(104d) Alginate Encapsulation for Large Scale Human Pluripotent Stem Cell Production

Human pluripotent stem cells (hPSCs) have the capability of differentiating into any somatic cell, providing a means to revolutionize healthcare by providing new routes for cell therapy based regenerative medicine. Due to such commercial and clinical applicability, hPSCs require more efficient bioprocessing. A primary limitation in hPSC propagation is its inability to survive as single cells; dissociation of cell-cell contact triggers the apoptotic pathway in hPSCs. Currently, prominent approaches utilize stirred suspension culture that maintains cell-cell contact and improves scalability. The mechanism of stirred-suspension culture, however, results in uncontrolled cell aggregation and large variation in aggregate size. In addition, bioreactor culture medium exposes hPSC aggregates to hydrodynamic stresses which might be detrimental to hPSC viability and phenotype.

In order to overcome these limitations, our group has developed a promising alternative manufacturing method by encapsulating hPSCs in alginate hydrogel modified with a peptide that mimics the cell-cell signaling and inhibits the apoptosis pathway of single cell hPSCs. This method has proven successful in maintaining fold expansion and high viability of hPSCs [1]. The hPSCs cultured in this substrate retained their pluripotency with high, sustained levels of OCT4 and NANOG gene and protein expression. In addition, hPSCs could be successfully differentiated spontaneously to the three germ layers and by induction into the definitive endoderm stage. Therefore, we hypothesize encapsulation in peptide-modified alginate would be a more efficient method for hPSC manufacturing over suspension culture. Currently, we are evaluating the effect of (i) configurations of encapsulated cells and (ii) alginate properties on the scalability of the designed system. We are encapsulating single cell as well as controlled micro-aggregates and evaluating their expansion potential. In addition, we are evaluating the alginate cross-linking concentration most supportive of hPSC growth and viability. In parallel, we are integrating our system with benchtop bioreactor and evaluating performance in comparison with stirred suspension culture. We expect our encapsulation system to support hPSC growth and expansion, in addition to protecting the cells from over-aggregation and bioreactor hydrodynamics.

References:

[1] Richardson, T.; Mathew, S.; Candielloe, J.; Goh, S.; Kumta, P.; Banerjee, I. Development of an Alginate Array Platform to Decouple the Effect of Multiparametric Perturbations on Human Pluripotent Stem Cells During Pancreatic Differentiation. Biotechnology Journal. 2018, 13, 1700099