Efficient and robust generation of functional hematopoietic cells from human pluripotent stem cells in albumin-free conditions.

Generation of human hematopoietic stem and progenitor cells (HSPCs) from pluripotent stem cells (PSCs) holds significant promise in disease modeling, drug screening, and the development of cell and gene therapies for various hematologic and nonhematologic disorders. However, efficient and consistent derivation of functional HSPCs remains a major challenge hindering their practical use in biomedical research and applications. One of the contributing factors is the use of human serum albumin (HSA), which has been widely regarded as essential for supporting ex vivo maintenance, expansion, and differentiation of human stem cells and their derivatives. The lot-to-lot variation of this critical reagent can contribute to inconsistent laboratory results and high manufacturing costs. In this study, we address the manufacturing and regulatory challenges associated with using HSA in HSPC differentiation and propose an approach using a caprolactam-based polymer as an alternative. Our differentiation method demonstrates robust HSPC generation across various embryonic and induced PSC lines and culture conditions, highlighting its adaptability and reliability. Importantly, PSC-derived HSPCs exhibit functional versatility in differentiation capacity into myeloid and lymphoid lineages, as validated through colony-forming assays as well as directed red blood cell and NK cell differentiations. These findings suggest that HSA is dispensable in HSPC differentiation and replacing it with synthetic polymers has the potential to mitigate lot-to-lot reagent variation, improve HSPC production consistency, lower manufacturing costs, and expedite clinical applications.