Interactions and Functions of p14ARF in Cancer

p14ARF 在癌症中的相互作用和功能

• 批准号: 6858441
• 负责人: RUTH A GJERSET
• 金额: $ 39.07万
• 依托单位: SIDNEY KIMMEL CANCER CENTER
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-01-01 至 2009-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6858441
• 关键词: SDS polyacrylamide gel electrophoresis; athymic mouse; crosslink; mass spectrometry; neoplasm /cancer genetics; neoplastic transformation; oncoproteins; p53 gene /protein; polymerase chain reaction; prostate neoplasms; protein protein interaction; protein structure function; tumor suppressor genes; western blottings; yeast two hybrid system

DESCRIPTION (provided by applicant): The broad, long-term objective of this study is to establish a rationale for improved therapies for cancer based on pl4ARF (ARF), whose gene is lost or silenced in some 40% of cancers. ARF plays a central role in cancer, attributable in part to its interaction with mdm2 and participation in the p53/mdm2 feedback mechanism, a DNA damage and stress response mechanism that is disrupted in most cancers. Loss of ARF destabilizes p53 as a result of unopposed mdm2-mediated degradation of p53, and impairs p53-mediated growth arrest or apoptosis. Loss of ARF may therefore compromise the outcome of p53-based therapies as well as conventional chemotherapies that use the p53 pathway to trigger apoptosis. In addition to this role in stabilizing p53, there is growing evidence for a broader array of activities and interactions of ARF that are less well understood, but that could be equally important for ARF function. For example, this laboratory has found that ectopic overexpression of ARF contributes to p53 protein accumulation through dual effects on protein synthesis and stabilization, and has observed that ARF can act independently of p53 in human tumor cells. This activity is enhanced by the presence of ARF C-terminal sequences, a region that is dispensable for mdm2 binding. The importance of ARF C-terminal sequences is further underscored by findings that cancer-associated C-terminal mutants of ARF can have altered physical properties and activities. It is therefore likely that important activities of ARF, relevant to the mechanism of cancer and to its treatment, lie outside of the p53/mdm2 feedback loop, and involve interactions with proteins other than mdm2, some of which require the ARF C-terminus. These additional activities and interactions, together with the involvement of ARF in the p53/mdm2 feedback loop, could be particularly relevant to cancers that frequently retain expression of wild-type p53 and may lose or deregulate ARF, such as prostate cancer, which is one focus of this study. These predictions will be tested through four Specific Aims designed to (1) identify protein-protein interactions of ARF or ARF 1b in prostate cancer cell lines and normal prostate epithelial cells, and identify possible changes accompanying cellular transformation using immobilized bacterial fusion proteins to select binding partners, followed by mass spectrometry, (2) elucidate the role of ARF in regulating translation of proteins other than p53, including the requirement for mdm2 and the ARF C-terminus. (3) Establish how ARF and ARF1b, alone and together with p53, contribute to growth suppression and to the therapy response of prostate epithelial cells (before and after cellular transformation), prostate cancer cells, and cancer cells of other origins, and the requirement for the ARF C-terminus, and (4) evaluate in nude mouse models of human and murine prostate cancer the anti-tumor activity of ARF, alone and together with p53 and chemotherapy. Through these studies the project will define the full range of ARF interactions and activities and their possible disruptions following cellular transformation, and will provide a rigorous pre-clinical evaluation in prostate cancer of the therapeutic potential of ARF-based therapies.

描述（由申请人提供）：这项研究的广泛长期目标是建立基于PL4ARF（ARF）改善癌症治疗方法的理由，PL4ARF（ARF）的基因在约40％的癌症中丢失或沉默。 ARF在癌症中起着核心作用，部分归因于其与MDM2的相互作用以及参与p53/MDM2反馈机制，这是大多数癌症中破坏的DNA损伤和应力反应机制。由于MDM2介导的p53的降解，ARF的损失破坏了p53的稳定，并损害了p53介导的生长停滞或凋亡。因此，ARF的丧失可能损害基于p53的疗法的结果以及使用p53途径触发凋亡的常规化学疗法。除了在稳定p53方面的作用外，还有越来越多的证据表明，广泛的ARF活动和相互作用的理解较少，但这对于ARF功能可能同样重要。例如，该实验室发现ARF的异位过表达通过对蛋白质合成和稳定的双重影响有助于p53蛋白的积累，并且观察到ARF可以在人类肿瘤细胞中独立于p53作用。 ARF C末端序列的存在增强了这种活性，ARF C末端序列是可用于MDM2结合的区域。 ARF C末端序列的重要性进一步强调了ARF的癌症相关的C末端突变体可以改变物理性质和活性。因此，与癌症机制及其治疗相关的ARF的重要活动可能位于p53/MDM2反馈回路之外，并且涉及与MDM2以外的蛋白质相互作用，其中一些需要ARF C-terminus。这些其他活动和相互作用，以及ARF参与p53/MDM2反馈回路，可能与经常保留野生型p53表达并可能失去或放松的ARF的癌症特别相关，例如前列腺癌，这是这项研究的重点。这些预测将通过四个特定目的进行测试，旨在（1）在前列腺癌细胞系中ARF或ARF 1B的蛋白质 - 蛋白质相互作用，并确定使用固定的细菌融合蛋白的细胞转化伴随细胞转化的可能变化，以选择结合疗法，然后选择了质量图3的作用。 MDM2和ARF C端的要求。 (3) Establish how ARF and ARF1b, alone and together with p53, contribute to growth suppression and to the therapy response of prostate epithelial cells (before and after cellular transformation), prostate cancer cells, and cancer cells of other origins, and the requirement for the ARF C-terminus, and (4) evaluate in nude mouse models of human and murine prostate cancer the anti-tumor activity of ARF, alone and together with p53和化学疗法。通过这些研究，该项目将定义各种ARF相互作用和活动及其在细胞转化后可能的破坏，并将在前列腺癌中对基于ARF的治疗的治疗潜力进行严格的临床前评估。