CLARIN 1 RETINAL FUNCTION AND THERAPEUTIC IMPLICATIONS FOR USH3

CLARIN 1 视网膜功能和 USH3 的治疗意义

• 批准号: 10006553
• 负责人: ASTRA DINCULESCU
• 金额: $ 37.5万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10006553
• 关键词: A/J Mouse; Address; Affect; Afferent Neurons; Aging; Amino Acids; Animal Model; Apical; Binding; Binding Proteins; Biological; Biological Assay; Blindness; Cell Death; Cell membrane; Cells; Cellular Structures; Chickens; Cilia; Clarin-1; Cochlea; Cochlear implant procedure; Code; Conflict (Psychology); Cultured Cells; Cytoskeleton; Data; Disease; Ear; Elements; Endoplasmic Reticulum; F-Actin; Genes; Goals; Grant; Hair Cells; Heat shock proteins; Hereditary Disease; Human; Individual; Knock-in Mouse; Knock-out; Knockout Mice; Labyrinth; Light; Mediating; Morphology; Mus; Mutation; N-Glycosylation Site; North America; Organelles; Outcome; Pathogenicity; Pathology; Patients; Pattern; Phenotype; Photoreceptors; Play; Property; Protein Isoforms; Proteins; Recombinants; Reporting; Research; Retina; Retinal Degeneration; Retinal Diseases; Retinitis Pigmentosa; Role; Sensory; Sensory Hair; Structure; Supplementation; Surface; Testing; Therapeutic; Therapeutic Effect; Usher Proteins; Usher Syndrome; Usher Syndrome Type 3; Variant; Vision; adeno-associated viral vector; arrestin 1; beta Actin; deafness; design; disease-causing mutation; endoplasmic reticulum stress; experimental study; gene replacement therapy; gene therapy; hearing impairment; in vivo; loss of function; mechanotransduction; member; mouse model; mutant; normal hearing; postnatal; preservation; prevent; prevent hearing loss; progressive hearing loss; promoter; restoration; retinal neuron; vector

ABSTRACT Usher syndrome (USH) is an inherited disorder responsible for more than half of combined deafness and blindness in humans. It affects both photoreceptors and cochlear hair cells, the sensory neurons of the retina and inner ear, respectively. Usher syndrome type III (USH3) is caused by mutations in clarin-1 (CLRN1) gene, leading to the gradual and permanent loss of both vision and normal hearing. Cochlear implantation alleviates to some degree the hearing impairment, but there is no treatment that prevents the progressive photoreceptor cell death that leads to blindness. Our goal is to develop an understanding of CLRN1 function in the retina so that a rational gene therapy approach for USH3 patients can be developed. Efforts to identify efficient therapeutic strategies for vision loss are hindered by the lack of USH3 animal models that mimic the human retinal disorder, and by conflicting reports on CLRN1’s endogenous cellular localization, with only one study showing its presence in photoreceptors. All current USH3 mouse models display a rapid hearing loss but normal retinal function and morphology. As part of a collaborative effort, we showed that CLRN1 gene replacement therapy using adeno-associated viral vectors (AAVs) can prevent hearing loss for at least 1 year following cochlear delivery in a mouse model of USH3, the first successful proof of concept experiment for this disorder. We also showed that retinal function is preserved following a similar gene therapy approach to treat the retina in a new mouse model of USH3, which displays a slow, progressive loss of retinal function with aging. These results suggest that postnatal CLRN1 delivery can prevent further loss of function in sensory cells, a particularly relevant issue for patients with more advanced stages of disease. Our central hypothesis is that CLRN1 plays an essential role in photoreceptor cells, structurally stabilizing the fragile connecting cilium region by interacting with other USH proteins and cytoskeletal elements. We address this hypothesis with three specific aims: 1) identify CLRN1’s endogenous retinal localization and the proteins that interact with CLRN1 in the mouse retina; 2) determine the retinal expression pattern and pathogenic effects of mutant CLRN1 proteins in-vivo; 3) test several gene therapy options in USH3 mouse models, targeting either the entire retina, or the photoreceptors alone. Completion of these goals would firmly identify CLRN1’s general function and, critically, serve to pinpoint which cells would need to be targeted for USH3 gene replacement therapy to prevent the loss of vision in patients.

抽象的 Usher综合征（USH）是一种遗传性疾病，负责超过一半的聋哑和 人类失明。它影响感光体和人工耳蜗细胞，即 视网膜和内耳。 Usher综合征III型（USH3）是由Clarin-1突变引起的 （CLRN1）基因，导致视力和正常听力的等级和永久丧失。 植入在某种程度上减轻了听力障碍，但是没有治疗可以阻止 导致失明的进行性光感受器细胞死亡。我们的目标是建立对 CLRN1在视网膜中的功能，因此可以开发为USH3患者的理性基因治疗方法。 缺乏USH3动物，阻碍了确定有效的视力丧失治疗策略的努力 模仿人类残留障碍的模型，以及有关CLRN1内源性细胞的报告 本地化，只有一项研究显示了它在感光体中的存在。所有当前的USH3鼠标模型 显示出迅速的听力损失，但残留功能和形态正常。作为协作努力的一部分，我们 表明使用腺相关病毒载体（AAV）的CLRN1基因替代疗法可以预防 在人工耳蜗交付后，在USH3的鼠标模型中，听力损失至少1年，这是第一个成功的 该疾病的概念验证实验。我们还表明，视网膜功能在 类似的基因疗法方法可以在新的USH3小鼠模型中治疗视网膜，该模型显示出缓慢的， 随着老化的视网膜功能的进行性丧失。这些结果表明，产后CLRN1交付可以 防止感觉细胞的进一步丧失功能，这是一个特别先进的患者的特别相关问题 疾病阶段。我们的中心假设是CLRN1在感光细胞中起着至关重要的作用， 通过与其他USH蛋白质相互作用和 细胞骨架元素。我们以三个具体目的解决了这一假设：1）确定CLRN1 内源性视网膜定位和与小鼠视网膜中CLRN1相互作用的蛋白质； 2）确定 突变体CLRN1蛋白在体内的视网膜表达模式和致病作用； 3）测试几个基因 USH3小鼠模型中的治疗选择，单独针对整个视网膜或光感受器。 这些目标的完成将严重确定CLRN1的一般功能，并批判性地确定 哪些细胞需要针对USH3基因替代疗法，以防止视力丧失 患者。