Wogonin inhibits tumor angiogenesis via degradation of HIF-1α protein

Abstract

Wogonin, a plant-derived flavone, has been shown recently to have antitumor effects. However, the mechanisms that wogonin inhibits tumor angiogenesis are not well known. In this study, we investigated the effects of wogonin on expression of hypoxia-inducible factor-1α (HIF-1α) and secretion of vascular endothelial growth factor (VEGF) in tumor cells. We found that wogonin decreased the expression of HIF-1α by affecting its stability and reduced the secretion of VEGF, which suppressed angiogenesis in cancer. Wogonin promoted the degradation of HIF-1α by increasing its prolyl hydroxylation, which depended on prolyl hydroxylase (PHD) and the von Hippel-Lindau tumor suppressor (VHL). Intriguingly, wogonin impeded the binding between heat-shock protein 90 (Hsp90) and HIF-1α. In addition, wogonin down-regulated the Hsp90 client proteins EGFR, Cdk4 and survivin, but did not affect the level of Hsp90. Wogonin also increased ubiquitination of HIF-1α and promoted its degradation in proteasome. We also found that wogonin could inhibit nuclear translocation of HIF-1α. Electrophoresis mobility shift assay (EMSA) showed that wogonin decreased the binding activity of exogenous consensus DNA oligonucleotide with HIF-1α in nuclear extracts from MCF-7 cells. Chromatin immunoprecipitation (ChIP) assay also revealed that HIF-1α directly binded to endogenous hypoxia-responsive element (HRE) and this binding was significantly decreased in MCF-7 cells treated with wogonin. Preliminary results indicated in vivo activity of wogonin against xenograft-induced angiogenesis in nude mice. Taken together, the results suggested that wogonin was a potent inhibitor of HIF-1α and provided a new insight into the mechanisms of wogonin against cancers.

Keywords: Angiogenesis; CAM; CHX; ChIP; DMOG; Degradation; ELISA; EMSA; HIF-1α; HRE; HUVEC; Hsp90; Hypoxia-inducible factor-1α (HIF-1α); ODDD; PHD; RACK1; VEGF; VHL; Wogonin; activated protein kinase C; chicken chorioallantoic membrane; chromatin immunoprecipitation assay; cycloheximide; dimethyloxalylglycine; electrophoretic mobility shift assays; enzyme linked immunosorbent assay; heat-shock protein 90; human umbilical vein endothelial cell; hypoxia-inducible factor-1α; hypoxic response element; prolyl hydroxylase; the oxygen-dependent degradation domain; the von Hippel–Lindau tumor suppressor; vascular endothelial growth factor.