MRP2, BSEP, and BCRP Transporter Polymorphisms and Chemotherapy Disposition

简介(由申请人提供)：该计划旨在为首席调查员(PI)提供机会，让其在两位高资历赞助商的直接监督下，获得分子生物学和临床药理学方面的知识和技能，以增强其发展成为一名成功的独立调查员的潜力。这项提案的范围将包括必要的课程作业、研讨会和新技术的实践经验，使PI能够在整个提案的五年内完成一项密集的研究提案。这项建议的总体科学目标是建立MRP2、BSEP和BCRP药物外排转运体功能多态与化疗倾向调节之间的联系。由于转运蛋白是药物处置过程的关键决定因素，因此将利用模型细胞系统中的功能研究来评估转运蛋白的活性并进行药代动力学分析。转运蛋白变体对蛋白质活性有显著的功能影响以及对化疗处置和毒性有重要影响的假说将通过两个特定的目标进行评估：第一，将利用体外重组痘苗病毒介导的HeLa细胞转染试验以及免疫印迹和共聚焦免疫荧光分析来评估总蛋白和细胞表面蛋白的表达，以确定MRP2、BCRP和BSEP变体的功能特征。其次，在特定目标1中确定的功能相关变体将在模型细胞系统中利用精确的功能分析进行研究，包括可诱导的基因表达极化稳定细胞系和内向外的膜小泡，以提供对功能改变机制、动力学分析、底物特异性和化疗介导的抑制的综合评估。药物处置和反应的个体差异是常用处方药的主要问题，但与化疗药物有关，因为疗效受到狭窄的治疗指数的限制。在许多化疗方案中，患者间的显著差异是常见的，显然，遗传因素在决定个人对药物治疗的反应方面发挥着至关重要的作用。通过优化治疗指数和降低与不良反应相关的死亡率和发病率，确定与抗癌药物处置改变相关的药物转运体基因可能会对肿瘤患者使用化疗药物产生重要的治疗后果。

DESCRIPTION (provided by applicant): The proposal seeks the opportunity for the Principal Investigator (PI) to gain knowledge and skills in molecular biology and clinical pharmacology under the direct supervision of two highly qualified sponsors to enhance his potential to develop into a successful independent investigator. The scope of this proposal will incorporate the necessary coursework, seminars, and hands-on experience in new techniques to enable the PI to complete an intensive research proposal over the full five years of the proposal. The overall scientific goal of this proposal is to establish the associations between functional polymorphisms in MRP2, BSEP, and BCRP drug efflux transporters and modulation of chemotherapeutic disposition. Because transporter proteins are critical determinants of the drug disposition process, functional studies in model cell systems will be utilized to evaluate transport activity and perform pharmacokinetic analyses. The hypothesis that transporter variants have significant functional consequences to protein activity and important implications for chemotherapy disposition and toxicity will be evaluated by 2 specific aims: First, MRP2, BCRP, and BSEP variants will be functionally characterized utilizing in vitro recombinant vaccinia-mediated transfection assays in HeLa cells and immunoblot and confocal immunofluoresecent analyses to evaluate total and cell surface protein expression. Second, functionally relevant variants identified in specific aim 1 will be studied in model cell systems utilizing precise functional assays, including inducible gene expression polarized stable cell lines and inside-out membrane vesicles, to provide a comprehensive evaluation of altered mechanisms of function, kinetic analysis, substrate specificity, and chemotherapy-mediated inhibition. Interindividual variation in drug disposition and response is a major concern for commonly prescribed drugs, but has particular relevance with regards to chemotherapeutic agents because efficacy is limited by narrow therapeutic indices. Significant interpatient differences are common with many chemotherapy regimens and it is apparent that genetic factors play vital roles in determining an individual's response to drug therapy. Defining drug transporter genotypes associated with altered disposition of anticancer agents may have significant therapeutic consequences for the use of chemotherapeutic agents in oncology patients by optimizing therapeutic indices and reducing mortality and morbidity associated with adverse effects.