Immunoregulation of Myelin Specific T Lymphocytes

描述(由申请人提供)： 项目摘要/摘要人类多发性硬化症(MS)需要新的和改进的药物，因为现有的治疗方法对疾病进程只有轻微的影响。目前的药物显示出比以前的一线治疗(干扰素和醋酸格拉替拉美钠)更好的疗效，但由于全球免疫抑制，大大增加了毒性和严重威胁生命的感染的风险。我们的实验室发现并正在开发部分MHC II类结构物(PMHC)作为治疗MS的可能的免疫治疗方法。pMHC含有MS风险因子的胞外结构域，与脑源性髓鞘少突胶质细胞糖蛋白(MOG)-35-55肽(pDR2/MOG-35-55)共价连接，可以逆转MOG肽诱导的DR2转基因小鼠的中枢神经系统炎症和临床症状。同样的结构(称为RTL1000)最近在多发性硬化症患者的1期安全性试验中成功使用。我们有强有力的临床前证据支持这样的想法，即RTL1000可以特异性地靶向与其他脑源性髓磷肽反应的肽特异性和旁观者T细胞。PMHC复合体的这种抑制活性需要将1‘1部分与主要在单核细胞上表达的特定受体结合。我们最近发现pMHC可以与MHCⅡ类分子不变链(CD74)特异性结合，下调其在单核细胞表面的表达，阻断巨噬细胞移动抑制因子(MIF)的炎症效应，巨噬细胞移动抑制因子是EAE和MS的关键致病细胞因子。我们进一步证明，DR-？1结构域是pDR2结构的主要CD74结合部分。此外，增加DR-1的浓度(在没有结合抗原肽的情况下)可以下调单核细胞上的CD74水平。这些发现表明，DR-1单独可能通过影响CD74的表达来调节MIF的活性，并以一种抗原肽非依赖性的方式治疗EAE。事实上，我们的初步实验表明DR-1对EAE有显著的治疗效果。这些数据具有重要的调控意义，能够开发一类新的治疗性DR-1衍生结构，用于治疗涉及MIF通过CD74信号转导的自身免疫和炎症情况。我们预计，DR-1通过其CD74结合区对CD74的调节将在广泛的炎症条件下提供显著的治疗效果，即使在致病T细胞的特异性尚未确定的情况下，并且在已知或强烈怀疑致病T细胞特异性的条件下，通过将特定抗原肽附着到DR-1部分可以提高治疗效果(例如。多发性硬化症的髓磷脂多肽)。此外，由于DR-1结构域存在于所有人类中，因此不会被认为是外来的，使用DR-1构建的治疗将不需要对潜在的接受者进行HLA筛查，并可用于治疗不表达HLA-DR2风险因子的多发性硬化症患者。我们的初步数据表明，EAE小鼠和MS受试者单核细胞上CD74水平显著增加，从而为发展pMHC结构提供了一个令人信服的理由，pMHC结构可以通过下调CD74表达来抑制MIF活性。在这项应用中，我们将检验这样的假设，即特定的DR-？1片段与单核细胞结合，减少CD74和MIF信号在细胞表面的表达，进而阻止细胞迁移到中枢神经系统和Teff细胞激活，导致EAE的临床和组织学症状逆转。为了解决这一假设，我们建议1)确定DR-1中的CD74结合区，并开发一个最佳的CD74结合的DR-1结构；2)评估DR-1结构对单核细胞和T细胞激活的抑制作用；以及3)确定DR-1结构对多肽特异性与非特异性(旁观者)T细胞特异性诱导的EAE的治疗效果。

DESCRIPTION (provided by applicant): PROJECT SUMMARY/ABSTRACT New and improved drugs are needed for human multiple sclerosis (MS) given that available treatments have only modest effects on the disease course. Current drugs demonstrate better efficacy than the previous first- line treatments (interferon-¿ and glatiramer acetate), but have greatly increased toxicity and risk of serious life- threatening infections due to global immunosuppression. A preferred treatment approach would be a safe and well-tolerated biologic with a potent ability to inhibit key disease-associated mechanisms involved in the pathogenesis of MS. Our laboratory discovered and is developing partial MHC class II constructs (pMHC) as a possible immunotherapy for MS. pMHC containing the extracellular domains of the MS risk factor, HLA-DR2, linked covalently to the encephalitogenic myelin oligodendrocyte glycoprotein (MOG)-35-55 peptide (pDR2/MOG-35-55) can reverse CNS inflammation and clinical signs of MOG-peptide-induced experimental autoimmune encephalomyelitis (EAE) in DR2 transgenic mice. The same construct (termed RTL1000) was recently used successfully in a Phase 1 safety trial in MS patients. We have strong pre-clinical evidence to support the idea that RTL1000 can specifically target both peptide-specific and bystander T-cells reactive to other encephalitogenic myelin peptides. This inhibitory activity of the pMHC complexes requires binding of the ¿1¿1 moiety to a specific receptor expressed predominantly on monocytes. We recently we discovered that pMHC can bind specifically to the invariant chain of MHC class II (CD74), down-regulating its expression on the monocyte cell surface and blocking the inflammatory effects of macrophage migration inhibitory factor (MIF) that is a key pathogenic cytokine for EAE and MS. We further demonstrated that the DR-¿1 domain is the major CD74 binding moiety of pDR2 constructs. Moreover, increased concentrations of DR-¿1 (in the absence of a bound antigenic peptide) could down-modulate CD74 levels on monocytes. These findings suggested that DR-¿1 alone might modulate MIF activity and treat EAE in an antigenic peptide-independent manner through its effects on CD74 expression. Indeed, our preliminary experiments demonstrated a significant treatment effect of DR-¿1on EAE. These data have important regulatory implications that enable the development of a new class of therapeutic DR-¿1-derived constructs for autoimmune and inflammatory conditions involving MIF signaling through CD74. We envision that modulation of CD74 by DR-¿1 through its CD74 binding region will provide significant therapeutic effects in a wide spectrum of inflammatory conditions even when the specificity of pathogenic T-cells has not been established, and that efficacy of treatment may be enhanced by attachment of specific antigenic peptides to the DR-¿1 moiety in conditions where pathogenic T- cell specificities are known or strongly suspected (eg. to myelin peptides in MS). Moreover, because the DR- ¿1 domain is present in all humans and thus would not be recognized as foreign, treatment using DR-¿1 constructs would not require HLA screening of potential recipients and could be used for treatment of MS subjects who do not express the HLA-DR2 risk factor. Our preliminary data suggest a significant increase in CD74 levels on monocytes in mice with EAE and subjects with MS, thus providing a compelling rationale for development off pMHC constructs that can inhibit MIF activity by down-regulating CD74 expression. In this application we will test the hypothesis that a specific segment of DR-¿1 binds to monocytes and decreases cell-surface expression of CD74 and MIF signaling that in turn blocks cell migration into the CNS and Teff-cell activation, resulting in reversal of clinical and histological signs of EAE. To address this hypothesis, we propose to 1) Identify the CD74-binding region within DR-¿1 and develop an optimal CD74-binding DR-¿1 construct; 2) Evaluate inhibitory effects of DR-¿1 constructs on monocyte and T-cell activation; and 3) Determine efficacy of DR-¿1 constructs for treatment of EAE induced by peptide-specific vs. non-specific (bystander) T-cell specificities.