Immunologic and Genetic Analysis to Understand the Pathogenesis of Primary Selective IgM Immunodeficiency (sIgMID)

原发性选择性免疫球蛋白（IG）M免疫缺陷（sIgMID）患者血清中存在孤立的IgM缺乏症，并且通常患有严重甚至危及生命的复发性感染。常见的临床表现包括上、下呼吸道感染、尿路感染和肠胃炎。在成年期，一些sIgMID患者发展为自身免疫性疾病。目前尚不清楚IgM缺乏如何导致这些症状。先前的研究表明，B细胞内在缺陷和T细胞功能缺陷可能有助于sIgMID。此外，遗传异常已被描述，但总体的发病机制sIgMID仍然知之甚少。我假设，sIgMID是一种异质性疾病，分层是关键，以了解发病机制。我们将首次分析约50例sIgMID患者的大型队列，目的是通过全面的免疫学和遗传学分析对疾病进行分层。这项研究有三个具体目标：（一） 我们希望对sIgMID患者进行全面的免疫学表型分析，包括分析B和T细胞亚群分布，根据活化状态、存活、凋亡、成熟和迁移表征B细胞亚群，分析IgM表达，以及体外刺激，包括与患者B细胞和健康供体T细胞的交叉刺激，反之亦然。（二） 我们希望对sIgMID患者及其未受影响的家庭成员进行全外显子组测序（和全基因组测序），以鉴定可能导致或易患sIgMID的单核苷酸变异和/或拷贝数变异。对于该分析，应根据目标（i）中出现的亚型对患者进行分组。（三） 为了验证新发现的候选变体并评估其生物学影响，我们希望从患者B细胞中产生淋巴母细胞样细胞系（LCL），并使用CRISPR/Cas9基因编辑将变体校正回参考。然后，我们将根据其扩增和产生IgM的能力来比较原始和校正的LCL。本研究的结果将使不同临床sIgMID表型与特定免疫缺陷和/或某些遗传变异相关成为可能。这种疾病分层将有助于诊断，并可能改善治疗。此外，我认为sIgMID是一种模型疾病，了解sIgMID的发病机制不仅可以阐明其他免疫缺陷，但它也将提供洞察一般免疫机制，如免疫球蛋白的产生，T细胞- B细胞相互作用和B细胞成熟和分化的调节。

Patients with primary selective immunoglobulin (Ig) M immunodeficiency (sIgMID) have an isolated IgM deficiency in serum and typically suffer from recurrent infections that can be severe or even life-threatening. Common clinical presentations include upper and lower respiratory tract infections, urinary tract infections and gastroenteritis. During adulthood, some sIgMID patients develop autoimmune diseases. It is not clear how IgM deficiency causes these symptoms. Previous studies indicate that B cell-intrinsic defects and defects in T cell function may contribute to sIgMID. Also genetic abnormalities have been described, but overall the pathogenesis of sIgMID remains poorly understood.I hypothesize that sIgMID is a heterogeneous disease and that stratification is key to understand the pathogenesis. For the first time, we will analyze a large cohort of ~50 sIgMID patients with the objective to stratify the disease by a comprehensive immunological and genetic analysis. This study has three specific aims: (i) We want to perform a comprehensive immunological phenotyping of sIgMID patients that comprises the analysis of B and T cell subset distribution, the characterization of B cell subsets according activation status, survival, apoptosis, maturation and migration, the analysis of IgM expression, as well as in vitro stimulation including cross-stimulation with patient B cells and healthy donor T cells and vice versa.(ii) We want to perform whole-exome sequencing (and whole-genome sequencing, respectively) of sIgMID patients and their unaffected family members to identify single nucleotide variants and/or copy number variants that may cause or predispose for sIgMID. For this analysis, patients shall be grouped according to subtypes emerging from aim (i).(iii) To validate the newly identified candidate variants and assess their biological impact, we want to generate lymphoblastoid cell lines (LCLs) from the patients B cells and use CRISPR/Cas9 gene editing to correct the variants back to reference. Then we will compare the original and corrected LCLs according to their capacity to expand and to produce IgM. The results from this study will make it possible to relate different clinical sIgMID phenotypes to specific immunological defects and/or certain genetic variants. Such disease stratification will facilitate diagnosis and could improve therapy. Furthermore, I see sIgMID as a model disease and understanding sIgMID pathogenesis could not only shed light on other immune deficiencies but it will also provide insight into general immune mechanisms such as regulation of immunoglobulin production, T cell - B cell interaction and B cell maturation and differentiation.