Abstract B044: Development of Tumour-specific CD146 CAR-T Cells for Targeted Cancer Immunotherapy of Solid Tumours

Abstract Background: In comparison to the success with hematologic cancers, CAR-T cells for solid cancers have been held back by significant “on-target/off-tumour” toxicities caused by the absence of specific target proteins. Although CD146 (aka MCAM) is a glycoprotein expressed on benign cells, including endothelial cells, pericytes and smooth muscle cells, a conformation has been identified enriched in tumours that we call tumour-specific (ts) CD146. Herein, we attempt to engender tumour specificity of CAR-T cells by targeting tsCD146 widely expressed in several solid tumours, notably melanoma and ovarian cancer. Methods: We assessed the CAR activity using flow cytometry by measuring CD69 expression on CAR-expressing Jurkat cells, an early activation marker, after co-culturing with CD146-positive and CD146-negative human melanoma and ovarian cancer cell lines. The effector functions of CAR-T cells have been evaluated both in vitro by the relative luciferase activity of target cells and ELISA for IFN-g and TNF-a secretion after co-culturing and in vivo by assessing the tumour growth by IVIS bioluminescent imaging. Results: Through multiple rounds of protein engineering, we have developed several CAR constructs to respond to a tumour-enriched CD146 isoform. Our CAR demonstrated antigen-dependent activation of Jurkat cells when co-cultured with CD146-positive human melanoma and ovarian cancer cell lines, comparable to a CAR targeting pan-CD146. CAR primary T cells co-cultured with these cell lines exhibited proportional killing activity and other effector functions. Upon assessment of anti-tumour activities of CAR-T cells in mice engrafted with WM266-4 cells, CAR-T-treated mice showed reduced tumour burden and prolonged survival compared to mice treated with untransduced T cells. Conclusions: We have successfully integrated the antigen-binding domain of an anti-tsCD146 antibody into a CAR scaffold. The ability of tsCD146 CAR primary T cells to target CD146-expressing cancerous cells provides a novel approach for targeted immunotherapy of solid tumours. Citation Format: Erez Uzuner, Holly Mole, Sevim Beyza GURLER, Chun Wai Wong, Macarena Lucia Fernandez Carro, Rotem Leshem, Kieran Sefton, Helen Tovey, Andrew Picken, Marcel Blot-Chabaud, Adam Hurlstone. Development of Tumour-specific CD146 CAR-T Cells for Targeted Cancer Immunotherapy of Solid Tumours [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr B044.