Systemic molecular therapy for muscular dystrophy

描述(由申请人提供)：这个项目的总体目标是解决杜氏肌营养不良症治疗发展中的一个关键的限速步骤：系统性基因“传递”。我们专注于解决这个问题的直接基因转移方法，特别是避免任何对慢性免疫抑制的依赖。我们基于来自多个实验室的令人信服的概念验证研究，在营养不良的小鼠中使用类似的腺相关病毒(AAV)载体，以及我们之前的发现，在逆行输注过程中强制外渗载体可以作为一种手段，在非营养不良的大型动物中有效地转导骨骼肌和心肌。我们假设，载体运输的潜在机制依赖于压力诱导的、可逆的静脉内皮细胞尺度依赖的屏障功能的改变。我们证明，这一假设机制与在深低温和大血管球囊闭塞的情况下，全身逆行输液过程中观察到的高效的全球基因转移到肌肉的模式是一致的。这些实验提出了几个额外的假设，这些假设应该导致将基因直接安全有效地转移到Duchenne肌营养不良犬模型的所有肌肉中。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接受不同剂量基因转移的幼崽的研究中。实验计划的成功完成将提供与躯体基因传递的生物学反应相关的一般信息，并在内皮完整性发生深刻但快速可逆的变化期间保护器官功能。它还将提供关于对人类最常见的单基因致命疾病之一--杜氏肌营养不良症--的系统基因治疗策略的合理设计的具体信息。

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.