Large Animal Model Efficacy Testing Is Needed Prior to Launch of a Stem Cell Clinical Trial
2017; Lippincott Williams & Wilkins; Volume: 121; Issue: 5 Linguagem: Inglês
10.1161/circresaha.117.311562
ISSN1524-4571
AutoresStephen E. Epstein, Dror Luger, Michael J. Lipinski,
Tópico(s)Viral Infectious Diseases and Gene Expression in Insects
ResumoHomeCirculation ResearchVol. 121, No. 5Large Animal Model Efficacy Testing Is Needed Prior to Launch of a Stem Cell Clinical Trial Free AccessArticle CommentaryPDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissionsDownload Articles + Supplements ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toSupplemental MaterialFree AccessArticle CommentaryPDF/EPUBLarge Animal Model Efficacy Testing Is Needed Prior to Launch of a Stem Cell Clinical TrialAn Evidence-Lacking Conclusion Based on Conjecture Stephen E. Epstein, Dror Luger and Michael J. Lipinski Stephen E. EpsteinStephen E. Epstein From the MedStar Heart and Vascular Institute, MedStar Washington Hospital Center, Washington, DC. , Dror LugerDror Luger From the MedStar Heart and Vascular Institute, MedStar Washington Hospital Center, Washington, DC. and Michael J. LipinskiMichael J. Lipinski From the MedStar Heart and Vascular Institute, MedStar Washington Hospital Center, Washington, DC. Originally published18 Aug 2017https://doi.org/10.1161/CIRCRESAHA.117.311562Circulation Research. 2017;121:496–498Stem cell therapeutics, as other therapeutic strategies, must surmount multiple requirements before reaching the phase of clinical trials. These requirements can be onerous in terms of time and funding and could, thereby, lead to abandonment of many promising therapeutics. One of these requirements is the perceived necessity of conducting large animal efficacy studies—especially pig studies—prior to initiating a clinical trial. The value of such a strategy, despite acceptance by experts, has never been demonstrated scientifically, raising the question of whether bias rather than evidence hampers the development of new therapeutic strategies.Of the questions facing investigators involved in developing new treatment strategies, one is common across disease disciplines and therapeutic approaches: What animal models should be used to determine whether the likelihood a therapeutic will be clinically successful is high enough to justify spending years of effort and tens, or even hundreds, of millions of dollars in clinical trials? A particularly vexing extension of this question is whether it is necessary to test efficacy in a large animal model prior to clinical trial launch.The purpose of this Viewpoint is to challenge the prevailing view, epitomized by the following quote: "Translational studies in large animal models (usually pigs) are rare because they are expensive, complex, time-consuming, technically demanding, slow, and usually not suitable for mechanistic investigations; nevertheless, because they are conducted in settings closer to the human situation than those found in rodent models, these studies are essential to justify the risks and costs of clinical trials."1This quote reflects current unofficial dogma that once efficacy of an intervention is established in mouse or other rodent models, efficacy needs confirmation in a large animal model—the most popular one being the pig.1–3 This is not a trivial conclusion because adequately powered pig studies are expensive from both a time and financial perspective and are beyond the capacity of most laboratories.4 Given these downsides, we need to ascertain how strong the evidence is that purports to validate this strategy.To be clear, we are not challenging the need of a large animal model when considering therapeutics for which a large animal model is patently necessary—for example, when testing an intravascular device such as a stent or a left ventricular assist device. These unquestionably need testing in animals with vessels and hearts comparable in size to those of humans. Acknowledging the complexity of the question and how different issues arise when using different therapeutics for a diverse array of diseases, we have focused on the issues surrounding a single therapeutic targeting a single disease category—the efficacy of stem cells in treating cardiovascular disease. We think, however, that the issues we raise apply to a broad range of therapeutics for use in a broad range of diseases. Also, we are not addressing the issue of animal models to test safety because this is a contentious issue about which many papers have been published.Disparities Between Animal Models of Disease and the Human Disease They are Supposed to ModelThe justifiably skeptical perspective relating to reliability of results obtained in murine models of disease to drive initiation of clinical trials derives, most compellingly, from the frequency with which positive results in such models have been followed by clinical trials failing to confirm efficacy. There are several possible reasons for this.Lack of Disease Complexity in Animal Models UsedOne such reason is that animal models of disease do not reflect the complexity