SPECIFIC INHIBITION OF HER2/NEU TRANSCRIPTION ELONGATION

HER2/NEU 转录延伸的特异性抑制

• 批准号: 6311254
• 负责人: SCOT W EBBINGHAUS
• 金额: $ 8.94万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-01-01 至 2002-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6311254
• 关键词: 3T3 cells; Adenoviridae; DNA binding protein; DNA footprinting; HeLa cells; alkylation; athymic mouse; chemical conjugate; chlorambucil; chromatin; disease /disorder model; gel mobility shift assay; gene expression; neoplasm /cancer genetics; neoplastic cell; nonsmall cell lung cancer; northern blottings; nuclear runoff assay; oligonucleotides; oncogenes; plasmids; transcription factor; transfection /expression vector

DESCRIPTION: The HER2/neu oncogene appears to play an important role in the initiation and progression of many types of human cancer, including approximately 25 percent of non-small cell lung cancer (NSCLC), the leading cause of death in both men and women in the U.S. The overall goal of this project is to find novel ways to specifically inhibit HER2/neu expression by developing oligonucleotide (ODN) - chlorambucil (CHL) conjugates that will bind in a site-specific manner to the HER2/neu gene by triplex DNA formation and lead to DNA alkylation at specific guanine bases by the CHL. Specifically, the work outlined in this application will accomplish the following goals: 1) Characterize the ability of chlorambucil-conjugated beta (natural) and alpha (nuclease resistant modification) anomeric ODNs to diret site-specific DNA alkylation in the HER-2/neu gene; 2) Characterize the ability of chlorambucil-conjugated ODNs to bind to the HER/neu gene and inhibit HER-2/neu gene transcription elongation in vitro and in a cDNA expression plasmid transfected into HeLa cells and NIH3T3 cells; 3) Demonstrate ODN binding and site-specific DNA modification in the chromatin of living NSCLC cells that express the HER-2/neu gene; 4) Characterize the ability of adenoviruses to mediate ODN uptake and nuclear localization in NSCLC cells when adenovirus-ODN complexes are formed with a chemical linker. 5) Determine the ability of optimally delivered ODN-CHL conjugates to inhibit HER-2/neu gene expression and reverse the malignant phenotype in tissue culture and rodent models of human NSCLC. The specific objectives of this application are designed to address the major obstacles to the successful development of an ODN-based site-specific DNA binding drug. The successful completion of these Specific Aims will lead to invaluable insights into the design of gene-specific DNA binding compounds. The development of a HER-2/neu gene specific anti-gene compound will provide a great deal of information about the role of the HER-2/neu gene in the initiation and progression of NSCLC and may lead to novel treatment approaches for HER-2/neu gene expressing cancers such as NSCLC.

描述：HER2/neu Oncogene似乎在 许多类型的人类癌症的启动和发展，包括 大约25％的非小细胞肺癌（NSCLC）是领先的 美国男性和女人的死亡原因总体目标 项目是找到新颖的方法来通过 开发寡核苷酸（ODN） - 将会 通过三个形成的位点特异性方式与HER2/NEU基因结合 并导致CHL在特定的鸟嘌呤碱基处进行DNA烷基化。 具体而言，本应用程序中概述的工作将完成 以下目标：1）表征氯化木偶联的能力 beta（天然）和α（抗核酸酶耐药修饰）的异构ODN HER-2/NEU基因中的Diret位点特异性DNA烷基化； 2）表征 氯化木偶联的ODN与HER/NEU基因结合的能力和 在体外和cDNA中抑制HER-2/NEU基因转录伸长 表达质粒转染到HeLa细胞和NIH3T3细胞中； 3） 在染色质中演示ODN结合和位点特异性DNA修饰 表达HER-2/neu基因的活细胞； 4）表征 腺病毒介导ODN吸收和核定位的能力 用化学接头形成腺病毒-ODN复合物时NSCLC细胞。 5）确定最佳传递的ODN-CHL偶联物的能力 抑制HER-2/NEU基因表达并逆转恶性表型 人NSCLC的组织培养和啮齿动物模型。 的特定目标 该应用程序旨在解决主要的障碍 成功开发基于ODN的位点特异性DNA结合药。 这 这些特定目标的成功完成将导致宝贵 对基因特异性DNA结合化合物的设计的见解。 这 HER-2/NEU基因特异性抗基因化合物的开发将提供 有关HER-2/NEU基因在中的作用的大量信息 NSCLC的启动和进展可能会导致新的治疗 HER-2/NEU基因表达癌症（如NSCLC）的方法。