The Dubowitz Neuromuscular Centre, UCL Great Ormond Street Institute of Child Health, London, UK.
Great Ormond Street Institute of Child Health Biomedical Research Centre, National Institute for Health Research, University College London, London, UK.
Skelet Muscle. 2024 Nov 29;14(1):28. doi: 10.1186/s13395-024-00360-4.
Antisense oligonucleotides (AON) represent a promising treatment for Duchenne muscular dystrophy (DMD) carrying out-of-frame deletions, but also show limitations. In a completed clinical trial golodirsen, approved by FDA to induce skipping of DMD gene exon 53 in eligible patients, we demonstrated increase in DMD expression and protein production, albeit with inter-patient variability.
Here, we investigate further the golodirsen mechanism of action using myotubes derived from MyoD transfected fibroblasts isolated from DMD patients at the baseline of the clinical trial SRP-4053.
We confirm golodirsen's selectivity and efficiency in removing only exon 53. For the first time in human cells, we revealed a significant reduction in the so called DMD "transcript imbalance", in golodirsen-treated DMD muscle cultures. The transcript imbalance is a unique DMD phenomenon characterized by non-homogeneous transcript expression along its entire length and responsible for the reduced stability of the transcript. Our in-vivo study also showed that the efficiency of exon skipping did not always correspond to a proportional restoration of the dystrophin protein. Predominant nuclear localization of the DMD transcript, observed in patients and animal models, persists even after exon skipping.
All these findings suggest challenges other than AON delivery for high level of protein restoration in DMD, highlighting the importance of investigating the biological mechanisms upstream of protein production to further enhance the efficiency of any AON treatment in this condition.
反义寡核苷酸 (AON) 为治疗带有移码缺失的杜氏肌营养不良症 (DMD) 提供了一种很有前途的治疗方法，但也存在局限性。在一项已完成的临床试验中，golodirsen 被 FDA 批准用于诱导符合条件的患者的 DMD 基因外显子 53 跳跃，我们证明了 DMD 表达和蛋白产物的增加，尽管存在个体间的差异。
在这里，我们使用从临床试验 SRP-4053 中的 DMD 患者的 MyoD 转染成纤维细胞中分离的肌管，进一步研究 golodirsen 的作用机制。
我们证实 golodirsen 选择性和效率仅能去除外显子 53。这是首次在人类细胞中，我们揭示了 golodirsen 处理的 DMD 肌肉培养物中所谓的 DMD“转录失衡”显著减少。转录失衡是一种独特的 DMD 现象，其特征是整个转录过程中的表达不均匀，并导致转录的不稳定性降低。我们的体内研究还表明，外显子跳跃的效率并不总是与肌营养不良蛋白的比例恢复成正比。在患者和动物模型中观察到的 DMD 转录物的主要核定位，即使在外显子跳跃后仍然存在。
所有这些发现表明，除了 AON 递送来提高 DMD 中的蛋白恢复水平之外，还存在其他挑战，这突出了研究蛋白产生上游的生物学机制的重要性，以进一步提高这种情况下任何 AON 治疗的效率。
