Screening Assays for Small Molecules Targeting Oncogenic eIF4E Expression

针对致癌 eIF4E 表达的小分子筛选试验

• 批准号: 8219966
• 负责人: Chunhong Yan
• 金额: $ 29.51万
• 依托单位: ALBANY MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-03-01 至 2015-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8219966
• 关键词: Actins; Address; Animals; Antineoplastic Agents; Antisense Oligonucleotides; Automobile Driving; BCL1 Oncogene; Biological Assay; Bioluminescence; CMV promoter; Cancer Cell Growth; Cell Line; Cell Survival; Cells; Chemicals; Chromatin; Clinical Trials; DNA; Dependovirus; Development; Disadvantaged; Distal; Down-Regulation; Drug Delivery Systems; Emerging Technologies; Environment; Eukaryotic Initiation Factor-4E; Eukaryotic Initiation Factors; Firefly Luciferases; Galactosidase; Gene Expression; Genes; Genetic Transcription; Genomics; Guidelines; Health; Human; Hyperactive behavior; Internal Ribosome Entry Site; Lead; Location; Luc Gene; Luciferases; Malignant Neoplasms; Measurement; Measures; Mediating; Messenger RNA; Methods; NIH Program Announcements; Oncogenic; PI3K/AKT; Pathway interactions; Pharmaceutical Preparations; Preclinical Drug Evaluation; Promoter Regions; Proteins; RNA Cap-Binding Proteins; Regulatory Element; Renilla Luciferases; Reporter; Reporter Genes; Research; Robotics; Screening procedure; Technology; Testing; Therapeutic; Therapeutic Agents; Therapeutic Intervention; Toxic effect; Transcript; Transgenic Organisms; Translations; Untranslated Regions; Vascular Endothelial Growth Factors; assay development; base; c-myc Genes; cancer cell; cancer therapy; cell growth; cellular engineering; cost; drug development; drug discovery; high throughput screening; homologous recombination; human FRAP1 protein; in vivo; inhibitor/antagonist; innovation; luminescence; neglect; novel; overexpression; promoter; recombinase; research clinical testing; response; small molecule; small molecule libraries; success; tumor growth

DESCRIPTION (provided by applicant): The eukaryotic translation initiation factor 4E (eIF4E) is frequently overexpressed in human cancers and contributes to cancer development and progression by selectively driving translation of genes that are essential for cancer cell growth and survival (e.g., c-myc, VEGF, BCL-2). In the past decades accumulating evidence has demonstrated that down-regulation of eIF4E expression in cancer cells can be an effective strategy for therapeutic intervention of cancer. Indeed, eIF4E antisense oligonucleotides have been shown to inhibit tumor growth without toxicity to host animals, and have entered clinical trials. However, because of the lack of a reliable assay that can be used in high-throughput drug screening, no small molecule that can inhibit eIF4E expression in cancer cells is currently available for clinical testing. This application is in response to the NIH Program Announcement PA-10-213, "Development of Assays for High-Throughput Screening for Use in Probe and Pre-therapeutic Discovery", and will address the imperative need of a high-throughput screening (HTS) assay for discovery of drug leads that target eIF4E expression in cancer cells. Specifically, our objective is to utilize emerging cell-engineering technologies to develop an innovative assay in which a bioluminescent reporter integrated in the native eIF4E gene locus is expressed under control of the endogenous eIF4E promoter. Such an assay is expected to overcome limitations of current reporter assays that are mostly based on cloned, transgenic promoters, which often lack essential distal/intronic cis-regulatory elements while residing in foreign chromatin environments. Therefore, the innovative assay would faithfully reproduce responses of the endogenous eIF4E gene to chemical treatments and thus be more reliable in high-throughput search for small molecules inhibitory for eIF4E expression. Accordingly, the specific aims of this project are: (1) to develop cell lines harboring a reporter gene in the native eIF4E gene locus for high-throughput drug screening, (2) to develop secondary assays for identification of false positives and prioritization of screening hits for further drug development. Completion of the proposed research will result in an innovative HTS assay useful for developing therapeutic agents that can cure cancer through inhibiting eIF4E expression. Therefore, the proposed studies will impact health of millions of people afflicted with cancer. This application will also provide a proof of concept for an innovative drug-discovery strategy that can be applied to search for agents targeting any given gene aberrantly expressed in cancer thereby significantly advancing targeted cancer therapy. PUBLIC HEALTH RELEVANCE: The objective of this project is to develop an innovative screening assay for use in discovery of therapeutic agents targeting eIF4E - a protein commonly overexpressed in human cancers. Ultimately, completion of the proposed research will lead to the development of novel cancer therapies, and thereby benefit millions of people afflicted with cancer.

描述（由申请人提供）：真核翻译起始因子4 E（eIF 4 E）经常在人类癌症中过表达，并通过选择性驱动癌细胞生长和存活所必需的基因（例如，c-myc、VEGF、BCL-2）。在过去的几十年中，越来越多的证据表明，下调癌细胞中eIF 4 E的表达可以是癌症治疗干预的有效策略。事实上，eIF 4 E反义寡核苷酸已经显示出抑制肿瘤生长而对宿主动物没有毒性，并且已经进入临床试验。然而，由于缺乏可用于高通量药物筛选的可靠测定，目前没有可抑制癌细胞中eIF 4 E表达的小分子可用于临床测试。该申请是对NIH计划公告PA-10-213“用于探针和治疗前发现的高通量筛选测定的开发”的响应，并将解决用于发现靶向癌细胞中eIF 4 E表达的药物先导物的高通量筛选（HTS）测定的迫切需要。具体而言，我们的目标是利用新兴的细胞工程技术，开发一种创新的检测方法，其中整合在天然eIF 4 E基因位点的生物发光报告基因在内源性eIF 4 E启动子的控制下表达。预期这种测定克服目前报道测定的局限性，所述报道测定主要基于克隆的转基因启动子，所述启动子通常缺乏必需的远端/内含子顺式调节元件，同时驻留在外源染色质环境中。因此，创新的测定将忠实地再现内源性eIF 4 E基因对化学处理的响应，从而在高通量搜索抑制eIF 4 E表达的小分子中更可靠。因此，本项目的具体目标是：（1）开发在天然eIF 4 E基因座中携带报告基因的细胞系，用于高通量药物筛选，（2）开发用于鉴定假阳性和筛选命中优先级的二级测定，用于进一步药物开发。拟议研究的完成将导致一种创新的HTS检测方法，可用于开发通过抑制eIF 4 E表达治愈癌症的治疗药物。因此，拟议的研究将影响数百万癌症患者的健康。该申请还将为创新药物发现策略提供概念证明，该策略可用于搜索靶向癌症中异常表达的任何给定基因的药物，从而显著推进靶向癌症治疗。 公共卫生相关性： 该项目的目标是开发一种创新的筛选方法，用于发现针对eIF 4 E（一种通常在人类癌症中过表达的蛋白质）的治疗药物。最终，完成拟议的研究将导致开发新的癌症疗法，从而使数百万癌症患者受益。