LAUSR

Despite new therapies, breast cancer remains one of the leading causes of cancer-related death in women worldwide. Part of the issue remains the inability of these therapies to target cancer stem cells (CSCs). These cells show unique resistance to chemotherapy and have enhanced capacity for self-renewal. Recently, our group has shown that Epithelial membrane protein-2, or EMP2, is expressed on CSCs defined by increased aldehyde dehydrogenase activity (ALDH+). Clinical data supports the role of EMP2 in advanced breast cancer as increased expression of the protein is observed in metastatic lesions compared to primary tumors and its expression is enhanced in chemotherapy resistant tumors. In parallel, EMP2 expression correlated with enhanced mammosphere formation, an accepted surrogate for cells with enriched stem cell capacity. We have recently developed a novel IgG1 monoclonal antibody to EMP2, which has been shown to reduce tumor load. As the antibody-antigen complex rapidly internalizes, we hypothesized that its anti-tumor effects could be enhanced through the creation of a fusion protein. Granzyme B (GrB) is a serine protease that plays a critical role in the body9s defense against viral infection and tumor development by initiating the apoptotic cascade via both caspase-dependent and -independent mechanisms. We developed novel fusion proteins composed of the anti-EMP2 backbone as the targeting moiety and granzyme B as the cytotoxic payload. Our initial design comprised of the active GrB fused to the IgG heavy chain (Fc) domain containing 2 single-chain antibodies against EMP2, a format that results in a longer plasma retention time. This new construct showed high and specific affinity for EMP2 (10−10M) using both flow cytometry and ELISA based formats. Moreover, treatment with these agents induced cell death to EMP2 positive cells, including those characterized by high ALDEFLUOR activity. Preclinical assessment of the anti-EMP2 immunotoxins showed no acute toxicity and acceptable plasma retention. Preliminary results using a 4T1 model system produced a significant reduction in tumor burden. Our results suggest that continued development of these agents may be successful at targeting advanced disease. Research supported, in part, by NCI R21 CA234642 and the Clayton Foundation for Research. Citation Format: Khalid A. Mohamedali, Kok Su-yin, Lawrence H. Cheung, Naina Dhawan, Michael G. Rosenblum, Madhuri Wadehra. Targeting breast cancer stem cells with anti-EMP2 immunotoxins delivering granzyme b [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 2891.