Therapeutic exon skipping as a treatment for Duchenne muscular dystrophy (DMD) has largely concentrated on the delivery of antisense oligomers to treat out-of-frame exon deletions. Here we report on the preclinical development of an adeno-associated virus (AAV)-encapsidated viral vector containing four copies of the noncoding U7 small nuclear RNA (U7snRNA), each targeted to either the splice donor or the splice acceptor sites of DMD exon 2. We have previously shown that delivery of this vector (scAAV9.U7.ACCA) to the Dup2 mouse model results in expression of full-length dystrophin from wild-type DMD mRNA, as well as an internal ribosome entry site (IRES)-driven isoform translated only in the absence of exon 2 (deletion exon 2 [Del2] mRNA). Here we present the data from a rigorous dose escalation toxicity study in nonhuman primates, encompassing two doses (3 × 10¹³ and 8 × 10¹³ vg/kg) and two time points (3 and 6 months postinjection). No evidence for significant toxicity was seen by biochemical, histopathologic, or clinical measures, providing evidence for safety that led to initiation of a first-in-human clinical trial.

中文翻译：

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接受临床相关剂量的 AAV9.U7snRNA 载体设计用于诱导 DMD 外显子 2 跳跃的非人灵长类动物缺乏毒性

治疗性外显子跳跃作为杜氏肌营养不良症 (DMD) 的一种治疗方法，主要集中在递送反义寡聚体以治疗框架外外显子缺失。在这里，我们报告了腺相关病毒 (AAV) 衣壳化病毒载体的临床前开发，该载体包含四个拷贝的非编码 U7 小核 RNA (U7snRNA)，每个拷贝都针对DMD外显子 2的剪接供体或剪接受体位点. 我们之前已经表明，将该载体 (scAAV9.U7.ACCA) 递送至 Dup2 小鼠模型会导致野生型DMD表达全长抗肌萎缩蛋白mRNA 以及内部核糖体进入位点 (IRES) 驱动的亚型仅在没有外显子 2 的情况下翻译（缺失外显子 2 [Del2] mRNA）。在这里，我们提供了来自非人灵长类动物的严格剂量递增毒性研究的数据，包括两个剂量（3 × 10 ¹³和 8 × 10 ¹³ vg/kg）和两个时间点（注射后 3 个月和 6 个月）。通过生物化学、组织病理学或临床测量，没有发现显着毒性的证据，提供了导致启动首次人体临床试验的安全性证据。