Strength of CAR signaling determines T cell versus ILC differentiation from pluripotent stem cells
Suwen Li, Chloe S. Wang, Amélie Montel‐Hagen, Ho-Chung Chen, Shawn Lopez, Olivia Zhou, Kristy Dai, Steven Tsai, William Satyadi, Carlos A. Botero, Claudia Wong, David Casero, Gay M. Crooks, Christopher S. Seet
Abstract
Generation of chimeric antigen receptor (CAR) T cells from pluripotent stem cells (PSCs) will enable advances in cancer immunotherapy. Understanding how CARs affect T cell differentiation from PSCs is important for this effort. The recently described artificial thymic organoid (ATO) system supports in vitro differentiation of PSCs to T cells. Unexpectedly, PSCs transduced with a CD19-targeted CAR resulted in diversion of T cell differentiation to the innate lymphoid cell 2 (ILC2) lineage in ATOs. T cells and ILC2s are closely related lymphoid lineages with shared developmental and transcriptional programs. Mechanistically, we show that antigen-independent CAR signaling during lymphoid development enriched for ILC2-primed precursors at the expense of T cell precursors. We applied this understanding to modulate CAR signaling strength through expression level, structure, and presentation of cognate antigen to demonstrate that the T cell-versus-ILC lineage decision can be rationally controlled in either direction, providing a framework for achieving CAR-T cell development from PSCs.