miR-466f-3p promotes pro-inflammatory activation of macrophages through NF-κB signaling and suppresses melanoma metastasis

BACKGROUND: Cancer metastasis remains a leading cause of cancer-related mortality, and there is an urgent need for innovative immunoregulatory strategies to suppress metastatic progression. Macrophages play a pivotal role in tumor immunity; however, the molecular mechanisms by which extracellular vesicles (EVs) regulate macrophage functional states in metastasis control remain incompletely understood. METHODS: In this study, we investigated the anti-metastatic mechanisms of induced pluripotent stem cell-derived extracellular vesicles (iPS-EVs) using a murine melanoma liver metastasis model. The contributions of adaptive and innate immunity were assessed using Rag2-deficient mice and clodronate liposome-mediated macrophage depletion. EV-associated microRNA profiling was conducted, and functional analyses were performed through in vivo metastasis assays, macrophage activation assays, gene expression analyses, and bioinformatic target prediction. RESULTS: Pretreatment with iPS-EVs significantly suppressed melanoma liver metastasis in vivo. This effect was largely maintained in Rag2-deficient mice, suggesting a limited role of adaptive immunity. In contrast, macrophage depletion completely abolished the anti-metastatic effect, indicating that macrophages are essential mediators. miRNA profiling revealed a high abundance of miR-466f-3p in iPS-EVs. EVs derived from F10 cells overexpressing miR-466f-3p exhibited similar anti-metastatic effects in vivo. Functional analysis showed that miR-466f-3p induced NF-κB activation and increased expression of IL-6 and Nos2 in macrophages, promoting a pro-inflammatory activation state. These findings suggested that macrophages exhibited a partial M1-like phenotype. Bioinformatic analysis suggested that multiple candidate molecules, including Commd6, may be involved in this regulation. CONCLUSION: These findings suggest that miR-466f-3p regulates innate immunity by promoting pro-inflammatory activation of macrophages and contributes to the suppression of metastasis. iPS-EVs represent a promising platform for EV-based immunotherapy targeting innate immune reprogramming to suppress cancer metastasis.