Engineering Multiple U7snRNA Constructs to Induce Single and Multiexon-skipping for Duchenne Muscular Dystrophy
Engineering Multiple U7snRNA Constructs to Induce Single and Multiexon-skipping for Duchenne Muscular Dystrophy
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DOI：
10.1038/mt.2012.26
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发表时间：
2012-06-01
影响因子：
12.4
通讯作者：
Davies, Kay E.
中科院分区：
文献类型：
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作者：
Goyenvalle, Aurelie;Wright, Jordan;Davies, Kay E.
杜氏肌营养不良症（DMD）是一种致命的肌肉萎缩性疾病引起的突变，在肌营养不良基因。反义介导的外显子跳跃是治疗DMD最有前途的方法之一，但仍然面临个性化医学挑战，因为DMD患者中发现的不同突变需要不同外显子的跳跃。然而，70%的DMD患者在中央基因组区域的突变丰富区域或“热点”中具有抗肌萎缩蛋白基因缺失。在这项研究中，我们开发了11种不同的U7小核RNA，穿梭反义序列，旨在掩盖参与外显子45至55剪接的关键元件。我们证明，这些构建体在DMD患者的成肌细胞体外和人DMD（hDMD）小鼠体内均诱导有效的外显子跳跃，并且它们可以组合成单个载体以实现至少3个外显子的多重跳跃。这些非常令人鼓舞的结果提供了使用编码多个U7小核RNA（U7snRNA）的腺相关病毒（AAV）载体可以实现有效的多外显子跳跃的原理证明，因此为DMD的临床治疗提供了非常有前途的工具。
Duchenne muscular dystrophy (DMD) is a fatal muscle wasting disorder caused by mutations in the dystrophin gene. Antisense-mediated exon skipping is one of the most promising approaches for the treatment of DMD but still faces personalized medicine challenges as different mutations found in DMD patients require skipping of different exons. However, 70% of DMD patients harbor dystrophin gene deletions in a mutation-rich area or "hot-spot" in the central genomic region. In this study, we have developed 11 different U7 small-nuclear RNA, to shuttle antisense sequences designed to mask key elements involved in the splicing of exons 45 to 55. We demonstrate that these constructs induce efficient exon skipping both in vitro in DMD patients' myoblasts and in vivo in human DMD (hDMD) mice and that they can be combined into a single vector to achieve a multi skipping of at least 3 exons. These very encouraging results provide proof of principle that efficient multiexon-skipping can be achieved using adeno-associated viral (AAV) vectors encoding multiple U7 small-nuclear RNAs (U7snRNAs), offering therefore very promising tools for clinical treatment of DMD.