Somatic Repeat Expansions as a Therapeutic Target for Trinucleotide Repeat Disorders

体细胞重复扩增作为三核苷酸重复疾病的治疗靶点

基本信息

批准号：
10605261
负责人：
Ricardo Mouro Pinto
金额：
$ 41.28万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-15 至 2027-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10605261
关键词：
Address Affect Alleles Antisense Oligonucleotides Biological Markers Biopsy Brain CAG repeat CRISPR therapeutics Candidate Disease Gene Cell model Cells Child Clustered Regularly Interspaced Short Palindromic Repeats Corpus striatum structure DNA Repair Gene DNA Repair Pathway DNA Repeat Expansion DNA Sequence Alteration Development Disease Disease Progression Drug Targeting Endonuclease I Exhibits Expanded DNA Repeat Fibroblasts Friedreich Ataxia Genes Genetic Goals Heart Human Huntington Disease Investigation Knock-out Knowledge Length Liver Mendelian disorder Methodology Methods Modeling Mus Mutation Nerve Degeneration Neurodegenerative Disorders Neuromuscular Diseases Neurons Oligonucleotides Onset of illness Parents Pathogenicity Pathway interactions Patients Phenotype Process RNA Splicing Reagent Reporting Research Role Sampling Spinal Ganglia System Testing Therapeutic Therapeutic Intervention Time Tissues Translating Trinucleotide Repeat Expansion Trinucleotide Repeats Validation base editor candidate validation clinical phenotype effective therapy efficacy testing endonuclease experimental study genome wide association study in vivo insight intergenerational minimally invasive mouse model new therapeutic target novel novel strategies novel therapeutics potential biomarker promoter stool sample therapeutic target tool

Address Affect Alleles Antisense Oligonucleotides Biological Markers Biopsy Brain CAG repeat CRISPR therapeutics Candidate Disease Gene Cell model Cells Child Clustered Regularly Interspaced Short Palindromic Repeats Corpus striatum structure DNA Repair Gene DNA Repair Pathway DNA Repeat Expansion DNA Sequence Alteration Development Disease Disease Progression Drug Targeting Endonuclease I Exhibits Expanded DNA Repeat Fibroblasts Friedreich Ataxia Genes Genetic Goals Heart Human Huntington Disease Investigation Knock-out Knowledge Length Liver Mendelian disorder Methodology Methods Modeling Mus Mutation Nerve Degeneration Neurodegenerative Disorders Neuromuscular Diseases Neurons Oligonucleotides Onset of illness Parents Pathogenicity Pathway interactions Patients Phenotype Process RNA Splicing Reagent Reporting Research Role Sampling Spinal Ganglia System Testing Therapeutic Therapeutic Intervention Time Tissues Translating Trinucleotide Repeat Expansion Trinucleotide Repeats Validation base editor candidate validation clinical phenotype effective therapy efficacy testing endonuclease experimental study genome wide association study in vivo insight intergenerational minimally invasive mouse model new therapeutic target novel novel strategies novel therapeutics potential biomarker promoter stool sample therapeutic target tool

