Structure Function Analysis of the Multi-specific Drug Transporter OCT1

多特异性药物转运蛋白OCT1的结构功能分析

• 批准号: 8814249
• 负责人: SANJAY K NIGAM
• 金额: $ 39.52万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2016-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8814249
• 关键词: ATP-Binding Cassette Transporters; Adult; Affect; Area; Avena sativa; Binding; Biological Assay; Biology; CCL21 gene; Carrier Proteins; Cations; Cells; Chemical Structure; Chemicals; Child; Collaborations; Computer Analysis; Computer Simulation; Crystallization; Crystallography; Data; Data Collection; Detergents; Drug Design; Drug Interactions; Drug Kinetics; Drug Transport; Drug toxicity; Epigenetic Process; Epithelium; Family; Future; Genetic; Genetic Polymorphism; Goals; Health; Homology Modeling; In Vitro; Integral Membrane Protein; Knowledge; Laboratories; Lead; Libraries; Ligands; Lipid Bilayers; Lipids; Mediating; Membrane Proteins; Metabolic; Metformin; Methods; Modeling; Molecular; Neonatal; Neurotransmitters; Oocytes; Organ; Organ Culture Techniques; Organic Anion Transporters; Organic Cation Transporter; Organic Cation Transporter 1; POU2F1 gene; Pathway interactions; Permeability; Pharmaceutical Preparations; Pharmacologic Substance; Phase; Production; Protein Family; Proteins; Publications; Publishing; Reaction; Research Design; Research Personnel; Resolution; Resources; Route; Running; Science; Slice; Structural Protein; Structure; Testing; Tissues; Toxic effect; Toxin; United States National Institutes of Health; Validation; Xenobiotics; base; chemical property; design; high throughput screening; in vivo; in vivo Model; inhibitor/antagonist; innovation; insight; interest; member; molecular dynamics; notch protein; pharmacophore; pressure; protein purification; screening; skills; small molecule; solute; structural biology; supercomputer; synchrotron radiation; three-dimensional modeling; uptake; virtual

DESCRIPTION (provided by applicant): Among the most highly expressed drug transporters in neonatal organs are the organic cation transporters (Octs), which transport drugs, neurotransmitters, metabolites and toxins. Many drug-drug interactions and metabolic derangements due to drugs are thought to occur at the level of the transporter, and certain polymorphisms can lead to drug toxicity. Oct1/SLC22A1 is the prototypical member of the SLC22A family of transporters and along with certain ABC transporters (e.g., MDR), Oats and Oatps, is one of the most important xenobiotic (phase III drug transporter) transporters in the body. The PI's lab identified Organic Anion Transporter 1 (Oat1/NKT/Slc22a6), which, along with Oct1 originally helped define the SLC22 transporter family. We have studied the in vivo function of several SLC22 transporters and performed detailed in vitro analysis and in silico modeling of substrate-transporter interactions. Nevertheless, detailed molecular analyses of substrate-transporter interactions-which are necessary to take a rational approach to diminishing adverse affects of drugs transported by Oct1-are limited by the absence of precise structural information. Here we propose to combine our knowledge of SLC22 transporter biology (and skill with in vitro, ex vivo and in vivo assays) together with the expertise of top-notch investigators in x-ray crystallography, protein and substrate modeling to investigate the molecular mechanisms of the key substrate-transporter interactions involved in Oct1-mediated organic cation xenobiotic handling. Although in this proposal we focus on Oct1, the results will also provide important information relevant to other SLC22 drug transporters. To our knowledge, we are among the very few groups in this field that has a proven ability to combine such diverse areas of expertise, including computationally biology, structural biology and in vivo/in vitro wet-lab biology and is one of the most innovative aspects of the proposed studies. We will crystallize and determine the high-resolution x-ray structure of Oct1 (SA1) in a continuing collaboration with Dr. Geoffrey Chang (involving both his laboratory and TransportPDB, a component of the NIH Protein Structural Initiative of which he is a founding member) to gain key insights into the structural basis of substrate binding and transport. We will use our documented expertise in ligand-based and transporter-based computational approaches (SA2) to identify the molecular determinants of substrates and the Oct1 protein mediating their interactions. Pharmacophore-based virtual screening of chemical structure libraries will be done, and identified structures will be used for molecular dynamic analyses to prioritize compounds. Identified compounds will be obtained and tested in wet-lab studies using in vitro, ex vivo and in vivo approaches well established in the PI's group (SA3). This multifaceted structural- computational-wet-lab strategy will result in major advances in understanding adverse drug reactions in children as well as adults and help move the drug transporter field forward.

描述(申请人提供)：在新生儿器官中表达最高的药物转运体是有机阳离子转运体(OCTS)，它运输药物、神经递质、代谢物和毒素。许多药物与药物的相互作用和药物引起的代谢紊乱被认为发生在转运蛋白的水平上，某些基因多态性可导致药物毒性。Oct1/SLC22A1是SLC22A转运蛋白家族的典型成员，与某些ABC转运蛋白(如MDR)、燕麦和OATP一起，是体内最重要的异源(III期药物转运蛋白)转运蛋白之一。PI的实验室发现了有机阴离子转运蛋白1(Oat1/NKT/Slc22a6)，它最初与Oct1一起帮助定义了SLC22转运蛋白家族。我们研究了几个SLC22转运蛋白的体内功能，并对底物-转运蛋白相互作用进行了详细的体外分析和电子模拟。然而，对底物-转运体相互作用的详细分子分析--采取合理的方法减少10月1日之前运输的药物的不良影响--由于缺乏准确的结构信息而受到限制。在这里，我们建议结合我们对SLC22转运体生物学的知识(以及体外、体外和体内分析的技能)，以及一流研究人员在X射线结晶学、蛋白质和底物建模方面的专业知识，来研究Oct1介导的有机阳离子异种生物处理中关键底物-转运体相互作用的分子机制。虽然在这项提案中我们重点关注10月1日，但结果也将提供与其他SLC22药物转运体相关的重要信息。据我们所知，我们是这一领域极少数被证明有能力将计算生物学、结构生物学和体内/体外湿实验室生物学等不同专业领域结合在一起的小组之一，也是拟议研究的最具创新性的方面之一。我们将与Geoffrey Chang博士(包括他的实验室和TransportPDB，他是NIH蛋白质结构倡议的创始成员之一)继续合作，结晶并确定Oct1(SA1)的高分辨率X射线结构，以获得对底物结合和运输的结构基础的关键见解。我们将使用我们在基于配体和基于转运体的计算方法(SA2)方面的书面专业知识来确定底物和介导它们相互作用的Oct1蛋白的分子决定因素。将对化学结构库进行基于药效团的虚拟筛选，并将识别出的结构用于分子动态分析，以确定化合物的优先顺序。已确定的化合物将在湿实验室研究中获得并测试，方法包括体外、体外和体内方法，这在Pi‘s组(SA3)中得到了很好的证实。这一多方面的结构-计算-湿实验室策略将在了解儿童和成人的不良药物反应方面取得重大进展，并有助于推动药物转运体领域的发展。