Abstract 3239: SMIC CAR T cells: CAR T with temporally-controlled, programmable cytokine signaling outputs

CAR T cell therapy has attained unprecedented success in the treatment of hematological malignancies. However, clinical benefit in solid tumor indications has been limited in part due to suppressive solid tumor microenvironments (TME) that inhibit T cell effector function and persistence. While the provision of cytokine support can help CAR T cells overcome suppressive TMEs, conventional approaches, such as combining CAR T therapy with systemically-administered cytokines/cytokine mimetics, or engineering CAR T cells to constitutively secrete cytokines can result in toxicities and adverse events. To mitigate these safety concerns, we designed Small Molecule-Inducible Cytotails (SMIC) that can be specifically activated by the clinically validated dimerizer drug, AP1903. SMIC is a chimeric fusion receptor comprising the FKBP(F36V) ectodomain, the transmembrane and JAK-activating domains of TpoR and one or more intracellular STAT-binding domains derived from the cytokine receptor (Cytotail) of interest. In response to AP1903, different SMIC designs resulted in downstream STAT activation that mimicked signaling of the parental cytokine receptor. Furthermore, two Cytotails could be fused in tandem to generate dual signaling outputs from a single receptor. Using 2nd generation CARs directed towards solid tumor targets, we demonstrated that SMIC CAR T cells bearing different Cytotail fusions could be programmed towards divergent phenotypes, such as those promoting expansion (IL7Rα Cytotail), memory maintenance (EGFR Cytotail) or effector function (IL12Rβ Cytotail). RNASeq analysis of SMIC CAR T cells bearing an IL7Rα.IL12Rβ tandem Cytotail (SMIC7.12) revealed near-identical gene expression profiles to control CAR T cells treated with exogenously supplemented IL-7 and IL-12, demonstrating that SMIC closely mimicked the signaling of native cytokine receptors. In the absence of target cells, AP1903 treatment alone was sufficient to expand SMIC7.12 CAR T cells more efficiently than exogenously supplemented IL-7 and IL-12, while preserving a juvenile memory phenotype. Compared to control CAR T cells, AP1903-activated SMIC7.12 enhanced CAR T cell cytotoxicity in vitro, and promoted CAR T cell activity in a subcutaneous tumor model. In conclusion, SMIC represents a novel chimeric cytokine receptor that can improve the potency and persistence of CAR T cells, while minimizing safety risks associated with cytokine co-therapy or constitutive cytokine secretion. Furthermore, fusion of different/combinatorial Cytotails to SMIC offers the flexibility for user-programmable signaling outputs, permitting control over CAR T cell phenotype and function. Citation Format: Regina Lin, Andrew Nager, Spencer Park, Janette Sutton, Zea Melton, Yi Zhang, Bijan Boldajipour, Tom Van Blarcom, Siler Panowski, Javier Chaparro-Riggers, Barbra Sasu. SMIC CAR T cells: CAR T with temporally-controlled, programmable cytokine signaling outputs [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3239.