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项目摘要

ABSTRACT Huntington’s disease (HD) and Friedreich ataxia (FA) are rare neurodegenerative diseases caused by expanded trinucleotide repeats (CAG and GAA, respectively) in the HTT and FXN genes, respectively, with larger alleles being associated with earlier disease onset and more severe clinical phenotypes. Despite these being single gene disorders, where the respective underlying genetic mutations have been known for over 20 years, there remains no cure or disease-modifying therapies, indicating that novel approaches are critical. A hallmark of most repeat expansion disorders is that the repeats are highly unstable, both intergenerationally (parent to child) and in somatic tissues, where the repeat expands progressively over time in a cell-/tissue-specific manner. Notably, in HD, medium-spiny neurons of the striatum, which succumb most severely to the effects of the HTT mutation, exhibit the most dramatic CAG expansions. Similarly, larger GAA repeat expansions have been reported in the heart and dorsal root ganglia of FA patients, where such tissues are most severely affected. These observations, together with growing evidence from GWAS and candidate gene association studies in HD patients, support the hypothesis that progressive repeat length increases in somatic tissues contribute to the pathogenic process. Thus, understanding the roles of disease modifiers in somatic repeat expansion may provide novel targets for therapeutic intervention directed at the repeat mutation itself. To that end, we have leveraged a CRISPR-based in vivo system, recently developed by us, to screen a number of candidate DNA repair genes and determine their role as potential modifiers of somatic CAG repeat instability in HD. Remarkably, this has resulted in the identification of novel genes, which when knocked out in the liver of HD mice, reduced CAG expansions and promoted contractions. We hereby propose a set of experiments aimed at: 1) Identifying non-invasive samples to study CAG repeat instability as a potential biomarker of disease, as well as developing novel long-read sequencing-based methodologies to more accurately size and quantify repeat instability; 2) Validating candidate modifier genes in a new model of CAG expansions using HD patient-derived fibroblasts, recently developed by us, as well as understand the potential adverse implications associated with inactivating such DNA repair genes. We also propose to investigate if such genes are equally involved in FA GAA expansions, using the same in vivo CRISPR platform and patient derived cellular models; 3) Development and testing of novel antisense oligonucleotide- and CRISPR-based therapeutic approaches targeting the repeat expansion process to suppress repeat expansions or actually promote contractions. This will lead to a better understanding of shared mechanisms across these diseases and potentially result in novel therapeutics that can be used in all repeat expansion disorders.

抽象的亨廷顿氏病（HD）和弗里德里希共济失调（FA）是罕见的神经退行性疾病通过在HTT和FXN基因中分别扩展的三核苷酸重复序列（CAG和GAA），分别较大的等位基因与较早的疾病发作和更严重的临床有关表型。尽管这些是单个基因疾病，其中相对基础遗传突变已有20多年了，尚无治愈或修改疾病的疗法，表明新方法至关重要。大多数重复扩张障碍的标志是重复次数是高度不稳定的，既有代间（父母对儿童）和在体细胞组织中，重复以细胞/组织特异性方式逐渐扩展。值得注意的是，在高清中纹状体的神经元最严重地屈服于HTT突变的作用，暴露了最戏剧性的CAG扩展。同样，据报道了较大的GAA重复扩展 FA患者的心脏和背根神经节，这些组织受到最严重影响。这些观察结果，以及GWAS和候选基因关联研究的越来越多的证据高清患者，支持以下假设有助于致病过程。这是了解疾病修饰剂在躯体中的作用重复扩展可能为针对重复突变的治疗干预提供新的靶标本身。为此，我们利用了一个基于CRISPR的体内系统，该系统最近由我们开发，用于筛选许多候选DNA修复基因，并确定其作为潜在修饰符的作用体细胞CAG在高清中重复不稳定性。值得注意的是，这导致了新基因的鉴定，当在HD小鼠的肝脏中撞倒时，CAG膨胀并促进了收缩。我们在此提出了一组针对以下实验：1）识别非侵入性样本以研究 CAG重复不稳定性作为潜在的疾病生物标志物，并发展了新的长阅读基于测序的方法更准确地大小并量化重复不稳定性； 2）验证使用HD患者衍生的成纤维细胞的新型CAG扩展模型中的候选修饰符基因，我们最近开发的，以及了解与灭活这种DNA修复基因。我们还建议调查此类基因是否同样涉及在FA GAA扩展中，使用相同的体内CRISPR平台和患者衍生的细胞模型； 3）新型反义寡核苷酸和基于CRISPR的疗法的开发和测试针对重复扩展过程以抑制重复扩展或实际促进的方法收缩。这将使人们更好地了解这些疾病中的共同机制，并可能导致新的疗法可用于所有重复扩张障碍。