Targeting Reticulin 4 (RTN4) Within Small Extracellular Vesicles Combats Metastasis and Reinforces Immunotherapy in Triple‐Negative Breast Cancer

ABSTRACT Small extracellular vesicles (sEV) are a class of natural vesicles rich in heterogeneous cargos with the great advantage of non‐invasive detection; these vesicles exhibit complex intercellular crosstalk and mediate important biological functions. However, the potential value of plasma sEV in clinical prognosis prediction of triple‐negative breast cancer (TNBC) and their biological functions have not been well elucidated. In this study, we isolated sEV from non‐metastatic and metastatic TNBC plasma samples. We found that the expression of reticulin 4 (RTN4) in metastatic patients was significantly higher than that in non‐metastatic patients. At the same time, clinical data showed that RTN4 was associated with poor prognosis and advanced‐stage TNBC patients. Subsequently, in vivo and in vitro assays showed that compared to RTN4 Low sEV, RTN4 high sEV significantly promoted tumour cell migration, invasion, epithelial‐mesenchymal transition (EMT) and lung metastasis, and upregulated the expression of PD‐L1 in tumour tissues and inhibited CD8 + T cell infiltration. Regarding mechanism research, we found that RTN4 within sEV drives tumour EMT and PD‐L1 expression by activating the NF‐κB signalling pathway. Further, through the combined treatment experiment of anti‐PD‐1 and anti‐RTN4, it was found that the combination of the two drugs was significantly superior to monotherapy in inhibiting tumour metastasis, EMT, and promoting CD8 + T cell infiltration. Our results highlight the molecular mechanism of sEV protein RTN4 in tumour progression and immune system regulation, indicating that RTN4 targeting and anti‐PD‐1 combined therapy have clinical potential. sEV protein RTN4 is a potential new prognostic marker for non‐invasive detection of TNBC and a new target for TNBC treatment.