Department of Neurology, University of Washington, Seattle, Washington, United States of America.
Department of Medicine, University of Washington, Seattle, Washington, United States of America.
PLoS Genet. 2020 Nov 11;16(11):e1009179. doi: 10.1371/journal.pgen.1009179. eCollection 2020 Nov.
Gene therapy approaches for DMD using recombinant adeno-associated viral (rAAV) vectors to deliver miniaturized (or micro) dystrophin genes to striated muscles have shown significant progress. However, concerns remain about the potential for immune responses against dystrophin in some patients. Utrophin, a developmental paralogue of dystrophin, may provide a viable treatment option. Here we examine the functional capacity of an rAAV-mediated microutrophin (μUtrn) therapy in the mdx4cv mouse model of DMD. We found that rAAV-μUtrn led to improvement in dystrophic histopathology & mostly restored the architecture of the neuromuscular and myotendinous junctions. Physiological studies of tibialis anterior muscles indicated peak force maintenance, with partial improvement of specific force. A fundamental question for μUtrn therapeutics is not only can it replace critical functions of dystrophin, but whether full-length utrophin impacts the therapeutic efficacy of the smaller, highly expressed μUtrn. As such, we found that μUtrn significantly reduced the spacing of the costameric lattice relative to full-length utrophin. Further, immunostaining suggested the improvement in dystrophic pathophysiology was largely influenced by favored correction of fast 2b fibers. However, unlike μUtrn, μdystrophin (μDys) expression did not show this fiber type preference. Interestingly, μUtrn was better able to protect 2a and 2d fibers in mdx:utrn-/- mice than in mdx4cv mice where the endogenous full-length utrophin was most prevalent. Altogether, these data are consistent with the role of steric hindrance between full-length utrophin & μUtrn within the sarcolemma. Understanding the stoichiometry of this effect may be important for predicting clinical efficacy.
使用重组腺相关病毒（rAAV）载体将微型（或微）肌营养不良蛋白基因递送到横纹肌的基因治疗方法已显示出显著进展。然而，一些患者对针对肌营养不良蛋白的免疫反应的潜在风险仍存在担忧。肌联蛋白是肌营养不良蛋白的发育同源物，可能提供一种可行的治疗选择。在这里，我们研究了 rAAV 介导的微肌联蛋白（μUtrn）治疗在 DMD 的 mdx4cv 小鼠模型中的功能能力。我们发现，rAAV-μUtrn 导致肌营养不良的组织病理学改善，并在很大程度上恢复了神经肌肉和肌肌腱接头的结构。胫骨前肌的生理研究表明，峰值力维持，比特定力部分改善。μUtrn 治疗的一个基本问题不仅是它是否可以替代肌营养不良蛋白的关键功能，而且全长肌联蛋白是否会影响较小的、高度表达的 μUtrn 的治疗效果。因此，我们发现 μUtrn 显著减小了相对全长肌联蛋白的肌节晶格间距。此外，免疫染色表明，肌营养不良病理生理学的改善主要受到快速 2b 纤维的有利纠正的影响。然而，与 μUtrn 不同，μDys（μDys）表达没有表现出这种纤维类型偏好。有趣的是，与 mdx4cv 小鼠相比，μUtrn 能够更好地保护 mdx:utrn-/- 小鼠中的 2a 和 2d 纤维，而 mdx4cv 小鼠中存在最丰富的内源性全长肌联蛋白。总的来说，这些数据与肌联蛋白全长与 μUtrn 之间在肌膜内的空间位阻作用一致。了解这种效应的化学计量可能对预测临床疗效很重要。
