Gene therapy in canine myotubular myopathy for clinical translation

描述（由申请人提供）：肌小管蛋白突变导致x连锁肌小管肌病（XLMTM），这是一种毁灭性的先天性肌肉疾病，可导致严重的肌肉无力和呼吸肌肉衰竭导致的过早死亡。只有支持性姑息治疗可用。我们已经证明，aav介导的基因替代挽救了肌小管蛋白缺乏小鼠的严重肌肉无力。新的数据显示，rAAV-MTM1单次全身治疗足以长期（至少一年）生存，并基本上完全改善肌小管蛋白缺乏小鼠的症状。然而，为了最终治疗XLMTM患者，必须采用可预测的大型动物模型来完善给药系统，评估关键的安全性参数，如宿主对载体和转基因的潜在免疫反应，并优化疗效测量。因此，我们开发了一个繁殖群体，其中受影响的雄性狗表现出与人类XLMTM直接相似的表型。在初步研究中，我们证实了局部基因替代疗法，在年轻狗的后肢肌肉内传递，改善肌小管蛋白缺乏骨骼肌的功能和病理。此外，在我们的群体中，第一个系统递送rAAV-MTM1的XLMTM犬在6个多月的时间里，四肢的力量和呼吸功能几乎保持正常，存活时间比任何未经治疗的突变犬长4个月。在小鼠和犬模型中的积极观察结果促使我们将重点放在系统性rAAV-MTM1基因治疗的开发上。我们假设适度水平的肌小管蛋白足以维持全身横纹肌的长期功能，包括重要的呼吸肌。我们建议利用犬类系统来验证这一假设，并优化载体选择（目标1）和剂量（目标2），同时评估潜在的安全问题。为此目的，我们的具体目标是：使用新型rAAV载体（血清型2i8）在犬模型中测试系统MTM1基因替代，该载体设计用于有效递送到骨骼肌，同时避免肝脏Aim 2。

DESCRIPTION (provided by applicant): Mutations in myotubularin cause X-linked myotubular myopathy (XLMTM), a devastating congenital muscle disorder causing severe muscle weakness and premature death from respiratory muscle failure. Only supportive, palliative care is available. We have shown that AAV-mediated gene replacement rescued severe muscle weakness in myotubularin-deficient mice. New data show that a single systemic treatment with rAAV-MTM1 sufficed for long-term (at least one year) survival and essentially complete amelioration of symptoms of mice with myotubularin-deficient muscles. However, for eventual therapy of XLMTM patients, it is imperative to employ a predictive large animal model to refine the delivery system, assess critical safety parameters such as the potential host immune response to vector and transgene, and optimize efficacy measurements. We, therefore, developed a breeding colony in which affected male dogs display a phenotype directly analogous to human XLMTM. In preliminary studies we confirmed that local gene replacement therapy, delivered intramuscularly in the hind limb of young dogs, improved both function and pathology in myotubularin-deficient skeletal muscles. Moreover, the first XLMTM dog in which rAAV-MTM1 was delivered systemically maintained nearly normal strength in all four limbs, and normal respiratory function for more than 6 months surviving > 4 months longer than any untreated mutant dog in our colony. The positive observations in the murine and canine models drive us to focus on the development of systemic rAAV-MTM1 gene therapy. We hypothesize that modest levels of myotubularin will suffice to sustain long-term functionality of striated muscles throughout the body, including the vital respiratory muscles. We propose to utilize the canine system to test this hypothesis and to optimize vector selection (Aim 1) and dosing (Aim 2), while assessing potential safety concerns. Towards this end our Specific Aims are: Aim 1. Test systemic MTM1 gene replacement in the canine model using a novel rAAV vector (serotype 2i8) engineered for effective delivery to skeletal muscle while avoiding the liver Aim 2. Determine dose-response relationships for safety, efficacy, and immune response parameters over a period of at least 32 weeks after systemic MTM1 gene replacement in young XLMTM dogs