CD34 cells CD34 cells are found in umbilical cord blood and bone marrow. They represent a small subset of MSCs. These cells are isolated based on the presence of a transmembrane glycoprotein as their surface characteristic. Clinical trials are underway in stroke patients [4]. 3.2
Umbilical cord blood Umbilical cord blood (UCB) is currently a popular source of adult stem cells being tested as a therapy for disease and injury. Numerous private and public banks have arisen in the USA and other parts of the world. The collection of umbilical cord blood is somewhat controversial in that various organizations, including the American Academy of Pediatrics [12], have questioned the utility of the collection and preservation in private banks. These concerns are based on the contention that there are few, if any, proven autologous therapies. To date, the main usage of these cells has been treatment of childhood diseases of the blood, although their experimental use for the treatment of cerebral palsy is currently under investigation. The minimum necessary dosage of cells for cell engraftment is usually considered to be 1 107 cells per kilogram. This includes the total nucleated cell fraction and not just stem cells. Thus, the child will ‘outgrow’ the available dose of autologous cells obtained at birth and available for transplant at a later date. Should autologous UCB be found efficacious for the treatment of acquired disorders, however, its usage would become wide spread. UCB has been used experimentally in brain injury models. Benefit of the treatment has been shown in a neonatal 3.3
hypoxic--ischemic rat model [13], adult rat stroke models [14-16], and a rat traumatic brain injury model [17]. On the other hand, Makinen et al. [18] did not find benefit with UCB in a rat stroke model. These were all acute studies. Multipotent adult progenitor cells Multipotent adult progenitor cells (MAPC) (Athersys) are derived from bone marrow as well as other tissue sources [19,20]. The phenotype consists of CD13+, fetal liver kinase 1 (Flk1)dim, c-kit-, CD44-, CD45-, MHC class I- and MHC class II-. These cells differentiate into mesenchymal cells, but also cells with visceral mesoderm, neuroectoderm and endoderm characteristics in vitro. They proliferate without senescence or loss of differentiation potential. We have used these cells in a rat model of neonatal hypoxic--ischemic injury, where cell administration results in improvement in behavioral outcome and neuronal sparing as determined by histology. We observed benefit in an acute model via both intracerebral and intravenous transplantation routes [21]. This was an important experiment in that we were able to show the efficacy of a safe and practical method of administration, that is intravenous. While some of the transplanted cells survived, and even displayed neuronal markers, the chief restorative feature was enhanced survival of endogenous neurons. We speculated this process was mediated by trophic factors, which would be most efficacious in the acute situation and perhaps less so in a chronic injury, as would be the case for cerebral palsy. Mays et al. [22] reported recent data from our group in a rat model of ischemic stroke. We demonstrated that immunosuppression was not required for allogeneic or xenogeneic cell mediated benefit. The studies noted that improvement with MAPC administration persisted at least as long as six months following acute treatment. Based on histological data, it was concluded that MAPC do not exert their benefit via cell replacement but more probably acted by trophic mechanisms. All of our work with MAPC is in acute studies, and once again we need to show improvement in a chronic injury model in order to supply pre-clinical evidence that would apply to cerebral palsy. 3.4
Induced pluripotent stem cells (IPS cells) Induced pluripotent stem cells (iPS cells) are now considered to be a substitute for embryonic stem cells [23]. The use of iPS cells has not yet been reported in any preclinical model of brain injury. It seems that the cells may be an ideal source for tissue repair, as they can be prepared from the patient’s own fibroblasts, eliminating considerations of rejection. However, there are a number of hurdles that will need to be cleared before this cell type would be available for clinical usage. First, the safety of the cells will need to be amply demonstrated in animal models. Do the cells form tumors? Are the viral agents used in the preparation of the cells a danger to the recipient? Are the cells effective in animal models? Robbins et al. [24] reviewed the use of these cells for transplantation and 3.5
Expert Opin. Biol. Ther. (2011) 11(4)
465
Update on stem cell therapy for cerebral palsy
concluded that reprogramming efficiency and safety considerations would need to be addressed before the initiation of clinical trials. Thus, while iPS cells seem quite promising, much work remains to be done at the basic translational science level before they can move into the clinic. Oligodendrocyte progenitor cells Oligodendrocyte progenitor cells (OPC) may be derived from fetal brain tissue [25], embryonic stem cells or iPS cells, the latter two via cell-differentiation protocols. Once again the problem in relation to the chronic nature of cerebral palsy is that the models of injury utilized in experimental animals are acute. OPC derived from human embryonic stem cells demonstrated some amelioration of function in rats undergoing traumatic spinal cord injury [26,27]. Keirstead et al. [28] used human embryonic-stem-cell-derived OPC in a rat model of spinal cord injury and compared the cells in an acute model versus a chronic model. Animals receiving the transplant seven days after the injury showed remyelination and improved motor ability compared with untreated animals; however the animals treated 10 months after the injury demonstrated no statistically significant improvement over control animals. This study underlines the potential difficulty of developing effective therapeutics in the chronic injury setting of the CNS. Tokumoto et al. [29] evaluated the ability of iPS cells derived from mouse embryonic fibroblasts to differentiate into oligodendrocytes and compared this with the differential ability of mouse embryonic stem cells (ESC). They found that intracellular factors inhibited the differentiation of iPS cells into mature oliogodendrocytes.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
3.6
Embryonic stem cells Embryonic stem cells (ESC) are certainly the most controversial type of stem cells. They are derived from embryos and generally require the destruction of that embryo. Consequently, there remain abiding ethical concerns about their use. In addition, the proliferative capacity of the cells and their potential for differentiation into many cell types makes the possibility of tumor formation quite real. Given that children receiving the cells would have many years in front of them, there would be ample time for tumor formation to occur. The animal models examined with ESCs are all in acute injuries. Zhang et al. [30] studied transplantation in a rat stroke model 24 h after the injury and found favorable postimplantation histological changes with survival of the transplanted cells, their migration and differentiation toward neural cell types. Liu et al. [31] reported that mesenchymal cells derived from ESCs lessened rat infarction volume, differentiated into neuronal and endothelial cells, and improved functional outcome when injected intravenously. Ma et al. [32] showed that embryonic-derived stem cells possessed the ability to migrate into the injury site and improve learning ability and memory fully eight months after the injury. Even though the benefit of the ESCs was long-lasting, the treatment was delivered in the acute phase after injury. 3.7
466
Fetal stem cells Finally, stem cells can be collected from fetal tissue. While the utility of these cells has not been widely explored in injury models, there are indeed indications of their potential. Aftab et al. [33] demonstrated that retinal progenitor cells from donor tissue of 16 -- 18 weeks gestational age were able to integrate into host retina and express rhodopsin. In other experiments cells from fetal brain transplanted acutely after hemorrhagic stroke displayed neuroprotecting anti-inflammatory capacity [34]. 3.8
4.
Experimental models
While cerebral palsy is caused by a number of conditions of which brain injury is a minor component, the models for cerebral palsy are generally based on some type of brain injury. The ideas for various therapies, therefore, are predicated on the notion that we can reverse the effects of the injury. Even though this may be the case for an acute injury, this theme does not apply to the many children with cerebral palsy whose condition arises from abnormalities of brain development. Our discussion in regard to the models of cerebral palsy is confined to the types of cerebral palsy arising from injury. Johnston et al. [35] have recently reviewed the available animal models and concluded that none are fully adequate. The Rice-Vannucci model [36] which combines unilateral carotid artery ligation with hypoxia in 7-day-old rat pups has been used for numerous studies on the cause and treatment of brain injury in the neonatal animal. These are studies of acute injury. The use of lipopolysaccaharide as a pretreatment to induce vulnerability to hypoxic--ischemic insult has added the important aspect of prenatal infection to the examination of the problem [37]. Girard et al. [38] showed that the combination of lipopolysaccharide exposure and hypoxic--ischemic injury in rats mimicked the motor deficits and neuropathological lesions seen in very premature infants. Their motor deficits were more persistent making this one of the more promising models for chronic injury. In view of the frequency of cerebral palsy occurring related to prematurity, the importance of white matter injury is an important consideration. Periventricular leukomalacia is the most frequent lesion in these patients. White matter lesions are not well-seen in rodent models, as the rodents have comparatively little white matter. In order to mimic the lesion seen in premature infants, several larger animal models have been developed which demonstrate white matter injury [35]. The perinatal rabbit model of cerebral palsy probably best fits the criterion of an injury producing motor disability. This model is produced by uterine ischemia [39-42] or by intrauterine administration of endotoxin [43]. However, these models do not appear to supply the chronic or long-lasting deficit we believe is required for satisfactory assessment.
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Carroll & Mays
Larger animal models, such as the sheep [44] or baboon [45], better reproduce the pathology seen in human infants. The pre-term baboon mimics the white matter neuropathology seen in premature human infants [45]. The expense of these methods, however, appears to be prohibitive for the number of animals required for an adequately powered study. One of the central problems in the development of stem cell therapies for cerebral palsy is still the lack of satisfactory experimental models. Ideally the model should include impairment of movement as a result of a brain injury. Secondly, the model should be one of chronic rather than acute injury. The more critical of these two factors actually is the need for a chronic or long-lasting injury. There have been numerous experimental treatments of acute injury models that have demonstrated success but none that have shown efficacy in a true, chronic model of injury. We and other investigators have shown that acute injuries are subject to repair by cell therapy, while the problem of chronic injury has been more resistant or neglected. The important feature that needs to be demonstrated is the capacity of the cell therapy to repair a chronic injury of any type. The type or location of the brain injury is comparatively less important than the need for a persistent, abnormal behavioral syndrome of some type in the animal. 5.
Possible mechanisms of action
One of the main ideas inherent in stem cell transplantation for cerebral palsy is that the stem cells would replace the cells of the damaged nervous system. Most reports dealing with adult stem cells show only a minimal survival of the transplanted cells with few, if any, of these cells displaying markers/functionality of nervous tissue [21,46,47]. It does not appear that replacement alone would be sufficient to account for improvement in the experimental situation. While embryonic or iPS cells may have somewhat greater potential for such replacement and transformation, the number of cells undergoing this process is quite limited in vivo. Even though there may be some replacement by transplanted cells, the cells often do not develop normal processes and may not function in neuronal circuitry [48]. Thus, cell replacement as an explanation for any improvement in the models is unlikely to be the case given the current state of our knowledge of the cell biology of stem cells. Another possibility is that the transplanted cells differentiate into astrocytes [48] or microglia. How this would assist in functional recovery is unclear. Bone-marrow-derived cells may participate in blood vessel regeneration by promoting adhesion of CXCR4-positive cells onto vascular endothelium [49], recruitment of endothelial progenitor cells [50], and in the formation of periendothelial vascular cells [51]. Borlongan et al. [52] have demonstrated that crude bone marrow may form endothelial cells in an animal model of stroke. A fourth set of ideas related to benefit is that the transplants induce a greater survival of intrinsic cells. We reported this
phenomenon in our neonatal hypoxic-ischemic model in animals treated with MAPC [21]. Mahmood et al. [53] used MSC injection to demonstrate that transplanted cells increased the expression of nerve growth factor and brain-derived neurotrophic factor after traumatic injury. This idea, for which the evidence seems strong, tends to restrict the benefit of stem cell transplantation to the acute post-injury period. Another possible mechanism of benefit is the effect of adult stem cells on splenic function during acute brain injury. In a stroke model Vendrame et al. showed that UCB lessened the splenic release of inflammatory cells and thereby protected the brain [54]. In support of this concept Walker et al. [55] demonstrated that the intravenous injection of MAPC after trauma blocked the normal splenic response to injury and improved outcome. These reports supported the idea that the spleen plays a role in adversely increasing the blood--brain barrier permeability and that the splenic response is blocked by adult stem cell therapy. Once again, this is a benefit only for the acute situation. 6.
Risks of treatment
The risks of stem cell therapy occur primarily with allogeneic transplants, which expose the recipient to graft-versus-host disease. Most reports of complications are in children undergoing hematopoietic stem cell transplantation for malignancies. These complications may relate in part to the fact that these children received radiation therapy, chemotherapy, or immunosuppressive medications in addition to the stem cell transplant. Herpes or cytomegalovirus infections may occur in these patients [56]. A variety of other medical complications are also reported in similar groups of patients [57,58]. Woodward et al. [59] reviewed 405 patients who received hematopoietic stem cell transplantation for a variety of disorders. Of these patients, 26 experienced some type of encephalopathy due to infection, organ failure, medication reaction, seizures, acute disseminated encephalomyelitis, thrombotic thrombocytopenic purpura or stroke. Herpes virus-6 encephalitis is also reported as a complication of unrelated umbilical cord transplant [60]. Clearly, we should not consider stem cell transplantation, particularly allogeneic, to be a benign procedure. Autologous transplantation may incur some of the same risks, particularly as the patients may be exposed to chemotherapy or infectious agents. The complications may relate significantly to the treatment accompanying transplantation or the site to which the transplant is delivered, such as into the cerebrospinal fluid. While adult stem cell transplants have been carried out in large numbers of cerebral palsy patients outside the US, there is no systematic reporting of complications. One would think that the route of administration, that is intravenous versus directly into the CNS, might be a key to the understanding of complications, but the reporting of routes and their complications are unavailable. Without question, the long-term complications are simply unknown.
Expert Opin. Biol. Ther. (2011) 11(4)
467
Update on stem cell therapy for cerebral palsy
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
7.
What’s needed next
We must have more knowledge of the biology and laboratory manipulation of the different types of stem cells. This area must include more work in the area of cell differentiation strategies. In addition we need to learn more about the effects of the various methods of stimulating intrinsic neural proliferation. A chronic, pre-clinical animal model is required for the study of the various competing cells types. The different cell types need to be compared in head-to-head competition. Controlled clinical trials are needed. These should be conducted with very specifically described patient groups, particularly more so than the current, on-going American trials. We must recognize that there are considerable differences among cerebral palsy patients, and therefore the patients need to be carefully matched for each study. This type of trial could only be achieved in a coordinated multiple-center paradigm. 8.
Conclusion
Current clinical trials in the use of stem cells for cerebral palsy are ongoing and incomplete. While there are a number of different cell types that are potential candidates as treatments, none have been shown to be effective in chronic animal models. Furthermore, available animal models do not adequately mimic cerebral palsy. Risks of the treatment are reported. More work on understanding the underlying beneficial biology of stem cells and the development and validation of more relevant animal models is required. 9.
Expert opinion
Stem cell therapy for cerebral palsy remains a frustrating area. Considering all the publicity about stem cells and
468
the fact that cell therapy is widely available outside the USA for a price, parents feel that surely the treatment must work. This view tends to be confirmed by preclinical reports of benefit in animal models of acute injury. Anecdotal reports of success, of which there are many, contribute little toward clarifying any benefit, but nevertheless encourage parents of cerebral palsy patients to seek the unproven therapy. There is no evidence as yet that stem cell therapy works in a chronic model of injury, as would be relevant to cerebral palsy. The problem remains difficult for several reasons: cerebral palsy is not a homogeneous disease, our knowledge of stem cell biology is in its infancy, the pre-clinical models are far from ideal, and various preclinical trials show efficacy in acute models leading to falsely raised hopes. We need a safe cell type that is effective in a chronic animal model of brain injury. Despite clinical use of stem cell treatment for cerebral palsy in many sites outside the USA, evidence of efficacy in a chronic animal model will be necessary before a clinical trial will be allowed in the USA using any type of allogeneic cell. We believe it would be inappropriate to conduct a clinical trial for cerebral palsy using allogeneic cells without safety and efficacy data in a chronic animal model. For the time being it may better to focus on the treatment of acute brain injuries with stem cells and thereby the improvement or prevention of cerebral palsy in this subset of patients.
Declaration of interest JE Carroll has received funding from Associazione Assistenza Figli Inabili Banca d’Italia, Cord Blood Registry, NINDS 5R42NS55606, RW Mays is an employee and stake holder at Athersys, Inc.
Expert Opin. Biol. Ther. (2011) 11(4)
Carroll & Mays
Bibliography
injury and upregulates interleukin-10. Expression in Macaca fascicularis. Brain Res 2010;1334:65-72
Papers of special note have beenhighlightes as either of interest () or of considerable interest () to readers. 1.
Patoine B. Media focus on ‘Miracle Cure’ for cerebral palsy pits science vs. hype. Ann Neurol 2009;66:A9-A11
2.
Cerebral palsy: hope through research. Bethesda, MD: National Institute of Neurological Disorders and Stroke, 2010. Available from: http://www.ninds. nih.gov/disorders/cerebral_palsy/ detail_cerebral_palsy.htm#154443104 [Last accessed 28 January 2011] A general summary of cerebral palsy.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
3.
Bartley J, Carroll JE. Stem cell therapy for cerebral palsy. Expert Opin Biol Ther 2003;3:541-9
4.
Bethesda, MD: clinicaltrials.gov. Available from: http://clinicaltrials.gov/
5.
Carroll JE, Borlongan C. Adult stem cell therapy for acute brain injury in children. CNS Neurol Disord Drug Targets 2008;7:361-9 A summary of the benefits of stem cells in acute injury.
.
6.
7.
8.
9.
10.
11.
Wang T, Tang W, Sun S, et al. Intravenous infusion of bone marrow mesenchymal stem cells improves brain function after resuscitation from cardiac arrest. Crit Care Med 2008;36(Suppl 11):S486-91 Kranz A, Wagner DC, Kamprad M, et al. Transplantation of placenta-derived mesenchymal stromal cells upon experimental stroke in rats. Brain Res 2010;1315:128-36 Zheng W, Honmou O, Miyata K, et al. Therapeutic benefits of human mesenchymal stem cells derived from bone marrow after global cerebral ischemia. Brain Res 2010;1310:8-16 Heile AM, Wallrapp C, Klinge PM, et al. Cerebral transplantation of encapsulated mesenchymal stem cells improves cellular pathology after experimental traumatic brain injury. Neurosci Lett 2009;463:176-81 Komatsu K, Honmou O, Suzuki J, et al. Therapeutic time window of mesenchymal stem cells derived from bone marrow after cerebral ischemia. Brain Res 2010;1334:84-92 Li J, Zhu H, Liu Y, et al. Human mesenchymal stem cell transplantation protects against cerebral ischemic
12.
.
13.
14.
15.
16.
17.
Section on Hematology and Section on Allergy/Immunology. Cord blood banking for potential future transplantation. Pediatrics 2007;119:165-70 Important recommendations from the Academy of Pediatrics. Pimental-Coelho PM, Magalhaes ES, Lopes LM, et al. Human cord blood transplantation in a neonatal rat model of hypoxic-ischemic brain damage: functional outcome related to neuroprotection in the striatum. Stem Cells Dev 2010;19:351-8 Vendrame M, Cassady J, Newcomb J, et al. Infusion of human umbilical cord blood cells in a rat model of stroke dose-dependently rescues behavioral deficits and reduces infarct volume. Stroke 2004;35:2390-5
22.
..
23.
..
24.
Mays RW, Borlongan CV, Yasuhara T, et al. Development of an allogeneic adherent stem cell therapy for treatment of ischemic stroke. J Exp Stroke Transl Med 2010;3:34-46 An extensive paper dealing with the development of MAPC therapy. Kazutoshi T, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 2006;126:663-76 An important basic paper. Robbins RD, Prasain N, Maier BF, et al. Inducible pluropotent stem cells: not quite ready for prime time? Curr Opin Organ Transplant 2010;15:61-7 A good summary on IPS cells.
Willing AE, Lixian J, Milliken M, et al. Intravenous versus intrastriatal cord blood administration in a rodent model of stroke. J Neurosci Res 2003;73:296-307
25.
Chen J, Sanberg PR, Li Y, et al. Intravenous administration of human umbilical cord blood reduces behavioral deficits after stroke in rats. Stroke 2001;32:2682-8
Wu C, Chang A, Smith MC, et al. beta4 tubulin identifies a primitive cell source for oligodendrocytes in the mammalian brain. J Neurosci 2009;29:7649-57
26.
Kerr CL, Letzen BS, Hill CM, et al. Efficient differentiation of human embryonic stem cells into oligodendrocyte progenitors for application in rat contusion model of spinal cord injury. Int J Neurosci 2010;120:305-13
27.
Sharp J, Frame J, Siegenthaler M, et al. Human embryonic stem cell derived oligodendrocyte progenitor cell transplants improve recovery after cervical spinal cord injury. Stem Cells 2010;28:152-63
28.
Keirstead HS, Nistor G, Bernal G, et al. Human embryonic stem cell-derived oligodendrocyte progenitor cell transplants remyelinate and restore locomotion after spinal cord injury. J Neurosci 2005;25:4694-705
29.
Tokumoto Y, Ogawa S, Nagamune T, et al. Comparison of efficiency of terminal differentiation of oligodendrocytes from induced pluripotent stem cells versus embryonic stem cells in vitro. J Biosci Bioeng 2010;109:622-8
Lu D, Sanberg PR, Mahmood A, et al. Intravenous administration of human umbilical cord blood reduces neurological deficit in the ratr after traumatic brain injury. Cell Transplant 2002;11:275-81
18.
Makinen S, Kekarainen T, Nystedt J, et al. Human umbilical cord blood cell do not improve middle cerebral artery occlusion in rats. Brain Res 2006;1123:207-15
19.
Jiang Y, Jahagirdar B, Reinhardt R, et al. Pluripotency of mesenchymal stem cells derived from adult marrow. Nature 2002;418:41-9
20.
Jiang T, Vaessen B, Lenvik T, et al. Multipotent progenitor cells can be isolated from postnatal murine bone marrow, muscle, and brain. Exp Hematol 2002;30:896-904
21.
.
intracerebrally delivered multipotent adult progenitor cells in neonatal hypoxic-ischemic rats. J Cereb Blood Flow Metab 2008;28:1804-10 This research demonstrates the equivalent benefit of intravenous administration.
Yasuhara T, Hara K, Maki M, et al. Intravenous grafts recapitulate the neurorestoration afforded by
Expert Opin. Biol. Ther. (2011) 11(4)
.
469
Update on stem cell therapy for cerebral palsy
30.
31.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
32.
33.
Zhang P, Li J, Liu Y, et al. Transplanted human embryonic neural stem cells survive, migrate, differentiate and increase endogenous nestin expression in adult rat cortical pre-infarction zone. Neuropathology 2009;29:410-21 Liu YP, Seckin H, Izci Y, et al. Neuroprotective effects of mesenchymal stem cells derived from human embryonic stem cells in transient focal cerebral ischemia in rats. J Cereb Blood Flow Metab 2009;29:780-91 Ma J, Wang Y, Yang J, et al. Treatment of hypoxic-ischemic encephalopathy in mouse by transplantation of embryonic stem cell derived cells. Neurochem Int 2007;51:57-65 Aftab U, Jiang C, Tucker B, et al. Growth kinetics and transplantation of human retinal progenitor cells. Exp Eye Res 2009;89:301-10
34.
Lee ST, Chu K, Jung KH, et al. Anti-inflammatory mechanism of travascular neural stme cell transplantation in haemorrhagic stroke. Brain 2008;131:616-29
35.
Johnston MV, Ferriero DM, Vannucci SJ, et al. Models of cerebral palsy: which ones are best? J Child Neurol 2005;20:984-7 An excellent review of animal models for cerebral palsy.
..
36.
.
37.
38.
39.
470
Rice JE, Vanucci RC, Brierley JB. The influence of immaturity on hypoxic-ischemic brain damage in the rat. Ann Neurol 1981;9:131-41 A basic model for hypoxic-ischemic injury in neonatal rats. Eklind S, Mallard C, Arvidsson P, et al. Lipopolysaccharide induces both a primary and a secondary phase of sensitization in the developing rat brain. Pediatr Res 2005;58:112-16 Girard S, Kadhim H, Beaudet N, et al. Developmental motor deficits induced by combined fetal exposure to lipopolysaccharide and early neonatal hypoxia/hypoxia ischemia: a novel animal model for cerebral palsy in very premature infants. Neuroscience 2009;158:673-82 Derrick M, Drobyshevsky A, Ji X, et al. Hypoxia-ischemia causes persistent movement deficits in a
50.
Derrick M, Drobyshevsky A, Ji X, et al. A model of cerebral palsy from fetal hypoxia-ischemia. Stroke 2007;38:731-5
Yamaguchi J, Kusano KF, Masuo O, et al. Stromal cell-derived factor-1 effects on ex vivo expanded endothelial progenitor cell recruitment for ischemic neovascularization. Circulation 2003;107:1322-8
51.
41.
Drobyshevsky A, Derrick M, Wyrwicz AM, et al. White matter injury correlates with hypertonia in an animal model of cerebral palsy. J Cereb Blood Flow Metab 2007;27:270-81
Rajantie I, Llmonen M, Alminaite A, et al. Adult bone marrow-derived cells recruited during angiogenesis comprise precursors for periendothelial vascular mural cells. Blood 2004;104:2084-6
52.
42.
Tan S, Drobyshevsky A, Jilling T, et al. Model of cerebral palsy in the perinatal rabbit. J Child Neurol 2005;20:972-9
43.
Saadani-Makki F, Kannan S, Lu X, et al. Intrauterine administration of endotoxin leads to motor deficits in a rabbit model: a link between prenatal infection and cerebral palsy. Am J Obstet Gynecol 2008;199:651, e1-e7
Borlongan CV, Lind DG, Dillon-Carter O, et al. Bone marrow grafts restore cerebral blood flow and blood brain barrier in stroke rats. Brain Res 2004;1010:108-16
53.
Mahmood A, Lu D, Chopp M. Intravenous administration of marrow stromal cells (MSCs) increasers the expression of growth factors in rat brain after traumatic brain injury. J Neurotrauma 2004;21:33-9
54.
Vendrame M, Gemma C, Pennypacker KR, et al. Cord blood rescues stroke-induced changes in splenocyte phenotype and function. Exp Neurol 2006;199:191-200
55.
Walker PA, Shinil KS, Jimenez F, et al. Intravenous multipotent adult progenitor cell therapy for traumatic brain injury: preserving the blood brain barrier via an interaction with splenocytes. Exp Neurol 2010;225:341-52
56.
Rubin J, Wide K, Rermberger M, et al. Acute neurological complications after hematopoietic stem cell transplantation in children. Pediatr Transplant 2005;9:62-7
57.
Uckan D, Cetin M, Yigitkanli I, et al. Life-threatening neurological complications after bone marrow transplantation in children. Bone Marrow Transplantation 2005;35:1-76
58.
Iguchi A, Kobayashi R, Yoshida M, et al. Neurological complications after stem cell transplantation in childhood. Bone Marrow Transplant 1999;24:647-52
59.
Woodward P, Helton K, McDaniel H, et al. Encephalopathy in pediatric patients after allogeneic hematopoietic stem cell transplantation is associated with a poor prognosis.
perinatal rabbit model of cerebral palsy: assessed by a new swim test. Int J Dev Neurosci 2009;27:549-57 40.
44.
Lotgering FK, Bishai JM, Struijk PC, et al. Ten-minute umbilical cord occlusion markedly reduces cerebral blood flow and heat production in fetal sheep. Am J Obstet Gynecol 2003;189:233-8
45.
Inder T, Neil J, Yoder B, et al. Patterns of cerebral injury in a primate model of preterm birth and neonatal intensive care. J Child Neurol 2005;20:965-7
46.
Reiss P, Zhang C, Saatman K, et al. Transplanted neural stem cells survive, differentiate, and improve neurological motor function after experimental traumatic brain injury. Neurosurgery 2002;51:1043-52
47.
48.
49.
Zhao LR, Duan WM, Reyes M, et al. Human bone marrow stem cells exhibit neural phenotypes and ameliorate neurological deficits after grafting into the ischemic brain of rats. Exp Neurol 2002;174:11-20 Kopen GC, Prockop DG, Phinney DG. Marrow stromal cells migrate throughout forebrain and cerebellum, and they differentiate into astrocytes after injection into neonatal mouse brains. Proc Natl Acad Sci USA 1999;96:10711-16 Peled A, Kollet O, Ponomayov T, et al. The chemokine SDF-1 activates the integrins LFA-1. VLA-4, and VLA-5 on immature human CD34+ cells: role in transendothelial/stromal migration and engraftment of NOD/SCID mice. Blood 2000;95:3289-96
Expert Opin. Biol. Ther. (2011) 11(4)
Carroll & Mays
Bone Marrow Transplant 2004;33:1151-7 60.
Chik KW, Chan PK, Li CK, et al. Human herpesvirus-6 encephalitis after unrelated umbilical cord blood transplant in children. Bone Marrow Transplant 2002;99:991-4
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Affiliation
James E Carroll†1 & Robert W Mays2 † Author for correspondence 1 Medical College of Georgia, Neurology, BG2000H, 1446 Harper Street, Augusta, GA 30912 USA E-mail:
[email protected] 2 Athersys, Inc., Regenerative Medicine, 3201 Carnegie Avenue, Cleveland, 44115-2634 USA
Expert Opin. Biol. Ther. (2011) 11(4)
471
Review
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
1.
Introduction
2.
Historical perspective
3.
The biology of GvT and GvHD and potential targets for DLI
4.
The role of Tregs
5.
The role of MRD in guiding
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
administration of DLI 6.
Efficacy of DLI in specific disease settings
7.
DLI in the pediatric setting
8.
General principles of DLI: effective cell dose, timing, toxicity and donor issues
9. 10.
Strategies to avoid DLI-associated toxicity Future therapeutic options and research imperatives in the field of DLI
11.
Conclusions
12.
Expert opinion
Claire Roddie & Karl S Peggs† †
UCL Cancer Institute, Department of Haematology, Paul O’Gorman Building, 72 Huntley Street, London, WC1E 6BT, UK
Introduction: Allogeneic hematopoietic stem cell transplantation (SCT) is the treatment of choice for many malignant hematological disorders. Following recent improvements in non-relapse-related mortality rates, relapse has become the commonest cause of treatment failure. Infusion of donor lymphocytes can potentially enhance immune-mediated antitumor activity and offers a salvage option for some patients. This paper reviews the current literature on the efficacy of this therapeutic strategy. Areas covered: The biology of adoptive cellular therapy with allogeneic immune cells to treat relapse across a spectrum of diseases in both the full intensity and reduced intensity hematopoietic SCT settings is explored. The review discusses the current limitations of the approach and reviews several new experimental strategies which aim to segregate the desired graft-versus-tumor effect from the deleterious effects of more widespread graft-versus-host reactivity. Expert opinion: Durable responses to DLI have been noted in chronic myeloid leukemia and responses have also been described in acute leukemia, multiple myeloma and chronic lymphoproliferative disorders. The new challenge in transplantation is to optimize DLI therapy in order to further improve patient outcomes. Keywords: allogeneic stem cell transplantation, DLI, graft-versus-host disease, relapse Expert Opin. Biol. Ther. (2011) 11(4):473-487
1.
Introduction
The efficacy of allogeneic stem cell transplantation (SCT) as a curative option for hematological malignancy is influenced by three factors: the underlying disease, the pre-transplant conditioning regimen and the graft-versus-tumor (GvT) effect mediated by donor leucocytes within the graft. The last two factors must be balanced against transplant related mortality (TRM). For example, despite delivering a reduction in relapse rates, further intensification of existing myeloablative (MA) conditioning chemo- and radio-therapy beyond current levels increases TRM and morbidity without improving overall survival (OS) [1,2]. Efforts to minimize treatment-related morbidity and mortality have focused on modulating conditioning protocols and improving supportive care. The introduction of reduced intensity conditioning (RIC) and non-MA transplants (the so-called ‘reduced toxicity’ transplants), which deliver lower TRM than conventional MA regimens, has revolutionized clinical practice, permitting allogeneic SCT in a population of previously ineligible patients [3,4]. The underlying principle of the RIC transplant is to provide sufficient immunosuppression to facilitate engraftment without the highly toxic, inflammatory ‘cytokine storm’ induced by conventional MA conditioning. These 10.1517/14712598.2011.554811 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
473
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
2.
Article highlights. .
.
.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
Response to donor lymphocyte infusion (DLI) is disease-dependent, with the best evidence in chronic phase chronic myeloid leukemia. Encouraging results have been obtained in indolent lymphoproliferative disorders, mantle cell lymphoma and Hodgkin’s lymphoma, but less so in acute lymphocytic leukemia, more aggressive non-Hodgkin’s lymphoma and multiple myeloma, which may benefit more from non-cellular therapies in the setting of relapse. Graft-versus-host disease (GvHD) and to a lesser degree marrow aplasia are the major side effects of DLI therapy. The escalating dose regimen administration schedule is one approach to minimizing GvHD. Promising advances in graft engineering (e.g. selection of T-cell subsets and selection of T cells directed against recipient minor histocompatability antigens) and gene therapy (e.g. TCR and chimeric antigen receptor constructs) may also help to tip the balance of immunity towards graft-versus-tumor effect and away from GvHD and improve clinical outcomes.
This box summarizes key points contained in the article.
transplants permit tumor eradication through facilitating the GvT effect more slowly post-transplant rather than directly through cytoreductive conditioning and for this reason may be more suited to the management of indolent hematologic diseases [5-7]. Due in part to the lower intensity conditioning of these procedures, relapse has become the leading cause of treatment failure [8-10]. The high uptake of RIC transplantation has resulted in the prevention and management of relapse becoming an increasingly prominent feature of clinical practice. One key approach has been the donor lymphocyte infusion (DLI) to enhance GvT responses [11]. One of the major problems faced by clinicians is the lack of explicit published data on the efficacy of DLI. The reasons for this are manifold. First, there is a reluctance to conduct clinical trials in this population due to the small patient numbers and often poor outcomes despite intervention. Second, studies often incorporate heterogeneous patient groups from which it is difficult to identify which factors are relevant to each disease entity. There is now international agreement on the need for collaborative multi-center studies and the need for a central database or sample repository to assess interventions in this area. An initiative was recently outlined in the National Cancer Institute First International Workshop on the Biology, Prevention and Treatment of Relapse after Allogeneic Haematopoietic Stem Cell Transplantation (2010) [10,12]. We provide a historical context for the use of DLI, discuss the biology underlying the GvT effect, review the available evidence on the utility of DLI across a range of hematological diseases and address the future of DLI along with developments in graft engineering. For the purposes of this review, we concentrate on T-cell rather than NK-cell therapies. 474
Historical perspective
Bidirectional immune reactivity (alloreactivity) between host and recipient T cells underlies many of the major complications following allogeneic SCT, namely graft failure and graft-versus-host disease (GvHD), but it is also responsible for the advantageous GvT effect [13]. Early evidence for GvT was based on clinical observation: complete remissions were observed in some patients with relapsed disease post-allograft in whom immunosuppression was withdrawn whilst in others GvHD appeared to protect against relapse [14,15]. Higher rates of relapse were observed in patients receiving syngeneic (i.e., from an identical twin donor) [16] or T-cell depleted transplants [17,18] and it was hypothesized that allogeneic T lymphocytes could be the active cell in the observed GvT effect. Preclinical studies revealed how specific donor T cells prevented growth of leukemia colonies in vitro and prevented development of acute myeloid leukemia in an in vivo immune-deficient mouse model of leukemia. Donor T cells active against leukemia cells were subsequently demonstrated to target recipient hematopoiesis-restricted minor histocompatability antigens (mHags) [19] and aberrantly or overexpressed ligands such as Proteinase 3 [20-22] in myeloid malignancies. These findings supported the concept of DLI as a therapeutic intervention, where isolation of donor T lymphocytes and their subsequent infusion to the recipient could hasten or intensify the GvT effect in the relapsed patient. In 1990, Kolb et al. published the first clinical study of DLI in patients with relapsed chronic myeloid leukemia (CML), which showed that infused donor buffy-coats in association with IFN-a could induce cytogenetic remissions [23]. Remarkably, DLI has since been shown to restore durable complete responses (CR) in up to 80% of these patients [24-33]. This success in CML prompted others to investigate the use of DLI across a range of hematological malignancies such as acute myeloid leukemia (AML), acute lymphocytic leukemia (ALL), non-Hodgkin’s lymphoma (NHL), multiple myeloma (MM) and Hodgkin’s lymphoma (HL) with variable success [34-38]. The role of DLI in these disorders remains poorly defined and response rates, optimal approaches and long-term outcomes are still unclear (refer to Table 1 for details of several of the larger trials of DLI therapy in a range of hematological diseases).
The biology of GvT and GvHD and potential targets for DLI
3.
The efficacy of allogeneic SCT derives largely from the allorecognition which permits donor cell engraftment and facilitates the GvT response. GvHD is an undesirable side effect of therapy and is thought to be initiated by tissue injury leading to activation and proliferation of alloreactive T cells, which then home to sites of inflammation and potentiate
Expert Opin. Biol. Ther. (2011) 11(4)
Roddie & Peggs
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Table 1. Efficacy and toxicity of DLIs in hematological malignancy (selected series). Disease/study author
Patient number
Cell dose ( 108/kg)
CR post DLI (%)
GvHD (acute > grade II/ chronic, extensive) (%)
CML/Kolb et al. [23] CML/van Rhee et al. [30] CML/Collins et al. [33] CML/Porter et al. [75] AML/Kolb et al. [32] AML/Shiobara et al. [35] AML/Schmid et al. [72] AML/Porter et al. [75] ALL/Kolb et al. [32] ALL/Collins et al. [33] ALL/Shiobara et al. [35] NHL/Russell et al. [76] NHL/Bloor et al. [78] NHL/Bishop et al. [80] HL/Peggs et al. [85] MM/Collins et al. [33] MM/van de Donk et al. [95] MM/Lokhorst et al. [96]
3 14 56 25 23 32 171 23 22 15 30 17 17 5 14 5 63 13
4.40 -- 7.40 0.70 -- 5.30 0.50 -- 3.62 0.005 -- 5.21z 0.10 -- 7.83z 0.01 -- 7.40z 0.001 -- 2.250 0.001 -- 31.8 0.30 -- 11 0.50 -- 6.20z 0.01 -- 11.3z 0.05 -- 1 0.01 -- 1 0.1 -- 1 0.01 -- 1 1.63 -- 5.53z 0.01 -- 3 0.01 -- 3.30
100 57 100 (Cy)/73 (H)/33 (A) 46 29 25 35 35 0 18 20 10 76 50 57 50 12 31
66.6 28a/28c 46a/21c* 28a/12c 41* 34a/33c* 43a/46c 35a/17c 41* 46a/32c* 34a/33c* 44a/89c 15a/31c 50 46a/32c* 22a/43c 62a/15c
*The studies marked encompass different disease entities and the incidence of GvHD reported relates to the whole study cohort rather than to individual diseases. z Indicates the mononuclear cell dose rather than the T-cell dose. a: Acute GvHD; A: Accelerated phase/blast crisis; ALL: Acute lymphocytic leukemia; AML: Acute myeloid leukemia; c: Chronic GvHD; CML: Chronic myeloid leukemia; CR: Complete response; Cy: Cytogenteic relapse; DLI: Donor lymphocyte infusion; GvHD: Graft-versus-host disease; H: Hematologic relapse; HL: Hodgkin’s lymphoma; MM: Multiple myeloma; NHL: Non-Hodgkin’s lymphoma.
tissue injury further. A major focus of transplant biology is to devise strategies to dissociate GvT from GvHD, with focus on both the antigen presenting cells (APCs) and the effector T cells. The mechanism(s) by which allorecognition occurs during the GvT response is not fully understood. Murine models of antigen presentation have shown that recipient APCs are crucial to initiation of GvT, whereas donor APCs appear not to be so critical [39,40]. Dendritic cells (DCs) are professional APCs which present antigen via both Class I and II MHC pathways to the adaptive immune system. Research is ongoing into methods of loading DCs with tumor antigen to enhance GvT activity and to try to develop methods to selectively eliminate DCs responsible for GvHD whilst promoting the expansion of those involved in GvT responses [10]. The GvT response is dependent on the ability of the graft to either induce immunity against tumor-specific neoantigens (probably a less common event) or to break tolerance and induce antigen-specific autoimmunity to overly- or aberrantly-expressed tumor-associated antigens such as Wilm’s tumor 1, Proteinase-3 and several mHags (HA-1, HA-2, HB-1 and BCL2A1) which are differentially expressed on hematopoietic cells [41-43]. Dissociation of GvT from GvHD might be achieved through the adoptive transfer of T cells directed against these antigens. Alternative approaches to dissociate GvT from GvHD have focused on potential differences in effector pathways. GvT and GvHD related tissue damage is thought to occur via different mechanisms: GvHD effectors generally utilize the
Fas:Fas-ligand cytolytic pathway, whilst GvT effectors use perforin-mediated cytotoxicity and possibly TNF-related apoptosis-inducing ligand with selective activity for malignant targets. Manipulation of these different mechanisms of cytotoxicity may facilitate enhancement of GvT and at the same time suppress GvHD [44]. 4.
The role of Tregs
Maintenance of transplant tolerance is dependent on the suppression of alloreactive donor T-cell clones by means of central deletion, clonal anergy and the inhibitory effects of regulatory T cells (Tregs). Tregs are naturally occurring clusters of differentiation (CD)4+ CD25+ forkhead box P3+ T cells that constitute ~ 1 -- 2% of the circulating CD4+ T-cell population. In the autologous setting, they help to prevent autoimmunity by dominantly suppressing the activity of autoreactive lymphocytes using a variety of mechanisms including the secretion of inhibitory cytokines such as IL-10 and TGF-b, and direct cell--cell contact inhibition [45,46]. In murine models, Tregs have been shown to prevent GvHD when co-infused with effector T cells, albeit with the potential to also suppress GvT [47,48]. It is, therefore, possible that infusions of ex vivo expanded Tregs given to patients with GvHD could ameliorate their symptoms and that depletion of Tregs from the stem cell (or DLI) product could enhance alloreactivity by ‘releasing the brake’ on the GvT effect.
Expert Opin. Biol. Ther. (2011) 11(4)
475
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
5. The role of MRD in guiding administration of DLI
The detection of minimal residual disease (MRD) may predict the likelihood of overt disease relapse in some diseases [49-52]. Monitoring for mixed chimerism (MC), that is, the return of recipient-derived hematopoietic cells is one potential approach. In classical MA conditioning, host hematopoiesis is replaced entirely by donor cells to create full donor chimerism (FDC). In this setting, the return of host cells (MC) is generally indicative of relapse with few exceptions. The potential importance of early intervention in terms of maximizing therapeutic efficacy is well established in CML and increasingly recognized in other disorders. For example, it was demonstrated in a retrospective analysis of patients with acute leukemia and myelodysplastic syndrome (MDS) who received DLI for relapse based on leukemia lineagespecific chimerism analysis following transplantation. The 3-year survival in those treated for molecular relapse was 42% compared to 16% in those treated for hematologic relapse, further suggesting that early intervention with DLI during molecular relapse rather than waiting for hematologic relapse has the potential to improve responses and survival [53]. In the RIC transplant setting, early MC is more common (particularly with T-cell depletion) with more gradual conversion to FDC over many months. Whether persistent MC, particularly when it is present only within the T cell lineage, is associated with a higher incidence of relapse and can thus be used as a basis for early intervention with DLI to achieve FDC and prevent relapse, remains controversial [54]. It is likely that the significance of MC differs according to whether T-cell depletion is incorporated as part of the conditioning regimen and remains stable or is increasing over time. Other methods of MRD detection, such as multiparametric flow cytometry (MFC) and PCR amplification of fusion transcripts and antigen receptor genes are used across a range of diseases. In many cases, they offer a much greater sensitivity than routine chimerism assays. However, except in the case of CML, these strategies have not yet been fully validated. Further detail is beyond the scope of this review, but the reader is referred to several studies of interest in specific disease settings [55-62]. 6.
Efficacy of DLI in specific disease settings
CML To date, most experience with DLI has been gained in CML. Post-transplant relapse rates are markedly higher for patients with advanced CML (accelerated phase (AP) or blast crisis (BC) compared with CML in first chronic phase (CMLCP)). Responses to DLI are often durable and are best in those with molecular relapse (90 -- 100%) followed by cytogenetic relapse (90%), hematological relapse in 6.1
476
CP (75%), relapse in AP/BC (36%) and worst in resistant disease (0%) [63]. Relapse following DLI represents a varied spectrum. The outlook for patients with extramedullary relapse is often poor and optimal treatments are yet to be defined [64]. Alternatively, patients who achieve hematological remission but remain molecularly or cytogenetically positive for bcr-abl (oncogenic fusion protein) can be successfully re-treated with DLI [65]. Disease persistence may be due to CML stem cells which do not express the maturation-associated antigens targeted by CD8+ mHag-specific cytotoxic T lymphocytes (CTLs) [66]. Work is ongoing to identify and eradicate CML stem cells. The role of additional agents in combination with DLI is unclear. There is evidence to suggest that IFN-a may potentiate the therapeutic efficacy of DLI such that lower total cell doses are required to achieve remission or that patients resistant to conventional DLI achieve responses [67,68]. Studies of cytoreductive chemotherapy plus DLI in patients with advanced CML have also been conducted, but outcomes have been disappointing [32]. The combination of DLI with a tyrosine kinase inhibitor (TKI) has been explored, although results have been conflicting. One study suggested more rapid remissions and improved OS and disease-free survival, particularly in the context of accelerated disease [69]. Other groups have obtained compelling preclinical data to suggest that the antiproliferative effect of TKI therapy affects both residual leukemia cells and tumor-responsive CTLs [70,71]. TKI therapy may, therefore, adversely affect the potentially curative immune effects of allogeneic transplantation and DLI. This would be an ideal area for a randomized controlled study. AML The probability of post-transplant relapse in AML has been variably reported as 20 -- 60% and the prognosis is generally poor. As a single agent, DLI is generally unable to induce remission in florid relapse. One study of DLI in combination with induction chemotherapy has revealed poor remission rates (15 -- 20%) and a low 2-year OS (15 -- 20%). Subgroup analysis revealed that chemo-responsive patients with favorable cytogenetics and low bulk disease at relapse did significantly better, with an OS of 56% at 2 years [72]. Combination regimens incorporating newer agents such as decitabine and azacitadine into DLI protocols are the subject of current clinical interest, but data are limited. Bearing in mind the poor outcomes in relapsed AML with current salvage modalities, a clinical study of prophylactic DLI in a cohort of AML patients was conducted and showed improved OS compared with case-matched controls [73]. Intervention based on MRD monitoring may reduce the exposure of patients to the risk of GvHD inherent in prophylactic strategies as previously discussed [53], and the results of combination of DLI with novel hypomethylating agents in this setting are eagerly awaited. 6.2
Expert Opin. Biol. Ther. (2011) 11(4)
Roddie & Peggs
These data require further maturation and validation, but are particularly relevant to a growing population of patients with AML or high risk MDS undergoing RIC transplantation. ALL Relapsed ALL has a poor prognosis, with only 7% of adult patients surviving 5 years [74]. DLI is rarely effective in florid relapse and one small study reports a CR rate of 10 -- 20% of limited duration in patients receiving matched sibling DLI [75]. The current research emphasis in ALL is on the prevention of relapse through optimization of up-front therapy and the development of new targeted therapies. Post-transplant cellular therapies have not yet realized their potential.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
6.3
NHL NHL comprises a heterogeneous group of histological diagnoses which can be divided clinically into indolent and aggressive groups. The curative potential of MA allogeneic transplantation is established in NHL, but is precluded in the majority of patients due to an unacceptably high TRM. The lower toxicity RIC strategies have facilitated the application of allogeneic transplantation and DLI in this group of patients. A significant GvT effect has been reported in indolent NHL following stem cell and DLI therapy, with less evidence supporting a strong effect in more aggressive histologies [76-80]. In our recent study of T cell depleted RIC transplantation in multiple relapsed follicular lymphoma, 13/82 patients relapsed, 10/13 received DLI and 9/10 experienced sustained remission. Overall, the incidence of GvHD was low, affecting < 20% of patients [77]. An overlapping study of DLI in ‘indolent NHL’ comprising 28 patients with either MC (n = 11) or relapsed disease ± MC (n = 17) demonstrated cumulative response rates of 91% in the MC cohort and 76% in the relapsed disease cohort, complicated by GvHD in 15 -- 31% [78]. Aggressive NHL is thought to be relatively poorly responsive to allogeneic SCT and DLI [76], but a recent study of 48 patients with relapsed diffuse large B-cell lymphoma undergoing RIC transplantation demonstrated progressionfree survival (PFS) and OS rates at 4 years of 48 and 47%, which improved further to 55 and 54% in those with chemo-sensitive disease before transplant. Overall, 12/48 patients received DLIs ± chemoimmunotherapy for relapse and 3/12 obtained durable remissions although the role of other therapies administered in close temporal approximation is difficult to evaluate [79,80]. Mantle cell lymphoma (MCL) is an aggressive NHL which generally responds poorly to treatment. In a study of RIC allogeneic transplantation in 70 patients with MCL, 15 relapsed post-transplant and 11/15 achieved CR with DLI [81]. This demonstrates that DLI is an effective salvage therapy in MCL, confirming the importance of GvT inferred 6.4
from the encouraging PFS survival rates delivered in T-cell replete RIC programs. Rituximab, a chimeric anti-CD20 mAb, is purported to augment the GvT effects of DLI by promoting antigen priming. Preclinical studies of rituximab-treated tumor cell lines demonstrated more effective alloantigen presentation [82]. Clinical experience remains largely anecdotal at present, though encouraging responses have been claimed and this is another area that would benefit greatly from a consolidated clinical study. HL Historically, poor risk HL patients rarely underwent allogeneic transplantation due to prohibitively high TRM [83]. RIC conditioning protocols have effectively reduced TRM and post-transplant survival outcomes have improved, but this has been complicated by high relapse rates (44 -- 81% at 2 -- 3 years) [84-86]. Published experience with DLI is relatively scarce. In many cases, response rates have been disappointing (around 30 -- 40%), and perhaps more significantly of limited duration. The experience following T-cell depleted transplantation seems qualitatively different. Our singleinstitution experience of 24 patients treated with DLI either alone (n = 14) or combined with debulking or consolidating chemo-radiotherapy (n = 10) demonstrated CR in 14 and partial responses (PR) in five patients (overall response 79%) [87]. The majority of responders developed GvHD. Responses were maintained in 11 patients at a median of 2.2 years from last DLI, and a further three died in CR of complications of GvHD. It seems likely that experience with DLI in relapsed HL will grow rapidly over coming years. 6.5
CLL Relapse following allogeneic SCT for CLL is reported in 20 -- 48% of patients and responds poorly to standard salvage chemotherapy. Reports on the use of DLI are limited, but show highly variable response rates (0 -- 60% CR) which are durable only in a minority [88-91]. The reasons why DLI often fails in CLL giving low durability responses is unclear. One group attempted to correlate clinical responses post-allograft with MRD kinetics by MFC and/or real time PCR and identified a pattern of an initial clinical GvT effect accompanied by a failure to completely eradicate MRD followed by subsequent frank relapse despite extensive chronic GvHD [92]. Possible mechanisms to explain this disease kinetic include clonal evolution, sanctuary sites, the development of tolerance and the presence of CLL stem cells which fail to be targeted by DLI. Unraveling this picture may elucidate further the pathophysiology underlying refractory and relapsing CLL. Novel immunological strategies to target CLL include the use of activated DLI (aDLI) which is addressed later in the article [93]. Combined therapy with DLI and Bi20 (FBTA05), a trifunctional, bispecific antibody targeting CD20 and CD3 which is thought to direct T cells towards 6.6
Expert Opin. Biol. Ther. (2011) 11(4)
477
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
CLL cells has been used in patients with refractory, tumor protein 53-mutated CLL. Transient clinical responses were observed, but disease recurrence was identified soon after cessation of therapy, despite further doses of DLI [94]. MM Relapse post-transplant is a significant problem in MM, affecting 50% of patients who achieve CR and 80% of those who do not achieve CR. Therapy with DLI gives overall response rates of 38% (CR = 19%, PR = 19%) complicated by high levels of acute GvHD (38% of patients) and chronic GvHD (42% of patients). A strong correlation exists between the likelihood of a favorable response to DLI and the development of GvHD [95]. Long-term survival is possible in 60% of patients who achieve CR with DLI whereas survival at 22 months is < 20% in those who achieve only PR post-DLI [96]. In one study of RIC transplantation in chemo-sensitive MM, only 2/20 patients achieved CR. In all, 14/20 received escalating-dose DLI for residual/progressive disease and 5/14 developed GvHD (which appeared to correlate with disease responses). Response durations were short (five were < 12 months) and progression often occurred despite persisting FDC, giving 2-year PFS rates of 30%. Dose escalation did not permit dissociation of the GvT from the GvHD effect [97]. Attempts are ongoing to define strategies to augment the efficacy of DLI using novel chemotherapy agents. The proteosome inhibitor bortezomib has been combined with DLI in animal models and found to reduce the incidence of GvHD whilst preserving the crucial GvT effect [98]. Lenalidomide and thalidomide can activate T cells and NK cells and may augment GvT activity [99].
