TCR V GENE REPERTOIRE IN MS AND SELECTIVE IMMUNOTHERAPY

MS 和选择性免疫治疗中的 TCR V 基因库

• 批准号: 2416309
• 负责人: LAWRENCE STEINMAN
• 金额: $ 24.02万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-12-01 至 1999-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2416309
• 关键词: B lymphocyte; T cell receptor; bacterial antigens; gene expression; gene mutation; gene rearrangement; human genetic material tag; human tissue; immunoglobulin genes; immunotherapy; laboratory rat; multiple sclerosis; myelin basic proteins; polymerase chain reaction; postmortem; receptor binding; site directed mutagenesis; tissue /cell culture; transfection

We have discovered VDJ beta and VJ alpha gene rearrangements in MS brain plaques that encode amino acid sequences identical to those seen in T cell clones from humans, mice and rats that have specificity for myelin basic protein (MBP) peptide 87-99. Using a high-efficiency expression system for TCR alpha and beta chains, we will rigorously prove that a major T cell response in the MS lesion is directed to MBPp87-99. We have developed potent T cell receptor antagonists for p87-99 that suppress ongoing paralysis in rodent models of EAE, and suppress proliferative and cytotoxic responses to p87-99 in humans. We will determine the putative contacts for amino acids in the TCR CDR3 regions with residues of p87-99. A core motif containing MBP87-91 (VHFFK) is found in this epitope. A number of microbes share sequence homology with this core motif-VHFFK, and stimulate MBPp87-99 specific T cells. We will determine whether microbial epitopes can trigger the transfected construct, as efficiently as the native MBP epitope. This would indicate how these T cells might become activated during infection, supporting the notion that self- reactivity arises from the molecular mimicry of self-antigens by microbes. Since T cells reactive to MBP p87-99 trigger EAE, we will test whether these microbial sequences either trigger EAE, or alternatively whether they serve as TCR or MHC antagonists and can block disease induction. Thus we will consider whether molecular mimics of MBP epitopes can induce disease or protect from pathology. We have demonstrated that a dominant antibody response found in the MS brain plaque and in MS cerebrospinal fluid is directed to an overlapping region of the MBP molecule from p86-99. This epitope is identical to the epitope restricted by HLA DR2 beta (HLA DRB1*1501), and overlaps with the epitope restricted by HLA DR2 alpha (HLA DRB5*0101). We will analyze immunoglobulin gene rearrangements in MS brain using RT-PCR, and in CSF using single cell PCR. We will see whether there are restricted Ig CDR3 motifs in MS. We will compare these CDR3 motifs to those found in murine monoclonal antibodies raised against the identical epitope p86-99. Our persistent goal is to develop selective immunotherapy for multiple sclerosis, a chronic disease of the central nervous system affecting approximately 250,000 Americans.

我们在MS脑中发现了VDJ β和VJ α基因重排 噬斑编码的氨基酸序列与T 来自人类、小鼠和大鼠的对髓磷脂具有特异性的细胞克隆 碱性蛋白（MBP）肽87-99。使用高效表达式 系统的TCR α和β链，我们将严格证明， MS损伤中的主要T细胞应答针对MBP β 87 -99。我们有 开发了针对p87-99的有效T细胞受体拮抗剂， 在EAE的啮齿动物模型中进行性瘫痪，并抑制增殖和 对人类p87-99的细胞毒性反应。我们将确定 TCR CDR 3区中的氨基酸与p87-99残基的接触。 在该表位中发现含有MBP 87 -91（VHFFK）的核心基序。一 许多微生物与该核心基序-VHFFK共享序列同源性， 并刺激MBP p87 -99特异性T细胞。我们将决定 微生物表位可以有效地触发转染的构建体 作为天然MBP表位。这将表明这些T细胞如何 在感染过程中被激活，支持自我- 反应性来自自身抗原的分子模拟， 微生物由于T细胞对MBP p87-99的反应性触发EAE，我们将测试 这些微生物序列是否触发EAE，或者 无论它们是作为TCR还是MHC拮抗剂， 诱导因此，我们将考虑MBP表位的分子模拟物是否 可以诱发疾病或防止病理。 我们已经证明，在MS中发现的显性抗体应答 脑斑块和MS脑脊液中的斑块是针对重叠的 MBP分子的p86-99区域。该表位与表位相同。 受HLA DR 2 β（HLA DRB 1 *1501）限制的表位，并与 受HLA DR 2 α（HLA DRB 5 *0101）限制的表位。我们将分析 使用RT-PCR在MS脑中和在CSF中的免疫球蛋白基因重排 使用单细胞PCR。我们将观察是否存在受限制的IG CDR 3 我们将比较这些CDR 3基序与在MS中发现的那些。 针对相同表位p86-99产生的单克隆抗体。我们 持续的目标是开发选择性免疫疗法， 硬化症，一种慢性中枢神经系统疾病， 大约25万美国人。