Laying a Foundation for Precision Medicine for Fuchs' Dystrophy

为福克斯营养不良症的精准医学奠定基础

基本信息

批准号：
10667612
负责人：
Venkateswara Vinod Mootha
金额：
$ 39.31万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-05-01 至 2025-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10667612
关键词：
Affect Age Aging Anterior Antisense Oligonucleotides Apoptosis Aqueous Humor Binding Biology Blindness Blood CUG repeat Cells Cornea Corneal Diseases Corneal Endothelium Corneal edema Country Cultured Cells DNA Repeat Expansion DNA Sequence Data Defect Degenerative Disorder Disease Disease Progression Disease model Dominant-Negative Mutation Edema Endothelial Cells Endothelium Event Expanded DNA Repeat Extracellular Matrix Extracellular Matrix Proteins Fibrosis Foundations Fuchs&apos Endothelial Dystrophy Future Gene Expression Genes Genetic Genetic Transcription Glaucoma Histologic Immune Inherited Keratoplasty Knock-in Laboratories Lead Length Leukocytes Link Medical Modeling Molecular Mus Myotonic Dystrophy Neural Crest Neurodegenerative Disorders Nuclear Oligonucleotides PF4 Gene Pathogenesis Pathway interactions Patients Population Pre-Clinical Model Prevalence Primary Open Angle Glaucoma Proteins Publishing RNA RNA Splicing Reporting Resources Risk Role Safety Sampling Somatic Mutation Specificity TCF4 haploinsufficiency Testing Therapeutic Tissues Toxic effect Trabecular meshwork structure Transcript Transcription Alteration Trinucleotide Repeats United States Up-Regulation Wild Type Mouse Work age related autosome biobank cell type cohort corneal epithelium drug testing empowerment gain of function genomic locus in vivo knock-down mouse model multidisciplinary overexpression postmitotic precision medicine premature prevent recruit senescence single-cell RNA sequencing therapeutic development transcription factor transcriptome transcriptome sequencing

项目摘要

PROJECT SUMMARY Fuchs’ endothelial corneal dystrophy (FECD) is an age-related degenerative disorder resulting in corneal edema and loss of vision. FECD occurs in 4% of whites over the age of 40 years and is the leading indication for corneal transplantation in the U.S. Over 70% of cases are caused by a CTG triplet repeat expansion in the TCF4 gene. Expanded CUG repeat RNA (CUGexp) transcripts expressed from this gene locus accumulate as nuclear foci in the corneal endothelium of patients. The CUGexp foci bind and functionally sequester the splicing factors MBNL1 and MBNL2 to trigger mis-splicing in FECD endothelial tissue. To examine the endothelial cell-type specificity for FECD, we will determine if somatic mutations in the post-mitotic corneal endothelium of FECD patients results in a larger triplet repeat expansion than in their blood. We will examine if other anterior segment cell-types including corneal epithelium, stromal keratocytes, and trabecular meshwork cells are prone to accumulation of CUGexp foci and associated molecular defects. To test the MBNL sequestration hypothesis, we will examine if the knockdown of MBNLs in healthy donor endothelial tissue is sufficient to recapitulate the mis-splicing and upregulation of extracellular matrix (ECM) genes found early in the disease course. In early and late FECD, we observed a marked overexpression of the cochlin gene that produces a secreted ECM protein also capable of recruiting immune cells. The cochlin protein was previously detected in the trabecular meshwork of patients with primary open angle glaucoma (POAG), and our preliminary data indicate that the protein is present in the aqueous humor and trabecular meshwork of FECD subjects. We will examine levels of the cochlin protein in corneal tissue, aqueous humor samples, and trabecular meshwork tissue of FECD patients for its possible contribution to corneal disease findings and the increased risk for glaucoma in these patients. Additionally, we will examine the prevalence of POAG in our large UTSW FECD cohort and determine if there is a correlation with the triplet repeat length. We will use single-cell RNA sequencing combined with IHC to identify the immune cells and their contribution to late-stage FECD. We have characterized the efficacy of 45 antisense oligonucleotides (ASOs) and duplex RNAs that block disease-causing CUGexp foci. Using patient-derived cells and tissue, we will examine their potency based on their ability to block foci formation and their specificity based on transcriptome-wide assessment of on- and off-target events. We will examine delivery, length of action, and safety using wild-type mice. To enable studies of disease biology and in vivo drug testing, we have generated what we believe is the first mouse model with a knock-in of expanded CTG repeats. In preliminary studies, we show that our knock-in strategy successfully recapitulates CUGexp foci formation in corneal endothelium. We will further characterize this murine model by aging these mice to examine for the transcriptional and histologic alterations of FECD disease, and then use these mice to test our lead CUG-repeat targeting oligonucleotides for therapeutic development.

