Inhibiting TGFβ Signaling in CAR T-Cells May Significantly Enhance Efficacy of Tumor Immunotherapy (2017)

Abstract The use of genetically engineered T cells expressing chimeric antigen receptors (CAR) against CD19 or BCMA to treat hematologic malignancies has demonstrated promising results in clinical trials. However, it is possible that the efficacy of CAR T cells could be limited in some clinical indications due to suppressive factors in the tumor microenvironment. Immunosuppressive cytokines (e.g., IL-6, IL-10 and TGFβ) provide a favorable environment for tumor growth and inhibit the tumoricidal activity of the endogenous T cells. In particular, transforming growth factor β (TGFβ) has been shown to suppress T-cell proliferation, inhibit the maturation of T helper cells and reduce their effector functions. Elevated levels of TGFβ have been detected in many human cancers, including multiple myeloma (MM). Here, we sought to modulate the activity of CAR T cell-based immunotherapy for MM by inhibiting TGFβ signaling in BCMA-specific CAR T cells (Budha et al., Cancer Cell, 2012). We evaluated two strategies to modulate the TGFβ pathway: (1) knocking out TGFβ Receptor II (TGFβRII) using CRISPR gene editing and (2) overexpressing a dominant negative TGFβRII in BCMA targeted CAR T cells. TGFβRII modulation inhibited TGFβ-dependent Smad2/3 phosphorylation. TGFβRII-modulated BCMA CAR T cells were insensitive to TGFβ-mediated inhibition of proliferation, cell cytotoxicity and cytokine production, both IFNg and granzyme B, in the context of primary and/or repeat antigen stimulation with BCMA+ve MM cell lines in vitro . We further evaluated the gene expression signatures of BCMA CAR T cells isolated from tumor and spleen in a NSG mouse xenograft model subcutaneously inoculated with myeloma cell line (RPMI8226). TGFβ pathway gene expression changes indicating TGFβ exposure were observed in BCMA CAR T cells isolated from tumor, but not spleen, suggesting that BCMA CAR T cells are encountering a TGFβ-enriched TME. However, both TGFβ pathway modulating strategies prevented the development of the TGFβ-induced gene expression phenotype. Together, these data suggest that inhibition of TGFβRII signaling may potentiate CAR T activity by directly inhibiting immunosuppressive effects of TGFβ in the tumor microenvironment. By specifically targeting TGFβ signaling in CAR T cells, it may be possible to avoid side effects associated with systemic administration of TGFβ antagonists. Gene editing to remove TGFβRII expression or overexpressing dominant negative TGFβ RII could modulate the activity of BCMA CAR T therapy in MM. Disclosures Vong: Juno Therapeutics: Employment, Equity Ownership. Nye: Juno Therapeutics: Employment, Equity Ownership. Hause: Juno Therapeutics: Employment, Equity Ownership. Clouser: Juno Therapeutics: Employment, Equity Ownership. Jones: Juno Therapeutics: Employment, Equity Ownership. Burleigh: Juno Therapeutics: Employment, Equity Ownership. Borges: Editas Medicine: Employment, Equity Ownership. Chin: Editas Medicine: Employment, Equity Ownership. Marco: Editas Medicine: Employment, Equity Ownership. Barrera: Editas Medicine: Employment, Equity Ownership. Da Silva: Editas Medicine: Employment, Equity Ownership. Harbinski: Editas Medicine: Employment, Equity Ownership. Giannoukos: Editas Medicine: Employment, Equity Ownership. Dhanapal: Editas Medicine: Employment, Equity Ownership. Jiang: Juno Therapeutics: Employment, Equity Ownership. Salmon: Juno Therapeutics: Employment, Equity Ownership. Wilson: Editas Medicine: Employment, Equity Ownership. Myer: Editas Medicine: Employment, Equity Ownership. Welstead: Editas Medicine: Employment, Equity Ownership. Bond: Juno Therapeutics: Employment, Equity Ownership. Sather: Juno Therapeutics: Employment, Equity Ownership.