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Cell-based myocardial repair: How should we proceed?
International Journal of
Cardiology ELSEVIER
International Journal of Cardiology 95 Suppl. 1 (2004) $8-S12
www.elsevier.com/locate/ijcard
Cell-based myocardial repair: How should we proceed? D o r i s A. Taylor P h D , Director, Centerfor Cardiovascular Repair, Departments of Physiology and Medicine, University of Minnesota, Minneapolis, MN 55455, USA
Abstract
Cell-based myocardial repair and regeneration heralds a new frontier in the treatment of cardiovascular disease. It provides an unprecedented opportunity to treat the underlying loss of cardiomyocytes that occurs after myocardial injury and that results in the cascade of events leading to heart failure. Yet, even as it progresses to the clinic, much remains to be understood about this technology. For example, controversies exist over the specific cells to be used, the cell dosages needed, how cells will impact the electrical activity of the myocardium, and even whether transplanted cells can actually improve myocardial function. We can perhaps answer these questions more quickly and more effectively - and thus benefit patients more rapidly - if we learn from the successes and failures of our gene therapy colleagues and take a prudent, step-wise approach from bench to bedside. To do so, we need only to promise what we can deliver, to do careful science, and then to deliver well on our promises. Although cellular cardiomyoplasty (cell transplantation for cardiac repair) shows great early clinical promise, its future as a new frontier in the treatment for cardiovascular disease will rest heavily on how we move forward in the next few years. Its success will heavily depend upon conducting carefully controlled, randomized double-blind clinical trials with appropriate endpoints, in the right patients. Choice of cell type, and mode of cell delivery, will also have to be considered, and may have to be matched to the patient. Irrespective of cell type, we can also be assured that cells offer both an opportunity for tissue repair and the potential for not yet understood outcomes. As with any frontier, there will be pitfalls and consequences to be considered that may surpass those of previous endeavors. But so too is the potential for previously unimagined success at treating the leading cause of death in the western world. In short, the promise for cardiovascular cell therapy is too great to be spoiled by ill-designed attempts that forget to account for both the natural propensities of cells and of the myocardium. © 2004 Published by Elsevier B.V. All rights reserved.
Keywords: Cell therapy; Myocardial repair; Clinical trials; Myoblasts; Stem cells
1. Background The idea is emerging that the adult heart, like other organs, contains a population of precursor, stem, or reserve cells, that are capable of some degree of endogenous repair when the myocardium is injured. However, it is clear in most cases, that after cardiac injury any demand for repair exceeds the capacity of the heart to respond. Thus, after acute myocardial injury, the damaged myocardium usually progresses through a cascade of events (shown schematically in Fig. 1) including inflammation, cardiocyte apoptosis, formation of a noncontracting fibrous scar, alteration in the workload of the surrounding myocardium (compensation) and, if the infarct is large enough, ultimately decompensation and deterioration to congestive heart failure (CHF). Current treatment options for these hearts undergoing this process include medical management, cardiac transplantation, mechanical circulatory assist devices (LVADs), or other experimental techniques * Correspondence address: 312 Church St SE, 7-105A BSBE, University of Minnesota, Minneapolis, MN 55455, USA. Tel.: +1-(612)-626-1416; fax: +1-(612)-626-1121. E-mail address." [email protected] 1389-9457/04/$ see front matter © 2004 Elsevier B.V. All rights reserved. doi: 10.1016/j.ijcard.2004.04.000
Cell type depends upon the injury
/ Hibernation Angiogenesis (Vascular cells) Bone marrow
MNC Angioblasts Stromal cells Growth factors Myoblasts +/- GF
Infarct/scar (Contractile cells) Skeletal myoblasts Cardiocytes/ cardiac stem cells Bone marrow
Fig. 1. Choosing a cell type for cardiac repair. Cell type choices likely depend on the injury and on the needed outcome (i.e. angiogenesis versus myogenesis).
(gene therapy, artificial hearts) all of which attempt to support the remaining healthy myocardium or, when that is not possible, to replace the damaged organ. In light of the limited efficacy and co-morbidity of these current treatment options, alternative, additional long-term therapeutic strategies are needed. Supplying cells capable of cardiac repair is one potential new therapeutic option.
D.A. Taylor~International Journal of Cardiology 95 Suppl. 1 (2004) $8-S12
2. The state of the art A cell transplantation technology called cellular cardiomyoplasty or more recently cellular cardiomyogenesis is the notion that if we can deliver stem or progenitor cells to the site of cardiac injury, we can restore blood flow and contractility to previously infarcted, scarred or dysfunctional heart. Preclinically, many types of cells have been transplanted into injured myocardium - including fetal cardiomyocytes [1], autologous skeletal myoblasts [2-4] smooth muscle cells [5], immortalized myoblasts [6], syngeneic skeletal myoblasts [7], fibroblasts [8], adult cardiac-derived cells [9], embryonic stem cells [10] and bone-marrow-derived stromal [11] or mononuclear cells [12,13]. Although several of these cell types may hold future promise as a therapeutic option, to date only skeletal myoblasts and bone-marrow-derived cells have been used in safety studies as a first step toward myocardial repair [13-16].
