The GOF Mutant p53 Beacon System

GOF 突变体 p53 信标系统

• 批准号: 9233416
• 负责人: Sumitra Deb
• 金额: $ 22.32万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-24 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9233416
• 关键词: Affect; Alleles; Alpha Cell; Animals; Biochemistry; Cancer Etiology; Cancerous; Cell Count; Cell Line; Cell physiology; Cell surface; Cells; Custom; Darkness; Defect; Development; Elements; Event; Evolution; Fluorescence; Gene Mutation; Genetic; Image; Invaded; Knowledge; Light; Location; Malignant Neoplasms; Monitor; Mus; Mutation; Neoplasm Metastasis; Pathway interactions; Phase; Population; Premalignant; Property; Proteins; Puromycin; Reporter Genes; Resistance; Screening for cancer; Stem cells; Streptavidin; System; Systems Development; TP53 gene; Testing; Therapeutic; Time; Transgenic Mice; cancer cell; cancer type; gain of function; genetic evolution; high throughput screening; improved; inhibitor/antagonist; luminescence; mutant; neoplastic cell; novel; novel therapeutics; promoter; small molecule; small molecule inhibitor; stem-like cell; therapeutic target; tool; tumor; tumor growth; tumor progression; vector

Project Summary We have developed a system in which Gain-Of-Function (GOF) mutant p53 makes living cells fluoresce and luminesce (GOF Mutant p53 Beacon System). This powerful tool allows us to instantly assess the presence or absence of mutant p53 in living cells and is not affected by wild-type p53. It also provides a means to assess activity and properties for a large variety of mutant p53 alleles in cell lines. We propose to test the utility of the Beacon System for assessing small molecule mutant p53 inhibitors and wild-type p53 reactivators. We will adapt the Beacon System for use in high throughput screening of inhibitors such that when mutant p53 is inhibited, the cells will fluoresce and luminesce. The utility of the system for identifying cellular modulators of mutant p53 will also be investigated by adapting the system so that when mutant p53 is inhibited, the cells become resistant to puromycin. The ultimate phase of developing this tool is to make transgenic mice in which cells that acquire p53 mutations fluoresce and luminesce. This advance should allow us to visualize any tumor growth in the living mouse as cells acquire p53 mutations and as the tumor cells grow, evolve, invade, and metastasize and establish glowing cell populations at new locations. The mouse system will also be adapted such that cells that acquire p53 mutations will express a cell surface streptavidin tag making it easy to purify the cells at various stages of cancer development to study cellular properties, stem-cell properties, genetic events, and the evolution of these events at early to late stages of cancer. Development of this system should create opportunities for a simple and rapid screening for cancer therapeutics that target mutant p53, one of the most prevalent cancer-causing dominant defects in all cancers. A mouse in which the tumor glows in the living mouse will provide a powerful real-time analysis of cancer progression. This tool will advance our knowledge of how tumors grow and tumor cells migrate, and how they respond to therapeutics.

项目摘要 我们已经开发了一个系统，在该系统中，功能获得（GOF）突变体p53使活细胞荧光和 Luminesce（GOF突变p53信标系统）。这种强大的工具使我们能够立即评估存在或 活细胞中没有突变体p53，不受野生型p53的影响。它还提供了评估的手段 细胞系中各种突变p53等位基因的活性和特性。我们建议测试 标准系统用于评估小分子突变体p53抑制剂和野生型p53重新激活器。我们将 调整信标系统以用于抑制剂的高吞吐量筛选，以便当突变p53为 抑制，细胞将荧光和发光。系统用于识别细胞调节剂的实用性 突变体p53也将通过调整系统来研究，以便在抑制突变体p53时，细胞 对毒素具有抗药性。开发此工具的最终阶段是使转基因小鼠在其中 获取p53突变荧光和发光的细胞。此进步应该使我们能够可视化任何肿瘤 活着的小鼠的生长随着细胞获得p53突变，并且随着肿瘤细胞的生长，进化，侵袭和 转移并在新位置建立发光的细胞种群。鼠标系统也将适应 这样，获得p53突变的细胞将表达细胞表面链霉亲和素标签，从而易于纯化 癌症发展各个阶段的细胞研究细胞特性，干细胞特性，遗传 事件，以及这些事件在癌症的早期至晚期的演变。 该系统的开发应该为癌症进行简单快速筛查的机会创造机会 靶向突变体p53的治疗剂是所有癌症中最普遍的引起癌症的主要缺陷之一。 肿瘤在活小鼠中发光的小鼠将提供对癌症的强大实时分析 进展。该工具将促进我们对肿瘤如何生长和肿瘤细胞迁移以及它们的了解 回应治疗学。