Towards Therapeutics for Neurodegenerative Diseases

描述（申请人提供）：神经退行性疾病包括阿尔茨海默病、帕金森病和朊病毒病，所有这些都是蛋白质加工障碍。为了开发蛋白质加工疾病的有效治疗方法，我们将朊病毒疾病作为模型系统。朊病毒疾病的研究是有利的，因为稳健的细胞和转基因（Tg）小鼠模型的疾病是可用的。这些细胞和小鼠系统有助于测量疾病进展和治疗干预。由于蛋白质加工疾病之间的共性，人们认为，在开发朊病毒疾病疗法方面学到的经验教训可能对影响更多人的阿尔茨海默病和帕金森病等疾病具有重要意义。朊病毒疾病的特征在于朊病毒蛋白质的细胞同种型（称为PrPc）错误折叠成引起疾病的同种型（称为PrPSc）。我们已经确定了关键点的复制PrPSc，可能适合于治疗性干预与小分子。这些包括底物PrPc的表达和定位;随后转化为PrPSc;以及PrPSc的细胞清除。在本申请中，我们提出产生新的筛选程序的化合物，可能作为治疗朊病毒疾病。我们的目标是开发用于PrPc、PrPSc蓄积和PrPSc清除的稳健的高通量系统（HTS）测定。这些体外筛选系统将用缺乏多药耐药性（mdr）基因la和B的新Tg小鼠补充，因此可以克服传统上与在CNS中获得大量浓度的测试化合物相关的问题。将mdrla和1b基因的敲除与支配PrPc表达的诱导型转基因系统相结合，将使我们能够测量PrPSc的清除率作为CNS中高浓度推定治疗剂的函数。除了有助于测试新化合物的功效外，这些新的Tg小鼠模型还将有助于验证我们的体外筛选靶点。因此，为了扩大在抗朊病毒药物发现有用的相关HTS检测的数量，我们还建议进行shRNA筛选，以确定参与朊病毒表达，复制和清除的非PrP基因。这项研究可能揭示朊病毒疾病的生物学，重要的是，导致开发新的HTS检测，以确定非PrP靶点的选择性配体，减少或抑制朊病毒疾病的进展。大量HTS测定的发展可能最终导致治疗化合物的医疗设备，其在协同使用时停止朊病毒的传播，从而停止疾病的进展。

DESCRIPTION (provided by applicant): The neurodegenerative diseases include Alzheimer's, Parkinson's and prion disease, all of which are disorders of protein processing. Toward developing effective therapies for the protein processing diseases, we have focused on prion disease as a model system. The study of prion disease is advantageous since robust cell and transgenic (Tg) mouse models of disease are available. These cell and mouse systems facilitate measurements of disease progression and therapeutic intervention. Due to the commonality amongst the protein processing diseases, it is thought that lessons learnt in developing therapies for prion disease may have important implications for disorders like Alzheimer's and Parkinson's diseases that affect vastly greater numbers of people. Prion diseases are characterized by the misfolding of the cellular isoform of the prion protein, designated PrPc, to the disease causing isoform, denoted PrPSc. We have identified critical points in the replication of PrPSc that may be suitable for therapeutic intervention with small molecules. These include, expression and localization of the substrate, PrPc; subsequent conversion to PrPSc; and cellular clearance of PrPSc. In this application we propose to generate new screening procedures for compounds that might serve as therapeutics for prion diseases. Our goal is to develop robust, high throughput system (HTS) assays for PrPc, PrPSc accumulation, and PrPSc clearance. These in vitro screening systems will be complemented with new Tg mice that lack the multi-drug resistance (mdr) genes la and b and hence, can overcome problems traditionally associated with achieving substantial concentrations of test compounds in the CNS. Coupling the knockout of the mdrla and 1b genes with an inducible transgene system that governs PrPc expression, will allow us to measure the clearance of PrPSc as a function of high concentrations of putative therapeutics in the CNS. Besides facilitating testing of the efficacy of novel compounds, these new Tg mouse models will also aid in validating our in vitro screening targets. So as to expand the number of relevant HTS assays useful in anti-prion drug discovery, we also propose to undertake a shRNA screen to identify non-PrP genes that are involved with prion expression, replication and clearance. This study may shed light on the biology of prion disease, and importantly, lead to the development of novel HTS assays to identify selective ligands for non-PrP targets that reduce or inhibit the progression of prion disease. The development of a substantial array of HTS assays may eventually lead to an armamentarium of therapeutic compounds, which when used in concert halt the propagation of prions and hence, the progression of disease.