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decrease the likelihood of tumor formation after cell transplantation [19]. By using myogenic (versus non-myogenic) cells, we increase the likelihood of regenerating contractile muscle in a previously infarcted region. Finally, by using relatively ischemia-resistant skeletal myoblasts rather than cardiocytes, we can obtain higher levels of cell engraftment and survival - in infarcted regions of the heart where cardiocytes perish [20]. Approximately 15 years of pre-clinical data have shown that autologous skeletal myoblasts can differentiate into striated muscle cells within the damaged myocardium [2] and that these cells can augment both diastolic and systolic myocardial performance in a number of animal species after both acute and chronic injury [2,7,21,22]. Based on these suggestions that regeneration of functional muscle in infarct is possible after myoblast implantation, clinical studies based have begun both in Europe and in the USA.
5. Clinical studies 3. The choice of cell type As indicated in Fig. 1, the choice of cell type may depend upon the injury. In conditions where chronic ischemia prevails, or where reperfusion is the primary objective, nonmyogenic bone-marrow-derived cells are a logical cell choice because these cells are thought to be primarily angiogenic. Under these conditions, bone-marrow mononuclear cells, bone-marrow or peripheral blood endothelial or vascular progenitor cells, angioblasts [17] or multi-potent adult progenitor cells [18] may be useful. However, in patients where restoration of contractile function is the clinical goal, such as those with end-stage heart failure or at early times post infarction when blood flow has been restored but cells have died, then delivering cells with contractile potential seems a more reasonable approach. Under these conditions, myoblasts appear the logical first choice although cardiac stem cells or other mesenchymal progenitors driven down a muscle lineage may also be of use.
4. Autologous skeletal myoblasts The first cells proposed clinically for cardiac repair were autologous skeletal muscle-derived progenitor cells called myoblasts. Using these cells to repair injured heart is straightforward. It requires obtaining a skeletal muscle biopsy, expanding a patient's cells in the laboratory and injecting those cells into the patient's heart either surgically or percutaneously. By using self-derived (autologous) skeletal myoblasts, we are able to overcome the major limitations associated with allogeneic cell-based treatments. Most notably, we overcome any shortage of donor tissue, the need for immunosuppression, and the ethical dilemma associated with the use of allogeneic or embryonic cells. By using primary cells rather than immortalized or totipotent stem cells, we
In the first clinical cell therapy trial Menasche and colleagues directly injected muscle-derived cells into the scarred region of the left ventricular in 10 patients undergoing concomitant coronary artery bypass grafting (CABG). In these patients over several years of follow-up, significant improvements in LVEF have been reported as have reports of increased regional wall motion in the areas where cells were injected. However, the concomitant CABG limits any functional interpretation of these studies. Nonetheless, these positive data provided a basis for initiation of a randomized controlled trial that was begun in 2003 (MAGIC trial). The results of that trial are eagerly anticipated. In a second myoblast study, cells were delivered as a sole therapy rather than as an adjunct to CABG [23]. In this safety study where patients were on average 6 to 7 years post infarct, trends toward increased myocardial perfusion and improvement in NYHA heart failure class I were also reported. Taken together these data suggest that myoblasts can be delivered late in the disease process (at least one month after injury and as long as 6 years), can engraft into infarcted regions of myocardium and survive for extended periods of time, can increase diastolic compliance, and improve myocardial wall thickening as well as contractility not only in the injured regions but in the surrounding myocardium. Although the pre-clinical and initial clinical data appear positive, myoblast CCM is not without potential limitations. For example, using autologous ceils necessitates sufficient time between injury and injection to grow cells in the laboratory. Yet, this waiting period may be right in line with nature's demand in that injecting cells too soon after an infarct seems to limit the number of cells that survive presumably due to the inflammatory response that occurs after an injury. Even though it is the most well-defined technique for myocardial regeneration, many important questions remain - questions that will need to be answered for any
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D.A. Taylor~International Journal of Cardiology 95 Suppl. 1 (2004) $8-S12
CCM technique. Probably the most important questions are those related to cell survival, integration, differentiation and functional effect. For example, if CCM is to be viewed as a long-term solution to myocardial injury, cells must be able to survive for years in heart; yet, the long-term fate of myoblasts is unclear. Similarly, it is not yet clear if and how myoblasts electrically integrate into surrounding myocardium and what impact integration may have on either function or rhythmicity. In fact, early clinical data suggest that myoblast transplantation may be associated with a transient period of electrical instability in a subset of patients who receive the cells. However, whether this reflects a natural response of the heart to multiple injections, is related to cell integration, differentiation or even death remains unclear. Finally, there is some debate as to whether myoblasts improve contraction or just prevent further deterioration of the injured heart. Although either may be desirable in patients after an acute myocardial injury, a better understanding of this process could help us know when to intervene. These debates are being resolved with more research and the future for myoblast transplantation appears promising. Furthermore, the understanding gained about myoblast CCM will provide a basis for developing second- and third-generation cells for cardiac repair.
