Mechanisms of disease in patients with I?B? mutations

 描述（由申请人提供）： 常染色体显性遗传性外胚层发育不良伴免疫缺陷（AD ED-ID）的特征是毛发稀疏、锥形齿、汗腺数量减少和对严重感染的易感性。这是由于IκBα中的杂合突变损害了Ser 32和Ser 26磷酸化以及受体信号传导后突变蛋白的后续降解。非降解突变蛋白在细胞质中积累并隔离NFκB的p50和p65亚基，通过显性负效应（DN）损害经典NFκB途径的活化。 我们对9例AD ED-ID患者的基因型-表型分析表明，IκBα点突变患者的疾病严重程度和NFκ B依赖性TLR驱动的细胞因子产生的受损程度显著低于N端截短患者。我们假设点突变体可能比截短突变体更稳定，从而在更高水平上积累，导致更强的DN效应和更严重的疾病。我们的分析还表明，尽管供体嵌合性良好，但AD ED-ID患者在造血干细胞移植（HSCT）后的结局较差。 我们已经创建了IκBα S32 I突变的杂合子敲入小鼠，该突变重现了AD ED-ID患者的表型，S32 I小鼠不能发生接触性超敏反应（CHS）或产生抗体。引人注目的是，该突变体完全缺乏次级淋巴器官，包括淋巴结和派伊尔集合淋巴结，并且不能形成生发中心（GC），这是先前在AD ED-ID患者中未识别的缺陷性非经典NFκB信号传导的两个典型特征。在突变体中，LTβ R驱动的由经典和非经典NFκB途径介导的趋化因子和粘附分子的诱导作用受损，非经典NFκB途径的组成部分p100的水平显著降低。IκBα突变-> Rag 2-/-，而WT->IκBα突变体，骨髓嵌合体形成适当的淋巴器官，发展CHS，形成GC。这些结果表明，有缺陷的结构细胞功能是IκBα突变患者免疫缺陷和HSCT不良结局的基础，纠正这种小生境可能是重建其免疫功能的关键。 我们提出的假设是，IκBα突变蛋白的稳定性与患者细胞中残留的NFκB活性和p100水平呈负相关，从而决定疾病的严重程度和HSCT的结果。我们将检验来自S32 IκBα突变小鼠的间充质干细胞（MSC）p100水平降低和对LTβR连接反应受损的假设，并评估WT MSC植入作为AD ED-ID临床前模型的潜在治疗策略。

 DESCRIPTION (provided by applicant): Autosomal dominant ectodermal dysplasia with immune deficiency (AD ED-ID) is characterized by sparse hair, conical teeth, reduced number of sweat glands and susceptibility to severe infections. It is due to heterozygous mutations in IκBα that impair Ser32 and Ser26 phosphorylation and subsequent degradation of the mutant protein following receptor signaling. The non-degraded mutant protein accumulates and sequesters the p50 and p65 subunits of NFκB in the cytoplasm, impairing activation of the canonical NFκB pathway through a dominant negative (DN) effect. Our genotype-phenotype analysis of 9 patients with AD ED-ID indicates that disease severity and impairment of NFκB-dependent TLR-driven cytokine production are significantly worse in patients with IκBα point mutations than in those with N-terminal truncations. We hypothesize that the point mutants may be more stable than the truncation mutants, and thereby accumulate at a higher level, resulting in a stronger DN effect and more severe disease. Our analysis also shows that patients with AD ED-ID have a poor outcome after hematopoietic stem cell transplantation (HSCT) despite good donor chimerism. We have created a knock-in mouse heterozygous for the IκBα S32I mutation that recapitulates the phenotype of patients with AD ED-ID, The S32I mouse fails to develop contact hypersensitivity (CHS) or produce antibodies. Strikingly, the mutant completely lacks secondary lymphoid organs including lymph nodes and Peyer's patches, and fails to form germinal centers (GCs), two features typical of defective non-canonical NFκB signaling that were not previously recognized in AD ED-ID patients. LTβR-driven induction of chemokines and adhesion molecules mediated by both canonical and non-canonical NFκB pathways was impaired, and the levels of p100, a component of the non-canonical NFκB pathway, were markedly diminished in the mutant. IκBα mutant->Rag2-/-, but not WT->IκBα mutant, bone marrow chimeras formed proper lymphoid organs, developed CHS and formed GCs. These results suggest that defective architectural cell function underlies the immunodeficiency and poor outcome of HSCT in patients with IκBα mutations, and that correction of this niche may be critical for reconstituting their immune function. We propose to test the hypothesis that the stability of the IκBα mutant protein inversely correlates with the residual NFκB activity and th level of p100 in the patient cells, and thus determines disease severity and the outcome of HSCT. We will test the hypothesis that mesenchymal stem cells (MSCs) from mice with the S32I IκBα mutation have reduced p100 levels and impaired response to LTβR ligation, and to evaluate implantation of WT MSCs as a potential therapeutic strategy in a preclinical model for AD ED-ID.