Molecular Characterization of Myelin-Specific T cells from MS Patients

MS 患者髓磷脂特异性 T 细胞的分子特征

基本信息

批准号：
8583032
负责人：
VIJAY K. KUCHROO
金额：
$ 31.64万
依托单位：
DANA-FARBER CANCER INST
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2018-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8583032
关键词：
Address Affinity Antigen Presentation Autoimmune Diseases Autoimmune Process Autoimmune Responses Autoimmunity Binding Biological Markers CD4 Positive T Lymphocytes Candidate Disease Gene Cells Clinical Trials Clonal Expansion Color Complex Cytokine Signaling Data Devices Disease Docking Event Frequencies Gene Expression Gene Expression Profile Genes Goals HLA-DR15 Higher Order Chromatin Structure Human Image Immunity Immunologic Monitoring Individual Institutes Instruction Label Libraries Ligand Binding Ligands Love MHC Class II Genes MHC binding peptide Molecular Multiple Sclerosis Myelin Myelin Basic Proteins Pathogenesis Pathway interactions Patients Peptide/MHC Complex Peptides Pharmaceutical Preparations Population Predisposition Process Property RNA Sequences Reaction Relapsing-Remitting Multiple Sclerosis Robotics Signal Transduction Signaling Molecule Staining method Stains Structure System T-Cell Activation T-Cell Receptor T-Lymphocyte Technology Therapeutic Time Transgenic Mice Virus Yeasts antimicrobial base cytokine genome-wide immunological synapse immunological synapse formation micromanipulator mouse model nano-string novel novel strategies peripheral blood prevent programs research study robotic device screening synaptogenesis therapeutic target transcription factor

项目摘要

Genome-wide analyses of MS susceptibility loci have emphasized the importance of MHC class II genes, strongly implicating antigen presentation to CD4 T cells as a key process in the pathogenesis of MS. However, it has been very difficult to study myelin-specific CD4 T cells from MS patients ex vivo, due to low T cell receptor affinities for the relevant peptide-MHC complexes. Therefore, two fundamental questions remain unresolved: First, which molecular properties distinguish myelin-specific T cells in patients with MS from those in healthy individuals? Second, are there distinguishing properties of myelin-specific and virusspecific T cells that could be exploited for therapeutic gain? In Aim 1, we will study TCR recognition by myelin-specific T cells from MS patients. Imaging experiments identified alterations in immunological synapse formation by myelin-specific compared to virus-specific T cells, and structural studies demonstrated unusual binding topologies by some myelin-specific TCRs isolated from MS patients. The Garcia lab recently developed a yeast peptide-MHC display approach to identify entirely novel peptide ligands for a given TCR and showed that one ofthe peptides was recognized with a non-conventional TCR topology and failed to induce signaling. This means that TCR binding topology can have a profound effect on signaling. This approach will be used to examine the irnpact of TCR topology on the function of myelin-specific T cells and determine whether administration of such peptides can prevent spontaneous disease in human TCR/MHC transgenic mice. In Aim 2, we will collaborate with Drs. Love and Hafler to study myelin and virus-specific T cells from MS patients and control subjects identified with the nanowell device. Antigenresponsive T cells will be isolated from nanowells with a robotic device, which enables detailed molecular characterization of myelin-specific T cell populations. The transcriptome of myelin-specific T cells from MS patients and control subjects will be examined, with an emphasis on the expression of genes associated with susceptibility to MS, transcription factors and cytokine signaling molecules. Clonal expansion of myelinspecific will be assessed by sequencing of TCR chains, and the frequency of myelin-reactive TCR sequences in peripheral blood and CSF T cells will be determined by lllumina sequencing ofthe TCR repertoire. These studies will determine whether myelin-specific T cells with particular cytokine proflles and/or gene expression programs are preferentially expanded in MS patients compared to healthy subjects.

MS 易感位点的全基因组分析强调了 MHC II 类基因的重要性，强烈暗示抗原呈递至 CD4 T 细胞是 MS 发病机制的关键过程。然而，由于细胞水平低，离体研究多发性硬化症患者的髓磷脂特异性 CD4 T 细胞非常困难。 T 细胞受体对相关肽-MHC 复合物的亲和力。因此，两个基本问题仍未解决：首先，哪些分子特性可以区分多发性硬化症患者的髓磷脂特异性 T 细胞来自健康人的那些？其次，髓磷脂特异性和病毒特异性是否有区别？可以利用 T 细胞获得治疗效果吗？在目标 1 中，我们将通过以下方式研究 TCR 识别：来自 MS 患者的髓磷脂特异性 T 细胞。影像学实验发现了免疫学的改变与病毒特异性 T 细胞相比，髓鞘特异性 T 细胞形成突触，结构研究证明从多发性硬化症患者中分离出的一些髓磷脂特异性 TCR 具有不寻常的结合拓扑。加西亚实验室最近开发了一种酵母肽-MHC 展示方法来鉴定全新的肽配体给定 TCR 并表明其中一种肽被非传统 TCR 拓扑识别，并且未能诱导信号传导。这意味着 TCR 结合拓扑结构可以对信号传导产生深远的影响。该方法将用于检查 TCR 拓扑结构对髓磷脂特异性 T 功能的影响。细胞并确定施用此类肽是否可以预防人类自发性疾病 TCR/MHC 转基因小鼠。在目标 2 中，我们将与 Drs. 合作。洛夫和哈夫勒研究髓磷脂和使用纳米井装置识别来自多发性硬化症患者和对照受试者的病毒特异性 T 细胞。抗原反应性 T 细胞将通过机器人设备从纳米孔中分离出来，从而能够获得详细的分子信息髓磷脂特异性 T 细胞群的表征。 MS 髓磷脂特异性 T 细胞的转录组将检查患者和对照受试者，重点是与相关基因的表达对 MS、转录因子和细胞因子信号分子的易感性。髓磷脂特异性克隆扩增将通过 TCR 链测序以及髓磷脂反应性 TCR 的频率进行评估外周血和脑脊液 T 细胞中的序列将通过 TCR 的 Illumina 测序来确定剧目。这些研究将确定具有特定细胞因子谱的髓磷脂特异性 T 细胞是否与健康受试者相比，MS 患者优先扩展和/或基因表达程序。