Scalable bioprocess for cardiomyocyte generation from human embryonic stem cells

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DESCRIPTION (provided by applicant): Myocardial infarction is a major cause of morbidity and mortality in the United States and most developed countries. Heart transplantation is an effective therapeutic modality in reconstituting the function of damaged heart. However, widespread application of this modality is severely limited due to the scarcity of organ donors and complications associated with the required immunosuppression. Cell therapies aiming at replacing infarcted heart muscle are highly desirable. Embryonic stem cells (ESCs) can serve as an inexhaustible source of cellular material for repairing damaged myocardium. Human ESCs (hESCs) have been shown to differentiate towards functional cardiomyocytes. Nonetheless, clinical realization of stem cell-based therapies for heart repair will require the production of ESC-derived cardiomyocytes in large numbers. Current methodologies entail the propagation and differentiation of ESCs in static cultures (e.g. dishes) which are challenging to scale-up. To that end, large cell quantities can be generated under tightly controlled culture conditions in scalable stirred-suspension bioreactors. We have discovered that mouse ESCs cultivated in a bioreactor can be expanded several fold and differentiate to multiple lineages. We hypothesize that hESCs cultured as aggregates in stirred-suspension vessels can also propagate to high concentrations. We propose to culture hESCs in a stirred bioreactor culture system and determine conditions which favor the growth of hESCs without compromising their viability. Furthermore, hESC differentiation to cardiomyocytes is carried out mainly while hESCs are organized as aggregates or embryoid bodies. Therefore, expansion of hESCs as aggregates in the bioreactor may be followed by switching to conditions directing the cells to adopt a cardiomyocyte fate. We will evaluate the cardiogenic potential of hESCs cultured as aggregates in the bioreactor. The resulting cells will be characterized for the expression of cardiomyocyte-associated genes/proteins and will be subjected to functional assays in vitro. This study will yield new information benefiting the development of bioprocesses for the generation of large quantities of hESC-derived cardiomyocytes suitable for infracted heart therapies. PUBLIC HEALTH RELEVANCE Myocardial infarction-induced heart failure is a prevailing cause of death in the United States and clinical therapies aiming at replacing or restoring damaged heart muscle are lacking. Stem cells with their extensive proliferative capacity and their ability to differentiate towards functional cardiomyocytes may serve as a renewable cellular source for regenerative heart therapies. This project seeks to further our understanding of the effects of bioreactor culture on the cardiogenic potential of human embryonic stem cells and to advance the bioprocess technology for the production of stem cell-derived cardiomyocytes in clinically relevant quantities.