Inhibition of Antigen Presentation in Multiple Sclerosis

描述（由申请人提供）：MHC II类区域是多发性硬化症（MS）和许多其他自身免疫性疾病的主要易感基因座，表明肽呈递给CD 4 T细胞在发病机制中至关重要。该项目的主要目标是开发针对MHCII类抗原呈递途径的治疗MS和其他自身免疫性疾病的新策略。这个项目是PI实验室和哈佛神经变性和修复中心（HCNR）之间的合作，该中心在我们的学术环境中建立了一个药物发现项目。在上一个资助期间，我们开发了一种新的真实的时间肽结合试验，并确定了几组小分子，调节肽结合的MHC II类。最有趣的一组化合物是由一种称为J10的小分子代表的，它可以加速肽加载超过70倍。这种小分子与HLA-DM具有功能相似性，HLA-DM是催化肽装载到晚期内体区室中的MHC II类分子上的蛋白质。基于MHC II类的治疗剂不能有效地加载到MHC分子上，因为它们在到达HLA-DM区室之前长时间暴露于内体蛋白酶。J10在宽的pH范围内是有活性的，这表明它可以使治疗剂在早期内体区室或细胞表面与MHC II类结合。新的数据表明，J10衍生物在体内是有活性的，并且它大大增强了肽展示。此外，J10催化剂与肽的C-末端的共价连接显著提高了体外和体内肽负载的效率。我们的假设是，基于MHC II类的治疗剂的功效可以用催化与MHC II类分子快速结合的小分子显著增强。下一个资助期的目标是确定其精确的作用机制，并在MS的人源化小鼠模型中测试治疗应用。在目标1中，我们将研究这些小分子加速肽加载的结构和功能机制。我们将通过定点诱变候选区域来绘制J10结合位点，并将J10与DR/肽复合物共结晶，通过X射线晶体学进行结构测定。在目的2中，我们将检查三种不同类别的基于MHC II类的治疗剂（克帕松、致耐受性自身肽、MHC II类抑制剂）的治疗功效是否可以通过J10的共同施用或共价连接来增加。该计划的药物化学部分将继续合成具有改进活性和药物样特性的衍生物。公共卫生相关性用于治疗自身免疫性疾病的许多治疗方法需要治疗剂与MHC II类分子的结合。一个主要问题是，这些化合物加载到MHC II类结合位点是低效的，因为加速肽加载的天然催化剂被限制在晚期内体区室。我们已经发现了小分子，大大加速肽结合MHC II类分子，现在将检查其作用机制以及在MS的人源化小鼠模型中的治疗应用。

DESCRIPTION (provided by applicant): The MHC class II region is a principal susceptibility locus for multiple sclerosis (MS) and many other autoimmune diseases, indicating that peptide presentation to CD4 T cells is critical in the pathogenesis. The major goal of this project is develop novel strategies for the treatment of MS and other autoimmune diseases that target the MHC class II antigen presentation pathway. This program is a collaborative effort between the PI's lab and the Harvard Center for Neurodegeneration and Repair (HCNR) that has established a drug discovery program within our academic environment. During the previous funding period, we developed a novel real time peptide binding assay and identified several groups of small molecules that modulate peptide binding by MHC class II. The most interesting group of compounds is represented by a small molecule termed J10 that accelerates peptide loading more than 70-fold. This small molecule has functional similarities with HLA-DM, the protein that catalyzes loading of peptides onto MHC class II molecules in a late endosomal compartment. MHC class II-based therapeutics are inefficiently loaded onto MHC molecules because they are exposed to endosomal proteases for extended periods of time before they reach the HLA-DM compartment. J10 is active over a broad pH range, indicating that it may enable MHC class II binding of therapeutics in early endosomal compartments or at the cell surface. New data demonstrate that a J10 derivative is active in vivo and that it substantially enhances peptide display. Furthermore, covalent attachment of the J10 catalyst to the C-terminus of peptides substantially increases the efficiency of peptide loading both in vitro and in vivo. Our hypothesis is that the efficacy of MHC class II-based therapeutics can be substantially enhanced with a small molecule that catalyzes rapid binding to MHC class II molecules. The goals for the next funding period are to define their precise mechanisms of action and to test therapeutic applications in humanized mouse models of MS. In Aim 1, we will investigate the structural and functional mechanisms by which these small molecules accelerate peptide loading. We will map the J10 binding site by site-directed mutagenesis of candidate regions and co-crystallize J10 with DR/peptide complexes for structure determination by X-ray crystallography. In Aim 2, we will examine whether the therapeutic efficacy of three different classes of MHC class II-based therapeutics (Copaxone, tolerogenic self-peptides, MHC class II inhibitors) can be increased by co- administration or covalent attachment of J10. The medicinal chemistry component of this program will continue to synthesize derivatives with improved activity and drug-like properties for these. PUBLIC HEALTH RELEVANCE A number of therapeutic approaches for the treatment of autoimmune diseases require binding of the therapeutic to MHC class II molecules. A major issue is that loading of these compounds into the MHC class II binding site is inefficient because the natural catalyst that accelerates peptide loading is confined to a late endosomal compartment. We have discovered small molecules that substantially accelerate peptide binding to MHC class II molecules and will now examine their mechanism of action as well as therapeutic applications in humanized mouse models of MS.