Generation of human induced pluripotent stem cell lines (NIHTVBi011-A, NIHTVBi012-A, NIHTVBi013-A) from autosomal dominant Hyper IgE syndrome (AD-HIES) patients carrying STAT3 mutation.
ABSTRACT: Autosomal dominant Hyper IgE syndrome (AD-HIES), a rare immune deficiency affecting fewer than one per million people, is caused by heterozygous deleterious mutations in STAT3. STAT3 signaling plays crucial roles in basic cellular functions affecting broad aspects of cellular homeostasis. Accordingly, in addition to immunological deficits, patients experience severe multisystem non-immunological features. Human induced pluripotent stem cells (hiPSC) are well established as in vivo disease models for various human pathologies. We describe the generation of iPSC from three AD-HIES patients. These iPSCs express pluripotency markers, differentiate into three germ layers, have normal karyotype and similar genome identity to parental cells.
Project description:Autosomal dominant HIES (AD-HIES) is a primary immunodeficiency caused by dominant negative mutations in STAT3 clustered in the DNA binding and SH2 domains. Although in vitro differences in mutational constructs are observed, clinical phenotypic correlates of these genetic changes have not been described. We reviewed the charts of 65 AD-HIES patients (DNA binding n=35; SH2 n=30), recorded the components of the NIH HIES clinical scoring system as well as brain and coronary artery abnormalities and analyzed data by mutation region in adults and children. Patients with SH2 domain mutations had increased frequency of high palate, broad inter-alar distance, upper respiratory tract infections and, in the pediatric sub-group, significant scoliosis. There was suggestion of increased mortality for patients with DNA binding mutations. Although subtle differences in phenotype were observed to depend on the STAT3 genotype, overall the clinical phenotypes were similar between individuals with DNA binding and SH2 domain mutations.
Project description:Autosomal dominant hyper IgE syndrome (AD-HIES), or Job's syndrome, is a primary immune deficiency caused by dominant-negative mutations in STAT3. Recurrent Staphylococcus aureus skin abscesses are a defining feature of this syndrome. A widely held hypothesis that defects in peripheral Th17 differentiation confer this susceptibility has never been directly evaluated. To assess the cutaneous immune response in AD-HIES, we induced suction blisters in healthy volunteers (HVs) and patients with AD-HIES and then challenged the wound with lethally irradiated bacteria. We show that cutaneous production of IL-17A and IL-17F was normal in patients with AD-HIES. Overproduction of TNF-? differentiated the responses in AD-HIES from HVs. This was associated with reduced IL-10 family signaling in blister-infiltrating cells and defective epithelial cell function. Mouse models of AD-HIES recapitulated these aberrant epithelial responses to S. aureus and involved defective epithelial-to-mesenchymal transition (EMT) rather than a failure of bacterial killing. Defective responses in mouse models of AD-HIES and primary keratinocyte cultures from patients with AD-HIES could be reversed by TNF-? blockade and by drugs with reported modulatory effects on EMT. Our results identify these as potential therapeutic approaches in patients with AD-HIES suffering S. aureus infections.
Project description:There are more than 7000 described rare diseases, most lacking specific treatment. Autosomal-dominant hyper-IgE syndrome (AD-HIES, also known as Job's syndrome) is caused by mutations in STAT3. These patients present with immunodeficiency accompanied by severe nonimmunological features, including skeletal, connective tissue, and vascular abnormalities, poor postinfection lung healing, and subsequent pulmonary failure. No specific therapies are available for these abnormalities. Here, we investigated underlying mechanisms in order to identify therapeutic targets. Histological analysis of skin wounds demonstrated delayed granulation tissue formation and vascularization during skin-wound healing in AD-HIES patients. Global gene expression analysis in AD-HIES patient skin fibroblasts identified deficiencies in a STAT3-controlled transcriptional network regulating extracellular matrix (ECM) remodeling and angiogenesis, with hypoxia-inducible factor 1? (HIF-1?) being a major contributor. Consistent with this, histological analysis of skin wounds and coronary arteries from AD-HIES patients showed decreased HIF-1? expression and revealed abnormal organization of the ECM and altered formation of the coronary vasa vasorum. Disease modeling using cell culture and mouse models of angiogenesis and wound healing confirmed these predicted deficiencies and demonstrated therapeutic benefit of HIF-1?-stabilizing drugs. The study provides mechanistic insights into AD-HIES pathophysiology and suggests potential treatment options for this rare disease.