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
6.7
7.
DLI in the pediatric setting
Compared to adults, children with hematologic malignancy show superior survival post-allogeneic SCT, but posttransplant relapse represents a significant problem [100]. The role of DLI in managing post-SCT relapse in pediatric hematologic malignancy is unclear, as most published data on the subject comprise small case series [101-103]. One important comparative analysis of outcomes in 49 children who received DLI for post-transplant relapse (18 ALL, 17 AML, 8 CML, 4 MDS, 2 juvenile myelomonocytic leukemia) and 1229 historical controls (no DLI) reported to the Centre for International Blood and Marrow Transplant Research demonstrated that DLI did not result in survival benefit for the majority of children treated [100], perhaps primarily reflecting the disease histologies represented rather than the age group of the patients. Several other studies have focused on strategies to try to prevent relapse. Bader et al. have defined how increasing MC after allogeneic SCT is an important prognostic factor for unfavorable outcome in children with ALL. They show that the probability of 3-year event-free survival (EFS) in 478
patients with FDC or low level MC is 66% compared with 23% for patients with increasing MC. They go on to show that of the increasing MC cohort, those who received immunotherapy had a 3-year EFS rate of 37% compared to 0% in those who did not receive immunotherapy and they propose that overt relapse could be prevented in a considerable group of patients through chimerism monitoring and early intervention with DLI in cases of increasing MC [104,105]. As in adults, post-transplant cellular therapies for ALL have not yet been fully optimized, although clinical studies of chimeric antigen receptors (CARs) targeting CD19 on leukemia cells have been established in pediatric ALL and results are awaited with interest. It is important to note that DLI is also an accepted treatment option for some non-malignant hematologic conditions such as severe aplastic anemia (SAA) and thalassemia major. Treatment failure is rare in SAA, but is mostly caused by graft rejection. Pediatric studies have suggested that early, low-dose DLI for increasing MC can prevent graft rejection whilst increasing the risk of GvHD [106]. It is recognized that MC and secondary graft failure are frequent complications of allogeneic SCT for thalassemia. Escalating doses of DLI in this setting have been shown to be safe but are only efficacious in restoring FDC in patients with low-risk relapse rather than in patients with high-risk relapse. For this reason, the authors suggest that DLI be commenced on detection of level 2 -- 3 chimerism (donor < 90%) [107].
General principles of DLI: effective cell dose, timing, toxicity and donor issues
8.
The best evidence addressing the optimal cell dose for DLI therapy has been established in CML. In one study, 68 patients received DLI in an escalating dose regimen (EDR) and the proportion of clinical responses increased with each increment in cell dose. Subgroup analysis showed that the effective cell dose in CML correlates with disease stage and donor type such that patients with cytogenetic or molecular relapse and those with unrelated donors respond to lower doses (< 107 CD3+ cells/kg) compared to patients with advanced CML or those with fully-matched sibling donors [108]. The optimal cell dose for most hematological malignancies is yet to be defined, but in some diseases such as MM there appears to be no clear correlation between CD3 cell dose and clinical response [109]. GvHD complicates DLI therapy in ~ 30% of patients and can be life threatening [110], although it is important to recognize that GvHD developing either following transplantation or DLI is a manifestation of alloreactivity which often acts as a marker for GvT activity. Therefore, although severe GvHD may compromise overall outcomes, less severe acute GvHD or limited chronic GvHD has been associated with superior outcomes in terms of reduced relapse risk. This impact was first recognized in CML, but has been described
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Roddie & Peggs
across many other pathologies now including acute leukemias, myeloma and a variety of lymphoma subtypes. GvHD risk does not appear to correlate with the underlying hematological malignancy being treated, but is positively correlated with the administered dose of DLI. In CML, a dose of 1 107/kg can induce complete donor chimerism and a potent graftversus-leukemia (GVL) effect, in some cases in the absence of clinical GvHD particularly if given at later time points following transplantation [31]. The time interval between SCT and DLI therapy appears to influence the likelihood of developing GvHD. A small dose of 1 105 T cells/kg can induce GvHD if administered on the day of transplant [111], yet a dose of 1 107 T cells/kg can be given at 12 months post-transplant without GvHD developing [31]. It is thought that the inflammatory cytokines produced in the immediate post-transplant period activate alloreactive donor T cells and lower the threshold for the emergence of GvHD. Changes in the frequency of recipientderived APCs over time may also have relevance to GvHD risk. Furthermore, the development of extrinsic mechanisms of peripheral T-cell tolerance (e.g., Tregs) at later time points may also modulate this risk. Other factors which make GvHD more likely to occur include donor sex mismatch (female donor to male recipient), advanced patient age and mismatch at the mHag level [112]. Aplasia is now a relatively infrequent complication of DLI. It is often transient, but in some cases may require hematopoietic stem cell rescue. It was reported historically in 15 -- 20% of treated CML patients with an associated mortality rate of ~ 5%. Aplasia is more common in hematological relapse of CML, possibly due to poor donor myeloid reserve, and is rarely reported in patients with exclusively cytogenetic or molecular relapse [33,113] or in those treated for low levels of recipient MC. Another important issue to consider when planning DLI is donor availability and willingness to undergo further leukopheresis. This process can bring significant delays in treatment and may compromise patient outcomes. Some groups advocate DLI collection at the outset to avoid delays, but the inherent difficulties with this approach include the need for additional donor leukopheresis, the financial burden of collecting DLI for all transplant patients and the handling and storage issues surrounding the DLI product once it is collected when there is a strong possibility that it will never be needed or used. It is the practice in our institution to prepare and store aliquots of DLI from the primary harvests of matched unrelated donors where the yield is in excess of the target dose of 4 106 CD34+ cells/kg necessary for transplantation. 9.
Strategies to avoid DLI-associated toxicity
The escalating dose regimen Administration of DLI as a single bolus of cells collected from a single leukopheresis and containing variable numbers of CD3+ T cells is referred to as a bulk dose regimen (BDR)
and this approach is associated with a high incidence of GvHD [33,75,114]. The EDR approach is fundamentally different in that the DLI product is quantitated for CD3+, CD4+ and CD8+ T-cell numbers and is then administered in multiple small aliquots with a dose escalation over time. In this way, the minimum cell dose needed to achieve disease remission is administered and with more modest cell doses, the likelihood of GvHD may be reduced [31]. One study in CML comparing BDR and EDR approaches demonstrated equivalent remission rates with both schedules, but a significantly lower incidence of GvHD in the EDR cohort [113]. It is critical when using the EDR schedule to allow an adequate interval between DLI doses to allow for assessment of response and toxicity. The optimum interval between doses is yet to be defined, but Dazzi et al. report that shorter intervals (rather than total cell dose) leads to a higher incidence of GvHD [113]. Mackinnon et al. recommend a minimum period of 3 months between escalating doses in clinically stable patients [31]. Suicide gene transfected donor T cells A potential solution to the undesirable alloreactivity of unmanipulated T cells is the transfection of donor T cells with a ‘suicide gene’ to permit their elimination in the event of GvHD. The ideal suicide gene is non-immunogenic, nontoxic in the quiescent state, but can efficiently trigger cell death once activated. Transfection of donor T cells with the herpes simplex virus 1-thymidine kinase (HSV-TK) gene has been described in several clinical trials. This strategy is purported to be safe and effective in controlling GvHD via ganciclovir-induced elimination of HSV-TK transfected T cells [115,116]. Problems with this approach include concerns over the possible adverse impact of this approach on GvT activity, the possible immunogenicity of the transfected cells which could promote their rapid clearance from the blood and alterations in the TK gene which lead to the expression of a non-functional protein [117,118]. These issues need to be addressed prior to this strategy being consolidated in clinical practice. Research is ongoing into other suicide genes such as chimeric Fas and caspase-9. Timed induction of these genes can lead to T-cell apoptosis and this may represent a promising non-viral alternative to HSV-TK [119,120]. 9.2
Cell selection/subsets With advances in available graft engineering techniques, DLI can now be tailored in attempts to tip the balance of immunity away from GvHD and towards GvT. Strategies to define the optimal cell type/combination to use, the cell activation status and the antigenic specificity are outlined below. 9.3
9.1
Selective depletion of alloreactive cells Alloreactive T cells, defined by their expression of activation-induced antigens such as CD25, CD69, 9.3.1
Expert Opin. Biol. Ther. (2011) 11(4)
479
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
CD137 or CD134 in response to exposure to host antigens, can be depleted from the DLI product with a view of reducing the incidence of GvHD. Co-incubation of donor lymphocytes with allogenic recipient stimulator cells followed by targeting with immunotoxin-conjugated antibodies specific for cell-surface activation markers or antibodies which can be sorted via immunomagnetic beads, permits separation of activated cells from DLI prior to infusion. Whether this approach reduces alloreactivity towards hematopoeisisrestricted mHags and reduces potential GvT activity remains to be clarified [121,122]. Manipulated DLI (CD8 T-cell depletion) CD8+ T cells are thought to be the primary mediators of GvHD in humans whilst CD4+ T cells are reported to contribute more to the GvT effect. A number of recent papers have focused on demonstrating how CD4 T lymphocytes are crucial to the development of the DLI-associated GvT effect. In one study, in vivo CD4 T-cell depletion abolished the antitumor effect of DLI which, by contrast, was not impacted by depletion of CD8 T cells. CD4 T cells clearly play an essential role in mediating early antitumor effects [123]. For this reason, a number of groups have explored CD8+ T-cell depletion as a strategy to reduce the incidence of GvHD. CD8+ T-cell depletion of the stem cell graft has been reported to reduce the risk of GvHD without a parallel increase in relapse rates [124]. This has also been confirmed in the DLI setting in CML [125,126]. A Phase I study of high-stringency immunomagnetic CD8 T-cell depletion of DLI was reported in which escalating doses of CD8 T-cell depleted DLI were given at 3-monthly intervals to patients with persistent disease or MC or disease. Responses were documented in 8/16 of the former and 5/11 of the latter. Five developed acute grade II -- IV GvHD and two died of GvHD-related complications. Clearly, GvHD remains a major problem despite CD8+ T-cell depletion and further studies are warranted to define the potential benefits and risks more clearly [127].
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
9.3.2
Future therapeutic options and research imperatives in the field of DLI
10.
T-cell engineering The main clinical imperative driving research in DLI biology remains how to shift the immunological balance of cellular immunotherapy away from GvHD towards GvT. Ideally, one would aim to select (and possibly expand) a highavidity tumor-reactive T-cell clone in vitro and then infuse it to the patient as DLI. Difficulties with this approach include identifying a high-avidity tumor-reactive clone in the first place (many have undergone central deletion due to the risk of autoimmunity of any high-avidity self-specific T cell) and also the deleterious impact of prolonged cell culture on subsequent T-cell persistence and function (terminal differentiation and exhaustion). Optimal cell culture 10.1
480
conditions are beyond the scope of this review, but are discussed in a review by Aqui and June [128]. T-cell engineering can potentially overcome the limitations of adoptive cellular therapies by introducing antigen receptors into T cells to re-direct their specificity. This potentially allows rapid generation of tumor-reactive T cells expressing either HLA-restricted, heterodimeric TCRs or CARs that recognize native cell-surface antigens. Initial clinical studies of TCR gene transfer have been described in metastatic melanoma using a high affinity MART (melanoma antigen recognized by T cells)-1 specific TCR. Tumor regression was reported in 30% of patients. Treatment was complicated by off-target toxicity associated with the destruction of melanocytes elsewhere in the body, but this responded to steroid therapy [129]. Clinical studies of first-generation CARs have been conducted in ovarian and renal cancers, lymphoma and neuroblastoma, although results to date have been somewhat disappointing and have highlighted potential issues with toxicity [130-133]. Second generation CARs often comprise an antibody binding motif and a CD28--CD3z dual signaling receptor which facilitates T-cell activation and expansion following stimulation. Studies of refined second generation CARs directed against CD19, CD20, CD23, CD33 and CD74 are awaited with interest. It is important to note that the development and safety monitoring of these new immunotherapies as advanced medicine therapy products are the subject of EU regulations, but this is beyond the scope of this review. aDLI Resistance to the therapeutic effects of DLI may occur due to failure of ‘in vivo’ activation of donor T cells. Several studies have reported that IL-2 stimulation of donor T cells (both in vivo and ex vivo) can induce clinical responses in patients who are resistant to DLI alone. Porter et al. showed that infusion of ‘ex vivo’ activated donor lymphocytes (using anti-CD3 and anti-CD28 coated beads) in patients with a range of hematological malignancies led to responses where conventional DLI had been disappointing. A total of 17 patients were evaluated and 8 achieved CR. Of those, 4/8 relapsed. The incidence of GvHD in this cohort compared favorably with that of conventional DLI [93]. Presently, Phase I and II studies of aDLI in CLL are underway at the University of Pennsylvania. 10.2
11.
Conclusions
Current experience provides evidence that DLI is the most consolidated approach to the management of disease relapse following allogeneic SCT. DLI is particularly effective in managing relapsed CML-CP, but encouraging clinical responses have also been reported in other hematological malignancies. The main drawback of DLI is GvHD which affects ~ 30% of all treated patients and can be life
Expert Opin. Biol. Ther. (2011) 11(4)
Roddie & Peggs
threatening. Rapid advances in our understanding of transplant biology have led to the development of strategies which aim to preserve the GvL effect whilst inhibiting the GvHD effect. The EDR approach, cell selection and genetic engineering strategies all offer the potential for refining DLI, but there is still a lack of definitive published data to guide our clinical practice. Ultimately, large, collaborative clinical trials are warranted.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
12.
Expert opinion
DLI will continue to have an important role in the management of disease relapse following allogeneic transplantation. Its potential has been demonstrated in a number of diseases, but attempts to standardize approaches within the context of prospective studies have been limited. There are a number of reasons for this, but growing international recognition of the importance of collaborative programs may help to define the most important outstanding questions and define appropriate therapeutic strategies either across disease subtypes (e.g., timing, dosing) or within specific disease histologies (e.g., the role of particular cytoreductive combinatorial approaches). Toxicity due to GvHD remains common and attempts to reduce this should be further evaluated in prospective studies. DLI administration via the EDR has been shown to minimize the GvHD risk in studies of CML with no apparently adverse impact on GvT function, and confirmation of similar efficacy in other diseases should be forthcoming within the next few years. Improvements in defining the risk of relapse following allogeneic transplantation are crucial for future studies and
may help to establish which patients are of sufficiently high risk to merit consideration for prophylactic DLI studies. Disease type and status at transplant taken together with MRD monitoring will be important factors in directing intervention. Elucidation of the impact of MC, potentially differing according to the transplantation platform used, will also require more concerted attention. It is likely that the role of graft manipulations will be further defined within the next 5 -- 10 years, including aDLI, T-cell subset selection, allodepletion and genetic modification. It will be critical to define the impact on GvT activity of any manipulation designed to reduce GvHD (e.g., Treg infusion). Parallel studies aimed at manipulating APCs may be equally important. The durability of DLI responses in many diseases and the factors which might influence this also merit further investigation, and perhaps there is a need to explore the concept of maintenance therapy (either in terms of DLI or combinations of newer pharmacological agents). One thing that becomes increasingly apparent when reviewing the literature on DLI is that there are many more questions than answers. The remarkable ability of the donor immune system to eradicate hematological tumors is undoubted, but taming this potential will require considerably more research within the context of larger collaborative networks.
Declaration of interest The authors declare no conflict of interest and have received no payment in preparation of this manuscript.
Expert Opin. Biol. Ther. (2011) 11(4)
481
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
Bibliography Papers of special note have been highlighted as either of interest () or of considerable interest () to readers. 1.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
2.
Aurer I, Gale RP. Are new conditioning regimens for transplants in acute myelogenous leukaemia better? Bone Marrow Transplantation 1991;7:255-61 Clift RA, Buckner CD, Appelbaum FR, et al. Allogeneic marrow transplantation in patients with chronic myeloid leukaemia in the chronic phase: a randomized trial of two irradiation regimens. Blood 1991;77:1660-5
3.
Niederwieser D, Lange T, Cross M, et al. Reduced intensity conditioning (RIC) haematopoietic cell transplants in elderly patients with AML. Best Pract Res Clin Haematol 2006;19(4):825-38
4.
Slavin S. New strategies for bone marrow transplantation. Cur Opin Immunol 2000;12:542-51
5.
Carella AM, Champlin R, Slavin S, et al. Mini-allografts: ongoing trials in humans. Bone Marrow Transplant 2000;25:345-50
6.
7.
8.
9.
10.
482
11.
12.
13.
14.
Laport GG, Sandmaier BM, Storer BE, et al. Reduced-intensity conditioning followed by allogeneic hematopoietic cell transplantation for adult patients with myelodysplastic syndrome and myeloproliferative disorders. Biol Blood Marrow Transplant 2008;14:246-55
19.
Aversa F, Tabilio A, Velardi A, et al. Treatment of high-risk acute leukemia with T-cell-depleted stem cells from related donors with one fully mismatched HLA haplotype. N Engl J Med 1998;339:1186-93
Bonnet D, Warren EH, Greenberg PD, et al. CD8(+) minor histocompatibility antigen-specific cytotoxic T lymphocyte clones eliminate human acute myeloid leukemia stem cells. Proc Natl Acad Sci USA 1999;96(15):8639-44
20.
Molldrem JJ, Clave E, Jiang YZ, et al. Cytotoxic T lymphocytes specific for a nonpolymorphic proteinase 3 peptide preferentially inhibit chronic myeloid leukemia colony-forming units. Blood 1997;90(7):2529-34
21.
Xue SA, Gao L, Hart D, et al. Elimination of human leukemia cells in NOD/SCID mice by WT1-TCR gene-transduced human T cells. Blood 2005;106:3062-7
22.
Molldrem JJ, Lee PP, Wang C, et al. Evidence that specific T lymphocytes may participate in the elimination of chronic myelogenous leukemia. Nat Med 2000;6:1018-23
23.
Kolb HJ, Mittermuller J, Clemm C, et al. Donor leukocyte transfusions for treatment of recurrent chronic myelogenous leukemia in marrow transplant patients. Blood 1990;76:2462-5 The original clinical paper describing the first use of DLI in CML.
Porter DL, Alyea EP, Antin JH, et al. NCI First International Workshop on the biology, prevention, and treatment of relapse after allogeneic hematopoietic stem cell transplantation: report from the committee on treatment of relapse after allogeneic hematopoietic stem cell transplantation. Biol Blood Marrow Transplant 2010;16(11):1467-503 Horowitz MM, Gale RP, Sondel PM, et al. Graft-versus-leukemia reactions after bone marrow transplantation. Blood 1990;75(3):555-62 Collins RH, Rogers ZR, Bennett M, et al. Hematologic relapse of chronic myelogenous leukemia following allogeneic bone marrow transplantation: apparent graft-versus-leukemia effect following abrupt discontinuation of immunosuppression. Bone Marrow Transplant 1992;10:391-5
Hegenbart U, Niederweser D, Sandmaier BM, et al. Treatment for acute myelogenous leukaemia by low-dose, total body, irradiation-based conditioning and hematopoietic cell transplantation from related and unrelated donors. J Clin Oncol 2006;24:444-53
15.
Bensinger WI. The current status of reduced-intensity allogeneic hematopoietic stem cell transplantation for multiple myeloma. Leukemia 2006;20:1683-9
16.
Gale RP, Horowitz MM, Ash RC, et al. Identical-twin bone marrow transplants for leukemia. Ann Intern Med 1994;120:646-52
17.
Marmont AM, Horowitz MM, Gale RP, et al. T-cell depletion of HLA-identical transplants in leukemia. Blood 1991;78:2120-30
18.
Apperley JF, Jones L, Hale G, et al. Bone marrow transplantation for patients with chronic myeloid leukaemia: T-cell depletion with Campath-1 reduces the incidence of graft-versus-host disease but may increase the risk of leukaemic
Bagicalupo A. Results of hematopoietic stem cell transplantation for hematologic malignancies. In: Hoffman R, Benz Jr EJ, Shattil SJ, et al. editors. Hematology: basic principles and practice. 4th edition. Churchill Livingstone, Edinburgh; 2004. p. 1713-25 Alyea EP, DeAngelo DJ, Moldrem J, et al. NCI First International Workshop on the biology, prevention and treatment
relapse. Bone Marrow Transplant 1986;1:53-66
of relapse after allogeneic hematopoietic cell transplantation: report from the committee on prevention of relapse following allogeneic cell transplantation for hematologic malignancies. Biol Blood Marrow Transplant 2010;16(8):1037-69
Higano CS, Brixey M, Bryant EM, et al. Durable complete remission of acute nonlymphocytic leukemia associated with discontinuation of immunosuppression following relapse after allogeneic bone marrow transplantation. A case report of a probable graft-versus-leukemia effect. Transplantation 1990;50:175-7
Expert Opin. Biol. Ther. (2011) 11(4)
..
24.
Cullis JO, Jiang YZ, Schwarer AP, et al. Donor leukocyte infusions for chronic myeloid leukemia in relapse after allogeneic bone marrow transplantation. Blood 1992;79:1379-81
25.
Bar BM, Schattenberg A, Mensink EJ, et al. Donor leukocyte infusions for chronic myeloid leukemia relapsed after allogeneic bone marrow transplantation. J Clin Oncol 1993;11:513-19
26.
Drobyski WR, Keever CA, Roth MS, et al. Salvage immunotherapy using donor leukocyte infusions as treatment for relapsed chronic myelogenous leukemia after allogeneic bone marrow transplantation: efficacy and toxicity of a defined T-cell dose. Blood 1993;82:2310-18
27.
Helg C, Roux E, Beris P, et al. Adoptive immunotherapy for recurrent CML after BMT. Bone Marrow Transplant 1993;12:125-9
28.
Hertenstein B, Wiesneth M, Novotny J, et al. Interferon-alpha and donor buffy
Roddie & Peggs
coat transfusions for treatment of relapsed chronic myeloid leukemia after allogeneic bone marrow transplantation. Transplantation 1993;56:1114-18 29.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
30.
31.
.
32.
33.
.
Porter DL, Roth MS, McGarigle C, et al. Induction of graft-versus-host disease as immunotherapy for relapsed chronic myeloid leukemia. N Engl J Med 1994;330:100-6 van Rhee F, Lin F, Cullis JO, et al. Relapse of chronic myeloid leukemia after allogeneic bone marrow transplant: the case for giving donor leukocyte transfusion before the onset of hematologic relapse. Blood 1994;83:3377-83 Mackinnon S, Papadopoulos EB, Carabasi MH, et al. Adoptive immunotherapy evaluating escalating doses of donor leukocytes for relapse of chronic myeloid leukemia after bone marrow transplantation: separation of graft-versus-leukemia responses from graft-versus-host disease. Blood 1995;86:1261-8 The first report on the Escalating Dose Regimen and its potential to reduce the incidence of DLI-related GvHD. Kolb HJ, Schattenberg A, Goldman JM, et al. Graft-versus-leukemia effect of donor lymphocyte transfusions in marrow grafted patients. European Group for Blood and Marrow Transplantation Working Party Chronic Leukemia. Blood 1995;86:2041-50 Collins RH, Shpilberg O, Drobyski WR, et al. Donor leukocyte infusions in 140 patients with relapsed malignancy after allogeneic bone marrow transplantation. J Clin Oncol 1997;15:433-44 The largest registry review of DLI for a range of haematological malignancies.
36.
37.
38.
39.
40.
Schaap N, Schattenberg A, Bar B, et al. Induction of graft-versus-leukemia to prevent relapse after partially lymphocyte-depleted allogeneic bone marrow transplantation by preemptive donor leukocyte infusions. Leukemia 2001;15(9):1339-46 van der Griend R, Verdonck LF, Petersen EJ, et al. Donor leukocyte infusions inducing remissions repeatedly in a patient with recurrent multiple myeloma after allogeneic bone marrow transplantation. Bone Marrow Transplant 1999;23(2):195-7 Verdonck LF, Petersen EJ, Lokhorst HM, et al. Donor leukocyte infusions for recurrent hematologic malignancies after allogeneic bone marrow transplantation: impact of infused and residual donor T cells. Bone Marrow Transplant 1998;22(11):1057-63 Shlomchik WD, Couzens MS, Tang CB, et al. Prevention of graft versus host disease by inactivation of host antigen-presenting cells. Science 1999;285(5426):412-15 Mapara MY, Kim YM, Wang SP, et al. Donor lymphocyte infusions mediate superior graft-versus-leukemia effects in mixed compared to fully allogeneic chimeras: a critical role for host antigen-presenting cells. Blood 2002;100(5):1903-9
41.
Bleakley M, Riddell SR. Molecules and mechanisms of the graft-versus leukaemia effect. Nat Rev Cancer 2004;4(5):371-80
42.
Mutis T, Verdijk R, Schrama E, et al. Feasibility of immunotherapy of relapsed leukemia with ex vivo-generated cytotoxic T lymphocytes specific for hematopoietic system-restricted minor histocompatibility antigens. Blood 1999;93:2336-41
34.
Lokhorst HM, Schattenberg A, Cornelissen JJ, et al. Donor leukocyte infusions are effective in relapsed multiple myeloma after allogeneic bone marrow transplantation. Blood 1997;90(10):4206-11
43.
Bonnet D, Warren EH, Greenberg PD, et al. CD8(+) minor histocompatibility antigen-specific cytotoxic T lymphocyte clones eliminate human acute myeloid leukemia stem cells. Proc Natl Acad Sci USA 1999;96:8639-44
35.
Shiobara S, Nakao S, Ueda M, et al. Donor leukocyte infusion for Japanese patients with relapsed leukemia after allogeneic bone marrow transplantation: lower incidence of acute graft-versus-host disease and improved outcome. Bone Marrow Transplant 2000;26(7):769-74
44.
Hsieh MH, Korngold R. Differential use of FasL- and perforin-mediated cytolytic mechanisms by T-cell subsets involved in graft-versus-myeloid leukemia responses. Blood 2000;96(3):1047-55
45.
Piccirillo CA, Shevach EM. Naturally-occurring CD4+ CD25+ immunoregulatory T cells: central players Expert Opin. Biol. Ther. (2011) 11(4)
in the arena of peripheral tolerance. Semin Immunol 2004;16:81-8 46.
Fehervari Z, Sakaguchi S. Development and function of CD25+CD4+ regulatory T cells. Curr Opin Immunol 2004;16:203-8
47.
Edinger M, Hoffmann P, Ermann J, et al. CD4+CD25+ regulatory T cells preserve graft-versus-tumor activity while inhibiting graft-versus-host disease after bone marrow transplantation. Nat Med 2003;9(9):1144-50
48.
Piccirillo CA, Shevach EM. Naturally-occurringCD4+CD25+ immunoregulatory T cells: central players in the arena of peripheral tolerance. Semin Immunol 2004;16:81-8
49.
Knechtli CJ, Goulden NJ, Hancock JP, et al. Minimal residual disease status as a predictor of relapse after allogeneic bone marrow transplantation for children with acute lymphoblastic leukaemia. Br J Haematol 1998;102:860-71
50.
Mackinnon S, Barnett L, Heller G, et al. Minimal residual disease is more common in patients who have mixed T-cell chimerism after bone marrow transplantation for chronic myelogenous leukemia. Blood 1994;83:3409-16
51.
Kroger N, Bacher U, Bader P, et al. NCI First International Workshop on the biology, prevention, and treatment of relapse after allogeneic hematopoietic stem cell transplantation: report from the committee on disease-specific methods and strategies for monitoring relapse following allogeneic stem cell transplantation. Part I: methods, acute leukemias, and myelodysplastic syndromes. Biol Blood Marrow Transplant 2010;16(9):1187-211
52.
Kroger N, Bacher U, Bader P, et al. NCI first international workshop on the biology, prevention, and treatment of relapse after allogeneic hematopoietic stem cell transplantation: report from the committee on disease-specific methods and strategies for monitoring relapse following allogeneic stem cell transplantation. part II: chronic leukemias, myeloproliferative neoplasms, and lymphoid malignancies. Biol Blood Marrow Transplant 2010;16(10):1325-46
53.
Sairafi D, Remberger M,Uhlin M, et al. “Leukemia lineage-specific chimerism analysis and molecular monitoring improve outcome of donor lymphocyte
483
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
54.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
55.
.
56.
57.
58.
59.
Peggs KS, Thomson K, Hart DP, et al. Dose-escalated donor lymphocyte infusions following reduced intensity transplantation: toxicity, chimerism, and disease responses. Blood 2004;103(4):1548-56 Kaeda J, O’shea D, Szydlo RM, et al. Serial measurement of BCR-ABL transcripts in the peripheral blood after allogeneic stem cell transplantation for chronic myeloid leukemia: an attempt to define patients who may not require further therapy. Blood 2006;107:4171-6 Excellent summary on the role of MRD monitoring post-transplant in CML patients. Hughes TP, Deininger MW, Hochhaus A, et al. Monitoring CML patients responding to treatment with tyrosine kinase inhibitors -- Review and recommendations for ‘harmonizing’ current methodology for detecting BCR-ABL transcripts and kinase domain mutations and for expressing results. Blood 2006;108:28-37 Stirewalt DL, Guthrie KA, Beppu L, et al. Predictors of relapse and overall survival in Philadelphia chromosome-positive acute lymphoblastic leukemia after transplantation. Biol Blood Marrow Transplant 2003;9:206-12 San Miguel JF, Vidriales MB, Lopez-Berges C, et al. Early immunophenotypical evaluation of minimal residual disease in acute myeloid leukemia identifies different patient risk groups and may contribute to postinduction treatment stratification. Blood 2001;98:1746-51
62.
63.
Kroger N, Zagrivnaja M, Schwartz S, et al. Kinetics of plasma-cell chimerism after allogeneic stem cell transplantation by highly sensitive real-time PCR based on sequence polymorphism and its value to quantify minimal residual disease in patients with multiple myeloma. Exp Hematol 2006;34:688-94 Raiola AM, Van Lint MT, Valbonesi M, et al. Factors predicting response and graft-versus-host disease after donor lymphocyte infusions: a study on 593 infusions. Bone Marrow Transplant 2003;31:687-93
64.
Ocheni S, Iwanski GB, Schafhausen P, et al. Characterisation of extramedullary relapse in patients with chronic myeloid leukemia in advanced disease after allogeneic stem cell transplantation. Leuk Lymphoma 2009;50(4):551-8
65.
Cummins M, Cwynarski K, Marktel S, et al. Management of chronic myeloid leukaemia in relapse following donor lymphocyte infusion induced remission: a retrospective study of the Clinical Trials Committee of the British Society of Blood & Marrow Transplantation (BSBMT). Bone Marrow Transplant 2005;36:1065-9
66.
67.
Mitterbauer G, Zimmer C, Fonatsch C, et al. Monitoring of minimal residual leukemia in patients with MLL-AF9 positive acute myeloid leukemia by RT-PCR. Leukemia 1999;13:1519-24
von dem Borne PA, van Luxemburg-Heijs SA, Heemskerk MH, et al. Molecular persistence of chronic myeloid leukemia caused by donor T cells specific for lineage-restricted maturation antigens not recognizing immature progenitor-cells. Leukemia 2006;20(6):1040-6 Posthuma EF, Marijt EW, Barge RM, et al. Alpha-interferon with very-low-dose donor lymphocyte infusion for hematologic or cytogenetic relapse of chronic myeloid leukemia induces rapid and durable complete remissions and is associated with acceptable graft-versus-host disease. Biol Blood Marrow Transplant 2004;10(3):204-12
60.
Provan D, Bartlett-Pandite L, Zwicky C, et al. Eradication of polymerase chain reaction-detectable chronic lymphocytic leukemia cells is associated with improved outcome after bone marrow transplantation. Blood 1996;88:2228-35
68.
Slavin S, Naparstek E, Nagler A, et al. Allogeneic cell therapy with donor peripheral blood cells and recombinant human interleukin-2 to treat leukemia relapse after allogeneic bone marrow transplantation. Blood 1996;87:2195-204
61.
Galimberti S, Benedetti E, Morabito F, et al. Prognostic role of minimal residual disease in multiple myeloma patients after non-myeloablative allogeneic
69.
Savani BN, Montero A, Kurlander R, et al. Imatinib synergizes with donor lymphocyte infusions to achieve rapid molecular remission of CML relapsing
484
after allogeneic stem cell transplantation. Bone Marrow Transplant 2005;36:1009-15
transplantation. Leukemia Res 2005;29:961-6
infusions”. Biol Blood Marrow Transplant 2010;16(12):1728-37
Expert Opin. Biol. Ther. (2011) 11(4)
70.
Chen J, Schmitt A, Chen B, et al. Imatinib impairs CD8+ T lymphocytes specifically directed against the leukemia associated antigen RHAMM/CD168 in vitro. Cancer Immunol Immunother 2007;56:849-61
71.
Gao H, Lee BN, Talpaz M, et al. Imatinib mesylate suppresses cytokine synthesis by activated CD4 T cells of patients with chronic myelogenous leukemia. Leukemia 2005;19:1905-11
72.
Schmid C, Labopin M, Nagler A, et al. Donor lymphocyte infusion in the treatment of first hematological relapse after allogeneic stem-cell transplantation in adults with acute myeloid leukemia: a retrospective risk factors analysis and comparison with other strategies by the EBMT Acute Leukemia Working Party. J Clin Oncol 2007;25(31):4938-45
73.
Schmid C, Schleuning M, Ledderose G, et al. Sequential regimen of chemotherapy, reduced-intensity conditioning for allogeneic stem-cell transplantation, and prophylactic donor lymphocyte transfusion in high-risk acute myeloid leukemia and myelodysplastic syndrome. J Clin Oncol 2005;23:5675-87
74.
Fielding AK, Richards SM, Chopra R, et al. Medical Research Council of the United Kingdom Adult ALL Working Party; Eastern Cooperative Oncology Group. Outcome of 609 adults after relapse of acute lymphoblastic leukemia (ALL); an MRC UKALL12/ECOG 2993 study. Blood 2007;109(3):944-50
75.
Porter DL, Collins RH Jr, Hardy C, et al. Treatment of relapsed leukemia after unrelated donor marrow transplantation with unrelated donor leukocyte infusions. Blood 2000;95(4):1214-21
76.
Russell NH, Byrne JL, Faulkner RD, et al. Donor lymphocyte infusions can result in sustained remissions in patients with residual or relapsed lymphoid malignancy following allogeneic haemopoietic stem cell transplantation. Bone Marrow Transplant 2005;36(5):437-41
77.
Thomson KJ, Morris EC, Milligan D, et al. T-cell-depleted reduced-intensity transplantation followed by donor leukocyte infusions to promote
Roddie & Peggs
graft-versus-lymphoma activity results in excellent long-term survival in patients with multiply relapsed follicular lymphoma. J Clin Oncol 2010;28(23):3695-700 78.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
79.
80.
81.
Bloor AJ, Thomson K, Chowdhry N, et al. High response rate to donor lymphocyte infusion after allogeneic stem cell transplantation for indolent non-Hodgkin lymphoma. Biol Blood Marrow Transplant 2008;14(1):50-8
Sureda A, Robinson S, Canals C, et al. Reduced-intensity conditioning compared with conventional allogeneic stem-cell transplantation in relapsed or refractory Hodgkin’s lymphoma: an analysis from the Lymphoma Working Party of the European Group for Blood and Marrow Transplantation. J Clin Oncol 2008;26(3):455-62
87.
Peggs KS, Kayani I, Edwards N, et al. Donor lymphocyte infusions modulate relapse risk in mixed chimeras and induce durable salvage in relapsed patients following T-cell--depleted allogeneic transplantation for Hodgkin’s lymphoma. J Clin Oncol 2011; In press
Thomson KJ, Morris EC, Bloor A, et al. Favorable long-term survival after reduced-intensity allogeneic transplantation for multiple-relapse aggressive non-Hodgkin’s lymphoma. J Clin Oncol 2009;27(3):426-32 Bishop MR, Dean RM, Steinberg SM, et al. Clinical evidence of a graft-versus-lymphoma effect against relapsed diffuse large B-cell lymphoma after allogeneic hematopoietic stem-cell transplantation. Ann Oncol 2008;19(11):1935-40
88.
Cook G, Smith GM, Kirkland K, et al. Clinical Trials Committee (CTC) of the British Society for Blood and Marrow Transplantation (BSBMT). Outcome following Reduced-Intensity Allogeneic Stem Cell Transplantation (RIC AlloSCT) for relapsed and refractory mantle cell lymphoma (MCL): a study of the British society for blood and marrow transplantation. Biol Blood Marrow Transplant 2010;16(10):1419-27
89.
Gribben JG, Zahrieh D, Stephans K, et al. Autologous and allogeneic stem cell transplantations for poor-risk chronic lymphocytic leukemia. Blood 2005;106(13):4389-96
90.
Ritgen M, Stilgenbauer S, von Neuhoff N, et al. Graft-versus-leukemia activity may overcome therapeutic resistance of chronic lymphocytic leukemia with unmutated immunoglobulin variable heavy-chain gene status: implications of minimal residual disease measurement with quantitative PCR. Blood 2004;104(8):2600-2
82.
Selenko N, Majdic O, Jager U, et al. Cross-priming of cytotoxic T cells promoted by apoptosis-inducing tumor cell reactive antibodies? J Clin Immunol 2002;22(3):124-30
83.
Gajewski JL, Phillips GL, Sobocinski KA, et al. Bone marrow transplants from HLA-identical siblings in advanced Hodgkin’s disease. J Clin Oncol 1996;14(2):572-8
84.
Peggs KS, Anderlini P, Sureda A. Allogeneic transplantation for Hodgkin lymphoma. Br J Haematol 2008;143(4):468-80
85.
86.
Peggs KS, Sureda A, Qian W, et al. UK and Spanish Collaborative Groups. Reduced-intensity conditioning for allogeneic haematopoietic stem cell transplantation in relapsed and refractory Hodgkin lymphoma: impact of alemtuzumab and donor lymphocyte infusions on long-term outcomes. Br J Haematol 2007;139(1):70-80
Delgado J, Pillai S, Benjamin R, et al. The effect of in vivo T cell depletion with alemtuzumab on reduced-intensity allogeneic hematopoietic cell transplantation for chronic lymphocytic leukemia. Biol Blood Marrow Transplant 2008;14(11):1288-97
94.
Buhmann R, Simoes B, Stanglmaier M, et al. Immunotherapy of recurrent B-cell malignancies after allo-SCT with Bi20 (FBTA05), a trifunctional anti-CD3 x anti-CD20 antibody and donor lymphocyte infusion. Bone Marrow Transplant 2009;43(5):383-97
95.
van de Donk NW, Kroger N, Hegenbart U, et al. Prognostic factors for donor lymphocyte infusions following non-myeloablative allogeneic stem cell transplantation in multiple myeloma. Bone Marrow Transplant 2006;37(12):1135-41
96.
Lokhorst HM, Wu K, Verdonck LF, et al. The occurrence of graft-versus-host disease is the major predictive factor for response to donor lymphocyte infusions in multiple myeloma. Blood 2004;103(11):4362-4
97.
Peggs KS, Mackinnon S, Williams CD, et al. Reduced-intensity transplantation with in vivo T-cell depletion and adjuvant dose-escalating donor lymphocyte infusions for chemotherapy-sensitive myeloma: limited efficacy of graft-versus-tumor activity. Biol Blood Marrow Transplant 2003;9(4):257-65
98.
Sun K, Welniak LA, Panoskaltsis-Mortari A, et al. Inhibition of acute graft-versus-host disease with retention of graft-versus-tumor effects by the proteasome inhibitor bortezomib. Proc Natl Acad Sci USA 2004;101(21):8120-5, Erratum in: Proc Natl Acad Sci USA. 2004;101(34):12777
99.
Lioznov M, El-Cheikh J Jr, Hoffmann F, et al. Lenalidomide as salvage therapy after allo-SCT for multiple myeloma is effective and leads to an increase of activated NK (NKp44(+)) and T (HLA-DR(+)) cells. Bone Marrow Transplant 2010;45(2):349-53
91.
Marks DI, Lush R, Cavenagh J, et al. The toxicity and efficacy of donor lymphocyte infusions given after reduced-intensity conditioning allogeneic stem cell transplantation. Blood 2002;100(9):3108-14
92.
Ritgen M, Bottcher S, Stilgenbauer S, et al.; German CLL Study Group. Quantitative MRD monitoring identifies distinct GVL response patterns after allogeneic stem cell transplantation for chronic lymphocytic leukemia: results from the GCLLSG CLL3X trial. Leukemia 2008;22(7):1377-86
100. Levine JE, Barrett AJ, Zhang MJ, et al. Donor leukocyte infusions to treat hematologic malignancy relapse following allo-SCT in a pediatric population. Bone Marrow Transplant 2008;42(3):201-5
Porter DL, Levine BL, Bunin N, et al. A phase 1 trial of donor lymphocyte infusions expanded and activated ex vivo via CD3/CD28 costimulation. Blood 2006;107(4):1325-31
101. Davies SM, Ramsay NK, Klein JP, et al. Comparison of preparative regimens in transplants for children with acute lymphoblastic leukemia. J Clin Oncol 2000;18:340-7
93.
Expert Opin. Biol. Ther. (2011) 11(4)
485
Donor lymphocyte infusion following allogeneic hematopoietic stem cell transplantation
102. Yoshimi A, Bader P, Matthes-Martin S, et al. European Working Group of MDS in Childhood (EWOG-MDS). Donor leukocyte infusion after hematopoietic stem cell transplantation in patients with juvenile myelomonocytic leukemia. Leukemia 2005;19:971-7
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
103. Ladon D, Pieczonka A, Jolkowska J, et al. Molecular follow up of donor lymphocyte infusion in CML children after allogeneic bone marrow transplantation. Appl Genet 2001;42:547-52 104. Bader P, Klingebiel T, Schaudt A, et al. Prevention of relapse in pediatric patients with acute leukemias and MDS after allogeneic SCT by early immunotherapy initiated on the basis of increasing mixed chimerism: a single center experience of 12 children. Leukemia 1999;13:2079-86 105. Bader P, Kreyenberg H, Hoelle W, et al. Increasing mixed chimerism is an important prognostic factor for unfavourable outcome in children with acute lymphoblastic leukemia after allogeneic stem-cell transplantation: possible role for pre-emptive immunotherapy? J Clin Oncol 2004;22:1696-705 106. Hoelle W, Beck JF, Dueckers G, et al. Clinical relevance of serial quantitative analysis of hematopoietic chimerism after allogeneic stem cell transplantation in children for severe aplastic anaemia. Bone Marrow Transplant 2004;33:219-23 107. Frugnoli I, Cappelli B, Chiesa R, et al. Escalating doses of donor lymphocytes for incipient graft rejection following SCT for thalassemia. Bone Marrow Transplant 2010;45(6):1047-51 108. Rezvani K, Rahemtulla A, Cummins M, et al. Response of CML to donor lymphocyte infusions (DLI): factors influencing the effective cell dose and implications for the definition of refractoriness. Blood 2000;96(S1):195 109. Bertz H, Burger JA, Kunzmann R, et al. Adoptive immunotherapy for relapsed multiple myeloma after allogeneic bone marrow transplantation (BMT): evidence for a graft-versus-myeloma effect. Leukemia 1997;11(2):281-3 110. Marks DI, Lush R, Cavenagh J, et al. The toxicity and efficacy of donor lymphocyte infusions given after reduced-intensity conditioning allogeneic
486
stem cell transplantation. Blood 2002;100(9):3108-14 111. Kernan NA, Collins NH, Juliano L, et al. Clonable T lymphocytes in T cell-depleted bone marrow transplants correlate with development of graft-v-host disease. Blood 1986;68:770-3 112. Dazzi F, Fozza C. Disease relapse after haematopoietic stem cell transplantation: risk factors and treatment. Best Pract Res Clin Haematol 2007;20(2):311-27 113. Dazzi F, Szydlo RM, Craddock C, et al. Comparison of single dose and escalating-dose regimens of donor lymphocyte infusion for relapse after allografting for chronic myeloid leukemia. Blood 2000;95:67-71 114. van Rhee F, Lin F, Cullis JO, et al. Relapse of chronic myeloid leukemia after allogeneic bone marrow transplant: the case for giving donor leukocyte transfusions before the onset of hematologic relapse. Blood 1994;83:3377-83 115. Bonini C, Ferrari G, Verzeletti S, et al. HSV-TK gene transfer into donor lymphocytes for control of allogeneic graft-versusleukemia. Science 1997;276(5319):1719-24 116. Verzeletti S, Bonini C, Marktel S, et al. Herpes simplex virus thymidine kinase gene transfer for controlled graft-versus-host disease and graft-versusleukemia: clinical follow-up and improved new vectors. Hum Gene Ther 1998;9(15):2243-51 117. Deschamps M, Mercier-Lethondal P, Certoux JM, et al. Deletions within the HSV-tk transgene in long-lasting circulating gene-modified T cells infused with a hematopoietic graft. Blood 2007;110:3842-52 118. Berger C, Flowers ME, Warren EH, et al. Analysis of transgene-specific immune responses that limit the in vivo persistence of adoptively transferred HSV-TK-modified donor T cells after allogeneic hematopoietic cell transplantation. Blood 2006;107:2294-302 119. Thomis DC, Marktel S, Bonini C, et al. A Fas-based suicide switch in human T cells for the treatment of graft-versus host disease. Blood 2001;97(5):1249-57 120. Straathof KC, Pule MA, Yotnda P, et al. An inducible caspase 9 safety switch for T-cell therapy. Blood 2005;105:4247-54
Expert Opin. Biol. Ther. (2011) 11(4)
121. Hartwig UF, Nonn M, Khan S, et al. Depletion of alloreactive donor T lymphocytes by CD95-mediated activation-induced cell death retains antileukemic, antiviral, and immunoregulatory T cell immunity. Biol BloodMarrow Transplant 2008;14:99-109 122. Wehler TC, Nonn M, Brandt B, et al. Targeting the activation-induced antigen CD137 can selectively deplete alloreactive T cells from antileukemic and antitumor donor T-cell lines. Blood 2007;109:365-73 123. Kamiryo Y, Eto M, Yamada H, et al. Donor CD4 T cells are critical in allogeneic stem cell transplantation against murine solid tumor. Cancer Res 2009;69(12):5151-8 124. Champlin R, Jansen J, Ho W, et al. Retention of graft-versus-leukemia using selective depletion of CD8-positive T lymphocytes for prevention of graft-versus-host disease following bone marrow transplantation for chronic myelogenous leukemia. Transplant Proc 1991;23:1695-6 125. Giralt S, Hester J, Huh Y, et al. CD8-depleted donor lymphocyte infusion as treatment for relapsed chronic myelogenous leukemia after allogeneic bone marrow transplantation. Blood 1995;86:4337-43 126. Alyea EP, Soiffer RJ, Canning C, et al. Toxicity and effcacy of defined doses of CD4(+) donor lymphocytes for treatment of relapse after allogeneic bone marrow transplant. Blood 1998;91:3671-80 127. Orti G, Lowdell M, Fielding A, et al. Phase I study of high-stringency CD8 depletion of donor leukocyte infusions after allogeneic hematopoietic stem cell transplantation. Transplantation 2009;88(11):1312-18 128. Aqui NA, June CH. Post-transplant adoptive T-cell immunotherapy. Best Pract Res Clin Haematol 2008;21(3):503-19 129. Morgan RA, Dudley ME, Wunderlich JR, et al. Cancer regression in patients after transfer of genetically engineered lymphocytes. Science 2006;314:126-9 130. Maher J, Brentjens RJ, Gunset G, et al. Human T-lymphocyte cytotoxicity and proliferation directed by a single chimeric TCRzeta/CD28 receptor. Nat Biotechnol 2002;20:70-5
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Roddie & Peggs
131.
