Characterizing the physiological and pharmacological roles of SLC22A24

表征 SLC22A24 的生理和药理作用

基本信息

批准号：
10595085
负责人：
KATHLEEN M GIACOMINI
金额：
$ 46.57万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
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/10595085
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 absorption 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 pharmacologic polarized cell preference response solute stable cell line steroid hormone uptake urinary

项目摘要

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.

表征 SLC22A24 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作用于细胞摄取和去除内源代谢物的药理学和许多处方尽管很重要，但大约五分之一的 SLC 转运蛋白没有已知的底物，并且是已知的。最近，我们的实验室通过对其中一种蛋白质 SLC22A24 进行了去孤儿处理。各种方法，其中包括全基因组关联信号的计算分析，同源建模、表达谱分析，以及重要的是稳定表达的细胞中的同位素摄取测定我们的研究表明，SLC22A24 是一种优先选择硫酸根的有机阴离子转运蛋白。最近的初步研究表明，转运蛋白也发挥着重要作用。拟议研究的主要目标是确定药物处置中的重要作用。 SLC22A24 在类固醇葡萄糖苷酸的处置中的生理和药理作用以及发现作为 SLC22A24 底物的其他内源代谢物和药物制剂。为了实现我们的目标，我们设计了三个具体目标，在目标 1 中，我们将利用细胞系研究来实现。确定 SLC22A24 在基底外侧膜或顶膜上的定位此外，我们还会发现。作为 SLC22A24 底物的处方药和其他代谢药物在目标 2 中，我们将使用 CRISPR/Cas9。方法创建在近曲小管中表达 SLC22A24 的转基因小鼠并使用小鼠来确定它在转运蛋白的药理学和生理底物的处置中的作用最后，在目标 3 中，我们。将在健康志愿者中进行关联研究，将 SLC22A24 的功能变异与血浆关联起来类固醇葡萄糖苷酸的水平和肾清除率采用的方法包括共聚焦显微镜，细胞中的同位素摄取和通量测定、小鼠中的 CRISPR/Cas9 方法、LC/MS/代谢组学分析我们假设这是在人类中进行的方法、关联分析和清除测定。对细胞、小鼠和健康志愿者进行全面的基因组、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估生理和心理的蓝图新分离的转运蛋白的药理作用。