Targeting Tumor Angiogenesis with G-Quadruplex Binders

描述（申请人提供）：血管内皮生长因子（VEGF）和缺氧诱导因子(HIF)-la在肿瘤血管生成中起关键作用。本研究的总体目标是探索一种新的治疗策略，通过小分子靶向VEGF和HIF-la基因启动子区形成的g -四重体结构，抑制VEGF和HIF-la基因的转录，从而阻止肿瘤血管生成。本课题的具体目的如下：(1)研究肿瘤细胞中VEGF和HIF-la基因的启动子区是否在体内形成g -四重体结构，并能被小分子靶向。(2)确定VEGF和HIF-la启动子区g -四重体结构在这些基因调控中的作用，以及g -四重体相互作用化合物如何调节这些基因的调控。(3)探索g -四重体相互作用药物作为一类抑制VEGF和HIF-la基因转录活性的新型抗血管生成药物的潜在应用。(4)在已知g -四plex相互作用药物的基础上，发现特异性抑制VEGF和HIF-la基因启动子活性介导的转录激活的新型g -四plex化合物或新的先导化合物。VEGF和HIF-la基因启动子区域g -四重体结构的形成将通过体内化学和酶探测技术进行研究。我们将使用突变分析和启动子报告分析来确定在这些基因的启动子区域形成的g -四重体结构的作用。利用多种RT-PCR、微阵列分析、ChIP分析、生物发光成像技术和血管生成实验，我们将评估g -四重体相互作用药物的体外和体内靶向选择性和抗血管生成活性。最后，无细胞生化分析和基于细胞的高通量筛选系统将用于鉴定在细胞培养系统中选择性下调VEGF或HIF-la表达的其他先导化合物。这些研究直接导致了G-四重体相互作用药物在人类肾癌临床前模型中的抗血管生成和抗肿瘤作用的评估。人类肾癌是一种肿瘤类型，其特征是由于von Hippel Lindau （VHL）肿瘤抑制基因的频繁丢失而导致HIF-la和VEGF异常过表达。

DESCRIPTION (provided by applicant): The vascular endothelial growth factor (VEGF) and hypoxia inducible factor (HIF)-la play a pivotal role in tumor angiogenesis. The overall objective of this proposal is to explore a new therapeutic strategy aimed at preventing tumor angiogenesis by repressing the transcription of VEGF and HIF-la genes through targeting G-quadruplex structures formed in the promoter region of these genes with small molecules. The specific aims of this proposal are as follows: (1) To investigate whether G-quadruplex structures are formed in vivo in the promoter regions of VEGF and HIF-la genes in tumor cells and can be targeted with small molecules. (2) To determine the role of G-quadruplex structures in VEGF and HIF-la promoter regions in the regulation of these genes, and how G-quadruplex interactive compounds modulate the regulation of these genes. (3) To explore the potential application of G-quadruplex-interactive agents as a new class of anti-angiogenic agents, which can repress the transcriptional activity of the VEGF and HIF-la genes. (4) To discover new classes of G-quadruplex compounds or new lead compounds based on known classes of G-quadruplex-interactive agents that specifically inhibit the transcriptional activation mediated by the promoter activity of VEGF and HIF-la genes. The formation of G-quadruplex structures in the promoter region of the VEGF and HIF-la genes will be studied by using in vivo chemical and enzymatic probing techniques. We will use mutation analysis and promoter reporter assays to determine the role of G-quadruplex structures formed in the promoter region of these genes. Utilizing multiple RT-PCR, microarray analysis, ChIP assay, bioluminescent imaging techniques, and angiogenesis assays, we will evaluate in vitro and in vivo target selectivity and antiangiogenic activity of G-quadruplex-interactive agents. Lastly, cell free biochemical assays and cell based high-throughput screening system will be utilized to identify other lead compounds that selectively downregulate either VEGF or HIF-la expression in a cell culture system. These studies lead directly to the evaluation of the anti-angiogenic and antitumor effects of G- quadruplex-interactive agents in a preclinical model of human kidney cancer, a tumor type characterized by the abnormal overexpression of HIF-la and VEGF due to the frequent loss of the von Hippel Lindau (VHL) tumor suppressor gene.