PMN-MDSCs-derived exosomal S100A9 drives breast cancer progression by enhancing cancer stemness and CXCL5-mediated metastatic potential

Abstract Polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) represent a critical subset of immunosuppressive cells within the tumor microenvironment. Their interaction with breast cancer stem cells (BCSCs) constitutes a vital link that drives the malignant progression of breast cancer, particularly in triple-negative breast cancer (TNBC). However, the precise molecular mechanisms through which these cells mediate BCSC-related stemness reprogramming and enhanced metastatic potential in breast cancer remain inadequately understood. In this study, PMN-MDSCs were isolated from mouse breast cancer solid tumor tissues. Functional characterization was conducted using flow cytometric phenotyping, nuclear morphology analysis, and T cell immunosuppressive function assays. In vivo tumorigenesis experiments confirmed their pro-tumor growth effects and enhanced tumor stem cell characteristics. Concurrently, the CD44 hi CD24 lo subpopulation of 4T1 cells was sorted to systematically verify their tumor stem cell properties and stemness plasticity. In vitro co-culture experiments demonstrated that PMN-MDSCs significantly increased the proportion of CD44 hi CD24 lo stem-like subpopulations in 4T1 cells, prompting non-stem cells to differentiate into stem cell-like cells. They also promoted STAT3 phosphorylation, upregulated the expression of stemness-related proteins and mesenchymal markers, downregulated the epithelial marker E-cadherin, and markedly enhanced CXCL5 secretion. These effects were effectively reversed by the stemness inhibitor Napabucasin. Transwell non-contact co-culture and exosome function assays further confirmed that PMN-MDSCs regulate cancer cell stemness in a paracrine manner. S100A9, a key effector molecule in their exosomes, induced CXCL5 secretion by activating the STAT3 pathway. CXCL5 subsequently activated both the ERK1/2 and STAT3 pathways through autocrine and paracrine mechanisms, enhancing tumor cell metastatic potential and establishing a positive feedback loop that sustains its own secretion. This loop effect was blocked by a CXCL5-neutralizing antibody. Single-cell sequencing data analysis and clinical sample validation of TNBC indicated significantly elevated infiltration of myeloid cells, as well as increased S100A9 and CXCL5 expression levels in cancer tissues, alongside a marked upregulation of the mRNA stemness index. The stemness index was closely associated with poor patient prognosis. In summary, this study reveals that PMN-MDSCs can activate the STAT3-CXCL5-ERK positive feedback regulatory axis via exosomal S100A9, synergistically enhancing breast cancer cell stemness and metastatic capacity. These findings provide a theoretical reference and potential intervention targets for targeting the tumor microenvironment to inhibit TNBC progression.