全外显子组测序鉴定自身免疫淋巴细胞增生综合征致病基因及其机制研究

Autoimmune lymphoproliferation syndrome (ALPS) is a rare disease, one type of primary immunodeficiency disorder (PID), which is characterized by impaired lymphocyte apoptosis, causing autoimmunity and lymphoproliferation. By far, the causal mutations of eight genes have been identified for ALPS, but the genetic defect remain unknown in around 1/3 of all ALPS patients, which causes a significant problem for the precision diagnosis and treatment. The applicant has successful experience in identifying the causal genetic defect of several PIDs. In this study, the applicant plans to identify the disease-causing mutation for ALPS with unknown genetic cause by using whole exome sequencing and biological experiments. In the preliminary study, RasGRP1 mutation was discovered in the ALPS patients by exome sequencing and bioinformatic analysis. It was further found to be loss-of-function mutation by biological experiment. Based on the previous findings that RasGRP1-deficient mice developed lymphoproliferative disorder and autoimmune phenotype, we hypothesize that the RasGRP1 mutation discovered in the ALPS patients contributes to the development of the disease. During the following study, the mechanisms underlying the disease development will be further investigated, including the effects of the mutation on TCR signaling pathway, corresponding molecular function, immune function and lymphocyte apoptosis. Through the finishing of the study, the genetic defect of these ALPS patients will be successfully identified. In addition, combining the human data yielded in this study and previous animal study results will clarify the roles of RasGRP1 in the host immune system. Taken together, the data obtained in this study have significant potential in clinical practice and will definitely contribute to the understanding of the function of RasGRP1.

自身免疫淋巴细胞增生综合征（ALPS）是由于机体淋巴细胞凋亡障碍而导致的一种罕见原发性免疫缺陷病（PID）。目前已发现8种致病基因。但仍有1/3患者病因不明，为针对性诊治带来严重阻碍。申请人在既往成功利用全外显子组测序辅助鉴定数种PID致病基因的基础上，拟结合外显子组测序与生物学实验来鉴定这部分病因不明ALPS患者的致病基因。前期测序及生物信息学分析已发现患者具有RasGRP1基因突变，并且预实验发现该突变为功能缺失性突变。根据既往动物实验发现RasGRP1缺陷小鼠发生自身免疫淋巴细胞增生性疾病，我们提出假设：RasGRP1基因突变导致人ALPS的发生。后续将进一步研究RasGRP1突变导致ALPS的致病机制以及对分子生物学与免疫功能的影响。本研究的实施，将成功鉴定这部分患者的致病基因，有助于实现精准诊疗；并且明确RasGRP1在机体免疫系统中的作用。因此本课题具有重要应用价值与科学意义。

自身免疫淋巴细胞增生综合征(ALPS)是由于机体淋巴细胞凋亡障碍而导致的一种罕见的原发性免疫缺陷病(PID)。目前已发现8种致病基因，但仍有1/3患者病因不明，为针对性诊治带来严重阻碍。本研究结合全外显子组测序与生物学实验成功鉴定了一个全新的ALPS致病基因RasGRP1。该基因突变为功能缺失突变，虽未能影响蛋白的表达水平，但抑制了T细胞TCR的信号传导，从而导致T细胞活化与增殖缺陷、以及活化诱导的T细胞凋亡缺陷。与经典ALPS不同，RasGRP1突变患者双阴性T细胞数量正常，而γδT细胞显著上升；且这些患者不但表现为自身免疫淋巴增生，还具有多重感染。该研究在全球首次发现报道了一种全新的ALPS致病基因RasGRP1，并被OMIM数据库收录。这是中国医生首次报道新型PID致病基因，从而实现了中国在该领域零的突破。本研究也明确了RasGRP1基因的免疫学功能、以及其在维持机体免疫稳态和耐受中的重要作用；另一方面，本研究成果直接向临床实践转化，指导患者精准诊断和针对性治疗，是践行精准医学的成功范例。