6. Bone marrow stem cells Bone-marrow-derived stem cells have received much attention of late as an ideal target for cardiac repair [11,12,24]. By their very nature, these multipotent stem cells respond to their microenvironment and develop a corresponding phenotype. This would suggest that in normal myocardium, undifferentiated stromal cells could become cardiocytes and integrate with surrounding host tissue. Data from several laboratories suggest this is feasible - although it appears to be a rare event [25]. But, it further suggests, as shown by Chiu and others [26], that when injected into injured or infarcted heart, stromal cells can develop characteristics of scar. More recent data from our group suggests that when injected throughout the scar and nearby healthy tissue, some stromal cells can differentiate into striated muscle albeit not cardiac muscle - and positively impact cardiac function [27]. Understanding this phenomenon and beginning to control it is an active area of investigation that must be better understood before stromal cells per se will be a viable target for myocardial repair. Within the last few years, data have emerged suggesting that lineage-negative (Lin-) c-kitP°s sub-populations of bone marrow cells can be derived and supplied in numbers sufficient to repopulate infarcted regions of myocardium with cardiocytes and vascular structures [12]. Although this is an extremely exciting development, these data have yet to be confirmed in mouse by other groups or in larger animal models. In fact, the original investigators have been unable to reproduce these data in non-human primates [28], and recent
data from two groups call into question the findings of the original study [29]. Furthermore, the functional impact of these cells even in the original study has not rigorously been evaluated. Translating these initial results into larger more physiologically relevant models of human cardiac disease is a critical next step in evaluating marrow-derived stem cell contribution to cardiac repair.
7. Clinical trials: TOPCARE-AMI and STAMM A clinical study using AC133+ bone marrow cells, which is a population containing both hematopoietic and multipotential stem cells, was initiated in 6 patients with prior myocardial infarction and concomitant coronary artery bypass grafting [16]. The cells were injected into the border zone of the infarct at a dose of 1-5x 106 cells and the function was measured 9-16 months after the surgery. None of the patients experienced ventricular arrhythmia, and the leftventricular fimction was enhanced in 4 patients and the infarct tissue perfusion was improved in 5 patients. Thus this study suggests that implantation of this population of stem cells increased angiogenesis without affecting safety in patients. In a similar study, TOPCARE-AMI, the benefits of implanting endothelial progenitor cells (EPC) or bonemarrow mononuclear cells were compared [13]. In this study cells were directly infused into coronary circulation of patients within 3-9 days of an acute myocardial infarction. Out of 20 patients, 11 received EPCs and 9 BMCs with both groups showing significant improvement in perfusion, ejection fraction and regional wall motion at 4 months. No difference was observed between the EPC and BMC groups. Taken together, these data illustrate the potential of bonemarrow cells to improve myocardial performance secondary to increasing cardiac perfusion. It also raises the hope for the potential of utilizing a multipotent stem cell for cardiac repair.
8. Cardiocytes and cardiac stem cells Although skeletal myoblasts appear to be the best first generation cell for use in myocardial repair, the intriguing question that remains is "what cell type will ultimately be best for myocardial repair?" At first glance, cardiocytes might seem the ideal target cell. Yet, several major obstacles remain to the use of these cells in vivo. First, for cardiocytes to be used for cell transplantation, they must be readily available as a cell source. Given their inability to replicate to a significant degree in vitro or in vivo, this remains unlikely at present. Second, for cardiocytes to survive in infarcted heart, they will require a vascular supply greater than that required for myoblast survival and one far greater than is available in infarct, which suggests that they will only be usable in conjunction with revascularization or angiogenesis. Until recently, the heart was thought to be devoid
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D.A. Taylor~InternationalJournal of Cardiology 95 Suppl. 1 (2004) $8-S12
Table 1 Comparison of myoblasts and bone-marrow-derivedmononuclear cells for CV repair a Cell Type
Dose
Patients
Delivery
Time after i n j u r y
Myoblasts
25-1200 million
Post M I / C H F
Intra-cardiac, >1 month to >6 years surgically or percutaneous
Reverseremodeling (myogenesis) Increased wall motion Increased contractility
Bone-marrow-derived stem/progenitor cells
2-250 million (includingEPCs as a small subset)
Acute MI/ refractory angina
Intra-coronary
Angiogenesis Increased perfusion Increased viability Increased wall motion
3-9 days
Outcomes
a Although these cell types are often compared in their ability to repair injured myocardium, these comparisons are not valid. As indicated cell type, cell dose, patient populations, delivery methods and time after injury are different. Thus it is not surprising that outcomes are different. Direct side by side comparisons are needed in similar patients or animal models before outcome can be attributed to cell type. of stem or progenitor cells capable of regenerating new cardiocytes. However, in the past several years several groups have demonstrated the purification of sca-l+c-kit+ cells from adult myocardium and showed that these cells, under specific defined conditions, can differentiate into cardiocytes in vitro [30-32]. The emergence of these cells opens another door in the armamentarium of cells available for cardiac repair. However, how these cells can be obtained, whether they can be grown to sufficient quantities in vitro or whether they can be mobilized in civo for endogenous repair remain completely unanswered questions.
