Zinc-Finger Nuclease- Mediated Repair of Disease-Causing Triplet CAG Repeats

锌指核酸酶介导的致病三联体 CAG 重复序列的修复

• 批准号: 8359421
• 负责人: David A Mittelman
• 金额: $ 19.26万
• 依托单位: VIRGINIA POLYTECHNIC INST AND ST UNIV
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8359421
• 关键词: Affect; Alleles; Animal Model; Benign; Biological Assay; CAG repeat; Cells; Cessation of life; Contracts; Corpus striatum structure; Disease; Etiology; Event; Exons; Future; Genes; Genetic; Genome; Genomics; Goals; Human; Huntington Disease; In Vitro; Individual; Inherited; Lead; Length; Mediating; Methods; Modeling; Mutation; Myotonic Dystrophy; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Onset of illness; Pathology; Patients; Population; Reagent; Reporting; Research; Rodent; Severities; Specificity; Spinocerebellar Ataxias; Testing; Therapeutic; Toxic effect; Trinucleotide Repeats; Triplet Multiple Birth; Zinc Fingers; base; improved; induced pluripotent stem cell; nervous system disorder; neurotoxicity; next generation; novel therapeutics; nuclease; polyglutamine; repaired; research study; therapy development; treatment strategy; validation studies

DESCRIPTION (provided by applicant): Huntington's disease (HD) is a variable onset neurodegenerative disorder caused by the expansion of a CAG repeat sequence in the first exon of the HTT gene. The repeat is polymorphic, especially in the striatum and cortex; and encodes a polyglutamine tract that is the pathological basis for neuronal toxicity and death. There is currently no cure for this devastating disorder. CAG repeat length in this dominant disorder principally determines the onset and severity of HD starting with repetitions that exceed 35-39 CAG repeats. Reducing neuronal toxicity and death by directly targeting the CAG repeat tract would be expected to reduce the severity or at least delay the onset of HD and other triplet repeat disorders. The proposed studies focus directly on the expanded CAG repeat tract as the target of therapy and seek to permanently disrupt or shrink the expanded CAG repeat at the genomic level. To achieve this goal, zinc finger nucleases (ZFNs) will be used to direct DSBs specifically to the expanded HTT repeat allele, with the intent of either shrinking the repeat to benign lengths or eliminating its expression through the induction of frame-shifting indels. In the first aim, this ZFN- based therapy will be tested in primary cells derived from a Huntington's patient. Next-generation sequencing will be used to determine the full spectrum of ZFN-induced changes to the HTT repeat. In conclusion, the proposed research will test key aspects of a ZFN-mediated treatment strategy for shrinking or disrupting expanded CAG repeat alleles that induce neurodegeneration and lead to Huntington's disease and other neurological disorders. PUBLIC HEALTH RELEVANCE: Expanded CAG/CTG repeat tracts are the genetic basis for more than a dozen inherited neurological disorders including Huntington's disease, myotonic dystrophy, and several spinocerebellar ataxias. Despite the multitude of pathologies underlying these disorders, they all share common etiology: the expansion of CAG/CTG repeats from short, benign tracts of about 30 repeats to longer, pathogenic lengths that can extend for several hundred repeats. We will validate a novel therapeutic strategy that targets this common feature-expanded repeats-in human patient cells and in animal models for Huntington's disease.

描述（由申请人提供）：亨廷顿氏病（HD）是由HTT基因的第一个外显子扩展引起的可变发作神经退行性疾病。重复是多态性的，尤其是在纹状体和皮层中。并编码一个聚谷氨酰胺道，这是神经元毒性和死亡的病理基础。目前无法治愈这种毁灭性疾病。 CAG重复长度在这种主要障碍中主要决定了HD的发作和严重程度，从重复超过35-39 CAG重复开始。预计通过直接靶向CAG重复道来降低神经元毒性和死亡，将减少严重程度或至少延迟HD和其他三重态重复​​疾病的发作。拟议的研究直接关注扩展的CAG重复道作为治疗的靶标，并试图在基因组水平上永久破坏或收缩扩展的CAG重复。为了实现这一目标，锌指核酸酶（ZFN）将用于将DSB专门引向扩展的HTT重复等位基因，其目的是将重复重复缩小到良性长度或通过诱导帧换档Indels的诱导来消除其表达。在 第一个目的是，这种基于ZFN的疗法将在来自亨廷顿患者的原代细胞中进行测试。下一代测序将用于确定ZFN诱导的HTT重复变化的全频谱。总之，拟议的研究将测试ZFN介导的治疗策略的关键方面，以缩小或破坏扩大的CAG重复等位基因，从而诱导神经变性并导致亨廷顿氏病和其他神经系统疾病。 公共卫生相关性：扩大的CAG/CTG重复区是十多种遗传性神经系统疾病的遗传基础，包括亨廷顿氏病，肌发育症和几种脊椎小脑共济失调。尽管这些疾病的基本病理众多，但它们都具有共同的病因：CAG/CTG重复的膨胀，从短的，良性的良性区域重复约30次重复到更长的致病长度，可以扩展数百个重复。我们将验证一种新型的治疗策略，该策略针对这种常见的特征膨胀的人类患者细胞以及亨廷顿氏病的动物模型。