Systemic molecular therapy for muscular dystrophy

描述（由申请人提供）：本项目的总体目标是解决杜氏肌营养不良症治疗开发中的关键限速步骤：系统性基因“递送”。我们专注于直接基因转移的方法来解决这个问题，特别是避免任何依赖于慢性免疫抑制。我们建立在来自多个实验室的在营养不良小鼠中使用类似腺相关病毒（AAV）载体的令人信服的概念验证研究以及我们先前的发现的基础上，即在逆行输注期间载体的强制外渗可以用作在非营养不良的大型动物中有效地使骨骼肌和心肌均变性的手段。我们推测，载体运输的潜在机制依赖于压力诱导的，可逆的小静脉内皮细胞的尺度依赖性屏障功能的改变。我们证明，这种假设的机制是一致的，与观察到的高效的全球模式的基因转移到肌肉在全身，逆行输注在设置深低温和球囊闭塞的大血管。提出的实验解决了几个额外的假设，应该导致安全和有效的基因转移直接到所有肌肉的杜氏肌营养不良症犬模型的方式。我们将同时测试几个假设的一系列措施的运动，呼吸和心肌功能的狗，然后应用新的知识在研究中的小狗随机接受基因转移在不同的剂量。实验计划的成功完成将提供与体细胞基因递送的生物学反应相关的一般信息，以及在内皮完整性发生深刻但快速可逆的变化期间保存器官功能。它也将提供有关的系统基因治疗策略的合理设计的具体信息，在一个最常见的单基因致死性疾病的人，杜氏肌营养不良症。公共卫生相关性：该项目通过建立在大型动物系统基因递送的最新进展，解决了肌营养不良症基因治疗发展中的中心限速步骤。我们使用一种新的方法来直接基因转移，以取代缺失的肌营养不良蛋白在运动，呼吸和心肌营养不良的狗，并使用前瞻性，随机研究，以评估治疗效果。

DESCRIPTION (provided by applicant): The overall aim of this project is to address a key rate-limiting step in the development of therapy for Duchenne Muscular Dystrophy: systemic gene "delivery". We focus on a direct gene transfer approach to this problem and specifically avoid any dependence on chronic immunosuppression. We build on compelling proof-of-concept studies from multiple labs using similar adeno-associated virus (AAV) vectors in dystrophic mice and our previous discovery that forced extravasation of vector during retrograde infusion can be used as a means to efficiently transduce both skeletal and cardiac muscle in non- dystrophic large animals. We hypothesize that the underlying mechanism of vector transport relies on pressure-induced, reversible alteration in the scale-dependent barrier function of the venular endothelium. We demonstrate that this hypothetical mechanism is consistent with the observed highly efficient global pattern of gene transfer to muscle during systemic, retrograde infusion in the setting of profound hypothermia and balloon occlusion of the great vessels. The experiments proposed address several additional hypotheses that should lead the way to safe and efficient gene transfer directly to all muscles in the canine model for Duchenne muscular dystrophy. We will concurrently test several hypotheses about serial measures of locomotive, respiratory and cardiac muscle function in the dog and then apply the new knowledge in the study of pups randomized to undergo gene transfer at varying doses. Successful completion of the experimental plan will provide general information relevant to the biological response to somatic gene delivery and the preservation of organ function during profound but rapidly reversible alterations in endothelial integrity. It will also provide specific information about the rational design of strategies for systemic gene therapy in one of the most common single-gene lethal diseases in man, Duchenne Muscular Dystrophy. PUBLIC HEALTH RELEVANCE: This project addresses the central rate-limiting step in the development of genetic treatments for the muscular dystrophies by building on recent progress in systemic gene delivery in the large animal. We use a novel approach to direct gene transfer to replace the missing dystrophin protein in locomotive, respiratory and cardiac muscle in the dystrophic dog, and use a prospective, randomized study to evaluate therapeutic efficacy.