Protein Knockout Technology/Molecular Analysis of Cancer

描述：(申请人提供) 泛素-蛋白酶体途径是细胞内主要的蛋白分解机制。 选择性地以细胞蛋白质为目标进行降解。蛋白质敲除， 我们最近开发的一项新技术，利用了 泛素蛋白水解酶系统指导降解，否则稳定 细胞蛋白质。这项研究计划的长期目标是将 蛋白质敲除系统作为一种快速分子分析工具来破译 细胞癌蛋白的功能及其作为药物的潜在用途 目标。此应用程序的目标是提高效率和 蛋白质敲除系统的特异性和评估其在 蛋白水解法去除已建立细胞中过表达的c-myc癌蛋白 白血病发生的培养和动物模型。理由是 工程底物对特定细胞蛋白的识别作用 泛素化机制的受体允许它们通过 泛素-蛋白酶体途径。研究计划是在以下方面制定的 基于强大的初步数据和许多实验室最近的研究 包括我们自己的。我们以独特的方式准备承担建议的 研究，因为我们有强有力的初步数据证明了 蛋白质敲除技术在稳定剂选择性降解中的应用 细胞蛋白质。申请的目标将通过以下方式实现 追求以下具体目标：(1)R21阶段：改善 蛋白质敲除系统的效率、特异性和递送。(2)R33 阶段：评价c-myc靶向降解在抑制致癌作用中的作用 已建立的细胞培养系统中的转化。(3)R33阶段：至 通过蛋白敲除对抗myc介导的小鼠肿瘤发生 白血病的发生。拟议的工作是创新的，因为它利用了 我们小组最近确定了泛素化机制的性质 还有其他人。我们期望这种方法将提供一个 在蛋白水平下调c-myc癌蛋白的有效手段 抑制c-myc介导的肿瘤转化。这些结果是 意义重大，因为预计它们将提供全面的评估 将蛋白质敲除技术作为一种简单且经济有效的分子 阐明癌症相关蛋白和基因功能的分析工具 途径，并验证它们作为治疗靶点的潜在用途 干预。

DESCRIPTION: (provided by applicant) The ubiquitin-proteasome pathway is a major cellular proteolysis machinery that selectively targets cellular proteins for degradation. Protein knockout, a new technology we recently developed, harnesses the specificity of the ubiquitin proteolytic system to direct the degradation of otherwise stable cellular proteins. The long-range goal of this research plan is to apply the protein knockout system as a rapid molecular analysis tool to decipher the function of cellular oncoproteins and to validate their potential use as drug targets. The objective of this application is to improve the efficiency and specificity of the protein knockout system and to assess its efficacy in the proteolytic removal of the overexpressed c-myc oncoprotein in established cell culture and animal models for leukemogenesis. The rationale is that recognition of specific cellular proteins by the engineered substrate receptors of the ubiquitination machinery allows for their degradation by the ubiquitin-proteasome pathway. The research plan has been formulated on the basis of strong preliminary data, and on recent studies by many laboratories including our own. We are uniquely prepared to undertake the proposed research, because we have strong preliminary data demonstrating the efficacy of the protein knockout technology in the selective degradation of stable cellular proteins. The objective of the application will be accomplished by pursuing the following specific aims: (1) R21 phase: To improve the efficiency, specificity and delivery of the protein knockout system. (2) R33 phase: To evaluate targeted c-myc degradation in the inhibition of oncogenic transformation in established cell culture systems. (3) R33 phase: To antagonize myc-mediated tumorigenecity by protein knockout in mouse models for leukemogenesis. The proposed work is innovative, because it capitalizes on the recently identified properties of the ubiquitination machinery by our group and by others. It is our expectation that this approach will offer an efficient means to downregulate the c-myc oncoprotein at the protein level and to inhibit c-myc-mediated neoplastic transformation. These results are significant, because they are expected to provide a comprehensive evaluation of the protein knockout technology as a simple and cost effective molecular analysis tool to elucidate the function of cancer-related proteins and genetic pathways, and to validate their potential use as targets for therapeutic intervention.