Regulation of VEGF in Tumors by Ras, EGF and PTEN

Ras、EGF 和 PTEN 对肿瘤中 VEGF 的调节

• 批准号: 6760221
• 负责人: Amit Maity
• 金额: $ 26.35万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6760221
• 关键词: SDS polyacrylamide gel electrophoresis; angiogenesis; biological signal transduction; cell line; enzyme activity; epidermal growth factor; gene expression; gene mutation; genetic regulation; growth factor receptors; guanine nucleotide binding protein; hypoxia; hypoxia inducible factor 1; messenger RNA; neoplastic process; northern blottings; oncogenes; phosphatidylinositol 3 kinase; point mutation; protein kinase; site directed mutagenesis; transfection /expression vector; tumor suppressor proteins; vascular endothelial growth factors; western blottings

DESCRIPTION (provided by applicant): Vascular endothelial growth factor (VEGF), a key mediator of angiogenesis, is often overexpressed in cancers. In many in vivo models, inhibition of VBGF function arrests tumor growth. While hypoxia has long been recognized to be a potent inducer of VEGF, VEGF can also be expressed in normoxia. Recent evidence indicates that angiogenesis can develop in tumor masses before they have grown to a size large enough to contain hypoxic regions, suggesting that angiogenic factors may be expressed by these tumors under normoxic conditions. In contrast to the induction of VEGF rnRNA under hypoxia whose mechanism is known to involve the hypoxia-inducible factor-1 (HIF-1), the upregulation of VEGF in normoxia is much less well understood. The overall aim of this grant is to study mechanisms of VEGF upregulation in normoxia by alterations commonly found in cancers: specifically, epidermal growth factor receptor (EGFR) activation and mutations in Ras and PTEN. VEGF mRNA levels and promoter activity in U87 human glioblastoma cells are increased by EGFR stimulation. Introduction of wild type PTEN into U87 cells, in which PTEN is inactivated, decreases VEGF mRNA levels and promoter activity. Specific Aim 1 will focus on defining the elements in this pathway, which appears to be PI(3) kinase dependent but independent of HIF- 1. H-ras transformation of Rat 1 fibroblasts leads to a six-fold increase in VEGF mRNA expression in normoxia. Furthermore, the level of HIF-1alpha protein is increased in normoxic Rat1-ras cells, an unexpected finding given that HIF-1alpha has traditionally been thought to only be induce under hypoxic conditions. The focus of Specific Aim 2 is to determine whether this increase in HIF- 1 alpha causes the increase in VEGF expression under normoxia. Aim 2 will also examine the signaling pathways that link Ras activation, HIF-1alpha and VEGF expression. These experiments will lead to a better understanding of HIF-1 alpha regulation, mechanisms of VEGF overexpression in cancers, and the effects of EGFR activation and Ras and PTEN mutations on gene expression.

描述(申请人提供)：血管内皮生长因子(血管内皮生长因子)是血管生成的关键介质，在癌症中经常过度表达。在许多体内模型中，抑制VBGF功能可以阻止肿瘤生长。虽然低氧一直被认为是血管内皮生长因子的有效诱因，但在常氧条件下也可以表达血管内皮生长因子。最近的证据表明，在肿瘤生长到足以容纳缺氧区之前，肿瘤中的血管生成就可以发展起来，这表明血管生成因子可能在常氧条件下由这些肿瘤表达。与已知的涉及缺氧诱导因子-1(HIF-1)的低氧诱导血管内皮生长因子rnRNA的机制不同，人们对常氧条件下血管内皮生长因子的上调知之甚少。这项拨款的总体目的是研究正常氧条件下通过癌症中常见的变化上调血管内皮生长因子的机制：具体地说，表皮生长因子受体(EGFR)的激活以及RAS和PTEN的突变。经EGFR刺激后，U87人脑胶质母细胞瘤细胞中的血管内皮生长因子基因表达水平和启动子活性均升高。将野生型PTEN导入失活PTEN的U87细胞，可降低VEGFmRNA水平和启动子活性。具体目标1将集中于确定该通路中的元件，该通路似乎是PI(3)激酶依赖的，但不依赖于HIF-1。H-ras转化大鼠1成纤维细胞后，在常氧条件下，VEGF mRNA的表达增加了6倍。此外，在正常缺氧的Rat1-ras细胞中，HIF-1α蛋白水平增加，这是一个意想不到的发现，因为传统上认为HIF-1α只有在低氧条件下才能诱导。具体目的2的重点是确定HIF-1α的这种增加是否导致常氧下血管内皮细胞生长因子表达的增加。AIM 2还将研究RAS激活、HIF-1α和VEGF表达之间的信号通路。这些实验将有助于更好地了解HIF-1α的调控，肿瘤中VEGF过表达的机制，以及EGFR激活和RAS和PTEN突变对基因表达的影响。