Tumor-derived exosomes promote angiogenesis via adenosine A2B receptor signaling.

RATIONALE: One hallmark of tumor-derived exosomes (TEX) is the promotion of cancer progression by stimulating angiogenesis. This study was performed to evaluate the role of adenosine receptors in TEX-induced angiogenesis. METHODS: TEX produced by UMSCC47 head and neck cancer cell line were isolated by mini size exclusion chromatography (mini-SEC). Enzymatic activity of ectonucleotidases CD39/CD73 carried by TEX was measured by HPLC. Adenosine content of TEX was measured by UPLC-MS/MS. Primary human macrophages were co-incubated with TEX or exosomes derived from the plasma of head and neck cancer patients and their marker expression profile was analyzed by flow cytometry. The macrophage secretome was analyzed by angiogenesis arrays. The in vitro angiogenic potential of TEX was evaluated in endothelial growth studies. Results were validated in vivo using basement membrane extract plug assays in A1R-/-, A2AR-/- and A2BR-/- rats. Vascularization was analyzed by hemoglobin quantification and immunohistology with vessel and macrophage markers. RESULTS: TEX carried enzymatically active CD39/CD73 and adenosine. TEX promoted A2BR-mediated polarization of macrophages toward an M2-like phenotype (p < 0.05) and enhanced their secretion of angiogenic factors. Growth of endothelial cells was stimulated directly by TEX and indirectly via macrophage-reprogramming dependent on A2BR signaling (p < 0.01). In vivo, TEX stimulated the formation of defined vascular structures and macrophage infiltration. This response was absent in A2BR-/- rats (p < 0.05). CONCLUSION: This report provides the first evidence for adenosine production by TEX to promote angiogenesis via A2BR. A2BR antagonism emerges as a potential strategy to block TEX-induced angiogenesis.