of the human disease they are meant to model. Therapeutic success might therefore be easier to achieve in the animal model. This, however, is common for any animal model of disease and, therefore, does not speak to any potential superiority of large versus small animal models for predicting outcomes in human therapeutic trials.Size, Lifespan, and Heart Rate Differences Between Mouse and Human ModelsAn obvious reason for disparity in outcomes is the possibility that certain disparities, such as size, lifespan, and baseline heart rates (500–600 bpm in mice) make responses of mice different from that of humans. As one size-related example, the number of stem cells injected per gram of tissue in the mouse easily exceeds what is possible to inject in humans or large animals by several orders of magnitude. Thus, greater distribution density throughout the heart and other tissues can be achieved with mice compared with pigs or humans. The prevailing assumption, fueled by these differences between mice versus pig and humans, is that pig data must more reliably mirror the potential for stem cells to improve clinical outcomes.1–3Differences in the Innate and Adaptive Immune Responses Between Mice and HumansDisparate therapeutic outcomes could also relate to differences in innate and adaptive immune responses between mice and humans.5,6 These differences have been partly explained by diverse evolutionary pressures caused by the different ecological niches each species occupied as they diverged somewhere between 65 and 75 million years ago.Immune/inflammatory response differences could lead to the conclusion that the effects of a therapeutic in murine models would, if the therapeutic depended at least partly on its effects on inflammatory responses, not reliably predict human outcomes. However, there are many redundancies of inflammatory pathways, so that differences in immune pathways do not inevitably predict different inflammatory outcomes. Our own perspective is that when considering differences in immune/inflammatory pathways, while it must be acknowledged that issues exist when extrapolating results from mouse studies to the clinical setting, it is likely that mechanistic insights and some reliability for predicting therapeutic outcomes can derive from carefully designed murine models.6Divergences From Human Immune/Inflammatory Responses: Is There Greater Similarity Between Pig and Man Versus Mouse and Man?The different immune/inflammatory pathways existing between mouse and man have fueled the conclusion that positive murine studies are unreliable predictors of human outcomes and, therefore, need confirmation by pig studies before clinical trial initiation.Because the array of reagents available to query immune/inflammatory pathways is much greater for mouse studies than for pig, the number of comparative studies between pig and human immune/inflammatory responses are fewer and less sophisticated than those performed in mice. However, there is reason to expect that major differences between human and pig responses do exist. Most evolutionary pressures leading to genetic and epigenetic alterations that enhance survival derive from developing innate and adaptive immune responses to prevent infection-induced fatality. Thus, just as it is easy to understand how different ecological niches, with their different evolutionary survival pressures, occupied by mice and humans lead to different inflammatory pathways, so too it would not be unexpected that the different ecological niches occupied by pig and human, with exposure to different pathogens, would cause their immune systems to evolve in different ways. In fact, such differences have been documented.7,8Studies Available Demonstrating Data Derived From Large Animal Studies Provide More Reliable Prediction of Outcome of Clinical TrialsNumerous murine studies have demonstrated positive results only to be followed by negative human trials. Although far fewer pig versus human studies have been performed, there are several porcine studies reporting positive results only to be followed by negative human trials.9–11 Thus, positive results of stem cell therapy in a murine or in a porcine model of disease do not necessarily predict success clinically.Conversely, it might seems reasonable to assume that the value of a negative preclinical study would be much greater that the value of a positive study (because of the belief that it is easier to treat a disease in an otherwise normal animal than in a patient). The problem, however, is that such studies do not exist. This is probably because a clinical trial, or even a pig study, is unlikely to be initiated if murine studies are negative (Online Table). We then need to ask—which is the critical issue addressed in this article—What are the data proving that large animal model testing is superior to small animal testing for predicting clinical outcomes and, therefore, are needed prior to the launch of a stem cell therapy trial?The simple answer is there are no data.Major Explanation for the Demonstrated Inability of Preclinical Trials to Predict Outcomes in Clinical