Cellular Phenotypes of Genetic Variants in Mucopolysaccharidosis

粘多糖贮积症遗传变异的细胞表型

• 批准号: 10638709
• 负责人: William J Buchser
• 金额: $ 45.15万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2028-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10638709
• 关键词: A549; Affect; Attenuated; Benign; Biochemical; Biological Assay; Biopsy; Birth; Cell Line; Cells; Cellular Morphology; Chimeric Proteins; Classification; Clinical; Complementary DNA; Consent; DNA; Data; Dermatan Sulfate; Diagnosis; Diagnostic; Diagnostic tests; Discrimination; Disease; Enzymes; Generations; Genes; Genetic Diseases; Genetic Variation; Genome engineering; Genomics; Glycosaminoglycans; Goals; Heparitin Sulfate; Human; Image; Impairment; In Vitro; Individual; Inherited; Intellectual functioning disability; Knock-out; Laboratories; Libraries; Link; Lysosomes; Machine Learning; Measurement; Measures; Medical; Medical Records; Membrane; Methods; Molecular; Morphology; Mucopolysaccharidoses; Mucopolysaccharidosis II; Neonatal Screening; Neurons; Pathogenicity; Patients; Phenotype; Population; Reporting; Research; Resolution; Sampling; Severities; Skin; System; Technology; Test Result; Testing; Transmembrane Domain; Universities; Variant; Washington; Whole Blood; cell type; diagnostic accuracy; disorder risk; experimental study; genetic testing; genetic variant; iduronate-2-sulfatase; improved; induced pluripotent stem cell; molecular diagnostics; novel; phenotypic data; polysulfated glycosaminoglycan; prevent; risk prediction; single cell sequencing; stem cells; variant of unknown significance

ABSTRACT There is presently an urgent need to develop methodical approaches to evaluate the function of genomic vari- ants. Clinical genomic testing is growing rapidly, and with it a larger-than-ever number of genetic variants that cannot be defined as either disease-causing or benign. The problem affects clinicians and their patients, who struggle to understand and interpret molecular diagnostic reports, the implications of the results, and how to manage their patients in the absence of definitive information. Systems that involve single cells to generate high-content, high-resolution functional data are of paramount importance to solving the problem posed by var- iants of uncertain significance. Here, we propose to utilize a novel cell-based platform that uses machine learn- ing to determine the combination of morphological phenotypes that define pathogenicity. We will apply the technology to the comprehensive functional assessment of variants in IDS, the gene responsible for Hunter syndrome. The core hypothesis outlined in this proposal is that experimental data measuring the direct func- tional effects of variants will inform accurate disease risk prediction. In addition, we hypothesize that an in vitro, cell-based assay based on morphological features will more accurately detect disease compared to existing biochemical testing using artificial substrates. We will perform a functional assay in a variant library using the RaftSeq pipeline in the Buchser laboratory, where a cellular phenotype will be established and then tested us- ing a second set of variants combined with rescue experiments. The results will inform variant classification in IDS molecular testing and improve diagnosis of individuals including those identified by low iduronate-2-sulfa- tase activity on newborn screening.

抽象的 目前，迫切需要开发有条不紊的方法来评估基因组变量的功能 蚂蚁。临床基因组检测正在迅速增长，并且它的遗传变异数量超过了 不能定义为引起疾病或良性。这个问题会影响临床医生及其患者 难以理解和解释分子诊断报告，结果的含义以及如何 在没有明确信息的情况下管理患者。涉及单元单元生成的系统 高结合，高分辨率功能数据对于解决var-提出的问题至关重要 具有不确定意义的IANT。在这里，我们建议利用一个基于细胞的新型平台，该平台使用机器学习 - 确定定义致病性的形态表型的组合。我们将应用 对ID中变体的全面功能评估的技术，负责猎人的基因 综合征。该提案中概述的核心假设是，测量直接func-的实验数据 变体的效果将为准确的疾病风险预测提供信息。此外，我们假设一个体外， 与现有 使用人工底物进行生化测试。我们将在变体库中使用该功能测定 布赫瑟实验室中的RaftSeq管道，在该实验室中将建立细胞表型，然后测试US- 第二组变体与救援实验相结合。结果将为变体分类提供信息 IDS分子测试并改善对个体的诊断，包括低iduronate-2-sulfa-鉴定的人 在新生儿筛查上的活动。