Wong, G. L.; Khan, M. S.; Manore, S.; Bindal, S.; Singh, R.; Lo, H.-W.

Mechanisms underlying breast cancer brain metastasis (BCBM) are still not well understood. Here, we identified that BCBM patient serum contained extracellular vesicles (EVs) with high levels of microRNAs (miRNAs)-107 and -425. Levels of miR-107 and miR-425 were elevated in brain metastases, and the elevation was associated with poor patient prognoses. Ectopic expression of miR-107 and miR-425 promoted mammospheres; inhibition of miR-425, but not miR-107, suppressed breast cancer mammosphere formation. EVs from miR-425-overexpressing breast cancer cells strongly activated astrocytes whereas their inhibitors abrogated the effect. Conditioned media from miR-425-activated astrocytes promoted mammospheres. Within astrocytes, miR-425 suppressed expression of transcription factor ZNF24, which downregulated CCL8 cytokine expression/secretion, leading to subsequent activation of astrocytes. We further determined the role of miR-425 in brain metastasis formation and observed that miR-425-overexpressing breast cancer cells exhibited significantly more aggressive growth in mouse brains compared to control cells. Immunohistochemistry and immunofluorescence analysis of mouse brain metastases revealed that miR-425 tumors exhibited significantly increased activation, intratumoral accumulation, and proliferation of astrocytes, and a decrease in ZNF24 expression compared to control tumors. Together, our findings demonstrate that breast cancer EV-derived miR-425 promotes BCBM via activating astrocytes in the brain microenvironment through the novel EV-miR-425-ZNF24-CCL8 signaling axis.