Elevated extracellular vesicular Nm23-H1 subdues the pro-migratory potential of breast cancer cell-derived extracellular vesicles

Metastasis is a key determinant in cancer mortality which is often associated with decreased levels of Nm23-H1, a well-established metastasis suppressor. Despite lacking a secretion signal peptide, Nm23-H1 has been reported to be present in the extracellular space and enclosed within extracellular vesicles (EVs). While the presence of Nm23-H1 proteins in EVs released by cancer cells has been observed through proteomics profiling, the role of vesicular Nm23-H1 remains unclear. Here, we investigated the function of vesicular Nm23-H1 using MDA-MB-231 (highly metastatic, low Nm23-H1) and MCF-7 (low/non-metastatic, high Nm23-H1) breast cancer cell models. Our findings confirm that Nm23-H1 is indeed encapsulated within EVs, and its levels can be manipulated through overexpression and knockdown approaches. Functional assays revealed that EVs derived from MDA-MB-231 cells that contained high levels of Nm23-H1 exhibit impaired pro-migratory properties, suggesting that vesicular Nm23-H1 may act as a metastasis suppressor. Furthermore, EVs with increased levels of Nm23-H1 altered the transcript levels of multiple cancer-related genes in recipient cells and stimulated type I interferon signaling through STAT1 phosphorylation. These results suggest the existence of an unconventional signaling pathway mediated by the uptake of EVs enriched with Nm23-H1, which may contribute to the anti-metastatic effect of Nm23-H1 in the tumor microenvironment. Additionally, our study demonstrates that elevated Nm23-H1 levels can impact the abundance of various other proteins encapsulated within breast cancer cell-derived EVs, such as SUSD2 (Sushi Domain Containing 2) which can also modulate metastasis. •Nm23 is a metastasis suppressor protein which can be encapsulated within EVs.•MDA-MB-231 breast cancer cells over-expressing Nm23 produced EVs with elevated Nm23.•Nm23-enriched EVs suppressed cell migration and altered gene transcription.•Nm23-enriched EVs stimulated type I interferon signaling through STAT1.•Elevated Nm23 can affect the abundance of other proteins (e.g., SUSD2) in EVs.