In this study, we describe both the individual and the cooperative mechanisms by which CD169+ macrophages (MP) and CD8+ T cells support an anti-tumor immune response against breast cancer tumors. Breast cancer is the second leading cause of death in women of all ages each year, thus a deeper understanding of the molecular determinants that promote tumor development and suppress the anti-tumor immune response is still greatly needed. Here we use high dimensional flow cytometry, functional assays, and new mouse models to investigate the MP - T cell anti-tumor partnership. With our collaborators, we developed a preclinical mouse model of breast cancer by genetically engineering mammary stem cells to harbor three cancer driver mutations that are frequently associated in human patients: functional loss of the tumor suppressor genes p53 and mixed-lineage leukemia protein 3, and constitutive activation of the PI3 kinase. To characterize the immune cells in this tumor model, we developed a powerful 30-color high dimensional spectral flow cytometry panel to track and phenotype most known immune cell subsets. We found that CD169+ MP rapidly infiltrated and outnumbered all other populations during tumor growth, suggesting that they play an important role in shaping the tumor microenvironment. Surprisingly, although current literature describes breast cancer MP to be highly immunosuppressive, our phenotypic analysis showed only low expression of the pro-tumor MP markers TREM2 and CD206, but extremely high expression of the anti-tumor markers FOLR2 and CD64. To further investigate if these MP promoted or prevented tumor growth, we selectively depleted CD169+ MP in tumor-bearing mice and observed significantly accelerated tumor growth compared to the non-depleted control group, suggesting that these MP promoted anti-tumor responses. MP depletion also affected tumor-infiltrating CD8+ T cells, which produced lower effector functions (e.g., secretion of IFNγ, CCL3, TNF), indicating that MP may also boost anti-tumor CD8+ T cell responses. To further test if CD8+ T cells control tumor growth, we depleted either CD8+ or CD4+ T cells in tumor-bearing mice and observed that only CD8+ but not CD4+ T cell depletion accelerated tumor growth compared to the control group. MP have been documented to provide type I IFN signals to CD8+ T cells via the triggering of the STING pathway. To explore this mechanism in our model, we created a CD169creSTINGflox/KO mouse in which MP lack functional STING. Tumors in these mice grew much faster compared to those in the control group, consistent with the hypothesis that STING sensing occurs in tumor MP and promotes anti-tumor effector CD8+ T cell responses. Overall, our data provide evidence for an unexpected anti-tumor role for CD169+ MP in breast tumors, presumably through enhancing anti-tumor effector CD8+ T cell responses.

Citation Format: Nicole Couturier, Marie Boutet, Kenta Nishitani, Wenjun Guo, Gregoire Lauvau. Unraveling the anti-tumor partnership of CD169+ macrophages and CD8+ T cells in a preclinical breast cancer model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3959.