cells Angiogenesis
Myogenesis
Injxn Rescue hibernating heart Grow/recruit new muscle Change Compliance Contraction wall stress improves improves Cytokine/neurohumoral status changes/ endogenous stem cells respond
Fig. 2. Possible effects of cells in injured myocardium. 9. C h a l l e n g e s to c e l l u l a r c a r d i o m y o p l a s t y
Several challenges remain if we are to successfully repair infarcted or failing myocardium with any type of cells. The majority of these challenges seem straightforward, but they are complicated by the extreme heterogeneity of cardiovascular disease. For example, the criteria for reproducible engraftment of large numbers of cells may be very different in the early post-MI patient where angiogenesis is the goal versus the patient with end-stage cardiac dysfunction where myogenesis is needed (Table 1). Similarly, the method of delivering cells (surgical versus endovascular), the concentration of cells delivered and a host of other criteria ranging from age to co-existing disease states have to be considered as cell transplantation emerges into clinical trials. Finally, the most significant challenges to cell repair of injured myocardium evolve from our attempts to do more than simply halt the progression of cardiovascular deterioration. Pre-clinical data suggest that growth factors, fibroblasts or any number of non-myogenic cells that increase perfusion may be able to slow deterioration or actually repair diastolic dysfunction [8,33]. Yet, if we wish to improve contractility of infarcted heart, we must implant cells that can electromechanically integrate into heart, thicken or contract in synchrony, and survive for extended periods of time (Fig. 2). This may very well involve more than simple cell engraftment and may ultimately depend upon combined
approaches where we used cells that can augment vessel growth and those that can regenerate muscle. It may involve combined cell- and tissue-engineering approaches where we create biologic devices in which we can guarantee nutrient delivery and blood supply to an infarcted region of tissue. Or it may involve cell- and gene-based approaches where we generate intelligent cells that can home only to the site of cardiovascular injury and not to other unwanted sites within the patient. Although these are not at the forefront of myocardial repair today, they are areas that may well form the next generation of cell-based products for cardiovascular repair.
10. C o n c l u s i o n
Cell transplantation for myocardial repair is here. Carefully evaluating this new modality provides an opportunity to define a new era in the treatment of cardiovascular disease. Combining genomics, tissue engineering and advanced imaging methods to find the best cell type(s), to understand the mechanism(s) by which cell engraftment improves function, to promote graft longevity, and to improve electromechanical graft integration are the next pre-clinical frontiers. Designing clinical trials that allow us to adequately evaluate the safety and efficacy of cell transplantation in patients with both acute and chronic cardiac disease but in the absence of complicating adjunctive therapies is crucial [34].
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D.A. Taylor~International Journal of Cardiology 95 SuppL 1 (2004) $8-S12
In short, basing clinical trials on good pre-clinical data in multiple animal species, carefully defining the cells we use, rigorously demonstrating safety and efficacy to find the best cell type for a given cardiac injury, and performing randomized controlled trials that can be compared worldwide are the tasks before us as a field. Accepting the challenge, and pushing back this frontier is our opportunity. Changing patient outcomes will be our prize. Yet, doing it improperly could relegate this promising field to failure before its potential is realized.
References [l] Koh GY, et al. Long-term survival of AT-1 cardiomyocyte grafts in syngeneic myocardium. Am J Physiol 1993;264:1727-1733. [2] Taylor DA, et al. Regenerating functional myocardium: improved performance after skeletal myoblast transplantation. Nat Med 1998;4:929-33. [3] Marelli D, et al. Cell transplantation for myocardial repair: an experimental approach. Cell Transpl 1992;1:383-390. [4] Kessler PD, Byrne BJ. Myoblast cell grafting into heart muscle: cellular biology and potential applications. Annu Rev Physiol 1999;61:219242. [5] Li R, et al. Smooth muscle cell transplantation into myocardial scar tissue improves heart function. J Mol Cell Cardiol 1999;31:513-522. [6] Robinson SW, et al. Arterial delivery of genetically labelled skeletal myoblasts to the murine heart: long-term survival and phenotypic modification of implanted myoblasts. Cell Transpl 1996;5:77-91. [7] Murray CE, et al. Skeletal myoblast transplantation for repair of myocardial necrosis. J Clin Invest 1996;98:2512-23. [8] Hutcheson KA, et al. Comparing the benefits of cellular cardiomyoplasty with skeletal myoblasts or dermal fibroblasts on myocardial performance. Cell Transpl 2000;9:359-368. [9] Li R, et al. Survival and function of bioengineered cardiac grafts. Circulation 1999;100(Suppl II):63-69. [10] Klug MA, et al. Purification of cardiac myocytes differentiated from embryonic stern cells for use in intracardiac grafting. Tissue Engineering. 1996. Taos, New Mexico: Keystone Symposia. [11] Wang JS, et al. Marrow stromal cells for cellular cardiomyoplasty: the importance of microenvironment for milieu dependent differentiation. Circulation 2000;102(Suppl II):683. [12] Orlic D, et al. Bone marrow cells regenerate infarcted myocardium. Nature 2001;410:701-705. [13] Assmus B, et al. Transplantation of progenitor cells and regeneration enhancement in acute myocardial infarction (TOPCARE-AMI). Circulation 2000;106:3009-3017. [14] Menasche P, et al. Myoblast transplantation for heart failure. Lancet 2001;357:279-280. [15] Menasche P. Cellular transplantation: hurdles remaining before widespread clinical use. Curt Opin Cardiol 2004; 19:154-161.
