Small molecule therapeutics for myotonic dystrophy type 1

总结 强直性肌营养不良1型（DM 1）是一种常染色体显性遗传疾病，由CTG扩增引起， 在DMPK基因的3'非翻译区的重复序列。DM 1的主要治疗靶点是 编码扩展的CUG重复序列（CUGexp）的突变体DMPK RNA，其形成毒性核内 患者细胞中的聚集体或CUGexp病灶。我们已经开发了一种基于RNA的筛选策略， 鉴定选择性调节DMPK CUGexp RNA而不影响正常的小分子 DMPK记录。在对2,500种化合物的试验性筛选中，我们鉴定出了一种原型小分子MDI 16， 其有效地逆转了患者细胞和HSALR小鼠中的关键DM 1病理特征 型号DM 1由于多个线索的鉴定大大提高了小分子 我们使用这种筛选策略从40，000种不同的药物样药物中鉴定出30种新的命中物， MSSR-UCLA文库中的小分子，该文库已经针对负债进行了广泛的过滤。这 与患者中的MDI 16相比，一组命中结果显示在减少CUGexp病灶方面具有更好的安全性和有效性 细胞重要的是，比较分析表明，我们的命中与反义核酸的表现不相上下。 针对CUGexp的寡核苷酸，并且优于发表于 文学作品在本申请中，我们提出通过功效排序来鉴定先导化合物， 我们的命中面板在逆转关键DM 1病理特征，包括 CUGexp病灶形成、RNA剪接异常、SHARP错误定位、CUGBP 1和GSK 3b升高 DM 1患者成肌细胞中的水平。将使用生物化学测定来确定作用机制 排名靠前的命中，将测试其在逆转DM 1骨骼肌疾病中的体内功效 在HSALR DM 1小鼠模型中。血脑屏障的命中率将在一个新的双测试- 在脑中表达CUGexp病灶的转基因小鼠模型中。由于没有有效的治疗方法， DM 1，候选治疗化合物的鉴定为这种衰弱的患者提供了希望。 疾病

Summary Myotonic dystrophy 1 (DM1) is an autosomal dominant disorder resulting from the expansion of a CTG repeat tract in the 3’ untranslated region of the DMPK gene. The primary therapeutic target in DM1 is the mutant DMPK RNA encoding expanded CUG repeats (CUGexp), which forms toxic intra-nuclear aggregates or CUGexp foci in patient cells. We have developed an RNA-based screening strategy to identify small molecules that selectively modulate the DMPK CUGexp RNA without affecting the normal DMPK transcript. In a pilot screen of 2,500 compounds we identified a prototype small molecule MDI16, which effectively reverses critical DM1 pathological features in both patient cells and in the HSALR mouse model of DM1. As identification of multiple leads greatly enhances the probability of a small molecule therapy for DM1, we used this screening strategy to identify 30 novel hits from 40,000 diverse drug-like small molecules of the MSSR-UCLA library, which has undergone extensive filtering against liabilities. This panel of hits show better safety and efficacy in reducing CUGexp foci when compared to MDI16 in patient cells. Importantly, comparative analysis demonstrates that our hits perform on par with antisense oligonucleotides directed against CUGexp and better than DM1 small molecule therapeutics published in the literature. In this application we propose to identify lead compounds by rank-ordering the efficacy, potency, selectivity and safety of our hit panel in reversing key DM1 pathological features including the formation of CUGexp foci, aberrant RNA splicing, SHARP mis-localization, elevated CUGBP1 and GSK3b levels in DM1 patient myoblasts. Biochemical assays will be used to determine the mechanism of action of top-ranked hits, which will be tested for their in vivo efficacy in reversing DM1 skeletal muscle disease in the HSALR DM1 mouse model. Blood brain barrier penetrance of the hits will be tested in a novel bi- transgenic mouse model that expresses CUGexp foci in the brain. As there is no effective treatment for DM1, the identification of candidate therapeutic compounds offers hope for patients with this debilitating disease.