Somatic Mutation in the Etiology of Multiple Sclerosis

项目摘要 自身免疫性疾病是一类超过80种慢性致残性疾病，其中免疫缺陷 系统引导身体攻击自身组织的正常成分(自身抗原)。自身免疫性疾病 仅在美国就有1470-2350万人受到影响(5%-8%)，使其成为最常见的 人类疾病。然而，尽管有相当多的研究致力于了解自身免疫，但 启动自身免疫反应的关键致病事件仍然缺乏连贯的解释。 自身免疫性疾病的家族聚集性表明疾病风险增加，遗传易感性， 是可以遗传的。但即使在风险遗传的环境中，疾病的发病也被发现发生在一个变量之后 延迟，这表明额外的随机过程是触发疾病发作的必要因素。这 这一现象让人想起在癌症中观察到的遗传易感性 然而，只有在额外的靶基因发生体细胞突变后，癌症才会发展。因此， 这项提案中描述的项目的目标是测试这样一个假设，即基因的体细胞突变 通过执行基于单细胞RNA序列来启动自身免疫需要调节免疫耐受性 多发性硬化症患者致病淋巴细胞的表达谱及序列分析 尽管在MS患者的中枢神经系统(CNS)中发现的炎性损害包括一个复杂的 T细胞、B细胞和巨噬细胞的混合物，克隆扩增、受体编辑和 在脑脊液中表达脑反应性抗体的体细胞突变记忆B细胞， 而B细胞耗竭治疗的戏剧性临床反应表明，自身反应性B细胞发挥着关键作用 在多发性硬化症病理中的作用，因此是拟议研究的重点。 提出了三个具体的目标：1.确定MS患者脑脊液中的致病B细胞是否携带体细胞 参与调节B细胞耐受性的基因突变，2.确定MS患者脑脊液中致病B细胞 患者表现出结构性或对激活信号的反应而改变的转录图谱，以及3. 研究在耐受相关信号通路上检测到的体细胞突变的功能相关性。 通过对单个脑脊液B细胞的mRNA序列和表达数据进行全基因组比对分析 我们希望从多发性硬化症患者中确定体细胞突变及其影响是否可以在致病 细胞。我们还将开发一种经过验证的单细胞rna-seq方法，用于进行类似的 在其他自身免疫性疾病和免疫反应环境中的研究。高级单曲的发展 细胞基因组方法学，将使我们能够探索体细胞突变和分化的潜在作用 多发性硬化症病因学中的基因表达将真正改变游戏规则。体细胞突变作用的证据 多发性硬化症的病因学和/或病理学将完全改变我们看待自身免疫性疾病发展的方式 并将对开发替代的个性化治疗方法具有广泛的影响。 OMB编号0925-0001/0002(08/12版批准至2015年8月31日)页面续格式页面

Project Summary Autoimmune diseases are a family of over 80 chronic, disabling illnesses in which defects in the immune system leads the body to attack normal components (self antigens) of its own tissues. Autoimmune disease affects 14.7 – 23.5 million people in the U.S. alone (5 – 8%), making it one of the most common classes of human diseases. However, despite considerable research being devoted to understanding autoimmunity, a coherent explanation of the critical causative events that initiate the autoimmune response is still lacking. Familial clustering of autoimmune disease indicates that an increased risk of disease, a genetic predisposition, can be inherited. But even in settings in which risk is inherited, disease onset is found to occur after a variable delay, suggesting that additional stochastic processes are necessary to trigger disease onset. This phenomenon is reminiscent of what has been observed in cancer where a genetic predisposition can be inherited, and yet cancer develops only after somatic mutations in additional target genes occur. Thus, the goal of the project described in this proposal is to test the hypothesis that somatic mutations in genes that regulate immune tolerance are required to initiate autoimmunity, by performing single cell RNA-seq-based expression profiling and sequence analysis of pathogenic lymphocytes from multiple sclerosis (MS) patients. Although the inflammatory lesions found in the central nervous system (CNS) of MS patients include a complex mixture of T cells, B cells and macrophages, the presence of clonally-expanded, receptor-edited and somatically-mutated memory B cells that express brain-reactive antibodies in the cerebrospinal fluid (CSF), and the dramatic clinical response to B cell depletion therapy, suggest that autoreactive B cells play a critical role in MS pathology, and are therefore the focus of the proposed studies. Three specific aims are proposed: 1. Determine if pathogenic B cells in the CSF of MS patients carry somatic mutations in genes involved in regulating B cell tolerance, 2. Determine if pathogenic B cells in the CSF of MS patients exhibit altered transcriptional profiles either constitutively or in response to activating signals, and 3. Investigate the functional relevance of somatic mutations detected on tolerance-related signaling pathways. Through the genome-wide comparative analysis of mRNA sequence and expression data of single CSF B cells from MS patients we expect to determine if somatic mutations and their effects can be identified in pathogenic cells. We will also have developed a validated single cell RNA-seq methodology for the conduct of similar studies in other autoimmune disease and immune response settings. The development of advanced single cell genomic methodologies that will allow us to explore the potential role of somatic mutations and differential gene expression in the etiology of MS will be a real game changer. Evidence for a role for somatic mutations in MS etiology and/or pathology would completely change the way we view autoimmune disease development and would have broad implications for the development of alternative personalized treatment approaches. OMB No. 0925-0001/0002 (Rev. 08/12 Approved Through 8/31/2015) Page Continuation Format Page