MODULATION OF CELLULAR CLEARANCE TO TREAT HUMAN DISEASE

描述(由申请人提供)：该项目的目标是生成一种基于细胞清除调节的全新治疗策略的原则证明数据。这一策略将在溶酶体储存疾病(LSD)上进行测试，LSD是一组50多种遗传性疾病，具有进行性、多系统的表型，主要影响儿童。目前LSD的现有治疗方法有很大的局限性。我们已经发现了一种转录因子TFEB，它控制着溶酶体和自噬小体的生物发生和功能(Sardiello等人)。《科学》2009；Settembre等人。科学，2011)，我们已经表明TFEB过表达促进LSD的细胞清除(Medina等人。戴夫。CELL，2011)。最近，我们证明了mTOR抑制剂促进TFEB核转位和活性(Settembre等人)。EMBO J.，2012)。在本项目中，我们将：1)在多发性硫酸酯酶缺乏症(MSD)小鼠模型中测试可诱导的TFEB过表达的治疗潜力。这种小鼠模型的高度严重性和广谱表型将使我们能够测试我们的治疗方法在多个组织上的效果；2)测试AAV介导的TFEB基因载体对Pompe病(PD)小鼠模型的治疗效果；Pompe病(PD)是一种主要累及肌肉和心脏的疾病。这将使我们能够比较 以TFEB为基础的方法，采用更传统的酶替代和基因替代疗法。我们已经通过病毒介导的TFEB在永生化成肌细胞的培养肌管中过表达以及通过肌肉注射AAV-TFEB在PD小鼠模型中的体内表达获得了非常令人鼓舞的初步数据；3)确定了促进TFEB活性的化合物。我们建立了一种基于TFEB核转位的高含量筛选方法。高含量药物筛查确定的阳性命中率将受到 一组体外二次分析和有希望的化合物将被应用于MSD和PD的小鼠模型。如果成功，拟议的方法将代表着LSD治疗模式的转变。与现有的针对单一疾病实体的治疗方法相反，我们基于细胞清除调节的方法可能会对许多LSD和常见的晚发型神经退行性疾病的治疗产生影响。

DESCRIPTION (provided by applicant): The goal of this project is to generate proof-of-principle data on a completely novel therapeutic strategy that is based on the modulation of cellular clearance. This strategy will be tested on lysosomal storage diseases (LSDs), a group of over 50 inherited diseases with a progressive, multisystemic phenotype that mostly affects children. Currently available therapies for LSDs have major limitations. We have discovered a transcription factor, TFEB, that controls the biogenesis and function of lysosomes and autophagosomes (Sardiello et al. Science 2009; Settembre et al. Science, 2011) and we have shown that TFEB overexpression promotes cellular clearance of LSDs (Medina et al. Dev. Cell, 2011). Recently, we demonstrated that mTOR inhibitors promote TFEB nuclear translocation and activity (Settembre et al. EMBO J., 2012). In this project, we will: 1) test the therapeutic potential of inducible TFEB overexpression in a mouse model of Multiple Sulfatase Deficiency (MSD). The high severity and broad spectrum of the phenotype of this mouse model will enable us to test the effects of our therapeutic approach on multiple tissues; 2) test the therapeutic efficacy of AAV- mediated gene delivery of TFEB in a murine model of Pompe disease (PD), a disorder that primarily involves muscles and heart. This will allow us to compare the efficacy of a TFEB-based approach with more traditional enzyme replacement and gene replacement therapies. We have already obtained very encouraging preliminary data by viral-mediated TFEB overexpression in cultured myotubes derived from immortalized myoblasts and in vivo in a PD mouse model by intramuscular injection of AAV-TFEB; 3) identify chemical compounds that promote TFEB activity. We have developed a high-content screening assay based on TFEB nuclear translocation. Positive hits identified by high-content drug screening will be subject to a panel of in vitro secondary assays and promising compounds will be administered to mouse models of MSD and PD. If successful, the proposed approach will represent a paradigm shift in the treatment of LSDs. Contrary to existing therapies, which are directed towards single disease entities, our approach based on the modulation of cellular clearance, may have an impact on the therapy of many LSDs and of common, late onset neurodegenerative diseases.