Testing Compounds that Increase SMN levels for Efficacy in Mouse Models of SMA

描述（由申请人提供）：脊髓性肌萎缩症（SMA）是一种常染色体隐性形式的肌肉萎缩症，大约每6000个活产婴儿中就有1个患有这种疾病，临床表现为运动神经元退化导致的进行性肌肉无力。SMA是由存活运动神经元（SMN）蛋白水平不足引起的，通常是由于SMN1基因的纯合缺失。一个几乎相同的复制基因SMN2不能防止SMA的发展，因为它的mRNA经历了选择性剪接，产生了截断的、不稳定的SMN蛋白。一小部分SMN2转录本包括外显子7，并编码与SMN1相同的SMN蛋白。SMA的严重程度与SMN蛋白水平相关，SMN2基因拷贝数低的婴儿在两年内死亡。患有轻度SMA的儿童有多个SMN2拷贝。这种剂量依赖性通过上调SMN2中全长SMN蛋白的表达，为治疗SMA创造了机会。我们实施了一项先导发现计划，以确定增加SMN蛋白水平的药物样化合物。使用一种新的基于细胞的报告实验，我们对大型化学多样性文库进行了高通量筛选（HTS）。我们已经在SMN蛋白表达的二次测试中验证了多次命中的活性，目前正在通过重点药物化学项目寻求三种化学上不同的支架的优化。目的1将测试从这些构效关系研究中产生的活性化合物的类药物特性、药代动力学和急性毒理学。Aim 2将在“7”SMA小鼠模型中检测最具活性和药理学上最合适的化合物，以确定化合物治疗是否会增加体内SMN蛋白，以及是否可以减轻疾病过程。目的3描述了一种组合方法来检查最有前途的化合物的功效。由于在基于细胞的HTS中发现的化合物似乎通过不同的机制刺激SMN水平，因此将通过评估这些导联的成对组合的生物活性来采取最大化SMN诱导和最小化毒性的治疗方法。这一提议整合了三个实验室在SMA分子生物学、神经退行性疾病新疗法的开发以及SMA小鼠模型药物试验方面的专业知识。该建议的预测结果是确定药理学上合适的化合物，增加SMA模型小鼠的SMN水平并恢复运动活动，可以推进到人体试验。最终目标是开发一种有效的药物治疗脊髓性肌萎缩症，这是婴儿死亡的主要遗传原因。

DESCRIPTION (provided by applicant): Spinal muscular atrophy (SMA) is an autosomal recessive form of muscular dystrophy that afflicts approximately 1 in 6,000 live births and clinically manifests as progressive muscle weakness due to deterioration of motor neurons. SMA results from insufficient levels of the survival motor neuron (SMN) protein, usually due to homozygous deletion of the SMN1 gene. A nearly identical copy gene, SMN2, fails to protect from development of SMA because its mRNA undergoes alternative splicing that produces a truncated, unstable SMN protein. A small fraction of the SMN2 transcripts include exon 7 and encode the same SMN protein as SMN1. SMA severity correlates with levels of the SMN protein, with those infants with low copy number of SMN2 genes dying within two years. Children with milder forms of SMA have multiple copies of SMN2. This dose-dependency creates the opportunity for treatment of SMA by up-regulating expression of the full-length SMN protein from SMN2. We implemented a lead discovery program to identify drug-like compounds that increase SMN protein levels. Using a novel cell-based reporter assay, we performed high-throughput screens (HTS) of large chemically diverse libraries. We have validated the activities of multiple hits in secondary tests for SMN protein expression and are presently pursuing the optimization of three chemically distinct scaffolds through a focused medicinal chemistry program. Aim 1 will test the drug-like properties, pharmacokinetics and acute toxicology of active compounds that emerge from these structure-activity relationship studies. Aim 2 will examine the most active and pharmacologically suitable compounds in the "7" SMA mouse model to determine whether compound treatment increases SMN protein in vivo and whether the disease process can be mitigated. Aim 3 describes a combinatorial approach to examine the efficacy of the most promising compounds. Since compounds identified in the cell-based HTS appear to act through different mechanisms to stimulate SMN levels, a treatment approach to maximize SMN induction and minimize toxicity will be undertaken by evaluating the biological activity of pair-wise combinations of these leads. This proposal merges the efforts of three laboratories with expertise in SMA molecular biology, development of novel therapies for neurodegenerative diseases, and performance of drug trials in murine models of SMA. The predicted outcome of this proposal is identification of pharmacologically suitable compounds that increase SMN levels and restore motor activity in SMA model mice that can be advanced to human trials. The ultimate goal is to develop an effective drug treatment of spinal muscular atrophy, the leading genetic cause of infant mortality. PUBLIC HEALTH RELEVANCE: Spinal muscular atrophy (SMA), a form of muscular dystrophy, is the leading genetic cause of infant death, and in less devastating forms leads to muscle weakness in children and adults. All forms of SMA result from insufficient levels of the protein called SMN. There is no treatment for SMA. We designed a strategy is to individually test very large collections of compounds for ability to specifically increase SMN protein levels. This grant application requests funds for characterizing these chemicals' drug-like properties and testing their efficacy in a mouse model of SMA. Success would engender their optimization followed by entry into human clinical trials.