[16] Stamm C, et al. Autologous bone-marrow stem-cell transplantation for myocardial regeneration. Lancet 2003;361:45-46. [17] Itescu S, Kocher AA, Schuster MD. Myocardial neovascularization by adult bone marrow-derived angioblasts: strategies for improvement of cardiomyocyte function. Heart Fail Rev 2003;8:253-258. [18] Jiang Y, et al. Pluripotency of mesenchymal stem cells derived from adult marrow. Nature 2002;418:41-49. [19] Tremblay JP, Roy B, Goulet M. Human myoblast transplantation: a simple model for tumorigenicity. Neuromuscul Disord 1991;1:341343. [20] Field L. Future therapy for cardiovascular disease. NHLBI Workshop Cell Transplantation: Future Therapy for Cardiovascular Disease 1998. Columbia, MD. [21] Atkins BZ, et al. Myogenic cell transplantation improves in uioo regional performance in infarcted rabbit myocardium. J Heart Lung Transpl 1999;18:1173-1180. [22] Chachques J, et al. Cellular therapy reverses myocardial dysfimction. American Association for Thoracic Surgery 80th Annual Meeting 2000. [23] Smits PC, et al. Catheter-based intramyocardial injection of antologous skeletal myoblasts as a primary treatment of ischemic heart failure: clinical experience with six-month follow-up. J Am Coll Cardiol 2003;42:2063-2069. [24] Toma C, Pittenger MF, Kessler PD. Adult human mesenchymal stem cells differentiate to a striated muscle phenotype following arterial delivery to the murine heart. Circulation 2000; 102(Suppl II):683. [25] Bittner RE, et al. Recruitment of bone-marrow-derived cells by skeletal and cardiac muscle in adult dystrophic mdx mice. Anat Embryol 1999;199:391-396. [26] Chiu RCJ, Zibaitis A, Kao RL. Cellular cardiomyoplasty: myocardial regeneration with satellite cell implantation. Ann Thorac Surg 1995;60:12-18. [27] Thompson RB, et al. Comparison of intracardiac cell transplantation: autologous skeletal myoblasts versus bone marrow cells. Circulation 2003;108(Suppl 1):II264-271. [28] Orlic D, et al. Cytokine mobilized CD34+ cells do not benefit rhesus monkeys following induced myocardial infarction. Blood 2002;100:28a-29a. Abstract. [29] Murry CE, et al. Haematopoietic stem cells do not transdifferentiate into cardiac myocytes in myocardial infarcts. Nature 2004;428:664-668. [30] Balsam LB, et al. Haematopoietic stem cells adopt mature haematopoietic fates in ischaemic myocardium. Nature 2004;428:668673. [31] Quaini F, et al. Cbimerism of the transplanted heart. N Engl J Med 2002;346:5-15. [32] Jackson KA, et al. Regeneration of ischemic cardiac muscle and vascular endothelium by adult stem cells. J Clin Invest 2001;107:13951402. [33] Atkins BZ, et al. Cellular cardiomyoplasty improves diastolic properties of injured heart. J Surg Res 1999;85:234-42. [34] Ellis M, Russell S, Taylor D. Translating cell transfer for cardiovascular disease to the bedside: a pre-clinical review and discussion of potential early trials. Card Vasc Regen 2000;3:197-203.
Cardiology ELSEVIER
International Journal of Cardiology 95 Suppl. 1 (2004) $8-S12
www.elsevier.com/locate/ijcard
Cell-based myocardial repair: How should we proceed? D o r i s A. Taylor P h D , Director, Centerfor Cardiovascular Repair, Departments of Physiology and Medicine, University of Minnesota, Minneapolis, MN 55455, USA
Abstract
Cell-based myocardial repair and regeneration heralds a new frontier in the treatment of cardiovascular disease. It provides an unprecedented opportunity to treat the underlying loss of cardiomyocytes that occurs after myocardial injury and that results in the cascade of events leading to heart failure. Yet, even as it progresses to the clinic, much remains to be understood about this technology. For example, controversies exist over the specific cells to be used, the cell dosages needed, how cells will impact the electrical activity of the myocardium, and even whether transplanted cells can actually improve myocardial function. We can perhaps answer these questions more quickly and more effectively - and thus benefit patients more rapidly - if we learn from the successes and failures of our gene therapy colleagues and take a prudent, step-wise approach from bench to bedside. To do so, we need only to promise what we can deliver, to do careful science, and then to deliver well on our promises. Although cellular cardiomyoplasty (cell transplantation for cardiac repair) shows great early clinical promise, its future as a new frontier in the treatment for cardiovascular disease will rest heavily on how we move forward in the next few years. Its success will heavily depend upon conducting carefully controlled, randomized double-blind clinical trials with appropriate endpoints, in the right patients. Choice of cell type, and mode of cell delivery, will also have to be considered, and may have to be matched to the patient. Irrespective of cell type, we can also be assured that cells offer both an opportunity for tissue repair and the potential for not yet understood outcomes. As with any frontier, there will be pitfalls and consequences to be considered that may surpass those of previous endeavors. But so too is the potential for previously unimagined success at treating the leading cause of death in the western world. In short, the promise for cardiovascular cell therapy is too great to be spoiled by ill-designed attempts that forget to account for both the natural propensities of cells and of the myocardium. © 2004 Published by Elsevier B.V. All rights reserved.
