Deep Phenotyping of Human Knockouts and Population Studies of the APOC3 Pathway

人类基因敲除的深度表型分析和 APOC3 通路的群体研究

• 批准号: 9902507
• 负责人: Daniel James Rader
• 金额: $ 67.9万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9902507
• 关键词: Address; Ankle; Apolipoprotein A-I; Apolipoproteins B; Apolipoproteins C; Atherosclerosis; Biological; Biological Markers; Biology; Catabolism; Cells; Cholesterol; Consanguinity; Coronary heart disease; Data; Disease; Dose; Event; Fatty acid glycerol esters; Functional disorder; Genes; Genetic; Genotype; Glucose; Heparin; Hepatic; Hepatocyte; Heterozygote; High Density Lipoprotein Cholesterol; High Density Lipoproteins; High Prevalence; Homeostasis; Homozygote; Human; Individual; Inflammation; Inflammatory; Insulin; Insulin Resistance; Isotopes; Kinetics; Knock-out; Knowledge; Label; Lipase; Lipids; Lipolysis; Lipoprotein (a); Lipoproteins; Liver Function Tests; Loss of Heterozygosity; Low-Density Lipoproteins; Measures; Mediating; Mediation; Metabolic; Metabolic Clearance Rate; Metabolic Diseases; Metabolic Pathway; Metabolism; Modeling; Mutation; Non-Insulin-Dependent Diabetes Mellitus; OGTT; Oral; Participant; Pathway interactions; Patients; Phenotype; Physiology; Plasma; Population; Population Study; Production; Proteins; Risk; Risk Factors; Role; Safety; Subgroup; Testing; Therapeutic; Thick; Thromboplastin; Tracer; Triglycerides; Uncertainty; Very low density lipoprotein; X-Ray Computed Tomography; apolipoprotein C-III; atherosclerosis risk; cardiometabolism; cardiovascular risk factor; cohort; extracellular; genomic locus; glucose metabolism; heart disease risk; human model; indexing; induced pluripotent stem cell; insight; lipid metabolism; lipoprotein lipase; liver function; loss of function; loss of function mutation; particle; rare variant; recruit; response; stable isotope; therapeutic target; uptake

Several recent studies indicate that loss of function (LoF) mutations in the apolipoprotein C-III (APOC3) gene offer protection from coronary heart disease (CHD) risk, possibly by reducing circulating triglyceride-rich lipoproteins (TRLs). Inhibition of the apoC-III pathway can provide an attractive therapeutic mechanism to lower CHD risk. There are however many outstanding uncertainties that exist, including impact of apoC-III deficiency on (i) lipoprotein kinetics; (ii) high density lipoprotein remodeling and functionality; (iii) non-lipid related pathways; (iv) the mechanisms by which apoC-III inhibition may reduce CHD risk, (v) the relative contribution of different lipoprotein- associated apoC-III levels on CHD; (vi) liver function and insulin resistance from a safety perspective. In this application, we propose studies to address these gaps by leveraging natural human models of apoC-III deficiency. We have identified the world’s first humans with homozygous APOC3 genetic deficiency who are from an isolated Pakistani village with a high prevalence of consanguinity. Recruitment of all inhabitants in this village (~5000 people) has already been completed; genotyping in a subset has already identified 113 APOC3 knockouts with complete apoC-III deficiency providing opportunities for detailed phenotyping. Specifically, in AIM-1, we will conduct deep phenotyping studies in 113 human APOC3 knockouts (n = 113) and in an equal number of heterozygotes and non-carriers to address the following: (i) role of apoC-III deficiency in modulating protein and lipid composition of TRLs to impact the activity of extracellular lipases; (ii) effect of apoC-III deficiency on subclasses of LDL; (iii) role of apoC-III in modulating Lp(a) levels and composition; and (iv) contribution of apoC-III deficiency to modulation of HDL composition and function. We will also address the consequences of genetic APOC3 deficiency on (iv) systemic lipid and glucose metabolism and a range of proteins related to inflammation and other pathways (v) dose-response association with atherosclerosis. In AIM-2, we will conduct lipoprotein kinetic studies using isotope tracers on 18 trios of APOC3 null homozygotes, heterozygotes, and non-carriers to evaluate the impact of apoC-III deficiency on kinetics of apoB- containing lipoproteins and production and clearance of HDL apoA-I. We will also generate human iPS cells from 5 APOC3 knockouts and 5 matched/related non-carriers and differentiate them into hepatocytes to evaluate (a) VLDL apoB and TG production (b) TRL and LDL uptake and (c) SRB1 mediated HDL uptake. In AIM-3, we will measure total plasma apoC-III and lipoprotein-associated (apoB lipoproteins, HDL, and Lp[a]) apoC-III in 5,000 participants from the EPIC-Norfolk cohort (2,500 with incident CHD events). These studies will enable assessment of: (i) disease mediation; (ii) CHD risk progressively adjusted for lipoprotein-associated apoC-III levels and other factors; (iii) dose-response association; (iv) relevance of apoC-III in subgroups (e.g., by T2D); (v) risk prediction beyond traditional CHD risk factors; and (vi) genetic loci associated with apoC-III levels.