Kershaw MH, Westwood JA, Parker LL, et al. A phase I study on adoptive immunotherapy using gene-modified T cells for ovarian cancer. Clin Cancer Res 2006;12:6106-15
132.
Lamers CH, Sleijfer S, Vulto AG, et al. Treatment of metastatic renal cell carcinoma with autologous T-lymphocytes genetically retargeted against carbonic anhydrase IX: first clinical experience. J Clin Oncol 2006;24:20-2
133.
Pule MA, Savoldo B, Myers GD, et al. Virus-specificTcells engineered to coexpress tumor-specific receptors: persistence and antitumor activity in individuals with neuroblastoma. Nat Med 2008;14:1264-70
Affiliation Claire Roddie1 FRCPath MRCP MBChB BSc (Hons) & Karl S Peggs†2 MD FRCPath MRCP MBChB † Author for correspondence 1 Clinical Research Fellow, UCL Cancer Institute, Department of Haematology, Paul O’Gorman Building, 72 Huntley Street, London, WC1E 6BT, UK 2 Senior Lecturer, UCL Cancer Institute, Department of Haematology, Paul O’Gorman Building, 72 Huntley Street, London, WC1E 6BT, UK Tel: +0207 679 6236; Fax: +0207 679 6222; E-mail:
[email protected]
Expert Opin. Biol. Ther. (2011) 11(4)
487
Review
Autologous cell therapy for cardiac repair Darryl R Davis* & Duncan J Stewart† 1.
Introduction
2.
Clinical trials with adult
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
autologous cell therapy 3.
Enhanced cell strategies
4.
Host effects
5.
Conclusions
6.
Expert opinion
*University of Ottawa Heart Institute, Ottawa, Ontario, Canada; and †Ottawa Hospital Research Institute, Ottawa, Ontario, Canada
Introduction: While new therapies have improved the prognosis of patients post acute myocardial infarction, many patients still suffer from irreversible damage and live with the debilitating consequences. However, with the advent of stem cell-based therapies, future treatments may enable us to harness the potential of autologous stem cells to prevent and even reverse heart damage. Areas covered: We outline the results of the early clinical trials using autologous cell therapy and highlight the hurdles and limitations that still need to be addressed. We also discuss new approaches that hold promise for developing the next generations of autologous cell therapy by exploring strategies to enhance their regenerative activity using biomaterials, genetic modification, optimal cell types and small molecule preconditioning. Expert opinion: Autologous cell therapy may be on the cusp of being widely adopted for the treatment of patients with large areas of myocardial damage. Techniques to enhance the activity and retention of autologous cell products may represent the next generation of this therapy. Keywords: cardiac stem cells, cell therapy, endothelial progenitor cells, mesenchymal stem cells, myocardial infarction, small molecules, somatic gene transfer Expert Opin. Biol. Ther. (2011) 11(4):489-508
1.
Introduction
Although there has been remarkable progress in the last several decades in the development of new pharmaceutical and interventional therapies for cardiac and vascular diseases, many patients still suffer from irreversible damage and live with the debilitating consequences, in particular heart failure (HF), which is emerging as a major challenge for health care systems worldwide. As the contractile function of the heart declines, so too does its ability to respond to medical therapies, leading eventually to the need for cardiac replacement with all the attendant costs and serious health consequences of living with a transplanted heart or mechanical device. With the explosion of new knowledge about the role of stem cells in tissue repair and regeneration, we are on the cusp of a new era in medicine, one in which we will be able to harness the potential of endogenous regenerative mechanisms to prevent and even reverse organ damage. The discovery of stem and progenitor cells, and the increasing understanding of their role not only in early embryonic development, but also organ homeostasis and repair throughout adult life, has led to the burgeoning field of cell therapy, which is being explored for a wide variety of medical problems, from spinal injury to diabetes. This review outlines the progress of first generation autologous stem cells and the hurdles that confront their ready translation to the clinical setting. The influence of the host factors that influence engraftment and regenerative potency will be examined. Techniques to enhance regeneration using biomaterials, genetic modification, optimal cell types and small-molecule preconditioning are being evaluated which offer tremendous promise to enhance the healing of injured myocardium. This review also looks forward at the future of autologous cell therapy for cardiac repair, 10.1517/14712598.2011.556615 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
489
Autologous cell therapy for cardiac repair
Article highlights. .
.
.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
.
Clinical trials demonstrate that even quite simple approaches to administer autologous cells yield positive results. While truly totipotent cells, such as embryonic stem cells and induced pluripotent stem cells, may have the greatest capacity to regenerate organs; safety concerns will significantly delay translation to clinical use. A host of bone-marrow-derived cell products have been developed for clinical use; however the main mechanism of benefit appears to be paracrine-mediated. Resident cardiac stem cells show great promise for autologous cardiac repair strategies as they posses both paracrine-mediated myocardial salvage and the capacity to differentiate into working myocardium. The next generation of autologous therapy will be engineered using combination cell therapy, improved mechanical retention, modified culture environments and somatic gene transfer.
This box summarizes key points contained in the article.
and assesses attempt to predict what will be required in order for this innovative strategy to be adopted widely in the management of patients that sustain large myocardial infarctions despite all modern reperfusion therapies. 2. Clinical trials with adult autologous cell therapy
Despite our incomplete understanding of the biology of stem and progenitor cells, a number of clinical trials have already been performed; many of which have rigorous design including randomization and blinding. The results of stem cell administration for the treatment of cardiac disease appear to show real promise. Even quite simple approaches have already yielded positive results for enhancing cardiac repair in the post myocardial infarction (MI) setting (Table 1) [1-30]. In most cases these trials have used unselected autologous bone marrow mononuclear cells (BMC), since these are routinely used for bone marrow transplantation in all large tertiary care centers. Moreover, several recent systematic reviews and metaanalyses have been performed of all randomized studies, totaling over 900 patients, and these support a highly significant, albeit modest, improvement in global left ventricular ejection fraction (LVEF), infarct area and end systolic volumes postMI [31-33]. Although it was initially thought that transdifferentiation of stem and progenitor cells into new vascular or cardiac tissue would be the predominant mechanism of restoration of structure and function, the results from both clinical and preclinical studies using BMCs indicate that the mechanism of this benefit is probably not due to direct cardiomyocyte replacement but rather to a variety of other effects which modulate cardiac repair [34,35]. These include enhancing neovascularization of the ischemic zone [36], 490
paracrine stimulation of endogenous cellular repair mechanisms [37,38] and modulation of immune responses with reduced fibrosis and scarring [39]. In the early post-MI setting, the delivery of BMCs may be effective in attenuating the initial inflammatory response, reducing scar formation and promoting more adaptive healing. However, in the setting of extensive damage and chronic organ failure (i.e., HF), it is likely that regeneration of new contractile myocardial elements either directly or indirectly will be required to improve contractility. In this case, it may be necessary to harness cells that have the capacity to transdifferentiate to cardiomyocytes, be they resident within the myocardium or harvested from other tissues, and both strategies are discussed below. 3.
Enhanced cell strategies
Optimal autologous cell types Although truly totipotent stem cells, such as embryonic stem cells or inducible pluripotent cells, demonstrate the greatest capacity for organ regeneration; inherent concerns about safety will probably impede their use in clinical cell therapies for the foreseeable future. Another class of bone marrow ‘stem’ cell is represented by the mesenchymal stromal/ stem cells (MSCs). These cells have been studied extensively over the last several decades and in addition to some capacity to differentiate to cardiac and vascular cell lineages [40], have important modulatory effects upon the host myocardium. Recently, tissue-resident stem cells have been identified in a number of adult organs including the brain [41] and heart [42]. As outlined below, these cells appear to have specific abilities to regenerate the cells of these tissues. 3.1
Autologous blood and bone marrow cells Bone marrow contains a variety of stem and progenitor cells including mesenchymal stem cells (MSCs) and endothelial progenitor cells (EPCs) [31,43]. However, progenitor or stem cells represent a very small proportion of bone marrow or circulating mononuclear cells (MNCs) (i.e., < 0.05%). Nonetheless, unselected MNCs appear to be able to improve cardiac function after delivery into the infarct-related artery, albeit modestly as reviewed above. While these benefits still need to be validated in a large, pivotal Phase III trial, it is also apparent that there may be tremendous opportunity to refine current cell therapy strategies to enhance efficacy by the selection or enrichment of cell populations with greater therapeutic activities, and the enhancement of regenerative activity of a given cell population. Most of the published clinical studies for cardiac cell therapy have used bone marrow or blood derived MNCs isolated by Ficoll gradient centrifugation, drawing on the established clinical expertise and infrastructure developed to support bone marrow transplantation as a routine clinical procedure in most tertiary hospital settings [1,2,21,22]. While expediting the road for translation to clinical application, it is highly improbable that unselected MNCs will ultimately prove to 3.1.1
Expert Opin. Biol. Ther. (2011) 11(4)
Davis & Stewart
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Table 1. Clinical trials of autologous bone marrow stem cells in the post myocardial infarction setting. Study
Design
N*
Cell type
Delivery
Days post MI
Outcome
Non-randomized trials Strauer et al. [1]
Primarily safety
10
BM-MNCs
IC
8±2
TOPCARE-AMI [2,3]
Primarily safety
59
IC
4.9 ± 1.5
Fernandez-Aviles et al. [4] Bartunek et al. [5]
Primarily safety Primarily safety
20 19
IC IC
13.5 ± 15.5 11.6 ± 1.4
BALANCE [6]
Efficacy
62
BM-MNCs, PB-EPCs BM-MNCs CD133+, BM-MNCs BM-MNCs
IC
7±2
STAR-heart [7]
Efficacy
191
BM-MNCs
IC
3102 ± 1168
Stroke volume"; infarct size#; wall motion"; perfusion" EF"; remodelling#; infarct size#; perfusion" EF"; ESV#; wall motion" EF"; perfusion"; myocardial viability" EF"; stroke volume index; mortality benefit EF"; stroke volume index; mortality benefit
Randomized controlled trials (RCTs) Randomized Chen et al. [8] Randomized Ge et al. [9] Randomized Ruan et al. [10] Randomized Huang et al. [11] Randomized Yao et al. [14]
34 10 9 20 24
BM-MSCs BM-MNCs BM-MNCs BM-MNCs BM-MNCs
IC IC IC IC IC
18.8 ± 0.5 <1 <1 <1 13 ± 8
Penicka et al. [15] BOOST [13,17,18] Janssen et al. [19,20]
17 30 33
BM-MNCs BM-MNCs BM-MNCs
IC IC IC
9 4.8 ± 1.3 <1
REPAIR-AMI [21-23]
Randomized Randomized Randomized, double blind Randomized
102
BM-MNCs
IC
4.3 ± 1.3
Meluzin et al. [16,25]
Randomized
44
BM-MNCs
IC
6.8 ± 0.3
ASTAMI [24,26] Suarez de Lezo et al. [27] FINCELL [28] REGENT [29]
Randomized Randomized Randomized Randomized
6.0 ± 1.3 7±2 <4 7
Randomized
BM-MNCs BM-MNCs BM-MNCs BM-MNCs CD34+ BM-MNcs BM-MNCs
IC IC IC IC
Traverse et al. [30]
50 10 39 80 80 40
IC
4.5
EF" EF"; perfusion" EF"; wall motion" EF"; infarct size# No change in systolic function; improved diastolic function No change in EF EF"(6 months only) No change in EF; infarct size#; regional systolic function" EF"; greater benefit in < 49% EF and > 5 days after MI EF" in a dose-dependent manner No benefit EF" EF" No benefit No change in EF; Improved LV end diastolic volume
*N: Number of patients receiving cells. BM-MNCs: Bone marrow-mononuclear cells; EF: Ejection fraction; ESV: End-systolic volume; IC: Intracoronary; LV: Left ventricle; MI: Myocardial infarction; PB: Peripheral blood.
be the most effective cell therapy product for cardiac and vascular repair. Although it is attractive to use antigenic cell surface markers to select subpopulations of cells with greater regenerative capacity, at this time there is no agreement as to what markers can be used to enrich the true ‘EPC’ fraction [5,44-48]. In addition, cell selection based on surface markers introduces major challenges in scaling up to manufacture sufficient cell numbers for effective clinical treatments given the very small proportion of MNCs which meet the most common ‘EPC’ definitions [44-46]. Indeed, the use of selected MNCs subsets (i.e., CD133+, CD34+) has not yielded substantially greater benefit compared with unselected MNCs in the limited number of clinical studies that have used this strategy [5,47,48]. An alternate strategy is to select regenerative cells from the MNC fraction based on their functional characteristics, in a manner analogous to the commonly used selection procedure for deriving mesenchymal stem cells or dendritic
cells. Specifically, the ability of subpopulations of MNCs to differentiate in attached culture in the presence of specific extracellular matrix components and growth and differentiation factors. There has been a wealth of experience in characterizing the regenerative activities of cultured-derived ‘EPCs’ in a variety of in vitro and in vivo models, which is briefly summarized below [49,50]. Culture modified endothelial progenitor cells (EPCs) So-called ‘early growth EPCs’ or ‘circulating angiogenic cells’ appear within 2 -- 3 days of MNC selection culture in the presence of endothelial growth factors (i.e., VEGF, IGF, EGF, etc.) [49,51]. While they express a number of endothelial characteristics (CD31, VEGFR2, tunica interna endothelial cell kinase (Tie2), eNOS, lectin binding and dioctadecyl3,3,3¢,3¢-tetramethylindo-carbocynanine perchlorate-labelled low-density lipoprotein (DI-LDL) uptake), they still retain monocyte (CD14) and leukocyte (CD45) markers. However, 3.1.1.1
Expert Opin. Biol. Ther. (2011) 11(4)
491
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Autologous cell therapy for cardiac repair
there is abundant in vitro and in vivo evidence that this cell population is highly angiogenic with the ability of these ‘early growth’ EPCs to restore perfusion in the ischemic hindlimb or improve cardiac function post MI having been soundly established [2,31,49,52]. Therapeutically, transplanted EPCs can provide stimulatory cytokines [53,54] and induce humoral effects that are sustained by the host tissues [55]. Therefore, these culture-modified MNCs may result in one or more of the following: i) the cells may modify the local milieu in order to increase recruitment of other circulating progenitor cells; ii) the injected cells may themselves be involved in producing growth factors and recruiting additional cellular and paracellular elements; or iii) the cells may differentiate and directly partake in new blood vessel growth. All these processes have been observed experimentally, however, it is believed that vascularization from paracrine/humoral factors and secondary recruitment of host stem/progenitor cells is probably the main mechanism leading to functional improvement [3,55-60]. Thus these cells have a number of properties that are quite favorable for their use in clinical therapy with extensive preclinical evidence demonstrating potent angiogenic activity in a number of relevant models [61-63] as well as ease of scalability of manufacturing for human studies. Late outgrowth EPCs More prolonged culture of blood or bone marrow-derived MNCs (i.e., > 1 -- 2 weeks) under the conditions described above gives rise to colony-like clusters or aggregates of highly proliferative cells, which then rapidly overgrow the dishes to produce uniform ‘cobblestone’ cultures of so-called ‘late outgrowth EPCs’ [31,64]. Unlike the early-growth EPCs described above, these cells have lost all leukocyte determinants and exhibit a robust endothelial phenotype. Indeed, apart from their markedly increased proliferative potential and possibly an increase in the expression of some markers of EC activation (i.e., E-selectin), the late growth cells are nearly indistinguishable from mature EC cultures. Thus, this population may be of great interest for regenerative medicine applications; however, to date they have been far less studied in relevant preclinical models. It has been suggested that they may act synergistically with early outgrowth EPCs, acting mainly to directly repair the vascular endothelium [65], however, they do not appear to share the same paracrine abilities of the ‘early growth’ cells, and in our experience, have not been effective in a head to head comparison of in vivo efficacy in the prevention of experimental pulmonary hypertension [66]. 3.1.1.2
Mesenchymal stromal (stem) cells (MSCs) MSCs represent an important population of bone marrowderived cells which have been studied extensively over several decades, and may offer certain advantages for specific regenerative medicine applications. MSCs are a type of nonhematopoietic, adult somatic stem cells that can be isolated from bone marrow [67] and extensively expanded in vitro [68]. In contrast to embryonic stem cells, which are capable 3.1.1.3
492
of differentiating into all cells of the body (i.e., are totipotent), MSCs have only limited in vivo differentiation and proliferation potential (i.e., are multipotent) [69]. However, they also exhibit important ‘immunomodulatory’ properties which probably contribute to their ability to reduce tissue damage and enhance repair [70]. Additionally, several recent studies have also shown that MSCs also stimulate the proliferation of other progenitor cell populations within target organs to promote endogenous repair [71-73]. Clinical trials utilizing MSCs for acute MI [8] or for graft-versus-host disease [74] have shown various levels of success, even though these results need to be confirmed in large and rigorously designed studies. Like EPCs, MSCs can be isolated directly from a patient’s own bone marrow, thereby avoiding complications involving the immune rejection of allogeneic tissue [75]; however, there is considerable evidence that they may be ‘immuneprivileged’, potentially permitting their use in allo-transplantation (i.e., cells from another individual) without the need for immunosuppressive therapy [76-80]. The use of allogeneic cells would make it feasible for MSCs to be delivered as an ‘off-the-shelf’ product for the treatment of acute (such as in the case of acute lung injury/acute respiratory distress syndrome (ALI/ARDS) patients) and chronic disorders. This unique feature makes MSC-based therapies very attractive for the treatment of acute organ injuries. While the clinical experience to date has demonstrated that autologous bone marrow or circulating stem/progenitor cell delivery is generally well tolerated, some preclinical reports have shown that intramyocardial injection of MSCs may differentiate into encapsulated structures with calcifications and ossifications [81,82]. This in vivo plasticity replicates in vitro observations that this highly proliferative cell source may permit phenotypic drift and the posibility of cancerous transformation [83]. Despite these findings, preclinical data has generally been encouraging with no signal reported of adverse events. But these findings may be a harbinger of the challenges that will confront the use of more pluripotent stem cells. Resident cardiac stem cells (CSCs) Ten years ago, prevailing dogma posited that the adult mammalian heart was incapable of self-regeneration after injury [84]. Adult cardiomyocytes were considered terminally-differentiated cells that have exited the cell cycle. This dogma was challenged by the discovery that the heart contains a reservoir of small cells that stain for stem cell markers, propagate in vitro and develop phenotypic features of heart cells after differentiation [85-92]. Retrospective studies using 14C birth dating of cells or chemotherapeutically labeled adult cells have since confirmed that cardiomyocyte turnover occurs in the adult human heart [93,94]. Given that CSCs are autologous and capable of forming all cardiac lineages, they represent logical candidates for cell therapy. Numerous markers have been proposed to identify resident CSCs for isolation and ex vivo amplification (Table 2) [85-88,95-100]. The first paper describing isolation of resident cardiac stem cells 3.1.2
Expert Opin. Biol. Ther. (2011) 11(4)
Davis & Stewart
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Table 2. Cell surface marker expression and stem cell characteristics of cardiac progenitor cells. Marker
Species
Source
Surface expression
Clonogenicity
Differentiation
Function benefit
c-kit [85]
Mouse/rat
Whole heart
Yes
IHC/coupling
Yes
sca-1 [86]
Mouse
Whole heart
Yes
IHC
Yes
abcg2 [88,95,96]
Mouse
Whole heart
--
IHC/coupling
--
SSEA-1 [87]
Rat
Ventricular
--
IHC
Yes
c-kit [97]
Human
Biopsy
Yes
IHC/coupling
Yes
Sca- [98-100]
Human
Atrial appendage
MDR1+, Sca-1+, CD45-, CD34-, VWF-, CD31-, vimentin-, linABCG2±, CD31±, CD38±, c-Kit-, Flk-1-, CD34-, CD45-, Flt-1Sca-1+, CD31+ c-Kit±, CD34±, CD45ABCG2±, c-Kit±, Sca-1±, Flk-1±, CD31-, CD45-, VE-cadhedrin-, SSEA4KDR-, CD45-, lin-, CD34-, CD133CD105+, CD31+ c-Kit±, CD45-, CD14-, CD34-, CD133-
Yes
IHC
Yes
ABCG2: ATP-binding cassette protein G2; Flk-1: Fetal liver kinase-1; Flt: Fms-like transcript; IHC: Immunochistochemistry; KDR: Kinase insert domain receptor; MDR-1: Multidrug resistance protein 1; Sca-1: Surface marker antigen-1; SSEA: Stage-specific embryonic antigen; VWF: Von Willebrand factor.
built upon previous work using isolated skeletal myoblasts [42]. In this work, the capacity of discreet subpopulations within myocardium to actively transport Hoechst dye defines a side population of cells on flow cytometry proved to be capable of multipotentiality [42,95] and myocardial repair [101]. This side population phenotype was later ascribed to the surface expression of the ATP-binding cassette protein G2 (ABCG2, a membrane transport protein mediating multi-drug resistance; CDw338) [95]. In humans, ABCG2 expression co-localizes with CD31+ cells, suggestive of a potential endothelial fate [102]. The most widely studied CSC marker, c-Kit, was first identified in histology sections from sex-mismatched transplanted adult human hearts [103]. This marker co-express with other stem cell markers (multidrug resistance protein 1 (MDR1) and surface marker antigen (Sca-1)) and has been shown to improve post MI function with differentiation into numerous cardiac lineages (cardiomyocytes, endothelial cells and smooth muscle cells) [104-107]. A Phase I clinical trial using antigenically purified sub-populations of c-Kit has begun to assess the feasibility and safety of intra-coronarydelivered CSC in chronic HF patients undergoing cardiac surgery [108]. Of note, studies using Sca-1 in mice have isolated a small CSC population capable of improving post-MI cardiac function [86]. These mouse cells partially expressed markers of endothelial/BMC origin (ABCG2, CD31, CD38) while not expressing markers of other stem cell (c-Kit, fetal liver kinase 1 (Flk-1)) or hematological (CD34, CD45, fms-like tyrosine kinase (Flt-1)) lineages. Several studies have explored the capacity of Sca-1+ cells isolated from human samples to improve cardiac function [98-100]. While the human epitope recognized by Sca-1 is not known, these studies found a homogenous cell population can be
isolated that partially expresses c-Kit and is capable of in vitro cardiac differentiation. Recently it has been shown that distinct subpopulations of CSCs can be isolated directly from cardiac tissue [109]. This technique simplifies culture methods by focusing on the primary cellular outgrowth product from cardiac samples, without recourse to antigenic sub-selection [87,97,110]. When samples of minced cardiac tissue are cultured, a lawn of flat cells emigrates spontaneously from the plated cardiac tissue. Within that lawn, clusters of CSCs emerge and proliferate. Using mild enzymatic dissociation, loosely-adherent cells surrounding the explant (termed cardiac outgrowth) can be serially harvested. This outgrowth contains complimentary sub-populations of cells expressing embryonic (stage-specific embryonic antigen 1 (SSEA-1)), stem cell-related (c-Kit, ABCG2), endothelial (CD34, CD31) and mesenchymal (CD90) antigens. Transitioning this direct outgrowth though three dimensional sphere culture has been shown to enhance both CSC content and potency [111]. Further expansion through monolayer culture provides a single cell product termed Cardiosphere Derived Cells (CDCs) that have been demonstrated to be clonogenic, multipotent and capable of self-renewal [112-114]. Recent studies of CDCs have demonstrated that in vivo these cells secrete VEGF, hepatocyte growth factor (HGF) and IGF-1 [115]. It has been estimated that direct cardiomyocyte and vascular transdifferentiation represents 20 -- 50% of the overall increase in cardiac tissue, suggesting that, like EPCs, indirect effects of CDCs on tissue preservation and/or recruitment of endogenous progenitors significantly contribute to therapeutic outcomes [114,115] Studies have focused on validating culture techniques [116-118] for broad clinical translation beginning with a recently started Phase I clinical trial [112,119,120].
Expert Opin. Biol. Ther. (2011) 11(4)
493
Autologous cell therapy for cardiac repair
Other autologous cell types for cellular cardiomyoplasty
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
3.1.3
A host of other autologous cell types have been cultured from diverse extra-cardiac progenitor sources for cellular cardiomyoplasty. Early experience with skeletal myoblasts demonstrated the challenging nature of this approach as myoblast transplantation was plagued by ventricular tachyarrhythmias and sudden cardiac death [121,122]. The pathogenesis of these arrhythmias is poorly understood, but may be related to the fact that skeletal muscle cells, unlike heart cells, are electrically isolated by the absence of appropriate gap junctions [123]. Adipose stem cells represent a promising extra-cardiac stem cell source that can be retrieved in high number from either liposuction aspirates or subcutaneous adipose tissue fragments [124,125]. Although adipose tissue represents a heterologous cell population, culture selects for a relatively homogenous cell population, enriched for cells expressing a stromal phenotype (CD13, CD29, CD73, CD90, CD133) [126-129]. In vitro, these cells can be easily be expanded and, like MSCs, can modestly differentiate into cardiomyocyte [130-133] and endothelial [134-137] lineages. Consistent with the MSC experience, adipose stem cells improve cardiac function [133,138-140] with limited evidence for in vivo differentiation [138,140,141], suggesting that paracrine-mediated effects mediate the majority of the beneficial effects seen with this cell type [142,143]. Based on their ease of culture and benign profile, adipose stem cells are currently under investigation for patients with ischemic heart disease [144] and acute MI [145]. Combination cell therapy Despite evidence that stem cell injection improves myocardial function, studies consistently demonstrate modest overall benefit, possibly related to very limited long-term engraftment and retention after intra-myocardial injection [146-150]. This finding is not surprising given that cells are injected into a hostile inflammatory milieu with poor mechanical and vascular support. Several studies have investigated the effect of transplanting complimentary cell types on myocardial function [99,151-153], based on the premise that cells supporting the surrounding host tissue through paracrine secretion may increase the retention and proliferative capacity of cells capable of forming true contractile cells. Early attempts focused upon angiogenic progenitor cells and skeletal myoblasts [151,152]. These studies demonstrated that while both cell types reduce scar size and improve ventricular function, combination therapy was superior. While intriguing, this data is not translatable into clinical trials given the problems of association of myoblasts with ventricular tachy-arrhythmias and sudden cardiac death [121,122]. Recently, a group from the Netherlands has taken this concept forward using combination therapy derived from the adult heart (i.e., epicardium-derived cells (EPDCs) and cardiac progenitor cells (CPCs)) [99]. These investigators demonstrated that co-transplantation improved post infarct 3.1.4
494
function compared with single-cell transplantation which itself was superior to vehicle controls. These findings using mixed cardiac derived cell populations (CPCs and EPDCs) mirror studies showing that purified human CPCs (c-Kit+) and mesenchymal cells (CD90+) are capable of independently improving post myocardial infarct cardiac function in an immunodeficient mouse model [154]. These promising results suggest synergies exist between multiple cell types, providing new directions in cardiac cell therapy using existing first generation cell types. Pre-conditioning using small molecules or enhanced culture techniques
3.2
The concept of introducing exogenous factors to promote endogenous repair is well established with many studies demonstrating benefit through intra-myocardial injection of cytokines (i.e., G-CSF, HGF), growth factors (IGF-1) or signaling proteins (Akt, proto-oncogene serine/threonineprotein kinase Pim-1, cardiotrophin 1 (CT-1)) in cardiac damage models [155-160]. However the potential clinical utility of these strategies is limited by their reliance on extracardiac cell types, lack of specificity, transient retention and requisite transduction with potentially oncogenic factors. Moreover, clinical trials of bone marrow mobilization using G-CSF and related factors have been disappointing because of lack of specificity and harmful pro-inflammatory effects [161]. Importantly, this work has provided tantalizing clues about the triggers underlying stem cell cycling. Recent work by a host of labs indicates that targeted manipulation of the very same pathways underlying cardiogenesis or pluripotency may significantly increase the overall number and potency of resident stem cells [158,162-167]. These insights have been used to enhance cells ex vivo prior to injection by the addition of protein factors or small molecules, such as AVE-9488 [168], PPARa agonists [169,170], statins [171,172] and TGF-b [116,167]. Alternatively, variations in culture techniques have been shown to alter the phenotype of cells without recourse to direct stimulation of potentially oncogenic pathways or genetic reprogramming. Conceptually, this approach has merit as the environments to which stem cells are injected for myocardial repair are considerably harsher than traditional culture conditions or natural stem cell niches [173-176]. Brief exposure to hypoxic conditions (2% O2) has been successfully used in several studies to enhance the retention and survival of cells within the low oxygen tension of infarct regions (as low as 0.2% O2) [177-180]. Interestingly, ‘normal’ cell culture conditions may be toxic to progenitor and stem cells as non-physiologic hyperoxic conditions (ambient atmospheric conditions, 20% O2) can promote oxidative DNA damage, genomic instability and premature cell senescence [181-184]. It follows that that culture of stem cells within physiological normal oxygen conditions (5% O2) improves cellular yield and potency [185-187]. A recent study has taken this observation to its natural conclusion by demonstrating that cell culture
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Davis & Stewart
conditions recapitulating the natural three dimensional stem cell niche/embroid state enhance adhesion molecule expression, cardiac progenitor cell content, in vivo cellular retention and regenerative potency of ex vivo-cultured cardiac stem cells [188]. These findings highlight the capacity of altered culture conditions to provide a superior cell product with the potential for ready translation to clinical use. Such ‘preconditioning’ has the advantage of simplicity but potentially suffers from lack of durability of effects once the cells are transplanted in vivo. An alternative approach involves direct genetic manipulation (i.e., gene transfer) or incorporation of cells into scaffolding matrices. As outlined below, the suitability of the specific strategy will depend on the specific pathway that is being targeted and other considerations that effect the efficacy of the approach (i.e., duration of effect), as well as safety, regulatory and ethical considerations. Genetic modification of stem cells Direct genetic engineering of stem cells has also been used to improve transplanted cell survival (b-Akt [189,190], colony stimulating factoer 1 (CSF-1) [191], cellular repressor of E1A stimulated genes (CREA1) [192], heme oxygenase 1 (HO-1) [193,194], survivin [195], IGF-1 [196,197], heat shock protein 20 (HSP20) [198], B cell leukaemia/lymphoma related protein2 (Bcl-2) [199], Pim-1 [200]), electrical integration (connexin 43 (Cx43)) [123], differentiation (TNF-a), homing/ migration (CXCR4 [201], monocyte chemotactic protein-3 (MCP-3) [202], eNOS [203]) and vasculogenesis (HGF [204], hypoxia-inducibel factor 1 (HIF-1) [205], VGEF [206], stromal cell-derived factor 1 (SDF-1) [207], bFGF [208]). Comparing these different approaches is problematic given variations in cell type (fetal myocytes, embryonic stem cells, skeletal myoblasts, mesenchymal stem cells and several cell types derived from the bone marrow), models of therapeutic benefit (ischemic hind limb, carotid injury, MI and pressure overload HF) and strategies for gene transfer to cells (viral, plasmid). Additionally as outlined in Table 3, these approaches have been broadly applied to increase retention and survival of cells whose primary mechanism of action involves paracrine-mediated survival rather than true cardiomyogenesis [123,189-200,203,207-230]. Notably, a promising study incorporates overexpression of the pro-survival factor Pim-1 into cardiogenic CSCs [200]. This approach is based on the observation that the benefits of CSC therapy occur even at generally low levels of cell engraftment [150,231,232]; which is presumably due to transplanted cell loss mediated by apoptosis. Pim-1, by enhancing the pro-survival pathways, resulted in better engraftment and long-term retention of CSCs [158,233]. Another approach targets the paracine profile of early outgrowth EPCs [234]. Given that reduced eNOS expression and NO production have been strongly implicated not only in endothelial dysfunction [235,236] but also in EPC dysfunction [237,238], measures to enhance eNOS production 3.3
represent a logical target for cellular therapy. Preliminary studies have demonstrated that overexpression of eNOS enhances the regenerative capacity of EPCs isolated from patients with ischemic cardiomyopathy [168], suggesting a synergistic effect between cell and gene therapy. Accordingly, the first clinical trial of genetically modified autologous stem cells (EPCs) targets has been initiated to target patients with significant LV dysfunction (left ventricuar ejection fraction (LVEF) £ 45%) early after a reperfused ST elevation MI (< 30 days) [239]. In this study, consenting patients will undergo apheresis to collect peripheral blood MNCs and plasma before randomization to one of three arms, receiving either Plasma-Lyte A (placebo), autologous endotheliallike, culture modified MNCs (E-CMMs), or autologous E-CMMs transfected with eNOS, by coronary injection to the infarct-related artery. After 2 -- 3 days, the early outgrowth EPCs will be transfected with a human eNOS-pVAX plasmid complexed with linear polyethylenimine. The final cell product (20 million eNOS-transfected or non-transfected EPCs) will be injected directly into the infarct-related artery distal to an inflated angioplasty balloon. The primary outcome measures is a change in global LVEF from baseline to 6 month follow-up as determined by cardiac MRI. Secondary efficacy measures include changes from baseline to 6 month followup in i) regional wall motion, wall thickening, and infarct volume as determined by cardiac MRI; ii) echocardiographic assessment of LVEF and ventricular volumes; iii) time to clinical worsening (death, hospitalization for angina, reinfarction); and iv) quality of life (the SF-36 and the Duke Activity Status Index). This study represents the first clinical trial of next generation of cell products and promises to provide an effective adjunctive to revascularization therapies in patients with large myocardial infarcts. Scaffolds, matricellular materials and mechanical retention
3.4
Remarkable progress has been made in engineering biomaterials with the aim of creating multiscale 3D architectures to promote tissue repair and regeneration [240-247]. Through the application of nanotechnology in medicine, it has become possible to design smart, multi-functional structures composed of polymeric materials incorporating suitable architectures and bioactive signaling factors, which can interact with the surrounding tissue environment and facilitate tissue regeneration [248,249]. Combining bioactive materials with stem cells increases both the differentiation potential and the secretion of ‘trophic’ factors involved in tissue repair and survival. Disrupting the normal matrix interactions and placing attached cells in suspension for injection into tissue or blood is a profoundly disruptive process which triggers rapid programmed cell death, or anoikis [250]. Tissue engineering offers the possibility of developing biomaterials to provide appropriate surfaces for integrin binding to improve cell survival, resulting in better retention and function of transplanted cells [251,252]. The design of synthetic materials that mimic
Expert Opin. Biol. Ther. (2011) 11(4)
495
496
SDF-1 overexpression [213,214]
VGEF overexpression [215-219]
HGF overexpression [220] Angiopoetin overexpression
MSCs
MSCs
MSCs MSCs
Expert Opin. Biol. Ther. (2011) 11(4)
CSF-1 overexpression [191]
SDF-1 overexpression [196]
SkM
SkM
CXCR4 overexpression [225,230]
Promotes angiogenesis Promotes angiogenesis
Adenovirus Adenovirus
Mediates recruitment of inflammatory cells to areas of inflammation CXCR4 mediates the homing of MSCs to damaged regions of damaged tissue
Retrovirus
Retrovirus, adenovirus
Plasmid
Plasmid
GATA-4 enhances the MSC secretome - thereby increasing cell survival and promoting post-infarction cardiac angiogenesis Promotes angiogenesis and metalloproteinase expression potentially leading to enhanced extracellular remodeling Improved post-MI myocardial function in rats Effect mediated by enhanced recruitment of progenitor cells
Promotes angiogenesis
Adenovirus, plasmid
Retrovirus
Increases engraftment and enhances MSC survival
Plasmid
Permits effective extracellular coupling with surrounding myocytes Protects cells from hypoxia and promotes dangiogenesis Promotes angiogenesis
Mechanism/rationale for targeting
MSCs over-expressing CCR1 accumulate in areas of damaged myocardium and improve myocardial function Expression of CXCR4 increases homing of genetically modified cells to infarct regions
Enhanced stem and progenitor cell migration to the heart providing improved post-MI angiogenesis and function recovery
Enhanced post-MI engraftment and function without arrhythmias noted
Enhanced post-MI engraftment and function without arrhythmias noted Enhanced vascularization in ischemic hind limb and corneal models Enhanced vascularization in ischemic hind limb model Improved post-MI myocardial function by preserving reversibly damaged myocytes and recruiting CSC-like cells Improves neo-vascularization and function in models of myocardial infarction, ischemic hind limb and pressure overload HF Improved post-MI function Co-expression with Akt reduces apoptosis and proves greater functional improvement thank either factor alone Enhanced hypoxia resistance, angiogenesis and functional recovery after myocardial infarction
Result
BAD: BCL2-associated agonist of cell death; Bax: Bcl2-associated X protein; Bcl-2: B-cell leukamia/lymphoma-associated protein 2; CPCs: Cardiac progenitor cells; CREG1: Cellular repressor of E1A-stimulated genes; CSCs: Cardiac stem cells; Cx43: Connexin 43; EPCs: Endothelial progenitor cells; FGF: Fibroblast growth factor; GATA-4: Gata binding proten 4; HF: Heart failure; HGF: Hepatocyte growth factor; HIF-1: Hypoxia-inducible factor; HO-1: Heme oxygenase 1; HSP20: Heat shock protein 20 kDA; hTERT: Human telomerase reverse transcriptase; MDR1: Multidrug resistance protein-1; MI: Myocardial infarction; MSCs: Mesenchymal stem cells; pim-1: Proto-oncogene serine/threonine-protein kinase; SDF-1: Stromal-cell-derived factor-1; SkM: Skeletal myoblasts.
MSCs
Enhanced retention MSCs CCR1 overexpression [224]
GATA-4 overexpression [223]
MSCs
[221,222]
VEGF overexpression [212]
EPCs
Adenovirus
Adenovirus
HIF-1 overexpression [210,211]
EPCs
Gene transfer method Plasmid
Genetic modification
Enhanced integration SkM Cx-43 overexpression [123,209]
Cell type
Table 3. Genetic modification of autologous stem cells.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Autologous cell therapy for cardiac repair
Expert Opin. Biol. Ther. (2011) 11(4)
Akt overexpression [189]
CREG1 overexpression [192]
HO-1 overexpression [193,194]
HSP20 overexpression [198]
IGF-1 overexpression [197]
Survivin overexpression [195]
MSCs
MSCs
MSCs
MSCs
MSCs
MSCs
Inhibits oligomerization of pro-apoptotic proteins Bax/Bak and subsequent membrane permealization Downstream mediator of AKT activation preventing apoptosis Inhibits apoptosis by inactivating the pro-apoptotic protein BAD and by increasing transcription of pro-survival genes (via activated NF-kB) Inhibits of apoptosis by activating Akt and degrading p53 Involved in the oxidative cleavage of heme and protects against apoptosis Restores function of pro-survival/angiogenic cytokines (Akt, FGF-2, IGF-1 and VEGF) Activates AKT pathway and inhibits apoptosis
Inhibits apoptosis by inhibiting Bax/Fas actions and possibly by inhibiting caspase activation
Adenovirus
Adenovirus
Lentivirus
Adenovirus
Adenovirus
Adenovirus
Retrovirus
Lentivirus
Retrovirus
Maintains telomerase activity and replication of telomeric DNA; permitting the indefinite division of cells Regulates endothelial function and protects against apoptosis
Mechanism/rationale for targeting
Adenovirus
Gene transfer method
Increased MSC survival and enhanced post MI function through increased secretion of growth factors (VEGF, FGF-2 and IGF-1) Enhances cell survival and recruitment of ckit+, MDR1+, CD31+ and CD34+ cells into the infarcted rat heart Enhanced cell survival, paracrine secretion (VEGF) and post-MI function
Protects against apoptosis, enhances cell survival and upregulates VEGF secretion
Enhanced post-MI cellular engraftment, persistence and functional improvement in mice Improved post-MI myocardial function in rats Effect mediated by paracrine effects [227]
Enhanced mitogenic activity, migratory activity, and cell survival using ischemic hind limb model eNOS enhanced endothelial reconstitution in a model of carotid injury while HO-1 had no effect Increased graft survival in ischemic rat myocardium
Result
BAD: BCL2-associated agonist of cell death; Bax: Bcl2-associated X protein; Bcl-2: B-cell leukamia/lymphoma-associated protein 2; CPCs: Cardiac progenitor cells; CREG1: Cellular repressor of E1A-stimulated genes; CSCs: Cardiac stem cells; Cx43: Connexin 43; EPCs: Endothelial progenitor cells; FGF: Fibroblast growth factor; GATA-4: Gata binding proten 4; HF: Heart failure; HGF: Hepatocyte growth factor; HIF-1: Hypoxia-inducible factor; HO-1: Heme oxygenase 1; HSP20: Heat shock protein 20 kDA; hTERT: Human telomerase reverse transcriptase; MDR1: Multidrug resistance protein-1; MI: Myocardial infarction; MSCs: Mesenchymal stem cells; pim-1: Proto-oncogene serine/threonine-protein kinase; SDF-1: Stromal-cell-derived factor-1; SkM: Skeletal myoblasts.
PIM-1 overexpression [200]
Bcl-2 overexpression [199]
Cardiomyoblasts (H9c2)
CPCs
eNOS and HO-1 overexpression [203]
hTERT overexpression [226]
Enhanced survival EPCs
EPCs
Genetic modification
Cell type
Table 3. Genetic modification of autologous stem cells (continued).
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Davis & Stewart
497
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Autologous cell therapy for cardiac repair
natural stem cell micro- and macroenvironments environments (i.e., niches) may potentially be a powerful tool both to understand and control stem cell function. Research in this field is still very limited, and artificial niches for stem cells have been developed mainly using synthetic proteins with applications mainly focused on neural stem cells [253]. Using patterned substrates, it can be shown that stem cells can ‘sense’ substrate features or topography at the micro- or nano-scale level, and react by changing cell shape (i.e., spreading) and organizing into multicellular physical patterns [254]. Recently, it has been demonstrated that a single cell encapsulation strategy can markedly increase the survival of MSCs by re-introducing cell--matrix interactions via integrin clustering and activation of the MAPK(ERK) signaling cascade [255]. These promising results demonstrate that bioengineered ‘micro-niches’ incorporating adhesion molecules provides a versatile tool for the enhancement of viability and targeted cellular engraftment. Mechanical approaches to enhance acute retention represent a complimentary means of increasing the potential of stem cell therapy for myocardial regeneration. Low initial engraftment of cell products has been attributed to back leak of injected cells due to myocardial contraction, clearance through venous or lymphatic drainage and/or washout by injection trauma [232,256-259]. Sealing the injection site with fibrin glue and reducing the ventricular rate have been shown to increase acute retention with resultant salutary benefits on myocardial contractility [232]. Similarly, labelling cells with superparamagnetic microspheres followed by intra-cardiac injection with a superimposed magnet [260] or intra-coronary implantation of metal stents [261-265] has been shown to enhance the acute retention and functional benefit of cell therapy. 4.
Host effects
The translation of cell-based approaches developed in otherwise healthy animals to therapeutic strategies is not straightforward as most patients suffering from acute MI have a history of multiple coronary artery disease (CAD) risk factors (RF) (diabetes, advanced age, smoking, hypertension and hypercholesterolemia) which have been shown to affect the number and function of circulating EPCs [31]. Clinical reports have also shown that patients with CAD and/or various RFs also show reduced numbers and function of circulating EPCs [266-271]. A clinical report by Vasa et al. showed that in patients with ischemic heart disease, there is an inverse correlation between the number of cardiovascular RFs and the number and migratory activity of EPCs [270]. More recently it has been shown that EPCs derived from older individuals [269], and type II diabetics without diagnosed heart disease [267], have impaired survival, proliferation and migration and reduced incorporation into vascular structures. In addition, BM-MNCs harvested from patients with ischemic cardiomyopathy have a profoundly reduced potential for neovascularization [268], suggesting that the dysfunction is not restricted to circulating 498
cells. This impaired ability of EPCs to contribute to neovascularization may reduce the efficacy of autologous cell delivery for therapeutic applications. 5.
Conclusions
Autologous cell therapy holds the hope of mending the broken heart. Cell therapy with multiple cell types (including those that do not differentiate into new muscle) appears to be beneficial. Also, significant functional improvements occur despite low levels of cell engraftment. New strategies (including more cardiogenic cell types, genetic modification, culture/small-molecule preconditioning and biomechanical engineering) promise to enhance these benefits and improvext generation of autologous cell therapies. 6.
Expert opinion
Over the last decade, a variety of promising autologous cell therapies (CDCs, CPCs, EPCs and MSCs) have been developed from a variety of tissue sources (adipose, bone marrow, blood and heart). The mechanisms of benefit underlying the majority of these cell types appears to be rescue of reversibly damaged myocardium, vascularization resulting from the action of paracrine/humoral factors and secondary recruitment of host stem/progenitor cells [3,55-60]. To date, only the more cardiogenic resident cardiac cell sources appear capable of efficient cardiomyocyte and vascular transdifferentiation [85,97,111,112,114,115]. Interestingly, only the aggregates of the distinct subpopulations differentiated from myocardial tissue appear capable of indirect effects on tissue preservation and/or recruitment of endogenous progenitors [115]. Autologous cell candidates from first-generation therapies are undergoing clinical trials, with second-generation cell products using modified/refined products just beginning clinical assesment [239]. The next fundamental advance in autologous cell candidates promises to be based on recent advances in cellular reprogramming, whereby it has become possible to generate embryonic-like cells from virtually any cell of the body [272]. These induced pluripotent stem (iPS) cells are capable of indefinite self-renewal while maintaining the ability to differentiate into all cell types [273-277]. This research has lead to a greater understanding of the cellular pathways that regulate the balance between pluripotency and differentiation. Not surprisingly, the key nuclear factors (i.e., octamer-binding protein 4 (Oct4), sex determining region Y-box 2 (Sox2), nanog) that drive this equilibrium towards pluripotency and away from differentiation are those that can be exploited to reprogram somatic cells [276,278]. Additional epigenic process significantly contribute to the complex balance between pluripotency and differentiation as pluripotent cell chromatin is transcriptionally more permissive while differentiation is accompanied by a transition to chromatin that is transcriptionally less active [279].
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Davis & Stewart
The rapid pace of this field has lead to the generation of iPS cells from a host of different cell lineages including human amniotic fluid, bone marrow, dermal fibroblasts, hair follicles, hematopoietic cells, neural stem cells and testis [274,280-287]. These cells have been shown to be almost identical to embryonic stem cells in terms of global gene expression, DNA methylation and histone modification [277,288,289]. Furthermore, it has been shown that cell types endogenously expressing low levels of the key reprogramming factors (i.e., neural stem cells and Sox2) require less genetic manipulation to become iPS cells [290]. This approach holds great promise in the generation of functional cell types that are reliable, scalable and capable of differentiating into an adult phenotype [272,291]. Intriguingly, an abbreviated protocol has recently been developed to generate cardiomyocyte-like cells from somatic cells though the introduction of factors that define
cardiogenesis (GATA binding protein 4 (Gata4), myocyte enhancer factor 2C (Mef2c), and T-box transcription factor 5 (Tbx5)) [292]. This data hints that the future of cardiac cell therapy may lie in the generation of autologous cardiomyocytes precursors from easily attainable and expandable sources (e.g., dermal fibroblasts). However, more study is needed to characterize and understand these approaches before they can be effectively (and safely) translated to the clinical setting.
Declaration of interest This paper has been sponsored by the Canadian Institute of Health Research, Heart and Stroke Foundation of Canada. The authors declare no conflict of interest and have received no payment in preparation of this manuscript.
Bibliography 1.
Strauer BE, Brehm M, Zeus T, et al. Repair of infarcted myocardium by autologous intracoronary mononuclear bone marrow cell transplantation in humans. Circulation 2002;106:1913-18
2.
Assmus B, Schachinger V, Teupe C, et al. Transplantation of progenitor cells and regeneration enhancement in acute myocardial infarction (TOPCARE-AMI). Circulation 2002;106:3009-17
3.
Schachinger V, Assmus B, Britten MB, et al. Transplantation of progenitor cells and regeneration enhancement in acute myocardial infarction: final one-year results of the TOPCARE-AMI Trial. J Am Coll Cardiol 2004;44:1690-9
4.
Fernandez-Aviles F, San Roman JA, Garcia-Frade J, et al. Experimental and clinical regenerative capability of human bone marrow cells after myocardial infarction. Circ Res 2004;95:742-8
5.
Bartunek J, Vanderheyden M, Vandekerckhove B, et al. Intracoronary injection of CD133-positive enriched bone marrow progenitor cells promotes cardiac recovery after recent myocardial infarction: feasibility and safety. Circulation 2005;112:I178-83
6.
7.
Yousef M, Schannwell CM, Kostering M, et al. The BALANCE Study: clinical benefit and long-term outcome after intracoronary autologous bone marrow cell transplantation in patients with acute myocardial infarction. J Am Coll Cardiol 2009;53:2262-9 Strauer BE, Yousef M, Schannwell CM. The acute and long-term effects of intracoronary stem cell transplantation in
cell transfer on diastolic function in patients after acute myocardial infarction: results from the BOOST trial. Eur Heart J 2006;27:929-35
191 patients with chronic heARt failure: the STAR-heart study. Eur J Heart Fail 2010;12:721-9 8.
