Functional Analysis of Triglyceride Regulator ApoA-V Using Natural Variants

使用天然变体进行甘油三酯调节剂 ApoA-V 的功能分析

• 批准号: 10245120
• 负责人: Sylvia Georgieva Stankov
• 金额: $ 4.6万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-16 至 2022-09-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10245120
• 关键词: APOA5 gene; Alanine Transaminase; Apolipoproteins; Apolipoproteins A; Apolipoproteins C; Arterial Fatty Streak; Atherosclerosis; Biochemical; Biomedical Research; Blood Vessels; Cause of Death; Cell surface; Cholesterol; Code; Coronary Arteriosclerosis; Development; Etiology; Event; Exhibits; Extrahepatic; Family; Fatty acid glycerol esters; Frequencies; Genes; Genetic study; Goals; Health; Heparin; High Density Lipoproteins; Human; Human Genetics; Hypertriglyceridemia; In Vitro; Injections; Investigation; Knock-out; Knockout Mice; LDL Cholesterol Lipoproteins; Lipase; Lipids; Lipoprotein Binding; Lipoproteins; Liver; Low Density Lipoprotein Receptor; Measures; Medicine; Molecular; Mus; Mutation; Myocardial Infarction; Nonesterified Fatty Acids; Olives - dietary; Oral; Outcome; Pathway interactions; Patients; Phenotype; Phospholipids; Plasma; Prevention; Process; Proprotein Convertases; Prospective Studies; Proteins; Recombinants; Residual state; Risk; Risk Factors; Role; Sampling; Stomach; Structure; Subtilisins; Techniques; Testing; Therapeutic; Tissues; Training; Triglyceride Metabolism; Triglycerides; Triolein; Variant; Work; absorption; base; biobank; career; case control; clinically relevant; design; disorder risk; effective therapy; exome sequencing; experience; experimental study; fast protein liquid chromatography; gain of function; genetic variant; genome wide association study; head-to-head comparison; in vitro Assay; in vivo; insight; interest; lipoprotein lipase; liver injury; loss of function; mouse model; novel; novel therapeutic intervention; overexpression; plasmid DNA; preference; segregation

Project Summary Coronary artery disease (CAD) is a leading cause of death worldwide. A major causal risk factor for CAD is elevated low-density lipoprotein cholesterol (LDL-C) levels. While reduction in LDL-C is a cornerstone of the prevention and treatment of CAD, many patients continue to experience CAD events despite highly effective LDL-C reduction, indicating important residual risk. Plasma triglycerides (TGs) are an independent predictor of CAD risk. TGs are strongly associated with CAD events even in statin-treated patients with low LDL-C levels, and are one of the strongest predictors of on-statin vascular risk. TGs are carried in TG-rich lipoproteins (TRLs). TRLs provide energy to extrahepatic tissues through the activity of cell-surface lipoprotein lipase (LPL), which hydrolyzes TGs in TRLs for the local absorption of the released free fatty acids. Recent large-scale human genetics studies have established that genetic variants associated with TG levels are also strongly associated with CAD. These genetic studies have especially pointed to the LPL pathway, and the apolipoprotein ApoA-V has been identified as a particularly interesting modulator of LPL. ApoA-V is primarily secreted from the liver and can exchange between high-density lipoproteins (HDLs) and TRLs. ApoA-V has been shown to enhance LPL activity, although the precise mechanism remains unclear. Human genetics have strongly supported ApoA-V’s role in TG metabolism: case-control, family-based sequencing studies, and an exome sequencing study have implicated several coding variants in hypertriglyceridemia (hyperTG), hyperchylomicronemia, and early myocardial infarction, respectively. Additionally, the hyperTG phenotype in apoa5 knockout mice can be suppressed by recombinant human ApoA-V or human APOA5 AAV. My work seeks to functionally characterize the effects of selected natural APOA5 variants, both in vitro and in vivo. Preliminary evidence supports 2 predicted loss-of-function (LoF) variants Q275X and Q305X and 3 ambiguous and/or potentially beneficial variants (predicted gain-of-function (GoF)) D37E, P215L, and T292I, that may be particularly informative for elucidating ApoA-V function. My first goal is to assess lipoprotein binding and LPL activity enhancement of selected APOA5 variants, as ApoA-V’s ability to bind lipoproteins is critical to localizing the protein at the TRL-LPL interface. I hypothesize that APOA5 predicted LoF variants decrease lipoprotein binding, blunt LPL activation, and increase plasma TG levels, while APOA5 predicted GoF variants increase lipoprotein binding, resulting in reciprocal phenotypes. My second goal is to determine APOA5 variant impacts on long-term TG metabolism and atherosclerosis. I hypothesize that atherosclerotic mouse models expressing APOA5 predicted LoF variants will have decreased TG clearance and augmented atherosclerotic progression while APOA5 predicted GoF variants will have reciprocal phenotypes. The proposed studies will provide substantial new insight into ApoA-V’s modulation of plasma TGs, a targetable CAD risk factor, that could guide development of rationally-designed therapeutics.

