Precision base editing for the treatment of motor neuron diseases

精准碱基编辑治疗运动神经元疾病

• 批准号: 10301562
• 负责人: Mandana Arbab
• 金额: $ 12.65万
• 依托单位: BROAD INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10301562
• 关键词: Address; Adult; Affect; Alleles; Amyotrophic Lateral Sclerosis; Animal Model; Antisense Oligonucleotides; Award; BCAR1 gene; Cell Death; Chimeric Proteins; Clinic; Communication; Complex; DNA Sequence Alteration; Data; Deaminase; Development; Disease; Disease model; Disease-Free Survival; Environment; Enzymes; Faculty; Foundations; Funding; Future; Genes; Genetic; Genome; Genomics; Genotype; Goals; Grant; Human; In Vitro; Individual; Injections; Institutes; Late-Onset Disorder; Learning; Libraries; Machine Learning; Mammalian Cell; Mentorship; Methods; Minor; Modeling; Molecular Abnormality; Motor Neuron Disease; Motor Neurons; Mus; Mutation; Neonatal; Neuraxis; Neurodegenerative Disorders; North America; Nucleotides; Other Genetics; Outcome; Pathogenicity; Patients; Peripheral; Phenotype; Point Mutation; Positioning Attribute; Procedures; Protocols documentation; Publishing; Research; Research Personnel; Route; Safety; Science; Serotyping; Spinal Muscular Atrophy; System; Techniques; Therapeutic; Therapeutic Intervention; Tissues; Translating; Treatment Efficacy; United States National Institutes of Health; Validation; Variant; Work; Writing; base; causal variant; community based research; data visualization; design; disease phenotype; disease-causing mutation; embryonic stem cell; experimental study; familial amyotrophic lateral sclerosis; genetic variant; genome editing; genomic locus; improved; in vivo; meetings; member; molecular pathology; molecular phenotype; mouse model; mutant; novel; predictive modeling; predictive test; programs; progressive neurodegeneration; screening; skills; success; symposium; therapeutic development; therapeutic genome editing; tool; transversion mutation

Motor neuron diseases (MNDs) such as spinal muscular atrophy (SMA) and amyotrophic lateral sclerosis (ALS) are a class of progressive neurodegeneration disorders, often caused by minor genetic abnormalities to which motor neurons are particularly vulnerable. Because each of these diseases is rare and the molecular pathologies so diverse and unresolved, it is hard to envision a therapeutic intervention tailored to each individually, or a single treatment to effectively treat them all. Current genome editing tools are easily programmable to target discrete genomic loci and affected tissues of MNDs are largely similar. Thus, a general genome editing therapeutic strategy for MNDs fitted to each mutation could meet this urgent need. Base editing tools can theoretically correct any C:G>T:A and A:T>G:C transition, yet the factors that determine efficiency and precision of base editing are not fully understood, and editing outcomes at a given locus are frequently unpredictable. For the development of base editing correction strategies at a great number of loci, screening through a multitude of base editor variants – currently any permutation of >10 deaminase enzymes and >15 Cas proteins, and counting – and sgRNA combinations for every target is prohibitive. A clear understanding of Cas protein, deaminase, and sequence determinants of editing outcomes is needed to facilitate the design of base editing strategies. We intend to develop a general workflow to design effective base editing strategies for causal MND SNPs and deliver these tools to MND affected tissues. SMA is a monogenic MND with a well-defined genetic cause, and animal models harboring the human causal gene that faithfully recapitulate disease phenotypes of SMA patients. Successful development of an effective and safe SMA genome editing treatment will assist the development of similar genome editing therapeutics for other genetic MNDs, including some forms of familial ALS. In this project we will (1) create a computational predictive model of base editing to facilitate the design of effective base editing strategies; (2) develop protocols to efficiently deliver base editing therapeutics to disease relevant tissues in mice to enable genome editing; and (3) use this pipeline to optimize a base editing therapeutic to rescue SMA in mice, and develop genome editing therapeutics for other causal MND mutations, focusing first on the most common single point mutation causal to ALS in North America, SOD1A4V. I will take advantage of the world-class genomics environment of the Broad Institute and the expertise in AAV-delivery at the Stanley Center for Psychiatric Research to realize these goals and develop skills that will help me to continue this work as an independent investigator. Through mentorship meetings and courses on grant-writing and data visualization I will improve my science-communication skills so that I may compete successfully for additional NIH R01 and R21 funding as a faculty member. By attending conferences and publishing my work I intend to establish myself as leader in the field of therapeutic genome editing for MND.

运动神经元病(MND)，如脊髓性肌萎缩症(SMA)和肌萎缩侧索硬化症(ALS)，是一类进行性神经退行性疾病，通常由运动神经元特别容易受到影响的轻微遗传异常引起。由于这些疾病中的每一种都很罕见，分子病理又如此多样和悬而未决，很难想象有一种针对每一种疾病的治疗干预措施，或者一种有效地治疗所有疾病的单一疗法。目前的基因组编辑工具很容易被编程来针对离散的基因组座位，并且受影响的MND组织基本上是相似的。因此，一种适用于每个突变的针对MND的通用基因组编辑治疗策略可以满足这一迫切需求。基本编辑工具理论上可以纠正任何C：G&gt；T：A和A：T&gt；G：C转换，但决定基本编辑效率和精度的因素尚未完全了解，并且在给定位置的编辑结果往往是不可预测的。对于在大量基因座上开发碱基编辑校正策略，通过大量碱基编辑变体-目前是&gt；10脱氨酶和&gt；15蛋白的任何排列和计数-以及每个靶标的sgRNA组合进行筛选是令人望而却步的。为了促进碱基编辑策略的设计，需要对编辑结果的CaS蛋白、脱氨酶和序列决定因素有一个清楚的了解。我们打算开发一个通用的工作流程，为因果MND SNP设计有效的碱基编辑策略，并将这些工具提供给受MND影响的组织。SMA是一种单基因MND，具有明确的遗传原因，并且动物模型含有人类因果基因，忠实地概括了SMA患者的疾病表型。有效和安全的SMA基因组编辑疗法的成功开发将有助于开发类似的基因组编辑疗法来治疗其他遗传性MND，包括某些形式的家族性ALS。在本项目中，我们将(1)创建碱基编辑的计算预测模型，以便于设计有效的碱基编辑策略；(2)开发协议，高效地向小鼠的疾病相关组织提供碱基编辑疗法，以实现基因组编辑；以及(3)利用这一管道来优化碱基编辑疗法，以挽救小鼠的SMA，并针对其他原因MND突变开发基因组编辑疗法，首先关注北美最常见的导致ALS的单点突变SOD1A4V。我将利用布罗德研究所世界级的基因组学环境和斯坦利精神病学研究中心在AAV交付方面的专业知识来实现这些目标，并发展技能，帮助我作为一名独立研究员继续这项工作。通过导师会议和关于拨款撰写和数据可视化的课程，我将提高我的科学交流技能，这样我就可以作为一名教员成功地竞争NIH R01和R21的额外资金。通过参加会议和发表我的工作，我打算在MND的治疗性基因组编辑领域确立自己的领导者地位。