Project description:<b>Introduction:</b> Hyper-IgE Syndrome (HIES) is a rare inborn error of immunity (IEI) characterized by a constellation of symptoms related to susceptibility to <i>Staphylococcal</i> skin and pulmonary infections, eczema, raised serum IgE (>2,000 IU/ml), craniofacial anomalies, and recurrent bone fractures. Data on HIES from the Indian subcontinent is scarce and restricted to small case series and case reports. This is the first compilation of national data on HIES. <b>Materials and Methods:</b> A total 103 cases clinically diagnosed and treated as HIES were analyzed from nine centers. Cases with clinical and/or molecular diagnosis of DOCK8 deficiency were not included. Patients were divided into two groups: group I for whom a heterozygous rare variant of STAT3 was identified, and group II, with clinical features similar to those of AD STAT3 deficiency, but without any genetic diagnosis. <b>Results:</b> Genetic diagnosis was available in 27 patients (26.2%) and all harbored rare variants in the STAT3 gene. Majority of these STAT3 HIES patients presented with recurrent skin abscesses (77.7%) or pneumonia (62.9%) or both (59.2%). Other features included eczema (37%), candidiasis (55.5%), facial dysmorphism (55.5%), recurrent fractures (11.1%), and retained primary teeth (7.4%). <i>Mycobacterial</i> infections were seen in a significant 18.5%. Mortality was seen in three subjects (11.1%). A similar trend in the clinical presentation was observed when all the 103 patients were analyzed together. Twenty percent of patients without a rare variant in the STAT3 gene had an NIH score of ?40, whereas, 51.9% of STAT3 HIES subjects had scores below the cut off of ?40. TH17 cell numbers were low in 10/11 (90.9%) STAT3 HIES tested. Rare variants observed were 8 in exon 21; 8 in exon 13; 3 in exon 10; 2 in exon 15, and one each in exon 6, 16, 17, 19, 22, and splice site downstream of exon 12. Seven variants were novel and included F174S, N567D, L404Sfs<sup>*</sup>8, G419 =, M329K, T714I, R518X, and a splice site variant downstream of exon 12. <b>Conclusions:</b> The report includes seven novel STAT3 variants, including a rare linker domain nonsense variant and a CC domain variant. <i>Mycobacterial</i> diseases were more frequent, compared to western literature.
Project description:Background:Hyper-IgE syndrome (HIES) caused by loss-of-function (LOF) mutations in STAT3 gene (STAT3 LOF HIES) is associated with dental and facial abnormalities in addition to immunological defects. The role of STAT3 in the pathogenesis of the dental/facial features is, however, poorly elucidated. Objectives:Since mechanism of cellular resorption of mineralized tissues such as bone and teeth are similar, we attempted to study the expression of genes involved in bone homeostasis in STAT3 LOF HIES. Methods:Peripheral blood mononuclear cells from healthy controls (HCs), STAT3 LOF HIES patients, STAT3-/- PC-3 cells and STAT3+/+ LNCaP cells were stimulated with IL-6 and quantitative PCR array was performed to study the relative mRNA expression of 43 pre-selected genes. PCR array finding were further evaluated after stattic induced STAT3 inhibition. Results:Osteopontin (OPN) gene was seen to be significantly upregulated after IL-6 stimulation in HC (mean fold change 18.6, p?=?0.01) compared with HIES subjects. Inhibition of STAT3 signaling by stattic followed by IL-6 stimulation abrogated the OPN response in HCs suggesting that IL-6-induced STAT3 signaling regulates OPN expression. Bioinformatics analysis predicted the presence of STAT3 response element TTCCAAGAA at position -2005 of the OPN gene. Conclusion:Regulation of OPN gene through IL-6-mediated STAT3 activation and its significant dysregulation in STAT3 LOF HIES subjects could make OPN a plausible candidate involved in the pathogenesis of dental/facial manifestations in HIES.
