A Unique Strategy To Arm Ipsc-Derived Car-T Cells To Perform Antibody-Directed Cellular Cytotoxicity To Facilitate Multi-Antigen Targeting.

Abstract Although chimeric antigen receptor (CAR)-T cells have been transformational for several liquid tumors, their effectiveness in treating a variety of solid tumors has been limited in part because the heterogeneity of tumor-associated antigens found in many bulky tumors represents a major obstacle to the development and the delivery of effective CAR-T cell therapies in cancer. Here we demonstrate that iPSC-derived CAR-T (CAR-iT) cells are an ideal off-the-shelf approach that can be engineered to perform antibody-dependent cell-mediated cytotoxicity (ADCC) to facilitate multi-antigen targeting with the introduction of various monoclonal antibodies (mAbs). Under this strategy, while the CAR is equipped to predominantly target antigen 1, targeting additional tumor associated antigens through the use of various mAbs can be leveraged through ADCC. To this end, CAR-iT cells were genetically edited at the iPSC stage to uniformly express a novel high-affinity variant (158V), non-cleavable CD16A (hnCD16) Fc receptor to recognize and bind to the Fc fragment of mAbs and facilitate ADCC. When combined with therapeutic mAbs targeting HER2, EGFR, or PDL1, the cytolytic efficacy of hnCD16+ CAR-iT cells was significantly enhanced compared to parental CAR-iT cells in several tumor models, including ovarian, HER2+ breast, and triple-negative breast cancers, underscoring the flexibility of a combined CAR and ADCC – mediated targeting approach. Importantly, although both CD16A and CAR signaling share common CD3ζ pathways, simultaneous activation did not appreciably attenuate the activation and effector function of CAR-iT cells. Moreover, hnCD16+ CAR-iT cells demonstrated enhanced control in an established triple negative breast cancer solid tumor xenograft model. The control of antigenically heterogenous tumors by hnCD16+ CAR-iT cells was also significantly improved when compared to parental CAR-iT cells. To confirm selectivity and avoidance of immunoglobulin interference with functional output, various in vitro cellular cytotoxicity assays were conducted with IgG1 to confirm that hnCD16 requires crosslinking to elicit ADCC while maintaining full potentiation of CAR function. Additional in vitro and in vivo studies are ongoing and will be presented. Collectively, the data demonstrates that CAR-iT cells can be further modified with hnCD16 Fc receptor to elicit ADCC and the combination of CAR and hnCD16 is a unique approach to tackling tumor heterogeneity often found in bulky tumors. Citation Format: Martin Hosking, Soheila Shirinbak, Joy Grant, Wen-I Yeh, Yijia Pan, Mochtar Pribadi, Amit Mehta, Amanda Yzaguirre, Philip Chu, Shohreh Sikaroodi, Angela Gentile, Bruce Walcheck, Jianming Wu, Anhco Nguyen, Bahram Valamehr. A unique strategy to arm iPSC-derived CAR-T cells to perform antibody-directed cellular cytotoxicity to facilitate multi-antigen targeting [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1497.