Carroll Veronica A, Ashcroft Margaret
Cell Growth Regulation and Angiogenesis Laboratory, Cancer Research UK Centre for Cancer Therapeutics, Institute of Cancer Research, Sutton, United Kingdom.
Cancer Res. 2006 Jun 15;66(12):6264-70. doi: 10.1158/0008-5472.CAN-05-2519.
Overexpression of hypoxia-inducible factors (HIF), HIF-1alpha and HIF-2alpha, leads to the up-regulation of genes involved in proliferation, angiogenesis, and glucose metabolism and is associated with tumor progression in several cancers. However, the contribution of HIF-1alpha versus HIF-2alpha to vascular endothelial growth factor (VEGF) expression and other HIF-regulated target genes under different conditions is unclear. To address this, we used small interfering RNA (siRNA) techniques to knockdown HIF-1alpha and/or HIF-2alpha expression in response to hypoxia, insulin-like growth factor (IGF)-I, or renal carcinoma cells expressing constitutively high basal levels of HIF-1alpha and/or HIF-2alpha due to loss of von Hippel-Lindau (VHL) function. We found that HIF-1alpha primarily regulates transcriptional activation of VEGF in response to hypoxia and IGF-I compared with HIF-2alpha in MCF-7 cells. We also observed a reciprocal relationship between HIF-1alpha and HIF-2alpha expression in hypoxia in these cells: HIF-2alpha siRNA enhanced HIF-1alpha-mediated VEGF expression in MCF-7 cells in response to hypoxia, which could be completely blocked by cotransfection with HIF-1alpha siRNA. In contrast, in renal carcinoma cells that constitutively express HIF-1alpha and HIF-2alpha due to loss of VHL function, we found that high basal VEGF, glucose transporter-1, urokinase-type plasminogen activator receptor, and plasminogen activator inhibitor-1 expression was predominantly dependent on HIF-2alpha. Finally, we showed that a newly identified small-molecule inhibitor of HIF-1, NSC-134754, is also able to significantly decrease HIF-2alpha protein expression and HIF-2alpha-regulated VEGF levels in renal carcinoma cells. Our data have important implications for how we target the HIF pathway therapeutically.
缺氧诱导因子（HIF）、HIF-1α和HIF-2α的过表达会导致参与增殖、血管生成和葡萄糖代谢的基因上调，并与多种癌症的肿瘤进展相关。然而，在不同条件下，HIF-1α与HIF-2α对血管内皮生长因子（VEGF）表达及其他HIF调控的靶基因的作用尚不清楚。为解决这一问题，我们运用小分子干扰RNA（siRNA）技术，针对缺氧、胰岛素样生长因子（IGF）-I，或因冯·希佩尔-林道（VHL）功能缺失而组成性高表达基础水平HIF-1α和/或HIF-2α的肾癌细胞，敲低HIF-1α和/或HIF-2α的表达。我们发现，与MCF-7细胞中的HIF-2α相比，HIF-1α主要在缺氧和IGF-I作用下调节VEGF的转录激活。我们还观察到这些细胞在缺氧时HIF-1α和HIF-2α表达之间存在相互关系：HIF-2α的siRNA增强了MCF-7细胞在缺氧时HIF-1α介导的VEGF表达，而与HIF-1α的siRNA共转染可完全阻断这一增强作用。相反，在因VHL功能缺失而组成性表达HIF-1α和HIF-2α的肾癌细胞中，我们发现较高的基础VEGF、葡萄糖转运蛋白-1、尿激酶型纤溶酶原激活物受体和纤溶酶原激活物抑制剂-1表达主要依赖于HIF-2α。最后，我们表明新发现的HIF-1小分子抑制剂NSC-134754也能够显著降低肾癌细胞中HIF-2α蛋白表达及HIF-2α调控的VEGF水平。我们的数据对如何在治疗上靶向HIF通路具有重要意义。
