High content screening to identify therapeutics for multiple sclerosis

描述（由申请人提供）：多发性硬化症（MS）是最常见的中枢神经系统（CNS）炎症性脱髓鞘疾病，是北美年轻人非创伤性神经功能障碍的主要原因。虽然MS的病因尚不清楚，但其发病的特点是免疫细胞浸润，少突胶质细胞和髓磷脂的破坏。5-15年后，大多数患者进入轴突变性阶段，逐渐出现不可逆转的神经功能衰退。目前的抗炎或免疫调节疗法在延缓疾病方面部分有效，但这些疗法并不能保证功能恢复。少突胶质细胞的替代及其随后的脱髓鞘轴突的再髓鞘再生可以阻止和逆转轴突损失和神经功能衰退。一种增加髓鞘再生的方法是通过治疗性刺激内源性修复。这种方法是可行的，因为在疾病早期的许多MS病变中，甚至在一些慢性病变中，确实会产生新的少突胶质细胞和自发的髓鞘再生，尽管内源性髓鞘再生最终会失败。本提案的假设是，选择的小分子可以作为髓鞘再生治疗刺激少突胶质细胞分化和髓鞘再生。最近的两项技术发展使鉴定和评价这种小分子成为可能。首先，优化了一种从小鼠身上获得高纯度原代少突胶质祖细胞（OPCs）的方法，该方法可作为筛选的起始材料。第二，一种高含量、基于细胞的表型筛选方法，以中等通量的方式自动对分化的少突胶质细胞进行评分。这个创新的平台将用于筛选小分子库，以确定可以促进OPC分化的hit。然后主屏幕上的假定命中

DESCRIPTION (provided by applicant): Multiple sclerosis (MS) is the most common inflammatory demyelinating disease of the central nervous system (CNS) and is the major cause of non-traumatic neurological disability in young adults in North America. Although the etiology of MS is not clear, its onset is characterized by infiltration of immune cells and the destruction of oligodendrocytes and myelin. After 5-15 years, most patients enter a phase of axonal degeneration and gradual, irreversible neurological decline. There is no cure for MS. Current anti- inflammatory or immunomodulatory therapies are partially effective in delaying the disease, but these therapies do not guarantee functional recovery. Replacement of oligodendrocytes and their subsequent remyelination of demyelinated axons could halt and reverse axonal loss and neurological decline. One approach to increase remyelination is through therapeutic stimulation of endogenous repair. This approach is feasible since generation of new oligodendrocytes and spontaneous remyelination do occur in many MS lesions during early stages of the disease, and even in some chronic lesions, although endogenous remyelination eventually fails. The hypothesis of this proposal is that selected small molecules can be used as remyelination therapeutics to stimulate differentiation of oligodendrocytes and remyelination. Two recent technological developments enable the identification and evaluation of such small molecules. First, an optimized method to produce a highly pure population of primary oligodendrocyte progenitor cells (OPCs) from the mouse that can be used as starting material for screening. Second, a high-content, cell-based phenotypic screening method to automatically score differentiated oligodendrocytes in a medium-throughput manner. This innovative platform will be used to screen a small molecule library to identify hits that can promote OPC differentiation. Putative hits from the primary screen will then be evaluated for their ability to activate the expression of oligodendrocyte-specific markers. Potency, cytotoxicity, cell-type selectivity and interaction with known signaling pathways that regulate oligodendrocyte generation will also be evaluated. The goal of this Phase I project is to identify the best small molecule drug candidate that can promote differentiation of OPCs for subsequent Phase II studies and development. Future Phase II projects will include detailed toxicological studies of the lead compound and in vivo testing in the mouse cuprizone model of demyelination. The long-term goal of this project is to develop therapeutic agents that will promote remyelination to reverse neurological decline in MS patients.