Genetic Approach to Therapy for DFNA9

DFNA9 的基因治疗方法

• 批准号: 10681990
• 负责人: Cynthia Casson Morton
• 金额: $ 77.41万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-20 至 2028-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10681990
• 关键词: Adult; Age; Alleles; Auditory system; Biological; Biology; Biopsy; Boston; CRISPR/Cas technology; Categories; Characteristics; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; DNA; Dermal; Development; Disease; Disease model; Early Intervention; Engineering; Enzymes; Equilibrium; Evaluation; Fibroblasts; Functional disorder; GJB2 gene; Gene Delivery; Gene Targeting; Genes; Genetic; Genetic Transcription; Goals; Guide RNA; Health; Hearing; Heterozygote; Histologic; Human; In Vitro; Individual; Injections; Knock-in; Knock-in Mouse; Laboratories; Labyrinth; Mediating; Mesenchymal; Messenger RNA; Methodology; Methods; Modeling; Mus; Mutation; Nonsense-Mediated Decay; Organism; Organoids; Pathogenicity; Pathology; Patients; Pediatric Hospitals; Phenotype; Plasmids; Pluripotent Stem Cells; Population; Procedures; Proteins; Protocols documentation; Reagent; Research; Resources; Safety; Sensorineural Hearing Loss; Severities; Site; Skin; Specificity; System; Technology; Temporal bone structure; Testing; Therapeutic; Therapeutic Intervention; Time; Tissue Sample; Tissues; Transcript; Transfection; Treatment Efficacy; Variant; cell type; deafness; design; design and construction; design verification; early onset; equilibration disorder; experimental study; gain of function; gain of function mutation; gene correction; gene discovery; gene therapy; genetic approach; hearing impairment; hereditary hearing loss; in vivo; induced pluripotent stem cell; inner ear development; innovation; insertion/deletion mutation; interest; mRNA Expression; mouse model; mutant; next generation sequencing; novel strategies; postnatal; preservation; prevent; progressive hearing loss; round window; safety assessment; success; tool development; transduction efficiency; translational goal; vector

ABSTRACT/PROJECT SUMMARY Impressive advances in gene discovery in the auditory system have occurred in the last decades, making specific targeted approaches for therapeutics realistic goals of great interest in the field of hearing and deafness. Hearing loss (HL) is an increasingly significant health problem in populations worldwide, with a substantial proportion due to genetic causes. Given the health burden and ongoing rapid discoveries, it is now essential to pursue new strategies for specific early interventions that could prevent or mitigate severity and progression of HL. One such disorder is DFNA9, an adult-onset sensorineural HL with balance dysfunction, caused by mutations in COCH, encoding cochlin, the most abundantly detected protein in the inner ear. This disease model is similar to and representative of the majority of genetic HL disorders with a dominant mode of inheritance and with a deleterious gain-of-function of the mutant protein. Aims 1 and 2 of this proposal involve utilization of the powerful and versatile CRISPR-Cas9 gene editing technology for specific targeting and disruption of the dominant missense pathogenic COCH variants p.G88E and p.A449T. We will utilize human fibroblasts from patients with these two variants, and derived pluripotent stem cells and organoids. These biological resources will serve as tools for the development of effective methods for allele-specific gene disruption of the COCH pathogenic variants, while leaving the normal allele intact and functional. The organoids will be assessed as a possible in vitro system for elucidating the biology of COCH aggregates pathognomonic of DFNA9 temporal bones. Furthermore, we will utilize two Coch knock-in (KI) mouse models with these variants for implementation of somatic tissue gene targeting in the inner ear. These approaches will establish methodologies for gene editing not only for DFNA9, but also for a broader category of other HL disorders with a dominant gain-of-function mechanism of pathology, with the ultimate goal of translation to clinical trials and therapeutic intervention.

摘要/项目摘要 在过去的几十年里，在听觉系统的基因发现方面取得了令人印象深刻的进展，使特定的 有针对性的治疗方法在听力和耳聋领域引起极大兴趣的现实目标。听证 损失(HL)是世界各地人口中一个日益严重的健康问题，其中相当大一部分是由于 与遗传因素有关。考虑到健康负担和持续的快速发现，现在至关重要的是寻求新的 可以预防或减轻HL严重程度和进展的特定早期干预策略。这样的一个 疾病是DFNA9，一种成人起病的感觉神经性HL伴平衡功能障碍，由Coch突变引起， 编码Cochlin的蛋白质是内耳中检测到最丰富的蛋白质。这种疾病模型类似于和 代表大多数遗传性HL疾病，具有显性遗传方式和有害的 突变蛋白的功能获得。 该提案的目标1和目标2涉及使用功能强大且功能多样的CRISPR-Cas9基因编辑 针对显性错义致病Coch变异体p.G88E的特异性靶向和干扰技术 和P.A449T。我们将利用来自这两个变异体患者的人成纤维细胞，并衍生出多能性。 干细胞和类器官。这些生物资源将成为开发有效方法的工具。 用于Coch致病变种的等位基因特异性基因中断，同时保持正常等位基因和 功能齐全。这些有机化合物将被评估为一种可能的体外系统，用于阐明COCH的生物学 聚集了DFNA9颞骨的病原体。此外，我们将使用两个可可敲入(KI)小鼠 利用这些变异体在内耳实施体细胞组织基因打靶的模型。这些 方法将不仅为DFNA9建立基因编辑方法，而且为更广泛的类别 其他以功能获得机制为主的HL障碍，最终目的是翻译 临床试验和治疗干预。