Non‐xenogeneic expansion and definitive endoderm differentiation of human pluripotent stem cells in an automated bioreactor. Issue 2 (16th December 2020)

• Record Type: Journal Article
• Authors: Jacobson, Elena F.; Chen, Zijing; Stoukides, Demetrios M.; Nair, Gopika G.; Hebrok, Matthias; Tzanakakis, Emmanuel S.
• Is Part Of: Biotechnology and bioengineering. Volume 118:Issue 2(2021)
• Page Start: 979
• Page End: 991
• Publication Date: 2020-12-16
• Subjects: definitive endoderm (DE) differentiation -- human pluripotent stem cells (hPSCs) -- pluripotent expansion -- stirred suspension bioreactor (SSB) -- xeno‐free; Biotechnology -- Periodicals; Bioengineering -- Periodicals; 660.6
• DOI: 10.1002/bit.27629
• Languages: English
• ISSNs: 0006-3592

Abstract

Abstract: Scalable processes are requisite for the robust biomanufacturing of human pluripotent stem cell (hPSC)‐derived therapeutics. Toward this end, we demonstrate the xeno‐free expansion and directed differentiation of human embryonic and induced pluripotent stem cells to definitive endoderm (DE) in a controlled stirred suspension bioreactor (SSB). Based on previous work on converting hPSCs to insulin‐producing progeny, differentiation of two hPSC lines was optimized in planar cultures yielding up to 87% FOXA2 + /SOX17 + cells. Next, hPSCs were propagated in an SSB with controlled pH and dissolved oxygen. Cultures displayed a 10‐ to 12‐fold increase in cell number over 5–6 days with the maintenance of pluripotency (>85% OCT4 + ) and viability (>85%). For differentiation, SSB cultures yielded up to 89% FOXA2 + /SOX17 + cells or ~ 8 DE cells per seeded hPSC. Specification to DE cell fate was consistently more efficient in the bioreactor compared to planar cultures. Hence, a tunable strategy is established that is suitable for the xeno‐free manufacturing of DE cells from different hPSC lines in scalable SSBs. This study advances bioprocess development for producing a wide gamut of human DE cell‐derived therapeutics. Abstract : A novel scalable method was developed for the xeno‐free expansion of human pluripotent stem cells (hPSCs) and their guided specification to definitive endoderm (DE) cells in automated stirred‐suspension bioreactors with pH and dissolved oxygen regulation. The generation is demonstrated of 1.7–0.8 million DE cells/ml of culture or approximately 8 DE cells per hPSC seeded. The findings provide a foundation for advancing the biomanufacturing of stem cell‐derived therapeutics.