Keywords: Cell therapy; Myocardial repair; Clinical trials; Myoblasts; Stem cells
1. Background The idea is emerging that the adult heart, like other organs, contains a population of precursor, stem, or reserve cells, that are capable of some degree of endogenous repair when the myocardium is injured. However, it is clear in most cases, that after cardiac injury any demand for repair exceeds the capacity of the heart to respond. Thus, after acute myocardial injury, the damaged myocardium usually progresses through a cascade of events (shown schematically in Fig. 1) including inflammation, cardiocyte apoptosis, formation of a noncontracting fibrous scar, alteration in the workload of the surrounding myocardium (compensation) and, if the infarct is large enough, ultimately decompensation and deterioration to congestive heart failure (CHF). Current treatment options for these hearts undergoing this process include medical management, cardiac transplantation, mechanical circulatory assist devices (LVADs), or other experimental techniques * Correspondence address: 312 Church St SE, 7-105A BSBE, University of Minnesota, Minneapolis, MN 55455, USA. Tel.: +1-(612)-626-1416; fax: +1-(612)-626-1121. E-mail address." [email protected] 1389-9457/04/$ see front matter © 2004 Elsevier B.V. All rights reserved. doi: 10.1016/j.ijcard.2004.04.000
Cell type depends upon the injury
/ Hibernation Angiogenesis (Vascular cells) Bone marrow
MNC Angioblasts Stromal cells Growth factors Myoblasts +/- GF
Infarct/scar (Contractile cells) Skeletal myoblasts Cardiocytes/ cardiac stem cells Bone marrow
Fig. 1. Choosing a cell type for cardiac repair. Cell type choices likely depend on the injury and on the needed outcome (i.e. angiogenesis versus myogenesis).
(gene therapy, artificial hearts) all of which attempt to support the remaining healthy myocardium or, when that is not possible, to replace the damaged organ. In light of the limited efficacy and co-morbidity of these current treatment options, alternative, additional long-term therapeutic strategies are needed. Supplying cells capable of cardiac repair is one potential new therapeutic option.
D.A. Taylor~International Journal of Cardiology 95 Suppl. 1 (2004) $8-S12
2. The state of the art A cell transplantation technology called cellular cardiomyoplasty or more recently cellular cardiomyogenesis is the notion that if we can deliver stem or progenitor cells to the site of cardiac injury, we can restore blood flow and contractility to previously infarcted, scarred or dysfunctional heart. Preclinically, many types of cells have been transplanted into injured myocardium - including fetal cardiomyocytes [1], autologous skeletal myoblasts [2-4] smooth muscle cells [5], immortalized myoblasts [6], syngeneic skeletal myoblasts [7], fibroblasts [8], adult cardiac-derived cells [9], embryonic stem cells [10] and bone-marrow-derived stromal [11] or mononuclear cells [12,13]. Although several of these cell types may hold future promise as a therapeutic option, to date only skeletal myoblasts and bone-marrow-derived cells have been used in safety studies as a first step toward myocardial repair [13-16].
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decrease the likelihood of tumor formation after cell transplantation [19]. By using myogenic (versus non-myogenic) cells, we increase the likelihood of regenerating contractile muscle in a previously infarcted region. Finally, by using relatively ischemia-resistant skeletal myoblasts rather than cardiocytes, we can obtain higher levels of cell engraftment and survival - in infarcted regions of the heart where cardiocytes perish [20]. Approximately 15 years of pre-clinical data have shown that autologous skeletal myoblasts can differentiate into striated muscle cells within the damaged myocardium [2] and that these cells can augment both diastolic and systolic myocardial performance in a number of animal species after both acute and chronic injury [2,7,21,22]. Based on these suggestions that regeneration of functional muscle in infarct is possible after myoblast implantation, clinical studies based have begun both in Europe and in the USA.
5. Clinical studies 3. The choice of cell type As indicated in Fig. 1, the choice of cell type may depend upon the injury. In conditions where chronic ischemia prevails, or where reperfusion is the primary objective, nonmyogenic bone-marrow-derived cells are a logical cell choice because these cells are thought to be primarily angiogenic. Under these conditions, bone-marrow mononuclear cells, bone-marrow or peripheral blood endothelial or vascular progenitor cells, angioblasts [17] or multi-potent adult progenitor cells [18] may be useful. However, in patients where restoration of contractile function is the clinical goal, such as those with end-stage heart failure or at early times post infarction when blood flow has been restored but cells have died, then delivering cells with contractile potential seems a more reasonable approach. Under these conditions, myoblasts appear the logical first choice although cardiac stem cells or other mesenchymal progenitors driven down a muscle lineage may also be of use.
