T helper differentiation of CAR T cells augments function and target cell killing

Abstract

Abstract Chimeric antigen receptor (CAR) T cell therapy involves genetically manipulating T cells to express an antigen receptor specific for tumor protein, resulting in a powerful anti-cancer therapeutic. However, trafficking to tumor sites and the suppressive nature of the tumor microenvironment have prevented effective treatment of solid tumors. Previous work has shown that antigen specific T helper (Th) subsets have variable tumor killing capabilities, however, it is not completely defined how different transcriptional differentiation programs effect the fitness and function of CAR T cells. It was hypothesized that differentiation of T cells transduced with CAR improves cancer cell killing in an antigen specific manner. To test this hypothesis, splenic CD4+ and CD8+ T cells were cultured under T cell activating (T0), Th2 (T2), or Th9 (T9) cell differentiation conditions and transduced with a 2nd generation CD19-specific CAR. Control cells were transduced with CD19-specific CAR lacking the intracellular domain. CAR T cells were co-cultured with CD19+ B cells and target cell killing was measured by flow cytometry. Preliminary results show that CAR T9 cells, but not T2 cells, kill target cells similar to the clinic relevant T0 cells. FACS analysis of T cells after co-culture with target cells revealed antigen-experience promoted production of lineage specific cytokines as well as granzymes A and B. T9 conditions maintained a larger proportion of CD8+ T cells than T2 conditions, suggesting that T9 conditions promote expansion of effector CD8+ T cells. These results suggest that the Th9 transcriptional program promotes target cell killing, transduction efficiency, and generates a better ratio of CD4:CD8 T cells for adoptive cell therapy.