Characterization of a Lysosomal Acid Lipase Variant Associated with Coronary Disease

 描述(由申请人提供)：在大规模的全基因组关联研究中，存在于近三分之一的人群中的溶酶体酸性脂肪酶(LIPA)基因座的两个紧密相连的常见内含子变体已被证明使冠状动脉疾病(CAD)的风险增加13-17%。LIPA介导胆固醇酯的水解，具有罕见功能丧失突变的患者会患上高胆固醇血症和冠心病。然而，这些常见的LIPA变异体是内含子的，与脂质异常无关，并导致单核细胞中LIPA转录增加，这一系列发现阻碍了进一步的机制理解。我们发现了一种以前未被识别的编码变体，它与内含子变体紧密连锁，与冠心病风险同样相关。我们推测，编码变异对LIPA酶活性和细胞胆固醇稳态有潜在影响，是心血管疾病的罪魁祸首。这一概念将通过评估编码变体对从已知和非已知冠心病患者队列中分离的循环单核细胞中LIPA mRNA、蛋白质和酶活性的水平的影响来进行评估。为了明确编码变体的含义，将研究在没有混淆内含体变体的情况下含有编码变体的诱导多能干细胞来源的单核/巨噬细胞。这个模型将允许确定分离中的编码变体是否改变了LIPA的mRNA、蛋白质和酶的活性。进一步的机制特征将包括对与CAD发病相关的关键脂质代谢表型的影响，包括胆固醇外流、脂质摄取和泡沫细胞形成。最后，鉴于编码变异体位于LIPA的信号肽区域，其对LIPA运输、成熟和分泌的影响将被评估为功能障碍的潜在机制。综上所述，这组高度平移的实验将为与CAD相关的一个重要的遗传风险等位基因提供第一个机械性的见解。

 DESCRIPTION (provided by applicant): Two tightly linked common intronic variants of the Lysosomal Acid Lipase (LIPA) locus, present in nearly one- third of the population, have been shown to increase the risk of coronary artery disease (CAD) by 13-17% in large-scale genome-wide association studies. LIPA mediates the hydrolysis of cholesteryl esters and patients with rare loss-of-function mutations develop hypercholesterolemia and CAD. However, these common LIPA variants are intronic, not associated with lipid abnormalities, and result in increased LIPA transcripts in monocytes, a constellation of findings that has prevented further mechanistic understanding. We have discovered a previously unrecognized coding variant in close linkage with the intronic variants that is equally associated with CAD risk. We hypothesize that the coding variant, with potential consequences to LIPA enzyme activity and cellular cholesterol homeostasis, is the culprit link with cardiovascular disease. This concept will be evaluated by assessing the effect of the coding variant on levels of LIPA mRNA, protein, and enzyme activity in circulating monocytes isolated from cohorts of patients with and without known CAD. In order to definitely implicate the coding variant, induced pluripotent stem cell derived monocytes/macrophages harboring the coding variant in the absence of the confounding intronic variants will be studied. This model will allow determination of whether the coding variant in isolation alters LIPA mRNA, protein, and enzyme activity. Further mechanistic characterization will include effects on crucial lipid metabolic phenotypes implicated in CAD pathogenesis including cholesterol efflux, lipid uptake, and foam cell formation. Finally, given that the coding variant lies in the signal peptide region of LIPA, its impact on LIPA trafficking, maturation, and secretion will be assessed as potential mechanisms for dysfunction. Taken together, this highly translational set of experiments will provide the first mechanistic insight ito an important genetic risk allele associated with CAD.