Yeast Assay for p53 Molecular Analysis in Bladder Cancer

膀胱癌中 p53 分子分析的酵母测定

• 批准号: 6878544
• 负责人: Ralph W. deVere White
• 金额: $ 13.33万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2006-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6878544
• 关键词: bladder neoplasm; cancer registry /resource; clinical research; diagnosis quality /standard; diagnostic tests; gene expression; gene mutation; genetic markers; human tissue; metastasis; molecular oncology; neoplasm /cancer diagnosis; neoplasm /cancer genetics; neoplastic process; nucleic acid quantitation /detection; p53 gene /protein; patient oriented research; prognosis; technology /technique development; tissue /cell preparation; tissue resource /registry; transitional cell carcinoma; yeasts

DESCRIPTION (provided by applicant): Bladder cancer is the fifth most frequently diagnosed tumor in the United States, with more than 57,400 new cases and 12,500 deaths annually. Eighty-five percent of these deaths come from the 25% of patients who present with muscle-invasive disease. Reduction in morbidity and mortality will come from improving outcome in this group of patients. Having rapid access to molecular analysis data and knowing its relationship to clinical outcome could be invaluable to patient management and treatment. The long-term objectives of this R21/R33 application are to 1) standardize the protocol for an archival-based yeast functional assay (aYFA) that will lead to broad clinical utilization, and 2) determine what clinical relevance molecular analysis may provide when detecting and evaluating p53 functionality in bladder cancer. A yeast functional assay is an existing methodology that detects p53 mutations in human cancers. In this assay, both p53 expression plasmids and reporter plasmids are co-transfected into yeast. The expressed wild-type (wt) p53 is able to bind to a p53-response element, whereas mutant p53 lacks the ability to bind. This binding and lack of binding leads to color differences in transactivational activity. Use of this current version of the yeast assay has one serious drawback: it was designed for the study of mRNA from cell lines and fresh-frozen tissue. Unfortunately, in large clinical trials, fresh-frozen tissue is rarely available for comparing genetic markers to patient outcome. Therefore, an archival-based yeast functional assay could prove to be extremely suitable for rapid, accurate, and treatment-relevant molecular analysis in a wider bladder cancer population. The specific aim of the R21 phase of this grant is to optimize the technology and methodology for a broader utilization of the archival-based yeast functional assay in the clinical setting. The specific aim of the R33 phase of this grant is to determine the clinical relevance of different p53 mutations (alterations and functionality) in bladder cancer. It is expected that the archival-based yeast functional assay technology will impact the molecular analysis of bladder cancer by proving to be both rapid and clinically relevant. The archival-based yeast functional assay technology is innovative in that it would give clinicians, regardless of location, the ability to obtain molecular analysis information that could prove valuable to patient outcome prior to actual treatment initiation. Currently, with fresh-frozen tissue required to perform the standard yeast assay, timely accessibility of individual patient molecular data for the clinician is virtually impossible to obtain. This makes the standard yeast assay less than optimal for widespread use and subsequently hampers future research endeavors. This grant proposes to compare p53 molecular analysis data and its relationship to patient clinical outcomes in bladder cancer patients. Three groups of patients that have already been reported on will form the basis of the R33 pilot study. The archival-based yeast functional assay technology will be used in testing bladder cancer specimens (cystectomy and TURBT) and then comparing their p53 status to patient clinical outcomes. The results obtained during the period of this grant will form the basis for future studies that should affect treatment strategies for this disease.

描述（由申请人提供）：膀胱癌是美国第五大最常诊断的肿瘤，每年有超过57，400例新发病例和12，500例死亡。其中85%的死亡来自25%的肌肉浸润性疾病患者。发病率和死亡率的降低将来自于这组患者结局的改善。快速获得分子分析数据并了解其与临床结果的关系对于患者管理和治疗可能是非常宝贵的。 这项R21/R33申请的长期目标是：1）标准化基于存档的酵母功能测定（aYFA）的方案，这将导致广泛的临床应用，2）确定在检测和评价膀胱癌中的p53功能时，分子分析可以提供什么样的临床相关性。 酵母功能测定是检测人类癌症中p53突变的现有方法。在该测定中，将p53表达质粒和报告质粒共转染到酵母中。表达的野生型（wt）p53能够结合p53-反应元件，而突变型p53缺乏结合的能力。这种结合和缺乏结合导致反式激活活性的颜色差异。目前使用的酵母检测方法有一个严重的缺点：它是为研究来自细胞系和新鲜冷冻组织的mRNA而设计的。不幸的是，在大型临床试验中，新鲜冷冻的组织很少用于比较遗传标记与患者结果。因此，基于存档的酵母功能测定可以证明非常适合在更广泛的膀胱癌人群中进行快速，准确和治疗相关的分子分析。 该资助的R21阶段的具体目标是优化技术和方法，以便在临床环境中更广泛地利用基于档案的酵母功能测定。R33阶段的具体目标是确定膀胱癌中不同p53突变（改变和功能）的临床相关性。预计基于存档的酵母功能测定技术将通过证明快速和临床相关性来影响膀胱癌的分子分析。基于档案的酵母功能测定技术是创新的，因为它将使临床医生，无论位置，获得分子分析信息的能力，可以证明有价值的病人的结果之前，实际的治疗开始。目前，由于需要新鲜冷冻的组织来进行标准酵母测定，临床医生几乎不可能及时获得个体患者的分子数据。这使得标准酵母测定法对于广泛使用来说不是最佳的，并且随后阻碍了未来的研究努力。 该基金旨在比较p53分子分析数据及其与膀胱癌患者临床结局的关系。已经报告的三组患者将构成R33试点研究的基础。基于存档的酵母功能检测技术将用于检测膀胱癌标本（cysteinase和TURBT），然后将其p53状态与患者临床结局进行比较。在该补助金期间获得的结果将构成未来研究的基础，这些研究将影响这种疾病的治疗策略。