Development of a scalable suspension culture for cardiac differentiation from human pluripotent stem cells

• Published in: Stem cell research Vol. 15; no. 2; pp. 365 - 375
• Main Authors: Chen, Vincent C., Ye, Jingjing, Shukla, Praveen, Hua, Giau, Chen, Danlin, Lin, Ziguang, Liu, Jian-chang, Chai, Jing, Gold, Joseph, Wu, Joseph, Hsu, David, Couture, Larry A.
• Format: Journal Article
• Language: English
• Published: England Elsevier B.V 01-09-2015; Elsevier
• Subjects: Actinin - metabolism; Cardiomyocyte differentiation; Cell Culture Techniques; Cell Differentiation; Cell Line; Gene Expression Regulation; GMP; Human pluripotent stem cells; Humans; Microscopy, Fluorescence; Myocytes, Cardiac - cytology; Pluripotent Stem Cells - cytology; Pluripotent Stem Cells - metabolism; RNA - chemistry; RNA - isolation & purification; Sequence Analysis, RNA; Suspension cell cultures; Troponin I - metabolism; Troponin T - metabolism; Wnt Signaling Pathway; Suspension cell cultures; GMP; Human pluripotent stem cells; Cardiomyocyte differentiation
• ISSN: 1873-5061; 1876-7753
• DOI: 10.1016/j.scr.2015.08.002

To meet the need of a large quantity of hPSC-derived cardiomyocytes (CM) for pre-clinical and clinical studies, a robust and scalable differentiation system for CM production is essential. With a human pluripotent stem cells (hPSC) aggregate suspension culture system we established previously, we developed a matrix-free, scalable, and GMP-compliant process for directing hPSC differentiation to CM in suspension culture by modulating Wnt pathways with small molecules. By optimizing critical process parameters including: cell aggregate size, small molecule concentrations, induction timing, and agitation rate, we were able to consistently differentiate hPSCs to >90% CM purity with an average yield of 1.5 to 2×109 CM/L at scales up to 1L spinner flasks. CM generated from the suspension culture displayed typical genetic, morphological, and electrophysiological cardiac cell characteristics. This suspension culture system allows seamless transition from hPSC expansion to CM differentiation in a continuous suspension culture. It not only provides a cost and labor effective scalable process for large scale CM production, but also provides a bioreactor prototype for automation of cell manufacturing, which will accelerate the advance of hPSC research towards therapeutic applications.
•We present a strategy to optimize cardiac differentiation in suspension for hiPSCs.•The matrix-free suspension platform integrates hPSC expansion and differentiation.•Cardiac production in suspension achieves >90% purity with 1L spinner flasks.•The production process in suspension is defined, scalable, and GMP compliant.