Controllable base editing therapy for DMD

DMD 的可控碱基编辑疗法

• 批准号: 10728698
• 负责人: Renzhi Han
• 金额: $ 59.95万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10728698
• 关键词: Adenine; Animal Model; Animals; Antigen-Presenting Cells; Basal lamina; Binding Sites; Biology; Birth; CRISPR/Cas technology; Capsid; Cardiac; Cardiomyopathies; Cells; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; Complex; Cytoskeleton; DNA; DNA Double Strand Break; DNA Sequence Alteration; Data; Degenerative Disorder; Dependovirus; Disease; Dose; Duchenne muscular dystrophy; Dystrophin; Elements; Embryo; Engineering; Event; Exons; Frequencies; Friends; Gene Expression; Gene Rearrangement; Genes; Genetic; Genomics; Germ Cells; Glycoproteins; Guide RNA; Heart failure; Human; Immune; Immune Tolerance; Immune response; Immune system; Immunity; Knowledge; Light; Longevity; Mediating; MicroRNAs; Modeling; Monitor; Mus; Muscle; Muscle function; Muscular Dystrophies; Muscular dystrophy cardiomyopathy; Mutation; Myocardial dysfunction; Myocardium; Myopathy; Nonsense Mutation; Oryctolagus cuniculus; Patients; Pharmaceutical Preparations; Play; Point Mutation; Positioning Attribute; Proteins; Reading Frames; Research; Residual state; Respiratory physiology; Role; Safety; Skeletal Muscle; Specificity; Steroids; Striated Muscles; System; TANK-binding kinase 1; TLR9 gene; Technology; Therapeutic; Transcript; Treatment Efficacy; Tumorigenicity; Viral Genome; Viral Packaging; Viral Vector; adeno-associated viral vector; base; base editing; base editor; boys; clinical translation; combat; design; disease-causing mutation; efficacy evaluation; experimental study; genome editing; genotoxicity; heart function; human disease; immunoengineering; immunogenic; immunomodulatory strategy; improved; in vivo; insight; male; manufacture; mortality; mouse model; mutant; novel; novel strategies; pharmacologic; precision medicine; repaired; respiratory; restoration; therapeutic development; therapeutic genome editing; transcriptomics; transduction efficiency; transgene expression; vector; ventilation

PROJECT SUMMARY/ABSTRACT Duchenne muscular dystrophy (DMD) is a lethal muscle degenerative disease with cardiomyopathy in over 90% of patients, and heart failure is a leading cause of mortality. DMD patients commonly harbor out-of-frame mutations which result in complete loss of dystrophin protein. While CRISPR-gene editing has been employed to delete the mutant exon for restoration of the dystrophin reading frame, this strategy raises potential safety concerns as it relies on repair of the double strand DNA breaks created by CRISPR/Cas9, which may cause unwanted large deletion, DNA rearrangement and viral vector integration. Developing novel approaches to precisely and safely correct the disease-causing mutations in striated muscles is in urgent need to combat DMD. Recent advances in base editors allow us to explore the feasibility of precise correction of genetic mutations in animal models of DMD without creating double strand DNA breaks. Our recent studies demonstrate it is highly efficient to correct a disease-causing point mutation in the striated muscles of a mouse model of DMD using systemic delivery of adeno-associated virus 9 (AAV9). This paves the way for clinical translation of in vivo base editing for DMD. However, novel strategies are in urgent need to solve several safety-related issues (e.g. the accumulation of off-target activities with persistent expression of base editing agents following AAV delivery; high dose of AAV utilized; host immune responses). Here we propose to develop a novel controllable base editing system (to reduce accumulation of off-target editing events and exposure of base editor to the host immune system) delivered in recently engineered myotropic AAV capsids (to increase muscle transduction efficiency with 10-fold less of AAV as compared to AAV9). The therapeutic efficacy and safety will be extensively investigated using a newly established DMD rabbit model, which faithfully recapitulates the clinical signs of muscular dystrophy and cardiomyopathy in human DMD. We will further engineer immune-friendly vectors and develop a novel immune-modulating strategy to mitigate the host immune responses to the in vivo AAV-delivered base editing therapy. Completion of the proposed studies will significantly advance our translational efforts to develop safer precision medicine for DMD.

项目概要/摘要 杜氏肌营养不良症 (DMD) 是一种致命的肌肉退行性疾病，90% 以上会出现心肌病 心力衰竭是患者死亡的主要原因。 DMD 患者通常存在框架外的问题 导致肌营养不良蛋白完全丧失的突变。虽然 CRISPR 基因编辑已被采用 为了删除突变外显子以恢复肌营养不良蛋白阅读框，该策略提高了潜在的安全性 担心，因为它依赖于 CRISPR/Cas9 产生的双链 DNA 断裂的修复，这可能会导致 不需要的大缺失、DNA 重排和病毒载体整合。开发新方法 对抗 DMD 迫切需要精确、安全地纠正横纹肌的致病突变。 碱基编辑器的最新进展使我们能够探索精确校正基因突变的可行性。 DMD 动物模型，不产生双链 DNA 断裂。我们最近的研究表明它是高度 使用该方法可有效纠正 DMD 小鼠模型横纹肌中的致病点突变 腺相关病毒 9 (AAV9) 的全身递送。这为体内基础的临床转化铺平了道路 DMD 编辑。然而，迫切需要新的策略来解决一些与安全相关的问题（例如 AAV 递送后，随着碱基编辑剂持续表达，脱靶活性不断累积； 使用高剂量的 AAV；宿主免疫反应）。在这里我们建议开发一种新型的可控底座 编辑系统（以减少脱靶编辑事件的积累以及碱基编辑器向宿主的暴露 免疫系统）在最近设计的肌向性 AAV 衣壳中传递（以增加肌肉转导 与 AAV9 相比，AAV 的效率低 10 倍）。治疗效果和安全性将受到广泛关注 使用新建立的 DMD 兔模型进行研究，该模型忠实地再现了 DMD 的临床症状 人类 DMD 中的肌营养不良症和心肌病。我们将进一步设计免疫友好型载体并 开发一种新的免疫调节策略来减轻宿主对体内 AAV 传递的免疫反应 碱基编辑疗法。完成拟议的研究将显着推进我们的转化工作 为 DMD 开发更安全的精准医疗。