Department of Medical Genetics, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB T6G2H7, Canada.
Laboratory of Biomedical Science, Department of Veterinary Medicine, College of Bioresource Sciences, Nihon University, Fujisawa, Kanagawa 252-0880, Japan.
Proc Natl Acad Sci U S A. 2022 Mar 1;119(9). doi: 10.1073/pnas.2112546119.
Duchenne muscular dystrophy (DMD) is primarily caused by out-of-frame deletions in the dystrophin gene. Exon skipping using phosphorodiamidate morpholino oligomers (PMOs) converts out-of-frame to in-frame mutations, producing partially functional dystrophin. Four single-exon skipping PMOs are approved for DMD but treat only 8 to 14% of patients each, and some exhibit poor efficacy. Alternatively, exons 45 to 55 skipping could treat 40 to 47% of all patients and is associated with improved clinical outcomes. Here, we report the development of peptide-conjugated PMOs for exons 45 to 55 skipping. Experiments with immortalized patient myotubes revealed that exons 45 to 55 could be skipped by targeting as few as five exons. We also found that conjugating DG9, a cell-penetrating peptide, to PMOs improved single-exon 51 skipping, dystrophin restoration, and muscle function in hDMDdel52; mice. Local administration of a minimized exons 45 to 55-skipping DG9-PMO mixture restored dystrophin production. This study provides proof of concept toward the development of a more economical and effective exons 45 to 55-skipping DMD therapy.
杜氏肌营养不良症（DMD）主要由肌营养不良蛋白基因的无义框缺失引起。使用磷酰胺酯键合的吗啉代寡聚物（PMO）进行外显子跳跃可将无义框转变为框内突变，从而产生部分功能的肌营养不良蛋白。有四种单外显子跳跃 PMO 被批准用于治疗 DMD，但每种药物仅能治疗 8%至 14%的患者，并且有些药物疗效不佳。另一方面，外显子 45 至 55 的跳跃可以治疗所有患者的 40%至 47%，并且与改善的临床结果相关。在这里，我们报告了用于外显子 45 至 55 跳跃的肽缀合 PMO 的开发。对永生化患者肌管的实验表明，通过靶向多达五个外显子，就可以跳过外显子 45 至 55。我们还发现，将穿透细胞肽 DG9 缀合到 PMO 上可以提高单外显子 51 的跳跃、肌营养不良蛋白的恢复以及 hDMDdel52 小鼠的肌肉功能。局部施用最小化的外显子 45 至 55 跳跃 DG9-PMO 混合物可恢复肌营养不良蛋白的产生。这项研究为开发更经济有效的外显子 45 至 55 跳跃 DMD 治疗方法提供了概念验证。
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