Characterization of a novel combined immunodeficiency with skeletal dysplasia

一种新型联合免疫缺陷伴骨骼发育不良的特征

基本信息

批准号：
8995190
负责人：
Luigi Daniele Notarangelo
金额：
$ 22.13万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-01-15 至 2016-09-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8995190
关键词：
Affect Architecture Autoantigens Autologous Biological Branchial arch structure CD4 Positive T Lymphocytes CD8B1 gene CRISPR/Cas technology Cartilage Cell surface Cells Data Defect Development Disease Disease model Dysplasia Elderly Eosinophilia Exfoliative Dermatitis FOXP3 gene Family Female Gene Mutation Generations Genes Hair Health Hematopoietic Stem Cell Transplantation Heparan Sulfate Biosynthesis Heparan Sulfate Proteoglycan Heparitin Sulfate Human IgE Immune Immune system Immunity Immunologic Deficiency Syndromes In Vitro Infant Infection Lead Limb Bud Link Lymphatic Diseases Lymphopenia Mesenchyme Missense Mutation Mitogens Molecular Mus Mutation Neural Crest Organogenesis Parents Patients Pectoral Peripheral Phenotype Play Proteoglycan Recurrence Regulatory T-Lymphocyte Reporting Role Serum Severe Combined Immunodeficiency Siblings Skeletal Development Syndrome System Systems Development T cell differentiation T-Cell Development T-Lymphocyte Technology Testing Therapeutic Thymic epithelial cell Thymus Gland Tissues Zebrafish base cell type congenital immunodeficiency disease phenotype exome sequencing gene correction glycosyltransferase induced pluripotent stem cell insight male member morphogens mouse model mutant novel polysulfated glycosaminoglycan skeletal skeletal dysplasia thymocyte thymus transplantation transcription factor

项目摘要

DESCRIPTION (provided by applicant): Immuno-osseous dysplasias (IOD) are a group of disorders characterized by immune deficiency and skeletal dysplasia. Schimke's disease (SIOD) and cartilage hair hypoplasia (CHH) represent the prototypic forms of these disorders, but several other cases of IOD with unknown molecular basis have been described. The immunodeficiency seen in IOD is often severe, and affects predominantly T lymphocytes. Immune dysregulation typical of Omenn syndrome (OS) has been reported in some patients with CHH. We have identified and studied two siblings (a male and a female) with rapidly progressive and fatal IOD. Both infants manifested T cell lymphopenia with markedly reduced number of CD8+ T cells, lack of proliferation to mitogens, hypogammaglobulinemia, increased serum IgE and eosinophilia. The elder sibling had features of OS, with erythroderma and expansion of oligoclonal and activated CD4+ T cells. No mutations were identified in the SMARCAL1 and RMRP genes, responsible for SIOD and CHH, respectively. Whole exome sequencing (WES) revealed that both siblings were homozygous, and both parents heterozygous, for a novel and drastic missense mutation (R339W) affecting a highly conserved residue of Exostosin-like 3 (EXTL3), a member of the exostosin (EXT) family of glycosyltransferases involved in heparan sulfate (HS) biosynthesis. HS are sulfated glycosaminoglycans that decorate cell surface and matrix proteoglycans. HS proteoglycans modulate the activity of morphogens that play a key role in skeletal development and T cell differentiation. Our overall hypothesis is that EXTL3 is critical for thymic development and that EXTL3 mutations in humans cause a novel form of IOD. In support of our hypothesis, our preliminary data demonstrate that EXTL3 is expressed by human thymocytes, thymic epithelial cells (TECs), and peripheral T cells. Moreover, mutation of Extl3 in zebrafish causes defects of limb bud development, and impaired thymic T cell development. To test our hypothesis, and to investigate the cellular basis of the immunodeficiency associated with EXTL3 mutations, we will generate and characterize mouse models with tissue-specific disruption of the Extl3 gene in neural crest mesenchyme cells, TECs, or thymocytes. All of these cell types produce HS and are critically involved in thymus organogenesis and T cell development. In parallel, in order to demonstrate the disease-causing effect of the EXTL3 p.R339W mutation identified in the affected siblings, we will perform disease modeling using patient-derived induced pluripotent stem cells (iPSCs). We will use the CRISPR/Cas9 technology to correct the EXTL3 p.R339W mutation in patient-derived iPSCs. In order to define the cell autonomous effect of the EXTL3 mutation, we will differentiate both mutant and gene-corrected iPSCs to TECs and T cells, and we will study HS biosynthesis under the same experimental conditions. We anticipate that this study will provide novel important insights on the biological role of HSPGs in immune system development and function, and will help define the optimal therapeutic approach to correct the immunodeficiency of this condition.

描述（由适用提供）：免疫 - 骨不典型增生（IOD）是一组具有免疫缺陷和骨骼发育不良的疾病。 Schimke病（SIOD）和软骨发育不全（CHH）代表了这些疾病的原型形式，但已经描述了其他几例具有未知分子基础的IOD病例。 IOD中看到的免疫缺陷通常很严重，主要影响T淋巴细胞。某些CHH患者已经报道了Omenn综合征（O）的典型免疫失调。我们已经确定并研究了两个具有快速进步和致命的iod的兄弟姐妹（男性和女性）。两名婴儿都表现出T细胞淋巴细胞减少，CD8+ T细胞数量明显减少，缺乏促分裂剂的增殖，低脂脂蛋白血症，血清IgE和嗜酸性粒细胞增多。老年兄弟姐妹具有OS的特征，具有红细胞和寡克隆和活化的CD4+ T细胞的扩展。在SMARCAL1和RMRP基因中分别鉴定出突变，分别负责SIOD和CHH。整个外显子组测序（WES）表明，两个兄弟姐妹都是纯合的，父母都是杂合的，用于一种新颖而又剧烈的错义突变（R339W），影响了雌酮蛋白样3（extl3）的高度保守的住所，exososin（Ext）（Extsososin（Ext）的glycososylesylesyltrantase sultrants sulfate sulfat sulflat sulfff insparan sulfff insparan sulfff insparan（extl3）。 HS是装饰细胞表面和基质蛋白聚糖的硫酸糖胺聚糖。 HS蛋白聚糖调节形态剂的活性，在骨骼发育和T细胞分化中起关键作用。我们的总体假设是，extl3对于胸腺发育至关重要，并且人类中的extl3突变引起了一种新型的IOD。为了支持我们的假设，我们的初步数据表明，extl3由人胸腺细胞，胸腺上皮细胞（TEC）和外周T细胞表达。此外，斑马鱼中ext3的突变会导致肢体发育的缺陷，并损害胸腺T细胞的发育。为了检验我们的假设，并研究了与extl3突变相关的免疫缺陷的细胞基础，我们将在神经rest间充质细胞，TEC或胸腺细胞中生成和表征extl3基因的组织特异性破坏的小鼠模型。所有这些细胞类型都会产生HS，并与胸腺器官发生和T细胞发育至关重要。同时，为了证明在受影响的兄弟姐妹中鉴定出的EXTL3 P.R339W突变的引起疾病的作用，我们将使用患者衍生的诱导多能干细胞（IPSC）进行疾病建模。我们将使用CRISPR/CAS9技术来纠正患者衍生的IPSC中的EXTL3 P.R339W突变。为了定义extl3突变的细胞自主效应，我们将将突变体和基因校正的IPSC分化为TEC和T细胞，我们将在相同的实验条件下研究HS生物合成。我们预计，这项研究将为HSPG在免疫系统开发和功能中的生物学作用提供新的重要见解，并有助于定义最佳的治疗方法，以纠正该疾病的免疫缺陷。