Extracellular vesicles from N2-like neutrophils enhance triple-negative breast cancer aggressiveness through an L-selectin-dependent protumoral signaling axis

Neutrophils are among the first immune cells recruited to the tumor microenvironment and may acquire either antitumor (N1) or protumor (N2) characteristics. We previously demonstrated that extracellular vesicles (EVs) released by melanoma and breast cancer cells induce an N2-like phenotype in neutrophils. In the present study, we examined whether EVs released by these N2-like neutrophils (N2-EVs) affect the behavior of MDA-MB-231 triple-negative breast cancer cells. Analysis of N2-EVs revealed the presence of L-selectin on their surface. Blocking L-selectin interactions with fucoidan reduced N2-EV binding to tumor cells, indicating a role for this adhesion molecule in EV-cell communication. Treatment of MDA-MB-231 cells with N2-EVs increased mitochondrial reactive oxygen species (ROS) production and activated the PI3K/AKT pathway. In addition, N2-EVs enhanced tumor cell migration and invasion, accompanied by increased NF-κB activation and MMP-9 expression. These effects were markedly reduced when L-selectin-mediated interactions were inhibited. Our findings demonstrate that EVs released by N2-like neutrophils actively modulate the malignant phenotype of triple-negative breast cancer cells and suggest that L-selectin contributes to this process. Together, these results provide evidence that tumor-educated neutrophils may reinforce protumor signaling through the release of EVs.