最近的几项研究表明，载脂蛋白C-III（APOC 3）基因的功能丧失（LoF）突变提供了 预防冠心病（CHD）风险，可能通过减少循环中富含磷脂酰肌醇的脂蛋白 （TRLs）。抑制apoC-III通路可以提供降低CHD风险的有吸引力的治疗机制。 然而，存在许多突出的不确定性，包括载脂蛋白C-III缺乏对（i） 脂蛋白动力学;（ii）高密度脂蛋白重塑和功能性;（iii）非脂质相关途径;（iv）脂蛋白代谢。 apoC-III抑制可降低CHD风险的机制，（v）不同脂蛋白的相对贡献， 相关的apoC-III水平对CHD的影响;（vi）从安全性的角度来看，肝功能和胰岛素抵抗。 在这个应用中，我们提出了研究，以解决这些差距，利用自然的人类模型， apoC-III缺乏。我们已经确定了世界上第一个纯合子APOC 3基因缺陷的人， 都来自巴基斯坦一个偏僻的村庄，那里有很多近亲。招募所有居民， 一个村庄（约5000人）的基因分型已经完成;一个子集的基因分型已经确定了113个APOC 3 完全apoC-III缺陷的敲除提供了详细表型分析的机会。 具体而言，在AIM-1中，我们将在113例人APOC 3敲除（n = 113）中进行深入的表型研究。 以及在相同数量的杂合子和非携带者中，以解决以下问题：（i）apoC-III缺乏在 调节TRL的蛋白质和脂质组成以影响细胞外脂肪酶的活性;（ii）apoC-III的作用 LDL亚类缺乏;（iii）apoC-III在调节Lp（a）水平和组成中的作用;和（iv） apoC-III缺乏对HDL组成和功能调节的贡献。我们还将讨论 遗传性APOC 3缺乏对（iv）全身脂质和葡萄糖代谢以及一系列蛋白质的影响 与炎症和其他途径有关（v）与动脉粥样硬化的剂量反应相关性。 在AIM-2中，我们将使用同位素示踪剂对18个APOC 3缺失的三联体进行脂蛋白动力学研究。 纯合子、杂合子和非携带者，以评估apoC-III缺乏对apoB- 含有脂蛋白和HDL apoA-I的产生和清除。我们还将从5个细胞中产生人类iPS细胞。 APOC 3敲除和5个匹配/相关的非携带者，并将其分化为肝细胞以评价（a） VLDL apo B和TG产生（B）TRL和LDL摄取和（c）SRB 1介导的HDL摄取。 在AIM-3中，我们将测量总血浆apoC-III和脂蛋白相关（apoB脂蛋白、HDL和 来自EPIC-Norfolk队列的5，000名参与者（2，500名发生CHD事件）中的Lp[a]）apoC-III。这些研究 将能够评估：（i）疾病调解;（ii）CHD风险逐步调整为脂蛋白相关 apoC-III水平和其他因素;（iii）剂量-反应相关性;（iv）亚组中apoC-III的相关性（例如，通过 T2 D）;（v）传统CHD风险因素以外的风险预测;和（vi）与apoC-III水平相关的遗传基因座。