Abstract A034: Rac-enhanced chimeric antigen receptor-macrophage (Race-CAR-M) immunotherapy for Triple Negative Breast Cancer
Authors: Mishra Abhinava K , Dhavan Pratik , Rodriguez Anthony , Montell Denise J 
Published: February 2025

Abstract
Triple-Negative Breast Cancer (TNBC) is an aggressive subtype lacking estrogen, progesterone, and HER2 receptors, limiting targeted therapy options. Conventional systemic therapies, such as chemotherapy, are non-specific, leading to significant toxicity. Despite the success of Chimeric Antigen Receptor T-cell (CAR-T) therapy in hematologic cancers, its use in solid tumors like TNBC is hindered by challenges such as limited T-cell infiltration, the immunosuppressive tumor microenvironment (TME), antigen heterogeneity and CAR-T-cell exhaustion. Macrophages can engulf pathogens, dying cells, and cancer cells and naturally infiltrate TME. An innovative approach involves engineering macrophages to express CAR that specifically target antigens on cancer cells, thereby creating CAR-Macrophages (CAR-Ms). CAR-M has several potential advantages over CAR-T such as more efficient tissue infiltration, tumor cell removal by engulfment, activation of broader adaptive immune responses, and reduced autoimmune risks. Early studies show that CAR-Ms can convert the immunosuppressive tumor microenvironment into a pro-inflammatory state, enhancing anti-tumor immunity. Initial clinical trials indicate CAR-Ms are safe and show some anti-tumor activity, but their efficacy in completely eradicating tumors is not fully established. We found that co-expression of an active form of Rac2 (Rac2E62K) enhances the ability of CAR-M to specifically recognize and engulf living target cancer cells. We propose that Rac-enhanced CAR-M (Race-CAR-M) has the potential to improve CAR-M efficacy against numerous cancers. To test the ability of Rac2E62K to enhance anti-tumor activity of macrophages in a more realistic 3D environment, we have adapted the chicken chorioallantoic membrane (CAM) assay to study tumor-macrophage interactions. This low-cost ethical model allows in vivo monitoring of angiogenesis and metastasis and can be used to test drug combination therapies with Race-CAR-Ms. Our ongoing work indicates that Rac2E62K macrophages cooperatively infiltrate the TNBC microenvironment and dramatically shrink tumors by engulfing target cancer cells. Rac2E62K macrophages limit secretion of proangiogenic Vascular Endothelial Growth Factor (VEGF) and reduce neoangiogenesis. In addition, nanoparticle encapsulated Olaparib produces additive effects with Rac2E62K macrophages in the CAM tumor xenograft model of TNBC. We are currently engineering Race-CAR-Ms for TNBC by incorporating Rac2E62K together with a CAR targeting TNBC specific tumor-associated antigens (TAA). Since Rac operates downstream of engulfment receptors in macrophages, our approach offers a tunable system for engineering CAR-M therapies against various cancers.
Citation Format: Abhinava K Mishra, Pratik Dhavan, Anthony Rodriguez, Denise J Montell. Rac-enhanced chimeric antigen receptor-macrophage (Race-CAR-M) immunotherapy for Triple Negative Breast Cancer [abstract]. In: Proceedings of the AACR IO Conference: Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2025 Feb 23-26; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2025;13(2 Suppl):Abstract nr A034.