In vitro study of normoxic epidermal growth factor receptor-induced hypoxia-inducible factor-1-alpha, vascular endothelial growth factor, and BNIP3 expression in head and neck squamous cell carcinoma cell lines: Implications for anti-epidermal growth factor receptor therapy

BackgroundWe previously showed that activation of epidermal growth factor receptor (EGFR) induces hypoxia inducible factor-1 (HIF-1) in head and neck squamous cell carcinoma (HNSCC) cells. In this study, we have furthered this by investigating the mechanism of HIF-1 activation by epidermal growth factor (EGF) and its association with the sensitivity to gefitinib. MethodsEGFR/HIF-1 signaling was tested by immunoblot, polymerase chain reaction (PCR), cell proliferation, and apoptosis assays. ResultsHIF-1 accumulated in cells overexpressing EGF and phosphorylated epidermal growth factor receptor (pEGFR), phosphatidylinositol-3-kinase (pPI3K), and mitogen-activated protein kinase (pMAPK). EGF-induced expression of HIF-1 and its targets, vascular endothelial growth factor (VEGF) and BNIP3, were blocked by gefitinib and PI3K-inhibitors and MAPK-inhibitors. HIF-1-siRNAs abrogated EGF-induced BNIP3 but not VEGF expression. Gefitinib inhibited cell proliferation and induced apoptosis more strongly in cells with constitutively active EGFR/HIF-1 signaling than in cells lacking activation of these pathways. HIF-1-siRNA treatment reduced sensitivity to gefitinib. ConclusionThe search for molecular predictors of sensitivity to gefitinib in HNSCC should be extended to the activation status of EGFR-downstream pathways, phosphorylated protein kinase B, pMAPK, and HIF-1. (c) 2014 Wiley Periodicals, Inc. Head Neck37: 1150-1162, 2015