Abstract 4875: B7-H3 (CD276) regulates the migration and infiltration of tumor-associated macrophages

B7-H3 (CD276), a member of the B7 family of immune checkpoint proteins, is expressed on tumor cells as well as various immune cells within the tumor microenvironment (TME), including macrophages. While evidence from numerous in vitro, in vivo, and ongoing clinical studies highlights its potential as a promising therapeutic target, the function and molecular mechanisms of B7-H3 in distinct stromal cell populations within the TME remain unclear. In this study, we investigated the roles of B7-H3 in tumor-associated macrophages (TAMs) using B7-H3 knockout mice, macrophage-tumor co-culture spheroids, and in vitro experiments. B7-H3 expression was significantly upregulated during the progressive stages of bone marrow-derived macrophage (BMDM) differentiation in vitro, supporting its functional role in macrophages. For in vivo studies, we generated B7-H3 knockout mice. Immunohistochemical staining and flow cytometric analysis of B16F10 melanoma tumor tissues subcutaneously implanted in B7-H3 knockout mice revealed a significant reduction in F4/80+ TAMs compared to tumors in wild-type mice. Similarly, anti-B7-H3 antibody treatment in 4T1 breast cancer-bearing mice reduced F4/80+ TAMs compared to control IgG treatment. However, tumor growth and the number of PCNA+ proliferating tumor cells were unaffected by either knockout or antibody-dependent inhibition of B7-H3 in vivo. Subsequent in vitro studies demonstrated that migration of BMDMs from B7-H3 knockout mice was impaired in both two-dimensional scratch-wound migration assays and a three-dimensional macrophage-tumor co-culture spheroid model. Furthermore, interferon-gamma treatment significantly enhanced the migration of BMDMs from wild-type mice, an effect that was completely blocked by a pharmacologic STAT1 inhibitor (fludarabine). Interestingly, BMDMs from B7-H3 knockout mice exhibited a significantly attenuated response to both interferon-gamma and STAT1 inhibition, indicating that B7-H3-dependent macrophage migration is mediated through the interferon-gamma and STAT1 signaling pathway. Collectively, our data indicate that B7-H3 regulates the migration and infiltration of TAMs into tumor tissues via the interferon-gamma/STAT1 signaling pathway. These findings suggest that B7-H3 is a promising therapeutic target for solid tumors such as melanoma and breast cancer. Citation Format: A-Ram Kang, Da Hyun Yun, Eun Jung Lee, Jong Dae Ji, Serkin Park. B7-H3 (CD276) regulates the migration and infiltration of tumor-associated macrophages [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4875.