Targeting CUG Expansions for the Treatment of Myotonic Dystrophy

描述(申请人提供)：强直性肌营养不良症1(DM1)是一种常见的神经肌肉疾病，由强直性蛋白激酶基因(DMPK)3‘-非翻译区的多CTG扩增引起。这种疾病无法治愈，也没有延缓疾病进展的治疗选择。现在有强有力的证据支持聚(CUG)RNA序列(转录后得到的聚CTG扩展)作为DM1的分子基础的有毒功能增益作用。聚(CUG)RNA结合并隔离重要的蛋白质，抑制它们的正常功能。在这些蛋白质中，最主要的是肌盲样蛋白1(MBNL1)，它介导了几个重要客户前-mRNAs的正确前-mRNAs剪接。由于聚(CUG)RNA对MBNL1的隔离，至少六种关键蛋白质的胎儿剪接变体在成人中产生，直接导致疾病症状。有相当多的遗传证据表明，破坏MBNL1-聚(CUG)RNA相互作用将逆转DM1。因此，这项建议的总体目标是识别和开发能够破坏聚(CUG)RNA-MBNL1相互作用并逆转DM1疾病表型的小分子。这种方法的关键是小分子必须以RNA为靶点，而不是MBNL1，因为MBNL1必须能够在细胞中正常发挥作用。聚(CUG)RNA是开发小分子结合剂的良好靶子，因为聚(CUG)形成了一个有结构的、稳定的发夹，而CUG重复序列不是细胞正常功能的一部分。因此，该RNA序列的特异性靶向应该会中断与MBNL1的相互作用，而不会影响其他基本的细胞过程。该建议的具体目的是：1)探索MBNL蛋白与聚(CUG)RNA的结合亲和力和特异性，2)鉴定聚(CUG)RNA的小分子结合物，3)在体外和强直性肌营养不良的细胞培养模型中评估干扰聚(CUG)RNA-MBNL1相互作用的化合物。通过追求这些目标，我们的目标是快速验证聚(CUG)RNA作为治疗DM1的靶点，并识别适合在DM1临床前模型中进行药物化学优化和评估的化合物。

DESCRIPTION (provided by applicant): Mytotonic dystrophy 1 (DM1) is a prevalent neuromuscular disorder that is caused by a poly-CTG expansion in the 3'-untranslated region of the myotonic protein kinase gene (DMPK). This disease cannot be cured, and there are no treatment options that delay disease progression. There is now strong evidence supporting a toxic gain-of-function role for the poly(CUG)RNA sequences (that result upon transcription of the poly-CTG expansion) as the molecular basis for DM1. The poly(CUG)RNA binds to and sequesters important proteins, inhibiting their normal function. Chief among these proteins is muscleblind-like protein 1 (MBNL1), which acts to mediate proper pre-mRNA splicing of several important client pre-mRNAs. As a result of the sequestration of MBNL1 by poly(CUG)RNA, fetal splice variants of at least six key proteins are produced in the adult, leading directly to the disease symptoms. There is considerable genetic evidence that disruption of the MBNL1-poly(CUG)RNA interaction will reverse DM1. Thus, the overall goal of this proposal is to identify and develop small molecules capable of disrupting poly(CUG)RNA-MBNL1 interactions and reversing the DM1 disease phenotype. The key to this approach is that the small molecules must target the RNA, rather than MBNL1, because MBNL1 must be able to act normally in the cell. The poly(CUG)RNA is a good target for the development of small molecule binders because poly(CUG) forms a structured, stable hairpin and CUG repeat sequences are not part of the normal functioning of the cell. Thus, specific targeting of this RNA sequence should disrupt interactions with MBNL1 without affecting other essential cellular processes. The specific aims of the proposal are: 1) to probe the binding affinity and specificity of MBNL proteins for poly(CUG)RNA, 2) the identification of small-molecule binders of poly(CUG)RNA, and 3) the assessment of compounds that disrupt the poly(CUG)RNA-MBNL1 interaction in vitro and in cell culture models of myotonic dystrophy. By pursuing these aims, our goal is to rapidly validate poly(CUG)RNA as a target for the treatment of DM1, and to identify compounds that will be appropriate for medicinal chemistry optimization and evaluation in pre-clinical models of DM1.