Defining novel pathways that arrest genetically unstable tetraploid cells

定义阻止遗传不稳定四倍体细胞的新途径

• 批准号: 8027587
• 负责人: NEIL J. GANEM
• 金额: $ 14.99万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-16 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8027587
• 关键词: Address; Advisory Committees; Agar; Applications Grants; Area; Award; Biochemical; Bioinformatics; Biological; Biological Assay; Candidate Disease Gene; Cell Cycle; Cell Cycle Arrest; Cell Cycle Progression; Cell Proliferation; Cell division; Cells; Cellular biology; Committee Members; Cytokinesis; Cytoskeleton; DNA Damage; Dana-Farber Cancer Institute; Data; Defect; Development; Environment; Enzymes; Faculty; Failure; Fluorescence Microscopy; Foundations; Future; G1 Arrest; Gene Expression Profiling; Genes; Goals; Growth; Housing; Human; Human Cell Line; In Vitro; Individual; Institution; Lead; MDM2 gene; Malignant Neoplasms; Mass Spectrum Analysis; Medial; Medical; Mentors; Microarray Analysis; Mitosis; Mutate; Nature; Pathway interactions; Pediatric Oncology; Phase; Positioning Attribute; Proliferating; Protein Biochemistry; Proteins; Publications; RNA Interference; Regulation; Research; Research Personnel; Research Training; Resources; Role; Schools; Screening Result; Screening procedure; Signal Pathway; Signal Transduction; Small Interfering RNA; Sorting - Cell Movement; Stress; System; Testing; Tetraploidy; Time; Training; Tumor Suppression; Tumor Suppressor Proteins; Ubiquitin; Work; base; cancer cell; cancer therapy; cancer type; career; experience; feeding; genome wide association study; genome-wide; in vitro activity; in vivo; innovation; interest; meetings; mouse model; novel; overexpression; prevent; programs; research study; response; tumor; tumorigenesis

DESCRIPTION (provided by applicant): Research: Tetraploid cells, which are a common byproduct of cell division failures, are genetically unstable and have the capacity to facilitate tumorigenesis. Consequently, cells possess p53-dependent tumor suppression mechanisms that limit the further proliferation of these cells by promoting a durable G1 cell cycle arrest. However, unlike other common cellular insults that activate p53 and promote G1 arrest, such as the DNA damage response, the mechanisms governing G1 arrest in response to tetraploidy remain largely unexplored. A major goal of this proposal is to uncover how cells 'sense' cellular changes associated with tetraploidy and then relay that information into the p53 pathway to prevent cell cycle progression. To address this fundamental question, I have developed a novel genome-wide RNAi screening assay to comprehensively identify proteins that are necessary to activate or maintain G1 cell cycle arrest after cytokinesis failure and the induction of tetraploidy. Results from this screen, in combination with complementary bioinformatic and biochemical approaches, will illuminate the nature of the stresses associated with tetraploidy: this includes identifying how these stresses are sensed, how they feed into signaling cascades that activate p53, and how they are ultimately overcome by cancer cells. Strong gene hits from this screen will be examined to determine if they are similarly required to activate G1 arrest in response to other common cellular defects, and whether they are commonly lost or mutated in human cancers. Ultimately, the most interesting candidate genes will be tested for tumor suppressive activities in vitro and in vivo. Overall, the aims described in this proposal have strong potential to identify the cellular defects associated with tetraploidy, as well as to uncover novel regulatory mechanisms of p53. Moreover, this work may also lead to the identification of new pathways that can be targeted by chemotherapeutics to reinforce G1 arrest in abnormal cancer cells. Candidate Career Goals: My long-term career goal is to obtain a tenure-track faculty position at a leading academic institution and establish a lab that is at the forefront of unraveling the many mysteries of cell cycle progression. In particular, I have noticed a great divide in the approaches used by cell biologists and cancer biologists to understand both the causes and consequences of cancer: in my lab, I plan on bridging this gap by combining my pre-existing cell biological expertise with the new training opportunities in genome-wide screening, bioinformatics, and transformation assays described in this proposal. Ultimately, my aim is to blend these two fields in order to take a unique multifaceted experimental approach to probe the detailed mechanisms governing cell cycle progression. The K99/R00 award will provide the protected time I need for focused advanced training in order to achieve this career goal. I expect the mentored phase of this proposal, which includes completing the genome-wide screen and characterizing novel pathways that regulate p53, to take 1-2 years and result in at least one high quality publication. The following independent phase of the award will then permit me to further explore the activation and regulation of these signaling pathways, as well as to uncover new tumor suppressor pathways. Together, these data will be used to justify future studies proposed in an R01 grant application that I expect to submit at the beginning of the third year of the independent phase. Environment: The Department of Pediatric Oncology at the Dana-Farber Cancer Institute (DFCI) and Harvard Medial School has an internationally recognized research program that houses a number of expert researchers in the areas of cell proliferation, tumorigenesis, and cancer cell biology. I have assembled a stellar mentoring and advisory committee (Dr. David Pellman, Dr. William Kaelin, Dr. Ron Depinho, Dr. Matt Meyerson, and Dr. William Hahn) that will greatly benefit my research and training experience. I will meet with members of this committee informally as needed for specific scientific or experimental advice, and formally twice a year to discuss the general progress of my project, identify key future directions, and plan for my transition to independence. In addition, the Harvard Medical Area has all of the necessary physical resources required to for me to complete the proposed training and research studies, including facilities for FACS sorting, mass spectrometry, microarray analysis, and genome-wide screening. PUBLIC HEALTH RELEVANCE: Human cells contain specific genes, called tumor suppressors, that are critically important to prevent the development of cancer. This proposal describes an innovative, genome-wide approach to identify novel tumor suppressors. Ultimately, the long-term goal of this work is to determine not only how cancers arise, but also to lay the foundation for the development of new anti-cancer therapies.

