SPP1+ Macrophage-T Cell Axis Orchestrates the Immune Microenvironment in Breast Cancer Bone Metastasis

Background Breast cancer is a highly common malignancy in women, with bone as its most frequent site of distant metastasis, significantly increasing mortality risk. Methods This study integrated single-cell RNA sequencing datasets (GSE266330 and GSE243526) to systematically characterize the dynamic evolution of the tumor microenvironment from normal tissue to primary tumor and bone metastasis in breast cancer. Results In total, 141,056 high-quality single cells were analyzed, identifying 11 major cell types. Myeloid cell and T cell proportions were significantly elevated in bone metastases, suggesting their pivotal role in metastatic progression. The bone metastasis-enriched Mac-SPP1 macrophage subpopulation exhibited M2 polarization, high metabolic activity, and a significant association with poor patient prognosis ( P = 4.9e-05). It highly expressed transcription factors such as HOXA9 and MAF, potentially promoting metastasis through regulation of an immunosuppressive network. Among T cells, the CD4-S100A9 and CD8-GZMK subpopulations were significantly expanded in bone metastases and exhibited high cytotoxicity, high exhaustion, and metabolic reprogramming. Pseudotime analysis showed that CD4-S100A9 cells were at the initial state of CD4+ T cell differentiation, whereas CD8-GZMK cells serve a key transitional node from effector memory toward exhaustion. Cell-cell communication analysis revealed that Mac-SPP1 interact specifically with these CD4/CD8 T cell subsets through the SPP1-integrin axis (e.g., SPP1-CD44) and the CD74-MIF/COPA signaling axis, potentially driving T cell functional exhaustion and immune evasion. Conclusion This study is the first to delineate the synergistic pro-metastatic role of the Mac-SPP1–CD4-S100A9/CD8-GZMK axis in breast cancer bone metastasis, providing a theoretical basis for developing therapeutic strategies targeting the immune microenvironment.