Recent studies have shown that stem cell memory T (T_SCM) cell-like properties are important for successful adoptive immunotherapy by the chimeric antigen receptor–engineered-T (CAR-T) cells. We previously reported that both human and murine-activated T cells are converted into stem cell memory-like T (iT_SCM) cells by coculture with stromal OP9 cells expressing the NOTCH ligand. However, the mechanism of NOTCH-mediated iT_SCM reprogramming remains to be elucidated. Here, we report that the NOTCH/OP9 system efficiently converted conventional human CAR-T cells into T_SCM-like CAR-T, “CAR-iT_SCM” cells, and that mitochondrial metabolic reprogramming played a key role in this conversion. NOTCH signaling promoted mitochondrial biogenesis and fatty acid synthesis during iT_SCM formation, which are essential for the properties of iT_SCM cells. Forkhead box M1 (FOXM1) was identified as a downstream target of NOTCH, which was responsible for these metabolic changes and the subsequent iT_SCM differentiation. Like NOTCH-induced CAR-iT_SCM cells, FOXM1-induced CAR-iT_SCM cells possessed superior antitumor potential compared with conventional CAR-T cells. We propose that NOTCH- or FOXM1-driven CAR-iT_SCM formation is an effective strategy for improving cancer immunotherapy.

Manipulation of signaling and metabolic pathways important for directing production of stem cell memory–like T cells may enable development of improved CAR-T cells.