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的生理和药理作用可溶性载体超家族（SLC）中的膜转运蛋白在生理和在细胞摄取和去除内源代谢物和许多处方的药理学毒品。尽管其重要性，但大约五分之一的SLC转运蛋白没有已知的底物，并且是被认为是“孤儿”蛋白。最近，通过应用各种方法，其中包括对全基因组关联信号的计算分析，同源性建模，表达分析以及重要的是稳定表达细胞中的同位素摄取测定 SLC22A24。我们的研究表明，SLC22A24是一种有机阴离子转运蛋白，偏爱硫酸盐和类固醇的糖苷结合物。最近的初步研究表明，转运蛋白也发挥在药物处置中的重要作用。拟议研究的主要目标是确定 Slc22a24在类固醇谷氨酸盐的处置中的生理和药物作用发现其他是SLC22A24底物的内源性代谢物和药物。为了实现我们的目标，我们设计了三个特定目标。在AIM 1中，我们将在细胞系中使用研究确定将SLC22A24定位到基底外侧或顶端膜上。此外，我们会发现处方药和其他代谢物是SLC22A24的底物。在AIM 2中，我们将使用CRISPR/CAS9 在近端管中创建表达SLC22A24的转基因小鼠的方法，并使用小鼠确定它在转运蛋白的药物和生理底物的处置中的作用。最后，在AIM 3中，我们将在健康志愿者中进行关联研究，以将SLC22A24中的功能变体与等离子体相关联类固醇谷化剂的水平和肾脏清除率。采用的方法将包括共聚焦显微镜，细胞中的同位素摄取和通量测定，小鼠CRISPR/CAS9方法，LC/MS/代谢组分析人类的方法，关联分析和清除确定。我们假设这个细胞，小鼠和健康志愿者中的全面基因组，代谢组和功能研究深层临床表型将成为系统评估生理和新脱键转运蛋白的药理作用。