A synthetic genomics approach for characterizing Age-Related Macular Degeneration-associated genetic variation

用于表征年龄相关性黄斑变性相关遗传变异的合成基因组学方法

• 批准号: 9889806
• 负责人: Jon Laurent
• 金额: $ 2.89万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2020-08-24
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9889806
• 关键词: 10q26; Affect; Age; Age related macular degeneration; Alleles; Biological Assay; Blindness; CRISPR/Cas technology; Cell Line; Cells; Chromatin; Chromosomes; Conflict (Psychology); DNA; DNA Resequencing; DNA biosynthesis; Data; Development; Diagnosis; Disease; Disease Progression; Disease susceptibility; Engineering; Environmental Risk Factor; Eye; Genes; Genetic Engineering; Genetic Variation; Genome; Genomic Segment; Genomic approach; Genotype; Goals; Haplotypes; Healthcare Systems; Heritability; Human; Human Cell Line; In Vitro; Libraries; Mammalian Cell; Metabolic Pathway; Methods; Mutation; Nature; Odds Ratio; Organism; Pathology; Patients; Phenotype; Population; Predisposition; Process; Property; Reporting; Research Design; Research Personnel; Resolution; Retina; Risk; Scheme; Sentinel; Site; Smoking; Structure of retinal pigment epithelium; Symptoms; Techniques; Technology; Tissues; Untranslated RNA; Variant; Work; Yeasts; age effect; base; causal variant; cell type; chromosomal location; combinatorial; complement pathway; design; gene therapy; genetic association; genetic risk factor; genome wide association study; genomic locus; healthspan; human tissue; in vivo; induced pluripotent stem cell; insertion/deletion mutation; new technology; novel; protective allele; risk variant; synthetic biology; synthetic genomics; therapeutic development; tool; yeast genome

Project Summary Age-related macular degeneration (AMD) is the primary cause of blindness in the developed world, and as populations age, AMD will become an increasingly large burden on our health care system. The eye represents one of the few tissues amenable to current gene therapy techniques, and thus identifying specific genetic changes giving rise to AMD would be a critical step in developing AMD-correcting gene therapies. Toward this end, recent genetic association studies have identified a genomic region that represents a major risk locus for AMD. The risk alleles associated with AMD in this region are characterized by significant and large odds ratios (>5), implicating this as a major determinant of disease susceptibility. The AMD-associated region lies at chromosomal location 10q26. The region encompasses all or part of three genes, HTRA1, ARMS2, and PLEKHA1, all of which are candidates to affect the retina and be a causal factor in AMD development. Further, the nature of genotyping data used in the association studies means that the associated SNPs are actually ‘sentinel’ SNPs representing a large haplotype block containing many other variants. It is therefore difficult to assess which SNP or indel is actually responsible for any functional effect contributing to the disease. A recent resequencing effort has identified at least 67 variants located within the AMD-associated locus at 10q26, any or many of which could be functional. Limited functional studies in the region have led to conflicting reports regarding which variant sites are functional, and which gene is the target of those sites, with some evidence supporting either or both of HTRA1 and ARMS2 as the key functional target. To efficiently assay and identify functional variants in such large genomic regions, the lab has developed new technologies for the assembly of very large DNA molecules. These techniques have been used to date for assembling megabase-sized yeast chromosomes, as well as myriad metabolic pathways and human gene loci. Further, methods for efficient incorporation of these large assemblies into the genome of various mammalian cell types have been described. The assembly strategy for these so-called ‘assemblons’ allows combinatorial incorporation of variable segments, thus enabling systematic functional analysis of single or multiple variants in parallel. The proposed study will utilize these technologies together to rapidly identify and characterize the in vitro and in vivo effects of AMD-associated variants in cells. Identification of the causal variants will enable targeted genotyping for AMD diagnosis and eventually development of targeted gene therapy to ameliorate AMD symptoms.

项目摘要 视网膜相关性黄斑变性（AMD）是发达国家失明的主要原因， 随着人口老龄化，AMD将成为我们医疗保健系统越来越大的负担。眼睛代表 少数几个组织之一，适合目前的基因治疗技术，从而确定特定的遗传 导致AMD的基因改变将是开发AMD纠正基因疗法的关键一步。朝着这个 最后，最近的遗传关联研究已经确定了一个基因组区域，代表了一个主要的风险位点， AMD.该地区与AMD相关的风险等位基因的特征是显著和较大的比值比 （>5），暗示这是疾病易感性的主要决定因素。 AMD相关区域位于染色体位置10q26。该区域包括三个区域的全部或部分 基因HTRA1、ARMS2和PLEKHA1，所有这些都是影响视网膜的候选者，并且是视网膜病变的致病因素。 AMD开发。此外，关联研究中使用的基因分型数据的性质意味着， 相关的SNP实际上是代表包含许多其他单倍型块的“哨兵”SNP。 变体。因此，很难评估哪个SNP或indel实际上负责任何功能效应 导致疾病。最近的一项重新测序工作已经确定了至少67个位于 10q26的AMD相关基因座，其中任何一个或多个可能是功能性的。功能研究有限， 关于哪些变异位点是功能性的，以及哪个基因是靶基因， 这些网站，有一些证据支持HTRA1和ARMS 2中的一个或两个作为关键功能目标。 为了有效地分析和鉴定如此大的基因组区域中的功能变体，该实验室开发了新的 组装超大型DNA分子的技术。这些技术迄今已被用于 组装巨型酵母染色体，以及无数的代谢途径和人类基因位点。 此外，用于将这些大组装体有效掺入各种哺乳动物的基因组中的方法 已经描述了细胞类型。这些所谓的“装配体”的装配策略允许组合 可变区段的并入，从而使得能够对单个或多个变体进行系统的功能分析， 并联拟议的研究将利用这些技术一起，以快速识别和表征在 细胞中AMD相关变体的体外和体内效应。鉴定致病变异体将使 用于AMD诊断的靶向基因分型和最终开发改善AMD的靶向基因疗法 症状