Analysis of the transcriptomic profiles for HER2 CD8+ CAR T cells-overexpressed FOXO1WT
T cell exhaustion and metabolic dysfunction induced by chronic antigen stimulation within the tumor microenvironment represent formidable obstacles in CAR T cell therapy for solid tumors. Previously, we developed a method to convert effector/exhausted CAR T cells that emerge from repeated antigen stimulation into stem cell memory-like CAR T (termed CAR-iTSCM) cells. These CAR-iTSCM cells exhibit expression of early memory markers and enhanced mitochondrial metabolism after quiescence under culture conditions containing IL-7, CXCL12, and the NOTCH-ligand. However, we noticed that this strategy was effective for CD19 CAR used for blood cancers, but not for HER2 CAR commonly used to treat solid tumors, due to tonic CAR signals. In this study, we developed a new culture condition for CAR iTSCM cells suitable for HER2 CAR and elucidated the mechanism of induction of stemness and mitochondrial fitness. We found that short-term resting condition with IL-7, CXCL12, and pan-tyrosine kinase inhibitor, Dasatinib can induce stem-like CAR T cells at the epigenetic level and enhance mitochondrial metabolism. We identified FOXO1 as a critical regulator in the process of epigenetic and metabolic reprogramming, and anti-tumor activity. We found that overexpression of FOXO1 leads to enhanced mitochondrial metabolism, efficient CAR T cell expansion, and potent anti-tumor effects through epigenetic modifications. These findings suggest that regulating the transcriptional activity of FOXO1 is a promising strategy for engineering effective CAR T cells for treating solid tumors. In this analysis, we aimed to investigate the effects of FOXO1WT overexpression on the transcriptomic profiles in 4-1BB-based HER2 CD8+ CAR T cells which exhibit tonic signals-induced exhaustion hallmarks. Overall design: We performed bulk RNA-seq analysis using HER2 CD8+ CAR T cells under four different conditions (Empty, FOXO1WT, 7.12.I, and 7.12.I.Das). Each CAR T cells were generated from different three healthy donors (n=3).