描述(申请人提供)：研究：四倍体细胞是细胞分裂失败的常见副产品，遗传不稳定，有促进肿瘤发生的能力。因此，细胞具有p53依赖的肿瘤抑制机制，通过促进持久的G1细胞周期停滞来限制这些细胞的进一步增殖。然而，与其他常见的激活P53并促进G1期停滞的细胞损伤不同，如DNA损伤反应，控制四倍体G1期停滞的机制在很大程度上仍未被探索。这项提议的一个主要目标是揭示细胞如何“感觉”与四倍体相关的细胞变化，然后将这些信息传递到p53途径，以防止细胞周期进展。为了解决这个根本问题，我开发了一种新的全基因组RNAi筛选试验，以全面鉴定在细胞质分裂失败和四倍体诱导后激活或维持G1细胞周期停滞所必需的蛋白质。这一筛选的结果，结合互补的生物信息学和生化方法，将阐明与四倍体相关的压力的性质：这包括确定这些压力是如何被感知的，它们如何进入激活p53的信号级联，以及它们最终是如何被癌细胞克服的。来自这个屏幕的强基因命中将被检查，以确定它们是否同样需要激活G1停滞，以回应其他常见的细胞缺陷，以及它们是否通常在人类癌症中丢失或突变。最终，最有趣的候选基因将在体外和体内进行肿瘤抑制活性测试。总体而言，该提案中描述的目标在识别与四倍体相关的细胞缺陷以及揭示p53的新调控机制方面具有很强的潜力。此外，这项工作还可能导致识别新的通路，这些通路可以作为化疗药物的靶点，以加强异常癌细胞的G1期停滞。 候选人的职业目标：我的长期职业目标是在一家领先的学术机构获得一个终身教职的职位，并建立一个处于揭开细胞周期进程诸多奥秘的前沿的实验室。特别是，我注意到细胞生物学家和癌症生物学家在理解癌症的原因和后果方面使用的方法存在巨大分歧：在我的实验室，我计划通过将我先前存在的细胞生物学专业知识与本提案中描述的全基因组筛查、生物信息学和转化分析方面的新培训机会相结合来弥合这一差距。最终，我的目标是将这两个领域结合起来，以便采取一种独特的、多方面的实验方法来探索控制细胞周期进程的详细机制。K99/R00奖将为我提供实现这一职业目标所需的有重点的高级培训所需的保护时间。我预计这项提案的指导阶段，包括完成全基因组筛选和表征调控p53的新途径，将需要1-2年的时间，并导致至少一篇高质量的文章发表。该奖项接下来的独立阶段将允许我进一步探索这些信号通路的激活和调节，以及发现新的肿瘤抑制通路。总之，这些数据将被用来证明R01拨款申请中提出的未来研究的合理性，我预计该申请将在独立阶段的第三年开始时提交。 环境：Dana-Farber癌症研究所(DFCI)和哈佛医学院的儿科肿瘤学系拥有一个国际公认的研究项目，其中包括细胞增殖、肿瘤发生和癌细胞生物学领域的多名专家研究人员。我已经组建了一个优秀的指导和咨询委员会(David Pellman博士、William Kaelin博士、Ron DePinho博士、Matt Meyerson博士和William Hahn博士)，这将极大地促进我的研究和培训经验。根据需要，我将与这个委员会的成员举行非正式会议，以获得具体的科学或实验建议，并每年正式两次，讨论我的项目的总体进展，确定关键的未来方向，并为我向独立的过渡制定计划。此外，哈佛医学区拥有我完成拟议的培训和研究所需的所有必要物理资源，包括FACS分类、质谱分析、微阵列分析和全基因组筛查的设施。 与公共卫生相关：人类细胞含有被称为肿瘤抑制因子的特定基因，这些基因对预防癌症的发展至关重要。这项提案描述了一种创新的、全基因组的方法来识别新的肿瘤抑制因子。归根结底，这项工作的长期目标不仅是确定癌症是如何产生的，而且还为开发新的抗癌疗法奠定基础。