SPECIFIC INHIBITION OF HER2/NEU TRANSCRIPTION ELONGATION

HER2/NEU 转录延伸的特异性抑制

• 批准号: 2465347
• 负责人: SCOT W EBBINGHAUS
• 金额: $ 10.08万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 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/2465347
• 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 癌基因似乎在 许多类型人类癌症的发生和进展，包括 约 25% 的非小细胞肺癌 (NSCLC) 是主要的癌症 美国男性和女性的死亡原因 该项目的总体目标 该项目旨在寻找特异性抑制 HER2/neu 表达的新方法 开发寡核苷酸 (ODN) - 苯丁酸氮芥 (CHL) 缀合物， 通过三链体 DNA 形成以位点特异性方式与 HER2/neu 基因结合 并导致 CHL 在特定鸟嘌呤碱基处进行 DNA 烷基化。 具体来说，本申请中概述的工作将完成 以下目标：1) 表征苯丁酸氮芥结合物的能力 β（天然）和 α（核酸酶抗性修饰）异头 ODN HER-2/neu 基因中的直接位点特异性 DNA 烷基化； 2）表征 苯丁酸氮芥结合的 ODN 与 HER/neu 基因结合的能力，以及 在体外和 cDNA 中抑制 HER-2/neu 基因转录延伸 表达质粒转染HeLa细胞和NIH3T3细胞； 3） 展示染色质中的 ODN 结合和位点特异性 DNA 修饰 表达 HER-2/neu 基因的活 NSCLC 细胞； 4）表征 腺病毒介导 ODN 摄取和核定位的能力 当腺病毒-ODN 复合物与化学接头形成时的 NSCLC 细胞。 5) 确定最佳递送的 ODN-CHL 缀合物的能力 抑制 HER-2/neu 基因表达并逆转恶性表型 人类非小细胞肺癌的组织培养和啮齿动物模型。 具体目标 该应用程序旨在解决实现这一目标的主要障碍 成功开发基于ODN的位点特异性DNA结合药物。 这 成功完成这些具体目标将带来宝贵的成果 深入了解基因特异性 DNA 结合化合物的设计。 这 HER-2/neu 基因特异性抗基因化合物的开发将提供 关于 HER-2/neu 基因在 NSCLC 的发生和进展并可能导致新的治疗方法 针对 HER-2/neu 基因表达癌症（例如 NSCLC）的方法。