Background: Despite the recent progress in cancer management, metastatic prostate cancer remains incurable. In the context of immunotherapy failure, we need to identify new immune biomarkers, to improve patient therapy, and explore their potential as therapeutic target. To this end, we screened the prognostic value of soluble immune checkpoints in the plasma of metastatic prostate cancer patients at diagnosis, and depicted their role in anti-tumor activities. Methods: Patients with hormone naive metastatic prostate cancer were prospectively included in the NK-Prostate study (NCT02963155). Blood collection was performed before treatment and patients were followed for 5 years. We determined by ELISA assay plasma levels the soluble form of PD-L1, Pan-BTN3A, BTN3A1, BTN2A1 and BTLA. Large extracellular vesicles (MVs) were pelleted at 10 000g by sequential ultracentrifugation. Results: 66 patients were included in the NK-Prostate with a median follow-up of 42.6 months. Only plasma levels of BTN2A1 were associated with overall survival > 50 months. Patients with enriched sBTN2A1 (soluble form of BTN2A1) presented poorer overall survival (HR for death 2.91 95%CI 1.33 – 6.33, p=0.0072) and progression-free survival than low-level patients (HR for progression 2.73 95%CI 1.48 – 5.05, p=0.0012). Prostate cancer cell line produced sBTN2A1, with 40% associated with extracellular vesicles. High level of BTN2A1+ microvesicles (MVs) were identified by spectral cytometry in patient’ plasma but not in healthy volunteer plasma, and deep phenotyping revealed that B cell-derived BTN2A1+ MVs were associated with shorter overall survival (HR 4.22 95%CI 1.68 – 10.57, p=0.0021). To evaluate the impact of BTN2A1+ MVs on γδ T cells, we generated BTN2A1 KO MVs, BTN2A1 overexpressed MVs, and BTN2A1 mutant overexpressed MVs (that do not interact with γδ TCR). We demonstrated that BTN2A1 overexpressed MVs could specifically interact with γδ T cell; and impaired their proliferation and cytotoxicity against prostate cancer cell lines (LNCaP, DU145 and PC3). Conclusions: Our study shows the negative impact of B cell derived BTN2A1+ MVs in hormone naïve metastatic prostate cancer. Plasma MVs phenotyping is an easy-to-perform test, accessible for all patients. Our results highlight the role of γδ T cell in the anti-tumor immunity in prostate cancer, making them an interesting target for immunotherapy in this pathology. Clinical trial information: NCT02963155.