Chen SL, Fang WW, Ye F, et al. Effect on left ventricular function of intracoronary transplantation of autologous bone marrow mesenchymal stem cell in patients with acute myocardial infarction. Am J Cardiol 2004;94:92-5
9.
Ge J, Li Y, Qian J, et al. Efficacy of emergent transcatheter transplantation of stem cells for treatment of acute myocardial infarction (TCT-STAMI). Heart 2006;92:1764-7
10.
Ruan W, Pan CZ, Huang GQ, et al. Assessment of left ventricular segmental function after autologous bone marrow stem cells transplantation in patients with acute myocardial infarction by tissue tracking and strain imaging. Chin Med J (Engl) 2005;118:1175-81
11.
12.
13.
Huang RC, Yao K, Zou YZ, et al. Long term follow-up on emergent intracoronary autologous bone marrow mononuclear cell transplantation for acute inferior-wall myocardial infarction. Zhonghua Yi Xue Za Zhi 2006;86:1107-10 Hendrikx M, Hensen K, Clijsters C, et al. Recovery of regional but not global contractile function by the direct intramyocardial autologous bone marrow transplantation: results from a randomized controlled clinical trial. Circulation 2006;114:I101-7 Schaefer A, Meyer GP, Fuchs M, et al. Impact of intracoronary bone marrow
Expert Opin. Biol. Ther. (2011) 11(4)
14.
Yao K, Huang R, Qian J, et al. Administration of intracoronary bone marrow mononuclear cells on chronic myocardial infarction improves diastolic function. Heart 2008;94:1147-53
15.
Penicka M, Horak J, Kobylka P, et al. Intracoronary injection of autologous bone marrow-derived mononuclear cells in patients with large anterior acute myocardial infarction: a prematurely terminated randomized study. J Am Coll Cardiol 2007;49:2373-4
16.
Panovsky R, Meluzin J, Janousek S, et al. Cell therapy in patients with left ventricular dysfunction due to myocardial infarction. Echocardiography 2008;25:888-97
17.
Wollert KC, Meyer GP, Lotz J, et al. Intracoronary autologous bone-marrow cell transfer after myocardial infarction: the BOOST randomised controlled clinical trial. Lancet 2004;364:141-8
18.
Meyer GP, Wollert KC, Lotz J, et al. Intracoronary bone marrow cell transfer after myocardial infarction: eighteen months’ follow-up data from the randomized, controlled BOOST (BOne marrOw transfer to enhance ST-elevation infarct regeneration) trial. Circulation 2006;113:1287-94
19.
Janssens S, Dubois C, Bogaert J, et al. Autologous bone marrow-derived stem-cell transfer in patients with ST-segment elevation myocardial
499
Autologous cell therapy for cardiac repair
clinical outcome, and serial changes in left ventricular function during 12-months’ follow-up. J Am Coll Cardiol 2008;51:674-6
infarction: double-blind, randomised controlled trial. Lancet 2006;367:113-21 20.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
21.
22.
23.
24.
25.
26.
500
Herbots L, D’hooge J, Eroglu E, et al. Improved regional function after autologous bone marrow-derived stem cell transfer in patients with acute myocardial infarction: a randomized, double-blind strain rate imaging study. Eur Heart J 2009;30:662-70 Erbs S, Linke A, Schachinger V, et al. Restoration of microvascular function in the infarct-related artery by intracoronary transplantation of bone marrow progenitor cells in patients with acute myocardial infarction: the doppler substudy of the reinfusion of enriched progenitor cells and infarct remodeling in acute myocardial infarction (REPAIR-AMI) trial. Circulation 2007;116:366-74
27.
28.
29.
Schachinger V, Erbs S, Elsasser A, et al. Intracoronary bone marrow-derived progenitor cells in acute myocardial infarction. N Engl J Med 2006;355:1210-21 Dill T, Schachinger V, Rolf A, et al. Intracoronary administration of bone marrow-derived progenitor cells improves left ventricular function in patients at risk for adverse remodeling after acute ST-segment elevation myocardial infarction: results of the Reinfusion of Enriched Progenitor cells And Infarct Remodeling in Acute Myocardial Infarction study (REPAIR-AMI) cardiac magnetic resonance imaging substudy. Am Heart J 2009;157:541-7 Lunde K, Solheim S, Aakhus S, et al. Exercise capacity and quality of life after intracoronary injection of autologous mononuclear bone marrow cells in acute myocardial infarction: results from the Autologous Stem cell Transplantation in Acute Myocardial Infarction (ASTAMI) randomized controlled trial. Am Heart J 2007;154:710-18 Meluzin J, Mayer J, Groch L, et al. Autologous transplantation of mononuclear bone marrow cells in patients with acute myocardial infarction: the effect of the dose of transplanted cells on myocardial function. Am Heart J 2006;152:975 e9-15 Lunde K, Solheim S, Forfang K, et al. Anterior myocardial infarction with acute percutaneous coronary intervention and intracoronary injection of autologous mononuclear bone marrow cells: safety,
30.
31.
Suarez de Lezo J, Herrera C, Pan M, et al. Regenerative therapy in patients with a revascularized acute anterior myocardial infarction and depressed ventricular function. Rev Esp Cardiol 2007;60:357-65 Huikuri HV, Kervinen K, Niemela M, et al. Effects of intracoronary injection of mononuclear bone marrow cells on left ventricular function, arrhythmia risk profile, and restenosis after thrombolytic therapy of acute myocardial infarction. Eur Heart J 2008;29:2723-32 Tendera M, Wojakowski W, Ruzyllo W, et al. Intracoronary infusion of bone marrow-derived selected CD34+CXCR4+ cells and non-selected mononuclear cells in patients with acute STEMI and reduced left ventricular ejection fraction: results of randomized, multicentre Myocardial Regeneration by Intracoronary Infusion of Selected Population of Stem Cells in Acute Myocardial Infarction (REGENT) Trial. Eur Heart J 2009;30:1313-21 Traverse JH, McKenna DH, Harvey K, et al. Results of a phase 1, randomized, double-blind, placebo-controlled trial of bone marrow mononuclear stem cell administration in patients following ST-elevation myocardial infarction. Am Heart J 2010;160:428-34 Ward MR, Stewart DJ, Kutryk MJ. Endothelial progenitor cell therapy for the treatment of coronary disease, acute MI, and pulmonary arterial hypertension: current perspectives. Catheter Cardiovasc Interv 2007;70:983-98
32.
Abdel-Latif A, Bolli R, Tleyjeh IM, et al. Adult bone marrow-derived cells for cardiac repair: a systematic review and meta-analysis. Arch Intern Med 2007;167:989-97
33.
Jiang M, He B, Zhang Q, et al. Randomized controlled trials on the therapeutic effects of adult progenitor cells for myocardial infarction: meta-analysis. Expert Opin Biol Ther 2010;10:667-80
34.
Murry CE, Soonpaa MH, Reinecke H, et al. Haematopoietic stem cells do not transdifferentiate into cardiac myocytes in
Expert Opin. Biol. Ther. (2011) 11(4)
myocardial infarcts. Nature 2004;428:664-8 35.
Hristov M, Erl W, Weber PC. Endothelial progenitor cells: mobilization, differentiation, and homing. Arterioscler Thromb Vasc Biol 2003;23:1185-9
36.
Martin-Rendon E, Brunskill S, Doree C, et al. Stem cell treatment for acute myocardial infarction. Cochrane Database Syst Rev 2008:CD006536
37.
Rubart M, Field LJ. Cardiac regeneration: repopulating the heart. Annu Rev Physiol 2006;68:29-49
38.
Urbich C, Aicher A, Heeschen C, et al. Soluble factors released by endothelial progenitor cells promote migration of endothelial cells and cardiac resident progenitor cells. J Mol Cell Cardiol 2005;39:733-42
39.
Narmoneva DA, Vukmirovic R, Davis ME, et al. Endothelial cells promote cardiac myocyte survival and spatial reorganization: implications for cardiac regeneration. Circulation 2004;110:962-8
40.
Toma C, Pittenger MF, Cahill KS, et al. Human mesenchymal stem cells differentiate to a cardiomyocyte phenotype in the adult murine heart. Circulation 2002;105:93-8
41.
Gage FH, Ray J, Fisher LJ. Isolation, characterization, and use of stem cells from the CNS. Annu Rev Neurosci 1995;18:159-92
42.
Hierlihy AM, Seale P, Lobe CG, et al. The post-natal heart contains a myocardial stem cell population. FEBS Lett 2002;530:239-43
43.
Smart N, Riley PR. The stem cell movement. Circ Res 2008;102:1155-68
44.
Asahara T, Masuda H, Takahashi T, et al. Bone marrow origin of endothelial progenitor cells responsible for postnatal vasculogenesis in physiological and pathological neovascularization. Circ Res 1999;85:221-8
45.
George J, Shmilovich H, Deutsch V, et al. Comparative analysis of methods for assessment of circulating endothelial progenitor cells. Tissue Eng 2006;12:331-5
46.
Romagnani P, Annunziato F, Liotta F, et al. CD14+CD34low cells with stem cell phenotypic and functional features are the major source of circulating
Davis & Stewart
myocardial balance of angiogenic cytokines. J Clin Invest 2006;116:1865-77
67.
Prockop DJ. Marrow stromal cells as stem cells for nonhematopoietic tissues. Science 1997;276:71-4
57.
Murry CE, Reinecke H, Pabon LM. Regeneration gaps: observations on stem cells and cardiac repair. J Am Coll Cardiol 2006;47:1777-85
68.
58.
Suuronen EJ, Price J, Veinot JP, et al. Comparative effects of mesenchymal progenitor cells, endothelial progenitor cells, or their combination on myocardial infarct regeneration and cardiac function. J Thorac Cardiovasc Surg 2007;134:1249-58
Dominici M, Le BK, Mueller I, et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy 2006;8:315-17
69.
Korbling M, Estrov Z. Adult stem cells for tissue repair -- a new therapeutic concept? N Engl J Med 2003;349:570-82
70.
Giordano A, Galderisi U, Marino IR. From the laboratory bench to the patient’s bedside: an update on clinical trials with mesenchymal stem cells. J Cell Physiol 2007;211:27-35
71.
Robinson SN, Ng J, Niu T, et al. Superior ex vivo cord blood expansion following co-culture with bone marrow-derived mesenchymal stem cells. Bone Marrow Transplant 2006;37:359-66
72.
Hatzistergos KE, Quevedo H, Oskouei BN, et al. Bone marrow mesenchymal stem cells stimulate cardiac stem cell proliferation and differentiation. Circ Res 2010;107:913-22
73.
Alakel N, Jing D, Muller K, et al. Direct contact with mesenchymal stromal cells affects migratory behavior and gene expression profile of CD133+ hematopoietic stem cells during ex vivo expansion. Exp Hematol 2009;37:504-13
74.
Ringden O, Uzunel M, Rasmusson I, et al. Mesenchymal stem cells for treatment of therapy-resistant graft-versus-host disease. Transplantation 2006;81:1390-7
75.
Baksh D, Song L, Tuan RS. Adult mesenchymal stem cells: characterization, differentiation, and application in cell and gene therapy. J Cell Mol Med 2004;8:301-16
76.
Koc ON, Day J, Nieder M, et al. Allogeneic mesenchymal stem cell infusion for treatment of metachromatic leukodystrophy (MLD) and Hurler syndrome (MPS-IH). Bone Marrow Transplant 2002;30:215-22
77.
Horwitz EM, Gordon PL, Koo WK, et al. Isolated allogeneic bone marrow-derived mesenchymal cells engraft and stimulate growth in children
endothelial progenitors. Circ Res 2005;97:314-22 47.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
48.
Klein HM, Ghodsizad A, Marktanner R, et al. Intramyocardial implantation of CD133+ stem cells improved cardiac function without bypass surgery. Heart Surg Forum 2007;10:E66-9 Manginas A, Goussetis E, Koutelou M, et al. Pilot study to evaluate the safety and feasibility of intracoronary CD133+ and CD133-CD34+ cell therapy in patients with nonviable anterior myocardial infarction. Catheter Cardiovasc Interv 2007;69:773-81
49.
Asahara T, Murohara T, Sullivan A, et al. Isolation of putative progenitor endothelial cells for angiogenesis. Science 1997;275:964-7
50.
Dimmeler S, Zeiher AM, Schneider MD. Unchain my heart: the scientific foundations of cardiac repair. J Clin Invest 2005;115:572-83
51.
52.
53.
54.
55.
56.
Ingram DA, Mead LE, Tanaka H, et al. Identification of a novel hierarchy of endothelial progenitor cells using human peripheral and umbilical cord blood. Blood 2004;104:2752-60 Kawamoto A, Tkebuchava T, Yamaguchi J, et al. Intramyocardial transplantation of autologous endothelial progenitor cells for therapeutic neovascularization of myocardial ischemia. Circulation 2003;107:461-8 Kamihata H, Matsubara H, Nishiue T, et al. Implantation of bone marrow mononuclear cells into ischemic myocardium enhances collateral perfusion and regional function via side supply of angioblasts, angiogenic ligands, and cytokines. Circulation 2001;104:1046-52
59.
60.
61.
Dobert N, Britten M, Assmus B, et al. Transplantation of progenitor cells after reperfused acute myocardial infarction: evaluation of perfusion and myocardial viability with FDG-PET and thallium SPECT. Eur J Nucl Med Mol Imaging 2004;31:1146-51 Jia L, Takahashi M, Yoshioka T, et al. Therapeutic potential of endothelial progenitor cells for cardiovascular diseases. Curr Vasc Pharmacol 2006;4:59-65
62.
Jujo K, Ii M, Losordo DW. Endothelial progenitor cells in neovascularization of infarcted myocardium. J Mol Cell Cardiol 2008;45:530-44
63.
Urbich C, Dimmeler S. Endothelial progenitor cells functional characterization. Trends Cardiovasc Med 2004;14:318-22
64.
Gulati R, Jevremovic D, Peterson TE, et al. Diverse origin and function of cells with endothelial phenotype obtained from adult human blood. Circ Res 2003;93:1023-5
Tse HF, Siu CW, Zhu SG, et al. Paracrine effects of direct intramyocardial implantation of bone marrow derived cells to enhance neovascularization in chronic ischaemic myocardium. Eur J Heart Fail 2007;9:747-53
65.
Cho HJ, Lee N, Lee JY, et al. Role of host tissues for sustained humoral effects after endothelial progenitor cell transplantation into the ischemic heart. J Exp Med 2007;204:3257-69
66.
Fazel S, Cimini M, Chen L, et al. Cardioprotective c-kit+ cells are from the bone marrow and regulate the
Ruel M, Suuronen EJ, Song J, et al. Effects of off-pump versus on-pump coronary artery bypass grafting on function and viability of circulating endothelial progenitor cells. J Thorac Cardiovasc Surg 2005;130:633-9
Hur J, Yoon CH, Kim HS, et al. Characterization of two types of endothelial progenitor cells and their different contributions to neovasculogenesis. Arterioscler Thromb Vasc Biol 2004;24:288-93 Ormiston ML, Slaughter GR, Deng Y, et al. The enzymatic degradation of hyaluronan is associated with disease progression in experimental pulmonary hypertension. Am J Physiol Lung Cell Mol Physiol 2010;298:L148-57
Expert Opin. Biol. Ther. (2011) 11(4)
501
Autologous cell therapy for cardiac repair
with osteogenesis imperfecta: implications for cell therapy of bone. Proc Natl Acad Sci USA 2002;99:8932-7
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
78.
Frank MH, Sayegh MH. Immunomodulatory functions of mesenchymal stem cells. Lancet 2004;363:1411-12
80.
Krampera M, Cosmi L, Angeli R, et al. Role for interferon-gamma in the immunomodulatory activity of human bone marrow mesenchymal stem cells. Stem Cells 2006;24:386-98
82.
Yoon YS, Park JS, Tkebuchava T, et al. Unexpected severe calcification after transplantation of bone marrow cells in acute myocardial infarction. Circulation 2004;109:3154-7 Breitbach M, Bostani T, Roell W, et al. Potential risks of bone marrow cell transplantation into infarcted hearts. Blood 2007;110:1362-9
83.
Rubio D, Garcia-Castro J, Martin MC, et al. Spontaneous human adult stem cell transformation. Cancer Res 2005;65:3035-9
84.
Chien KR, Olson EN. Converging pathways and principles in heart development and disease: CV@CSH. Cell 2002;110:153-62
85.
Beltrami AP, Barlucchi L, Torella D, et al. Adult cardiac stem cells are multipotent and support myocardial regeneration. Cell 2003;114:763-76
86.
Oh H, Bradfute SB, Gallardo TD, et al. Cardiac progenitor cells from adult myocardium: homing, differentiation, and fusion after infarction. Proc Natl Acad Sci USA 2003;100:12313-18
87.
Ott HC, Matthiesen TS, Brechtken J, et al. The adult human heart as a source for stem cells: repair strategies with embryonic-like progenitor cells. Nat Clin Pract Cardiovasc Med 2007;4(Suppl 1):S27-39
88.
89.
502
90.
Le BK. Immunomodulatory effects of fetal and adult mesenchymal stem cells. Cytotherapy 2003;5:485-9
79.
81.
Biochem Biophys Res Commun 2008;371:615-20
Pfister O, Oikonomopoulos A, Sereti KI, et al. Role of the ATP-binding cassette transporter Abcg2 in the phenotype and function of cardiac side population cells. Circ Res 2008;103:825-35 Yamahara K, Fukushima S, Coppen SR, et al. Heterogeneic nature of adult cardiac side population cells.
Bu L, Jiang X, Martin-Puig S, et al. Human ISL1 heart progenitors generate diverse multipotent cardiovascular cell lineages. Nature 2009;460:113-17
91.
Xaymardan M, Cimini M, Fazel S, et al. c-Kit function is necessary for in vitro myogenic differentiation of bone marrow hematopoietic cells. Stem Cells 2009;27:1911-20
92.
Cimini M, Fazel S, Zhuo S, et al. c-kit dysfunction impairs myocardial healing after infarction. Circulation 2007;116:I77-82
93.
Bergmann O, Bhardwaj RD, Bernard S, et al. Evidence for cardiomyocyte renewal in humans. Science 2009;324:98-102
94.
Kajstura J, Urbanek K, Perl S, et al. Cardiomyogenesis in the adult human heart. Circ Res 2010;107:305-15
95.
Martin CM, Meeson AP, Robertson SM, et al. Persistent expression of the ATP-binding cassette transporter, Abcg2, identifies cardiac SP cells in the developing and adult heart. Dev Biol 2004;265:262-75
96.
97.
Pfister O, Mouquet F, Jain M, et al. CD31- but not CD31+ cardiac side population cells exhibit functional cardiomyogenic differentiation. Circ Res 2005;97:52-61 Bearzi C, Rota M, Hosoda T, et al. Human cardiac stem cells. Proc Natl Acad Sci USA 2007;104:14068-73
98.
Goumans MJ, de Boer TP, Smits AM, et al. TGF-beta1 induces efficient differentiation of human cardiomyocyte progenitor cells into functional cardiomyocytes in vitro. Stem Cell Res 2007;1:138-49
99.
Winter EM, van Oorschot AA, Hogers B, et al. A new direction for cardiac regeneration therapy: application of synergistically acting epicardium-derived cells and cardiomyocyte progenitor cells. Circ Heart Fail 2009;2:643-53
100. Smits AM, van VP, Metz CH, et al. Human cardiomyocyte progenitor cells differentiate into functional mature cardiomyocytes: an in vitro model for studying human cardiac physiology and pathophysiology. Nat Protoc 2009;4:232-43 101. Oyama T, Nagai T, Wada H, et al. Cardiac side population cells Expert Opin. Biol. Ther. (2011) 11(4)
have a potential to migrate and differentiate into cardiomyocytes in vitro and in vivo. J Cell Biol 2007;176:329-41 102. Meissner K, Heydrich B, Jedlitschky G, et al. The ATP-binding cassette transporter ABCG2 (BCRP), a marker for side population stem cells, is expressed in human heart. J Histochem Cytochem 2006;54:215-21 103. Quaini F, Urbanek K, Beltrami AP, et al. Chimerism of the transplanted heart. N Engl J Med 2002;346:5-15 104. Beltrami AP, Barlucchi L, Torella D, et al. Adult cardiac stem cells are multipotent and support myocardial regeneration. Cell 2003;114:763-76 105. Linke A, Muller P, Nurzynska D, et al. Stem cells in the dog heart are self-renewing, clonogenic, and multipotent and regenerate infarcted myocardium, improving cardiac function. Proc Natl Acad Sci USA 2005;102:8966-71 106. Urbanek K, Cesselli D, Rota M, et al. Stem cell niches in the adult mouse heart. Proc Natl Acad Sci USA 2006;103:9226-31 107. Tang XL, Rokosh G, Sanganalmath SK, et al. Intracoronary administration of cardiac progenitor cells alleviates left ventricular dysfunction in rats with a 30-day-old infarction. Circulation 2010;121:293-305 108. Bolli R. Cardiac stem cell infusion in patients with ischemic cardiomyopathy (SCIPIO, NCT00474461). Bethesda, MD: clinicaltrials.gov, 2010. Available from: http://clinicaltrials.gov/ct2/show/ NCT00474461 [Last accessed 27 January 2011] 109. Davis DR, Kizana E, Terrovitis J, et al. Isolation and expansion of functionally-competent cardiac progenitor cells directly from heart biopsies. J Mol Cell Cardiol 2010;49:312-21 110. Tang YL, Shen L, Qian K, Phillips MI. A novel two-step procedure to expand cardiac Sca-1+ cells clonally. Biochem Biophys Res Commun 2007;359:877-83 111. Messina E, De Angelis L, Frati G, et al. Isolation and expansion of adult cardiac stem cells from human and murine heart. Circ Res 2004;95:911-21 112. Smith RR, Barile L, Cho HC, et al. Regenerative potential of
Davis & Stewart
cardiosphere-derived cells expanded from percutaneous endomyocardial biopsy specimens. Circulation 2007;115:896-908 113.
114.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
115.
116.
117.
118.
119.
120.
121.
Davis DR, Zhang Y, Smith RR, et al. Validation of the cardiosphere method to culture cardiac progenitor cells from myocardial tissue. PLoS ONE 2009;4:e7195 Davis DR, Smith RR, Marban E. Human cardiospheres are a source of stem cells with cardiomyogenic potential. Stem Cells 2010;28:903-4 Chimenti I, Smith RR, Li TS, et al. Relative roles of direct regeneration versus paracrine effects of human cardiosphere-derived cells transplanted into infarcted mice. Circ Res 2010;106:971-80 Chakravarty T, Matsushita S, Terrovitis J, et al. Thymosin Beta-4 (TB4) enhances cardiosphere-derived cardiac stem cell (CDC)-mediated cardioprotection by promoting CDC migration and angiogenesis via Akt phosphorylation [abstract 3611]. Circulation 2009;120:S836 Lee S-T, White AJ, Matsushita S, et al. Direct intramyocardial injection of autologous cardiosphere-derived cells yields consistent engraftment and improves cardiac function in a porcine model of ischemic cardiomyopathy [abstract 3976]. Circulation 2009;120:S898 Zhang Y, Abraham RM, Marban E. Dedifferentiation of mammalian cardiomyocytes into cardiac stem cells [abstract 3875]. Circulation 2009;120:S877 CArdiosphere-Derived aUtologous Stem CElls to Reverse ventricUlar dySfunction (CADUCEUS). Bethesda, MD: clinicaltrials.gov 2009. Available from: http://clinicaltrials.gov/ct2/show/ NCT00893360 [Last accessed 27 January 2011] Johnston PV, Sasano T, Mills K, et al. Engraftment, differentiation, and functional benefits of autologous cardiosphere-derived cells in porcine ischemic cardiomyopathy. Circulation 2009;120:1075-83 Menasche P, Hagege AA, Vilquin JT, et al. Autologous skeletal myoblast transplantation for severe postinfarction left ventricular dysfunction. J Am Coll Cardiol 2003;41:1078-83
122.
Smits PC, van Geuns RJ, Poldermans D, et al. Catheter-based intramyocardial injection of autologous skeletal myoblasts as a primary treatment of ischemic heart failure: clinical experience with six-month follow-up. J Am Coll Cardiol 2003;42:2063-9
133. Valina C, Pinkernell K, Song YH, et al. Intracoronary administration of autologous adipose tissue-derived stem cells improves left ventricular function, perfusion, and remodelling after acute myocardial infarction. Eur Heart J 2007;28:2667-77
123.
Abraham MR, Henrikson CA, Tung L, et al. Antiarrhythmic engineering of skeletal myoblasts for cardiac transplantation. Circ Res 2005;97:159-67
124.
Gimble JM, Katz AJ, Bunnell BA. Adipose-derived stem cells for regenerative medicine. Circ Res 2007;100:1249-60
134. Planat-Benard V, Silvestre JS, Cousin B, et al. Plasticity of human adipose lineage cells toward endothelial cells: physiological and therapeutic perspectives. Circulation 2004;109:656-63
125.
126.
127.
Fraser JK, Wulur I, Alfonso Z, Hedrick MH. Fat tissue: an underappreciated source of stem cells for biotechnology. Trends Biotechnol 2006;24:150-4 Katz AJ, Tholpady A, Tholpady SS, et al. Cell surface and transcriptional characterization of human adipose-derived adherent stromal (hADAS) cells. Stem Cells 2005;23:412-23 Gronthos S, Franklin DM, Leddy HA, et al. Surface protein characterization of human adipose tissue-derived stromal cells. J Cell Physiol 2001;189:54-63
128.
Mitchell JB, McIntosh K, Zvonic S, et al. Immunophenotype of human adipose-derived cells: temporal changes in stromal-associated and stem cell-associated markers. Stem Cells 2006;24:376-85
129.
Lindroos B, Boucher S, Chase L, et al. Serum-free, xeno-free culture media maintain the proliferation rate and multipotentiality of adipose stem cells in vitro. Cytotherapy 2009;11:958-72
130.
131.
132.
van DA, Niessen HW, Zandieh DB, et al. Differentiation of human adipose-derived stem cells towards cardiomyocytes is facilitated by laminin. Cell Tissue Res 2008;334:457-67 Bai X, Pinkernell K, Song YH, et al. Genetically selected stem cells from human adipose tissue express cardiac markers. Biochem Biophys Res Commun 2007;353:665-71 Song YH, Gehmert S, Sadat S, et al. VEGF is critical for spontaneous differentiation of stem cells into cardiomyocytes. Biochem Biophys Res Commun 2007;354:999-1003
Expert Opin. Biol. Ther. (2011) 11(4)
135. Madonna R, Geng YJ, De CR. Adipose tissue-derived stem cells: characterization and potential for cardiovascular repair. Arterioscler Thromb Vasc Biol 2009;29:1723-9 136. Miranville A, Heeschen C, Sengenes C, et al. Improvement of postnatal neovascularization by human adipose tissue-derived stem cells. Circulation 2004;110:349-55 137. De FF, Tirino V, Desiderio V, et al. Human CD34+/CD90+ ASCs are capable of growing as sphere clusters, producing high levels of VEGF and forming capillaries. PLoS ONE 2009;4:e6537 138. Cai L, Johnstone BH, Cook TG, et al. IFATS collection: human adipose tissue-derived stem cells induce angiogenesis and nerve sprouting following myocardial infarction, in conjunction with potent preservation of cardiac function. Stem Cells 2009;27:230-7 139. Wang L, Deng J, Tian W, et al. Adipose-derived stem cells are an effective cell candidate for treatment of heart failure: an MR imaging study of rat hearts. Am J Physiol Heart Circ Physiol 2009;297:H1020-31 140. Bai X, Yan Y, Song YH, et al. Both cultured and freshly isolated adipose tissue-derived stem cells enhance cardiac function after acute myocardial infarction. Eur Heart J 2010;31:489-501 141. Schenke-Layland K, Strem BM, Jordan MC, et al. Adipose tissue-derived cells improve cardiac function following myocardial infarction. J Surg Res 2009;153:217-23 142. Sadat S, Gehmert S, Song YH, et al. The cardioprotective effect of mesenchymal stem cells is mediated by IGF-I and VEGF. Biochem Biophys Res Commun 2007;363:674-9
503
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Autologous cell therapy for cardiac repair
143. Rehman J, Traktuev D, Li J, et al. Secretion of angiogenic and antiapoptotic factors by human adipose stromal cells. Circulation 2004;109:1292-8
and improves cardiac function in chronic ischemic heart failure. J Thorac Cardiovasc Surg 2006;132:1321-8
144. Fernandez-Aviles F, Perrin E. A randomized clinical trial of adipose-derived stem cells in treatment of non revascularizable ischemic myocardium (PRECISE, NCT00426868). Bethesda, MD: clinicaltrials.gov 2010. Available from: http://clinicaltrials.gov/ct2/ show/NCT00426868?term= NCT00426868&rank=1 [Last accessed 27 January 2011]
152. Guarita-Souza LC, Carvalho KA, Woitowicz V, et al. Simultaneous autologous transplantation of cocultured mesenchymal stem cells and skeletal myoblasts improves ventricular function in a murine model of Chagas disease. Circulation 2006;114:I120-4
145. Serruys P. Randomized clinical trial of adipose-derived stem cells in the treatment of pts with st-elevation myocardial infarction (APOLLO, NCT00442806). Bethesda, MD: clinicaltrials.gov 2007. Available from: http://clinicaltrials.gov/ct2/ show/NCT00442806?term= NCT00442806&rank=1 [Last accessed 27 Januarry 2011] 146. Kinnaird T, Stabile E, Burnett MS, et al. Marrow-derived stromal cells express genes encoding a broad spectrum of arteriogenic cytokines and promote in vitro and in vivo arteriogenesis through paracrine mechanisms. Circ Res 2004;94:678-85 147. Ziegelhoeffer T, Fernandez B, Kostin S, et al. Bone marrow-derived cells do not incorporate into the adult growing vasculature. Circ Res 2004;94:230-8 148. Popa ER, Harmsen MC, Tio RA, et al. Circulating CD34+ progenitor cells modulate host angiogenesis and inflammation in vivo. J Mol Cell Cardiol 2006;41:86-96 149. Terrovitis J, Kwok KF, Lautamaki R, et al. Ectopic expression of the sodium-iodide symporter enables imaging of transplanted cardiac stem cells in vivo by single-photon emission computed tomography or positron emission tomography. J Am Coll Cardiol 2008;52:1652-60 150. Li Z, Lee A, Huang M, et al. Imaging survival and function of transplanted cardiac resident stem cells. J Am Coll Cardiol 2009;53:1229-40 151. Bonaros N, Rauf R, Wolf D, et al. Combined transplantation of skeletal myoblasts and angiopoietic progenitor cells reduces infarct size and apoptosis
504
153. Nakanishi C, Yamagishi M, Yamahara K, et al. Activation of cardiac progenitor cells through paracrine effects of mesenchymal stem cells. Biochem Biophys Res Commun 2008;374:11-16 154. Smith RR, Chimenti I, Marban E. Unselected human cardiosphere-derived cells are functionally superior to c-kit- or CD90-purified cardiosphere-derived cells [abstract 3417]. Circulation 2008;118:S420 155. Calvillo L, Latini R, Kajstura J, et al. Recombinant human erythropoietin protects the myocardium from ischemia-reperfusion injury and promotes beneficial remodeling. Proc Natl Acad Sci USA 2003;100:4802-6 156. Urbanek K, Rota M, Cascapera S, et al. Cardiac stem cells possess growth factor-receptor systems that after activation regenerate the infarcted myocardium, improving ventricular function and long-term survival. Circ Res 2005;97:663-73 157. Dawn B, Guo Y, Rezazadeh A, et al. Postinfarct cytokine therapy regenerates cardiac tissue and improves left ventricular function. Circ Res 2006;98:1098-105 158. Muraski JA, Rota M, Misao Y, et al. Pim-1 regulates cardiomyocyte survival downstream of Akt. Nat Med 2007;13:1467-75 159. Wollert KC, Chien KR. Cardiotrophin-1 and the role of gp130-dependent signaling pathways in cardiac growth and development. J Mol Med 1997;75:492-501 160. Kristensen DM, Kalisz M, Nielsen JH. Cytokine signalling in embryonic stem cells. APMIS 2005;113:756-72 161. Zohlnhofer D, Dibra A, Koppara T, et al. Stem cell mobilization by granulocyte colony-stimulating factor for myocardial recovery after acute
Expert Opin. Biol. Ther. (2011) 11(4)
myocardial infarction: a meta-analysis. J Am Coll Cardiol 2008;51:1429-37 162. Engel FB, Schebesta M, Duong MT, et al. p38 MAP kinase inhibition enables proliferation of adult mammalian cardiomyocytes. Genes Dev 2005;19:1175-87 163. Engel FB, Hsieh PC, Lee RT, Keating MT. FGF1/p38 MAP kinase inhibitor therapy induces cardiomyocyte mitosis, reduces scarring, and rescues function after myocardial infarction. Proc Natl Acad Sci USA 2006;103:15546-51 164. Sato N, Meijer L, Skaltsounis L, et al. Maintenance of pluripotency in human and mouse embryonic stem cells through activation of Wnt signaling by a pharmacological GSK-3-specific inhibitor. Nat Med 2004;10:55-63 165. Tseng AS, Engel FB, Keating MT. The GSK-3 inhibitor BIO promotes proliferation in mammalian cardiomyocytes. Chem Biol 2006;13:957-63 166. Gude N, Muraski J, Rubio M, et al. Akt promotes increased cardiomyocyte cycling and expansion of the cardiac progenitor cell population. Circ Res 2006;99:381-8 167. Li TS, Hayashi M, Ito H, et al. Regeneration of infarcted myocardium by intramyocardial implantation of ex vivo transforming growth factor-beta-preprogrammed bone marrow stem cells. Circulation 2005;111:2438-45 168. Sasaki K, Heeschen C, Aicher A, et al. Ex vivo pretreatment of bone marrow mononuclear cells with endothelial NO synthase enhancer AVE9488 enhances their functional activity for cell therapy. Proc Natl Acad Sci USA 2006;103:14537-41 169. Okayasu T, Tomizawa A, Suzuki K, et al. PPARalpha activators upregulate eNOS activity and inhibit cytokine-induced NF-kappaB activation through AMP-activated protein kinase activation. Life Sci 2008;82:884-91 170. Yakubu MA, Nsaif RH, Oyekan AO. Peroxisome proliferator-activated receptor alpha activation-mediated regulation of endothelin-1 production via nitric oxide and protein kinase C signaling pathways in piglet cerebral microvascular endothelial cell culture. J Pharmacol Exp Ther 2007;320:774-81
Davis & Stewart
171.
172.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
173.
174.
Ma FX, Chen F, Ren Q, Han ZC. Lovastatin restores the function of endothelial progenitor cells damaged by oxLDL. Acta Pharmacol Sin 2009;30:545-52 Behfar A, Yamada S, Crespo-Diaz R, et al. Guided cardiopoiesis enhances therapeutic benefit of bone marrow human mesenchymal stem cells in chronic myocardial infarction. J Am Coll Cardiol 2010;56:721-34 Chacko SM, Khan M, Kuppusamy ML, et al. Myocardial oxygenation and functional recovery in infarct rat hearts transplanted with mesenchymal stem cells. Am J Physiol Heart Circ Physiol 2009;296:H1263-73 Cipolleschi MG, Dello SP, Olivotto M. The role of hypoxia in the maintenance of hematopoietic stem cells. Blood 1993;82:2031-7
175.
Csete M. Oxygen in the cultivation of stem cells. Ann NY Acad Sci 2005;1049:1-8
176.
Khan M, Kutala VK, Vikram DS, et al. Skeletal myoblasts transplanted in the ischemic myocardium enhance in situ oxygenation and recovery of contractile function. Am J Physiol Heart Circ Physiol 2007;293:H2129-39
177.
Wang JA, Chen TL, Jiang J, et al. Hypoxic preconditioning attenuates hypoxia/reoxygenation-induced apoptosis in mesenchymal stem cells. Acta Pharmacol Sin 2008;29:74-82
178.
Wang JA, He A, Hu X, et al. Anoxic preconditioning: a way to enhance the cardioprotection of mesenchymal stem cells. Int J Cardiol 2009;133:410-12
179.
180.
181.
182.
Grayson WL, Zhao F, Bunnell B, Ma T. Hypoxia enhances proliferation and tissue formation of human mesenchymal stem cells. Biochem Biophys Res Commun 2007;358:948-53 Chacko SM, Ahmed S, Selvendiran K, et al. Hypoxic preconditioning induces the expression of prosurvival and proangiogenic markers in mesenchymal stem cells. Am J Physiol Cell Physiol 2010;99:C1562-70
down-regulation of p16 and extracellular signal regulated kinase. Biochem Biophys Res Commun 2010;391:1471-6 183.
184.
185.
186.
187.
188.
Ames BN, Shigenaga MK, Hagen TM. Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci USA 1993;90:7915-22 Wang F, Thirumangalathu S, Loeken MR. Establishment of new mouse embryonic stem cell lines is improved by physiological glucose and oxygen. Cloning Stem Cells 2006;8:108-16 Fehrer C, Brunauer R, Laschober G, et al. Reduced oxygen tension attenuates differentiation capacity of human mesenchymal stem cells and prolongs their lifespan. Aging Cell 2007;6:745-57 Li TS, Cheng K, Malliaras K, et al. Expansion of human cardiac stem cells in physiological oxygen improves cell production efficiency and potency for myocardial repair. Cardiovasc Res 2011;89:157-65 Li T-S, Lee S-T, Matsushita S, et al. Cardiospheres recapitulate a niche-like microenvironment rich in stemness and cell-matrix interactions, rationalizing their enhanced functional potency for myocardial repair. Stem Cells 2010;28:2088-98
189.
Mangi AA, Noiseux N, Kong D, et al. Mesenchymal stem cells modified with Akt prevent remodeling and restore performance of infarcted hearts. Nat Med 2003;9:1195-201
190.
Bock-Marquette I, Saxena A, White MD, et al. Thymosin beta4 activates integrin-linked kinase and promotes cardiac cell migration, survival and cardiac repair. Nature 2004;432:466-72
191.
Li TS, Marban E. Physiological levels of reactive oxygen species are required to maintain genomic stability in stem cells. Stem Cells 2010;28:1178-85 Jin Y, Kato T, Furu M, et al. Mesenchymal stem cells cultured under hypoxia escape from senescence via
van Gent DC, Hoeijmakers JH, Kanaar R. Chromosomal stability and the DNA double-stranded break connection. Nat Rev Genet 2001;2:196-206
192.
Aharinejad S, Abraham D, Paulus P, et al. Colony-stimulating factor-1 transfection of myoblasts improves the repair of failing myocardium following autologous myoblast transplantation. Cardiovasc Res 2008;79:395-404 Deng J, Han Y, Yan C, et al. Overexpressing cellular repressor of E1A-stimulated genes protects
Expert Opin. Biol. Ther. (2011) 11(4)
mesenchymal stem cells against hypoxia- and serum deprivation-induced apoptosis by activation of PI3K/Akt. Apoptosis 2010;15:463-73 193. Zeng B, Chen H, Zhu C, et al. Effects of combined mesenchymal stem cells and heme oxygenase-1 therapy on cardiac performance. Eur J Cardiothorac Surg 2008;34:850-6 194. Zeng B, Ren X, Lin G, et al. Paracrine action of HO-1-modified mesenchymal stem cells mediates cardiac protection and functional improvement. Cell Biol Int 2008;32:1256-64 195. Fan L, Lin C, Zhuo S, et al. Transplantation with survivin-engineered mesenchymal stem cells results in better prognosis in a rat model of myocardial infarction. Eur J Heart Fail 2009;11:1023-30 196. Elmadbouh I, Haider HK, Jiang S, et al. Ex vivo delivered stromal cell-derived factor-1alpha promotes stem cell homing and induces angiomyogenesis in the infarcted myocardium. J Mol Cell Cardiol 2007;42:792-803 197. Haider HK, Jiang S, Idris NM, Ashraf M. IGF-1-overexpressing mesenchymal stem cells accelerate bone marrow stem cell mobilization via paracrine activation of SDF-1alpha/ CXCR4 signaling to promote myocardial repair. Circ Res 2008;103:1300-8 198. Wang X, Zhao T, Huang W, et al. Hsp20-engineered mesenchymal stem cells are resistant to oxidative stress via enhanced activation of Akt and increased secretion of growth factors. Stem Cells 2009;27:3021-31 199. Kutschka I, Kofidis T, Chen IY, et al. Adenoviral human BCL-2 transgene expression attenuates early donor cell death after cardiomyoblast transplantation into ischemic rat hearts. Circulation 2006;114:I174-80 200. Fischer KM, Cottage CT, Wu W, et al. Enhancement of myocardial regeneration through genetic engineering of cardiac progenitor cells expressing Pim-1 kinase. Circulation 2009;120:2077-87 201. Zhang D, Fan GC, Zhou X, et al. Over-expression of CXCR4 on mesenchymal stem cells augments myoangiogenesis in the infarcted myocardium. J Mol Cell Cardiol 2008;44:281-92
505
Autologous cell therapy for cardiac repair
202. Schenk S, Mal N, Finan A, et al. Monocyte chemotactic protein-3 is a myocardial mesenchymal stem cell homing factor. Stem Cells 2007;25:245-51
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
203. Kong D, Melo LG, Mangi AA, et al. Enhanced inhibition of neointimal hyperplasia by genetically engineered endothelial progenitor cells. Circulation 2004;109:1769-75 204. Song MB, Yu XJ, Zhu GX, et al. Transfection of HGF gene enhances endothelial progenitor cell (EPC) function and improves EPC transplant efficiency for balloon-induced arterial injury in hypercholesterolemic rats. Vascul Pharmacol 2008;51:205-13 205. Jiang M, Wang B, Wang C, et al. Angiogenesis by transplantation of HIF-1alpha modified EPCs into ischemic limbs. J Cell Biochem 2008;103:321-34 206. Iwaguro H, Yamaguchi JI, Kalka C, et al. Endothelial progenitor cell vascular endothelial growth factor gene transfer for vascular regeneration. Circulation 2002;105:732-8 207. Zhang M, Mal N, Kiedrowski M, et al. SDF-1 expression by mesenchymal stem cells results in trophic support of cardiac myocytes after myocardial infarction. FASEB J 2007;21:3197-207 208. Tateishi K, Ashihara E, Honsho S, et al. Human cardiac stem cells exhibit mesenchymal features and are maintained through Akt/GSK-3beta signaling. Biochem Biophys Res Commun 2007;352:635-41 209. Roell W, Lewalter T, Sasse P, et al. Engraftment of connexin 43-expressing cells prevents post-infarct arrhythmia. Nature 2007;450:819-24 210. Jiang M, Wang B, Wang C, et al. In vivo enhancement of angiogenesis by adenoviral transfer of HIF-1alpha-modified endothelial progenitor cells (Ad-HIF-1alphamodified EPC for angiogenesis). Int J Biochem Cell Biol 2008;40:2284-95 211. Ben-Shoshan J, Schwartz S, Luboshits G, et al. Constitutive expression of HIF-1alpha and HIF-2alpha in bone marrow stromal cells differentially promotes their proangiogenic properties. Stem Cells 2008;26:2634-43 212. Iwaguro H, Yamaguchi J, Kalka C, et al. Endothelial progenitor cell vascular endothelial growth factor gene transfer
506
for vascular regeneration. Circulation 2002;105:732-8 213. Zhang M, Mal N, Kiedrowski M, et al. SDF-1 expression by mesenchymal stem cells results in trophic support of cardiac myocytes after myocardial infarction. FASEB J 2007;21:3197-207 214. Unzek S, Zhang M, Mal N, et al. SDF-1 recruits cardiac stem cell-like cells that depolarize in vivo. Cell Transplant 2007;16:879-86 215. Wang X, Hu Q, Mansoor A, et al. Bioenergetic and functional consequences of stem cell-based VEGF delivery in pressure-overloaded swine hearts. Am J Physiol Heart Circ Physiol 2006;290:H1393-405 216. Yang F, Cho SW, Son SM, et al. Genetic engineering of human stem cells for enhanced angiogenesis using biodegradable polymeric nanoparticles. Proc Natl Acad Sci USA 2010;107:3317-22 217. Gao F, He T, Wang H, et al. A promising strategy for the treatment of ischemic heart disease: mesenchymal stem cell-mediated vascular endothelial growth factor gene transfer in rats. Can J Cardiol 2007;23:891-8 218. Matsumoto R, Omura T, Yoshiyama M, et al. Vascular endothelial growth factor-expressing mesenchymal stem cell transplantation for the treatment of acute myocardial infarction. Arterioscler Thromb Vasc Biol 2005;25:1168-73 219. Lin H, Shabbir A, Molnar M, et al. Adenoviral expression of vascular endothelial growth factor splice variants differentially regulate bone marrow-derived mesenchymal stem cells. J Cell Physiol 2008;216:458-68 220. Duan HF, Wu CT, Wu DL, et al. Treatment of myocardial ischemia with bone marrow-derived mesenchymal stem cells overexpressing hepatocyte growth factor. Mol Ther 2003;8:467-74 221. Jiang S, Haider HK, Idris NM, et al. Supportive interaction between cell survival signaling and angiocompetent factors enhances donor cell survival and promotes angiomyogenesis for cardiac repair. Circ Res 2006;99:776-84 222. Shujia J, Haider HK, Idris NM, et al. Stable therapeutic effects of mesenchymal stem cell-based multiple gene delivery for
Expert Opin. Biol. Ther. (2011) 11(4)
cardiac repair. Cardiovasc Res 2008;77:525-33 223. Li H, Zuo S, He Z, et al. Paracrine factors released by GATA-4 overexpressed mesenchymal stem cells increase angiogenesis and cell survival. Am J Physiol Heart Circ Physiol 2010;299:H1772-8 224. Huang J, Zhang Z, Guo J, et al. Genetic modification of mesenchymal stem cells overexpressing CCR1 increases cell viability, migration, engraftment, and capillary density in the injured myocardium. Circ Res 2010;106:1753-62 225. Zhang D, Fan GC, Zhou X, et al. Over-expression of CXCR4 on mesenchymal stem cells augments myoangiogenesis in the infarcted myocardium. J Mol Cell Cardiol 2008;44:281-92 226. Murasawa S, Llevadot J, Silver M, et al. Constitutive human telomerase reverse transcriptase expression enhances regenerative properties of endothelial progenitor cells. Circulation 2002;106:1133-9 227. Gnecchi M, He H, Liang OD, et al. Paracrine action accounts for marked protection of ischemic heart by Akt-modified mesenchymal stem cells. Nat Med 2005;11:367-8 228. Gnecchi M, He H, Noiseux N, et al. Evidence supporting paracrine hypothesis for Akt-modified mesenchymal stem cell-mediated cardiac protection and functional improvement. FASEB J 2006;20:661-9 229. Mirotsou M, Zhang Z, Deb A, et al. Secreted frizzled related protein 2 (Sfrp2) is the key Akt-mesenchymal stem cell-released paracrine factor mediating myocardial survival and repair. Proc Natl Acad Sci USA 2007;104:1643-8 230. Cheng Z, Ou L, Zhou X, et al. Targeted migration of mesenchymal stem cells modified with CXCR4 gene to infarcted myocardium improves cardiac performance. Mol Ther 2008;16:571-9 231. Terrovitis J, Kwok KF, Lautamaki R, et al. Ectopic expression of the sodium-iodide symporter enables imaging of transplanted cardiac stem cells in vivo by single-photon emission computed tomography or positron emission tomography. J Am Coll Cardiol 2008;52:1652-60
Davis & Stewart
232.
233.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
234.
Terrovitis J, Lautamaki R, Bonios M, et al. Noninvasive quantification and optimization of acute cell retention by in vivo positron emission tomography after intramyocardial cardiac-derived stem cell delivery. J Am Coll Cardiol 2009;54:1619-26 Muraski JA, Fischer KM, Wu W, et al. Pim-1 kinase antagonizes aspects of myocardial hypertrophy and compensation to pathological pressure overload. Proc Natl Acad Sci USA 2008;105:13889-94 Taljaard M, Ward MR, Kutryk MJ, et al. Rationale and design of enhanced angiogenic cell therapy in acute myocardial infarction (ENACT-AMI): the first randomized placebo-controlled trial of enhanced progenitor cell therapy for acute myocardial infarction. Am Heart J 2010;159:354-60
235.
Landmesser U, Drexler H. The clinical significance of endothelial dysfunction. Curr Opin Cardiol 2005;20:547-51
236.
Sherman DL, Loscalzo J. Endothelial dysfunction and cardiovascular disease. Cardiologia 1997;42:177-87
237.
238.
239.
240.
241.
242.
Aicher A, Heeschen C, Mildner-Rihm C, et al. Essential role of endothelial nitric oxide synthase for mobilization of stem and progenitor cells. Nat Med 2003;9:1370-6 Guthrie SM, Curtis LM, Mames RN, et al. The nitric oxide pathway modulates hemangioblast activity of adult hematopoietic stem cells. Blood 2005;105:1916-22 Stewart DJ. Enhanced Angiogenic Cell Therapy -- Acute Myocardial Infarction Trial (ENACT-AMI; NCT00936819). Bethesda, MD: clinicaltrials.gov, 2009. Available from: http://clinicaltrials. gov/ct2/show/NCT00936819 [Last accessed 28 January 2011] Yim EK, Leong KW. Proliferation and differentiation of human embryonic germ cell derivatives in bioactive polymeric fibrous scaffold. J Biomater Sci Polym Ed 2005;16:1193-217 Battista S, Guarnieri D, Borselli C, et al. The effect of matrix composition of 3D constructs on embryonic stem cell differentiation. Biomaterials 2005;26:6194-207 Zimmermann WH, Melnychenko I, Wasmeier G, et al. Engineered heart tissue grafts improve systolic and diastolic
function in infarcted rat hearts. Nat Med 2006;12:452-8 243.
