High-throughput genetic & small-molecule screening for therapeutic modifiers

高通量遗传

• 批准号: 7938795
• 负责人: KEVIN P. CAMPBELL
• 金额: $ 91.03万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7938795
• 关键词: Affect; Affinity; Applications Grants; Area; Arenavirus; Basic Science; Biological Assay; Birth; Candidate Disease Gene; Cell Culture Techniques; Cell Line; Cell Surface Proteins; Cells; Collaborations; Collection; Communities; Cultured Cells; Degenerative Disorder; Development; Diagnosis; Disease; Dystroglycan; Extracellular Matrix Proteins; Future; Gene Expression; Gene Mutation; Generations; Genes; Genetic; Genetic Counseling; Glycoproteins; Goals; Health Benefit; Hereditary Disease; Histone Deacetylase Inhibitor; In Vitro; Individual; Inherited; Intervention; Knock-in Mouse; Laboratories; Laminin; Link; Luciferases; Mediating; Mission; Modeling; Modification; Mus; Muscle; Muscle Weakness; Muscular Dystrophies; Mutation; National Institute of Neurological Disorders and Stroke; Neuraxis; Pathology; Patients; Peptides; Pharmaceutical Preparations; Post-Translational Protein Processing; Preclinical Drug Evaluation; Prevention; Process; Processed Genes; Proteins; RNA Interference; Reporter; Research; Research Institute; Research Support; Screening procedure; Skeletal Muscle; Speed; Stroke; Therapeutic; Therapeutic Agents; Therapeutic Intervention; Time; Validation; alpha Dystroglycan; base; cohort; congenital muscular dystrophy; design; extracellular; gene discovery; glycosylation; glycosyltransferase; human disease; improved; in vivo; insight; meetings; mouse model; muscular dystrophy mouse model; nervous system disorder; new therapeutic target; novel; novel therapeutics; public health relevance; receptor; small molecule; therapeutic development; therapeutic evaluation; therapeutic target; tool; treatment strategy; wasting

DESCRIPTION (provided by applicant): Dystroglycan is a widely expressed transmembrane glycoprotein that acts as a high- affinity receptor for both extracellular matrix proteins containing laminin-G domains and certain arenaviruses. Secondary dystroglycanopathies encompass a collection of muscular dystrophies characterized by impaired post-translational processing of dystroglycan. Profound muscle weakness and wasting as well as potential central nervous system impariment are typical pathologies associated with secondary dystroglycanopathies. Causative mutations for these disorders are found in known or putative glycosyltransferases that participate in the O-glycosylation of alpha- dystroglycan, a modificaiton required for functionality. Despite extensive efforts to understand the genetics and pathology of these diseases, the genetic causes of over half of these cases remain a mystery. Furthermore, there are no treatments available for patients. This proposal outlines approaches to elucidate remaining genetic causes of dystroglycanopathies, discover and validate small molecule and peptide effectors of dystroglycan glycosylation and provide the muscular dystrophy field with improved mouse models to sustain rapid future progress. These goals will be successfully met through collaboration with the Schultz laboratory at The Scripps Research Institute. The objective of Specific Aim 1 is to identify novel dystroglycanophy genetic loci using both high-throughput in vitro complementation and knockdown screens. Elucidation of new candidate genes will offer new opportunities for improved genetic diagnosis, new viable therapeutic targets and a better understanding of dystroglycan post-translational processing. Specific Aim 2 is designed to identify novel small molecule and secreted peptide effectors of dystroglycan glycosylation in a cell culture based, high-throughput manner. This unbiased approach will provide new directions for the development of therapeutic interventions for muscular dystrophy. Specific Aim 3 is targeted at both validation of previously and newly identified therapeutic strategies and the development of conditional and knockdown mouse models of dystroglycanopathy. The new mouse models will better capture both the genetic and phenotypic complexity of dystroglycanophies than the currently available cohort of mouse models. These models will serve the muscular dystrophy research community in efforts to explain the cellular mechanism and to develop viable treatment strategies for each genetic cause of dystroglycanopathy. These studies will provide the muscular dystrophy research field with improved tools and progress towards suitable means of improving dystroglycan function. This research meets the challenge of the National Institute of Neurological Disorders and Stroke mission statement to support "research on the causes, prevention, diagnosis, and treatment of neurological disorders and stroke, and supports basic research in related scientific areas". PUBLIC HEALTH RELEVANCE: Muscular dystrophies are a diverse set of inherited diseases characterized by progressive skeletal muscle weakness and wasting. Dystroglycan, a cell surface protein, requires extensive modification to serve as a link between the intracellular and extracellular cellular support network in muscle such that, when disrupted, it results in several forms of muscular dystrophy. This proposal is designed to identify new gene mutations that can cause these types of muscular dystrophy, discover small molecules that can improve dystroglycan function, develop needed mouse models of the disease and to validate both newly identified and currently known treatment strategies.