Project description:RATIONALE:Hyper-IgE syndrome (HIES) is a rare primary immunodeficiency presenting as two forms including autosomal dominant HIES (AD-HIES) and autosomal recessive HIES (AR-HIES), which are mainly caused by mutations in STAT3 and DOCK8, respectively. To date, only about 500 cases have been reported worldwide including 37 cases in China. The spectrum and prevalence of mutations and molecular pathogenesis in HIES remain poorly understood. PATIENT CONCERNS:Here we reported two Chinese children presenting clinical manifestations of HIES. DIAGNOSIS:Based on medical history, clinical manifestations, and laboratory findings, a diagnosis of HIES was made for both children. Targeted next-generation sequencing (NGS) identified a novel heterozygous deletion of 15 bp (c.1960_1974del, p.G654_D658del or alternatively c.1966_1980del, and p.G656_D660del), and a recurrent missense mutation (c.1144C>T, p.R382W) in STAT3 in the two patients, respectively. INTERVENTIONS:The two patients have been given the successful treatment of skin infections with cefaclor. OUTCOMES:Both patients have been under follow-up for more than 6 months, with no signs of recurrent infections. LESSONS:Our results extend the spectrum of STAT3 mutations associated with ADHIES and highlight the value of targeted NGS in confirming diagnosis of genetic disorders.
Project description:Dominant negative mutations in the transcription-factor STAT3 underlie the rare primary immunodeficiency Job's syndrome. Allogeneic Hematopoietic Stem Cell Transplantation (HSCT) has shown promise in correction of the underlying immunological defect, with one report suggesting HSCT can prevent development of wider connective tissue complications. Here, we report the case of a 26 year old male who developed an acute ST-elevation myocardial infarction due to coronary artery ectasia and thrombosis, occurring despite pediatric allogeneic HSCT for STAT3-HIES and a predicted 10-year conventional cardiovascular risk of 0.1%. Vasculopathy associated with STAT3-HIES may persist or arise following HSCT and can precipitate life-threatening complications. This has implications for counseling and vascular surveillance, and highlights the need for further studies to determine the risk, pathogenesis, and optimal management of the vasculopathy associated with STAT3-HIES.
Project description:BACKGROUND AND AIMS:Autosomal dominant hyper-IgE (AD-HIES) is a primary immunodeficiency caused by mutations in STAT3. Elevated levels of IgE, an ineffective immune response, connective tissue abnormalities, and coronary arterial dilation and tortuosity characterize AD-HIES. To date, coronary artery evaluation in AD-HIES patients has been limited to lumenography measurements. Direct in vivo coronary vessel wall (VW) imaging may allow for better interrogation of coronary vessel abnormalities. The goal of this prospective study was to evaluate the coronary VW of AD-HIES patients using Magnetic Resonance Imaging (MRI) and histology. VW image findings were compared in healthy subjects and subjects with coronary atherosclerotic disease (CAD). METHODS:A total of 28 subjects (10 with AD-HIES, 8 healthy, 10 with CAD) were studied by coronary VW MRI imaging. Additionally, a post-mortem coronary artery from one VW imaged AD-HIES patient was examined. RESULTS:Coronary VW in AD-HIES was thicker than in healthy controls but not significantly different from VW thickness in CAD subjects. AD-HIES coronaries showed increased VW area compared to healthy controls and CAD subjects. On histology, the AD-HIES coronary artery had findings consistent with atherosclerotic plaque, but had minimal luminal narrowing, deficient adventitia thickening and absence of both internal and external elastic laminae. CONCLUSIONS:This is the first study to demonstrate subclinical coronary atherosclerosis in AD-HIES patients on VW imaging by MRI. Histologic evaluation confirmed the presence of atherosclerosis with lack of supportive adventitial thickening and elastic components. These findings suggest mechanisms for coronary dilation in AD-HIES and thereby help direct clinical management.
