Therapeutic strategies for mitigating loss of retinal ganglion cells in familial dysautonomia

减轻家族性自主神经功能障碍患者视网膜神经节细胞丢失的治疗策略

基本信息

批准号：
10093053
负责人：
Frances Lefcort
金额：
$ 18.64万
依托单位：
MONTANA STATE UNIVERSITY - BOZEMAN
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-01 至 2023-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10093053
关键词：
5&apos Splice Site Age Automobile Driving Autonomic nervous system Blindness Blood Brain Cell Survival Cessation of life Clinic Clinical Clinical Research Cytoprotection Data Defect Diet Disease Embryo Enhancers Exons Familial Dysautonomia Family Generations Genes Genetic Goals Human Hybrids In Vitro Individual Introns Kinetins Knockout Mice Life Mediating Messenger RNA Methods Modeling Modification Mus Mutate Mutation Nervous system structure Neurodegenerative Disorders Neurons Oral Pathway interactions Patients Penetration Peripheral Phenotype Point Mutation Pre-Clinical Model Preclinical Testing Proteins Publishing Quality of life RNA Splicing Reporting Retina Retinal Degeneration Retinal Dystrophy Retinal Ganglion Cells Sensory Spinal Muscular Atrophy Splice-Site Mutation Technology Teenagers Testing Therapeutic Tissues Transgenes Transgenic Mice Translations Treatment Protocols Viral Wild Type Mouse Work base cell type clinical investigation compare effectiveness conditional knockout disease phenotype effectiveness evaluation effectiveness testing exon skipping ganglion cell gene correction gene therapy improved in vivo insight intravitreal injection mRNA Precursor macula mouse model mutant nervous system disorder novel optic nerve disorder pre-clinical prevent promoter protein complex recessive genetic trait restoration retinal ganglion cell degeneration retinal neuron reverse genetics small molecule targeted treatment tool

项目摘要

PROJECT SUMMARY Given the recent FDA approval of targeted AAV gene therapy platforms and of small-molecule splicing modulators as treatments for genetic neurological disorders, our goal is to apply these powerful technologies to prevent the progressive optic neuropathy and blindness that develops in patients with the genetic recessive disease, Familial dysautonomia (FD). FD results from a splice site mutation in intron 20 of the gene ELP1 (formerly called IKBKAP). As a consequence of the mis-splicing, exon 20 is variably skipped, the mutant mRNA degraded, resulting in reduced levels of the encoded protein, Elp1.. Interestingly, the ability to splice the mutated pre-mRNA varies according to tissue type, with neurons least capable of splicing the mutated pre- mRNA. While the majority of the clinical deficits are due to the devastation of the sensory and autonomic nervous systems, as patients enter their teens, their macular retinal ganglion cells progressively die, manifesting as visual loss. Mouse that are null for Elp1 are embryonic lethal so the field has, until now, taken two distinct strategies to generate mouse models to investigate FD: (i) generation of conditional knock-out mice (CKO) using cell-type specific cre-driven promoters; and (ii) transgenic mice that contain the human FD ELP1 splicing mutation. The former approach has generated mouse models that recapitulate the FD optic neuropathy that results from the progressive death of retinal ganglion cells. These mice are an excellent pre- clinical model for testing the effectiveness of gene therapy for preventing the progressive demise of retinal ganglion cells (Aim 1A). However this model does not lend itself to testing the effectiveness of splicing enhancer compounds since it lacks the FD splicing mutation. The latter approach has culminated in the generation of transgenic mice that include copies of the human FD ELP1 mutated gene. These mice are asymptomatic unless they are crossed to a hypomorph or null background mouse, but these compound mice are typically too sick to investigate consistently. Here we will make a new “hybrid” line by crossing in the human FD ELP1 mutated gene into our retina-specific CKO line (Pax6-cre;Elp1flox/flox) to overcome these major challenges to the field. In so doing, we will generate a single mouse model that manifests the human FD optic neuropathy, in an otherwise healthy background, and contains the splice site mutation, which can be used to test a variety of therapeutic approaches (Aim1A, B). The overall aim of this proposal is to assess and compare two methods for restoring normal levels of the Elp1 protein in this new model mouse retinae using: (i) AAV2- mediated gene therapy (gene reintroduction) of the wild type Elp1 gene injected intravitreously, and (ii) a novel splicing enhancer compound that has been shown to promote the inclusion of exon 20 in the mutant FD gene in the retina, delivered orally through diet. Our goal is to test which method best mitigates the death of retinal ganglion cells in addition to interrogating whether a combination of both methods (Aim 1C) will have additive effects on promoting the survival of retinal ganglion cells, given they work via two distinct pathways.

