Suppression of tumor angiogenesis by metformin treatment via a mechanism linked to targeting of HER2/HIF-1α/VEGF secretion axis

Anti-angiogenesis is currently considered as one of the major antitumor strategies for its protective effects against tumor emergency and later progression. The anti-diabetic drug metformin has been demonstrated to significantly inhibit tumor angiogenesis based on recent studies. However, the mechanism underlying this anti-angiogenic effect still remains an enigma. In this study, we investigated metformin-induced inhibitory effect on tumor angiogenesis in vitro and in vivo. Metformin pretreatment significantly suppressed tumor paracrine signaling-induced angiogenic promotion even in the presence of heregulin (HRG)-beta 1 (a co-activator of HER2) pretreatment of HER2(+) tumor cells. Similar to that of AG825, a specific inhibitor of HER2 phosphorylation, metformin treatment decreased both total and phosphorylation (Tyr 1221/1222) levels of HER2 protein and significantly reduced microvessel density and the amount of Fitc-conjugated Dextran leaking outside the vessel. Furthermore, our results of VEGF-neutralizing and -rescuing tests showed that metformin markedly abrogated HER2 signaling-induced tumor angiogenesis by inhibiting VEGF secretion. Inhibition of HIF-1 alpha signaling by using RNAi or YC-1, a specific inhibitor of HIF-1 alpha synthesis, both completely diminished mRNA level of VEGF and greatly inhibited endothelial cell proliferation promoted by HER2+ tumor cell-conditioned medium in both the absence and presence of HRG-beta 1 pretreatment. Importantly, metformin treatment decreased the number of HIF-1 alpha nucleus positive cells in 4T1 tumors, accompanied by decreased microvessel density. Our data thus provides novel insight into the mechanism underlying the metformin-induced inhibition of tumor angiogenesis and indicates possibilities of HIF-1 alpha-VEGF signaling axis in mediating HER2-induced tumor angiogenesis.