Trials of Stem Cell TherapyAdequacy of Design of Preclinical StudiesThere is another, possibly most important, reason for why animal stem cell studies are such poor predictors of human trial outcomes: most preclinical studies are poorly designed and inadequately powered. A particularly egregious issue is that most preclinical studies use too few animals in each study arm, so that false-positive outcomes can often occur by chance alone. This is virtually the rule in pig studies; because of expense, relatively few pigs are included in a study—usually <10/group. Yet farm pigs are genetically heterogenous, just as humans—and much more so than mice within a given species. It is, therefore, virtually certain that such small numbers of animals will not provide reproducible, accurate results. Yet, these are the studies that provide the basis for initiating clinical studies requiring years of effort and tens of millions of dollars. Given that mice are relatively inexpensive, it might be assumed that murine studies are often underpowered. However, underpowering is also the rule in murine studies. This issue of underpowering, in the context of interanimal heterogeneity, was reported many years ago by one of the authors12 and is illustrated more recently in the table contained in the Editorial Comment of Bolli and Ghafghazi.1This problem of poor study design is compounded by the fact that study designs rarely have an embedded requirement for confirming a positive result. The need to confirm positive results is scientifically essential. However, replication to determine reproducibility is rarely performed—usually for practical and logistic reasons.Poor science is, as we and others think,4 the cause of many of the disparities we observe between murine and human study results. This problem was recognized and eloquently articulated in the article dedicated to forming a consortium dealing with the issue of nonreproducible preclinical results of putative infarct-sparing interventions4: "…the translational failures that have plagued the field of cardioprotection for the past 40 years have been caused by the fact that most 'positive' preclinical studies have not been reproducible and have not been conducted with sufficient rigor…"Ironically, reproducibility is perhaps the most valid reason to repeat a positive murine study in the pig. A positive well-designed murine study, followed by a positive well-designed pig study, satisfies the scientific need for replication of positive results. However, it is likely that reliability for predicting human trial outcome would probably be achieved equally as well if the murine model results were replicated in a second experiment using a murine model.Proposed Strategy for Enhancing the Ability of Preclinical Murine Models to Predict Clinical Outcomes of InterventionsIs there a strategy that, when used in murine testing, will more reliably predict human trial outcomes than is now the case? A strategy we recently adopted assumes greater reliability will ensue if a study with positive outcomes (1) is replicated and (2) is combined with parallel studies identifying a mechanism that reasonably explains the positive outcome results.Thus, we demonstrated13 that intravenous administration of mesenchymal stem cells significantly improved left ventricular function in mice with an acute myocardial infarction that had large infarcts; the capacity to improve LV function was replicated in separate experiments in mice with ischemic cardiomyopathy. We further demonstrated that one of the mechanisms responsible for the improved LV function was a mesenchymal stem cell–induced systemic anti-inflammatory effect. We think that replicable efficacy, combined with demonstration of a responsible mechanism, provides a stronger likelihood of predicting clinical success of the intervention than if just efficacy in a single experiment in a mouse model were demonstrated. However, this perspective awaits the results of a pivotal clinical trial.ConclusionThe proper study of man, as has been said, is man—not mice and not pigs. However, studies of these models have supplied critically important information about how humans might respond to a therapeutic. Most importantly, when an intervention improves outcome in a well-designed trial in any animal model, this proof of concept is a critical piece of evidence leading to further studies. Possibly, well-designed large-animal studies will better predict human trial outcomes than data from small animals—however, evidence proving this is lacking. The mouse also has the advantage because of the diverse reagents available to test molecular pathways and because it is relatively inexpensive to study large numbers of animals, of providing a model that can be used both to test large numbers of animals and to derive mechanistic insights relating to the therapeutic being studied.In this Viewpoint article, we address specifically stem cell therapy. However, we think our conclusions relate to a broad range of therapeutic interventions. It is our contention that it is not which animal model is being tested—mouse or pig—that provides