244.
245.
246.
Bhang SH, Lim JS, Choi CY, et al. The behavior of neural stem cells on biodegradable synthetic polymers. J Biomater Sci Polym Ed 2007;18:223-39 Gelain F, Bottai D, Vescovi A, Zhang S. Designer self-assembling peptide nanofiber scaffolds for adult mouse neural stem cell 3-dimensional cultures. PLoS ONE 2006;1:e119 Wang F, Guan J. Cellular cardiomyoplasty and cardiac tissue engineering for myocardial therapy. Adv Drug Deliv Rev 2010;62:784-97 Wang H, Zhou J, Liu Z, Wang C. Injectable cardiac tissue engineering for the treatment of myocardial infarction. J Cell Mol Med 2010;14:1044-55
247.
Zimmermann WH. Embryonic and embryonic-like stem cells in heart muscle engineering. J Mol Cell Cardiol 2010: published online 30 October 2010, doi:10.1016/j.yjmcc.2010.10.027
248.
Boudriot U, Dersch R, Goetz B, et al. Electrospun poly-L-lactide nanofibres as scaffolds for tissue engineering. Biomed Tech (Berl) 2004;49:242-7
249.
Zhang YZ, Venugopal J, Huang ZM, et al. Characterization of the surface biocompatibility of the electrospun PCL-collagen nanofibers using fibroblasts. Biomacromolecules 2005;6:2583-9
250.
Zvibel I, Smets F, Soriano H. Anoikis: roadblock to cell transplantation? Cell Transplant 2002;11:621-30
251.
Giraud MN, Armbruster C, Carrel T, Tevaearai HT. Current state of the art in myocardial tissue engineering. Tissue Eng 2007;13:1825-36
252.
Suuronen EJ, Sheardown H, Newman KD, et al. Building in vitro models of organs. Int Rev Cytol 2005;244:137-73
253.
Liu CY, Westerlund U, Svensson M, et al. Artificial niches for human adult neural stem cells: possibility for autologous transplantation therapy. J Hematother Stem Cell Res 2003;12:689-99
254.
Khor HL, Kuan Y, Kukula H, et al. Response of cells on surface-induced nanopatterns: fibroblasts and mesenchymal progenitor cells. Biomacromolecules 2007;8:1530-40 Expert Opin. Biol. Ther. (2011) 11(4)
255. Karoubi G, Ormiston ML, Stewart DJ, Courtman DW. Single-cell hydrogel encapsulation for enhanced survival of human marrow stromal cells. Biomaterials 2009;30:5445-55 256. Brenner W, Aicher A, Eckey T, et al. 111In-labeled CD34+ hematopoietic progenitor cells in a rat myocardial infarction model. J Nucl Med 2004;45:512-18 257. Kraitchman DL, Tatsumi M, Gilson WD, et al. Dynamic imaging of allogeneic mesenchymal stem cells trafficking to myocardial infarction. Circulation 2005;112:1451-61 258. Zhou R, Thomas DH, Qiao H, et al. In vivo detection of stem cells grafted in infarcted rat myocardium. J Nucl Med 2005;46:816-22 259. Goussetis E, Manginas A, Koutelou M, et al. Intracoronary infusion of CD133+ and CD133--CD34+ selected autologous bone marrow progenitor cells in patients with chronic ischemic cardiomyopathy: cell isolation, adherence to the infarcted area, and body distribution. Stem Cells 2006;24:2279-83 260. Cheng K, Davis D, Li T-S, et al. Magnetic targeting enhances engraftment and functional benefit of iron-labeled cardiosphere-derived cells in myocardial infarction. Circ Res 2010;106:1570-81 261. Consigny PM, Silverberg DA, Vitali NJ. Use of endothelial cells containing superparamagnetic microspheres to improve endothelial cell delivery to arterial surfaces after angioplasty. J Vasc Interv Radiol 1999;10:155-63 262. Pislaru SV, Harbuzariu A, Gulati R, et al. Magnetically targeted endothelial cell localization in stented vessels. J Am Coll Cardiol 2006;48:1839-45 263. Polyak B, Fishbein I, Chorny M, et al. High field gradient targeting of magnetic nanoparticle-loaded endothelial cells to the surfaces of steel stents. Proc Natl Acad Sci USA 2008;105:698-703 264. Singh JP. Enabling technologies for homing and engraftment of cells for therapeutic applications. JACC Cardiovasc Interv 2009;2:803-4 265. Kyrtatos PG, Lehtolainen P, Junemann-Ramirez M, et al. Magnetic tagging increases delivery of circulating progenitors in vascular injury. JACC Cardiovasc Interv 2009;2:794-802
507
Autologous cell therapy for cardiac repair
266. Urbich C, Dimmeler S. Risk factors for coronary artery disease, circulating endothelial progenitor cells, and the role of HMG-CoA reductase inhibitors. Kidney Int 2005;67:1672-6
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
267. Tepper OM, Galiano RD, Capla JM, et al. Human endothelial progenitor cells from type II diabetics exhibit impaired proliferation, adhesion, and incorporation into vascular structures. Circulation 2002;106:2781-6 268. Heeschen C, Lehmann R, Honold J, et al. Profoundly reduced neovascularization capacity of bone marrow mononuclear cells derived from patients with chronic ischemic heart disease. Circulation 2004;109:1615-22 269. Heiss C, Keymel S, Niesler U, et al. Impaired progenitor cell activity in age-related endothelial dysfunction. J Am Coll Cardiol 2005;45:1441-8 270. Vasa M, Fichtlscherer S, Aicher A, et al. Number and migratory activity of circulating endothelial progenitor cells inversely correlate with risk factors for coronary artery disease. Circ Res 2001;89:E1-7 271. Verma S, Kuliszewski MA, Li SH, et al. C-reactive protein attenuates endothelial progenitor cell survival, differentiation, and function: further evidence of a mechanistic link between C-reactive protein and cardiovascular disease. Circulation 2004;109:2058-67 272. Yamanaka S. Pluripotency and nuclear reprogramming. Philos Trans R Soc Lond B Biol Sci 2008;363:2079-87 273. Park IH, Arora N, Huo H, et al. Disease-specific induced pluripotent stem cells. Cell 2008;134:877-86 274. Dimos JT, Rodolfa KT, Niakan KK, et al. Induced pluripotent stem cells generated from patients with ALS can be differentiated into motor neurons. Science 2008;321:1218-21 275. Ebert AD, Yu J, Rose FF Jr, et al. Induced pluripotent stem cells from a
508
spinal muscular atrophy patient. Nature 2009;457:277-80 276. Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 2006;126:663-76 277. Wernig M, Meissner A, Foreman R, et al. In vitro reprogramming of fibroblasts into a pluripotent ES-cell-like state. Nature 2007;448:318-24 278. Loh YH, Ng JH, Ng HH. Molecular framework underlying pluripotency. Cell Cycle 2008;7:885-91 279. Meshorer E, Misteli T. Chromatin in pluripotent embryonic stem cells and differentiation. Nat Rev Mol Cell Biol 2006;7:540-6 280. Li C, Zhou J, Shi G, et al. Pluripotency can be rapidly and efficiently induced in human amniotic fluid-derived cells. Hum Mol Genet 2009;18:4340-9 281. Kunisato A, Wakatsuki M, Kodama Y, et al. Generation of induced pluripotent stem (iPS) cells by efficient reprogramming of adult bone marrow cells. Stem Cells Dev 2009;19:229-38 282. Amoh Y, Kanoh M, Niiyama S, et al. Human hair follicle pluripotent stem (hfPS) cells promote regeneration of peripheral-nerve injury: an advantageous alternative to ES and iPS cells. J Cell Biochem 2009;107:1016-20 283. Golestaneh N, Kokkinaki M, Pant D, et al. Pluripotent stem cells derived from adult human testes. Stem Cells Dev 2009;18:1115-26 284. Park IH, Arora N, Huo H, et al. Disease-specific induced pluripotent stem cells. Cell 2008;134:877-86 285. Takahashi K, Tanabe K, Ohnuki M, et al. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 2007;131:861-72 286. Yu J, Vodyanik MA, Smuga-Otto K, et al. Induced pluripotent stem cell lines
Expert Opin. Biol. Ther. (2011) 11(4)
derived from human somatic cells. Science 2007;318:1917-20 287. Huangfu D, Osafune K, Maehr R, et al. Induction of pluripotent stem cells from primary human fibroblasts with only Oct4 and Sox2. Nat Biotechnol 2008;26:1269-75 288. Maherali N, Sridharan R, Xie W, et al. Directly reprogrammed fibroblasts show global epigenetic remodeling and widespread tissue contribution. Cell Stem Cell 2007;1:55-70 289. Okita K, Ichisaka T, Yamanaka S. Generation of germline-competent induced pluripotent stem cells. Nature 2007;448:313-17 290. Eminli S, Utikal J, Arnold K, et al. Reprogramming of neural progenitor cells into induced pluripotent stem cells in the absence of exogenous Sox2 expression. Stem Cells 2008;26:2467-74 291. Irion S, Nostro MC, Kattman SJ, Keller GM. Directed differentiation of pluripotent stem cells: from developmental biology to therapeutic applications. Cold Spring Harb Symp Quant Biol 2008;73:101-10 292. Ieda M, Fu JD, Delgado-Olguin P, et al. Direct reprogramming of fibroblasts into functional cardiomyocytes by defined factors. Cell 2010;142:375-86
Affiliation Darryl R Davis*1 MD & Duncan J Stewart†2 MD †, *Author for correspondence 1 University of Ottawa Heart Institute, Ottawa, Ontario, K1Y 4W7, Canada 2 Ottawa Hospital Research Institute, 501 Smyth Road, Ottawa, Ontario, K1H 8L6, Canada Tel: +1 613 739 6686; Fax: +1 613 739 6294; E-mail:
[email protected]
Review
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Platelet rich plasma therapies for sports muscle injuries: any evidence behind clinical practice? 1.
Introduction
2.
Methods
3.
Results
4.
Discussion
5.
Conclusion
6.
Expert opinion
Isabel Andia†, Mikel Sa´nchez & Nicola Maffulli †
Research Department, Osakidetza Basque Health Service, 48170 Zamudio, Spain
Introduction: At present, no drugs are available to hasten restoration of muscle function after injury. Platelet-rich plasma (PRP) therapies may help athletes by promoting muscle regeneration. Areas covered: This is a systematic review assessing the evidence base for PRP therapies in the management of muscle injuries. A computerized literature search, citation tracking and hand searching for original studies assessing the effect of PRP therapies on skeletal muscle cell biology, skeletal muscle repair, or regeneration in animals or humans was performed. No randomized trials have studied the merits of PRP injections for muscle healing. Clinical studies indicated that PRP therapies may enhance muscle repair after strain or contusion, and laboratory data indicated that they can enhance diverse aspects of myogenesis. However muscle injuries present a complicated picture that includes many components other than muscle cells, such as blood vessels, connective tissue and neural components. Expert opinion: The field is relevant but under-researched. No PRP formulation has yet displayed proven solid evidence for the stimulation of healing and recovery after sports muscle injuries. Therefore, major issues, including standardization of formulations and application procedures, need to be addressed to inform clinical studies before recommending best practice guidelines. Keywords: platelet-rich plasma, regeneration, skeletal muscle, sport injuries Expert Opin. Biol. Ther. (2011) 11(4):509-518
1.
Introduction
Muscle injuries resulting from extrinsic or intrinsic mechanisms are extremely common in sports, accounting for about 35 -- 45% of all injuries [1], with contact sports and sports that require the production of large eccentric forces presenting the highest risk [2,3]. The vulnerability of athletes [4] to strains and contusions represents a substantial problem for professional players and their clubs. Such injuries involve significant time lost from training and competition. Given the increasing demands of training and competitions, treatment modalities able to accelerate recovery from muscle injuries without adversely affecting recurrence rate whilst minimizing scarring are of paramount consequence. At present, no drugs have been proven to hasten the restoration of muscle function after injury. Therefore, in the absence of any available evidence-based treatments, injection therapies may be an important option to help professional athletes [5]. Among the injected agents are Traumeel (a homeopathic formulation), Actovegin (an amino acid mixture) [6-8] and autologous serum [9] or plateletrich plasma (PRP) [10]. PRP involves the use of the patients’ own proteins to restore tissue integrity and function. Initially, PRP therapies were developed to treat cutaneous ulcers [11], but an increased understanding of the biological properties
10.1517/14712598.2011.554813 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
509
PRP therapies for sports muscle injuries
Article highlights. .
.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
.
In clinical management of muscle injuries, the current hypothesis is that intramuscular injections of platelet-rich plasma (PRP) deliver supraphysiological concentrations of growth factors and cytokines to the injured site, influencing cell migration, proliferation, differentiation or fusion and ultimately enhancing muscle regeneration. Given the biocompatibility of using the patient’s own proteins, safety is guaranteed, simplifying translation from the laboratory to the patient. However the rapidity of translation has sparked debate regarding the level of evidence of clinical benefit needed to introduce PRP technologies in the sports medicine setting. No randomized trials have tested PRP injections in muscle healing, and our systematic search identified only four clinical reports, all of them level 3 or 4 observational studies. Moreover, although laboratory research typically informs clinical studies in this area, in this field basic science experiments were performed simultaneously or even after clinical applications rather than the other way round. The field is relevant but under-researched. Muscle injuries present a complicated picture that includes many components other than muscle cells, such as blood vessels, connective tissue and neural components. PRP therapies are exceptional in that they largely target multiple regenerative processes because of their ability to secrete high levels of the chemokines, cytokines and growth factors, which are required to control activities of different cell types.
This box summarizes key points contained in the article.
of platelets [12,13] and the realization of their healing potential extended the applications to other medical problems [14,15]. Moreover, given the biocompatibility of using the patient’s own proteins, safety is guaranteed, simplifying translation from the laboratory to the patient. PRP therapies may influence muscle regeneration by acting on the satellite cells [16], whose activities are controlled by growth factors and other cytokines, including IGFs, hepatocyte growth factor (HGF), VEGF, basic fibroblastic growth factor (bFGF) or angiopoietin type I (ANGPT-1), plasmin and urokinase plasminogen (uPA) [17-24]. In clinical management of muscle injuries, the current hypothesis is that intramuscular injections of PRPs deliver supraphysiological concentrations of the above-mentioned factors [25-27] at the injured site, influencing cell migration, proliferation, differentiation or fusion and ultimately enhancing muscle regeneration [28]. Assuming this knowledge, PRP therapies hold promise for accelerating muscle healing and returning the elite athlete to competition earlier. However, the rapidity of translation has sparked debate regarding the level of evidence of clinical benefit needed to introduce PRP technologies in the sports medicine setting. We performed an electronic systematic search using comprehensive sources and focusing on the use of PRP in 510
the management of muscle injuries. To gain a more complete understanding from a scientific and medical point of view, we have covered the entire health research spectrum, and, using pre-specified criteria, we have included all potentially relevant articles from laboratory and clinical research. The field is relevant to orthopaedic sports medicine, but under-researched: we aim to define the current status of our knowledge concerning PRP and muscle healing, a necessary task to guide future research efforts and to identify potential implications. 2.
Methods
Search strategy The search strategy had two main components. First, in terms of treatment, we searched using all current names that describe this therapy modality, that is, platelet rich plasma (PRP), platelet-rich fibrin matrix (PRFM), autologous fibrin, autologous conditioned serum (ACS), platelet concentrate (PC), platelet gel (PG), autologous growth factors (AGF), plasma or preparation rich in growth factors (PRGF) and platelet releasate or lysate (PL). The applied search strategy (Table 1) covers all variants of the treatment in review, including materials containing leukocytes such as leukocyte-platelet rich plasma (L-PRP), platelet-leukocyte-rich plasma (P-LRP) or platelet-leukocyte gels (PLG). Secondly, we searched for the target, combining the following terms: skeletal muscle injury, strain or contusion and skeletal muscle healing, repair or regeneration. The applied search strategy in Medline and EMBASE using the OVID platform is displayed in Table 1. Via the Web of Science, searches combining the above key words were performed in the Science Citation Index Expanded (SCIEXPANDED) from 1899-present and in the Conference Proceedings Citation Index - Science (CPCI-S) from 1990 to the present (the first week of October, 2010). Google Scholar was also searched. All seemingly relevant articles and reviews were screened for meaningful references and the retrieved article references were further examined for additional publications. 2.1
Criteria for study consideration and data extraction
2.2
Studies were eligible if they provided specific information related to the effects of PRP therapies (including ACS) in skeletal muscle and if they were original studies assessing the effect of PRP-therapies on skeletal muscle cell biology, skeletal muscle repair or regeneration in animals or humans. Studies focusing on the repair of non-skeletal muscle were not considered. There were no language or data restrictions. Studies were identified by two authors independently. From the included studies, the following data were extracted: study design (descriptive or controlled, laboratory studies, in vitro or in vivo or clinical experimentation), sample type (cell line, primary culture, animal species, number of animals, target population, number of patients), type of PRP product,
Expert Opin. Biol. Ther. (2011) 11(4)
Andia, Sa´nchez & Maffulli
Table 1. Search strategy in EMBASE 1980 to 2010 Week 41, Ovid MEDLINE 1959 to October Week 1 2010, Ovid MEDLINE Daily Update October 15, Ovid MEDLINE in process & other non-indexed citations. No
Search strategy
1 2
(Plasma adj3 (growth factor* or relasate)).mp ((thrombocyte* or platelet*) adj3 (plasma or concentrate* or gel or fibrin* or lysate*)).mp ((Autologous or endogenous or autogenous) adj3 (serum or blood)).mp OR/1 -- 3 (note: combination of terms related to product) (Musc* adj5 (heal* or injur* or strain* or contus* or regener* or repair*)).mp AND/4 -- 5 Remove duplicates from 6
3 4
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
5 6 7
*Truncation, adj3: words in either order between 3 words, mp: title, original title, abstract, subject heading word, name of substance word.
anatomical location of the injured muscle, outcome measures and principal conclusions. Articles that focused on satellite cells treated with PRP for tissue engineering were excluded. 3.
Results
Systematic search for PRP and muscle: identified articles
3.1
Eligibility of the studies based on titles, abstracts and fulltext articles was assessed as shown in Figure 1. Numerous reviews and opinion papers highlight the relevance of PRP treatments in orthopedics and sports medicine [28-37]. However, no randomized trials have tested PRP injections in muscle healing, and our search identified only four clinical reports [9,10,38,39], all of them level 3 or 4 observational studies. Three of these were published reports [9,38,39], and the other was an oral presentation [10]. Other relevant articles were three laboratory experimental reports, two in vivo [40,41] and one in vitro [42] (Tables 2 and 3). All papers were published in English. 3.2
Description of the studies Clinical studies
3.2.1
Wright-Carpenter [9] assessed the effects of ACS injections in a non-blinded, non-randomized case control study. ACS is an autologous liquid serum conditioned by incubation of whole blood with glass beads; it contains signaling proteins that include IL-1b, TNF-a, IL-7, fibroblast growth factor-2 (FGF-2), IL-1 receptor antagonist (IL-1Ra), HGF, platelet derived growth factor (PDGF-AB), TGFb-1 and IGF-1. The experimental group treated with ACS included 17 patients, while the control group, which was analyzed retrospectively, included 11 patients who had received Traumeel/ Actovegin(3:2). Traumeel is a homeopathic formulation containing both botanical and mineral ingredients in
homeopathic concentrations. It is purported to suppress the release of inflammatory mediators and stimulate the release of anti-inflammatory cytokines. Actovegin is a deproteinized calf blood hemodialysate consisting of a physiological mix of amino acids. The rest ice compression elevation protocol was employed for initial care in both groups. The severity of the tear, which was scored as grade 2 with detection of bleeding on MRI, was similar for all control and experimental groups [43]. Most tears were located in the hamstring and adductor muscles (12 in the experimental group and 9 in the control group). The injected volumes (5 ml) were identical in both groups. The injection technique and post-injury treatment are described well. The mean number of treatments per patient was 5.4 in the ACS group and 8.3 in the reference group. The main outcome measured was the time needed to resume full sporting activities. Return to competition was decided after isokinetic strength assessment. The experimental group returned to competition after 16.6 days, while the control group took 22.3 days; in addition, MRI scans taken at 16 days in both groups confirmed that regression of the edema/bleeding was faster in the ACS group. Both treatments were safe. At the 2nd World Congress of Regenerative Medicine, Sanchez et al. [10] reported the application of leukocyte-free PRP [44] in 21 muscle injuries of different severities and at different anatomical locations; small tears progressed well with a single application, while more severe tears required two or three ultrasound-guided injections. The injected volume depended on tear severity. These athletes, who played in first division teams of the Spanish Soccer League, resumed normal training activities in half the time needed by matched historical controls. Using the same leukocyte-free PRP preparation, Wee et al. [38] reported good outcomes (1 week to return to pre-injury activities) after three weekly ultrasoundguided injections to treat adductor longus strain in a professional bodybuilder. Objective measurements, such as swelling or manual muscle testing, were not reported. Pain is always mentioned, but the visual analogue scale or analgesic consumption were not reported as outcome measures. Recently, Hamilton et al. [39] reported buffered L-PRP injection in a grade II hamstring strain injury and daily physiotherapy program. Seventeen days after injury, the patient had full range of motion and was pain free in maximal contraction consistent with MRI demonstrating complete resolution. In vivo controlled laboratory studies Myogenesis relies upon satellite cell activation, proliferation, migration to the site of injury, differentiation, fusion with existing damaged muscle or other satellite-cell-derived myocytes and maturation (increased myofiber diameter). Thus, to assess the progress of muscle regeneration from a biological perspective, researchers measure the number of activated satellite cells, molecular markers of cell differentiation (i.e., RNA or proteins) or the diameter of regenerating myofibers. Using this strategy, two separate research teams [40,41] used syngeneic 3.2.2
Expert Opin. Biol. Ther. (2011) 11(4)
511
PRP therapies for sports muscle injuries
Potentially relevant articles based on search terms: n = 158 Search strategy in EMBASE 1980 to 2010 week 41, Ovid MEDLINE® 1959 to October week 1 2010, Ovid MEDLINE Daily Update October 15, Ovid MEDLINE in process and other non-indexed citations
Articles excluded after screening titles (n = 125)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Potentially relevant articles retrieved for evaluation (n = 33)
Excluded after evaluation of abstract: narrative review, opinion papers (n = 25); tendon/ligament (n = 3)
Search via Web of Science Clinical reports (n = 1) Included studies: Clinical studies (n = 4) Controlled laboratory studies (n = 3) Descriptive laboratory study (n = 1)
Search in Scholar Google Congress communications: Clinical studies (n = 1) Controlled laboratory study (n = 1)
Figure 1. Flow diagram of the systematic literature research. A total of 125 articles were excluded as the title or the abstract clearly indicated that they were not relevant, and 28 articles were further excluded because they were narrative reviews or opinion papers or evaluate tendon/ligament.
animals (mice or rats) to test the therapeutic effects of ACS and L-PRP, respectively. In 2004, Wright-Carpenter et al. [40] applied ACS in a contusion injury model by injecting 10 µl at 2, 24 and 48 h after injury. The control group was treated with the same volume of saline. The number of activated cells was assessed at 30 and 48 h after contusion, and the size of regenerating myofibers was measured in histological samples on days 0, 2, 4, 6, 7, 8, 14, 21, 28 and 35, assessing the progress of regeneration. The number of activated satellite cells was higher in ACS-treated contusions at 30 and 48 h. Moreover, larger regenerating myofibers were observed in the ACS group by days 7 -- 8; however, by day 14, there were no differences between groups. These results indicated that ACS hastens muscle regeneration after contusions by promoting earlier initiation of the activation and/or recruitment of satellite cells and by achieving earlier fusion. Hammond et al. [41] described the effects of leukocyteplatelet-rich plasma (L-PRP) in the treatment of strains. The authors induced either weak or severe strains (by single or multiple repetitions) in the tibialis anteriorior of syngeneic rats and injected 100 µl of L-PRP at days 0, 3, 512
5 and 7. The control group was treated identically but with PPP. Muscle regeneration was assessed by molecular measurements of myogenin and MyoD, measuring both mRNA and protein levels. In addition, myogenesis was assessed by quantification of centrally-nucleated fibers (widely accepted as a marker for muscle regeneration) peaking 2 weeks after injury in the PRP-treated group. Functional recovery was evaluated by torque measurements at days 3, 5, 7, 14 and 21. In weak strains, PRP ameliorated the force loss at day 3, while in more severe strains PRP improved the contractile function at days 7 and 14, and shortened the recovery time from 21 days to 14 days. The authors concluded that L-PRP injections hasten functional recovery and that myogenesis was probably the mechanism underlying this acceleration. In a poster communication [45], muscle lacerations treated with leukocyte-depleted PRP in a sheep model showed enhanced regeneration when compared to platelet-poor plasma. Thus, three independent in vivo studies of different methodological values have assessed the effects of three different autologous preparations injected in three different injury models: contusions, strains and lacerations (Table 3).
Expert Opin. Biol. Ther. (2011) 11(4)
IV
IV
IV
Strain/ hamstring One injection Buffered L-PRP Case report /recreational athlete Hamilton et al. 2010 [39]
ACS: Autologous conditioned serum; L-PRP: Leukocyte platelet rich plasma; PRP: Platelet rich plasma.
Return to competition No re-injuries Return to competition Pain Safety Return to pre-injury activities MRI evaluation Pain Safety Strain or contusion /Different locations Strain/Adductor longus Case-series n = 20, historical controls/professional athletes, Case report /professional athlete Sanchez et al. 2005 (Oral presentation) [10] Wee et al. 2009 (Letter) [38]
Faster return to competition Safety Acceleration of healing Reduced pain Safety Return to pre-injury activities at 3 weeks Full range of motion and MRI resolution at day 17
III Strain/ Hamstring and adductor(most)
ACS versus Traumeel/Actovegin ACS:5.4 injections/patient Control group: 8.3 injections/patient One to three injections Pure-PRP Three injections Pure-PRP Case-control n = 16 (experimental) n = 11 (control group) /recreational athletes Wright-Carpenter et al. 2004 [9] (Original article)
Faster regression of oedema Faster return to competition Safety Regression of the oedema Strength (isokinetic test) Return to competition
Mechanism/ location Treatment Study design/target population
Outcome measures
In vivo controlled laboratory studies Ranzato et al. [42] evaluated proliferation and motility in C2C12 mouse myoblasts treated with platelet lysates. Due to technical difficulties associated with isolating and maintaining cultures of primary satellite cells, immortalized cell lines are frequently used as satellite cell models. C2C12 is a commonly used cell line, isolated from clonal cultures derived from the thigh muscles of 2-month-old C3H mice 70 h after crush injury. To obtain a platelet lysate, platelet pellets are washed, repeatedly frozen and thawed and finally centrifuged to eliminate debris. A 20% platelet lysate was used in these experiments. The authors used a scratch wound model and chemotaxis assays. In scratch models, the wound healing space is reduced by both migration and proliferation of cells. In the chemotaxis model, on the other hand, the effect of migration does not overlap with proliferation. The results showed increased proliferation and motility, and the latter effect was more evident [42]. This is relevant given the isolation and relatively sparse distribution of satellite cells in uninjured tissues; proliferation and directional motility are both required to reach large populations of activated myoblasts at a site of focal injury. This study also provided a mechanistic explanation by demonstrating that activation of p38 and PI3K was involved in the myogenic program (differentiation) in cell motility. Taken together, these studies contained too many variables regarding the product (ACS, L-PRP, pure PRP and platelet lysates), the method of application (variable number of injections, volume, frequency), the type of injury (contusion, strain or laceration), anatomical location and severity. Moreover, although laboratory research typically informs clinical studies in this area, in this field basic science experiments were performed simultaneously or even after clinical applications rather than the other way round. 3.2.3
Clinical studies (type of article)
Table 2. Platelet rich plasma therapies to treat muscle injuries: clinical studies.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Principal results
Level of evidence
Andia, Sa´nchez & Maffulli
4.
Discussion
Although the management of sports injuries with PRP injections has been advocated since 2003 [46], this strategy has not yet been tested in clinical trials dealing with muscle injuries. In reviewing the published work on PRP therapies for muscle injuries, we found only one peer-reviewed clinical study [9] in recreational athletes, and it contained important methodological limitations, such as a lack of blinding, retrospective controls, incomplete reporting and a lack of objective measurements. The absence of studies may impress clinical researchers. This is not so extraordinary for muscle sport injuries, as their management is based largely on experimental studies or empirical evidence. Even when considering the clinical evidence base for the universally-accepted early management of soft tissue injuries, that is ice (also known as cooling or cryotherapy), after meta-analyses [47], conclusions and recommendations were greatly limited and guidelines continue to be formulated on an empirical basis. This presumably reflects not only the importance of key details of the application procedures, such as the interaction of the
Expert Opin. Biol. Ther. (2011) 11(4)
513
514
ACS: Autologous conditioned serum; L-PRP: Leukocyte platelet rich plasma; myoD: Myoblast determination protein; PPP: Platelet-poor plasma; PRP: Platelet-rich plasma.
Enhanced proliferation and motility Motility more central than proliferation P38 and PI3K drive cell migration Percentage wound closure rate Proliferation Chemotaxis Scratch wound closure Antibody blockade Controlled laboratory/ mouse myoblasts C2C12 Ranzato et al. 2009 [42]
Platelet lysate Inhibitors of MAPK signalling ERK, p38, PI3K
Principal conclusions Effect measures Assay type Design/Cell type Cell cultures
Treatment
Controlled laboratory Sheep, n = 4 Controlled laboratory syngenic mice, n = 108 Carda, et al. 2005 [45] Wright-Carpenter et al. 2004 [40]
Pure-PRP versus PPP One application ACS versus saline Three injections
Lacerations over the back Contusion gastrocnemius
Histology: number of activated satellite cell, fiber diameter
Enhanced structural outcome with PRP Enhanced satellite cell activation and larger fiber diameter with ACS
Enhanced functional recovery Stimulation of myogenesis
Percentage of maximal torque mRNA: MyoD and myogenin Histology: centrally nucleated fibers Qualitative histology Strain Tibialis anterior Controlled laboratory/ syngenic rats, n = 72 Hammond et al. 2009 [41]
L-PRP versus PPP Four injections
Injury/anatomical location Treatment Study design/animal Animal studies
Table 3. Platelet rich plasma and muscle repair: laboratory studies.
Outcome measures
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Principal conclusions
PRP therapies for sports muscle injuries
cooling surface with the tissue, but also the major hurdles for developing adequate clinical trials, which include large variations with regard to injury severity and affected muscle groups, non-specificity of reported symptoms, concomitant treatments, allocation of elite athletes into randomized controlled trials and outcome measurements independent of patient motivation. PRP injection is a form of management of muscle injuries that can be considered for clinical practice. However, it is hard to recommend it as best practice, first because it is based on scarce level III -- IV studies and the recommendations of expert opinion; second because PRP therapies are unclear regarding the best formulation for muscle injuries. Essentially, there are two different liquid formulations gathered under the same PRP terminology. To differentiate and define those, two descriptive terms have been proposed [48]: L-PRP, which contains fivefold to -- eightfold more platelets and more leukocytes than peripheral blood; in contrast, P-PRP avoids leukocytes, and has a moderate increase in platelet count (1.5 -- 2.5-fold above baseline). It is not known whether muscle injuries treated with L-PRP or P-PRP progress in different ways. Theoretically, L-PRP may mimic the initial phase of inflammation in which a high number of neutrophils infiltrate the injured site; the interactions of neutrophils with platelets can induce a hyperactive leukotactic response of circulating neutrophils toward the injury site. Neutrophils may exacerbate tissue damage via several different mechanisms (i.e., secreting pro-inflammatory cytokines such as TNF-a, IFN-g, IL-6 or IL-1b) that cause matrix destruction through the production of MMP-1, -3 and -13. Moreover, neutrophils secrete high concentrations of a number of cytolytic and cytotoxic chemicals, such as oxygen radicals and hydrochlorous acid [49]. Consequently, interaction of activated neutrophils with the damaged tissue can and does intensify muscle damage, which is known to be the secondary injury related to the inflammatory response [50]. Indeed, research shows that hindering neutrophil infiltration can result in reduced overall muscle damage [51]. Hence, assuming their probable dissimilarities in neutrophil chemotaxis and activation, L-PRP and P-PRP might be critically different in regulating the complex innate immune response and subsequent healing outcome. To gain further information about those critical differences in the early healing phase, both formulations should be compared, preferably using large-animal models and adequate outcome measurements. Proponents of PRP therapies in muscle applications may offer several arguments in their defense. First, medicine is dynamic, and it is worthwhile to exploit the therapeutic value of an otherwise safe technology that has the potential to benefit patients, as shown in other clinical applications [52,53], even if it will probably be refined as laboratory and clinical research are conducted. Second, while in recreational athletes muscle injuries may recover uneventfully in a matter of weeks, professional athletes need urgent solutions because they must return to higher levels of performance and activities in a
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Andia, Sa´nchez & Maffulli
shorter time. Third, knowledge of repair mechanisms supports the biochemical basis of adding supraphysiological concentrations of growth factors to injured tissues [54]. There are important insights arising from research that may help in understanding the clinical potential of PRPs and their suitability as a therapeutic tool in muscle injuries. Muscle injuries certainly present a complicated picture that includes many components other than muscle cells, such as blood vessels (endothelial cells and pericytes), connective tissue (fibroblasts) and neural components (motor neuron, Schwann cells) [55-57]. In addition, PRP therapies are exceptional in that they largely target multiple regenerative processes because of their ability to secrete high levels of the chemokines, cytokines and growth factors, which are required to control activities of different cell types. It is not known which are the key factors, but several growth factors abundant in PRPs have been extensively studied in muscle regeneration [24]. For example, HGF is the primary component of crushed muscle extract [58], and it is currently the most probable candidate for initiating regeneration by satellite cell activation via c-met receptors. HGF promotes activation, proliferation, differentiation and chemotaxis. IGF-I and -II each increase following muscle injury and promote myoblast proliferation and myofiber differentiation as well as enhancing muscle cell survival and hypertrophy under tissue-specific circumstances. While not as intensively studied as HGF and IGFs, VEGF, bFGF and ANGPT-1 appear to have potential in regeneration by inducing muscle angiogenesis [55]. Also less well-recognized, brain derived neurotrophic factor (BDNF) is a relevant component of PRPs with an important role in regulating satellite cell function and regeneration, as shown in vivo [18]. BDNF has been known since the early 1990s in sports research because, of all neurotrophins, it is the most susceptible to systemic regulation by exercise and physical activity; it is also known because of its metabotropic activity [59]. When axonal communication with the muscle cell body is interrupted by injury, Schwann cells produce neurotrophic factors, such as nerve growth factor (NGF) and BDNF [60]. Thus, additional increases in BDNF in the context of PRPs may help in the progressive recovery of neural communication [61-64]. On the other hand, the presence of relatively high concentrations of TGF-b1 prompts the question of whether PRPs may favor healing by fibrosis instead of regeneration. Both platelets and leukocytes secrete TGF-b1, and there is good reason to think that boosting TGF-b1 levels might induce excessive accumulation of fibrotic tissue [16,65]. However, molecular combinations may be antagonistic or even suppressive, and the combined effect of TGF-b1 with other PRPsecreted molecules on collagen synthesis was weaker than that of TGF-b1 in isolation [66]. Then again, with the release of proteases, such as plasmin or thrombin, PRPs may fuel the fibrinolytic activity required for myogenesis [23]. For example, IGF-binding proteins (IGFBPs) still need to be cleaved to deliver bioactive IGF to its receptor and stimulate cell
activities. Unraveling the protease-induced activation of IGFs, HGF or TGF-b1 may be the key to understanding some PRP actions needed to optimize PRP formulations. Thus, in complex systems such as PRPs, the major challenge is to disentangle the relative effects of the components and to understand how they influence given cell activities. Indeed, the PRP story has turned out to be immensely more complex that it seemed at first. 5.
Conclusion
According to the findings of this review, no PRP formulation has yet displayed proven solid evidence for the stimulation of healing and recovery after sports muscle injuries. Pilot clinical studies along with empirical experience indicate that PRP therapies may enhance muscle repair after strain or contusion. Laboratory data indicate that such treatments can enhance myogenesis. However, the fundamental principles governing when and how PRP therapies can be usefully employed in muscle injuries are emerging at a slow pace. The key to attain standardization and improved formulations will be the identification of crucial elements in these preparations. Given our rudimentary knowledge of the mechanism of action of the PRPs, it remains uncertain how best to use this technology to affect early healing, and produce improved and accelerated functional recovery. 6.
Expert opinion
Currently, the use of PRPs in elite athletes and ensuing discussion in the media has fueled clinical demand outpacing basic and clinical research and hindering progress on such therapies. The ease of use and lack of fear of side and adverse effects involved with PRPs is detrimental, as this allows practitioners to use it frequently without guidelines such as timing of treatment, number and technique of injections or volume; frequently, the personal experience of the practitioner is the only source of evidence to substantiate practice. Failure to understand the mechanism of action of PRPs frustrates efforts to develop best formulations regarding the optimal platelet concentration. Earlier, in oral and maxillofacial surgery, a minimum four-fold to fivefold increase in the number of platelets was considered necessary to produce a therapeutic effect [67]. In retrospect, it is obvious that such an assertion was inappropriate, and not supported by basic science [68,69]. The best PRP formulation for muscle injuries will be clearer after research efforts have provided a comprehensive description of the relations between PRP components, healing mechanisms and functional outcome. In particular, several critical questions about how to optimize PRP therapies should be a high priority for researchers. First, to standardize PRP formulations, research must identify key elements in these preparations. For example, it is relevant to establish differences between pure platelet-rich plasma and leukocyte-platelet concentrates regarding tissue damage
Expert Opin. Biol. Ther. (2011) 11(4)
515
PRP therapies for sports muscle injuries
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
exacerbation [50-57]. In addition, the optimal balance between plasma myogenic factors, such as IGFs and HGF, and platelet-secreted angiogenic or chemotactic factors needs clarification. In fact, platelets are the major source of chemotactic factors such as platelet factor 4 (PF-4) which, in cooperation with PDGF and CXCL7, activates fibroblast migration. Second, to identify the best timing for application, the implications of physicochemical temporal conditions of the tissue (i.e., pH, NO and oxygen) should be evaluated. Indeed, most injured tissues, which are under hypoxic conditions, shift to normoxia after angiogenesis; thus, the biological and clinical effects of PRPs under these circumstances may differ. Moreover, which cells or biological events PRPs target in each temporal phase of repair is unknown [54]. Furthermore, if
reduction of scarring is a plausible goal, it would involve identifying the actions of TGFb-1 in this context. These questions need to be addressed to standardize the formulations and procedures for application. Because of the safety of these products, basic science, clinical discovery and patient-oriented research should be interdependent rather than successive steps. The substantial challenges of incorporating such research into clinical care must be pursued if the potential of PRPs is to be realized.
Declaration of interest The authors state no conflict of interest and have received no payment for the preparation of this manuscript.
Bibliography 1.
2.
3.
4.
5.
6.
7.
8.
516
Mummery WK, Schofield G, Spence JC. The epidemiology of medically attended sport and recreational Injuries in Queensland. J Sci Med Sport 2002;5:307-20 King DA, Hume PA, Milburn PD, Guttenbeil D. Match and training injuries in rugby league a review of published studies. Sports Med 2010;40(2):163-78
Olympic games. Clin J Sport Med 2009;19(1):33-8 9.
10.
Schmikli SL, Backx FJG, Kemler HJ, van Mechelen W. National Survey on Sports Injuries in the Netherlands: target populations for sports injury prevention programs. Clin J Sport Med 2009;19:101-6 Yard EE, Schroeder MJ, Fields SK, et al. The epidemiology of United States high school soccer injuries, 2005-2007. Am J Sports Med 2008;36:1930-7 Orchard JW, Best TM, Mueller-Wohlfahrt H-W, et al. The early management of muscle strains in the elite athlete: best practice in a world with a limited evidence basis. Br J Sports Med 2008;42:158-9 Pfister A, Koller W. Treatment of fresh muscle injury. Spotverletz Sportschaden 1990;4:41-4 Schneider C, Schneider B, Hanisch J, van Haselen R. The role of a homoeopathic preparation compared with conventional therapy in the treatment of injuries: an observational cohort study. Compl Ther Med 2008;16:22-7 Tsitsimpikou C, Tsiokanos A, Tsarouhas K, et al. Medication use by athletes at the Athens 2004 summer
Wright-Carpenter T, Klein P, Schaferhoff P, et al. Treatment of muscle injuries by local administration of autologous conditioned serum: a pilot study on sportsmen with muscle strains. Int J Sports Med 2004;25:588-93 Sanchez M, Anitua E, Andia I. Application of autologous growth factors on skeletal muscle healing. Presented at 2nd World Congress on Regenerative Medicine, May 18 -- 20, 2005, Leipzig, Germany. Available from: http:// www.plateletrichplasma.com/pdf/ Orthopedic-PRP/Sports%20Medicine/ 66-SanchezRegMed2005.pdf [Last accessed 21 January 2011]
11.
Margolis DJ, Kantor J, Santanna J, et al. Effectiveness of platelet releasate for the treatment of diabetic neuropathic foot ulcers. Diabetes Care 2001;24:483-8
12.
Anitua E, Andia I, Ardanza B, et al. Autologous platelets as a source of proteins for healing and tissue regeneration. Thromb Haemost 2004;91:4-15
13.
Nurden AT, Nurden P, Sanchez M, et al. Platelets and wound healing. Front Biosci 2008;13:3832-48
14.
Anitua E, Sanchez M, Nurden AT, et al. New insights into and novel applications for platelet-rich fibrin therapies. Trends Biotechnol 2006;24:227-34
15.
Anitua E, Sanchez M, Orive G, Andia I. The potential impact of the preparation rich in growth factors (PRGF) in different medical fields. Biomaterials 2007;28:4551-60
Expert Opin. Biol. Ther. (2011) 11(4)
16.
Huard J, Li Y, Fu FH. Current concepts review -- muscle injuries and repair: current trends in research. J Bone Joint Surg-Am Vol 2002;84A:822-32
17.
Abou-Khalil R, Mounier R, Chazaud B. Regulation of myogenic stem cell behaviour by vessel cells. Cell Cycle 2010;9:1-5
18.
Clow C, Jasmin BJ. Brain-derived neurotrophic factor regulates satellite cell differentiation and skeletal muscle regeneration. Mol Biol Cell 2010:21:2182-90
19.
Cornelison DDW. Context matters: in vivo and in vitro influences on muscle satellite cell activity. J Cell Biochem 2008;105(3):663-9
20.
Kasemkijwattana C, Menetrey J, Bosch P, et al. Use of growth factors to improve muscle healing after strain injury. Clin Orthop Rel Res 2000;370:272-85
21.
Menetrey J, Kasemkijwattana C, Day CS, et al. Growth factors improve muscle healing in vivo. J Bone Joint Surg-Br Vol 2000;82B:131-7
22.
Smith CW, Klaasmeyer JG, Woods TL, Jones SJ. Effects of IGF-I, IGF-II, bFGF and PDGF on the initiation of mRNA translation in C2C12 myoblasts and differentiating myoblasts. Tissue Cell 1999;31:403-12
23.
Suelves M, Lopez-Alemany R, Lluis F, et al. Plasmin activity is required for myogenesis in vitro and skeletal muscle regeneration in vivo. Blood 2002;99:2835-44
24.
Ten Broek RW, Grefte S, Von den Hoff JW. Regulatory factors
Andia, Sa´nchez & Maffulli
and cell populations involved in skeletal muscle regeneration. J Cell Physiol 2010;224:7-16 25.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
26.
Anitua E, Sanchez M, Orive G, Andia I. Delivering growth factors for therapeutics. Trends Pharmacol Sci 2008;29:37-41 Li JJ, Huang YQ, Basch R, Karpatkin S. Thrombin induces the release of angiopoietin-1 from platelets. Thromb Heamost 2001;85:204-6
27.
Bendinelli P, Matteuci E, Dogliotti G, et al. Molecular basis of anti-inflammatory action of platelet rich plasma on human chondrocytes. Mechanism of NF-kappaB inhibition via HGF. J Cell Physiol 2010;225:757-66
28.
Sanchez M, Anitua E, Orive G, et al. Platelet-rich therapies in the treatment of orthopaedic sport injuries. Sports Med 2009;39:345-54
29.
Alsousou J, Thompson M, Hulley P, et al. The biology of platelet-rich plasma and its application in trauma and orthopaedic surgery. A review of the literature. J Bone Joint Surg Br 2009;91B:987-96
30.
31.
Creaney L, Hamilton B. Growth factor delivery methods in the management of sports injuries: the state of play. Br J Sports Med 2008;42:314-20 Foster TE, Puskas BL, Mandelbaum BR, et al. Platelet-rich plasma from basic science to clinical applications. Am J Sports Med 2009;37:2259-72
32.
Hall MP, Band PA, Meislin RJ, et al. Platelet-rich plasma: Current concepts and application in sports medicine. J Am Acad Orthop Surg 2009;17:602-8
33.
Harmon KG. Muscle injuries and PRP: what does the science say? Br J Sports Med 2010;44:616-17
34.
35.
36.
Linklater JM, Hamilton B, Carmichael J, et al. Hamstring injuries: anatomy, imaging, and intervention. Sem Musculoskeletal Radiol 2010;14:131-61 Ljungqvist A, Schwellnus MP, Bachl N, et al. International Olympic Committee consensus statement: molecular basis of connective tissue and muscle injuries in sport. Clin Sports Med 2008;27:231-9 Mishra A, Woodall J, Vieira A. Treatment of tendon and muscle using platelet-rich plasma. Clin Sports Med 2009;28:113-25
37.
Reider B. Proceed with caution. Am J Sports Med 2009;37:2099-100
38.
Wee LL, Lee DYH, Soon MYH. Plasma rich in growth factors to treat adductor longus tear. Ann Acad Med Singapore 2009;38:733-4
39.
40.
41.
Hamilton B, Knez W, Eirale C, Chalabi H. Platelet enriched plasma for acute muscle injury. Acta Orthop Belg 2010;76:443-8 Wright-Carpenter T, Opolon P, Appell HJ, et al. Treatment of muscle injuries by local administration of autologous conditioned serum: animal experiments using a muscle contusion model. Int J Sports Med 2004;25:582-7 Hammond JW, Hinton RY, Curl LA, et al. Use of autologous platelet-rich plasma to treat muscle strain injuries. Am J Sports Med 2009;37:1135-42
42.
Ranzato E, Balbo V, Boccafoschi F, et al. Scratch wound closure of C2C12 mouse myoblasts is enhanced by human platelet lysate. Cell Biol Int 2009;33:911-17
43.
Jarvinen TAH, Jarvinen TLN, Kaariainen M, et al. Muscle injuries: optimising recovery. Best Pract Res Clin Rheumatol 2007;21:317-31
49.
Tiidus PM. Skeletal muscle damage and repair: classic paradigms and recent developments. J Musculoskeletal Pain 2010;18(4):396-402
50.
Tidball JC. Inflammatory processes in muscle injury and repair. Am J Physiol Regul Integr Comp Physiol 2005;288:R345-353
51.
Tuomi H, Best TM. The inflammatory response: friend or enemy for muscle injury? Br J Sports Med 2003;37:284-6
52.
Dougherty EJ. An evidence-based model comparing the cost-effectiveness of platelet-rich plasma gel to alternative therapies for patients with nonhealing diabetic foot ulcers. Adv Skin Wound Care 2008;21:568-75
53.
Peerbooms JC, Sluimer J, Bruijn DJ, Gosens T. Positive effect of an autologous platelet concentrate in lateral epicondylitis in a double-blind randomized controlled trial platelet-rich plasma versus corticosteroid injection with a 1-year follow-up. Am J Sports Med 2010;38:255-62
54.
Andia I, Sanchez M, Maffulli N. Tendon healing and platelet-rich plasma. Exp Op Biol Therap 2010;10:1415-26
44.
Sanchez M, Anitua E, Azofra J, et al. Comparison of surgically repaired achilles tendon tears using platelet-rich fibrin matrices. Am J Sports Med 2007;35:245-51
55.
Christov C, Chretien F, Abou-Khalil R, et al. Muscle satellite cells and endothelial cells: Close neighbors and privileged partners. Mol Biol Cell 2007;18:1397-409
45.
Carda C, Mayordomo E, Enciso M, et al. Structural effects of the application of a preparation rich in growth factors on muscle healing following acute surgical lesion. Poster presentation at the 2nd International conference on Regenerative medicine. Int J Artificial Organs 2005;28(4):363
56.
Kandler B, Fischer MB, Watzek G, Gruber R. Platelet-released supernatant increases matrix metalloproteinase-2 production, migration, proliferation, and tube formation of human umbilical vascular endothelial cells. J Periodontol 2004;75:1255-61
57.
46.
Sanchez M, Azofra J, Anitua E, et al. Plasma rich in growth factors to treat an articular cartilage avulsion: a case report. Med Sci Sports Exerc 2003;35:1648-52
Smith C, Kruger MJ, Smith RM, Myburgh KH. The inflammatory response to skeletal muscle injury. Illuminating complexities. Sports Med 2008;38:947-69
47.
Bleakley C, McDonough S, MacAuley D. The use of ice in the treatment of acute soft-tissue injury - A systematic review of randomized controlled trials. Am J Sports Med 2004;32:251-61
58.
Bischoff R. A satellite cell mitogen from crushed adult muscle. Develop Biol 1986;115:140-7
59.
Knaepen K, Goekint M, Heyman EM, Meeusen R. Neuroplasticityexercise-induced response of peripheral brain-derived neurotrophic factor. Sports Med 2010;40:765-801
60.
Notterpek L. Neurotrophins in myelination: a new role for a puzzling receptor. Trends Neurosci 2003;26:232-4
48.
