Targeting the PMP22 Protein to Develop Leads Against Charcot-Marie-Tooth Disease

项目摘要 腓骨肌萎缩症（CMTD）是一种使人衰弱的遗传性周围神经病变，其发病率为1：2500 人类CTMD病理学的标志是严重缺陷的PNS髓鞘。目前没有 对这种疾病的药物治疗或治愈。超过75%的CMTD病例是由以下原因引起的 由于三体性导致的野生型（WT）外周髓鞘蛋白（PMP 22）的雪旺细胞过表达， PMP 22低表达（WT/null情况）或遗传显性杂合子（WT/突变体） 改变PMP 22蛋白序列的突变。已知WT PMP 22可以正确折叠并与 只有20%的效率。我们的基本原理是， PMP 22的表达水平或体内折叠效率具有有效治疗的潜力 剂.减少蛋白质表达的药物可能被证明对最常见的 由基于三体的过表达引起的CMTD形式（1A型）。另一方面，药物作为 折叠校正剂或表达增强剂可能证明对由以下引起的CMTD形式有效： PMP 22的低表达和/或错误折叠。该项目的具体目标是建立 药物发现计划的基础，为最常见的 Charcot-Marie-Tooth病（CMTD）是由遗传变异引起的， 编码外周髓磷脂蛋白22（PMP 22）（一种四面体膜蛋白）的PMP 22基因。 这些目标是： 目标1.开发可在高通量筛选（HTS）中实施的初步测定 鉴定改变野生型（WT）和疾病突变体的总表达的小分子的模式 PMP 22的形式，或增强蛋白质的细胞表面膜运输。 目标二。进行初步筛选，以发现和表征调节总 在一些实施方案中，本发明的组合物可提高PMP 22的表达水平和/或增强PMP 22细胞表面运输效率。确定命中 浓度-响应（效价）曲线。 目标3：使用施旺细胞进行命中的二次筛选，并在一个实验室中测试命中功效。 髓鞘形成测定。进行生物物理研究，以测试铅化合物是否以某种方式起作用， 涉及直接结合到PMP 22蛋白，并且还测试它们对PMP 22蛋白稳定性的影响。 该项目的成功完成将提供一系列先导化合物， 为了将来药物化学优化先导化合物结构， 可用的CMTD小鼠和大鼠模型以及临床前试验。

Project Summary Charcot-Marie-Tooth Disease (CMTD) is a debilitating hereditary peripheral neuropathy that afflicts 1:2500 humans. The hallmark of CTMD pathology is severely defective PNS myelin. There is currently no pharmacological treatment or cure for this disease. Over 75% of all cases of CMTD are caused by Schwann cell overexpression of wild type (WT) peripheral myelin protein (PMP22) due to trisomy, underexpression of PMP22 (for the WT/null case), or genetically dominant heterozygous (WT/mutant) mutations that alter the PMP22 protein sequence. WT PMP22 is known to properly fold and traffic with only 20% efficiency under healthy conditions. Our driving rationale is that small molecules that tune either the expression levels or in vivo folding efficiency of PMP22 have the potential to be effective therapeutic agents. Drugs that reduce expression of the protein are likely to prove effective against the most common form of CMTD (Type 1A) caused by trisomy-based overexpression. On the other hand, drugs that act as folding correctors or expression enhancers are likely to prove effective for forms of CMTD caused by underexpression and/or misfolding of PMP22. The Specific Aims of this project are designed to establish the foundations for a drug discovery program to develop therapeutic compounds for the most common forms of Charcot-Marie-Tooth Disease (CMTD), which are caused by genetic variations that impact the PMP22 gene that encodes peripheral myelin protein 22 (PMP22), a tetraspan membrane protein. These Aims are: Aim 1. Develop a primary assay that can be implemented in high throughput screening (HTS) mode to identify small molecules that alter the total expression of wild type (WT) and disease mutant forms of PMP22, or that enhance the cell surface membrane trafficking of the protein. Aim 2. Conduct primary screens to discover and characterize lead compounds that modulate total expression levels of PMP22 and/or that enhance PMP22 cell surface trafficking efficiency. Determine hit concentration-response (potency) curves. Aim 3. Conduct a secondary screen of hits using Schwann cells and test for hit efficacy in a myelination assay. Conduct biophysical studies to test whether lead compounds act in a way what involves direct binding to the PMP22 protein and also test for their impact on PMP22 protein stability. Successful completion of this project will provide a series of lead compounds that will be ready for future medicinal chemistry optimization of lead compound structures, efficacy testing in available mouse and rat models of CMTD, and pre-clinical testing.