HEPATIC DRUG TRANSPORTERS IN DRUG DISPOSITION

本申请的总体目标是进一步了解 载体介导的过程中的作用，负责摄取 外源性物质从体循环进入肝脏-一个关键的第一步 在其肝脏消除中起作用。许多人的肝脏 最近已经克隆了基底外侧膜转运蛋白， 在功能上以内源性 化合物如胆汁酸。我们克隆了两个新的人类 与大鼠器官序列同源的肝脏药物转运蛋白 阳离子转运蛋白（rOCT 1）和人有机阴离子转运蛋白 （hOATP）。此外，我们还发现了一种突变（Phe 83- > Ser 83）在人肝钠依赖性胆汁酸转运蛋白中， hNTCP，由单个T->C取代产生，其具有显著的 减少运输活动。我们的假设是 转运系统识别临床使用的各种药物， 功能或活动的可变性在以下方面具有重要意义： 药物消除和治疗。 因此，建议进行研究， 通过个体功能表达的特定假设 运输机 具体目标1中的拟议研究将测试 假设我们新克隆的转运蛋白hOATP 2和hNLT， 代表生物化学特征的多特异性胆汁酸 转运蛋白和肝特异性有机阳离子转运蛋白。 我们还将定义突变hNTCP的转运动力学 转运蛋白，以评估该突变是否改变底物亲和力。 此外，将检验hNTCP运输的假设， 在对牛磺熊去氧胆酸的治疗反应中起关键作用 酸（胆汁淤积性肝病）和奥曲肽（癌症 化疗）。具体目标2侧重于定点突变 和同源嵌合体构建方法， 更好地理解和识别赋予运输商 活性和底物特异性。这些研究将提供重要的 关于人类肝脏药物转运系统的见解，并增强 我们对这些过程在药物处置中的作用的理解。

The overall objective of this application is to further our understanding of the role of carrier-mediated processes responsible for the uptake of xenobiotics from the systemic circulation into the liver -a critical first step in their hepatic elimination. A number of human hepatic basolateral membrane transporters have recently been cloned and functionally characterized with respect to the transport of endogenous compounds such as bile acids. We have cloned two novel human hepatic drug transporters with sequence homologies to the rat organic cation transporter (rOCT1), and the human organic anion transporter (hOATP). In addition, we have identified a mutation (Phe83- >Ser83)in the human hepatic sodium-dependent bile acid transporter, hNTCP, resulting from a single T->C substitution, that has a marked reduction in its transport activity. It is our hypothesis that these transport systems recognize a variety of drugs in clinical use, and that variability in function or activity has important implications in terms of drug elimination and therapy. Therefore, studies are proposed to test specific hypothesis through functional expression of individual transporters. The proposed studies in Specific Aim 1 will test the hypothesis that our newly cloned transporters, hOATP2 and hNLT, represent the biochemically characterized multispecific bile acid transporter, and a liver specific organic cation transporter, respectively. We will also define the transport kinetics of the mutant hNTCP transporter to assess whether this mutation alters substrate affinity. Additionally, the hypothesis will be tested that transport by hNTCP plays a pivotal role in the therapeutic response to tauroursodeoxycholic acid (in cholestatic liver disease) and to octreotide (in cancer chemotherapy). Specific Aim 2 focusses on site-directed mutagenesis and homologous chimera construction methods that will be used to better understand and identify key domains conferring transporter activity and substrate specificity. These studies will provide important insights regarding human hepatic drug transport systems, and enhance our understanding of the role of such processes in drug disposition.