Immunogenetic Metabolomics Reveals Key Enzymes That Modulate CAR T-cell Metabolism and Function

Immune evasion is a critical step of cancer progression that remains a major obstacle for current T cell-based immunothera-pies. Hence, we investigated whether it is possible to genetically reprogram T cells to exploit a common tumor-intrinsic evasion mechanism whereby cancer cells suppress T-cell function by gen-erating a metabolically unfavorable tumor microenvironment (TME). In an in silico screen, we identified ADA and PDK1 as metabolic regulators. We then showed that overexpression (OE) of these genes enhanced the cytolysis of CD19-specific chimeric antigen receptor (CAR) T cells against cognate leukemia cells, and conversely, ADA or PDK1 deficiency dampened this effect. ADA-OE in CAR T cells improved cancer cytolysis under high concentrations of adenosine, the ADA substrate, and an immuno-suppressive metabolite in the TME. High-throughput transcrip-tomics and metabolomics analysis of these CAR T cells revealed alterations of global gene expression and metabolic signatures in both ADA-and PDK1-engineered CAR T cells. Functional and immunologic analyses demonstrated that ADA-OE increased pro-liferation and decreased exhaustion in CD19-specific and HER2-specific CAR T cells. ADA-OE improved tumor infiltration and clearance by HER2-specific CAR T cells in an in vivo colorectal cancer model. Collectively, these data unveil systematic knowledge of metabolic reprogramming directly in CAR T cells and reveal potential targets for improving CAR T-cell therapy.

Automated summary

Immune evasion is a major obstacle in T cell-based immunotherapies for cancer. This study investigated the possibility of genetically reprogramming T cells to overcome tumor-induced suppression of T-cell function through metabolic reprogramming. The overexpression of ADA and PDK1 enhanced the cytolytic activity of CAR T cells against leukemia cells, while deficiency of ADA or PDK1 dampened this effect. ADA overexpression improved cancer cytolysis under high concentrations of adenosine in the tumor microenvironment. These findings provide insight into metabolic reprogramming in CAR T cells and reveal potential targets for improving CAR T-cell therapy.