Exercise-induced extracellular vesicles delay tumor development by igniting inflammation in an immunologically cold triple-negative breast cancer mouse model.

Abstract

BACKGROUND Preclinical studies demonstrate that exercise reduces tumor incidence and growth. Rapid release of extracellular vesicles (EVs) during exercise suggests their potential role as mediators of exercise-induced systemic effects and physiological adaptation. This study investigated the impact of exercise-induced plasma EVs on tumor growth and immune tumor microenvironment in murine models of triple-negative breast cancer (TNBC): EO771 (a C57BL/6-derived TNBC cell line) and 4T1 (a BALB/c-derived TNBC cell line). METHODS Size exclusion chromatography was used to isolate exercise-induced EVs from plasma of healthy female mice (BALB/c and C56BL/6, n = 30 per strain) that underwent ten 30-min moderate-intensity treadmill running sessions over 2 weeks. Nanoparticle tracking analysis, Western blot, and electron microscopy confirmed the presence of EVs in the samples. Tumor-bearing mice (n = 72 per strain) were administered with exercise-induced EVs before or/and after tumor implantation. Local and systemic immune responses were assessed using flow cytometry, enzyme-linked immunosorbent assay (ELISA), and quantitative polymerase chain reaction (qPCR). RESULTS Administration of exercise-induced EVs, particularly before tumor implantation, significantly suppressed tumor growth and reduced tumor burden in both TNBC models. In EO771, endpoint tumor volumes were 278-330 mm³ in treated groups compared to 799 mm³ in untreated (p < 0.0001), while in 4T1, treated groups showed volumes of 287-564 mm³ vs. 696 mm³ in untreated (p = 0.0002). Notable differences in tumor-infiltrating lymphoid and myeloid cell subpopulations indicated immunomodulatory effects of exercise-induced EVs, particularly in the 4T1 model, where their continuous administration significantly increased intratumoral cluster of differentiation 8 (CD8) T lymphocyte proportion (5.77% vs. 0.90% in untreated, p < 0.0001). Similarly, in the EO771 model, exercise-induced EVs administered before tumor implantation led to a marked rise in intratumoral CD8 T lymphocytes (2.24% vs. 1.08% in untreated, p = 0.0181). CONCLUSION Our findings indicate that exercise-induced EV treatment elicits a pro-inflammatory antitumor immune response, suggesting a shift of immunologically cold TNBC tumors towards a more inflamed phenotype associated with better outcomes. Our study supports the further investigation of EVs as modulators of antitumor immunity and their potential utility in enhancing the efficacy of immunotherapy.