Abstract 4009: Discovery of inhibitory CAR target DSG1 for damping NECTIN4 on-target off-tumor toxicity in iPSC-derived CAR-T cell therapy

Abstract

Abstract Century Therapeutics is developing NECTIN4 targeted induced Pluripotent Stem Cell (iPSC) derived Chimeric Antigen Receptor (CAR) T cell therapy. NECTIN4 targeted antibody drug conjugate enfortumab vedotin, while generally well tolerated, causes severe skin adverse events in some patients, thought to be driven by on-target off-tumor toxicity against NECTIN4 displaying skin keratinocytes. We include a safety switch in our first clinical-stage iPSC-derived CAR-NK cell therapy, CNTY-101, which provides an antibody target for elimination of the engineered cells should that be necessary. An additional method for reducing the incidence and severity of this type of adverse event is through the incorporation of an inhibitory CAR, also known as a NOT-gate, directed against antigen expressed on normal skin keratinocytes, but the optimal surface protein to target with such an inhibitory CAR for a NECTIN4 targeted therapy is unknown. Here, we identify DSG1 (Desmoglein-1) as a top inhibitory CAR target for NECTIN4 targeted CAR-T cell therapy by conducting a multi-omics analysis of public single-cell RNAseq, bulk RNAseq, and protein microarray immunohistochemistry datasets. We find that DSG1 is constitutively and uniformly displayed on the surface of skin keratinocytes in the layers of epidermis where they display NECTIN4, and DSG1 gene expression in normal skin is uniformly high across individuals. Conversely, DSG1 mRNA and protein are rarely expressed in NECTIN4 expressing cancer indications including bladder, breast, non-small cell lung, ovarian, and pancreatic. Further, autoimmune antibodies specific to DSG1 have been described, suggesting that DSG1 is targetable by a CAR in situ. Finally, DSG1 is not expressed by our iPSC-derived CAR-T cells. While NECTIN4 expression is greatest in skin keratinocytes, epithelial cells in other tissues express NECTIN4 at lower levels. In this study, we compare the expression levels of NECTIN4 to targets associated with tissue and cell-type specific on-target off-tumor toxicity in primary CAR-T cell clinical trials. Our findings support the strategy of DSG1-directed inhibitory CAR incorporation into NECTIN4 specific CAR-T cell therapeutic candidates. Citation Format: Matthew S. Hall, Christopher M. Dower, Michael Miller, John Wheeler, Jill Carton, Ohad Manor, Hyam Levitsky. Discovery of inhibitory CAR target DSG1 for damping NECTIN4 on-target off-tumor toxicity in iPSC-derived CAR-T cell therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4009.