Regulation of small extracellular vesicle production by sirtuin 1 as a determinant of breast cancer metastasis

Metastatic breast cancer remains a major clinical challenge and the primary cause of breast cancer-related mortality. Increasing evidence points to a role for tumor-derived small extracellular vesicles (EVs) secreted by cancer cells in preparing distant sites for metastasis. Sirtuin 1 (SIRT1), a metabolic and epigenetic regulator frequently downregulated in advanced breast cancer, influences key cellular processes, including lipid metabolism and autophagy, both of which are implicated in EV biogenesis. However, the mechanisms by which SIRT1 loss alters small EV production and function to promote breast cancer metastasis remain unclear. In this study, we investigated how SIRT1 expression influences the production and cargo of small EVs released by breast cancer cells, and how these changes affect aggressiveness. First, using triple-negative breast cancer (TNBC) cell lines with genetically altered SIRT1 levels, we observed that SIRT1 loss increased small EV secretion. Second, incubation of recipient breast cancer cells in vitro with small EVs derived from SIRT1-deficient cells enhanced their migration and invasiveness to a greater extent than with small EVs from SIRT1-proficient control cells. In vivo , inhibition of SIRT1 by genetic or small molecule approaches led to the development of larger tumors in both orthotopic transplantation and MMTV-PyMT mouse mammary tumor models. Moreover, animals preconditioned with small EVs derived from SIRT1-deficient cells, followed by tail vein injection of mammary tumor cells, showed increased lung colonization and metastatic outgrowth compared to those preconditioned with EVs derived from control cells, suggesting that SIRT1 regulation of EV cargo contributes to shaping a pro-metastatic microenvironment. In conclusion, this work identifies a novel link between SIRT1 status in breast cancer cells and their small EV-mediated signaling potential. Our findings position SIRT1 as a key regulator of small EV content and underscore the therapeutic potential of targeting SIRT1-dependent pathways that drive small EV-mediated metastasis in breast cancer.