IL-15-mediated reduction of mTORC1 activity preserves the stem cell memory phenotype of CAR-T cells and confers superior antitumor activity

Abstract

Efforts to improve the quality and fitness of chimeric antigen receptor (CAR)-engineered T cells, through CAR design or manufacturing optimizations, are critical to further enhance the potency of this promising therapy. A critical parameter influencing the effectiveness of CAR-T cell therapy is the T cell differentiation status of the final product, with less-differentiated, less-exhausted CAR-T cells being more therapeutically effective. In the current study, we demonstrate that CAR-T cells expanded in IL-15 alone (CAR-T/IL-15), even after extended ex vivo expansion, preserve a less-differentiated stem cell memory (Tscm) phenotype, defined as CD62L+ CD45RA+ CCR7+ as compared to cells cultured in IL-2 (CAR-T/IL-2). CAR-T/IL-15 cells exhibited reduced expression of exhaustion markers, higher anti-apoptotic properties, and increased proliferative capacity upon antigen challenge. Furthermore, CAR-T/IL-15 cells exhibited decreased mTORC1 activity, global reduction in expression of glycolytic enzymes and improved mitochondrial fitness. In fact, CAR-T/IL-2 cells cultured in rapamycin (mTORC1 inhibitor) shared similar phenotypic features as CAR-T/IL-15 suggesting that IL-15-mediated reduction of mTORC1 activity is responsible for preserving the Tscm phenotype. Importantly, CAR-T/IL-15 promoted superior antitumor responses in vivo in comparison to CAR-T/IL-2. Interestingly, inclusion of additional cytokines with IL-15, either IL-7 and/or IL-21, reduced the beneficial effects of IL-15 for CAR-T phenotype and antitumor potency. Taken together, our findings show that IL-15 preserves the Tscm phenotype by improving their metabolic fitness, and therefore has great potential for the future application of adoptive T cell therapy.