Project description:The hyper IgE syndrome (HIES) is characterized by abscesses, eczema, recurrent infections, skeletal and connective tissue abnormalities, elevated serum IgE, and diminished inflammatory responses. It exists as autosomal-dominant and autosomal-recessive forms that manifest common and distinguishing clinical features. A majority of those with autosomal-dominant HIES have heterozygous mutations in signal transducer and activator of transcription (STAT)-3 and impaired T(H)17 differentiation.To elucidate mechanisms underlying different forms of HIES.A cohort of 25 Turkish children diagnosed with HIES were examined for STAT3 mutations by DNA sequencing. Activation of STAT3 by IL-6 and IL-21 and STAT1 by IFN-alpha was assessed by intracellular staining with anti-phospho (p)STAT3 and -pSTAT1 antibodies. T(H)17 and T(H)1 cell differentiation was assessed by measuring the production of IL-17 and IFN-gamma, respectively.Six subjects had STAT3 mutations affecting the DNA binding, Src homology 2, and transactivation domains, including 3 novel ones. Mutation-positive but not mutation-negative subjects with HIES exhibited reduced phosphorylation of STAT3 in response to cytokine stimulation, whereas pSTAT1 activation was unaffected. Both patient groups exhibited impaired T(H)17 responses, but whereas STAT3 mutations abrogated early steps in T(H)17 differentiation, the defects in patients with HIES with normal STAT3 affected more distal steps.In this cohort of Turkish children with HIES, a majority had normal STAT3, implicating other targets in disease pathogenesis. Impaired T(H)17 responses were evident irrespective of the STAT3 mutation status, indicating that different genetic forms of HIES share a common functional outcome.
Project description:Heterozygous in-frame mutations in coding regions of human STAT3 underlie the only known autosomal dominant form of hy- per IgE syndrome (AD HIES). About 5% of familial cases remain unexplained. The mutant proteins are loss-of-function and dominant-negative when tested following overproduction in re- cipient cells. However, the production of mutant proteins has not been detected and quantified in the cells of heterozygous pa- tients. We report a deep intronic heterozygous STAT3 mutation, c.1282-89C>T, in 7 relatives with AD HIES. This mutation creates a new exon in the STAT3 complementary DNA, which, when over- expressed, generates a mutant STAT3 protein (D427ins17) that is loss-of-function and dominant-negative in terms of tyrosine phos- phorylation, DNA binding, and transcriptional activity. In immor- talized B cells from these patients, the D427ins17 protein was 2 kDa larger and 4-fold less abundant than wild-type STAT3, on mass spectrometry. The patients’ primary B and T lymphocytes responded poorly to STAT3-dependent cytokines. These findings are reminiscent of the impaired responses of leukocytes from other patients with AD HIES due to typical STAT3 coding muta- tions, providing further evidence for the dominance of the mutant intronic allele. These findings highlight the importance of sequenc- ing STAT3 introns in patients with HIES without candidate variants in coding regions and essential splice sites. They also show that AD HIES-causing STAT3 mutant alleles can be dominant-negative even if the encoded protein is produced in significantly smaller amounts than wild-type STAT3. Overall design: Examination of gene expression in EBV-B cells from healthy subjects and AD HIES patients stimulated with IL-21, IL-6, IL-23, IL-10, and IFNa