描述（由申请人提供）：肌营养不良聚糖是一种广泛表达的跨膜糖蛋白，其作为含有层粘连蛋白-G结构域的细胞外基质蛋白和某些沙粒病毒的高亲和力受体。继发性肌营养不良症包括一系列以肌营养不良症翻译后加工受损为特征的肌营养不良症。严重的肌肉无力和消耗以及潜在的中枢神经系统损伤是与继发性肌营养不良症相关的典型病理。这些疾病的致病突变存在于已知或推定的糖基转移酶中，这些糖基转移酶参与α-肌营养不良聚糖的O-糖基化，这是功能所需的修饰。尽管人们为了解这些疾病的遗传学和病理学做出了广泛的努力，但其中一半以上病例的遗传原因仍然是个谜。此外，患者没有可用的治疗方法。该提案概述了阐明肌营养不良症剩余遗传原因的方法，发现和验证肌营养不良症糖基化的小分子和肽效应物，并为肌营养不良领域提供改进的小鼠模型以维持未来的快速进展。通过与斯克里普斯研究所的舒尔茨实验室合作，这些目标将成功实现。具体目标 1 的目标是利用高通量体外互补和敲低筛选来鉴定新的营养不良遗传位点。新候选基因的阐明将为改进基因诊断、新的可行治疗靶点以及更好地理解肌营养不良聚糖翻译后加工提供新的机会。具体目标 2 旨在以基于细胞培养的高通量方式鉴定肌营养不良聚糖糖基化的新型小分子和分泌肽效应物。这种公正的方法将为肌营养不良症治疗干预措施的发展提供新的方向。具体目标 3 旨在验证先前和新确定的治疗策略，以及开发肌聚糖病的条件性和敲除小鼠模型。与目前可用的小鼠模型相比，新的小鼠模型将更好地捕捉肌营养不良症的遗传和表型复杂性。这些模型将为肌营养不良症研究界服务，努力解释细胞机制，并为肌营养不良症的每种遗传原因制定可行的治疗策略。这些研究将为肌营养不良研究领域提供改进的工具，并在改善肌营养不良聚糖功能的合适方法方面取得进展。这项研究满足了国家神经疾病和中风研究所使命宣言的挑战，即支持“神经疾病和中风的病因、预防、诊断和治疗的研究，并支持相关科学领域的基础研究”。 公众健康相关性：肌营养不良症是一系列不同的遗传性疾病，其特征是进行性骨骼肌无力和萎缩。肌营养不良症聚糖是一种细胞表面蛋白，需要进行广泛的修饰才能充当肌肉中细胞内和细胞外细胞支持网络之间的联系，因此，当它被破坏时，会导致多种形式的肌营养不良症。该提案旨在识别可能导致这些类型的肌营养不良症的新基因突变，发现可以改善肌营养不良症功能的小分子，开发所需的疾病小鼠模型，并验证新发现的和目前已知的治疗策略。