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Cellular and Genetic Defects in Keratoconus

圆锥角膜的细胞和遗传缺陷

基本信息

批准号：
10584762
负责人：
Shukti Chakravarti
金额：
$ 62.51万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-06-10 至 2027-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10584762
关键词：
Abbreviations Address Adherent Culture Adhesions Adolescent Affect Animal Model Antioxidants Biological Biological Assay Biological Markers Biomechanics Blindness Blood Cells CENPF gene CRISPR/Cas technology Candidate Disease Gene Cell Culture Techniques Cell Differentiation process Cell Survival Cell physiology Cells Cellular Stress Cicatrix Classification Clinical Clustered Regularly Interspaced Short Palindromic Repeats Code Collagen Complex Cornea Corneal dystrophy Cytoskeleton DNA Data Databases Defect Development Diagnosis Disease Disease model Disease susceptibility Early Diagnosis Early treatment Epithelial Cells Etiology Extracellular Matrix Extracellular Matrix Proteins Eye diseases Family Foundations Future Gene Frequency Genes Genetic Genetic Predisposition to Disease Goals Human Individual Keratoconus Keratoplasty Lead Liquid substance Measurable Measures Mediating Minor Modeling Mutate Mutation Organoids Oxidative Stress Pathogenesis Pathogenicity Patients Penetrance Peripheral Blood Mononuclear Cell Persons Physiological Predisposition Production Protein Secretion Proteomics Publishing Regulation Risk Shapes Sorting Stratification Stromal Cells Testing Therapeutic Agents Thick Thinness Tissues Untranslated RNA Variant Vision Work alternative treatment biobank biomarker identification comparison control corneal epithelium crosslink driving force exome exome sequencing functional disability gene network genetic architecture genome sequencing genome wide association study genome-wide high risk improved induced pluripotent stem cell insight migration next generation sequencing oxidative damage programs response transcriptomics translational impact treatment strategy whole genome

项目摘要

Keratoconus (KC), a common corneal dystrophy that affects young people, causes progressive thinning, scarring and loss of corneal shape, which can ultimately lead to loss of vision. Crosslinking of collagens in the cornea can stiffen and delay its weakening, but severe cases require corneal transplantation. Although KC has a strong genetic component, its etiology is complex, polygenic and multifactorial. There is an urgent need to understand its etiology for developing early diagnosis and treatment strategies for KC. To address this, our competitive renewal application focuses on identifying cellular defects, biomarkers and the genetic causes of KC. Beyond obvious familial KC, the vast majority are isolated where disease likely results from rare pathogenic coding sequence variants and genome-wide common noncoding variants that increase one's susceptibility. Elucidating the underlying genetic defects in these “isolated KC” requires a range of biological evidence. Our recent studies and preliminary data provide this biological foundation for the current proposal. First, by whole- exome sequencing of KC families, we identified rare pathogenic variants in genes related to cell stress, cytoskeleton and extracellular matrix (ECM), which are now prioritized as candidate genes and networks for the isolated KC studies. Second, our transcriptomic and proteomic characterizations of KC and control donor corneas identified significant dysregulation in the NRF2-antioxidant program that is crucial for corneal cell survival and its functions. Finally, we developed corneal cell culture models that mimic key KC features, from oxidative stress to ECM insufficiency, and assays to measure these. We further developed the first cornea organoids from human induced pluripotent stem cells that will allow functional studies of genes and therapeutic agents in a physiological, cornea-like setting and in organoid-derived epithelial and stromal cell cultures. Importantly, this approach will yield cell culture disease models from genetically defined patient blood cells. These cell culture disease surrogates are particularly important, as there are no animal models that can capture the polygenic complexity of KC. In Aim 1 we will assess potential NRF2-regulated antioxidants as tear fluid biomarkers for KC, and investigate this network in corneal cell cultures. In Aim 2 we will identify rare pathogenic variants and common noncoding variants that increase disease susceptibility in isolated KC cases using the 1000Genome and the UK Biobank databases as controls. In Aim 3 we will functionally test the concept that a rare pathogenic variant (e.g., our published c.G12982A HSPG2), will cause cellular disease surrogates when CRISPR-edited into cells derived from KC individuals with high polygenic and not controls with low polygenic scores. Our findings will lead to potential anti-oxidant biomarkers, development of NRF2- activators for KC treatments, genetically defined KC cell culture models and insights into the complex genetic architecture of KC. Our studies are highly relevant to the goals of the NEI in understanding the complex genetics of eye diseases, treatments and reversing vision loss.

圆锥角膜（KC）是一种影响年轻人的常见角膜营养不良，会导致角膜逐渐变薄，