强直性肌营养不良1型(DM1)的症状是多系统和危及生命的。神经肌肉疾病植根于DM1蛋白激酶(DMPK)基因中的非编码CTG微卫星扩增,在转录时,物理隔离剪接调节蛋白的肌肉盲样(MBNL)家族。蛋白质和重复之间发生的高亲和力结合使MBNL蛋白质无法执行其转录后剪接调节,从而导致与疾病症状直接相关的下游分子效应,例如肌强直和肌肉无力。在这项研究中,我们基于先前证明的证据,表明miRNA-23b和miRNA-218的沉默可以增加DM1细胞和小鼠中的MBNL1蛋白。这里,我们在DM1肌肉细胞中使用blockmiR反义技术,3D小鼠来源的肌肉组织,和体内小鼠阻断这些微小RNA的结合位点,以增加MBNL翻译成蛋白质而不与微小RNA结合。BlockmiRs通过挽救错误剪接显示出治疗效果,MBNL亚细胞定位,和高度特异性转录组表达。阻断miR在3D小鼠骨骼组织中具有良好的耐受性,不诱导免疫应答。在体内,候选blockmiR还增加Mbnl1/2蛋白并挽救握力,拼接,和组织学表型。
The symptoms of Myotonic Dystrophy Type 1 (DM1) are multi-systemic and life-threatening. The neuromuscular disorder is rooted in a non-coding CTG microsatellite expansion in the DM1 protein kinase (DMPK) gene that, upon transcription, physically sequesters the
Muscleblind-like (MBNL) family of splicing regulator proteins. The high-affinity binding occurring between the proteins and the repetitions disallow MBNL proteins from performing their post-transcriptional splicing regulation leading to downstream molecular effects directly related to disease symptoms such as myotonia and muscle weakness. In this study, we build on previously demonstrated evidence showing that the silencing of miRNA-23b and miRNA-218 can increase MBNL1 protein in DM1 cells and mice. Here, we use blockmiR antisense technology in DM1 muscle cells, 3D mouse-derived muscle tissue, and in vivo mice to block the binding sites of these microRNAs in order to increase MBNL translation into protein without binding to microRNAs. The blockmiRs show therapeutic effects with the rescue of mis-splicing, MBNL subcellular localization, and highly specific transcriptomic expression. The blockmiRs are well tolerated in 3D mouse skeletal tissue inducing no immune response. In vivo, a candidate blockmiR also increases Mbnl1/2 protein and rescues grip strength, splicing, and histological phenotypes.