Breast Cancer-Derived Exosomes Promote Tumor Immune EscapeThrough the miR-374a-5p/ARRDC3 Axis

This study aimed to investigate the molecular mechanism by which breast cancer-derived exosomes promote tumor immune escape through the miR-374a-5p/ARRDC3 axis, and to explore its potential as anovel target for breast cancer immunotherapy. In the study, qRT-PCR (quantitative real-time PCR) was used to detect the expression levels of miR-374a-5p and ARRDC3 in breast cancer cells. Exosomes were isolated from MDAMB-231 cells and co-cultured with recipient MDA-MB-231 cells for 48 h. MDA-MB-231 cells were divided intosix groups: control group, exosome-negative control (Exo-NC) group, negative control inhibitor (NC inhibitor)group, miR-374a-5p inhibitor group, miR-374a-5p inhibitor+shRNA negative control (sh-NC) group, and miR374a-5p inhibitor+sh-ARRDC3 group. Cell proliferation, migration, and invasion were evaluated by colony formation, wound healing, and Transwell assays, respectively. Co-culture with CD8⁺ T cell was performed to assess CD8⁺T cell apoptosis and cytotoxic activity against tumor cells. Cytokine levels in the supernatant were determined byELISA, and PD-L1 expression was detected by qRT-PCR and Western blot. Compared with the normal mammaryepithelial cell line MCF-10A, miR-374a-5p expression was significantly upregulated, while ARRDC3 expression was downregulated in breast cancer cell lines (MDA-MB-231, Hs578T, MDA-MB-468, and MDA-MB-453)(P<0.05). Analysis of the GEPIA database also revealed significantly reduced ARRDC3 expression in breast cancertissues (P<0.05). Compared with the Con group, the expression of miR-374a-5p, the number of plate clones, thescratch repair rate, the number of penetrating cells, and the apoptosis rate of CD8⁺ T cells in the Exo-NC groupand the NC inhibitor group were significantly increased (P<0.05), and the levels of TNF-α, IFN-γ and IL-4, andthe expression level of ARRDC3 were significantly decreased (P<0.05). Compared with Con group, the expression of miR-374a-5p, the number of plate clones, the rate of scratch repair, the number of transmembrane cells andthe apoptosis rate of CD8⁺ T cells in Exo-NC group and NC inhibitor group were significantly increased (P<0.05),and the levels of TNF-α, IFN-γ and IL-4, the killing rate of CD8+ T cells and the expression level of ARRDC3 weresignificantly decreased (P<0.05). Compared with Exo-NC group and NC inhibitor group, miR-374a-5p expression, plate clone number, scratch repair rate, transmembrane cell number and CD8+ T cell apoptosis rate in miR374a-5p inhibitor group were significantly decreased (P<0.05), and TNF-α, IFN-γ and IL-4 levels, CD8+ T cellkilling rate and ARRDC3 expression level were significantly increased (P<0.05). Compared with the miR-374a-5pinhibitor+sh-NC group, the expression level of ARRDC3, the levels of TNF-α, IFN-γ and IL-4, and the killing rateof CD8⁺ T cells in the miR-374a-5p inhibitor+sh-ARRDC3 group were significantly decreased (P<0.05), and theplate clone number, scratch repair rate, transmembrane cell number, CD8⁺ T cell apoptosis rate and PD-L1 expression level were significantly increased (P<0.05). In summary, BC-derived Exo can up-regulate the expression ofPD-L1 through the miR-374a-5p/ARRDC3 axis, inhibit the activity of CD8+ T cells in the tumor microenvironment,and promote the immune escape of cancer cells. 

CSTR: 32200.14.cjcb.2026.06.0003