4. Autologous skeletal myoblasts The first cells proposed clinically for cardiac repair were autologous skeletal muscle-derived progenitor cells called myoblasts. Using these cells to repair injured heart is straightforward. It requires obtaining a skeletal muscle biopsy, expanding a patient's cells in the laboratory and injecting those cells into the patient's heart either surgically or percutaneously. By using self-derived (autologous) skeletal myoblasts, we are able to overcome the major limitations associated with allogeneic cell-based treatments. Most notably, we overcome any shortage of donor tissue, the need for immunosuppression, and the ethical dilemma associated with the use of allogeneic or embryonic cells. By using primary cells rather than immortalized or totipotent stem cells, we
In the first clinical cell therapy trial Menasche and colleagues directly injected muscle-derived cells into the scarred region of the left ventricular in 10 patients undergoing concomitant coronary artery bypass grafting (CABG). In these patients over several years of follow-up, significant improvements in LVEF have been reported as have reports of increased regional wall motion in the areas where cells were injected. However, the concomitant CABG limits any functional interpretation of these studies. Nonetheless, these positive data provided a basis for initiation of a randomized controlled trial that was begun in 2003 (MAGIC trial). The results of that trial are eagerly anticipated. In a second myoblast study, cells were delivered as a sole therapy rather than as an adjunct to CABG [23]. In this safety study where patients were on average 6 to 7 years post infarct, trends toward increased myocardial perfusion and improvement in NYHA heart failure class I were also reported. Taken together these data suggest that myoblasts can be delivered late in the disease process (at least one month after injury and as long as 6 years), can engraft into infarcted regions of myocardium and survive for extended periods of time, can increase diastolic compliance, and improve myocardial wall thickening as well as contractility not only in the injured regions but in the surrounding myocardium. Although the pre-clinical and initial clinical data appear positive, myoblast CCM is not without potential limitations. For example, using autologous ceils necessitates sufficient time between injury and injection to grow cells in the laboratory. Yet, this waiting period may be right in line with nature's demand in that injecting cells too soon after an infarct seems to limit the number of cells that survive presumably due to the inflammatory response that occurs after an injury. Even though it is the most well-defined technique for myocardial regeneration, many important questions remain - questions that will need to be answered for any
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D.A. Taylor~International Journal of Cardiology 95 Suppl. 1 (2004) $8-S12
CCM technique. Probably the most important questions are those related to cell survival, integration, differentiation and functional effect. For example, if CCM is to be viewed as a long-term solution to myocardial injury, cells must be able to survive for years in heart; yet, the long-term fate of myoblasts is unclear. Similarly, it is not yet clear if and how myoblasts electrically integrate into surrounding myocardium and what impact integration may have on either function or rhythmicity. In fact, early clinical data suggest that myoblast transplantation may be associated with a transient period of electrical instability in a subset of patients who receive the cells. However, whether this reflects a natural response of the heart to multiple injections, is related to cell integration, differentiation or even death remains unclear. Finally, there is some debate as to whether myoblasts improve contraction or just prevent further deterioration of the injured heart. Although either may be desirable in patients after an acute myocardial injury, a better understanding of this process could help us know when to intervene. These debates are being resolved with more research and the future for myoblast transplantation appears promising. Furthermore, the understanding gained about myoblast CCM will provide a basis for developing second- and third-generation cells for cardiac repair.
6. Bone marrow stem cells Bone-marrow-derived stem cells have received much attention of late as an ideal target for cardiac repair [11,12,24]. By their very nature, these multipotent stem cells respond to their microenvironment and develop a corresponding phenotype. This would suggest that in normal myocardium, undifferentiated stromal cells could become cardiocytes and integrate with surrounding host tissue. Data from several laboratories suggest this is feasible - although it appears to be a rare event [25]. But, it further suggests, as shown by Chiu and others [26], that when injected into injured or infarcted heart, stromal cells can develop characteristics of scar. More recent data from our group suggests that when injected throughout the scar and nearby healthy tissue, some stromal cells can differentiate into striated muscle albeit not cardiac muscle - and positively impact cardiac function [27]. Understanding this phenomenon and beginning to control it is an active area of investigation that must be better understood before stromal cells per se will be a viable target for myocardial repair. Within the last few years, data have emerged suggesting that lineage-negative (Lin-) c-kitP°s sub-populations of bone marrow cells can be derived and supplied in numbers sufficient to repopulate infarcted regions of myocardium with cardiocytes and vascular structures [12]. Although this is an extremely exciting development, these data have yet to be confirmed in mouse by other groups or in larger animal models. In fact, the original investigators have been unable to reproduce these data in non-human primates [28], and recent
data from two groups call into question the findings of the original study [29]. Furthermore, the functional impact of these cells even in the original study has not rigorously been evaluated. Translating these initial results into larger more physiologically relevant models of human cardiac disease is a critical next step in evaluating marrow-derived stem cell contribution to cardiac repair.
7. Clinical trials: TOPCARE-AMI and STAMM A clinical study using AC133+ bone marrow cells, which is a population containing both hematopoietic and multipotential stem cells, was initiated in 6 patients with prior myocardial infarction and concomitant coronary artery bypass grafting [16]. The cells were injected into the border zone of the infarct at a dose of 1-5x 106 cells and the function was measured 9-16 months after the surgery. None of the patients experienced ventricular arrhythmia, and the leftventricular fimction was enhanced in 4 patients and the infarct tissue perfusion was improved in 5 patients. Thus this study suggests that implantation of this population of stem cells increased angiogenesis without affecting safety in patients. In a similar study, TOPCARE-AMI, the benefits of implanting endothelial progenitor cells (EPC) or bonemarrow mononuclear cells were compared [13]. In this study cells were directly infused into coronary circulation of patients within 3-9 days of an acute myocardial infarction. Out of 20 patients, 11 received EPCs and 9 BMCs with both groups showing significant improvement in perfusion, ejection fraction and regional wall motion at 4 months. No difference was observed between the EPC and BMC groups. Taken together, these data illustrate the potential of bonemarrow cells to improve myocardial performance secondary to increasing cardiac perfusion. It also raises the hope for the potential of utilizing a multipotent stem cell for cardiac repair.
