Using transcriptomics and ex vivo organotypic models to discover mechanisms of APOL1-associated podocytopathies

使用转录组学和离体器官型模型来发现 APOL1 相关足细胞病的机制

• 批准号: 10590895
• 负责人: Agustin Gonzalez-Vicente
• 金额: $ 13.22万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-18 至 2027-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10590895
• 关键词: APOL1 gene; Address; African American; African American population; African ancestry; Alleles; Architecture; Area; Bioinformatics; Biological Assay; Biology; Black American; CRISPR/Cas technology; Cell Line; Cell physiology; Chemicals; Chronic Kidney Failure; Classification; Clinical; Complex; Cox Proportional Hazards Models; Data; Development; Disease; Early Diagnosis; Experimental Models; Gene Expression; Gene Expression Profile; Genes; Genetic; Genetic Transcription; Genetic Variation; Genotype; Goals; Health; Human; Individual; Investigation; Kidney; Kidney Diseases; Laboratories; Maps; Measures; Mentors; Mentorship; Modeling; Molecular; Mutation; Names; Nephrology; Nephrotic Syndrome; Outcome; Participant; Pathogenicity; Pathway Analysis; Pathway interactions; Patients; Persons; Phenotype; Population; Primates; Principal Investigator; Production; Prognosis; Prognostic Marker; Research; Research Personnel; Research Proposals; Reverse Transcription; Risk; Rodent; Science; Specialized Epithelial Cell; Stress; Technology; Testing; Therapeutic; Training; Transgenic Mice; Translating; Variant; career development; cell type; cohort; design; differential expression; genetic association; genetic variant; high risk; high risk population; in silico; in vitro testing; inhibitor; insight; laboratory experience; large datasets; mouse model; novel; novel therapeutics; patient oriented research; podocyte; preservation; programs; risk variant; screening; skills; social health determinants; stable cell line; stem cells; therapeutic target; tool; transcriptome; transcriptome sequencing; transcriptomics; translational scientist

ABSTRACT Chronic Kidney Disease (CKD) is common among African American (AA) patients. The excess risk for CKD in this population is partially explained by genetic variations in the APOL1 gene (named G1 and G2) that are unique to African ancestral populations. Understanding the molecular basis for the association between genetic variants and the risk for kidney disease is an important goal in biomedical science. The APOL1 gene is unique to humans and a few primates creating limitations to research approaches using conventionally available experimental models. This career development research proposal is designed to train a promising early stage investigator to address these issues by using organotypic kidney models derived from human inducible stem cells (iPSC) in conjunction with state-of-the-art bioinformatics to interrogate human patient data and explore therapeutics. The current proposal is designed to test the hypothesis that APOL1 kidney disease risk variants drive transcriptional differences that can be identified by the integration of glomerular transcriptomes from people with APOL1-associated kidney disease and ex vivo models, to gain insight into APOL1 function in health and disease. Key preliminary data developed by the applicant demonstrates that a variant-dependent APOL1 transcriptional signatures identified in human glomeruli are conserved in mouse models expressing APOL1 variants. The applicant will acquire new skills in two general areas: 1) expertise in the use of novel ex vivo culture models for mechanistic studies, and 2) computational and bioinformatics analysis of large datasets. These skills will be acquired through mentorship, didactics and a pragmatic research program divided into three specific aims: 1) Define APOL1 risk genotype-associated transcriptional phenotypes in podocytes derived from isogenic cell lines homozygous for the G0, G1 and G2 alleles that are anchored to human glomerular gene expression; 2) Anchor APOL1 variants-associated podocyte transcriptional phenotypes to NEPTUNE glomerular gene co-expression modules and clinical outcomes, and identity compounds that could modulate such phenotypes and associated subcellular processes, and 3) Test the chemical perturbagens’ ability to reverse the APOL1 risk genotype transcriptional phenotype and identify kidney disease mechanisms. Completion of this proposal will provide insights into APOL1 biology and function that could be translated to clinical technology by providing early diagnosis and prognosis tools, identification of therapeutic targets and patient derived renal models. Additionally this proposal aims to establish the Principal Investigator (PI) as an independent translational scientist in nephrology. The plan includes mentored training in patient oriented research, extensive wet-lab training in the production of organotypic models from iPSC lines as well as comprehensive coursework in bioinformatics.

摘要 慢性肾脏病(CKD)在非裔美国人(AA)患者中很常见。慢性肾脏病的超额风险 这一群体的部分原因是APOL1基因(命名为G1和G2)的遗传变异是独特的 非洲祖先的人口。了解遗传变异之间关联的分子基础 肾脏疾病的风险是生物医学科学的一个重要目标。APOL1基因是人类独有的 一些灵长类动物使用传统的实验方法对研究方法造成限制 模特们。这份职业发展研究提案旨在培养有前途的早期调查员，以 通过使用源自人类诱导干细胞(IPSC)的器官型肾脏模型来解决这些问题 与最先进的生物信息学相结合，询问人类患者数据并探索治疗方法。这个 目前的提案旨在测试APOL1肾病风险变量推动 通过肾小球转录本的整合可以识别转录差异 APOL1相关肾脏疾病患者和体外模型，以了解APOL1的功能 在健康和疾病方面。申请人开发的关键初步数据表明，依赖于变种的 在人肾小球中发现的APOL1转录信号在小鼠模型中是保守的 APOL1变异体。申请者将在两个一般领域获得新的技能：1)使用新奇经验的专业知识 用于机械学研究的活体培养模型，以及2)大数据集的计算和生物信息学分析。 这些技能将通过指导、教学和分成三个部分的实用研究计划获得 具体目标：1)确定来源于以下来源的足细胞中的APOL1风险基因相关转录表型 定位于人肾小球基因的G0、G1和G2等位基因纯合子等基因细胞系 表达；2)锚定APOL1变异体相关足细胞转录表型与海王星肾小球 基因共表达模块和临床结果，以及可能调节这种结果的识别化合物 表型和相关的亚细胞过程，以及3)测试化学扰动剂逆转 APOL1风险基因型转录表型和确定肾脏疾病的机制。完成这项工作 提案将提供对APOL1生物学和功能的见解，这些见解可以通过以下方式转化为临床技术 提供早期诊断和预后工具，确定治疗目标和患者来源的肾脏 模特们。此外，这项提议旨在将首席调查员(PI)设立为独立的翻译 肾脏病科学家。该计划包括以患者为中心的研究指导培训、广泛的湿法实验室 从IPSC生产线生产有机模型的培训以及综合课程工作 生物信息学。