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位点，表明肽向CD4 T细胞的呈递在发病机制中至关重要。该项目的主要目标是开发针对MHC II类抗原递呈途径的治疗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）之间的合作成果，该中心在我们的学术环境中建立了一个药物发现项目。在之前的资助期间，我们开发了一种新的实时肽结合试验，并确定了几组调节MHC II类肽结合的小分子。最有趣的一组化合物是一种被称为J10的小分子，它能将肽负载加速70倍以上。这种小分子在功能上与HLA-DM相似，HLA-DM是一种催化肽在内体晚期区室装载到MHC II类分子上的蛋白质。基于MHC ii类的治疗药物不能有效地装载到MHC分子上，因为它们在到达HLA-DM隔室之前暴露于内体蛋白酶较长时间。J10在很宽的pH范围内具有活性，这表明它可能使MHC II类药物在早期内体室或细胞表面结合。新的数据表明，J10衍生物在体内具有活性，并显著增强了肽的显示。此外，J10催化剂与肽的c端共价附着，大大提高了肽在体外和体内的负载效率。我们的假设是，基于MHC II类的治疗方法的疗效可以通过催化与MHC II类分子快速结合的小分子大大增强。下一个资助期的目标是确定它们的确切作用机制，并测试在ms人源化小鼠模型中的治疗应用。在Aim 1中，我们将研究这些小分子加速肽装载的结构和功能机制。我们将通过候选区域的定点诱变来绘制J10的结合位点，并将J10与DR/肽复合物共结晶，通过x射线晶体学来确定结构。在Aim 2中，我们将研究三种不同类型的MHC II类药物（Copaxone，耐受性自肽，MHC II类抑制剂）的治疗效果是否可以通过J10的共给药或共价附着来提高。该计划的药物化学成分将继续合成具有改进活性和类似药物性质的衍生物。许多自身免疫性疾病的治疗方法需要将治疗药物与MHC II类分子结合。一个主要问题是，将这些化合物装载到MHC II类结合位点是低效的，因为加速肽装载的天然催化剂局限于内体晚期腔室。我们已经发现了可以大大加速肽与MHC II类分子结合的小分子，现在将研究它们的作用机制以及在MS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.