Dose-dependent bidirectional effects of gemcitabine in 4 T1 breast tumors are associated with angiogenesis and PMN-MDSC-linked immunosuppression.

Metronomic chemotherapy is designed to leverage low-dose scheduling to remodel the tumor microenvironment, yet subtherapeutic exposure may also activate tumor-supportive programs. Here, we investigated whether Gemcitabine (GEM) exhibits a dose-dependent bidirectional phenotype in an immunocompetent 4 T1 breast cancer model and defined accompanying vascular and immune changes. Across a graded GEM dosing range, low-dose exposure promoted tumor progression, increased angiogenesis-associated markers CD31 and laminin, elevated circulating pro-angiogenic cell signatures marked by CD61 and VEGFR2, increased Ki67 positivity, and reduced apoptosis. In contrast, higher-dose exposure suppressed tumor growth, reduced vascular markers, shifted tumors toward lower proliferation and higher apoptosis, and was accompanied by increased ALT and AST. Immune profiling showed that low-dose exposure expanded intra-tumoral PMN-MDSCs and increased the PMN/M-MDSC ratio while decreasing the CD8/Treg ratio, whereas higher-dose exposure reversed these trends. Multiplex mediator profiling highlighted VEGF-A, GM-CSF, G-CSF, CCL2, CXCL1, IL-10, and TGF-β, and correlation analyses linked pro-angiogenic and granulocytic cues to vascular density, PMN-MDSC dominance, T cell imbalance, and tumor burden. These findings define, in an immune-intact 4 T1 model, a dose window in which GEM shifts from tumor suppression to tumor promotion in association with angiogenesis-related remodeling and functionally relevant myeloid immunosuppression, underscoring the need for biomarker-guided calibration of metronomic gemcitabine strategies and for validation in metastatic settings.