Liver-directed somatic gene correction rAAV system of regulatable Cas9/sgRNA

可调节Cas9/sgRNA的肝脏定向体细胞基因校正rAAV系统

• 批准号: 9322551
• 负责人: Wen Xue
• 金额: $ 47.74万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9322551
• 关键词: Adult; Alleles; Animal Model; Basic Science; Biotechnology; CRISPR/Cas technology; Capsid; Clinical; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Development; Disease; Future; Gene Mutation; Genes; Glutamates; Goals; Guide RNA; Hepatocyte; Hereditary Disease; Homologous Gene; Human; Knock-in Mouse; Laboratories; Length; Leukocyte Elastase; Liver; Lung; Lung diseases; Lysine; Maps; Mediating; Methods; Molecular; Mus; Mutation; Nature; Nonhomologous DNA End Joining; Patients; Phenotype; Point Mutation; Production; Protease Inhibitor; Pulmonary Emphysema; RNA Interference; Recombinant adeno-associated virus (rAAV); Respiratory physiology; Safety; Serotyping; Serum; Somatic Cell; Staphylococcus aureus; System; Testing; Therapeutic; Therapeutic Studies; Tissues; Translating; Work; Yin; adeno-associated viral vector; alpha 1-Antitrypsin; alpha 1-Antitrypsin Deficiency; arm; aspergillopepsin II; c new; clinical application; clinically relevant; deep sequencing; design; gene correction; gene function; gene therapy; genome editing; homologous recombination; improved; in vivo; in vivo Model; innovation; knock-down; mouse model; mutant; novel; novel strategies; pre-clinical; preclinical study; repaired; restoration; small hairpin RNA; tool; vector

Project Summary/Abstract – Project 3: The ability to correct disease gene mutations in vivo has broad potential utility for both therapy and basic research. CRISPR/Cas9 is a powerful RNA-guided tool for genome editing. Our recent discovery that CRISPR/Cas9 delivery can cure genetic disease in adult mouse liver provided proof-of-concept of gene correction therapy. This subproject will interact synergistically with all other Projects and Cores of this tPPG to develop new rAAV CRISPR tools to treat alpha- 1 antitrypsin deficiency (AATD). The main goal of this proposal is to establish a pre-clinical rAAV paradigm for CRISPR-mediated correction of AAT deficiency in mouse models carrying the mutant human AAT gene. The impact of this project is to develop somatic gene correction using rAAV systems to (1) maximize efficiency and safety of CRISPR delivery, (2) maximize the rate of homologous recombination for gene correction, and (3) efficiently correct AAT mutation in the liver to treat lung disease in mice. The development of safe and effective delivery vehicles and genome editing tools to correct AAT deficiency will guide future clinical trials for CRISPR- mediated gene therapy for AAT lung disease. Because AAV serotypes can target a wide range of tissues, our approach has broad basic research and clinical applications beyond AATD. Project 3 has three Aims that focus on different aspects of liver-directed somatic AAT correction: Aim 1: Develop liver-directed rAAV vehicles to maximize efficiency and precision of Z-AAT genome-editing in mice. Aim 2: Investigate rAAV HDR templates to correct Z-AAT mutation in mouse liver. Aim 3: Explore Z-AAT correction in mouse models in vivo to treat the lung disease.

项目摘要/摘要-项目3： 体内纠正疾病基因突变的能力对这两种疾病都有广泛的潜在用途 治疗和基础研究。CRISPR/Cas9是一个强大的RNA引导的基因组编辑工具。 我们最近发现CRISPR/Cas9传递可以治愈成年小鼠肝脏的遗传病 提供了基因矫正治疗的概念验证。这一子项目将协同互动。 与该tPPG的所有其他项目和核心一起开发新的rAAV CRISPR工具，以治疗阿尔法- 1抗胰蛋白酶缺乏(AATD)。这项建议的主要目标是建立临床前rAAV CRISPR介导的纠正携带AAT基因小鼠模型中AAT缺乏的范例 突变的人类AAT基因。这个项目的影响是发展体细胞基因校正使用 RAAV系统以(1)最大限度地提高CRISPR交付的效率和安全性，(2)最大限度地提高 同源重组用于基因校正，以及(3)有效地校正AAT突变 用肝治疗小鼠肺部疾病。发展安全有效的运输工具和 纠正AAT缺乏症的基因组编辑工具将指导CRISPR的未来临床试验- AAT肺病的介导性基因治疗因为AAV血清型的靶标范围很广 在组织方面，我们的方法在AATD之外具有广泛的基础研究和临床应用。 项目3有三个目标，分别关注肝脏导向的体细胞AAT矫正的不同方面： 目的1：开发肝脏导向的rAAV载体以最大限度地提高Z-AAT的效率和精度 小鼠的基因组编辑。目的2：研究rAAV HDR模板以纠正Z-AAT突变 老鼠的肝脏。目的：探讨Z-AAT在小鼠体内校正治疗肺部疾病的作用。