Dohan Ehrenfest DM, Rasmusson L, Albrektsson T. Classification of platelet concentrates: from pure platelet-rich plasma (P-PRP) to leucocyte- and platelet-rich fibrin (L-PRF). Trends Biotechnol 2009;27:158-67
Expert Opin. Biol. Ther. (2011) 11(4)
517
PRP therapies for sports muscle injuries
61.
62.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
63.
Elgazzar RF, Mutabagani MA, Abdelaal SE, Sadakah AA. Platelet rich plasma may enhance peripheral nerve regeneration after cyanoacrylate reanastomosis: a controlled blind study on rats. Int J Oral Maxillofac Surg 2008;37:748-55 Farrag TY, Lehar M, Verhaegen P, et al. Effect of platelet rich plasma and fibrin sealant on facial nerve regeneration in a rat model. Laryngoscope 2007;117:157-65
64.
Sariguney Y, Yavuzer R, Elmas C, et al. Effect of platelet-rich plasma on peripheral nerve regeneration. J Reconstr Microsurg 2008;24(3):159-67
65.
Li Y, Foster W, Deasy BM, et al. Transforming growth factor-beta1 induces the differentiation of myogenic
518
cells into fibrotic cells in injured skeletal muscle: a key event in muscle fibrogenesis. Am J Pathol 2004;164:1007-19
Ding XG, Li SW, Zheng XM, et al. The effect of platelet-rich plasma on cavernous nerve regeneration in a rat model. Asian J Androl 2009;11:215-21 66.
Anitua E, Sanchez M, Nurden AT, et al. Reciprocal actions of platelet-secreted TGF-beta1 on the production of VEGF and HGF by human tendon cells. Plast Reconstr Surg 2007;119:950-9
67.
Marx R. Platelet-rich plasma (PRP): what is PRP and what is not PRP? Implant Dent 2001;10:225-8
68.
Weibrich G, Hansen T, Kleis W, et al. Effect of platelet concentration in platelet-rich plasma on peri-implant bone regeneration 2004;34:665-71
69.
Yamaguchi R, Terashima H, Yoneyama S, et al. Effects of platelet rich plasma on intestinal anastomotic healing rats: PRP concentration is a key factor. J Surg Res 2010:published online 2 November 2010, doi:10.1016/j. jss.2010.10.001
Expert Opin. Biol. Ther. (2011) 11(4)
Affiliation
Isabel Andia†1 PhD, Mikel Sa´nchez2 MD & Nicola Maffulli3 MD PhD † Author for correspondence 1 Research Department, Osakidetza Basque Health Service, B Arteaga 107, 48170 Zamudio, Spain 2 Unidad de Cirugı´a Artrosco´pica, UCA ‘Mikel Sa´nchez’, Clı´nica USP-La Esperanza, c/La Esperanza 3, 01002 Vitoria-Gasteiz, Spain 3 Queen Mary University of London, Barts and the London School of Medicine and Dentistry, Center for Sports and Exercise Medicine, Mile End Hospital, 275 Bancroft Road, London E1 4 DG, UK
Review
1.
Introduction
2.
First-line medical treatment of metastatic colorectal cancer
3.
Conclusion
4.
Expert opinion
Cytotoxic triplets plus a biologic: state-of-the-art in maximizing the potential of up-front medical treatment of metastatic colorectal cancer Fotios Loupakis†, Chiara Cremolini, Marta Schirripa, Gianluca Masi & Alfredo Falcone
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
†
University of Pisa, Department of Oncology, Transplants and New Technologies in Medicine, Pisa, Italy
Introduction: Up-front treatment of metastatic colorectal cancer (mCRC) has progressively become more complex during last few years. Nowadays, treatment options range from monotherapies with biologics or traditional chemotherapeutic agents to intensive combinations of chemotherapy plus targeted drugs. Areas covered: This review deals with the results of the most recent firstline trials in the medical treatment of mCRC, with a special focus on recently closed or ongoing trials of intensive concomitant combinations of all active cytotoxics plus a biologic agent. Expert opinion: Combinations of three cytotoxic drugs plus a biologic are under clinical investigation and therefore should not be recommended for routine use, nevertheless preliminary results are promising. The main challenges for the future are not only to demonstrate the real clinical usefulness of intensive approaches, but also to gain the ability of defining prior to treatment which patients will benefit most on the basis of clinical and molecular elements. Keywords: bevacizumab, cetuximab, chemotherapy, colorectal cancer, panitumumab Expert Opin. Biol. Ther. (2011) 11(4):519-531
1.
Introduction
In the last 20 years the median overall survival (OS) of patients with metastatic colorectal cancer (mCRC) has increased from 8 -- 12 months to 18 -- 24 months thanks to the introduction of irinotecan, oxaliplatin and monoclonal antibodies, such as the anti-EGFR cetuximab and panitumumab and the anti-VEGF bevacizumab, and the development of integrated treatment strategies to achieve metastases resection [1]. The availability of this wide variability of therapeutic options offers new possibilities of up-front treatment, but a subsequent question arises: which is the best therapeutic strategy for each patient according to the aim of the treatment, the biology and the clinical characteristics of both tumor and patient? Both sequential and combination chemotherapy can be employed in the treatment of mCRC. The commonly accepted decisional algorythm suggests reserving most active up-front regimens to patients with potentially resectable metastases, in order to achieve secondary resection and long-term disease control. On the other hand, patients with never resectable, widespread disease and no options of further resectability are candidates to receive less intensive regimens and even single agent fluropyrimidine, with the objective of prolonging survival without affecting quality
10.1517/14712598.2011.552882 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
519
Cytotoxic triplets plus a biologic in mCRC
Article highlights. .
.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
.
.
A fluoropyrimidine-based doublet as up-front chemotherapy backbone for the addition of a biologic is a standard-of-care for the vast majority of metastatic colorectal cancer patients. Three-cytotoxics regimen 5-fluorouracil/folinic acid, oxaliplatin, and irinotecan (FOLFOXIRI) might represent a preferrable option when aiming to induce relevant tumor shrinkage, such as for potentially resectable disease or for never resectable, widespread, biologically aggressive, symptomatic disease. The efficacy of biologics in the up-front setting is well-established. The indication of anti-EGFR monoclonal antibodies is restricted to KRAS wild-type patients. Molecular predictors, able to drive the therapeutic decision in KRAS wild-type patients, are currently under investigation. The safety and activity of four-drug regimens, combining the three cytotoxics with a biologic agent, have been recently evaluated in early clinical trials with promising results. Other Phase II and III studies are currently ongoing. Future studies might explore the strategy of concentrating the greatest cytoreductive activity by adopting an intensive four-drug regimen in a short initial phase of the up-front treatment, followed by a maintenance period until progression.
This box summarizes key points contained in the article.
of life [2]. Nevertheless, in our opinion, the use of combined first-line regimens might apply to a wider range of clinical scenarios, so that, for example, an intensive up-front treatment might be the preferred option also for patients with symptomatic and widespread, aggressive disease, while the sequential approach should be actually reserved for patients unfit for combinations, due to age or relevant comorbidities. However, this way of modeling doesn’t fit the complexity of clinical practice, so that the choice of the intensity of the up-front chemotherapy might be, nowadays, influenced by multiple clinical and biological considerations. Moreover, such increasing complexity is further complicated by both the availability of biological drugs and the soaring insights into molecular determinants. In fact, in the last few years, clinical trials have shown that treatment with two cytotoxics in association with the antiEGFR [3-5] or the anti-VEGF [6,7] monoclonal antibodies (mAbs) is safe, active and effective, but at the moment, we don’t have a scientific demonstration concerning the best chemoterapeutic regimen to be associated with a biologic, nor the best biologic to be associated with chemotherapy. The biomolecular characterization of mCRC has gained a determinant role: the presence of KRAS mutation is a well-ascertained predictive factor of resistance to cetuximab [3,4,8,9] and panitumumab [5,10] and the presence of BRAF mutation defines a subgroup of patients with extremely bad prognosis [9,11,12]. 520
Hence, in the era of target therapies and of molecular selection, one wonders which settings might be the most suitable for an intensive first-line therapy as a three-drug regimen and ongoing clinical trials are evaluating the opportunity for adding biological agents to the triplet. The objective of the present review is to rapidly summarize evidence from literature about the efficacy of up-front combined regimens, with particular regard to the triplet, and to secondly explore the potential role of the association of a biologic to such an active regimen, in an attempt to define the clinically and molecularly selected population, that might benefit as more as possible from an intensive up-front strategy.
First-line medical treatment of metastatic colorectal cancer
2.
The choice of up-front chemotherapy: two is (nearly always) better than one
2.1
A nodal issue in the choice of the up-front treatment for mCRC patients concerns how intensive the chemotherapy regimen should be. It has been firstly demonstrated [13] and then confirmed [14] by Grothey et al., that the exposure to all cytotoxics during the course of the disease determines a significant improvement in OS. The striking positive correlation of the percentage of patients treated with 5-fluorouracil, oxaliplatin and irinotecan at some point of their disease with OS strongly supports the strategy of making all active agents available to patients, in order to maximize their survival. All Phase III randomized trials, conducted with the aim to compare sequential with combined approaches, achieved analogous results. The capecitabine, irinotecan, and oxaliplatin in advanced colorectal cancer (CAIRO) trial [15] randomized 820 patients to receive first-line capecitabine, followed by second-line irinotecan and third-line capecitabine plus oxaliplatin (sequential strategy) or first-line capecitabine plus irinotecan, followed by second-line capecitabine plus oxaliplatin (combination strategy). The Fluorouracil, Oxaliplatin, and CPT-11 (irinotecan): Use and Sequencing (FOCUS) trial [16] randomized 2135 mCRC patients, not amenable to curative strategy, to first-line single-agent 5-fluorouracil (FU) until failure followed by single-agent irinotecan (strategy A) or 5-FU until failure followed by combination therapy (strategy B), or up-front combination chemotherapy (strategy C). In both strategy B and C, patients were further divided in a 1:1 ratio to receive 5-FU plus oxaliplatin or 5-FU plus irinotecan as combination treatment. The study, initially launched to establish the superiority of one of the strategies in terms of 2 year-OS, was then amended as a consequence of positive results of trials of 5-FU-based irinotecan [17-19] or oxaliplatin doublets [20-22] versus 5-FU monotherapy and, before completion of accrual, a supplementary analysis was planned to examine the non-inferiority of strategy B in comparison with strategy C. The main results of the CAIRO and FOCUS trials are summarized in Table 1.
Expert Opin. Biol. Ther. (2011) 11(4)
Loupakis, Cremolini, Schirripa, Masi & Falcone
Table 1. Comparisons of staged versus combination approaches in FOCUS and CAIRO studies: main measures of outcome. Strategy
FOCUS (2135 patients) First line 5FU A = 710 patients; B = 712 patients
Response rate PFS (months)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
OS (months) Exposure to 3 cytotoxic agents Safety QoL overall
CAIRO (820 patients)
First line combination C-ir = 356 patients; C-ox = 357 patients
28% (A, B)
49% (C-ir); 57% (C-ox) p < 0.001z 6.3 (A, B) 8.5 (C-ir); 8.7 (C-ox) p < 0.001z 13.8 (A); 15.1 (B) 16.7 (C-ir); 15.4 (C-ox) C-ir versus A HR = 0.84; p = 0.01§ 16% (A); 19% (B) 33% No relevant differences No difference
Sequential Strategy 410 patients
Combination strategy 410 patients
20%
41% p < 0.0001
5.8 16.3 36%
7.8 HR = 0.77; p = 0.0002 17.4 HR = 0.92; p = 0.328 53% No relevant differences No difference
*A: single-agent 5-fluorouracil (5-FU) until failure followed by single-agent irinotecan; B: 5-FU until failure followed by combination therapy (5-FU with oxiplatin or irinotecan); C: up-front combination chemotherapy (5-FU with oxiplatin (C-ox)or irinotecan (C-ir). z for both C-ir versus A + B and C-ox versus A + B. § mOS for C (C-ir + C-ox) = 15.9; p = 0.02 for A versus C (level of significance for multiple comparisons = p < 0.01.
In both studies the up-front combined approach did not produce an advantage in OS, except for patients treated with first-line 5-FU and irinotecan in strategy C of the FOCUS trial, compared with those randomly assigned to strategy A (hazard ratio (HR): 0.84, 95% CI 0.73 -- 0.96, p = 0.01). On the other hand, the up-front combined approach warranted significantly higher objective response rate (RR) (41 versus 20%, p < 0.0001 in the CAIRO trial; 49 -- 57% versus 28%, p < 0.001 in the FOCUS trial) and progression free survival (PFS) (7.8 versus 5.8 months, HR: 0.77, p = 0.0002 in the CAIRO trial; 8.7 versus 6.3 months for first-line 5-FU plus oxaliplatin versus 5-FU monotherapy, p < 0.001 and 8.5 versus 6.3 months for first-line 5-FU plus irinotecan versus 5-FU monotherapy, p < 0.001 in the FOCUS trial). Although such trials did not demonstrate a better OS for patients receiving up-front combined chemotherapy, thus leading authors to suggest the staged approach as a valid choice for patients with extensive disease, some crucial points probably need deeper analysis. First of all it should be noted that both trials reported median survivals in the range between 13.9 and 17.4 months, that is relevantly shorter than expected even for combination arms, if compared with other contemporaneous experiences with first-line doublets. Secondly, one of the most attractive facets of a staged approach certainly lies in the possibility of limiting treatment-related toxicities, thus allowing guarantee of the preservation of a better quality of life. However, in both trials while oxaliplatin- and irinotecan-related adverse events were obviously more frequent among patients in the combination arms, no significant differences in the occurrence of grade
3 -- 4 toxicities over all lines, or in the incidence of deaths due to toxicity was evidenced between staged and combined strategies. Also in terms of quality of life, no significant differences in the perception of health status were detected. Thirdly, the percentage of patients exposed to all three cytotoxics was considerably lower in sequential arms in both trials (36 versus 53% in CAIRO and 16 and 19% versus 33% in the FOCUS trial). Such a finding is easily explained by considering that approximately only 50 -- 70% of patients starting a line of therapy will be suitable to receive a next-line treatment. Last, but not least, the item of patients’ selection deserves to be emphasized. As underlined by authors themselves [23], both CAIRO and FOCUS trials included poor prognosis patients’ populations. Patients with liver-only metastases, with a chance to achieve secondary resection, were excluded from FOCUS and underrepresented in CAIRO. Such consideration explains the short survivals registered in both trials and widens the way to the crucial importance of the choice of the best strategy when planning mCRC patients’ treatment from the very beginning. In the light of data from the CAIRO and FOCUS trials, it appears, therefore, mandatory to choose a combined up-front treatment for patients with marginally or potentially resectable metastases. However, taking into account both the low percentage of patients exposed to all three drugs, which is linearly related to OS, and the absence of a clear advantage in terms of safety and quality of life for the sequential strategy, combination therapy might be considered as a reasonable standard of care also for the majority of never-resectable patients, thus reserving first-line monotherapy to patients clearly unfit for combination therapy, due to age and/or
Expert Opin. Biol. Ther. (2011) 11(4)
521
Cytotoxic triplets plus a biologic in mCRC
comorbidities or with a reluctant attitude to accepting an increased risk of toxicity in a first-line treatment. The choice of up-front chemotherapy: when three is better than two
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
2.2
A diametrically opposite approach to mCRC consists of the combined administration of all active cytotoxics as up-front treatment. The schedule developed by the Gruppo Oncologico Nord Ovest (GONO) [24,25] allowed to achieve, in a Phase III randomized trial, remarkable results in terms of both activity and efficacy, with an acceptable increase in adverse events, that did not compromise treatment’s feasibility and safety [26]. Compared with first-line Folinic Acid, Fluorouracil & Irinotecan (FOLFIRI), up-front GONO-5-FU/folinic acid, oxaliplatin, and irinotecan (FOLFOXIRI) resulted in higher RR (60 versus 34%, p < 0.0001), PFS (median PFS: 9.5 versus 6.6 months, HR: 0.59, p < 0.001) and OS (median OS: 23.4 versus 16.7 months, HR: 0.74, p = 0.026) in a population of 244 untreated mCRC patients, deemed initially unresectable. The superior activity for the experimental arm resulted in a significantly higher rate of secondary resections. Of patients treated with GONO-FOLFOXIRI 15% underwent radical surgery on metastases, compared with 6% of patients treated with FOLFIRI (p = 0.033). Such percentages rise to 36 and 12% respectively, considering patients with liver-only metastases. Another Phase III randomized trial, coordinated by the Hellenic Oncology Research Group (HORG) [27], compared the three-drug regimen HORG-FOLFOXIRI to FOLFIRI in a population of 285 untreated mCRC patients. Although the trial failed to demonstrate any superiority for the experimental arm, some improvements for the triplet were reported in terms of RR (43 versus 33.6%, p = 0.168), time to progression (TTP) (median TTP: 8.4 versus 6.9 months, HR: 0.83, 95% CI 0.64 -- 1.08, p = 0.17) and OS (median OS: 21.5 versus 19.5 months, HR: 0.86, 95% CI 0.64 -- 1.16, p = 0.337). The secondary resection rate was higher among patients treated with HORG-FOLFOXIRI (10 versus 4%, p = 0.08). However, at least two main facets should be taken into account when globally interpreting results. Firstly, different schedules of triple-drug regimens were adopted in these studies: while in GONO-FOLFOXIRI, 5-FU was administered at a dosage of 3200 mg/m2 as a continuous 48-h infusion and 5-FU bolus was abolished, in HORG-FOLFOXIRI, 5-FU was administered both on days 2 and 3 as continuous 22-h infusion at a dosage of 600 mg/m2/day and as bolus at a dosage of 400 mg/m2/day. Irinotecan and oxaliplatin planned doses were considerably higher in the GONOFOLFOXIRI versus HORG-FOLFOXIRI (irinotecan: 165 versus 150 mg/m2; oxaliplatin: 85 versus 65 mg/m2). Secondly, study populations were selected according to slightly but relevantly different criteria. While patients aged > 75 years, as well as patients aged 70 -- 75 and with a performance status (PS) ‡ 1, were not included in GONO trial, all 522
patients aged ‡ 18 and with PS £ 2 were eligible for the HORG trial. As a potential consequence of such discrepancies, in the HORG trial, the compliance to FOLFOXIRI was significantly worse compared with FOLFIRI, with a higher percentage of courses being delayed (8.3 versus 14%, p = 0.04) and dose reductions (7 versus 3%, p = 0.001). Both dose reductions and treatment delays were more frequent in the group of elderly patients and, among them, in those treated with HORG-FOLFOXIRI [28]. Despite these major criticisms, in the HORG trial the three-drug combination showed a trend toward better TTP and RR and a more than doubled secondary resection rate. In the metanalysis by Golfinopoulos et al. [29], GONO and HORG trials have been comprehensively analyzed, demonstrating that the addition of oxaliplatin to first-line 5-FU and irinotecan provided a significant advantage in terms of both PFS (HR: 0.73, 95% CI 0.55 -- 0.95) and OS (HR: 0.79, 95% CI 0.63 -- 0.98). With particular regard to secondary surgery, the pooled analysis of patients treated with GONO-FOLFOXIRI in two Phase II and in the Phase III trial revealed that 37 out of 196 initially unresectable patients achieved radical resection of metastases, with a ‘rescue rate’ of 19% [30]. Notably, after a median follow up of 67 months, in the group of radically resected patients, 5-year and 8-year survivals were 42 and 33% respectively. At 5 years, 29% patients were free of disease. For patients undergoing secondary hepatic resection, an increasing amount of evidence supports the relevance of pathologic complete response (pCR) as a meaningful endpoint, significantly related to longer survival [31,32]. In particular, in the series presented by Adam et al. [33], 29 out of 767 (4%) patients with liver metastases, who underwent radical liver resection after systemic chemotherapy, achieved pCR. Three- and five-year survivals for patients with pCR were 91 and 76%, respectively, and were significantly higher when compared with patients without pCR (61 and 45%, respectively, p = 0.004). Ten-year survivals were 68 and 29% with and without pCR, respectively. After a median follow-up of 52.2 months, less recurrences occurred in patients with pCR (41 versus 62% for patients without pCR, p = 0.03). A noteworthy percentage (11%) of pCRs was reported in the group of 37 patients who underwent secondary R0 resection after FOLFOXIRI. Such results strongly support the choice of FOLFOXIRI regimen as a very active ‘conversion’ therapy, able to induce relevant tumoral shrinkage and, therefore, particularly appropriate for patients with potentially resectable metastases, in order to provide them not only a chance of resection [34], but also a chance of cure [35,36], absolutely unexpected until a few years ago. Nevertheless, the setting of patients with potentially or marginally resectable metastases does not represent the sole context, in which the choice of a highly active regimen might represent the preferred option.
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Loupakis, Cremolini, Schirripa, Masi & Falcone
In fact, in the Phase III GONO trial, excluding from the analysis patients who had undergone radical surgery on metastases, the FOLFOXIRI arm retained a significant advantage in PFS (median PFS: 9.5 months versus 6.6 months; HR: 0.59, 95% CI 0.45 -- 0.76, p < 0.0001) and a trend toward longer OS (median OS: 20.2 months versus 15.9 months, HR: 0.80, 95% CI 0.61 -- 1.05, p = 0.12) [37]. With regard to the wide spectrum of clinically and biologically different diseases included in the definition of ‘never resectable’ mCRC, a significant tumoral shrinkage should represent the major aim also in those patients with widespread, aggressive, symptomatic diseases, sometimes responsible for a deep deterioration of patients’ quality of life. In fact, although clinical trials typically enroll a small percentage of poor-prognosis patients, Sargent et al. [38] have observed in a pooled analysis of nine studies in the firstline metastatic setting, that the relative benefit of combination regimens was the same in PS 2 patients compared with PS 0 -- 1 patients, both in terms of survival (HR: 0.89, 95% CI 0.82 -- 0.96, p = 0.003 in PS 0 -- 1 patients and HR: 0.79, 95% CI 0.62 -- 0.99, p = 0.04 in PS 2 patients, with p = 0.18 for PS--treatment interaction) and likelihood of response (odds ratio (OR): 2.71, 95% CI 2.34 -- 3.14, p = 0.0001 in PS 0 -- 1 and OR: 2.85, 95% CI 1.61 -- 5.02, p = 0.0003 in PS 2, with a p = 0.71 for a PS--treatment interaction). Although PS 2 patients presented an increased risk of toxicities and 60-day mortality, it should be noted that a significant treatment--PS interaction is lacking in toxicity analysis. Consistently with above reported results, in the Phase III GONO trial, among patients defined as ‘high-risk’ according to the Kohne scoring system, those treated with FOLFOXIRI achieved significantly longer PFS (median PFS: 8.3 versus 4.4 months, HR: 0.44, 95% CI 0.24 -- 0.82) and longer, although not significantly, OS (median OS: 14.1 versus 11.7 months, HR: 0.78, 95% CI 0.43 -- 1.41) [37]. Nowadays, the relevance of molecular determinants in driving therapeutic decisions is well-recognized. KRAS mutations burst on the scene of mCRC, leading not only to a more accurate selection of patients to be treated with anti-EGFR monoclonal antibodies, but also to launching a sort of ‘molecular revolution’, with a strong influence on physicians’ mental attitudes. At the same time, a considerable amount of data have firstly suggested and then corroborated the awful effect of BRAF mutation as a poor prognostic factor for mCRC patients [11,39]. A soaring amount of data about tumors’ biology suggests that BRAF-mutated tumors represent an almost homogenous group with peculiar genotypic and also phenotypic features [40-44], altogether responsible for their aggressive behavior and poor prognosis. As a consequence of these attainments of knowledge, beside clinical and biochemical parameters, such as PS, number of metastatic sites, white blood cell count, serum alkaline phosphatase and lacticum dehydrogenase, hemoglobin levels and time to metastases, the knowledge of BRAF mutational status
has acquired an important meaning as a relevant tool to better estimate tumor aggressiveness. Though in the absence of specific data from prospective trials, the choice of an intensive and very active first-line regimen might represent an appropriate option also for patients with BRAF-mutated disease. Therefore, the presence of major clinical, but also molecular prognostic indicators of marked aggressiveness might influence the choice of the treatment toward an intensive first-line regimen, such as FOLFOXIRI, with the aim of rapidly reducing tumor burden, thus potentially improving patients’ symptoms and prolonging survival. Increasing complexity: the achievement of biologics
2.3
Three biologic agents have entered the clinic for the treatment of mCRC during the last five years. All them are monoclonal antibodies belonging to two distinct classes: anti-EGFRs (cetuximab and panitumumab) and anti-VEGF (bevacizumab). Summarizing the main findings of various clinical studies [4,6,7,45,46] we can argue that VEGF inhibition with bevacizumab is an effective strategy in combination with chemotherapy in the first- and second-line, while the anti-EGFRs have been demonstrated to be active and efficacious even in the third-line as monotherapy in chemorefractory patients. Looking at the clinical effect of these molecules, especially in terms of OS, it is evident that it hasn’t been as huge as expected while their costs are certainly not negligible considering both side-effects and economic expenses. Notwithstanding these relatively small clinical achievements, the study and the use of these drugs has dramatically changed our way of looking to mCRC from a biomolecular point of view [47]. It is nowadays globally accepted [2,48,49] that mCRC should be categorized according to the mutational analysis of the KRAS oncogene since this feature precludes patients from deriving any benefit with anti-EGFRs. The first assessment of KRAS mutations as predictors of resistance to cetuximab derives from a small retrospective analysis [50]. This initial suggestion has been verified and confirmed in randomized Phase III studies aimed at demonstrating anti-EGFRs’ efficacy across different lines of treatment [3,4,8,10]. The Cetuximab combined with irinotecan in first-line therapy for metastatic colorectal cancer (CRYSTAL) first-line trial of FOLFIRI plus or minus cetuximab, aimed at verifying an improvement in PFS with the addition of the mAb. The experimental arm gained a 15% relative risk reduction for progression (HR: 0.85, 95% CI 0.72 -- 0.99, p = 0.048) in the intention-totreat population. Looking at the results according to KRAS mutational status it is clearly evident that only wild-type patients benefited (HR for PFS: 0.68; 95% CI 0.50 -- 0.94, p = 0.02) while for KRAS-mutant patients there was no advantage (HR: 1.07, 95% CI 0.71 -- 1.61, p = 0.75) [4]. Similar results have been obtained in the Phase II randomized Oxaliplatin and Cetuximab in First-Line Treatment of mCRC (OPUS) trial that compared folinic acid leucovorin and
Expert Opin. Biol. Ther. (2011) 11(4)
523
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Cytotoxic triplets plus a biologic in mCRC
oxiplatin (FOLFOX)-4 plus cetuximab to chemotherapy alone. Objective response rate (ORR) was the primary endpoint. In the overall population the addition of cetuximab marginally improved the ORR (46 versus 36%, OR: 1.52, 95% CI 0.975 -- 2.355, p = 0.064), but among KRAS-wildtype patients the difference was quite wider (61 versus 37%, OR:2.54, 95% CI 1.238 -- 5.227, p = 0.011) while KRAS mutant patients even experienced a detrimental effect of the anti-EGFR (33 versus 49%, OR: 0.507, 95% CI 0.223 -1.150, p = 0.106) [3]. Analogous results have been obtained with the combination of panitumumab with FOLFOX in a recently published Phase III randomized trial [51]. The pooled analysis of the two above mentioned studies with cetuximab provided a clear estimation of the effect of adding it to first-line chemotherapy in KRAS-wild-type patients: EGFR inhibition doubled the odds of achieving a response (OR: 2.16, 95% CI 1.64 -- 2.86, p < 0.0001), significantly reduced the risk of disease progression by 34% (HR: 0.66, 95% CI 0.55 -- 0.80, p < 0.0001) and improved OS (HR: 0.81, 95%CI 0.69 -- 0.94, p = 0.006) [9]. This last finding in terms of OS is strikingly similar to that obtained in a meta-analysis of five randomized first-line trials looking at the efficacy of bevacizumab in addition to chemotherapy in molecularly unselected patients (HR: 0.81, 95% CI 0.73 -- 0.90, p < 0.0001) [52]. It should be noted in fact that bevacizumab exerts its effects independently from KRAS mutational status [53]. In the meta-analysis, even in terms of PFS the results with the anti-VEGF in the overall population are similar to those obtained with cetuximab in KRAS-wildtype patients (HR: 0.61, 95% CI 0.45 -- 0.83, p = 0.002), but the overall effect of bevacizumab in improving tumoral shrinkage is less evident (OR: 1.41, 95%CI 0.92 -- 2.15, p = 0.11) and doesn’t reach statistical significance. This last observation coupled with another retrospective analysis comparing the waterfall plot of two randomized Phase II studies [54] of capecitabine (Xeloda) plus oxaliplatin (XELOX) or capecitabine plus irinotecan (XELIRI) with cetuximab [55] or bevacizumab [56] led some authors to suggest anti-EGFR inhibition as a more productive strategy when response is the primary objective [57]. In fact, in that analysis the activity of cetuximab as measured by the average and the median reduction in tumor burden (ARTB and MRTB) seemed to be greater than that of the anti-VEGF (ARTB 38.9 versus 29.9%, MRTB 35 versus 30%; p = 0.009). In the absence of head-to-head comparisons of bevacizumab with the antiEGFRs all retrospective and indirect conclusions should be interpreted cautiously and first-line decision-making should be driven also by clinical considerations that take into account toxicity profiles, patients’ preferences, and the global stategy to be pursued across subsequent lines [58]. In oncology practice the most reliable and unquestionable end-point to which all efforts are aimed at is and shall be always overall survival. The increasing complexity of the clinical scenario of mCRC patients cleared the way for new end-points potentially indirectly related to OS, that could be 524
important for research but still don’t have sure clinical implications. It has been suggested, for example, that bevacizumab may improve the pathologic response in liver metastases [59,60]. On the other hand recent studies questioned the reliability of traditional response rate as good estimator of bevacizumab’s activity and the radiological evaluation of tumor response seems to be more accurate when derived from the observation of lesions’ morphological changes according to new evaluation criteria [61]. In the attempt to further refine our knowledge about mCRC the search for other useful molecular markers progressively gained more and more attention, especially after the unforeseen demonstration that the concomitant use of upfront anti-EGFRs with bevacizumab plus chemotherapy is futile [62,63] and the question of which antibody may be better for KRAS-wild-type patients came to the fore. Researchers looked at effectors of EGFR signaling pathway other than RAS as possible predictors to maximize the benefit from EGFR inhibitors. Most interesting results derived from BRAF mutational status. Such mutations are mutually exclusive with those of KRAS. After the first description of the possible role as negative predictor of anti-EGFRs’ activity for the BRAF-mutated gene [64], also other confirmatory experiments were conducted [65]. The vast majority of data was derived from patients in advanced lines of treatment. The widest analysis has been recently published by De Roock et al., who collected samples from 649 chemorefractory patients treated with cetuximab plus chemotherapy at 11 European centers. Among 350 KRAS-wild-type patients assessed for response, BRAF mutants had a significantly lower response rate (8.3% (2 out of 24) versus 38.0% (124 out of 326), OR: 0.15, 95% CI 0.02 -- 0.51, p = 0.0012), shorter PFS (median 8 versus 26 weeks; HR: 3.74, 95% CI 2.44 -- 5.75, p < 0.0001) and OS (median 26 versus 54 weeks; HR: 3.03, 95% CI 1.98 -- 4.63, p < 0.0001) [66]. No data from the randomized trials of anti-EGFR monotherapy versus best supportive care with respect to BRAF mutational status have been published yet. As regards first-line of treatment, given the relative rarity of BRAF mutation, data from randomized trials of chemotherapy plus or minus anti-EGFRs are inconclusive due to their low statistical power [67]. Many other trials looked at the pure prognostic effect of BRAF mutational status independently from treatment with anti-EGFRs. As mentioned elsewhere in this review all data consistently confirm that BRAF-mutant tumors have an extremely bad prognosis in the metastatic phase. Due to the strength of such observations and their possible implications special attention has been recently given to the molecular profile of BRAF-mutant CRC, suggesting that such tumors have a specific gene expression pattern especially when BRAF mutation occurs in microsatellite-stable tumors [42]. As suggested in the previous paragraph, in the near future clinicians will want to know what would be the best approach to face this new category of aggressive mCRC with traditional treatments whilst waiting for new targeted drugs.
Expert Opin. Biol. Ther. (2011) 11(4)
Loupakis, Cremolini, Schirripa, Masi & Falcone
Triplet plus biologics: preliminary findings and ongoing trials
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
2.4
The clinical relevance of up-front combined regimens, with particular reference to the results of the GONO-FOLFOXIRI regimen and to the more and more extensive amount of data confirming targeted therapies’ safety and efficacy, have been interpreted as promising foundations for developing even more intensive combination schedules, including threedrug regimens, combined with a biologic agent. Although no conclusive findings are available to date, here we look into preliminary evidence and current experiences. A list of ongoing trials is included in Table 2. A Phase II trial (FOIB study), conducted by the GONO group, has assessed the safety and activity of the combination of GONO-FOLFOXIRI regimen plus bevacizumab in previously untreated, unresectable mCRC patients [68]. According to a Phase II single-stage Fleming design, assuming a null hypothesis of 10 months-progression free rate (10m-PFR) of 50% and an alternative hypothesis of 10m-PFR of 70%, with alpha and beta-errors of 0.05 and 0.10, the experimental treatment would have been judged to be promising if at least 33 patients, out of 53 evaluable, had been free of progression at 10 months. At a median follow-up of 28.8 months, 42 (74%) out of 57 treated patients were actually free of progression at 10 months, with a median PFS of 13.1 months and a median OS of 30.9 months (Figure 1). In terms of activity, promising results were reported, with a RR of 77% and a disease control rate of 100%. Such a considerable activity translated into a radical resection rate of 26%, rising to 40% among patients with liver-only metastases. A pCR was observed in the 20% of patients who underwent radical resection. The safety profile was absolutely consistent with expected toxicities and no unforeseen adverse events were reported. Such results gain higher prominence when considering clinical characteristics of the study population, including a high percentage of patients with synchronous metastases (86%) and a relatively underrepresented proportion of patients with liver-only disease (53%). Consistently with such poor prognostic indicators, a higher than expected frequency of BRAF mutations (18%) was reported. The interpretation of study results according to KRAS and BRAF mutational status revealed no differences in terms of both PFS and OS between patients with KRAS-wild-type and KRAS-mutated tumors, or, even more relevantly, between patients with BRAF wild-type and BRAF mutated tumors. Also patients with BRAF-mutated tumors, in fact, achieved meaningful results in terms of survival (median PFS: 12.8 months and median OS: 23.8 months), which did not significantly differ from results for the BRAF-wild-type subgroup (HR for PFS: 0.89, 95% CI 0.41 -- 1.91, HR for OS: 0.76, 95% CI 0.26 -- 2.21). Though keeping in mind the small sample size and the retrospective nature of considerations about the BRAF-mutated subgroup, one could argue that the adoption of an intensive
regimen might allow containment of the aggressive behavior of these peculiar diseases. Based on promising results of the Phase II FOIB trial, a Phase III multicenter study, comparing FOLFOXIRI plus bevacizumab versus FOLFIRI plus bevacizumab (Triplet plus Bevacizumab (TRIBE) trial), is currently ongoing [69]. Preliminary safety data from the first 150 enrolled patients confirmed the safe profile reported in the FOIB trial, without revealing unexpected toxicities or significant differences between arms, except for oxaliplatin-specific adverse events [70]. Such forthcoming results will probably clarify the effect of the up-front triplet plus bevacizumab, potentially throwing light also on clinical and molecular features, useful for defining the population more likely to benefit from such an intensive regimen. A Phase II randomized trial is currently evaluating the safety and efficacy of first-line FOLFOX plus bevacizumab and FOLFOXIRI plus bevacizumab in a population of mCRC patients with liver-only disease, in order to explore the potentiality of such regimen in a ‘neoadjuvant’ setting [71]. Similarly, another Phase II trial, that adopts surgically complete resectability as the primary endpoint, is evaluating the activity of FOLFOXIRI plus bevacizumab in a population of mCRC patients with not optimally resectable liver or lung metastases [72]. A less extensive amount of data has been published until now about the safety and efficacy of the combination of a three-drug regimen with an anti-EGFR monoclonal antibody. A Phase I dose-escalating study has explored the safety of first-line FOLFOXIRI (according to the GONO schedule) plus cetuximab, revealing neutropenia and diarrhea as the most common treatment-related toxicities and identifying 125 mg/m2 as the recommended irinotecan dose to be further investigated in following experiences. Besides treatment’s feasibility, the study attested treatment’s activity reporting a promising RR of 75% in a cohort of 20 mCRC patients, not selected for EGFR expression or for KRAS mutational status [73]. A group of 42 molecularly unselected, initially unresectable mCRC patients have been treated with the first-line triplet (according to a schedule differing from GONO-FOLFOXIRI only by a higher planned dose of irinotecan) in a Phase II trial [74]. According to preliminary results, 22 out of 37 evaluable patients achieved a response, with a RR of 82% and a disease control rate of 97%. The feasibility of the treatment’s schedule was demonstrated by the acceptable safety profile, that included diarrhea and neutropenia as the most common grade 3 -- 4 toxicities, with awaited frequencies. Final efficacy data are not available yet. Another Phase II trial is currently ongoing, evaluating the combination of HORG-FOLFOXIRI plus cetuximab in a population of unresectable mCRC patients, with no molecular selection criteria [75]. The combination of a chrono-modulated schedule of three-drug chemotherapy with cetuximab has been tested in
Expert Opin. Biol. Ther. (2011) 11(4)
525
526 Phase III, Open-label, Two-arms, Randomized
NCT00719797
A Phase III randomized trial of FOLFOXIRI + BEVACIZUMAB versus FOLFIRI + BEVACIZUMAB as first-line treatment for metastatic colorectal cancer TRIBE [69]
Phase II, Open-label, Single-arm Phase II, Open-label, Single-arm, Non-Randomized, Active Control
NCT01126866
NA
NCT00689624
Phase II Study on Curative Resectability of Not Optimally Resectable Liver and/or Lung Metastases From Colorectal Carcinoma (CRC) Under Intensified Chemotherapy (FOLFOXIRI/ Bevacizumab) [72]
Phase II trial of FOLFOXIRI plus Panitumumab as fist-line treatment for KRAS- and BRAF-wild-type metastatic colorectal cancer TRIP
A Triplet Combination With Irinotecan Plus Oxaliplatin, Continuous Infusion 5-Fluorouracil And Leucovorin Plus Cetuximab As First Line Treatment In Metastatic Colorectal Cancer. A Pilot Phase II Trial [75]
Phase II, Open-label, Single-arm
NCT00778102
A Multicentre Randomized Phase II Study to Assess the Safety and Resectablity in Patients With Primarily Unresectable Liver Metastases Secondary to Colorectal Cancer Receiving Treatment With 5-FU, Leucovorin, Oxaliplatin and Bevacizumab With or Without Irinotecan as First Line Treatment [71]
Phase II, Open-label, Two-arms, Randomized
Study design
Clinical Trials.gov ID
Name
Expert Opin. Biol. Ther. (2011) 11(4)
Patients with unresectable mCRC
Patients with unresectable mCRC
Patients with not optimally resectable mCRC (liver and/or l ung metastases only)
Patients with liver only mCRC
Patients with unresectable mCRC
Main inclusion criteria
Table 2. Ongoing clinical trials evaluating intensive treatments of three-chemoterapeutic agents plus a biologic.
Response rate
Response rate
Surgical resectability
Surgical resectability
PFS
Primary end point
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Recruiting
Recruiting
Recruiting
Recruiting
Recruiting
Status
Greece
Italy
Germany
Austria, France, Spain
Italy
Location
Cytotoxic triplets plus a biologic in mCRC
Loupakis, Cremolini, Schirripa, Masi & Falcone
A. Progression free survival (%)
100
75
50 Number of patients: 57 Number of events: 47 Median follow up: 28.8 months
25
Median progression-free survival: 13.1 months 0 12
24
36
24
36
Time (months) B. 100
Overall survival (%)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
0
75
50 Number of patients: 57 Number of events: 26 Median follow up: 28.8 months
25
Median survival: 30.9 months 0 0
12 Time (months)
Figure 1. Progression-free A. and overall survival B. curves of the Phase II FOIB trial (GONO-FOLFOXIRI plus bevacizumab).
TRIBE trial are awaited with great interest, in order to really understand the effect of the addition of oxaliplatin to firstline FOLFIRI plus bevacizumab. Some urgent issues need to be handled, to really carve out the most appropriate space for these regimens in clinical practice. The availability of targeted agents might represent a very appealing chance to restrict the use of chemotherapy to a short, initial period, in which the achievement of a meaningful tumoral shrinkage represents the major goal of the therapeutic strategy. In this regard, the choice of an intensive up-front chemotherapy, whose activity might even be optimized by the combination with a biologic drug, would be extremely appropriate. Results of the Phase III randomized Maintenance Bevacizumab after Induction Therapy in Metastatic Colon cancer (MACRO) trial, that compared XELOX plus bevacizumab until disease progression with XELOX plus bevacizumab administered for 6 cycles and followed by maintenance with bevacizumab alone, suggest that the earlier discontinuation of chemotherapy does not substantially impair treatment’s efficacy in terms of both PFS (HR: 1.11, 95% CI 0.89 -- 1.37) and OS (HR:1.04, 95% CI 0.81 -- 1.32) [77]. The adoption of an intensive first-line treatment as a four-drug regimen would therefore allow concentrate of the greatest cytoreductive activity to a limited period, thus potentially allowing patients to be spared of a part of the chemorelated toxicities. Moreover, to really optimize the adoption of such regimens in clinical practice, efforts in translational research will be indispensable, not only to identify molecular markers, to be potentially applied as tools to better orient therapeutic choices, but also to acquire an essential amount of knowledge, to disclose the ‘backstage’ of diseases with extremely different behaviors. 4.
a Phase II trial (preoperative chemotherapy for hepatic resection (POCHER) trial) reporting a response rate of 79% and a resection rate of 63% in a population of 43 unresectable mCRC patients with liver-only metastases [76]. The GONO group is currently conducting a Phase II trial, with the aim of evaluating the safety and activity of the combination of GONO-FOLFOXIRI plus panitumumab in a population of unresectable mCRC patients with KRAS and BRAF wild-type tumors. The adoption of these molecular inclusion criteria allows maximization of treatment efficacy, by exploring its potential in a population with favorable predictors of disease’s sensitivity to anti-EGFR mAbs. Conversely, the exclusion of patients with BRAF-mutated tumors does not permit detection of a mild activity of such an intensive strategy in this poor-prognosis subgroup. 3.
Conclusion
Ongoing trials will certainly provide further insights into ‘four-drug’ regimens. In particular, efficacy results from
Expert opinion
The choice of the up-front treatment of mCRC patients is nowadays an intriguing challenge for medical oncologists, made complex and crucial not only by the availability of the wide spectrum of therapeutic options, but also by the influence of this first choice on the following steps of the global therapeutic strategy. In our opinion, while waiting for new drugs, to further improve results achieved until now, CRC research should focus on three main topics: 1) The optimization of the global strategy; 2) The refinement of the use of available cytotoxic and targeted agents; 3) The identification of molecular tools, able to drive therapeutic decisions. First of all, in the definition of the global strategy for treatment, modern oncologists can take advantage of an amount of locoregional and systemic approaches, whose best integration still needs to be established. Moreover, in the era of targeted
Expert Opin. Biol. Ther. (2011) 11(4)
527
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Cytotoxic triplets plus a biologic in mCRC
agents, the possibility of alternating induction phases, maintenance periods and chemotherapy ‘holidays’ further widens the range of practicable techniques. For all these reasons, nowadays, the global strategy for treatment deserves itself a prominent role, thus promoting a shift in medical oncologists’ mental attitudes from the concept of a ‘step by step’ designed therapeutic route to a ‘continuum of care, [78], whose foundations are laid during the up-front decisions. Secondly, the more and more extensive adoption of biologic drugs in current practice, as well as the growing amount of results from clinical trials, underline the need for a deeper awareness of the best integration of available biologic drugs in the therapeutic route. Having ascertained the detrimental effect of the double inhibition of VEGF and EGFR, despite encouraging preclinical and early clinical evidence, it would be reasonable to investigate the use of biologics in a sequential manner. In KRAS-wild-type patients, for example, the evaluation of the early inhibition of EGFR could be suggested with the aim of achieving the best tumor shrinkage, followed by the anti-VEGF for best exploiting the efficacy of the latter as maintenance. In this complex scenario, also the choice of intensive treatments, like four-drug regimens, should be weighted up by a global point of view focusing on the
objective of concentrating the greatest cytoreduction in an initial short phase of treatment, to be then maintained by less intensive regimens while considering the possibility of a ‘re-induction’ phase with all active drugs. This approach will obviously overcome the fixed scheme of lines of treatment with the traditional endpoint of PFS, instead looking at the objective of extending the time to strategy failure. Finally, in order to deliver to each patient the best therapeutic option, not only in terms of first-line regimen, but also in terms of global strategy across all the lines of treatment, an essential aid is awaited from the attempts of translational research to disclose the phenotypic and genotypic features of molecularly and clinically different diseases. The pharmacodynamic approach might be useful to provide insights into mechanisms of intrinsic and acquired resistance, thus contributing to building a comprehensive ‘continuum of molecular characterization’, able to drive the therapeutic ‘continuum of care’.
Declaration of interest The authors declare no conflict of interest and have received no payment in preparation of this manuscript.
Bibliography 1.
2.
3.
4.
5.
528
Kopetz S, Chang GJ, Overman MJ, et al. Improved survival in metastatic colorectal cancer is associated with adoption of hepatic resection and improved chemotherapy. J Clin Oncol 2009;27:3677-83 NCCN Clinical Practice Guidelines in Oncology (NCCN GuidelinesTM). Colon Cancer. Version 1.2011. Fort Washington, PA: NCCN.org. 2011. Available from: http://www.nccn. org/professionals/physician_gls/pdf/colon. pdf [Last accessed 14 January 2011] Bokemeyer C, Bondarenko I, Makhson A, et al. Fluorouracil, leucovorin, and oxaliplatin with and without cetuximab in the first-line treatment of metastatic colorectal cancer. J Clin Oncol 2009;27:663-71
6.
Hurwitz H, Fehrenbacher L, Novotny W, et al. Bevacizumab plus irinotecan, fluorouracil, and leucovorin for metastatic colorectal cancer. N Engl J Med 2004;350:2335-42
7.
Saltz LB, Clarke S, Diaz-Rubio E, et al. Bevacizumab in combination with oxaliplatin-based chemotherapy as first-line therapy in metastatic colorectal cancer: a randomized Phase III study. J Clin Oncol 2008;26:2013-19
8.
9.
Van Cutsem E, Kohne CH, Hitre E, et al. Cetuximab and chemotherapy as initial treatment for metastatic colorectal cancer. N Engl J Med 2009;360:1408-17 Peeters M, Price TJ, Cervantes A, et al. Randomized Phase III study of panitumumab with fluorouracil, leucovorin, and irinotecan (FOLFIRI) compared with FOLFIRI alone as second-line treatment in patients with metastatic colorectal cancer. J Clin Oncol 2010;28:4706-13
Karapetis CS, Khambata-Ford S, Jonker DJ, et al. K-ras mutations and benefit from cetuximab in advanced colorectal cancer. N Engl J Med 2008;359:1757-65 Bokemeyer C, Kohne C, Rougier P, et al. Cetuximab with chemotherapy (CT) as first-line treatment for metastatic colorectal cancer (mCRC): analysis of the CRYSTAL and OPUS studies according to KRAS and BRAF mutation status. J Clin Oncol 2010;28(15s):abstract 3506
10.
Amado RG, Wolf M, Peeters M, et al. Wild-type KRAS is required for panitumumab efficacy in patients with metastatic colorectal cancer. J Clin Oncol 2008;26:1626-34
11.
Souglakos J, Philips J, Wang R, et al. Prognostic and predictive value of
Expert Opin. Biol. Ther. (2011) 11(4)
common mutations for treatment response and survival in patients with metastatic colorectal cancer. Br J Cancer 2009;101:465-72 12.
Tol J, Nagtegaal ID, Punt CJ. BRAF mutation in metastatic colorectal cancer. N Engl J Med 2009;361:98-9
13.
Grothey A, Sargent D, Goldberg RM, Schmoll HJ. Survival of patients with advanced colorectal cancer improves with the availability of fluorouracil-leucovorin, irinotecan, and oxaliplatin in the course of treatment. J Clin Oncol 2004;22:1209-14
14.
Grothey A, Sargent D. Overall survival of patients with advanced colorectal cancer correlates with availability of fluorouracil, irinotecan, and oxaliplatin regardless of whether doublet or single-agent therapy is used first line. J Clin Oncol 2005;23:9441-2
15.
Koopman M, Antonini NF, Douma J, et al. Sequential versus combination chemotherapy with capecitabine, irinotecan, and oxaliplatin in advanced colorectal cancer (CAIRO): a phase III randomised controlled trial. Lancet 2007;370(9582):135-42
16.
Seymour MT, Maughan TS, Ledermann JA, et al. Different strategies
Loupakis, Cremolini, Schirripa, Masi & Falcone
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
of sequential and combination chemotherapy for patients with poor prognosis advanced colorectal cancer (MRC FOCUS): a randomised controlled trial. Lancet 2007;370:143-52 17.
Saltz LB, Cox JV, Blanke C, et al. Irinotecan plus fluorouracil and leucovorin for metastatic colorectal cancer. Irinotecan Study Group. N Engl J Med 2000;343:905-14
18.
Douillard JY, Cunningham D, Roth AD, et al. Irinotecan combined with fluorouracil compared with fluorouracil alone as first-line treatment for metastatic colorectal cancer: a multicentre randomised trial. Lancet 2000;355:1041-7
19.