8. Cardiocytes and cardiac stem cells Although skeletal myoblasts appear to be the best first generation cell for use in myocardial repair, the intriguing question that remains is "what cell type will ultimately be best for myocardial repair?" At first glance, cardiocytes might seem the ideal target cell. Yet, several major obstacles remain to the use of these cells in vivo. First, for cardiocytes to be used for cell transplantation, they must be readily available as a cell source. Given their inability to replicate to a significant degree in vitro or in vivo, this remains unlikely at present. Second, for cardiocytes to survive in infarcted heart, they will require a vascular supply greater than that required for myoblast survival and one far greater than is available in infarct, which suggests that they will only be usable in conjunction with revascularization or angiogenesis. Until recently, the heart was thought to be devoid
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Table 1 Comparison of myoblasts and bone-marrow-derivedmononuclear cells for CV repair a Cell Type
Dose
Patients
Delivery
Time after i n j u r y
Myoblasts
25-1200 million
Post M I / C H F
Intra-cardiac, >1 month to >6 years surgically or percutaneous
Reverseremodeling (myogenesis) Increased wall motion Increased contractility
Bone-marrow-derived stem/progenitor cells
2-250 million (includingEPCs as a small subset)
Acute MI/ refractory angina
Intra-coronary
Angiogenesis Increased perfusion Increased viability Increased wall motion
3-9 days
Outcomes
a Although these cell types are often compared in their ability to repair injured myocardium, these comparisons are not valid. As indicated cell type, cell dose, patient populations, delivery methods and time after injury are different. Thus it is not surprising that outcomes are different. Direct side by side comparisons are needed in similar patients or animal models before outcome can be attributed to cell type. of stem or progenitor cells capable of regenerating new cardiocytes. However, in the past several years several groups have demonstrated the purification of sca-l+c-kit+ cells from adult myocardium and showed that these cells, under specific defined conditions, can differentiate into cardiocytes in vitro [30-32]. The emergence of these cells opens another door in the armamentarium of cells available for cardiac repair. However, how these cells can be obtained, whether they can be grown to sufficient quantities in vitro or whether they can be mobilized in civo for endogenous repair remain completely unanswered questions.
cells Angiogenesis
Myogenesis
Injxn Rescue hibernating heart Grow/recruit new muscle Change Compliance Contraction wall stress improves improves Cytokine/neurohumoral status changes/ endogenous stem cells respond
Fig. 2. Possible effects of cells in injured myocardium. 9. C h a l l e n g e s to c e l l u l a r c a r d i o m y o p l a s t y
Several challenges remain if we are to successfully repair infarcted or failing myocardium with any type of cells. The majority of these challenges seem straightforward, but they are complicated by the extreme heterogeneity of cardiovascular disease. For example, the criteria for reproducible engraftment of large numbers of cells may be very different in the early post-MI patient where angiogenesis is the goal versus the patient with end-stage cardiac dysfunction where myogenesis is needed (Table 1). Similarly, the method of delivering cells (surgical versus endovascular), the concentration of cells delivered and a host of other criteria ranging from age to co-existing disease states have to be considered as cell transplantation emerges into clinical trials. Finally, the most significant challenges to cell repair of injured myocardium evolve from our attempts to do more than simply halt the progression of cardiovascular deterioration. Pre-clinical data suggest that growth factors, fibroblasts or any number of non-myogenic cells that increase perfusion may be able to slow deterioration or actually repair diastolic dysfunction [8,33]. Yet, if we wish to improve contractility of infarcted heart, we must implant cells that can electromechanically integrate into heart, thicken or contract in synchrony, and survive for extended periods of time (Fig. 2). This may very well involve more than simple cell engraftment and may ultimately depend upon combined
approaches where we used cells that can augment vessel growth and those that can regenerate muscle. It may involve combined cell- and tissue-engineering approaches where we create biologic devices in which we can guarantee nutrient delivery and blood supply to an infarcted region of tissue. Or it may involve cell- and gene-based approaches where we generate intelligent cells that can home only to the site of cardiovascular injury and not to other unwanted sites within the patient. Although these are not at the forefront of myocardial repair today, they are areas that may well form the next generation of cell-based products for cardiovascular repair.
10. C o n c l u s i o n
Cell transplantation for myocardial repair is here. Carefully evaluating this new modality provides an opportunity to define a new era in the treatment of cardiovascular disease. Combining genomics, tissue engineering and advanced imaging methods to find the best cell type(s), to understand the mechanism(s) by which cell engraftment improves function, to promote graft longevity, and to improve electromechanical graft integration are the next pre-clinical frontiers. Designing clinical trials that allow us to adequately evaluate the safety and efficacy of cell transplantation in patients with both acute and chronic cardiac disease but in the absence of complicating adjunctive therapies is crucial [34].
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In short, basing clinical trials on good pre-clinical data in multiple animal species, carefully defining the cells we use, rigorously demonstrating safety and efficacy to find the best cell type for a given cardiac injury, and performing randomized controlled trials that can be compared worldwide are the tasks before us as a field. Accepting the challenge, and pushing back this frontier is our opportunity. Changing patient outcomes will be our prize. Yet, doing it improperly could relegate this promising field to failure before its potential is realized.
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