Extracellular vesicles of cancer cells induce FOXP3+ fibroblasts and facilitate tumor invasion via the Wnt3-β-catenin pathway.

Forkhead-box-protein P3 (FOXP3) is a key transcription factor in T regulatory cells (Tregs). However, its expression and significance in non-immune stromal cells in the tumor microenvironment remain unclear. Here, we demonstrated FOXP3 expression in stromal fibroblasts of mouse and human gastrointestinal tumors. Immunohistological examination revealed FOXP3 expression in αSMA collagen I myofibroblasts. In the mouse omentum inoculated with gastric cancer cells, cytokeratin/CD45/FoxP3 stromal cells were identified via flow-cytometry, and high FOXP3 expression was noted in fibroblasts surrounding the tumor glands, where CD8 T cells were exclusively infiltrated. Extracellular vesicles (EVs) from mouse gastric cancer cells upregulated Foxp3 transcription in fibroblasts, which partly depends on increase of transcription factors including NFAT1 and c-Rel, and activation of TGF-β and STAT5 pathways. In FOXP3 fibroblasts, immunosuppressive cytokines including IL-10 and CCL2 were upregulated. FOXP3 overexpression in NIH/3T3 fibroblasts enhanced Wnt3a-induced β-catenin responses, accompanied by cell growth and tumor invasion in mice stomach. As the mechanism, FOXP3 induced CDH11 expression in fibroblasts, which augmented the Wnt3/β-catenin pathway, and blocking of CDH11 suppressed tumor invasion mediated by FOXP3 fibroblasts. Our results suggest that cancer cell-derived EVs regulate FOXP3 expression in stromal fibroblasts, attenuating antitumor immunity, and facilitating tumor invasion.