Kohne CH, van Cutsem E, Wils J, et al. Phase III study of weekly high-dose infusional fluorouracil plus folinic acid with or without irinotecan in patients with metastatic colorectal cancer: European Organisation for Research and Treatment of Cancer Gastrointestinal Group Study 40986. J Clin Oncol 2005;23:4856-65
20.
de Gramont A, Figer A, Seymour M, et al. Leucovorin and fluorouracil with or without oxaliplatin as first-line treatment in advanced colorectal cancer. J Clin Oncol 2000;18:2938-47
21.
Giacchetti S, Perpoint B, Zidani R, et al. Phase III multicenter randomized trial of oxaliplatin added to chronomodulated fluorouracil-leucovorin as first-line treatment of metastatic colorectal cancer. J Clin Oncol 2000;18:136-47
22.
23.
24.
25.
Goldberg RM, Sargent DJ, Morton RF, et al. A randomized controlled trial of fluorouracil plus leucovorin, irinotecan, and oxaliplatin combinations in patients with previously untreated metastatic colorectal cancer. J Clin Oncol 2004;22:23-30 Koopman M, Seymour MT, Punt CJ. The CAIRO and FOCUS studies: which lesson is to be learned? Oncologist 2009;14:192-3; author reply 194-6 Falcone A, Masi G, Allegrini G, et al. Biweekly chemotherapy with oxaliplatin, irinotecan, infusional Fluorouracil, and leucovorin: a pilot study in patients with metastatic colorectal cancer. J Clin Oncol 2002;20:4006-14 Masi G, Allegrini G, Cupini S, et al. First-line treatment of metastatic colorectal cancer with irinotecan, oxaliplatin and 5-fluorouracil/leucovorin
(FOLFOXIRI): results of a phase II study with a simplified biweekly schedule. Ann Oncol 2004;15(12):1766-72 26.
27.
28.
Falcone A, Ricci S, Brunetti I, et al. Phase III trial of infusional fluorouracil, leucovorin, oxaliplatin, and irinotecan (FOLFOXIRI) compared with infusional fluorouracil, leucovorin, and irinotecan (FOLFIRI) as first-line treatment for metastatic colorectal cancer: the Gruppo Oncologico Nord Ovest. J Clin Oncol 2007;25:1670-6 Souglakos J, Androulakis N, Syrigos K, et al. FOLFOXIRI (folinic acid, 5-fluorouracil, oxaliplatin and irinotecan) vs FOLFIRI (folinic acid, 5-fluorouracil and irinotecan) as first-line treatment in metastatic colorectal cancer (MCC): a multicentre randomised phase III trial from the Hellenic Oncology Research Group (HORG). Br J Cancer 2006;94:798-805 Vamvakas L, Athanasiadis A, Karampeazis A, et al. Clinical outcome of elderly patients with metastatic colorectal cancer treated with FOLFOXIRI versus FOLFIRI: Subgroup analysis of a randomized phase III trial from the Hellenic Oncology Research Group (HORG). Crit Rev Oncol Hematol 2010;76:61-70
29.
Golfinopoulos V, Salanti G, Pavlidis N, Ioannidis JP. Survival and disease-progression benefits with treatment regimens for advanced colorectal cancer: a meta-analysis. Lancet Oncol 2007;8:898-911
30.
Masi G, Loupakis F, Pollina L, et al. Long-term outcome of initially unresectable metastatic colorectal cancer patients treated with 5-fluorouracil/ leucovorin, oxaliplatin, and irinotecan (FOLFOXIRI) followed by radical surgery of metastases. Ann Surg 2009;249:420-5
31.
Tanaka K, Takakura H, Takeda K, et al. Importance of complete pathologic response to prehepatectomy chemotherapy in treating colorectal cancer metastases. Ann Surg 2009;250:935-42
32.
Blazer DG III, Kishi Y, Maru DM, et al. Pathologic response to preoperative chemotherapy: a new outcome end point after resection of hepatic colorectal metastases. J Clin Oncol 2008;26:5344-51
Expert Opin. Biol. Ther. (2011) 11(4)
33.
Adam R, Wicherts DA, de Haas RJ, et al. Complete pathologic response after preoperative chemotherapy for colorectal liver metastases: myth or reality? J Clin Oncol 2008;26:1635-41
34.
Folprecht G, Grothey A, Alberts S, et al. Neoadjuvant treatment of unresectable colorectal liver metastases: correlation between tumour response and resection rates. Ann Oncol 2005;16:1311-19
35.
Adam R, Wicherts DA, de Haas RJ, et al. Patients with initially unresectable colorectal liver metastases: is there a possibility of cure? J Clin Oncol 2009;27:1829-35
36.
Tomlinson JS, Jarnagin WR, DeMatteo RP, et al. Actual 10-year survival after resection of colorectal liver metastases defines cure. J Clin Oncol 2007;25:4575-80
37.
Masi G, Vasile E, Loupakis F, et al. Randomized trial of two induction chemotherapy regimens in metastatic colorectal cancer: an updated analysis. J Natl Canc Inst 2011;103:21-30
38.
Sargent DJ, Kohne CH, Sanoff HK, et al. Pooled safety and efficacy analysis examining the effect of performance status on outcomes in nine first-line treatment trials using individual data from patients with metastatic colorectal cancer. J Clin Oncol 2009;27:1948-55
39.
Roth A, Klingbiel D, Yan P, et al. Molecular and clinical determinants of survival following relapse after curative treatment of stage II--III colon cancer (CC): results of the translational study on the PETACC 3-EORTC 40993-SAKK 60-00 trial. J Clin Oncol 2010;28(15s):abstract 3504
40.
Deng G, Bell I, Crawley S, et al. BRAF mutation is frequently present in sporadic colorectal cancer with methylated hMLH1, but not in hereditary nonpolyposis colorectal cancer. Clin Cancer Res 2004;10:191-5
41.
Samowitz WS, Sweeney C, Herrick J, et al. Poor survival associated with the BRAF V600E mutation in microsatellite-stable colon cancers. Cancer Res 2005;65:6063-9
42.
Tejpar S, Popovici V, Delorenzi M, et al. Mutant KRAS and BRAF gene expression profiles in colorectal cancer: results of the translational study on the PETACC 3-EORTC 40993-SAKK 60-00 trial. J Clin Oncol 2010;28(15s):abstract 3505
529
Cytotoxic triplets plus a biologic in mCRC
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
43.
Aust DE, Lutz MP, Mauer M, et al. Lessons from PETACC 2: No prognostic impact of KRAS-/BRAF-status in stage III colon cancer treated with adjuvant 5-FU monotherapy. J Clin Oncol 2010;28(15s):abstract 3591
44.
Tran B, Kopetz S, Tie J, et al. Differences in sites of metastatic disease and outcomes observed in patients with BRAF mutant colorectal cancers. J Clin Oncol 2010;28(15s):abstract 3592
45.
Giantonio BJ, Catalano PJ, Meropol NJ, et al. Bevacizumab in combination with oxaliplatin, fluorouracil, and leucovorin (FOLFOX4) for previously treated metastatic colorectal cancer: results from the Eastern Cooperative Oncology Group Study E3200. J Clin Oncol 2007;25:1539-44
46.
47.
48.
49.
50.
51.
52.
530
Cunningham D, Humblet Y, Siena S, et al. Cetuximab monotherapy and cetuximab plus irinotecan in irinotecan-refractory metastatic colorectal cancer. N Engl J Med 2004;351:337-45
53.
Hurwitz HI, Yi J, Ince W, et al. The clinical benefit of bevacizumab in metastatic colorectal cancer is independent of K-ras mutation status: analysis of a Phase III study of bevacizumab with chemotherapy in previously untreated metastatic colorectal cancer. Oncologist 2009;14:22-8
54.
Arnold D, Hinke A, Reinacher-Schick AC, et al. Waterfall plot analysis of XELOX or XELIRI with cetuximab or bevacizumab in patients with advanced colorectal cancer (ACRC): Combined analysis of two randomized first-line phase II trials of the AIO CRC study group. J Clin Oncol 2008;26(Suppl):abstract 4067
55.
Baselga J, Rosen N. Determinants of RASistance to anti-epidermal growth factor receptor agents. J Clin Oncol 2008;26:1582-4 Van Cutsem E, Nordlinger B, Cervantes A. Advanced colorectal cancer: ESMO Clinical Practice Guidelines for treatment. Ann Oncol 2010;21(Suppl 5):v93-7 Allegra CJ, Jessup JM, Somerfield MR, et al. American Society of Clinical Oncology provisional clinical opinion: testing for KRAS gene mutations in patients with metastatic colorectal carcinoma to predict response to anti-epidermal growth factor receptor monoclonal antibody therapy. J Clin Oncol 2009;27:2091-6 Lievre A, Bachet JB, Le Corre D, et al. KRAS mutation status is predictive of response to cetuximab therapy in colorectal cancer. Cancer Res 2006;66:3992-5 Douillard JY, Siena S, Cassidy J, et al. Randomized, Phase III trial of panitumumab with infusional fluorouracil, leucovorin, and oxaliplatin (FOLFOX4) versus FOLFOX4 alone as first-line treatment in patients with previously untreated metastatic colorectal cancer: the PRIME study. J Clin Oncol 2010;28:4697-705 Wagner AD, Arnold D, Grothey AA, et al. Anti-angiogenic therapies for
metastases treated with XELOX/ FOLFOX. Ann Surg Oncol 2010;17:2059-65
metastatic colorectal cancer. Cochrane Database Syst Rev 2009(3):CD005392
56.
57.
von Weikersthal FL, Moosmann N, Vehling-Kaiser U, et al. Cetuximab plus capecitabine/irinotecan (XELIRI) versus cetuximab plus capecitabine/oxaliplatin (XELOX) as first line treatment for patients with metastatic colorectal cancer (crc): a randomized trial of the AIO CRC study group. Ann Oncol 2006;17(Suppl 9):abstr3270 Schmiegel WH, Reinacher-Schick A, Freier W, et al. Comparable safety and response rate with bevacizumab in combination with capecitabine/oxaliplatin (CapOx/Bev) versus capecitabine/ irinotecan (CapIri/Bev) in advanced CRC (mCRC): A randomized phase II study of the AIO GI tumor study group. J Clin Oncol 2007;25(18S):abstract 4034 Adam R, Haller DG, Poston G, et al. Toward optimized front-line therapeutic strategies in patients with metastatic colorectal cancer -- an expert review from the International Congress on Anti-Cancer Treatment (ICACT) 2009. Ann Oncol 2010;21:1579-84
58.
Grothey A. EGFR Antibodies in Colorectal Cancer: where do they belong? J Clin Oncol 2010;28:4668-70
59.
Ribero D, Wang H, Donadon M, et al. Bevacizumab improves pathologic response and protects against hepatic injury in patients treated with oxaliplatin-based chemotherapy for colorectal liver metastases. Cancer 2007;110:2761-7
60.
Klinger M, Tamandl D, Eipeldauer S, et al. Bevacizumab improves pathological response of colorectal cancer liver
Expert Opin. Biol. Ther. (2011) 11(4)
61.
Chun YS, Vauthey JN, Boonsirikamchai P, et al. Association of computed tomography morphologic criteria with pathologic response and survival in patients treated with bevacizumab for colorectal liver metastases. JAMA 2009;302:2338-44
62.
Hecht JR, Mitchell E, Chidiac T, et al. A randomized Phase IIIB trial of chemotherapy, bevacizumab, and panitumumab compared with chemotherapy and bevacizumab alone for metastatic colorectal cancer. J Clin Oncol 2009;27:672-80
63.
Tol J, Koopman M, Cats A, et al. Chemotherapy, bevacizumab, and cetuximab in metastatic colorectal cancer. N Engl J Med 2009;360:563-72
64.
Di Nicolantonio F, Martini M, Molinari F, et al. Wild-type BRAF is required for response to panitumumab or cetuximab in metastatic colorectal cancer. J Clin Oncol 2008;26:5705-12
65.
Loupakis F, Ruzzo A, Cremolini C, et al. KRAS codon 61, 146 and BRAF mutations predict resistance to cetuximab plus irinotecan in KRAS codon 12 and 13 wild-type metastatic colorectal cancer. Br J Cancer 2009;101:715-21
66.
De Roock W, Claes B, Bernasconi D, et al. Effects of KRAS, BRAF, NRAS, and PIK3CA mutations on the efficacy of cetuximab plus chemotherapy in chemotherapy-refractory metastatic colorectal cancer: a retrospective consortium analysis. Lancet Oncol 2010;11:753-62
67.
Van Cutsem E, Lang I, Folprecht G, et al. Cetuximab plus FOLFIRI: final data from the CRYSTAL study on the association of KRAS and BRAF biomarker status with treatment outcome. J Clin Oncol 2010;28(15s):abstract 3570
68.
Masi G, Loupakis F, Salvatore L, et al. Bevacizumab with FOLFOXIRI (irinotecan, oxaliplatin, fluorouracil, and folinate) as first-line treatment for metastatic colorectal cancer: a phase 2 trial. Lancet Oncol 2010;11:845-52
69.
Combination Chemotherapy and Bevacizumab as First-Line Therapy in Treating Patients With Metastatic Colorectal Cancer. Bethesda, MD:
Loupakis, Cremolini, Schirripa, Masi & Falcone
Dose-escalation study in previously untreated patients with metastatic colorectal cancer. Proceedings 2010 Gastrointestinal Cancers Symposium. Abstract 480. Available from: http://www.asco.org/ASCOv2/ Meetings/Abstracts? &vmview=abst_detail_view& confID=72&abstractID=2265 [Last accessed 14 January 2011]
clinicaltrials.gov, 2008. Available from: http://www.clinicaltrials.gov/ct2/ show/NCT00719797?term= NCT00719797&rank=1 [Last accessed 14 January 2011] 70.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
71.
72.
73.
Falcone A, Loupakis F, Cupini S, et al. FOLFOXIRI plus bevacizumab (BV) versus FOLFIRI plus BV as first-line treatment of metastatic colorectal cancer (MCRC): preliminary safety results of the phase III randomized TRIBE study by the Gruppo Oncologico Nord-Ovest (GONO). J Clin Oncol 2010;28(15s):abstract 3543
74.
A Study of Avastin (Bevacizumab) in Combination With mFOLFOX-6 or FOLFOXIRI in Patients With Metastatic Colorectal Cancer. Bethesda, MD: clinicaltrials.gov, 2008. Available from: http://www.clinicaltrials.gov/ct2/ show/NCT00778102?term= NCT00778102&rank=1 [Last accessed 14 January 2011] Curative Resectability of Not Optimally Resectable Liver and/or Lung Metastases From Colorectal Carcinoma (CRC) Under Intensified Chemotherapy (APRIORI). Bethesda, MD: clinicaltrials. gov, 2010. Available from: http://www. clinicaltrials.gov/ct2/show/ NCT01126866?term= NCT01126866&rank=1 [Last accessed 14 January 2011] Folprecht G, Schuette K, Stoehlmacher J, et al. Cetuximab and FOLFOXIRI:
Ychou M, Desseigne F, Thezenas S, et al. Preliminary results of a multicentre phase II trial evaluating cetuximab in combination with FOLFIRINOX (LV5FU + Irinotecan + Oxaliplatin) as first line treatment of metastatic colorectal cancer (mCRC) patients. Proceedings 2009 Gastrointestinal Cancers Symposium. Abstract 450. Available from: http://www.asco.org/ ASCOv2/Meetings/Abstracts?& vmview=abst_detail_view&confID= 63&abstractID=10577 [Last accessed 14 January 2011]
75.
Irinotecan/Oxaliplatin/5-Fluorouracil/ Leucovorin/Cetuximab As First Line Treatment In Colorectal Cancer. Bethesda, MD: clinicaltrials.gov, 2008. Available from: http://www. clinicaltrials.gov/ct2/show/ NCT00689624?term= NCT00689624&rank=1 [Last accessed 14 January 2011]
76.
Garufi C, Torsello A, Tumolo S, et al. POCHER (preoperative chemotherapy
Expert Opin. Biol. Ther. (2011) 11(4)
for hepatic resection) with cetuximab (Cmab) plus CPT-11/5-fluorouracil (5-FU)/leucovorin(FA)/oxaliplatin (L-OHP) (CPT-11-FFL) in unresectable colorectal liver metastases (CLM). J Clin Oncol 2009;27(Suppl):abstract e15020 77.
Tabernero J, Aranda E, Gomez A, et al. Phase III study of first-line XELOX plus bevacizumab (BEV) for 6 cycles followed by XELOX plus BEV or single-agent (s/a) BEV as maintenance therapy in patients (pts) with metastatic colorectal cancer (mCRC): The MACRO Trial (Spanish Cooperative Group for the Treatment of Digestive Tumors [TTD]). J Clin Oncol 2010;28(15s):abstract 3501
78.
Goldberg RM, Rothenberg ML, Van Cutsem E, et al. The continuum of care: a paradigm for the management of metastatic colorectal cancer. Oncologist 2007;12(1):38-50
Affiliation
Fotios Loupakis†1,2, Chiara Cremolini2, Marta Schirripa2, Gianluca Masi1,2 & Alfredo Falcone1,2 † Author for correspondence 1 University of Pisa, Department of Oncology, Transplants and New Technologies in Medicine, Pisa, Italy E-mail:
[email protected] 2 Azienda Ospedaliero-Universitaria Pisana, Polo Oncologico, Pisa, Italy
531
Review
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
The impact of biologic response modifiers on hepatitis B virus infection 1.
Introduction
Matthew B Carroll
2.
Discussion
301 Fisher Avenue, Keesler AFB, MS, USA
3.
Expert opinion
Introduction: The biologic response modifiers are a diverse group of medications that have emerged over the last decade. They target pro-inflammatory cytokines or cell surface molecules that drive illnesses such as rheumatoid arthritis. Despite the greater control afforded they have also ushered in a new spectrum of side effects. As the same immunologic machinery that helps control infections such as HBV contributes to the pathogenesis of rheumatologic diseases, persistence or reactivation of the virus remains an evolving concern. Areas covered: A systemic literature review was performed using the PubMed and Medline databases (1996 to January 2010) searching for the index term ‘Hepatitis B’ combined with the terms ‘tumor necrosis factor’, ‘B cell’, ‘rituximab’, ‘IL-1’, ‘anakinra’, ‘IL-6’, ‘tocilizumab’, ‘CTLA-4’, and ‘abatacept’. All relevant articles in English were reviewed and secondary references of interest were also retrieved. This paper addresses the role of the various cytokines and cluster of differentiation molecules in controlling HBVinfection and the currently known effect that the biologic response modifiers have on viral control by the host immune response. Expert opinion: The risk of HBV reactivation is greatest in HBsAg positive patients. These patients should start antiviral therapy one week before receiving a biologic response modifier. The risk of HBV reactivation in HBsAg negative patients appears very low but when HBsAb titers are low use of rituximab or TNF-a antagonists may increase the risk of reactivation. Keywords: ankylosis spondylitis, B cells, cytokines, hepatitis B virus, rheumatoid arthritis, tumor necrosis factor Expert Opin. Biol. Ther. (2011) 11(4):533-544
1.
Introduction
Chronic infection with HBV remains a significant public health problem affecting more than 350 million people worldwide [1]. The incidence of chronic hepatitis B virus (HBV) infection is disproportionately higher in areas of the world such as Asia, sub-Saharan Africa, and the Amazon river basin of South America [2]. In these areas where the prevalence can exceed 8% perinatal infection is the most frequent mode of transmission. While the perinatal mode of HBV transmission does not lead to acute hepatitis it does result in the establishment of chronic infection which carries a 15 -- 40% lifetime risk of developing liver failure, cirrhosis or hepatocellular carcinoma. In the USA and western Europe, the prevalence of HBV infection is less than 2% as spread of the virus is facilitated by sexual contact and injected drug use [1,2]. Patients chronically infected with HBV may present in one of four phases, but transition through all phases may not occur in everyone [3]. Those exposed to the virus perinatally or during their childhood develop immune tolerance. This stage is characterized by the presence of the hepatitis B envelope antigen (HBeAg) and high serum levels of HBV DNA with mild to no inflammatory changes noted on liver biopsy [3]. While these patients are at a low risk of progressing to cirrhosis or hepatocellular 10.1517/14712598.2011.554810 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
533
The impact of biologic response modifiers on hepatitis B virus infection
Article highlights. .
.
.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
.
.
Chronic infection with hepatitis B virus progresses through several stages, each of which carries different risks of reactivation. Overlap in cells and cytokines recruited to control chronic HBV infection also play a role in the pathophysiology of rheumatological diseases such as rheumatoid arthritis. The risk of reactivation of chronic or latent viral and mycobacterial infection varies with each biologic response modifier. Stratification of patients chronically infected with hepatitis B who will be treated with a biologic response modifier for a rheumatologic disease should undergo evaluation by a hepatologist or infectious disease specialist and serological testing for the presence of the virus and associated liver injury. It is recommended to initiate antiviral therapy 1 week before and continue for 6 months after treatment with a biologic response modifier.
This box summarizes key points contained in the article.
carcinoma, they should be monitored for progression to the immune clearance stage. This stage marks maturation of the host immune response to the virus and is characterized by the presence of HBeAg, elevated serum levels of HBV DNA, elevation in serum alanine aminotransferase (ALT) levels, and necroinflammatory activity on liver biopsy [3]. When HBeAg seroconversion occurs a patient enters into the inactive hepatitis B surface antigen (HBsAg) carrier stage. Inactive carriers form the largest group of patients chronically infected with HBV [3]. These patients have little to no HBV DNA detectable in their serum and little to no inflammatory activity on liver biopsy however 20 -- 30% can have episodic or sustained HBV reactivation, which can fuel progressive liver damage [3]. Progression to this fourth or reactivation phase of chronic HBV infection can occur spontaneously or during immune suppression [3]. Separate from these four phases is occult HBV infection. Occult HBV infection is marked by the absence of HBsAg and HBV DNA in the serum; however, low-level HBV replication may persist with HBV DNA found in the liver [4]. Hepatitis B core antibodies (HBcAb) with or without hepatitis B surface antibodies (HBsAb) are detectable in the serum [4]. Immunosuppression may lead to HBV reactivation in these patients [4]. The key to containment and eradication of HBV is a robust immune response. While such a response is complex and recruits multiple cells and cytokines, the innate arm of host immunity is activated first. It eradicates infected hepatocytes and releases cytokines, which inhibit viral replication [5]. Though not directly infected within hours of initial HBV infection liver macrophages (Ku¨pffer cells) recognize the virus and activate pathways releasing pro-inflammatory cytokines such as IL-6, TNF-a, IL-1b and IL-8 [6]. The rapid release of 534
IL-6 controls HBV gene expression and replication at a transcriptional level [6]. Infected hepatocytes release IFN-a and b which activates NK cells [7]. NK cells eliminate HBVinfected hepatocytes and also release IFN-g and TNF-a which inhibits further viral replication without triggering hepatocyte destruction [7]. Dendritic cells are also activated during initial HBV infection [7]. These cells capture viral particles through Toll-like receptors, secrete cytokines such as IFN-a, TNF-a, IL-12, and IL -10 which help polarize naı¨ve T-cells and process antigens for presentation to T-cells via MHC molecules [7]. The adaptive arm of host immunity is then recruited as elimination of HBV infection and ultimately disease resolution depends on a robust polyclonal T-cell response [5]. Mature dendritic cells migrate from the liver to lymph nodes to activate T-cells. As facilitated by various pro-inflammatory cytokines and the interaction between the T-cell receptor and MHC--antigen complex, T-cell activation occurs [7]. Cytotoxic CD8+ T-cells continue the generation of pro-inflammatory cytokines. Naı¨ve CD4+ T-cells differentiate under the direction of B-cells and cytokines into both a TH1 phenotype, which generates cytokines such as IL-2, IFN-g and TNF-a, thus enhancing the host cytotoxic response and a TH2 phenotype releasing IL-4 IL-10, and IL-12 to boost the host humoral response to HBV [7]. When the host immune response is unable to eradicate the virus and chronic infection results, HBV-specific T-cell responses gradually wane but the humoral response is sustained and vigorous [8]. By their nature chronic inflammatory rheumatological diseases such as rheumatoid arthritis (RA) and the seronegative spondyloarthropathies represent aberrant manifestations of the host immune response [9]. A majority of the same cells and cytokines that organize the host immune response to infections such as HBV are activated and chronically recruited, thus contributing to the pathogenesis of these rheumatological diseases. Several decades of research have convincingly demonstrated that the excessive production of pro-inflammatory cytokines such as IL-1, IL-6 and TNF-a are critical to the initiation and sustainment of these diseases [9,10]. Attempts at refining pharmacological therapies to more effectively treat rheumatological diseases have lead to the emergence of a class of medications known as biologic response modifiers [11]. This diverse group of medications has the unifying goal of targeting a specific cytokine, cellmembrane-bound molecule, or lymphocyte vital to the induction and/or perpetuation of the immune response. Over the last 10 years multiple biological response modifiers have emerged to treat chronic inflammatory rheumatological diseases. Some antagonize cytokines such as TNF-a (infliximab, etanercept, adalimumab, certolizumab and golimumab), mimic the natural action of the IL-1 receptor antagonist (anakinra), or antagonize the IL-6 receptor (tocilizumab). Others inhibit cell signaling pathways or promote cell cytolysis such as the antagonist of the cluster of differentiation (CD) 80/86 molecules (abatacept) and an anti-CD20 monoclonal antibody (rituxmab) respectively. Having the ability to target
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Carroll
molecules or cells of the immune response the biologic response modifiers have made stopping the clinical, serological and radiographic manifestations an attainable goal, especially when administered in conjunction with traditional disease-modifying medications such as methotrexate [11]. While the biologic response modifiers have ushered in a new era of control over chronic inflammatory rheumatologic diseases, they have also been associated with a new spectrum of adverse events. A longstanding concern associated with the use of these agents has been the risk of infection. As a class the biologic response modifiers appear to be associated with higher rates of infection and serious infection, although this risk may only be elevated for a brief period (16 weeks) after starting some of these agents [12-14]. The reactivation of latent or opportunistic bacterial, viral or fungal infections have also been problematic. This issue gained attention shortly after the introduction of the TNF-a antagonists when post-marketing surveillance demonstrated higher rates of reactivation of Mycobacterium tuberculosis even though clinical trials did not indicate such a risk existed [11,13,15]. Opportunistic infection with atypical Mycobacteria, Histoplasmosis, Listeria, Aspergillus, Nocardia, and Cytomegalovirus have also been associated with the use of TNF-a antagonists [11,13]. Amongst the other biologic response modifiers treatment with rituximab has been associated with reactivation of the JC virus (manifesting as progressive multifocal leukoencephalopathy) and abatacept has been associated with higher rates of herpes simplex infections [12,16,17]. Anakinra does not consistently appear to be associated with such infections [12,16,17]. The effect that the biologic response modifiers have on the host immune response to contain and/or eradicate HBV is an evolving area of interest. While a growing body of data suggests that select biological response modifiers may be associated with an increased risk of HBV reactivation, this risk is not as well defined for the other therapies in this group [18]. The purpose of this review is to examine the current literature and summarize the data that exists about the effect that biologic response modifiers prescribed by rheumatologists may have on patients chronically infected with HBV. Also incorporated in the review of each biologic response modifier on chronic HBV infection are the currently known effects that polymorphisms of the various cytokines and cell signaling pathways have on control and eradication of HBV. 2.
Discussion
TNF-a antagonists TNF-a is a cytokine composed of three identical 17 kDa units and forms part of the initial cytokine response when cells are exposed to a diverse number of infectious, physical, and immunological stimuli [19-22]. Initial secretion of TNF-a comes from preformed stores cleaved from membrane-bound TNF-a residing on inflammatory or antigen-presenting cells (APC) such as macrophages. Subsequent release reflects new synthesis as 2.1
TNF-a is inserted into the cell membrane and then cleaved [9]. TNF-a triggers a host of diverse effects on the immune system and can amplify its own synthesis [9,11]. Soluble TNF-a increases expression of adhesion molecules, stimulates release of other pro-inflammatory cytokines such as IL-1 and IL-6, and can induce apoptosis [9,11]. While the majority of TNF-a synthesis is by monocytes and macrophages under various circumstances other cells may produce TNF-a such as hepatocytes and T-cells [19,22,23]. In RA, cells containing TNF-a are found in the synovial lining, juxta-articular blood vessels, and at the cartilage--pannus junction [19]. Levels of TNF-a are often much greater in the synovial space as compared with the serum in RA patients [20]. Local production of TNF-a directly stimulates osteoclast formation and leads to the characteristic bone resorption identified radiographically [19]. The TNF-a (TNFA) allele is found on chromosome 6 in the class III region of the MHC [24,25]. Genetic polymorphisms that lead to lower amounts of TNF-a secretion have been associated with an increased risk of progression to chronic HBV infection [24,26]. Multivariate analyses of Korean, Caucasian German and Chinese populations have demonstrated that the -238GA haplotype is associated with a higher risk of progressing to chronic HBV infection [24,27-29]. Other polymorphisms such as the -308GG haploytpe [30,31], the -857CC haplotype [27,28,32], and combination -308G/-238 G homozygotes [30] have also been associated with an increased risk of developing chronic HBV infection. Lower levels of TNF-a attributable to these polymorphisms have several adverse effects on the host immune system response to HBV: the cytokine cascade initiated and sustained by TNF-a is less potent [33], hepatocyte clearance via Fas--Fas-ligand mediated apoptosis is decreased [34], and CD8+ T-cell responses are dampened by the relative imbalance between TNF-a and higher levels of IFN-g [26,35]. An evolving body of literature suggests that patients with chronic HBV infection treated with TNF-a antagonists without concomitant antiviral therapy are at increased risk of experiencing reactivation of the virus. In patients who are HBsAg-positive a review of 35 case reports suggested that treatment with infliximab, in the absence of concomitant antiviral therapy, was associated with the highest rates of developing clinically symptomatic hepatitis, greater than a twofold increase in aspartate aminotransferase (AST)/ALT, greater than 1000-fold increase in HBV DNA levels, and risk of death [18]. The authors of this review recommended that patients who are HBsAg-positive and would benefit from treatment with a TNF-a antagonist should be started on antiviral therapy 1 -- 2 weeks prior to initiating such therapy [18,36]. Such patients should remain on antiviral therapy for 6 months after ceasing TNF-a antagonist therapy as immune reconstitution could lead to a flare of HBV [37]. Close serologic monitoring of trends in AST/ALT and HBV DNA was also recommended and infliximab was suggested as a second-line agent to treat the inflammatory disease of patients who were HBsAg-positive [18]. A consensus statement
Expert Opin. Biol. Ther. (2011) 11(4)
535
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
The impact of biologic response modifiers on hepatitis B virus infection
from the American College of Rheumatology (ACR) in 2008 stated that use of TNF-a antagonists was contraindicated in patients with HBV infection who had ChildPugh Class B or C liver disease regardless of the concomitant use of antiviral agents [38]. A consensus statement from the European League Against Rheumatism recommended that patients should be screened for HBV infection prior to starting TNF-a antagonists [39]. In patients who are HBsAg-negative but HBcAb-positive the risk of HBV reactivation appears very low when treatment with a TNF-a antagonist is started. While several case reports [40-42] and a retrospective case series of 88 Korean patients followed over 6 ½ years [43] suggested that some patients had serological evidence and occasionally clinical evidence of HBV reactivation, two recent prospective cohort studies comprising 88 European patients reported no cases of HBV reactivation [44,45]. In a case report [41] and one of these prospective studies [45] a decrease in HBsAb titers were observed in some of the patients. It was recommended in the prospective study that a decrease in HBsAb titers, especially in patients starting TNF-a antagonists with baseline low HBsAb titers, should have close monitoring for evidence of HBV reactivation [45]. Routine antiviral prophylaxis has not been recommended in patients who are HBsAgnegative but HBcAb-positive when treated with TNF-a antagonists [44]. Close clinical and serologic follow-up of such patients would nonetheless be prudent. IL-1 receptor antagonists IL-1 is a 17-kDa protein secreted mainly by monocytes and macrophages and has inflammatory effects, which include the induction of IL-6 and COX-2 [9,16]. It can also be produced by endothelial cells, B-cells and activated T-cells [9]. IL-1 can bind to two types of cell surface receptors. Type I IL-1 receptors (IL-1R) have a cytoplasmic tail, which facilitates intracellular signaling, whereas Type II IL-1R bind IL-1 without signal transduction [9]. A naturally produced IL-1 receptor antagonist (IL-1RA) binds Type I IL-1R with high affinity, competing with IL-1 and its ability to activate target cells [9]. Anakinra is almost identical to the nonglycosylated form of the naturally occurring IL-1RA except for an additional N-terminal methionine [16]. In RA the IL-1RA is found at lower levels in inflamed joints than would be expected [16]. Genetic studies of select populations suggest that certain polymorphisms of the IL-1 receptor antagonist gene (IL-1RN) may impart resistance to the development of chronic infection with HBV. Intron 2 of the IL-1RN gene is an 86 base pair variable number tandem repeat located in the regulatory region of the IL-1 genes and has potential functional importance by modulating IL-1 protein production [46,47]. Five different alleles of varying repeating lengths have been identified (alleles 1 through 5) [47]. In a study of 190 mainland Chinese patients with chronic HBV infection a lower number had allele 2 of IL-1RN [46]. The authors 2.2
536
suggested that allele 2 imparted greater resistance to HBV infection as they were able to generate a more robust response to clear the virus through increased IL-1b production [46]. However, a study of 80 Iranian patients did not demonstrate a similar finding, though the authors suggest that the absence of a correlation in their population could have been due to the smaller number of patients in the study [47]. To date no reports of HBV reactivation have been published of patients being treated with anakinra. An observational study of the safety of anakinra in 2006 did not report any cases of HBV reactivation with use of the medication [48]. The most recent package label for the medication available through the FDA does not comment on any known relationship to the reactivation of HBV infection [49]. The manufacturer of anakinra reported no cases of reactivation of HBV to date [50]. IL-6 receptor antagonists IL-6 is a 20-kDa protein predominantly synthesized and secreted by cells of macrophage lineage and T-cells, although synthesis and secretion of this cytokine can occur in other tissues throughout the body [51]. It is a pleiotropic cytokine with a diverse array of biological activities and regulates critical cell functions such as proliferation, differentiation and gene activation [51,52]. IL-6 controls the response of both the innate [6] and adaptive arms of host immunity in response to an infection as well as regulating the acute phase response [6,52,53]. It also induces T-cell proliferation and promotes the differentiation of cytotoxic T-cells. IL-6 exerts its numerous effects through interactions with membrane-bound IL-6 receptor (IL-6R) and the recruitment of gp130, a protein that is ubiquitous to most cells [51]. Soluble forms of IL-6R and gp130 modulate the systemic effects of IL-6 [51]. In RA serum levels of IL-6 are elevated and overproduction contributes to the pathogenesis of this illness [52]. IL-6 promotes and sustains inflammation in the synovium through leukocyte recruitment and contributes to joint destruction through endothelial cell, synovial fibroblast and osteoclast activation [52]. The constitutional and systemic effects seen in RA reflect the activity of IL-6 as a pyrogen [52]. Tocilizumab is the sole monoclonal antibody currently available which targets the IL-6 receptor to antagonize the effects of IL-6 in RA. In chronic HBV infection, serum IL-6 levels have correlated with the extent of hepatocyte damage and the development of cirrhosis [54,55]. IL-6 secretion is increased during acute exacerbations of chronic HBV infection [55]. Additionally, elevated serum IL-6 levels antedated the development of hepatocellular carcinoma in patients chronically infected with HBV [56]. Despite these observations, several studies across multiple ethnic groups have failed to establish a link between IL-6 gene promoter polymorphisms and outcomes of chronic infection although one study of an European population suggested a link between IL-6 -174 G/C and the course of chronic HBV infection [5,26,57-59]. While this study focused on the role of the polymorphism IL-6 -174, a Korean 2.3
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Carroll
study found no relationship with two other polymorphisms (positions -572 and -597) [58]. Given the role of IL-6 in acute exacerbations of chronic HBV infection it has been suggested that therapeutic neutralization of IL-6 could pose a risk in chronic HBV infected patients [6]. To date two case reports, both of Japanese patients chronically infected with HBV who were subsequently treated with tocilizumab, have been published [60,61]. The first case was a 60 year old female who had seropositive RA resistant to multiple disease-modifying agents [60]. She was found to be in the immune tolerant phase (HBeAgpositive, elevated HBV DNA level and normal ALT) while receiving tocilizumab. She received tocilizumab for more than 5 years without an exacerbation of hepatitis, so she was placed on entecavir and continued on tocilizumab [60]. The second case was a 40 year old male with adult onset Still’s disease complicated by amyloid A amyloidosis [61]. His inflammatory arthritis was also refractory to multiple disease modifying agents [61]. During the course of treating his arthritis with immunosuppressive medications he had a mild persistent elevation in ALT and evidence of chronic active hepatitis B infection (HBsAg-positive, HBeAg-positive, and HBV DNA over 107 copies/ml) [61]. The patient was started on entecavir (0.5 mg daily) with his HBV DNA level falling below the limit of detection. He was then started on tocilizumab (8 mg/kg every 4 weeks) [61]. No reactivation of his arthritis or his chronic HBV infection was observed [61]. The Japan College of Rheumatology in 2009 published guidelines about the use of tocilizumab in HBV-infected patients [62]. As Japan has had access to tocilizumab for almost a decade, recommendations were made to exclude patients with active HBV infection from receiving this medication [62]. Anti-CD20 therapy The dynamic role of B-cells in coordinating the host immune response to a pathogen extends well beyond that of antibody production [63]. B-cells are also efficient APCs, up to 1000 times more potent in processing and presenting antigens than other APCs such as macrophages or dendritic cells [63]. B-cells are very effective in presenting antigens when found at low concentrations [63]. B-cells also provide key costimulatory signals for CD4+ cells thus activating cellular immunity and dictating the extent of the initial expansion these cells undergo. Additionally B-cells can secrete or respond to the release of cytokines such as IL-6, IL-1, IFN-g and TNF-a. Such cytokines not only establish or enhance a proinflammatory state, they also exert a regulatory role on future T-cell functions [63]. One of the most immunogenic proteins of HBV is the hepatitis B core antigen (HBcAg) [64]. HBcAg at very low concentrations can activate numerous naı¨ve B-cells through crosslinking of their surface immunoglobulin receptors [64]. This T-cell-independent activation helps initiate production of HBcAb IgM antibodies. HBcAg can also bind to non-HBcAg-specific B-cells akin to the immune activation 2.4
observed to a superantigen [64]. B-cells acting in the capacity of APC phagocytize HBcAg as well as HBsAg. These antigens are then processed and presented to cognate CD4+ cells with appropriate costimulatory molecules to recruit the host cellular immune response [8,64]. As the host immune response contains initial HBV infection they produce increasing quantities of HBsAb [2]. It has been observed that B-cell depletion leads to decreased HBsAb serum titers with an increase in HBV DNA and HBV reactivation [65]. Rituximab is a chimeric monoclonal antibody with affinity for the CD20 molecule. The CD20 molecule is neither shed nor internalized and the main mechanism of action of rituximab is through antibody dependent cell-mediated cytotoxicity [66]. The CD20 molecule is widely expressed on B-cells, ranging from pre-B-cells to those later in differentiation, but it is absent from plasma cells [66]. It was originally developed and later approved for the treatment of B-cell non-Hodgkin’s lymphoma as used in conjunction with cyclophosphamide, hydroxydaunorubicin, vincristine and prednisone (CHOP) chemotherapy [39,65]. It has since been approved for use with methotrexate in the treatment of moderate to severe RA in patients with an inadequate response to TNF-a inhibitors [39]. A growing body of literature supports the risk of HBV reactivation with rituximab therapy [65]. From the oncology literature rituximab has the ability to induce HBV reactivation whether administered alone or with CHOP chemotherapy [65]. The risk of this reactivation is increased in males and with very low HBsAb titers [65]. The prevalence of such reactivation varies between 20 and 55% in HBsAg-positive patients [67], thus the recommendation for HBsAg-positive patients who need rituximab is to start antiviral prophylaxis [65,67]. Resistance to some antiviral therapies, specifically lamivudine, may decrease with some chemotherapy regimens but increases when concomitant steroids or fludaravine are administered [65]. For patients who are HBsAg-negative a prevalence for HBV reactivation of 3% has been reported [65]. In HBsAgnegative, HBcAb-positive, HBsAb-negative patients, regardless of HBV DNA levels at the start of rituximab therapy, HBV reactivation appears to be a rare occurrence. Such reactivation however has been associated with prolonged use of chemotherapy, lower efficacy of chemotherapy against lymphoma, and death from HBV hepatitis [65]. A case mortality rate of 30 -- 38% associated with HBV reactivation has been observed in this group of patients, thus antiviral prophylaxis has also been recommended [65]. In HBcAb-negative, HBsAb-positive, HBsAb-positive patients close clinical and serological monitoring for HBV reactivation has been recommended. It has been proposed that monitoring levels of HBsAb titers with HBV DNA levels may provide early clues to HBV reactivation [65]. When HBsAb titers fall, HBV DNA levels can rise and so will the risk of HBV reactivation [65]. The authors of one review recommended starting lamivudine when HBsAb titers declined to or started below 300 mIU/ml with close
Expert Opin. Biol. Ther. (2011) 11(4)
537
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
The impact of biologic response modifiers on hepatitis B virus infection
monitoring of trends in HBsAb titers and HBV DNA levels for titers that were otherwise stable, although they did note that cases existed where HBV reactivation occurred even with high (over 1000 mIU/ml) HBsAb titers [65]. When lamivudine therapy is started, in the absence of resistance, such therapy should be continued for 6 months after the last cycle of chemotherapy [37,65]. Two case reports have been published that describe HBV reactivation in patients chronically infected with the virus who were subsequently treated with rituximab for a rheumatologic illness. Both patients were HbsAg-positive [68,69]. One patient had severe RA [68] and was treated with several TNF-a antagonists sequentially as well as cyclosporine, abatacept and steroids. During her treatment with TNF-a antagonists she was found to have chronic HBV infection [68]. She was started on lamivudine and tolerated other diseasemodifying agents with a low but detectable HBV DNA level [68]. One month after receiving rituximab the patient had clinical manifestations of HBV reactivation with an increase in AST/ALT and dramatic increase in HBV DNA levels [68]. Tenofovir was added to lamivudine and one month later the patient’s elevated AST/ALT normalized [68]. The other case was a patient with ankylosing spondylitis unable to tolerate multiple disease-modifying agents who later was found to be chronically infected with HBV [69]. Lamivudine prophylaxis was started 3 months prior to receiving rituximab [69]. Despite significant improvement in her ankylosing spondylitis and no elevation in AST/ALT after starting treatment with rituximab she did have an asymptomatic increase in HBV DNA about 5 months after her dose was administered [69]. The 2008 practice guidelines from the ACR recommended against starting or resuming rituximab in patients chronically infected with HBV who were Child-Pugh class B or C regardless of whether or not they were receiving antiviral therapy, the same contraindication given for abatecept and the TNF-a antagonists [38]. Anti-CD80/86 therapy One of the bridges that links innate immunity with adaptive immunity is the stimulation of T-cells by APC. T-cell stimulation requires two signals from the APC. The first signal occurs when the antigen processed by the APC is presented in a MHC molecule, which interacts with a T-cell receptor that recognizes the antigen. The second signal is a costimulatory signal. The best characterized costimulatory signal involves the interaction between the CD80 (B7-1) and CD86 (B7-2) molecules on the APC and the CD28 molecule on the T-cell [70]. Expression of CD80 and CD86 is enhanced by the presence of various microbes as well as by the cytokines released in response to microbial invasion. Most CD4+ T-cells and about half of CD8+ T-cells constitutively express CD28. Signals working through CD28 can enhance the production of multiple cytokines (specifically IL-2), promote energy metabolism, and turn on cell cycle progression [70]. The development and survival of regulatory 2.5
538
T-cells (Treg), cells with the critical role of inhibiting immune responses, have also been linked to CD80/CD86 and CD28 signaling. Cytotoxic T-lymphocyte antigen-4 (CTLA-4) is a regulatory protein similar to CD28 in structure which undergoes inducible production after T-cell activation [70]. It exists in both a membrane-bound and soluble form, with the latter enabling the molecule to exert effects beyond direct cell-to-cell interaction. CTLA-4 extinguishes T-cell responses by inhibiting production of IL-2 and arresting cell cycle progression. It is constitutively expressed on Treg cells [71]. Abatacept is a soluble fusion protein which links the CTLA-4 extracellular domain to the Fc region of IgG molecule. Through the inhibition of the costimulatory signaling of T-cells in RA, abatacept has demonstrated clinical efficacy in multiple trials [12,70]. Recent studies have demonstrated that some CTLA-4 polymorphisms influence the ability of the immune response to clear HBV infection [71]. A study of several hundred Caucasian and African American patients in the USA linked the presence of the +49G polymorphism of CTLA-4 to an increased chance of HBV clearance [71]. It has been proposed that this outcome is attributable to the altered polarity of the CTLA-4 molecule (as compared with that produced by the +49A polymorphism) [71]. Additionally the +49G polymorphism has been associated with T-cells that express less membrane-bound CTLA-4, proliferate under conditions of suboptimal activation, and are less responsive to CTLA-4 directed inhibition [71]. These effects of the +49G polymorphism were noted regardless of the -1722C polymorphism, a polymorphism associated with decreased CTLA-4 production due to altered transcriptional activity in the gene promoter region [71]. In this same study the presence of the +6230A polymorphism was associated with HBV persistence, an outcome ascribed to decreased T-cell responsiveness related to levels of soluble CTLA-4 [71]. A similar relationship between these CTLA-4 polymorphisms and outcome of HBV infection was noted in an Iranian study of 51 patients chronically infected with HBV [72]. Beyond the effect on clearance of HBV the +49G polymorphism of CTLA-4 has been shown in males from a Han Chinese population to have a protective effect against the development of hepatocellular carcinoma [73]. To date no case reports have been published of reactivation of HBV infection in patients receiving abatacept. The manufacturer of abatacept has not received any reports of cases of HBV reactivation in patients receiving the medication [74]. A recent therapy and safety management publication recommended screening for HBV prior to starting abatacept as the safety of using abatacept in such a patient has not been established [75]. As noted earlier the ACR in 2008 stated that it was contraindicated to start or resume abatacept in patients chronically infected with HBV with Child-Pugh Class B or C regardless of whether concomitant antiviral prophylaxis was given [38].
Expert Opin. Biol. Ther. (2011) 11(4)
Carroll
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Table 1. Risk of HBV reactivation in patients chronically infected with HBV treated with biologic response modifiers for a rheumatologic disease*. Type of biologic response modifier
Medications
HBsAg+ patients
HBsAg- and HBcAb+ patients
TNF-a antagonists
Risk of reactivation exists (aggregate data from 35 case reports [18]).
IL-1 receptor antagonist IL-6 receptor antagonist
infliximab etanercept adalimumab certolizumab golimumab anakinra tocilizumab
Little to no risk of reactivation (case reports, retrospective case series, and prospective case series [40-45]) Increased risk associated with lower HBsAb titers [41,45]. No information No information
Anti-CD20 therapy
rituximab
Anti-CD80/86 therapy
abatacept
No information No cases of reactivation (two case reports [60,61]). 20 -- 55% prevalence reported (data from oncology literature [65,67] and two case reports in rheumatological literature [68,69]). No information
Rare to 3% prevalence reported (data from oncology literature [65]). Increased risk associated with lower HBsAb titers [65]. No information
*In the absence of antiviral therapy
3.
Expert opinion
The role each of the biologic response modifiers has on the risk of HBV reactivation in chronic infection varies and in some circumstances remains to be determined. A summary of the aforementioned known risk of HBV reactivation based on HBsAg status in patients concomitantly treated with biologic response modifiers is summarized in Table 1. Integrating expert opinions, societal practice guidelines, data from clinical trials and known pathophysiology it would be reasonable to refer all patients with known HBsAg positivity or detectable levels of HBV DNA to a hepatologist or infectious disease specialist [39] for further evaluation with serious consideration of starting antiviral therapy before treatment with a biologic response modifier. Initiation of antiviral therapy at least 1 week before treatment with a biologic response modifier is prudent as data from a randomized study evaluating HBsAg positive lymphoma patients treated with chemotherapy demonstrated lower HBV reactivation rates, lower rates of hepatitis, and longer survival in the group treated with lamivudine a week prior to chemotherapy [36,37,76]. Concomitant glucocorticoid use in patients chronically infected with HBV on a biologic response modifier should be done cautiously as oncology experience with rituximab suggest this predisposes to an increased risk of virus reactivation and fosters antiviral resistance [65,77]. If a patient who is actively being treated with a biologic response modifier develops clinical or serologic evidence of HBV infection, prompt consultation with a hepatologist or infectious disease expert and institution of an antiviral agent appropriate for detected resistance patterns and HBV DNA levels should be instituted. Therapy with the biologic response modifier should be held until clinical improvement, a decrease in HBV DNA, improvement in
ALT and durable compliance with antiviral therapy can be demonstrated. It is unclear as to how long this period should be. Prompt institution of antiviral therapy appears needed to minimize the effects generated by an immune system that reconstitutes towards the end of the last dose of intravenously administered biologic response modifiers such as infliximab and rituximab [18,65,76] Patients that do not have clinical or serological evidence of HBV reactivation should, in the opinion of the author, have monitoring performed once every 4 -- 8 weeks. Serologic monitoring should at a minimum include an ALT, HBV DNA levels, and HBsAb titer. The development of resistance to antiviral therapy should be considered in a previously stable patient on a biologic response modifier and a nucleot(s)ide analogue who presents with clinical hepatitis or serologic evidence of HBV reactivation. Upon cessation of a biologic response modifier antiviral therapy should be continued for at least 6 months as reconstitution of the immune system may occur, an issue more likely to arise after treatment with infliximab or rituximab [18,65,76]. The practice guidelines from the American Association for the Study of Liver Diseases recommend continuing antiviral therapy for 6 months after cessation of immunosuppression [77] whereas guidelines from the European Association for the Study of the Liver recommend a 12 month course [4]. A summary of these recommendations is provided in Table 2. For patients that are HbsAg-negative but HbcAb-positive without detectable HBV DNA in the serum it appears reasonably safe to start a biologic response modifier without the prior initiation of antiviral therapy, with the exception of rituximab. Treatment without concomitant antiviral therapy may not be appropriate when receiving rituximab, as HBV reactivation in lymphoma patients treated with rituximab and chemotherapy has resulted in a refractory
Expert Opin. Biol. Ther. (2011) 11(4)
539
The impact of biologic response modifiers on hepatitis B virus infection
Table 2. Recommendations and their rationale for implementing a biologic response modifier (BRM) in patients chronically infected with HBV Issue
Recommendation
Rationale
Prior to start of BRM
.
