Tumor associated macrophages-programmed core immunosuppressive microenvironments via Galectin-1 in breast cancer

There has been considerable interest in therapeutic targeting of tumor-associated macrophages (TAMs) mainly by depleting or repolarizing them toward anti-tumoral phenotypes. However, a core mediator co-opted for TAMs-mediated remodeling of tumor microenvironment remains poorly understood. Here, to dissect TAMs-mediated early microenvironment, we performed single-cell RNA sequencing of TAMs-enriched and -depleted mouse breast tumor at the early stages of progression, before detectable change in tumor growth appeared. We found that early intratumoral accumulation of TAMs endowed cancer cells with invasive properties to promote early dissemination and dampened antitumoral T cell responses. Accordingly, TAMs-depleted tumor using clodronate displayed a decreased epithelial-to mesenchymal transition (EMT) signature of cancer cells and harbored a distinct transcriptome associated with CD8+ T cells activation. Furthermore, we identified a role of Galectin-1 (Gal-1) as a molecular checkpoint in TAMs-induced CD8+ T cell exhaustion. Gal-1 inhibition reversed immune suppression via reinvigoration of CD8+ T cells, thus impairing tumor growth and boosting the efficacy of immune checkpoint inhibitor in a breast tumor model. These results provide a comprehensive insight into the early-programmed microenvironment by TAMs and reveal an immune evasion mechanism that could be targeted by Gal-1 to unleash antitumor immune responses. Overall design: To establish orthotopic implantation model, 4T1-Luc2 cells were orthotopically implanted into the fourth mammary fat pad of 8-week-old female BALB/c mice. Depletion of macrophages was performed by intraperitoneal injection of clodronate or vehicle liposome (Encapsula Nano Science) at days 3, 5 and 7 after 4T1 implantation. The cell suspensions were diluted in nuclease-free water to achieve a targeted cell count of 10,000. Cell suspension was mixed with master mix and loaded with Single Cell 3'V3.1 Gel Beads and Partitioning Oil into a chromium Next GEM chip G. RNA transcripts from single cells were uniquely barcoded and reverse-transcribed within droplets.