Mechanisms of Genetic Instability and Tumor Suppression

遗传不稳定性和肿瘤抑制机制

• 批准号: 7467347
• 负责人: Geoffrey Myles Wahl
• 金额: $ 50.79万
• 依托单位: SALK INSTITUTE FOR BIOLOGICAL STUDIES
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-07-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7467347
• 关键词: Affect; Binding; Biochemical; Cancer Patient; Cell Nucleus; Cells; Chromatin; Complex; Condition; Cytoplasm; DNA Damage; Enzymes; Feedback; Genetic; Genotype; Goals; Grant; Growth; Human; In Vitro; Kinetics; Laboratories; Malignant Neoplasms; Mediating; Modeling; Molecular; Molecular Genetics; Mus; Mutation; Normal Cell; Numbers; Oncogenes; Oncogenic; Output; Pathway interactions; Protein p53; Proteolysis; Regulation; Research Design; Role; Screening procedure; Small Interfering RNA; Stress; Structure-Activity Relationship; TP53 gene; Testing; Therapeutic; Transcriptional Activation; Transfection; Tumor Suppression; Tumor Suppressor Proteins; attenuation; base; c-myc Genes; cancer cell; cancer therapy; design; improved; in vivo; mouse model; mutant; stoichiometry; tumor; ubiquitin ligase

DESCRIPTION (provided by applicant): The tumor suppressor pathway involving the P53 transcriptional regulator is inactivated in almost all human cancers. In many cancers this is due to attenuation of wild type P53 function by aberrant expression of its negative regulators Hdm2 and HdmX. Mouse models and in vitro transfection studies show that the control of P53 stability and functional output are very sensitive to small changes in the concentrations of its proposed regulatory factors. Therefore, precise quantification of the intracellular concentrations of P53 and its negative regulators including H/Mdm2, H/Mdmx and the deubiquitylating enzyme HAUSP are critical for developing accurate models of P53 regulation. The four Specific Aims of this grant employ quantitative biochemical strategies, mouse molecular genetics, and siRNA functional analyses and screens to elucidate the molecular mechanisms that regulate the P53 tumor suppressor pathway in normal and cancer cells prior to and following stress. Specific Aims 1 and 2 quantify P53 and its regulators in the cytoplasm, nucleus, and on chromatin. The studies are designed to test a new model for P53 regulation in which P53-mediated activation of the H/Mdm2 ubiquitin ligase acts in a positive feedback loop to eliminate H/Mdmx, the primary P53 transcriptional antagonist. Specific Aim 3 uses mouse models to test the positive feedback loop hypothesis, and to analyze structure-function relationships in Mdm2 and MdmX. Specific Aim 4 proposes to examine the mechanisms by which activated oncogenes can inactivate the P53 pathway in the substantial fraction of tumors that express wild type P53. It also utilizes an siRNA screen to identify new regulators that mediate proteasomal degradation of P53, Hdm2 and HdmX. Cancer clearly results from activating mutations in oncogenes that accelerate growth or increase survival, and inactivating mutations that disable tumor suppressors such as P53. However, we now understand that the P53 gene is normal in almost 50% of cancers, and that oncogenic mutations disable P53 function. Thus, activating P53 in such cancers could provide a huge opportunity to improve cancer treatment for a substantial number of cancer patients. Achieving this goal requires understanding how P53 is regulated in normal cells to deduce how it is inactivated in tumors. Only then can we develop therapies to target the genetic defects that disable P53 in a particular tumor. This proposal is designed to provide such information.

描述（由申请人提供）：涉及p53转录调节剂的肿瘤抑制途径在几乎所有人类癌症中都灭活。在许多癌症中，这是由于野生型p53功能的衰减造成的，其阴性调节剂HDM2和HDMX的表达异常。小鼠模型和体外转染研究表明，对p53稳定性和功能输出的控制对其拟议调节因素的浓度的微小变化非常敏感。因此，精确定量p53的细胞内浓度及其负调节剂，包括H/MDM2，H/MDMX和去偶联酶Hausp对于开发p53调节的准确模型至关重要。该赠款的四个特定目的采用定量的生化策略，小鼠分子遗传学以及siRNA功能分析和筛选，以阐明在压力之前和遵循压力之前和之后癌细胞中调节正常和癌细胞中p53肿瘤抑制途径的分子机制。具体目的1和2在细胞质，核和染色质上量化了p53及其调节剂。该研究旨在测试p53调节的新模型，其中p53介导的H/MDM2泛素连接酶的激活作用于阳性反馈回路，以消除原发性p53转录拮抗剂H/MDMX。特定目标3使用小鼠模型来测试正反馈循环假设，并分析MDM2和MDMX中的结构 - 功能关系。具体目标4提出了检查激活的癌基因可以使p53途径在大量表达野生型p53的肿瘤中灭活的机制。它还利用siRNA屏幕来识别介导p53，hdm2和hdmx的蛋白酶体降解的新调节剂。癌症显然是由于激活了加速生长或增加生存的癌基因中的突变而导致的，以及使抑制肿瘤抑制子（如p53）的灭活突变。但是，我们现在了解到，几乎50％的癌症中p53基因是正常的，并且致癌突变禁用了p53功能。因此，在这种癌症中激活p53可以为大量癌症患者提供巨大的机会来改善癌症治疗。实现此目标需要了解正常细胞中p53的调节方式，以推断出肿瘤中的p53。只有这样，我们才能开发疗法来靶向在特定肿瘤中禁用p53的遗传缺陷。该建议旨在提供此类信息。