Systemic molecular therapy for muscular dystrophy

描述（由申请人提供）：该项目的总体目标是解决杜氏肌营养不良症治疗发展中的关键限速步骤：系统性基因“传递”。我们专注于直接的基因转移方法来解决这个问题，并特别避免任何对慢性免疫抑制的依赖。我们基于多个实验室在营养不良小鼠中使用类似的腺相关病毒（AAV）载体进行的令人信服的概念验证研究，以及我们之前的发现，即逆行输注期间载体的强制外渗可以作为有效转导非营养不良大型动物骨骼和心肌的一种手段。我们假设，载体运输的潜在机制依赖于压力诱导的，可逆的改变，在规模依赖的静脉内皮屏障功能。我们证明，这种假设的机制与观察到的在深度低温和大血管球囊闭塞的情况下，在全身、逆行输注过程中基因向肌肉转移的高效全局模式是一致的。提出的实验解决了几个额外的假设，这些假设应该引导安全有效的基因直接转移到犬杜氏肌营养不良模型的所有肌肉。我们将同时测试关于狗的运动、呼吸和心肌功能的一系列测量的几个假设，然后将新知识应用于随机进行不同剂量基因转移的幼崽的研究。实验计划的成功完成将为在内皮完整性发生深刻但迅速可逆的改变时体细胞基因传递和器官功能保存的生物学反应提供一般信息。它还将提供关于合理设计系统性基因治疗策略的具体信息，以治疗人类最常见的单基因致死性疾病之一杜氏肌营养不良症。公共卫生相关性：该项目以大型动物体内系统基因传递的最新进展为基础，解决了肌肉萎缩症基因治疗发展中的中心限速步骤。我们采用一种新的直接基因转移方法来替代营养不良犬运动肌、呼吸肌和心肌中缺失的营养不良蛋白，并采用前瞻性、随机研究来评估治疗效果。

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.