Optimizing and validation of gene therapy vectors to treat limb girdle muscular dystophy

治疗肢带型肌营养不良症的基因治疗载体的优化和验证

• 批准号: 10400144
• 负责人: JEFFREY S CHAMBERLAIN
• 金额: $ 49.6万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10400144
• 关键词: Affect; Animal Model; Antibodies; Area; Binding Sites; Calpain; Cardiac; Cardiotoxicity; Clinical Trials; Code; Codon Nucleotides; Collaborations; Complementary DNA; Development; Diagnosis; Disease; Dose; Duchenne muscular dystrophy; Dystrophin; Exercise Tolerance; Fiber; Gene Delivery; Gene Expression; Gene Transduction Agent; Gene therapy trial; Genes; Goals; Heart; Heart Block; Human; International; Knockout Mice; Knowledge; Limb structure; Limb-Girdle Muscular Dystrophies; Longitudinal Studies; MicroRNAs; Modification; Mus; Muscle; Muscle Weakness; Muscular Atrophy; Muscular Dystrophies; Mutation; Myocardium; Neuromuscular Diseases; Outcome; Outcome Measure; Pathogenesis; Patients; Peptide Hydrolases; Phenotype; Physiological; Property; Proteins; Series; Skeletal Muscle; Testing; Therapeutic; Therapeutic Effect; Therapeutic Intervention; Time; Toxic effect; Transgenes; Treatment Efficacy; Universities; Untranslated Regions; Validation; Variant; Viral Vector; Washington; Wasting Syndrome; Western Blotting; Wheelchairs; Work; adeno-associated viral vector; design; differential expression; experience; gene therapy; gene therapy clinical trial; in vitro testing; in vivo; laser capture microdissection; member; micro-dystrophin; mini-dystrophin; mitochondrial dysfunction; mouse model; novel; overexpression; pre-clinical; safety testing; therapeutic evaluation; vector

PROJECT SUMMARY We aim to develop a gene therapy for limb girdle muscular dystrophy type 2A (LGMD2A), an autosomal recessive (AR) muscle wasting disorder due to mutations in CAPN3. LGMD2A is considered to be the most prevalent of the AR LGMDs and yet there is currently no treatment for patients, who are usually wheelchair dependent a decade after diagnosis. We and others have shown that overexpression of CAPN3 can be accomplished in skeletal muscle without toxicity; a finding which makes the feasibility of gene therapy for LGMD2A a realistic goal; however, LGMD2A is unique from most other LGMDs and this fact warrants careful development of gene therapy vectors. One consideration is that CAPN3 cannot be expressed in the heart, due to cardiac toxicity, which is not the case with Duchenne muscular dystrophy and other LGMDs. Although a few pre-clinical, proof of concept studies have successfully accomplished AAV-Capn3 overexpression in mice, there has not been a systematic optimization of any gene therapy construct for humans with LGMD2A, especially one that considers the relative skeletal muscle vs cardiac gene expression issues. Furthermore, because LGMD2A preferentially impacts slow fibers, it is critical that the therapeutic construct that is ultimately carried into clinical trials is optimized for slow fiber expression. Because each patient can only be dosed one time, it is imperative that the construct used for gene delivery is ideal. In this application, we will use an iterative and systematic approach to optimize these vectors. We have assembled an expert, collaborative team with deep experience in the area of regulatory cassettes, AAV vector development and LGMD2A pathogenesis and calpainopathy animal models. The team will work together to generate and test the safety and efficacy of a series of AAV vectors to develop a treatment for LGMD2A. These vectors will be optimized for slow fiber expression while avoiding cardiac toxicity. One team member, Dr. Hauschka, is largely credited with creating the vast majority of regulatory cassettes being used in the current AAV-gene therapy trials for Duchenne muscular dystrophy. Dr. Chamberlain designed and optimized the first micro and mini dystrophins, which formed the basis for all constructs currently in gene therapy clinical trials for DMD. Drs. Spencer and Kramerova generated numerous mouse models that have led to the identification of outcome measures useful for testing therapeutic interventions for LGMD2A. Dr. Cannon, is an internationally known muscle physiologist who will carry out physiological assessments of contractile function. The team will apply their extensive and cumulative knowledge of LGMD2A, AAV vectors, regulatory cassettes and mouse muscle testing to create this gene therapy for LGMD2A.

项目概要 我们的目标是开发一种针对 2A 型肢带型肌营养不良症 (LGMD2A) 的基因疗法，这是一种常染色体隐性遗传病 (AR) 由于 CAPN3 突变导致的肌肉萎缩症。 LGMD2A 被认为是最普遍的 AR LGMD，但目前尚无针对通常依赖轮椅的患者的治疗方法 诊断后十年。我们和其他人已经证明 CAPN3 的过度表达可以在 骨骼肌无毒性；这一发现使 LGMD2A 基因治疗的可行性成为现实 目标;然而，LGMD2A 与大多数其他 LGMD 不同，这一事实需要仔细开发基因 治疗载体。一种考虑是 CAPN3 由于心脏毒性而不能在心脏中表达， 但杜氏肌营养不良症和其他 LGMD 的情况并非如此。尽管有一些临床前证明 概念研究已成功实现AAV-Capn3在小鼠体内的过表达，但目前还没有 对 LGMD2A 人类的任何基因治疗结构进行系统优化，尤其是考虑到 相对的骨骼肌与心脏基因表达问题。此外，由于 LGMD2A 优先 影响慢纤维，因此最终进入临床试验的治疗结构至关重要 针对慢纤维表达进行了优化。由于每位患者只能服药一次，因此必须 用于基因传递的构建体是理想的。在此应用程序中，我们将使用迭代和系统的方法来 优化这些向量。我们组建了一支在以下领域拥有丰富经验的专家协作团队 调节盒、AAV 载体开发和 LGMD2A 发病机制和钙蛋白酶病动物模型。 该团队将共同生成并测试一系列AAV载体的安全性和有效性，以开发 LGMD2A 的治疗。这些载体将针对慢纤维表达进行优化，同时避免心脏毒性。 Hauschka 博士是一名团队成员，他因创造了绝大多数监管盒而受到赞誉。 用于当前杜氏肌营养不良症的 AAV 基因治疗试验。张伯伦博士设计并 优化了第一个微型和迷你抗肌营养不良蛋白，这构成了目前基因治疗中所有构建体的基础 DMD 的临床试验。博士。 Spencer 和 Kramerova 制作了许多小鼠模型，这些模型导致了 确定可用于测试 LGMD2A 治疗干预措施的结果测量。卡农博士是一位 国际知名的肌肉生理学家将对收缩功能进行生理评估。 该团队将运用他们在 LGMD2A、AAV 载体、监管盒方面广泛积累的知识 和小鼠肌肉测试来创建 LGMD2A 基因疗法。