项目摘要 冠状动脉疾病（CAD）是全球死亡的主要原因。主要因果风险因素 CAD的低密度脂蛋白胆固醇（LDL-C）水平升高。降低LDL-C是一个基石 在预防和治疗CAD中，许多患者继续体验CAD事件的情况很高 有效的LDL-C减少，表明重要的残留风险。血浆甘油三酸酯（TGS）是独立的 预测CAD风险。 TGS即使在他汀类药物治疗的患者中，TG也与CAD事件密切相关 LDL-C水平，是状态蛋白血管风险的有力预测指标之一。 TG在TG富含TG中携带 脂蛋白（TRL）。 TRL通过细胞表面脂蛋白的活性为肝外组织提供能量 脂肪酶（LPL），它在TRL中水解TGS，用于局部滥用释放的游离脂肪酸。最近的 大规模的人类遗传学研究已经确定，与TG水平相关的遗传变异也是 与CAD密切相关。这些遗传研究尤其指出了LPL途径， 载脂蛋白APOA-V已被确定为LPL的特别有趣的调节剂。 ApoA-V是主要的 从肝脏分泌，可以在高密度脂蛋白（HDL）和TRL之间交换。 apoa-v有 尽管精确的机制尚不清楚，但我们被证明可以增强LPL活性。人遗传学有 强烈支持APOA-V在TG代谢中的作用：病例对照，基于家庭的测序研究和一个 外显子测序研究已在高甘油三项血症（Hypertg）中实施了几种编码变体， 分别高质粒度血症和早期心肌梗塞。另外，在 重组人apoA-V或Human ApoA5 AAV可以抑制ApoA5敲除小鼠。 我的作品旨在在功能上表征选定的天然ApoA5变体的影响，都在体外 和体内。初步证据支持2个预测功能丧失（LOF）变体Q275X和Q305X和3 模棱两可和/或潜在有益的变体（预测功能获得（GOF））D37E，P215L和T292I， 对于阐明APOA-V功能，这可能特别有用。我的第一个目标是评估脂蛋白 所选apoA5变体的结合和LPL活性增强，因为ApoA-V结合脂蛋白的能力为 对于将蛋白质定位在TRL-LPL界面中至关重要。我假设ApoA5预测了LOF变体 降低脂蛋白结合，钝LPL激活并增加血浆TG水平，而ApOA5 预测的GOF变体增加了脂蛋白结合，从而导致相互表现型。我的第二个目标 是为了确定对长期TG代谢和动脉粥样硬化的APOA5变体影响。我假设这一点 表达APOA5预测LOF变体的动脉粥样硬化小鼠模型将改善TG 清除和增强的动脉粥样硬化进展，而ApoA5预测GOF变体将具有 相互表型。拟议的研究将为Apoa-V的调制提供大量的新见解 等离子体TGS是一种可靶向的CAD危险因素，可以指导理性设计的治疗的发展。