Induced Pluripotent Stem Cells and Erythrocyte Production

Abstract SCI-38

Induced pluripotent stem cells (iPSCs) are somatic cells that have been turned into embryonic-like stem cells by forced expression of factors critical for establishing pluripotency. Because iPSCs can be differentiated into any type of cell in the human body, including hematopoietic cells, they are seen as a logical alternative source of red blood cells (RBCs) for transfusion. In addition, the unlimited expansion potential of iPSCs makes it easy to adopt iPSC technology for RBC biomanufacturing. iPSCs can be generated from any type of donor, including O/Rh-negative universal donors and donors with very rare blood phenotypes, which makes it possible to generate blood products to accommodate virtually all patient groups. We have developed an approach for generating large quantities of RBCs from iPSCs by inducing them to differentiate into CD34⁺CD43⁺ hematopoietic progenitors in coculture with OP9 stromal cells, followed by selective expansion of erythroid cells in serum-free media with erythropoiesis-supporting cytokines. Erythroid cultures produced by this approach consist of leukocyte-free populations of CD235a⁺ RBCs with robust expansion potential and long (up to 90 days) life spans. In these cultures, up to 1.8×10⁵ RBCs can be generated from a single iPSC. Similar to embryonic stem cells, iPSC-derived RBCs express predominantly embryonic and fetal hemoglobin, with very little adult hemoglobin. It is already feasible to adopt iPSC technologies for producing cGMP-grade RBCs using defined animal-product-free differentiation conditions. However, the induction of the complete switch from embryonic to fetal and adult hemoglobin, as well as the terminal maturation and enucleation of iPSC-derived erythroid cells, remains a significant challenge. We recently identified at least three distinct waves of hematopoietic progenitors with erythroid potential in iPSC differentiation cultures. The characterization of erythroid cells produced from these waves of hematopoiesis may help to define populations with definitive erythroid potential and facilitate the production of erythrocytes from iPSCs. Additional critical steps toward translating iPSC-based RBC technologies to the clinic include the development of bioreactor-based-technology for further scaling-up of cell production, and evaluation of the therapeutic potential and safety of human pluripotent stem cell-derived blood cells in animal models. Overall, the manufacturing of RBCs provides several advantages. It can improve the continuity of the blood supply, minimize/eliminate the risk of infection transmission, reduce the incidence of hemolytic and nonhemolytic transfusion reactions, and provide an opportunity to generate RBCs that fit specific clinical needs by using genetically engineered iPSCs or iPSCs with rare blood groups.