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

基本信息

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
项目类别：
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

项目摘要

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 (APOC3) 基因中的功能丧失 (LoF) 突变会导致可能通过减少循环中富含甘油三酯的脂蛋白来预防冠心病 (CHD) 风险 (TRL) 抑制 apoC-III 通路可以提供一种有吸引力的治疗机制来降低 CHD 风险。然而，存在许多突出的不确定性，包括 apoC-III 缺陷对 (i) 的影响脂蛋白动力学；(ii) 高密度脂蛋白重塑和功能；(iii) 非脂质相关途径； 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apoC-III 水平与CHD 的关系；(vi) 从安全角度看肝功能和胰岛素抵抗。在此应用中，我们提出研究通过利用自然人类模型来解决这些差距我们发现了世界上第一例患有纯合 APOC3 遗传缺陷的人类。来自巴基斯坦一个偏僻的村庄，这里的所有居民都是血缘关系。村庄（约 5000 人）已完成；子集中的基因分型已鉴定出 113 个 APOC3 完全 apoC-III 缺陷的基因敲除为详细表型分析提供了机会。具体来说，在 AIM-1 中，我们将对 113 个人类 APOC3 敲除（n = 113）进行深度表型研究并在相同数量的杂合子和非携带者中解决以下问题：(i) apoC-III 缺陷在调节 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；以及 (iv) 我们还将讨论 apoC-III 缺陷对 HDL 组成和功能调节的影响。遗传性 APOC3 缺陷对 (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。在 AIM-2 中，我们将使用同位素示踪剂对 18 个 APOC3 null 三重奏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 缺陷对 apoB- 动力学的影响含有脂蛋白以及 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。 APOC3 敲除和 5 个匹配/相关非携带者并将其分化为肝细胞以进行评估 (a) VLDL apoB 和 TG 产生 (b) TRL 和 LDL 摄取以及 (c) SRB1 介导的 HDL 摄取。在 AIM-3 中，我们将测量总血浆 apoC-III 和脂蛋白相关蛋白（apoB 脂蛋白、HDL 和 Lp[a]) apoC-III 对来自 EPIC-Norfolk 队列的 5,000 名参与者（其中 2,500 名患有冠心病事件）进行了这些研究。将能够评估：(i) 疾病调节；(ii) 逐步调整脂蛋白相关的冠心病风险 apoC-III 水平和其他因素；(iii) 剂量反应关联；(iv) apoC-III 在亚组中的相关性（例如，通过 (v) 传统冠心病风险因素之外的风险预测；以及 (vi) 与 apoC-III 水平相关的基因位点。