Enhancing Anti-Tumor Activity of CAR T Cells Through FOXO1-Mediated Stemness and Mitochondrial Fitness

Loss of stemness and mitochondrial dysfunction induced by chronic antigen stimulation represents formidable obstacles in chimeric antigen receptor (CAR) T cell therapy for solid tumors. However, the regulation of stemness and mitochondrial fitness in CAR T cells remains unclear. Here we found that short-term resting conditions with IL-7, CXCL12, and the pan-tyrosine kinase inhibitor, Dasatinib, could reprogram CAR T cells, which exhibit tonic signal-dependent exhaustion hallmarks, into stem-like CAR T cells through epigenetic remodeling and enhance mitochondrial metabolism, thereby resulting in augmented antitumor activity against solid tumors. We identified forkhead box protein O1 (FOXO1) as a critical regulator of the epigenetic and metabolic reprogramming as well as antitumor activity. Moreover, it was found that FOXO1WT overexpression led to enhanced mitochondrial metabolism, efficient CAR T cell expansion, and potent antitumor effects through epigenetic modifications. We demonstrated the central role of FOXO1 in both stemness and mitochondrial metabolism in CAR T cells. These findings suggest that regulating FOXO1 is a promising strategy for engineering CAR T cells to treat solid tumors.