项目摘要鉴于FDA最近获得了目标AAV基因治疗平台和小分子剪接的批准调节剂作为遗传神经系统疾病的治疗方法，我们的目标是将这些强大的技术应用于预防遗传隐性患者发展的进行性视神经病变和失明疾病，家族性障碍（FD）。 FD是由基因ELP1内含子20中的剪接位点突变引起的（以前称为Ikbkap）。由于错过的拼写，外显子20被多样跳过，突变体 mRNA降解，导致编码蛋白的水平降低，ELP1。根据组织类型进行突变的mRNA品种，神经元无法剪接突变的前的前神经元 mRNA。虽然大多数临床缺陷是由于感觉和自主神经的破坏神经系统，当患者进入青少年时，它们的黄斑视网膜神经节细胞逐渐死亡，表现为视觉损失。 ELP1无效的鼠标是胚胎致死的产生小鼠模型来研究FD的两种不同的策略：（i）有条件敲除小鼠的产生（CKO）使用细胞类型特异性CRE驱动的启动子；（ii）包含人FD ELP1的转基因小鼠剪接突变。前一种方法生成了鼠标模型，可以概括FD光学元件神经病是由于视网膜神经节细胞进行性死亡而导致的。这些老鼠是一个极好的预 - 测试基因疗法预防残留逐渐消失的临床模型神经节细胞（AIM 1A）。但是，该模型不适合测试剪接的有效性增强剂化合物由于缺乏FD剪接突变。后来的方法最终达到了转基因小鼠的产生，包括人FD ELP1突变基因的副本。这些老鼠是除非将它们越过hydomorph或null背景小鼠，否则无症状，但是这些复合小鼠通常太病了，无法持续调查。在这里，我们将通过越过人FD ELP1突变基因进入我们的视网膜特异性CKO系（PAX6-CRE； ELP1FLOX/FLOX）以克服这些主要对该领域的挑战。这样，我们将生成一个显示人类FD光学的单个鼠标模型神经病，在原本健康的背景下，并包含剪接部位突变，可用于测试各种治疗方法（AIM1A，B）。该提案的总体目的是评估和比较在这种新模型小鼠视网膜中恢复ELP1蛋白正常水平的两种方法使用：（i）AAV2- 静脉注射的野生型ELP1基因的介导基因治疗（基因重新引入），以及（ii）一种新颖的剪接增强剂化合物已被证明促进了外显子20在突变体FD基因中的包含在视网膜中，通过饮食口服。我们的目标是测试哪种方法最能减轻视网膜死亡神经节细胞除了询问是否会添加两种方法（AIM 1C）的组合促进视网膜神经节细胞存活的影响，鉴于它们通过两种不同的途径起作用。

项目成果

期刊论文数量（1）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Reduction of retinal ganglion cell death in mouse models of familial dysautonomia using AAV-mediated gene therapy and splicing modulators.

DOI：
10.1038/s41598-023-45376-w
发表时间：
2023-10-30
期刊：
SCIENTIFIC REPORTS
影响因子：
4.6
作者：
Schultz, Anastasia;Cheng, Shun-Yun;Kirchner, Emily;Costello, Stephanann;Miettinen, Heini;Chaverra, Marta;King, Colin;George, Lynn;Zhao, Xin;Narasimhan, Jana;Weetall, Marla;Slaugenhaupt, Susan;Morini, Elisabetta;Punzo, Claudio;Lefcort, Frances
通讯作者：
Lefcort, Frances