Combinational pharmacotherapies for neuronal abnormalities in Down syndrome

描述(由申请人提供)：唐氏综合症(DS)是导致智力残疾(ID)的最常见的遗传原因，发病率约为750例活产儿中的一例。虽然智商可能很轻，但平均智商在40-50之间。在美国，患有DS的人口目前估计为约350,000人，随着平均预期寿命的增加，这一数字还在继续增加。因此，针对DS患者认知缺陷的药物治疗将产生重大影响。目前，人们热衷于在挽救DS小鼠模型Ts65Dn学习/记忆缺陷的基础上，启动ID在DS中的临床试验。然而，有三个因素可能会限制成功。首先，Ts65Dn在161个人类21号染色体蛋白基因中只有88个是三体的；非三体的HSA21同源基因包括一些导致L/M缺陷和突触可塑性受损的基因，当它们过度表达时。这些基因在DS中的过度表达将影响表型的分子基础，并可能影响药物反应。其次，人类群体中的遗传异质性也可能改变表型和药物反应。第三，一种药物可能不足以纠正导致人类认知缺陷复杂性的许多分子和细胞扰动。为了解决这三个问题，我们建议使用培养的人类原代神经元并测试药物的组合。与对照组相比，DS胎儿脑神经细胞培养显示突触密度降低，线粒体功能降低，活性氧水平升高。在目标1中，我们将分别用八种不同的药物治疗DS胎儿脑神经细胞培养，以拯救这些异常，每一种药物都已被证明或预测对Ts65Dn或DS有效。在目标2中，我们将使用一种新的实验/计算方法，反馈系统控制(FSC)，来测试相同药物的组合对每一种DS异常的挽救作用。目前，确定治疗疾病的药物/剂量组合是基于高通量随机药物筛选或临床环境中的反复试验。相比之下，FSC提供了一种系统的搜索范例，它将细胞系统中的实验测试与使用计算机算法相结合，以寻找生物有效的药物/剂量组合。关键的是，FSC已经表明，在广泛的药物/剂量范围内，生物反应是平稳的。因此，搜索迅速收敛到最佳读数，只需要测试200-300种药物/剂量组合，而不是数十万种。在目标3中，我们将把FSC扩展到Cascade FSC，以优化同时抢救多个DS异常的药物组合。我们提出的系统的重要优势是：i)它使用了一个完整的人类HSA21三体神经元模型，ii)作为一个细胞系统，它允许快速筛选许多药物，iii)药物的组合在抢救多路径异常方面更有可能有效，因此在拯救导致多个、复杂的认知障碍的电路方面更有效，iv)组合中每种药物的浓度可能会低于任何药物单独使用时所需的浓度；这将减少潜在的负面相互作用和非靶向效应；v)只需要测试200-300种药物/剂量组合，而不是数十万种。这项应用的目标是确定一种药物组合的最佳剂量，这些药物共同拯救DS神经元中存在的多种异常。在未来的研究中，最终的融合药物组合将在皮质神经元培养中用于修复其他异常，在DS来源的神经分化诱导多能干细胞(IPSCs)中进行测试，并作为最终的临床前试验评估，在用于修复L/M缺陷的完整DS小鼠模型中进行测试。这一努力是一种新的方法，可以增加DS认知缺陷的临床试验既安全又有效的可能性。

DESCRIPTION (provided by applicant): With an incidence of approximately one in 750 live births, Down syndrome (DS) is the most common genetic cause of intellectual disability (ID). Although ID can be mild, the average IQ is ~40-50. In the United States, the population of people with DS is currently estimated at ~350,000 and continues to increase as the average life expectancy increases. Pharmacotherapies for cognitive deficits in DS would, therefore, have a significant impact. Currently, there is enthusiasm for initiating clinical trials for ID in DS base on rescue of learning/memory deficits in one mouse model of DS, the Ts65Dn. However, three factors may limit success. First, the Ts65Dn is trisomic for only 88 of 161 human chromosome 21 (HSA21) protein genes; non-trisomic HSA21 orthologs include some that cause L/M deficits and impaired synaptic plasticity when they are over expressed. Over expression of these genes in DS will affect the molecular basis of the phenotype and, possibly, drug responses. Second, genetic heterogeneity in the human population may also alter phenotype and drug responses. Third, a single drug may be insufficient to correct the many molecular and cellular perturbations that contribute to the complexity of human cognitive deficits. To address these three issues, we propose to use cultured human primary neurons and test combinations of drugs. Relative to controls, neuronal cultures from DS fetal brain show decreased synapse density, decreased mitochondrial function, and increased levels of reactive oxygen species. In Aim 1, we will test DS fetal brain neuronal cultures for rescue of these abnormalities by separately treating with eight different drugs, each of which has been shown or is predicted to be effective in the Ts65Dn or DS. In Aim 2, we will use a novel experimental/computational method, Feedback System Control (FSC), to test combinations of the same drugs for rescue of each DS abnormality. Currently, identifying drug/dose combinations to treat disease is based on high-throughput random drug screenings or trial-and-error testing in a clinical setting. In contrast, FSC provides a systematic search paradigm that combines experimental testing, in a cell system, with use of a computer algorithm to find biologically effective drug/dose combinations. Critically, FSC has shown that biological responses are smooth over a wide range of drugs/doses. Therefore the search rapidly converges to an optimal readout, requiring testing of only 200-300 drug/dose combinations, not hundreds of thousands. In Aim 3, we will extend FSC to Cascade FSC to optimize the drug combination for simultaneous rescue of multiple DS abnormalities. The important advantages of the system we propose are that i) it uses a complete human trisomy HSA21 neuronal model, ii) as a cell system, it allows rapid screening of many drugs, iii) a combination of drugs is more likely to be effective in rescuing multiple pathway abnormalities and therefore in rescuing circuits underlying multiple, complex cognitive failures, iv) the concentrations required for each drug in the combination likely will be lower tha those required when any drug is used alone; this will decrease potential negative interactions and off target effects; and v) only 200-300 drug/dose combinations need to be tested, not hundreds of thousands. The goal of this application is to identify the optimal doses of a combination of drugs that together rescue multiple abnormalities present in DS neurons. In future studies, the final converged drug combination will be tested in cortical neuronal cultures for rescue of additional abnormalities, in DS-derived neurally differentiated induced pluripotent stem cells (iPSCs), and, as a final preclinical trial evaluation, in a complete DS mouse model for rescue of L/M deficits. This effort is a novel approach to increase the probability that clinical trials for cognitive deficits in DS will be both safe and effective.