NGS in Large CAD Families: In-Depth Identification of Rare Risk Genomic Variants

摘要冠状动脉疾病（CAD）是世界范围内导致死亡的主要原因。遗传因素对CAD的发展有重要影响。因此，该项目的长期目标是确定CAD的新遗传和分子决定因素/标记。为了实现这一目标，我们花费了超过10年的时间来确定和获取24个非常大的多代家族（GeneQuest II，平均谱系大小=16）的数据。这已经成为一个独特的和非常有价值的资源，发现易感基因和基因组变异，赋予冠心病的风险。我们已经完成了408个多态性标记的全基因组连锁扫描，这些标记在GeneQuest II家族中每10 cM覆盖整个人类基因组，并在染色体3q28和7p22.3上鉴定了两个高度显著的CAD位点以及其他四个显著位点。早在90年代，我们还建立了另一个具有良好特征的428个家族性早发CAD的美国队列（GeneQuest，平均系谱大小=5）。同样的3q28 CAD位点在GeneQuest中也显示出高度显著的连锁。全基因组下一代测序（NGS）已成为鉴定复杂疾病易感基因的一种使能技术。因此，我们建议采用一种创新的综合策略，将24个GeneQuest II家族的全基因组NGS和全基因组连锁分析相结合，以确定与CAD相关的基因组变异。24个GeneQuest II家族中所有受影响的家族成员将进行全基因组NGS，并鉴定出新的罕见基因组变异。私有变异将通过与疾病家族的简单共分离来确定它们是否为致病突变。其他罕见变异将在24个大型GeneQuest II家族中使用基于家族的罕见变异关联研究来分析与CAD的关联，该研究包括基因或功能区域的多种变异以及来自多种变异的单倍型。将在复制群体（428个GeneQuest家族）中验证阳性关联。我们按照以下顺序对罕见变异进行优先排序：(1)连锁峰下的罕见变异；(2) GWAS识别的CAD位点上或附近的罕见变异；(3)连锁峰或GWAS位点外的罕见变异。生物信息学分析和相关功能/表达研究将用于确定与CAD相关的变异是否影响附近基因的功能或表达。这些研究将导致识别新的基因组变异，赋予冠心病的风险，并揭示冠心病发病机制的新的遗传/分子途径。

DESCRIPTION (provided by applicant): NGS in Large CAD Families: In-Depth Identification of Rare Risk Genomic Variants Abstract Coronary artery disease (CAD) is the leading cause of death worldwide. Genetic factors contribute significantly to the development of CAD. The long-term objective of this project is thus to identify novel genetic and molecular determinants/markers for CAD. To achieve this goal, we have spent more than 10 years of extensive efforts to identify and acquire data for 24 very large, multigenerational families (GeneQuest II, mean pedigree size=16). This has become a unique and highly valuable resource for discovering susceptibility genes and genomic variants that confer risk of CAD. We have completed a genome-wide linkage scan with 408 polymorphic markers that cover the entire human genome by every 10 cM in GeneQuest II families, and identified two highly significant CAD loci on chromosome 3q28 and 7p22.3 and four other significant loci. Back in the 90s, we also had established another well-characterized US cohort of 428 CAD families with familial, early onset CAD (GeneQuest, mean pedigree size=5). The same 3q28 CAD locus showed a highly significant linkage in GeneQuest, too. Whole genome next generation sequencing (NGS) has become an enabling technology to identify susceptibility genes for complex diseases. Thus, we propose to employ an innovative, integrated strategy that combines whole genome NGS and genome-wide linkage analysis in the 24 GeneQuest II families to identify genomic variants associated with CAD. All affected family members in the 24 GeneQuest II families will be subjected to whole genome NGS, and novel rare genomic variants will be identified. Private variants will be characterized by simple co- segregation with disease i families to determine whether they are disease-causing mutations. Other rare variants will be analyzed for association with CAD in the 24 large GeneQuest II families using family-based rare variant association studies that incorporate multiple variants in a gene or a functional region as well as haplotypes from multiple variants. Positive associations will be validated in the replication population (428 GeneQuest families). We prioritize rare variants in the following succeeding order: (1) Rare variants under linkage peaks; (2) Rare variants at or near CAD loci identified by GWAS; (3) Rare variants outside of linkage peaks or GWAS loci. Bioinformatics analysis and relevant functional/expression studies will be used to determine whether variants associated with CAD affect the function or expression of nearby genes. These studies should lead to identification of new genomic variants that confer risk of CAD and uncover novel genetic/molecular pathways for the pathogenesis of CAD.