项目概要福克斯内皮性角膜营养不良 (FECD) 是一种与年龄相关的退行性疾病，导致角膜水肿 40 岁以上的白人中有 4% 会出现 FECD，这是角膜的主要指征。在美国，超过 70% 的移植病例是由 TCF4 基因中的 CTG 三联体重复扩增引起的。从该基因位点表达的扩展 CUG 重复 RNA (CUGexp) 转录物在 CUGexp 病灶结合并功能性隔离剪接因子 MBNL1。和 MBNL2 触发 FECD 内皮组织中的错误剪接以检查内皮细胞类型特异性。对于 FECD，我们将确定 FECD 患者有丝分裂后角膜内皮是否发生体细胞突变我们将检查是否存在其他前段细胞类型。包括角膜上皮、角膜基质细胞和小梁网细胞，容易积聚为了检验 MBNL 隔离假说，我们将检查是否存在 CUGexp 病灶和相关分子缺陷。健康供体内皮组织中 MBNL 的敲低足以重现错误剪接和在早期和晚期 FECD 中，我们发现细胞外基质 (ECM) 基因的上调。观察到 cochlin 基因显着过度表达，该基因产生分泌的 ECM 蛋白，也能够先前在患有该病的患者的小梁网中检测到了cochlin蛋白。原发性开角型青光眼（POAG），我们的初步数据表明该蛋白质存在于我们将检查 FECD 受试者的房水和小梁网中的耳蜗蛋白水平。 FECD 患者的角膜组织、房水样本和小梁网组织，以了解其可能的情况此外，我们还对角膜疾病的发现以及这些患者患青光眼的风险增加做出了贡献。将检查我们大型 UTSW FECD 队列中 POAG 的患病率，并确定是否存在相关性我们将使用单细胞RNA测序结合IHC来鉴定免疫。细胞及其对晚期 FECD 的贡献我们已经表征了 45 反义的功效。使用患者来源的细胞阻断致病性 CUGexp 病灶的寡核苷酸 (ASO) 和双链体 RNA。和组织，我们将根据它们阻止病灶形成的能力及其基于的特异性来检查它们的效力我们将检查传递、作用时长和对靶标和脱靶事件的全转录组评估。为了进行疾病生物学和体内药物测试的研究，我们已经生成了使用野生型小鼠的安全性。我们认为这是第一个敲入扩展 CTG 重复序列的小鼠模型。表明我们的敲入策略成功地重现了角膜内皮中的 CUGexp 病灶形成。通过老化这些小鼠来检查转录和组织学，进一步表征该小鼠模型 FECD 疾病的改变，然后使用这些小鼠来测试我们的领先 CUG 重复靶向寡核苷酸治疗的发展。

项目成果

期刊论文数量（12）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Instability of TCF4 Triplet Repeat Expansion With Parent-Child Transmission in Fuchs' Endothelial Corneal Dystrophy.

Fuchs 内皮性角膜营养不良中 TCF4 三联体重复扩增与亲子传递的不稳定性。

DOI：
10.1167/iovs.18-24119
发表时间：
2018
期刊：
Investigative ophthalmology & visual science
影响因子：
4.4
作者：
Saade,JoannaS;Xing,Chao;Gong,Xin;Zhou,Zhengyang;Mootha,VVinod
通讯作者：
Mootha,VVinod

Fuchs' Endothelial Corneal Dystrophy and RNA Foci in Patients With Myotonic Dystrophy.

DOI：
10.1167/iovs.17-22350
发表时间：
2017-09-01
期刊：
Investigative ophthalmology & visual science
影响因子：
4.4
作者：
Mootha VV;Hansen B;Rong Z;Mammen PP;Zhou Z;Xing C;Gong X
通讯作者：
Gong X

Transcriptomic meta-analysis reveals ERRα-mediated oxidative phosphorylation is downregulated in Fuchs' endothelial corneal dystrophy.

DOI：
10.1371/journal.pone.0295542
发表时间：
2023
期刊：
PloS one
影响因子：
3.7
作者：
通讯作者：

Congenital Corneal Endothelial Dystrophies Resulting From Novel De Novo Mutations.

DOI：
10.1097/ico.0000000000000670
发表时间：
2016-02
期刊：
Cornea
影响因子：
2.8
作者：
Cunnusamy K;Bowman CB;Beebe W;Gong X;Hogan RN;Mootha VV
通讯作者：
Mootha VV

A novel H395R mutation in MKKS/BBS6 causes retinitis pigmentosa and polydactyly without other findings of Bardet-Biedl or McKusick-Kaufman syndrome.

MKKS/BBS6 中的一种新的 H395R 突变会导致色素性视网膜炎和多指症，而没有其他 Bardet-Biedl 或 McKusick-Kaufman 综合征的发现。

DOI：
发表时间：
2016
期刊：
Molecular vision
影响因子：
2.2
作者：
Hulleman,JohnD;Nguyen,Annie;Ramprasad,VL;Murugan,Sakthivel;Gupta,Ravi;Mahindrakar,Avinash;Angara,Ravi;Sankurathri,Chandrasekhar;Mootha,VVinod
通讯作者：
Mootha,VVinod