[Objective] Increased infiltration of CD8+ T cells and decreased infiltration of M2 macrophages are favorable prognostic factors in high-grade serous ovarian cancer (HGSOC). CD276 (B7H3) is a transmembrane co-inhibitory protein belonging to the B7 family. CD276 expression is suppressed in CD8+ T cell-rich, PD-L1-high immune reactive tumors. This suggests that CD276 plays immunosuppressive roles in addition to acting as a molecular brake for countering the T cell activation. In this study, we aimed to elucidate the role of CD276 in HGSOC microenvironment toward M2 macrophages. [Methods] CD276 expression was evaluated in HGSOC clinical cases (n=62) by immunohistochemistry tests. The correlation between survival and the presence of tumor-infiltrating immune cells, including M2 macrophages, was assessed. CD276 knockout murine ovarian cancer cell lines (HM1 and ID8) were established and used for experiments. [Results] In HGSOC patients, higher CD276 expression associated with poor prognosis (p<0.001). CD276 expression positively correlated with the number of CD206+ M2 macrophages (Pearson's r=0.29, p=0.02), but not with that of CD8+ T cells. Although CD276 knockout did not affect in vitro proliferation of HM1 and ID8 cell lines, CD276 knockout tumors were less progressive than control tumors in immunocompetent mouse models (p<0.001). Lower infiltration of CCR2+/CD206+ M2 macrophages and higher infiltration of IFN-γ+ CD8+ T cells was observed in the knockout tumors (p<0.001 and P=0.02, respectively). The difference in progression between CD276 knockout tumors and control tumors was negated when tumor cells were inoculated into immunodeficient nude mice. RNA sequencing and protein analyses revealed decreased levels of CCL2, the ligand of CCR2, in CD276 knockout cells and tumors. In vitro, monocyte chemotaxis was reduced by CD276 knockout cells' supernatant (p<0.001), and the difference was partially negated with CCR2 antagonist pretreatment. Furthermore, CCR2 blocking in vivo reduced tumor-infiltrating M2 macrophages and tumor volume in the CD276 control (p=0.048 and p=0.02, respectively), but not in the CD276 knockout group, suggesting that CCL2-CCR2 axis-induced M2 macrophages partially contribute to CD276-mediated tumor progression. [Conclusion] CD276 contributes to an unfavorable immunosuppressive tumor microenvironment driven by M2 macrophages via the CCL2-CCR2 axis in HGSOC.

Citation Format: Taito Miyamoto, Ryusuke Murakami, Masayo Ukita, Mana Taki, Koji Yamanoi, Kaoru Abiko, Ken Yamaguchi, Junzo Hamanishi, Takashi Kobayashi, Masaki Mandai. CD276 in tumor cells contributes to the M2 macrophage-rich immunosuppressive tumor microenvironment via upregulating the CCL2-CCR2 axis in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2746.