Screen all patients at high risk for HBV infection prior to immunosuppression [77] Check HBsAg and HBsAb and consider vaccination if seronegative [77] Seek consultation with a hepatologist or infectious disease specialist for patients that have chronic HBV infection
.
HBsAg (+) patient–start antiviral therapy appropriate for detected resistance patterns and HBV DNA level at least 1 week prior to initiation of BRM . Review risk and benefits of using rituximab in this group of patients
.
HBsAg (-) but HBcAb (+) patients without detectable HBV DNA----initiation of BRM appears reasonably safe but: . Review risk and benefits of using rituximab in this group of patients Check HBsAb titers
.
. .
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Preventing reactivation upon initiation of BRM
.
.
.
. .
.
.
Recommendation made by the AASLD based on multiple time series and uncontrolled studies [77] Recommendation made by the AASLD based on randomized controlled trials [77] Recommendation to seek consultation is based on expert opinion [39] Recommendation to start antiviral therapy 1 week prior to initiation of biologic is based on a randomized trial [18,36,37,76] Case report data from rheumatology literature suggests HBV reactivation may still occur in HBsAg (+) patients treated with rituximab on antiviral therapy [68,69] HBV reactivation in HBsAg (-) but HBcAb (+) patients treated with rituximab have experienced a refractory hepatitis with a 38% mortality rate. [65] HBsAg (-) but HBcAb (+) patients with low titers of HBsAb prior to start of TNF-a antagonist or rituximab appear at higher risk of HBV reactivation [41,45,65]
Monitoring for reactivation during treatment with BRM
Check ALT, HBV DNA, and HBsAb titers every 4 -- 8 weeks regardless of whether patient is taking antiviral therapy
Recommendation is based on the opinion of the author
Development of reactivation during treatment with BRM
. Stop BRM and seek consultation with a hepatologist or infectious disease expert . Initiate or change antiviral therapy to mitigate risk of immune reconstitution as BRM is cleared . Minimize patient exposure to glucocorticoids as risk of reactivation and antiviral resistance increases (data from patients treated with rituximab) [65,77]
Development of antiviral resistance should be considered in a previously stable patient who presents with clinical hepatitis or serologic evidence of HBV reactivation
Cessation of BRM
. Patient on antiviral therapy: Continue therapy for 6 months after . Patient not on antiviral therapy: Consider close clinical and serologic monitoring for HBV reactivation based on the dosing interval of the BRM
. .
Immune reconstitution as BRM is cleared may trigger HBV reactivation [18] The AASLD recommends continuing antiviral therapy for 6 months after immunosuppresion ends; the EASL recommends continuing antiviral therapy for 12 months after immunosuppression ends [4,77]
AASLD: American Association for the Study of Liver Diseases; HBsAg: Hepatitis B surface antigen; HBsAb: Hepatitis B surface antibody; HBcAb: Hepatitis B core antibody; ALT: Alanine aminotransferase; EASL: European Association for the Study of the Liver.
hepatitis with a 38% mortality rate [65]. The risk of HBV reactivation when treated with a biologic response modifier may also depend on the HBsAb levels. Patients with persistently low HBsAb levels treated with TNF-a antagonists or rituximab may have an elevated risk of future HBV reactivation. Consultation with a hepatologist or infectious disease expert before starting a biologic response modifier in a HBsAg-negative but HbcAb-positive patients would be prudent [39]. Again, in the opinion of the author, clinical and serologic monitoring for HBV reactivation should be performed every 4 -- 8 weeks after biologic response modifier therapy is started. These recommendations are again summarized in Table 2. 540
While all of the currently available literature links clinical and/or serological reactivation as a possible outcome in patients chronically infected with HBV treated with a biologic response modifier without antiviral therapy, it should be considered that some medications in this class may in the future demonstrate a beneficial effect on controlling the inflammation generated by the host immune response. As an example, though the long-term effects remain unknown, short (less than 1 year) courses of TNF-a antagonists are reasonably well tolerated and even a potential treatment adjunct in patients infected with HCV [39]. No significant changes in HCV RNA or increases in aminotransferases are typically observed in patients with chronic HCV
Expert Opin. Biol. Ther. (2011) 11(4)
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Carroll
infection exposed to TNF-a antagonists [39]. Clinical experience with rituximab in treating patients with HCV-related cryoglobulinemic vasculitis is growing and thus far appears to be reasonably efficacious without significant hepatic toxicity [39]. Drawing on known immunologic mechanisms the costimulatory molecule antagonist CTLA-4 is known to dampen the host immune response to an immunological challenge by inducing and maintaining T-cell tolerance [70]. In the context of chronic HBV infection CTLA-4 could play a role in attenuating the inflammation generated during the host response to viral reactivation. Abatacept, as a CTLA-4 fusion molecule, may thus have a tolerable or even have a beneficial effect on the host immune response during HBV reactivation. The demonstration of a benefit when biologic response modifiers modulate the host immune response in patients chronically infected with HBV awaits further study.
with HBV reactivation (with elevated AST/ALT, HbsAgpositive, or with detectable HBV DNA) need to defer or stop treatment with a biologic response modifier, seek input from a hepatologist or infectious disease expert and start antiviral therapy appropriate for detected resistance patterns and serum viral load. Antiviral therapy should continue during biologic response modifier treatment and continue for 6 months after the treatment has been stopped. This will mitigate the risk of an immune reconstitution response, which can occur off the biologic response modifier. For patients who are HBsAg-negative but HBcAb-positive the risk of HBV reactivation while treated with a biologic response modifier appears fairly low, although it appears dependent on the medication used. Consultation with a hepatologist or infectious disease expert prior to the initiation of a biologic response modifier and close clinical and serologic monitoring would be prudent.
Conclusion Infection with HBV remains a significant public health issue worldwide. The same host immune system that may successfully fight and eradicate infection with HBV also participates in the pathogenesis of chronic inflammatory rheumatological diseases. Biologic response modifiers are a diverse group of medications currently at the disposal of rheumatologists to control rheumatological diseases such as RA and the seronegative spondyloarthropathies. The effect that these medications have on the host control of HBV infection when treated for a rheumatological illness is an evolving area of interest. Integrating case reports and case series, expert recommendations and societal consensus statements patients with clinical hepatitis or serological evidence consistent
Acknowledgements
3.1
I am indebted to Jennifer Carroll for her critical review of this manuscript and dedicate this to my daughter Abigail.
Declaration of interest The author has no financial support or other benefits from commercial sources to disclose. The author has received no pharmaceutical or industry support in writing this manuscript. The author wishes to state that the views expressed in this article are his and do not reflect the official policy or position of the United States Air Force, Department of Defense or the US Government.
Expert Opin. Biol. Ther. (2011) 11(4)
541
The impact of biologic response modifiers on hepatitis B virus infection
Bibliography Furst DE. The risk of infections with biologic therapies for rheumatoid arthritis. Semin Arthritis Rheum 2010;39:327-46
26.
Ben-Ari Z, Mor E, Papo O, et al. Cytokine gene polymorphisms in patients infected with hepatitis B virus. Am J Gastroenterol 2003;98:144-50
Shi YH, Shi CH. Molecular characteristics and stages of chronic hepatitis B virus infection. World J Gastroenterol 2009;15:3099-105
15.
Keane J, Gershon S, Wise RP, et al. Tuberculosis associated with infliximab, a tumor necrosis factor alpha-neutralizing agent. N Engl J Med 2001;345:1098-104
27.
EASL Clinical Practice Guidelines: management of chronic hepatitis B. J Hepatol 2009;50:227-42
16.
Olsen NJ, Stein CM. New drugs for rheumatoid arthritis. N Engl J Med 2004;350:2167-79
Li HQ, Li Z, Liu Y, et al. Association of --238G/A and --857C/T polymorphisms of tumor necrosis factor-alpha gene promoter region with outcomes of hepatitis B virus infection. Biomed Environ Sci 2006;19:133-6
28.
Ribeiro CSS, Visentainer JEL, Moliterno RA. Association of cytokine genetic polymorphisms with hepatitis B infection evolution in adult patients. Mem Inst Oswaldo Cruz 2007;102:435-40
17.
Li HQ, Li Z, Liu Y, et al. Association of polymorphism of tumor necrosis factor-alpha gene promoter region with outcome of hepatitis B virus infection. World J Gastroenterol 2005;11:5213-17
29.
Kim YJ, Lee HS, Yoon JH, et al. Association of TNF-alpha promoter polymorphisms with the clearance of hepatitis B virus infection. Hum Mol Genet 2003;12:2541-6
30.
Cheong JY, Cho SW, Hwang IL, et al. Association between chronic hepatitis B virus infection and interleukin-10, tumor necrosis factor-alpha gene promoter polymorphisms. J Gastro Hepatology 2006;21:1163-9
31.
Niro GA, Fontana R, Gioffreda D, et al. Tumor necrosis factor gene polymorphisms and clearance or progression of hepatitis B virus infection. Liver Int 2005;25:1175-81
32.
Du T, Guo XH, Zhu XL, et al. Association of TNF-alpha promoter polymorphisms with the outcomes of hepatitis B virus infection in Chinese Han population. J Viral Hepat 2006;13:618-24
33.
Fang JWS, Shen WW, Meager A, et al. Activations of the tumor necrosis factor-alpha system in the liver in chronic hepatitis B virus infection. Am J Gastroenterol 1996;91:748-53
34.
Rehermann B. Intrahepatic T-cells in hepatitis B: viral control versus liver cell injury. J Exp Med 2000;191:1263-8
35.
Su F, Schneider RJ. Hepatitis B virus HBx protein sensitizes cells to apoptotic killing by tumor necrosis factor alpha. Proc Natl Acad Sci USA 1997;94:8744-9
36.
Lau GK, Yiu HH, Fong DY, et al. Early is superior to deferred pre-emptive lamivudine therapy for hepatitis B patients undergoing chemotherapy. Gastroenterology 2003;125:1742-9
Lok ASF. Chronic hepatitis B. N Engl J Med 2002;346:1682-3
2.
Dienstag JL. Hepatitis B virus infection. N Engl J Med 2008;359:1486-500
3.
4.
5.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
14.
1.
6.
7.
8.
Hosel M, Quasdorff M, Wiegmann K, et al. Not interferon, but interleukin-6 controls early gene expression in hepatitis B virus infection. Hepatology 2009;50:1773-82 Larrubia JR, Benito-Martinez S, Miquel-Plaza J, et al. Cytokines -- their pathogenic and therapeutic role in chronic viral hepatitis. Rev Esp Enferm Dig 2009;101:343-51 Ganem D, Prince AF. Hepatitis B virus infection -- natural history and clinical consequences. N Engl J Med 2004;350:1118-29
9.
Choy EHS, Panayi GS. Cytokine pathways and joint inflammation in rheumatoid arthritis. N Engl J Med 2001;344:907-16
10.
Rasheed Z, Haqqi TM. Update on targets of biologic therapies for rheumatoid arthritis. Curr Rheumatol Rev 2008;4:246-60
11.
Giles JT, Bathon JM. Serious infections associated with anticytokine therapies in the rheumatic diseases. J Int Care Med 2004;19:320-34
12.
Nurmohamed MT. Newer biological agents in the treatment of rheumatoid arthritis. Drugs 2009;69:2035-43
13.
Fleischmann RM, Iqbal I, Stern RL. Considerations with the use of biological therapy in the treatment of rheumatoid arthritis. Expert Opin Drug Saf 2004;3:391-403
542
18.
Sibilia J, Westhovens R. Safety of T-cell co-stimulation modulation with abatacept in patients with rheumatoid arthritis. Clin Exp Rheumatol 2007;25(Suppl 46):S46-56 Carroll MB, Forgione MA. Use of tumor necrosis factor alpha inhibitors in hepatitis B surface antigen-positive patients: a literature review and potential mechanisms of action. Clin Rheumatol 2010;29:1021-9
19.
Weaver AL. Differentiating the new rheumatoid arthritis biologic therapies. J Clin Rheumatol 2003;9:99-114
20.
Case JP. Old and new drugs used in rheumatoid arthritis: part 2. Am J Therapeutics 2001;8:163-79
21.
Penn H. Biologic therapies in autoimmune diseases. Clin Med 2006;6:105-8
22.
Gonzalez-Amaro R, Garcia-Monzon C, Garcia-Buey L, et al. Induction of tumor necrosis factor alpha production by human hepatocytes in chronic viral hepatitis. J Exp Med 1994;179:841-8
23.
Roux CH, Brocq O, Breuil V, et al. Safety of anti-TNF-alpha therapy in rheumatoid arthritis and spondyloarthropathies with concurrent B or C chronic hepatitis. Rheumatology 2006;45:1294-7
24.
25.
Hohler T, Kruger A, Gerken G, et al. A tumour necrosis factor-alpha (TNF-alpha) promoter polymorphism is associated with chronic hepatitis B infection. Clin Exp Immunol 1998;111:579-82 Wilson AG, Symons JA, McDowell TL, et al. Effects of a polymorphism in the human tumor necrosis factor alpha promoter on transcriptional activation. Proc Natl Acad Sci USA 1997;94:3195-9
Expert Opin. Biol. Ther. (2011) 11(4)
Carroll
37.
38.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
39.
Calabrese LH, Zein NN, Vassilopoulos D. Hepatits B virus reactivation with immunosuppressive therapy in rheumatic diseases: assessment and preventive strategies. Ann Rheum Dis 2006;65:983-9
receptor antagonist genes in patients with chronic hepatitis B in Iran. World J Gastroenterol 2006;12:5044-7 48.
den Broeder AA, de Jong E, Franssen MJAM, et al. Observational study on efficacy, safety, and drug survival of anakinra in rheumatoid arthritis patients in clinical practice. Ann Rheum Dis 2006;65:760-2
Saag K, Teng GG, Patkar NM, et al. American College of Rheumatology 2008 recommendations for the use of nonbiologic and biologic disease-modifying antirheumatic drugs in rheumatoid arthritis. Arthritis Rheum 2008;59:762-84
49.
Ferri C, Govoni M, Calabrese L. The A, B, Cs of viral hepatitis in the biologic era. Curr Opin Rheumatol 2010;22:443-50
50.
Carroll, Matthew B (matthew.
[email protected]). ‘Anakinra and HBV.’ Email to Pilstrom Bjorn of Biovitrium (
[email protected]) 5 Aug 2010
Kineret. Washington DC: Drugs@FDA. 2004. Available from: http://www. accessdata.fda.gov/scripts/cder/drugsatfda/ index.cfm?fuseaction=Search.DrugDetails [Last accessed 13 January 2011]
40.
Raftery G, Griffiths B, Kay L, et al. Chronic viral hepatitis and TNF-alpha blockade. Rheumatology 2007;46:1381
51.
41.
Montiel PM, Solis JA, Chirinos JA, et al. Hepatitis B virus reactivation during therapy with etanercept in an HBsAg negative and anti-HBs-positive patient. Liver Int 2008;28:718-20
Heinrich PC, Behrmann I, Muller-Newen G, et al. Interleukin-6-type cytokine signalling through the gp130/Jak/STAT pathway. Biochem J 1998;334:297-314
52.
42.
Madonia S, Orlando A, Scimeca D, et al. Occult hepatitis B and infliximab-induced HBV reactivation [letter]. Inflamm Bowel Dis 2007;13:508-9
Hushaw LL, Sawaqed R, Sweis G, et al. Critical appraisal of tocilizumab in the treatment of moderate to severe rheumatoid arthritis. Ther Clin Risk Manag 2010;6:143-52
53.
Ataseven H, Bahcecioglu IH, Kuzu N, et al. The levels of ghrelin, leptin, TNF-alpha, and IL-6 in liver cirrhosis and hepatocellular carcinoma due to HBV and HDV infection. Mediators Inflamm 2006;2006(4):78380
43.
Kim YJ, Bae SC, Sung YK, et al. Possible reactivation of potential hepatitis B virus occult infection by tumor necrosis factor-alpha blocker in the treatment of rheumatic diseases. J Rheumatol 2010;37:346-50
44.
Caporali R, Bobbio-Pallavicini F, Atzeni F, et al. Safety of tumor necrosis factor alpha blockers in hepatitis B virus occult carriers (hepatitis B surface antigen negative/anti-hepatitis B core antigen positive) with rheumatic diseases. Arthritis Care Res 2010;62:749-54
54.
55.
Kuo TM, Hu C, Chen YL, et al. HBV replication is significantly reduced by IL-6. J Biomed Sci 2009;16:41-9
Charpin C, Guis S, Colson P, et al. Safety of TNF-blocking agents in rheumatic patients with serology suggesting past hepatitis B state: results from a cohort of 21 patients. Arthritis Res Ther 2009;11:179-84
56.
Wong VW, Yu J, Cheng AS, et al. High serum interleukin-6 level predicts future hepatocellular carcinoma development in patients with chronic hepatitis B. Int J Cancer 2009;124:2766-70
57.
Li C, Zhi-Xin C, Li-Juan Z, et al. The association between cytokine gene polymorphisms and the outcomes of chronic HBV infection. Hepatol Res 2006;36:158-66
58.
Park BL, Lee HS, Kim YJ, et al. Association between interleukin 6 promoter variants and chronic
45.
46.
Zhang P-A, Li Y, Xu P, et al. Polymorphisms of interleukin-1B and interleukin-1 receptor antagonist genes in patients with chronic hepatitis B. World J Gastroenterol 2004;10:1826-9
47.
Ranjbar M, Alizadeh AHM, Hajilooi M, et al. Polymorphisms of interleukin-1R
Liu Q, Feng GX, Lin YL, et al. Detection of interleukin-6 and -12 in of hepatitis B patients and its clinical significance. Di Yi Jun Yi Da Xue Xue Bao 2001;21:858-9
Expert Opin. Biol. Ther. (2011) 11(4)
hepatitis B progression. Exp Mol Med 2003;35:76-82 59.
Fabris C, Toniutto P, Bitetto D, et al. Gene polymorphism at the interleukin 6 --174 G>C locus affects the outcome of chronic hepatitis B. J Infection 2009;59:144-5
60.
Nagashima T, Minota S. Long-term tocilizumab therapy in a patient with rheumatoid arthritis and chronic hepatitis B. Rheumatology 2008;47:1838-40
61.
Kishida D, Okuda Y, Onishi M, et al. Successful tocilizumab treatment in a patient with adult-onset Still’s disease complicated by chronic active hepatitis B and amyloid A amyloidosis. Mod Rheumatol 2011: published onlone 8 October 2010. doi: 10.1007/s10165-010-0365-8
62.
Koike R, Harigai M, Atsumi T, et al. Japan College of Rheumatology 2009 guidelines for the use of tocilizumab, a humanized anti-interleukin-6 receptor monoclonal antibody, in rheumatoid arthritis. Mod Rheumatol 2009;19:351-7
63.
Stasi R. Rituximab in autoimmune hematologic diseases: not just a matter of B-cells. Seminars in Hematology 2010;47:170-9
64.
Lazdina U, Cao T, Steinbergs J, et al. Molecular basis for the interaction of the hepatitis B virus core antigen with the surface immunoglobulin receptor on naive B-cells. J Virol 2001;75:6367-74
65.
Tsutsumi Y, Yamamoto Y, Tanaka J, et al. Prevention of hepatitis B virus reactivation under rituximab therapy. Immunotherapy 2009;1:1053-61
66.
Cooper N, Arnold DM. The effect of rituximab on humoral and cell mediated immunity and infection in the treatment of autoimmune diseases. Br J Haematol 2010;149:3-13
67.
Tsutsumi Y, Kanamori H, Mori A, et al. Reactivation of hepatitis B virus with rituximab. Expert Opin Drug Saf 2005;4:599-608
68.
Pyrpasopoulou A, Douma S, Vassiliadis T, Reactivation of chronic hepatitis B virus infection following rituximab administration for rheumatoid arthritis. Rheumatol Int 2009: published on line 13 September 2009, doi: 10.1007/s00296-009-1202-2
69.
Rodriguez-Escalera C, Fernandez-Nebro A. The use of
543
The impact of biologic response modifiers on hepatitis B virus infection
rituximab to treat a patient with ankylosing spondylitis and hepatitis B. Rheumatology 2008;47:1732-3 70.
71.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
72.
544
73.
Sharpe AH, Abbas AK. T-Cell costimulation -- biology, therapeutic potential, and challenges. N Engl J Med 2006;355:973-5 Thio CL, Mosbruger TL, Kaslow RA, et al. Cytotoxic T-lymphocyte antigen 4 gene and recover from hepatitis B virus infection. J Virol 2004;78:11258-62 Alizadeh AHM, Hajilooi M, Ranjbar M, et al. Cytotoxic T-lymphocyte antigen 4 gene polymorphisms and susceptibility to chronic hepatitis B. World J Gastroenterol 2006;12:630-5
74.
75.
Gu X, Qi P, Zhou F, et al. +49G>A polymorphism in the cytotoxic T-lymphocyte antigen-4 gene increases susceptibility to hepatitis B-related hepatocellular carcinoma in a male Chinese population. Hum Immunol 2010;71:83-7 Levine, Loree. “Information you requested from Bristol-Myer-Squibb.” Message to Matthew B. Carroll. 12 August 2010. E-mail. Pham T, Claudepierre P, Constantin A, et al. Abatacept therapy and safety management. Joint Bone Spine 2009;76(Suppl 1):S1-56
Expert Opin. Biol. Ther. (2011) 11(4)
76.
Nathan DM, Angus PW, Gibson PR. Hepatitis B and C virus infections and anti-tumor necrosis factor-alpha therapy: guidelines for clinical approach. J Gastroenterol Hepatol 2006;21:1366-71
77.
Lok ASF, McMahon BJ. Chronic hepatitis B. Hepatology 2007;45:507-39
Affiliation Matthew B Carroll MD FACP FACR 301 Fisher Avenue, Keesler AFB, MS 39534, USA Tel: +1 228 376 3629; Fax: +1 228 376 0105; E-mail:
[email protected]
Drug Evaluation
Kinoid of human tumor necrosis factor-alpha for rheumatoid arthritis Introduction
2.
Overview of the market
3.
Chemistry and preparation of the TNF-kinoid
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Luca Semerano, Eric Assier, Laure Delavalle´e & Marie-Christophe Boissier†
1.
4.
Pharmacodynamics
5.
Pharmacokinetics
6.
Effects in animal models
7.
Safety and tolerability
8.
Conclusions
9.
Expert opinion
†
University of Paris-13, Sorbonne Paris-Cite´, EA4222, Li2P, 74 rue Marcel Cachin, Bobigny, France
Introduction: Anti-TNF-a drugs have dramatically changed treatment of rheumatoid arthritis (RA) in terms of both clinical control and articular damage prevention. Despite this, they hold some important drawbacks, such as frequent therapeutic failures and high costs. Anti-TNF-a active immunization, with a therapeutic vaccine against TNF-a, is a promising alternative antiTNF-a targeting strategy, potentially devoid of treatment limitations of some of current anti-TNF blocking agents. Areas covered: This review covers the preclinical proof-of-concept of antiTNF-a vaccination with the kinoid of human TNF-a (TNFK) and analyzes the body of evidence forming the rationale for the application of this strategy in RA and other TNF-a-dependent diseases. We describe the theoretical bases of anti-TNF-a active immunization and of experimental data supporting the applicability of TNFK to human disease in terms of both safety and efficacy. Expert opinion: Based on preclinical efficacy and safety data supporting its feasibility in a Phase I -- II trial in Crohn’s disease, anti-TNF-a vaccination with TNFK has entered the phase of clinical development and promises to be a valuable anti-TNF-a targeting strategy in human disease. The focus is made in the first clinical trial in RA (Phase II) on the efficacy in active RA patients having developed antibodies against anti-TNF mAbs. Keywords: anti-cytokine vaccination, anti-TNF-a, kinoid, rheumatoid arthritis, TNFK Expert Opin. Biol. Ther. (2011) 11(4):545-550
1.
Introduction
Rheumatoid arthritis (RA) is the most common inflammatory rheumatic disease with a prevalence ranging from 0.3 to 1.5% in different populations [1]. It is characterized by an invasive synovial proliferation that leads to joint damage with pain and loss of function, with precocious disability [2]. RA patients have associated co-morbidities leading to a mortality estimated at almost twofold that of general population [3]. RA is, therefore, a huge public health problem resulting in high direct and indirect costs for the community [4]. 2.
Overview of the market
TNF-a-targeting agents brought a revolution in the treatment of RA, providing unheard of results in terms of disease clinical control and prevention of RA structural damage and consequent disability. TNF-a can be targeted with mAbs or their fragments (infliximab (IFX), adalimumab, golimumab, certolizumab) or with fusion products carrying a TNF-a soluble receptor (etanercept). Anti-TNF-a drugs first opened the perspective of a successful cytokine-targeting strategy in RA. Sales of the four anti-TNF-a agents on the market in 2008 (adalimumab, IFX, etanercept and certolizumab pegol) reached $16 billions. By 2014, analysts forecast the entire 10.1517/14712598.2011.566856 © 2011 Informa UK, Ltd. ISSN 1471-2598 All rights reserved: reproduction in whole or in part not permitted
545
Kinoid of human TNF-a
Box 1. Drug summary. Drug name Phase Indication Pharmacology
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Route of administration Pivotal trials
Kinoid of human TNF-a Phase II clinical trial, pre-registration Rheumatoid arthritis Active immunization (vaccination) against the pro-inflammatory cytokine TNF-a Intramuscular TNFK001 (http://clinicaltrials.gov/ ct2/show/NCT00808262) TNFK003 (http://www.controlled-trials. com/mrct/trial/772671/TNFK003)
class of anti-TNF drugs to generate a $25 billion market, with growth driven by new entrants and continuing demand for the incumbents (source: EvaluatePharma) [5]. In 2008, TNF-a inhibitors accounted for 80% of RA drug sales in the US, France, Germany, Italy, Spain, the UK and Japan (source: Pharmacor) [6] within a market that, for all biological therapies for RA, was estimated at $7 billion in 2007 (source: Datamonitor Research Store) [7]. Current TNF-a targeting strategies have nevertheless shown several drawbacks as far as safety, efficacy and costs are concerned. Despite the good safety/efficacy profile in selected patients, the overall risk of infection and possibly neoplasm is increased in RA patients treated with antiTNF-a mAbs compared to classic DMARDs [8]. Primary and secondary failures are not infrequent; moreover, < 50% of responder patients in clinical trials attained disease remission [9]. The treatment with anti-TNF blocking agents has high costs for the community [10]. While some of these drawbacks such as the increased risk of infection and neoplasm are presumably related to the blockade of TNF-a itself, others, such as the high production costs, and the risk of antidrug antibody (ADA) production with possible loss of efficacy and side effects, are proper to current anti-TNF-a agents, especially mAbs [11], and might be possibly overcome by alternative anti-TNF strategies. An alternative way to target TNF-a is active immunization, where a TNF-a derivative can be used as the immunogen to develop an anti-TNF-a active immunotherapy consisting in a vaccine [12]. The immunogen must be capable of disrupting B cell, but not T cell, tolerance to TNF-a, thereby eliciting the production of high titers neutralizing antibodies [13]. This strategy allows the production of polyclonal autologous anti-TNF-a antibodies potentially bypassing the risk of an anti xeno- or allogenic antibody response. Refining of ADA detection techniques allowed in fact detecting ADA in up to 40 and 30% of IFX and adalimumab treated patients, respectively [11]. The presence of ADA is associated with low trough drug levels, infusion-related reactions (for IFX) and therapeutic failure [14]. Active immunization offers then the possibility of overcoming this limitation. 546
The direct costs for anti-TNF blocking agents, together with the costs of drug administration, monitoring and side effect management, result in a heavy economical burden for the community [15], while the active immunization strategy might potentially be a less expensive alternative. Finally, the longer persistence of detectable anti-TNF-a antibody titers induced by active anti-TNF-a immunization draws a less cumbersome administration scenario for the patient, with possibly higher treatment acceptance.
Chemistry and preparation of the TNF-kinoid
3.
The preclinical proof-of-concept of active anti-TNF-a immunization with a compound called kinoid of human TNF-a (TNFK) has been established in a TNF-a-dependent animal model, the human TNF-a (hTNF-a) transgenic mice (TTG mice) [13,16,17] (Box 1). This has led to subsequent testing of TNFK in a Phase I clinical trial in Crohn’s disease. A Phase II clinical trial in previously anti-TNF-a treated RA patients having developed ADA is currently ongoing. TNFK belongs to a family of cytokine derivatives capable of acting as anti-cytokine vaccines called ‘kinoids’ [18]. Their name and preparation recalls those of the toxoids, detoxicated but still immunogenic products, derived from bacterial toxins by formalin treatment at 37 C for several days. At the beginning of the 1980s, a detoxication procedure using glutaraldehyde instead of formaldehyde was described for the preparation of fully atoxic polymerized antigens with high immunogenicity [19]. This technology with either glutaraldehyde or formaldehyde was then applied to cytokines in order to convert them into derivatives devoid of biological activity but capable, when administered in animals, of inducing anti-cytokine antibodies. These derivatives were called kinoids [20]. TNFK is a heterocomplex of inactivated hTNF-a and a carrier, the keyhole limpet hemocyanin (KLH). KLH is a heterogeneous copper-containing respiratory protein isolated from the mollusk Megathura crenulata belonging to a group of non-heme proteins called hemocyanins. It consists of two subunits isoforms with a molecular mass of 390 103 and 360 103 D, originating, respectively, two different oligomeric aggregates, KLH1 and KLH2. The molecular mass of the oligomers ranges from 4,500,000 to 13,000,000. Due to its large size and its numerous epitopes KLH is capable of inducing a substantial immune response; its abundance of lysine residues for haptens coupling, with a high hapten: carrier protein ratio, increases the likelihood of generating hapten-specific antibodies [21]. For preparing the heterocomplex, glutaraldehyde is used to couple hTNF-a to the KLH carrier protein. KLH, and then glutaraldehyde, are added to a solution of hTNF-a treated with dimethylsulfoxide, in a mixture of 1 molecule of KLH and 40 molecules of hTNF-a. After 45 min incubation at 4 C, the preparation is dialyzed against the working buffer and then treated with formaldehyde for 6 days at 37 C.
Expert Opin. Biol. Ther. (2011) 11(4)
Semerano, Assier, Delavalle´e & Boissier
Concentration and duration of aldehyde treatments have been adapted for hTNF-a in order to obtain a strong and persistent inactivation of its biological activity. The unreacted aldehyde is quenched by addition of glycine (0.1 M), leading to complex stabilization. The excess aldehyde is eliminated by dialysis against Dulbecco’s phosphate buffer solution (PBS) [13].
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
4.
Pharmacodynamics
It is assumed that TNFK is a heterocomplex in which KLH provides T epitopes and bears at its surface a high density of hTNF-a preserved B-epitopes. The aim of carrier proteins is to promote carrier-specific T-cell help to a B-cell polyclonal response [21]. Given that a high number of hTNF-a molecules are covalently bound to KLH, kinoid immunocomplexes will present a high density of hTNF-a antigens in their native conformation to the antibody-producing B cells to crosslink specific B-cell receptors [18]. TTG mouse, expressing hTNF-a as a self antigen, is the only relevant model to study TNF-induced anti-hTNF-a antibody production [13]. In all immunized mice in different study protocols, immunization with TNFK induced specific anti-hTNF-a antibodies as detected by ELISA [13,16,17]. In a protocol where mice received three injections of TNFK at days 0, 7 and 28, these antibodies, tested at day 122 after TNFK first injection, appeared to belong mainly to IgG1 (52%) and IgG2a (48%), with negligible amounts of IgG3, IgM and IgE [13]. Purified IgG from hyperimmune sera exhibited a high affinity for hTNF-a with Kd values ranging from 5 10-8 to 10-10 M and were able to block its interaction with the high affinity TNFRI (Kd of 0.6 nM) [22], resulting in undetectable circulating hTNF-a in immunized mice. Anti-hTNF-a antibodies have a neutralizing anti-TNF-a effect as confirmed both in vitro by L929 cytotoxicity assay, showing cytotoxicity inhibition by hyperimmune sera at dilutions up to 10-4, and in vivo, where purified IgG from sera of immunized mice prevented TNF-a-galactosamine lethal shock in recipient mice [13]. 5.
Pharmacokinetics
TNFK is mixed at a 1:1 ratio with the PBS and administered intramuscularly with the adjuvant ISA51 (Seppic, France). The latter is similar to Freund’s incomplete adjuvant and is composed of a mix of mineral oil and a surfactant of the mono-oleate family; it is currently used in immunotherapy of cancer and infectious diseases [23]. ISA51 is used in a 1:1 ratio with the mix TNFK--PBS to obtain a water-in-oil emulsion [18]. Different administration schedules have been tested in mice, involving two (at days 0 and 7), three (at days 0, 7 and 28) or four injections with dose regimens varying from 5 to 30 µg of TNFK [13,16,17].
Whatever the exact administration schedule, all immunization protocols were able to induce anti-hTNF-a antibodies in TTG mice. In a three injection scheme (30 + 30 + 7 µg at days 0, 7 and 28), anti hTNF-a antibodies were detectable at first bleeding as soon as 5 weeks after TNFK first injection [16]; they peaked at 6 -- 8 weeks after first injection [13], with a > 50% decline within 16 weeks. In a protocol with three injections of TNFK 4 µg at days 0, 7 and 28, a TNFK boost given 12 weeks after the TNFK first injection induced a significant increase in neutralizing anti-hTNF-a antibodies as soon as 3 weeks after the boost [17]. TNFK was first administered in humans in a Phase I -- II open-label dose escalation study on 13 patients with moderate to active Crohn’s disease, the TNFK001 study (http:// clinicaltrials.gov/ct2/show/NCT00808262). The administration schedule consisted of three injections of TNFK at days 0, 7 and 28 at doses of 60, 180 and 360 µg. Four patients received a fourth boost dose at 6 months. In all immunized patients, anti-TNF-a antibodies were detected, with a peak in titers between the fourth and the fifth week after first TNFK injection, and a 50% reduction within 12 weeks. The boost at 6 month resulted in a new peak in antibody titers 3 -- 4 weeks later [24]. As far as RA is concerned, a dose-finding Phase II clinical trial is currently ongoing in RA patients previously treated with anti-TNF agents having developed ADA. The primary goal of this trial is to demonstrate that active immunization with TNFK is able to induce polyclonal anti-TNF-a antibodies in RA patients previously treated with anti-TNF-a mAbs who underwent a secondary therapeutic failure (i.e., loss of clinical response) and have developed ADA. Among the inclusion criteria of these patients having an active RA is the positivity of antibodies against a TNF antagonist at screening or on a sample taken since discontinuation of IFX and/or adalimumab (http://www.controlled-trials.com/mrct/trial/772671/ TNFK003). 6.
Effects in animal models
TNFK immunization has proven its efficacy in the spontaneous arthritis of TTG mice thereby posing the rationale for its use in RA. When given before arthritis development, TNFK markedly reduced the clinical severity of arthritis and resulted in less histological joint inflammation and destruction compared to control mice [13,16]. In an experimental three injection protocol (days 0, 7 and 28), a highly significant difference in clinical and histological score was already evident when animals were sacrificed 6 weeks after the first injection, compared to controls. TNFKimmunized animals showed mild histological inflammation and no histological destruction. The co-administration of methotrexate did not change the results [16]. When, with the same experimental protocol, the observation was prolonged up to 17 weeks, arthritis onset
Expert Opin. Biol. Ther. (2011) 11(4)
547
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
Kinoid of human TNF-a
happened to be delayed by 9 weeks compared to controls and still low clinical and histological scores were found in immunized mice. The therapeutic efficacy, its duration and the effect of a TNFK boost were better evaluated in a subsequent experiment more resembling to a human disease scenario, as TTG mice were immunized after spontaneous arthritis onset [17]. In 12 weeks follow-up after TNFK immunization, arthritis was dramatically ameliorated, and clinical scores did not differ from those of mice treated with weekly IFX at a dose of 1 mg/kg over the same time period. These findings were corroborated by histology, showing low inflammation and no sign of cartilage destruction in immunized animals. The observation was prolonged to 30 weeks after TNFK first injection in order to study the duration of clinical effect and the kinetics of TNFK-induced anti-hTNF-a antibodies. After the initial amelioration, arthritis clinical score in immunized mice started to increase from week 12 after first injection to the end of the experiment. This trend was reversed by a TNFK boost given at week 12, before clinical degradation ensued. The worsening in clinical control of arthritis coincided with a decrease in anti-hTNF-a antibody titers, while the TNFK boost triggered a significant increase in antibody titers 3 weeks after its administration. Mild histological scores of joint inflammation, destruction and cartilage degradation at the end of the experiment confirmed the long-term prevention of structural damage of TNFK immunization. 7.
iv)
v)
vi)
Safety and tolerability
Some major safety issues are raised by the novel anti-TNF-a approach of active immunotherapy, namely: i) The delivered TNF-a must be devoid of toxicity but still be immunogenic, and this is the case of the TNFK heterocomplex, where aldehyde treatment results in a hTNF-a derivative satisfying these requirements. In all experiments conducted with TNFK, no short-term toxicity linked to its administration and ascribable to hTNF-a activity-related toxicity was detected [13,16,17]. This was the case even in the limited experience in humans. ii) The anti-TNF-a vaccination must result in rupture of B-cell but not of T-cell tolerance (i.e., vaccination must not induce memory, T cells capable of recognizing the native cytokine). In fact, the persistence of a T-cell population sensitized against a selfcytokine would result in a localized cellular response in its site of production. iii) This issue was addressed in an animal study where 6 -- 8 weeks old TTG mice received three injections of TNFK (days 0, 7, 28 ± a boost at day 90) and were followed up for 120 days after the first injection. Our group showed that the splenocytes 548
vii)
viii)
8.
from TNFK-immunized TTG mice did not trigger any cell-mediated immune response to self hTNF-a, as tested by T-cell proliferation and IL-2 and IFN-g production in culture supernatants, whatever the administration regimen of TNFK [13]. The only detectable cellular response was against KLH. Conversely in Balb/C mice, a TNFK-induced anti-hTNF-a cellular response was detected when hTNF-a (a heterologous antigen for this strain) was administered. In TNFK001 study in Crohn’s disease patients, stimulation of PBMCs of immunized patients with TNF-a failed to induce proliferation. The rupture of B-cell tolerance must be reversible. Our group demonstrated that when TTG mice were immunized with TNFK before spontaneous arthritis appearance, anti-hTNF-a antibodies peaked 6 -- 8 weeks after TNFK first injection and had a > 50% antibody titers decline within 12 -- 16 weeks. This kinetics is ascribable to short life of B-cell memory in the absence of a specific T-cell help [13]. A long-term study where immunized TTG mice were monitored up to 30 weeks after TNFK first injection immunization confirmed the same results [17]. A similar kinetics, albeit with the limitation of study design and sample size, seems to be confirmed in humans, based on the results of TNFK001 study. In the 13 immunized patients anti-TNF-a antibody titers were markedly reduced, and sometimes no longer detectable, within 12 -- 15 weeks after first injection. A raise in the levels of TNF-a induced by other stimuli (infections, tumors) must not elicit the production of anti-TNF antibodies after TNFK immunization. This was demonstrated in a study where monthly administration of hTNF-a to TTG mice failed to induce any raise of anti-hTNF-a antibodies [17]. Ideally, the ‘physiological’ activity of hTNF-a in normal tissues should be conserved (see points ii, iii and iv).
Conclusions
An important preclinical body of evidence (not inferior to that which first led to test a monoclonal anti-TNF-a antibody in 10 RA patients in 1992) supports the feasibility of antiTNF-a active immunization in TNF-a-dependent human diseases. The efficacy in TTG mice spontaneous arthritis, the relevant model for TNF-a inhibition, strongly suggests its potential application in RA. The reversibility of antiTNF-a antibody levels increase and the absence of memory T-cells induction are both arguments in favor of a good safety profile. The first results of an open-label study in Crohn’s
Expert Opin. Biol. Ther. (2011) 11(4)
Semerano, Assier, Delavalle´e & Boissier
disease are consistent with animal data regarding the kinetics of antibodies induction and decrease, and a good tolerance is suggested. A dose-finding randomized trial, ongoing at the present time in RA, will presumably provide more relevant safety and efficacy information determining whether or not TNFK will access the Phase III of clinical development.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
9.
Expert opinion
We are presently at an early phase of clinical development, as Phase II studies are ongoing in RA and Phase I -- II in Crohn’s is not ended, yet. The expert opinion is consequently based on proof-of-concepts experiments in preclinical and pharmacodynamics studies in mice that allow formulating some hypotheses. The active immunotherapy with TNFK aims to reversibly vaccinate against TNF-a. Unlike the already marketed antiTNF-a agents, one can suppose that using TNFK could have advantages in terms of simplicity and frequency of injections. The effect would probably be quite durable after each injection (several weeks or months). Moreover, TNFK treatment is not concerned by a possible reduction of effect due to ADA. These antibodies, found in up to 40% of IFXtreated and in 30% of adalimumab-treated patients, reduce the therapeutic efficacy of the drugs and are responsible of therapeutic failures and adverse reactions. So, the ADApositive patient might be a specific clinical situation in which TNFK administration could be warranted. Another advantage is a lower economic burden for the community as the costs of production of the kinoid would be presumably lower than those of current anti-TNF-a agents. Cost reductions are currently requested in developed countries and appear as a necessary condition for treating TNF-a-dependent diseases with targeted treatments in developing countries. The access for the patients to expensive
biological therapies is strongly limited in many countries by health authorities or other third party payers, and the choice of treatment will be more and more influenced by costeffectiveness analyses. In this scenario, a less expensive alternative providing ‘value’ and ‘value for money’ in RA treatment would certainly be welcomed. If the safety and efficacy data suggested by animal models are confirmed by ongoing human clinical studies, it is conceivable that TNFK will have a considerable impact on RA treatment strategies. Not only TNFK promises to be a direct competitor of passive anti-TNF-a immunotherapies, but also future scenarios might be conceived, including combination or sequential treatment with both passive and active TNF-a-targeting strategies. The reversibility of anti-TNF-a vaccination with TNFK and lack of induction of immunological memory versus the native cytokine are the key conditions for a favorable benefit:risk ratio. All preclinical studies show a bell curve of anti-TNF-a antibodies levels and preliminary results in humans confirm this point. The administration of TNF-a to TNFK-vaccinated animals fails to induce an anti-TNF-a response and, in addition, the persistence of residual levels of active TNF-a is probably sufficient to protect the host against infection and tumors. Nevertheless, all these safety considerations, based on animal models data, will have to be confirmed in ongoing and future clinical trials in humans.
Declaration of interest M-C Boissier has been a consultant for Neovacs, Inc. and his laboratory has received research grants from Neovacs, Pfizer, UCB Pharma and Roche. This manuscript was written without any interactions outside co-authors. The other authors declare no conflict of interest.
Expert Opin. Biol. Ther. (2011) 11(4)
549
Kinoid of human TNF-a
Bibliography 1.
2.
Expert Opin. Biol. Ther. Downloaded from informahealthcare.com by HINARI on 03/26/11 For personal use only.
3.
Alamanos Y, Voulgari PV, Drosos AA. Incidence and prevalence of rheumatoid arthritis, based on the 1987 American College of Rheumatology criteria: a systematic review. Semin Arthritis Rheum 2006;36(3):182-8 Detaille SI, Heerkens YF, Engels JA, et al. Common prognostic factors of work disability among employees with a chronic somatic disease: a systematic review of cohort studies. Scand J Work Environ Health 2009;35(4):261-81 Myasoedova E, Gabriel SE. Cardiovascular disease in rheumatoid arthritis: a step forward. Curr Opin Rheumatol 2010;22(3):342-7
4.
Franke LC, Ament AJ, van de Laar MA, et al. Cost-of-illness of rheumatoid arthritis and ankylosing spondylitis. Clin Exp Rheumatol 2009;27(4 Suppl 55):S118-23
5.
EvaluatePharma. Therapeutic focus -- anti-TNF market shows no sign of suppression. Available from: http:// www.evaluatepharma.com/Universal/ View.aspx?type=Story&id=185386& isEPVantage=yes 2009
6.
7.
8.
Koulinska I, Vasilakis-Scaramozza C, Crowell K. Rheumatoid Arthritis (Event Driven). Available from: http:// decisionresources.com/Products-andServices/Report.aspx?r=pcorim04xx 2010
12.
Bongartz T, Sutton AJ, Sweeting MJ, et al. Anti-TNF antibody therapy in rheumatoid arthritis and the risk of serious infections and malignancies: systematic review and meta-analysis of rare harmful effects in randomized controlled trials. JAMA 2006;295(19):2275-85 Sfikakis PP. The first decade of biologic TNF antagonists in clinical practice: lessons learned, unresolved issues and future directions. Curr Dir Autoimmun 2010;11:180-210
10.
Russell AS. Quality-of-life assessment in rheumatoid arthritis. Pharmacoeconomics 2008;26(10):831-46
Bartelds GM, Wijbrandts CA, Nurmohamed MT, et al. Anti-infliximab and anti-adalimumab antibodies in relation to response to adalimumab in infliximab switchers and anti-tumour necrosis factor naive patients: a cohort study. Ann Rheum Dis 2010;69(5):817-21 Capini CJ, Bertin-Maghit SM, Bessis N, et al. Active immunization against murine TNFalpha peptides in mice: generation of endogenous antibodies cross-reacting with the native cytokine and in vivo protection. Vaccine 2004;22(23-24):3144-53
13.
Le Buanec H, Delavallee L, Bessis N, et al. TNFalpha kinoid vaccination-induced neutralizing antibodies to TNFalpha protect mice from autologous TNFalpha-driven chronic and acute inflammation. Proc Natl Acad Sci USA 2006;103(51):19442-7
14.
Radstake TR, Svenson M, Eijsbouts AM, et al. Formation of antibodies against infliximab and adalimumab strongly correlates with functional drug levels and clinical responses in rheumatoid arthritis. Ann Rheum Dis 2009;68(11):1739-45
15.
Datamonitor. Rheumatoid Arthritis Market Forecast. Available from: http:// www.datamonitor.com/store/Product/ rheumatoid_arthritis_market_forecast? productid=IMHC0215 2009
9.
550
11.
Chen YF, Jobanputra P, Barton P, et al. A systematic review of the effectiveness of adalimumab, etanercept and infliximab for the treatment of rheumatoid arthritis in adults and an economic evaluation of their cost-effectiveness. Health Technol Assess 2006;10(42):iii-iv, xi-xiii, 1-229
16.
Delavallee L, Le Buanec H, Bessis N, et al. Early and long-lasting protection from arthritis in tumour necrosis factor alpha (TNFalpha) transgenic mice vaccinated against TNFalpha. Ann Rheum Dis 2008;67(9):1332-8
17.
Delavallee L, Semerano L, Assier E, et al. Active immunization to tumor necrosis factor-alpha is effective in treating chronic established inflammatory disease: a long-term study in a transgenic model of arthritis. Arthritis Res Ther 2009;11(6):R195
18.
Bizzini B, Drouet B, Zagury D, et al. Kinoids: a family of immunogens for active anticytokine immunotherapy
Expert Opin. Biol. Ther. (2011) 11(4)
applied to autoimmune diseases and cancer. Immunotherapy 2010;2(3):347-65 19.
Relyveld EH, Ben-Efraim S. Preparation of vaccines by the action of glutaraldehyde on toxins, bacteria, viruses, allergens, and cells. Methods Enzymol 1983;93:24-60
20.
Bizzini B, Achour A. “Kinoids”: the basis for anticytokine immunization and their use in HIV infection. Cell Mol Biol (Noisy-le-grand) 1995;41(3):351-6
21.
Harris JR, Markl J. Keyhole limpet hemocyanin (KLH): a biomedical review. Micron 1999;30(6):597-623
22.
Yang L, Lindholm K, Konishi Y, et al. Target depletion of distinct tumor necrosis factor receptor subtypes reveals hippocampal neuron death and survival through different signal transduction pathways. J Neurosci 2002;22(8):3025-32
23.
Aucouturier J, Ascarateil S, Dupuis L. The use of oil adjuvants in therapeutic vaccines. Vaccine 2006;24(Suppl 2):S2-44-5
24.
Benji P, Burger F, Kruger D, et al. Therapeutic immunization against TNF with a TNF-kinoid in Crohn’s disease patients is safe and induces anti-TNF antibodies. In: Presented at: GASTRO2009 International Meeting UEGW/WCOG; 20 -- 25 November 2009; London, UK
Affiliation Luca Semerano1,2, Eric Assier2, Laure Delavalle´e2 & Marie-Christophe Boissier†1,2 † Author for correspondence 1 Assistance Publique-Hoˆpitaux de Paris (AP-HP), Hoˆpital Avicenne, Rheumatology Department, 125 rue de Stalingrad, 93000 Bobigny, France 2 University of Paris-13, Sorbonne Paris-Cite´, EA4222, Li2P, 74 rue Marcel Cachin, 93000 Bobigny, France E-mail:
[email protected]
