A serum- and feeder-free system to generate CD4 and regulatory T cells from human iPSCs
Helen Fong, Matthew Mendel, John Jascur, Laeya A. Najmi, Ken Kim, Garrett Lew, Swetha Garimalla, Suruchi Schock, Jing Hu, Andres Gordillo Villegas, Anthony Conway, Jason D. Fontenot, Simona Zompì
Abstract
iPSCs can serve as a renewable source of a consistent edited cell product, overcoming limitations of primary cells. While feeder-free generation of clinical grade iPSC-derived CD8 T cells has been achieved, differentiation of iPSC-derived CD4sp and regulatory T cells requires mouse stromal cells in an artificial thymic organoid. Here we report a serum- and feeder-free differentiation process suitable for large-scale production. Using an optimized concentration of PMA/Ionomycin, we generated iPSC-CD4sp T cells at high efficiency and converted them to Tregs using TGFβ and ATRA. Using genetic engineering, we demonstrated high, non-viral, targeted integration of an HLA-A2 CAR in iPSCs. iPSC-Tregs ± HLA-A2-targeted CAR phenotypically, transcriptionally and functionally resemble primary Tregs and suppress T-cell proliferation in vitro. Our work is the first to demonstrate an iPSC-based platform amenable to manufacturing CD4 T cells to complement iPSC-CD8 oncology products and functional iPSC-Tregs to deliver Treg cell therapies at scale.