the most important information predicting outcome in human stem cell trials. Rather, it is the rigorous design of adequately powered studies and demonstrated reproducibility of positive results in either large or small animal models of disease that will best indicate whether it is reasonable to undertake the large expenditure in funds and time to initiate clinical trials. Even then, however, preclinical studies provide only a probability—not a certainty—of a successful clinical outcome.DisclosuresS.E. Epstein is a consultant to and holds equity interest in CardioCell, LLC. The other authors report no conflicts.FootnotesThe opinions expressed in this article are not necessarily those of the editors or of the American Heart Association.The online-only Data Supplement is available with this article at http://circres.ahajournals.org/lookup/suppl/doi:10.1161/CIRCRESAHA.117.311562.Correspondence to Stephen E. Epstein, MD, MedStar Heart and Vascular Institute, MedStar Washington Hospital Center, 110 Irving St, NW, Washington, DC 20010. E-mail [email protected]References1. Bolli R, Ghafghazi S. Cell therapy needs rigorous translational studies in large animal models.J Am Coll Cardiol. 2015; 66:2000–2004. doi: 10.1016/j.jacc.2015.09.002.CrossrefMedlineGoogle Scholar2. Harding J, Roberts RM, Mirochnitchenko O. 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Intravenously delivered mesenchymal stem cells: systemic anti-inflammatory effects improve left ventricular dysfunction in acute myocardial infarction and ischemic cardiomyopathy.Circ Res. 2017; 120:1598–1613. doi: 10.1161/CIRCRESAHA.117.310599.LinkGoogle Scholar Previous Back to top Next FiguresReferencesRelatedDetailsCited ByRomeo F, Mazurek R, Sakata T, Mavropoulos S and Ishikawa K (2022) Device‐Based Approaches Targeting Cardioprotection in Myocardial Infarction: The Expanding Armamentarium of Innovative Strategies, Journal of the American Heart Association, 11:23, Online publication date: 6-Dec-2022. Coulon S, Schuman J, Du Y, Bahrani Fard M, Ethier C and Stamer W (2022) A novel glaucoma approach: Stem cell regeneration of the trabecular meshwork, Progress in Retinal and Eye Research, 10.1016/j.preteyeres.2022.101063, (101063), Online publication date: 1-Apr-2022. Silvis M, van Hout G, Fiolet A, Dekker M, Bosch L, van Nieuwburg M, Visser J, Jansen M, Timmers L and de Kleijn D (2021) Experimental parameters and infarct size in closed chest pig LAD ischemia reperfusion models; lessons learned, BMC Cardiovascular Disorders, 10.1186/s12872-021-01995-7, 21:1, Online publication date: 1-Dec-2021. Tsikopoulos K, Sidiropoulos K, Kitridis D, Moulder E, Ahmadi M, Drago L and Lavalette D (2021) Preventing Staphylococcus aureus stainless steel‐associated infections in orthopedics. A systematic review and meta‐analysis of animal literature , Journal of Orthopaedic Research, 10.1002/jor.24999, 39:12, (2615-2637), Online publication date: 1-Dec-2021. Mahfouzi S, Safiabadi Tali S and Amoabediny G (2021) Decellularized human-sized pulmonary scaffolds for lung tissue engineering: a comprehensive review, Regenerative Medicine, 10.2217/rme-2020-0152, 16:8, (757-774), Online publication date: 1-Aug-2021. Petnehazy O, Donko T, Ellis R, Csoka A, Czeibert K, Baksa G, Zucker E, Repa K, Takacs A, Repa I and Moizs M (2021) Creating a cross‐sectional, CT and MR atlas of the Pannon minipig, Anatomia, Histologia, Embryologia, 10.1111/ahe.12657, 50:3, (562-571), Online publication date: 1-May-2021. Ruane-O'Hora T and Markos F (2021) Platelets Do Not Alter Flow-Mediated Dilation or Arterial Conduction in vivo, Journal of Vascular Research, 10.1159/000516045, 58:4, (231-236), . Ticha P, Pilawski I, Yuan X, Pan J, Tulu U, Coyac B, Hoffmann W and Helms J (2020) A novel cryo-embedding method for in-depth analysis of craniofacial mini pig bone specimens, Scientific Reports, 10.1038/s41598-020-76336-3, 10:1, Online publication date: 1-Dec-2020. Childs P, Reid S, Salmeron-Sanchez M and Dalby M (2020) Hurdles to uptake of mesenchymal stem cells and their progenitors in therapeutic products, Biochemical Journal, 10.1042/BCJ20190382, 477:17, (3349-3366), Online publication date: 18-Sep-2020. López‐Beas J, Guadix J, Clares B, Soriano‐Ruiz J, Zugaza J and Gálvez‐Martín P (2020) An overview of international regulatory frameworks for mesenchymal stromal cell‐based medicinal products: From laboratory to patient, Medicinal Research Reviews, 10.1002/med.21659, 40:4, (1315-1334), Online publication date: 1-Jul-2020. Khalil A, Jaenisch R and Mooney D (2020) Engineered tissues and strategies to overcome challenges in drug development, Advanced Drug Delivery Reviews, 10.1016/j.addr.2020.09.012, 158, (116-139), . August 18, 2017Vol 121, Issue 5 Advertisement Article InformationMetrics © 2017 American Heart Association, Inc.https://doi.org/10.1161/CIRCRESAHA.117.311562PMID: 28819039 Originally publishedAugust 18, 2017 Keywordsstem cellsclinical trialsanimal modelsresearchswinePDF download Advertisement SubjectsAnimal Models of Human DiseaseCell TherapyClinical StudiesInflammationMechanisms
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