Fig 2: CTLA4-related gene expression in TN-BCSCs. A Expression heatmap of the DEGs between TN-BCSCs and B.C.s (CD44− and CD24+). The red indicates the upregulated expression, and the green indicates the downregulated expression. B Intersection between DEGs in TN-BCSC and CTLA4-related genes (CTLA4 Gene Term). C Expression distribution plot of CTLA4-related DEGs in TN-BCSCs in eight cell subsets. D Violin plots of FBXO9, CHI3L1 and MYC. E The correlation between CHI3L1 and CTLA4 expression in B.C. was obtained from the TIMER2.0 database. The result is corrected by tumor purity. F and G The CHI3L1 expression in BCSCs obtained from MDA-MB-231 and MDA-MB-453 cells and the parental cells of BCSCs as measured by RT-qPCR and Western blot. H The expression of CHI3L1 in the supernatant of conditioned medium of BCSCs obtained from MDA-MB-231 and MDA-MB-453 cells and the parental cells of BCSCs as determined by ELISA. ns p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001. Cell experiments were repeated three times

Fig 3: In vivo verification of the involvement of TN-BCSCs in immune escape of TNBC through CHI3L1/MAF. A Growth of MDA-MB-231 and BCSCs-231 orthotopic B.C. transplanted tumors. The tumor growth curve drawn based on the relative bioluminescence intensity is on the left. The bioluminescence intensity and tumor size at the end of the experiment are shown on the right side. B and C Detection of CHI3L1, MAF and CTLA4 expression in tumor tissues by RT-qPCR and Western blot. D and E, Detection of CTLA4+ T cell proportion and CD8+ T cytotoxicity (GzmB and Ki67) in tumor tissues by flow cytometry (n = 8). F Growth of MDA-MB-453 orthotopic B.C. transplanted tumor. The tumor growth curve drawn based on the relative bioluminescence intensity is on the left. The bioluminescence intensity and tumor size at the end of the experiment are shown on the right side (n = 8). G Flow cytometry to detect the proportion of BCSCs in MDA-MB-453 orthotopic B.C. transplanted tumors in mice treated with hrCHI3L1 or sh-MAF-1 (n = 8). H and I, Detection of CTLA4+ T proportion and CD8+ T cytotoxicity in MDA-MB-453 orthotopic B.C. transplanted tumor in mice treated with hrCHI3L1 or sh-MAF-1 by flow cytometry. ns p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001

Fig 4: Schematic illustration of the molecular mechanism by which TN-BCSC-derived CHI3L1 promotes immune escape of TNBC via the MAF/CTLA4 axis. TNBC mainly contains 8 cell subsets, among which TN-BCSCs exist in the epithelial cell subgroup. As a characteristic gene expressed in TN-BCSCs, CHI3L1 is related to TNBC cell stemness. It upregulates the expression of CTLA4 in T cells through MAF, thereby inhibiting the cytotoxicity of CD8+ T cells and producing immunosuppression. CTLA4+ T cells promote the stemness phenotype of TNBC cells by secreting S100A4 through positive feedback