Characterizing the physiological and pharmacological roles of SLC22A24

表征 SLC22A24 的生理和药理作用

• 批准号: 10371025
• 负责人: KATHLEEN M GIACOMINI
• 金额: $ 49.96万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10371025
• 关键词: Alleles; Biological; Biological Assay; Biology; Blood; CRISPR/Cas technology; Cell Line; Cell membrane; Cells; Cellular Assay; Computer Analysis; Confocal Microscopy; Drug Kinetics; Drug Prescriptions; Drug Transport; Evaluation; Excision; Exhibits; Expression Profiling; Family; Gene Frequency; Genetic Polymorphism; Genomics; Glucuronides; Goals; Homology Modeling; Hormones; Human; Human Biology; In Vitro; Individual; Isotopes; Kidney; Knock-in; Laboratories; Ligands; Liquid substance; Measures; Membrane Transport Proteins; Methods; Methotrexate; Mus; Nutrient; Organic Anion Transporters; Orphan; Orthologous Gene; Participant; Pharmaceutical Preparations; Pharmacology; Physiological; Physiology; Plasma; Play; Population; Proteins; Proximal Kidney Tubules; Renal clearance function; Research; Role; Signal Transduction; Steroids; Sulfate; Testing; Therapeutic; Transgenic Mice; Variant; Xenobiotics; analytical method; apical membrane; basolateral membrane; clinical phenotype; design; drug disposition; genetic association; genome-wide; healthy volunteer; liquid chromatography mass spectrometry; member; metabolomics; mouse model; novel; polarized cell; preference; response; solute; stable cell line; steroid hormone; uptake; urinary

Characterizing the Physiological and Pharmacological Roles of SLC22A24 Membrane transporters in the solute carrier superfamily (SLC) play critical roles in physiology and pharmacology serving in the cellular uptake and removal of endogenous metabolites and many prescription drugs. In spite of their importance, about one-fifth of SLC transporters have no known substrates and are considered “orphan” proteins. Recently, our laboratory de-orphaned one of these proteins, SLC22A24, by applying various methods, which included computational analyses of genomewide association signals, homology modeling, expression profiling, and importantly, isotopic uptake assays in cells stably expressing SLC22A24. Our studies revealed that SLC22A24 is an organic anion transporter with a preference for sulfate and glucuronide conjugates of steroids. Recent preliminary studies suggest that the transporter also plays an important role in drug disposition. The major goals of the proposed research are to determine the physiological and pharmacological roles of SLC22A24 in the disposition of steroid glucuronides and to discover other endogenous metabolites and pharmacologic agents that are substrates of SLC22A24. To achieve our goals, we have designed three specific aims. In aim 1, we will employ studies in cell lines to determine the localization of SLC22A24 to the basolateral or apical membranes. In addition, we will discover prescription drugs and other metabolites that are substrates of SLC22A24. In aim 2, we will use CRISPR/Cas9 methods to create transgenic mice expressing SLC22A24 in the proximal tubule and use the mice to determine its role in the disposition of pharmacologic and physiologic substrates of the transporter. Finally, in aim 3, we will conduct association studies in healthy volunteers to associate functional variants in SLC22A24 with plasma levels and renal clearance of steroid glucuronides. Methods employed will include confocal microscopy, isotopic uptake and flux assays in cells, CRISPR/Cas9 methods in mice, LC/MS/ metabolomic analytical methods, association analyses and clearance determinations in humans. We postulate that this comprehensive genomic, metabolomic and functional studies in cells, mice and in healthy volunteers including deep clinical phenotyping will serve as a blueprint for systematic evaluation of the physiological and pharmacological roles of a newly de-orphaned transporter.

SLC 22 A24的生理和药理作用 溶质载体超家族（SLC）中的膜转运蛋白在生理学和生物学中起着关键作用， 药理学服务于细胞摄取和去除内源性代谢物， 毒品尽管它们很重要，但大约五分之一的SLC转运蛋白没有已知的底物， 被认为是“孤儿”蛋白质。最近，我们的实验室通过以下方法使这些蛋白质之一SLC 22 A24去孤儿化： 应用各种方法，包括全基因组关联信号的计算分析， 同源性建模、表达谱分析，以及重要的是，稳定表达 SLC22A24.我们的研究表明，SLC 22 A24是一种对硫酸盐有偏好的有机阴离子转运蛋白 和类固醇的葡糖苷酸缀合物。最近的初步研究表明，转运蛋白也起着重要的作用。 药物处置的重要作用。拟议研究的主要目标是确定 SLC 22 A24在类固醇葡萄糖醛酸苷处置中的生理学和药理学作用， 发现作为SLC 22 A24底物的其他内源性代谢物和药理学试剂。 为了实现我们的目标，我们制定了三个具体目标。在目标1中，我们将采用细胞系研究， 确定SLC 22 A24在基底膜或顶膜的定位。此外，我们还将发现 处方药和其他代谢物是SLC 22 A24的底物。在目标2中，我们将使用CRISPR/Cas9 建立在近端小管中表达SLC 22 A24的转基因小鼠的方法，并使用小鼠来确定 其在转运蛋白的药理学和生理学底物的处置中的作用。在目标3中，我们 将在健康志愿者中进行关联研究，以将SLC 22 A24中的功能变体与血浆中的 类固醇葡萄糖醛酸苷的水平和肾脏清除率。所采用的方法将包括共焦显微镜， 细胞中的同位素摄取和通量测定，小鼠中的CRISPR/Cas9方法，LC/MS/代谢组学分析 方法、关联分析和人体清除率测定。我们假设， 在细胞、小鼠和健康志愿者中进行的全面基因组学、代谢组学和功能研究，包括 深入的临床表型分析将作为系统评价生理和 新的去孤儿转运蛋白的药理作用。