AAV vector delivery to skeletal muscle, platform for therapeutic protein delivery

综合征（SIDS）或合并心脏和骨骼肌病。这些疾病的治疗主要包括饮食控制，并且迄今为止远未达到最佳效果。最近开发的重组腺相关病毒（rAAV）载体的肝细胞和肌纤维的高效转导FAO疾病的研究提供了新的工具。具体而言，我们的实验室已经产生了表达FAO酶的rAAV载体，所述FAO酶的缺乏导致肌病，例如短链酰基CoA脱氢酶（SCAD）和长链酰基CoA脱氢酶（LCAD）。来自缺乏这些酶的患者的人类细胞系是可用的，并且存在这两种疾病的突变小鼠模型。我们建议利用表达FAO酶的rAAV载体，试图解开FAO疾病的病理生物学，并更好地定义这些疾病的分子或细胞为基础的治疗终点。这将在三个具体目标中实现：（1）评估细胞群体或器官内有限百分比的SCAD缺陷或LCAD缺陷细胞的遗传校正可影响脂肪酸氧化的生物化学校正的程度。(2)确定受体结合和进入是否是rAAV在完整哺乳动物肝脏或肌束中稳定转导的限制性步骤。(3)检验以下假设：在LCAD和VLCAD缺乏症中观察到的肝脏病理学继发于毒性代谢物的蓄积，而不是肝细胞内的主要能量衰竭。(4)为了确定在宫内或新生儿IV注射后广泛载体递送后，小鼠中FAO缺陷的整体表型校正是否更有效。从这些研究中获得的信息也可以用于指导其他器官定向治疗的可行性，可能包括干细胞移植。

Syndrome (SIDS) or with a combined cardiac and skeletal myopathy. Treatment of these disorders has consisted primarily of dietary manipulation and has been far less than optimal to this point. The recent development of recombinant adeno-associated virus (rAAV) vectors for highly efficient transduction of hepatocytes and myofibers present new tools for the study of FAO disorders. Specifically, our laboratory has produced rAAV vectors expressing FAO enzymes whose deficiency results in myopathy, such as short-chain acyl CoA dehydrogenase (SCAD) and long-chain acyl CoA dehydrogenase (LCAD). Human cell lines from patients deficient in these enzymes are available, and mutant mouse models exist for both of these disorders. We propose to utilize rAAV vectors expressing FAO enzymes in an attempt to unravel the pathobiology of FAO disorders and to better define endpoints for molecular or cell-based therapies of these disorders. This will be accomplished in three specific aims: (1) To assess the extent to which genetic correction of a limited percentage of SCAD deficient or LCAD deficient cells within a cell population or organ can affect biochemical correction of fatty acid oxidation. (2) To determine whether receptor binding and entry are limiting steps for stable transduction by rAAV in a intact mammalian liver or muscle bundle. (3) To test the hypothesis that the liver pathology observed in LCAD and VLCAD deficiencies are secondary to the accumulation of toxic metabolites as opposed to primary energy failure within hepatocytes. (4) To determine whether global phenotypic correction of FAO deficiency in mice is more effective after widespread vector delivery after intrauterine or neonatal IV injection. The information gained from these studies could also be used to guide the feasibility of other organ-directed therapies, potentially including stem cell transplantation.