STAT1 Gain-of-function Mutations Research Articles

STAT1 gain of function mutation impairs immune response to viral infections

Abstract The coordinated effort of the immune system in host defense is dependent upon the proper release and interpretation of different cytokine signals. Several examples of monogenic diseases targeting the JAK-STAT pathway reveal there is still a critical need to understand basic principles of cytokine signaling and cytokine output. For example, patients with STAT1 gain-of-function (GOF) mutations exhibit a type 1/IFN-gamma bias that antagonizes a type 3/IL-17 immune response important for controlling fungal infection, yet patients also, paradoxically, exhibit chronic and sometimes lethal viral infections. Using a novel conditional knock in STAT1-GOF mouse model, we demonstrate that STAT1-GOF mice respond poorly to viral infections that are easily controlled by WT mice, such as MCMV and LCMV-Armstrong. High-dimensional flow cytometry and scRNA-seq reveal an impaired NK and CD8 T cell effector response in STAT1-GOF mice that is not simply due to an exhaustion phenotype. Instead, STAT1-GOF mice respond poorly to viral infections due to an unexpected defect in IFN-gamma production early during the innate immune response, leading to a cytokine storm, immunopathology, and an impaired adaptive immune response. scATAC/RNA-seq uncovers altered usage of different STAT complexes in STAT1-GOF mice, leading to a misinterpretation of early innate-stimulating cytokines. These results provide insight into why patients with interferonopathies or elevated interferons respond poorly to viral infections and highlight the importance of properly tuning STAT1 during an immune response. R.L.P. was supported by a Postdoctoral Research Associate (PRAT) fellowship from the National Institute of General Medical Sciences (NIGMS), award number 1Fi2GM137942-01

STAT3 gain-of-function mutations connect leukemia with autoimmune disease by pathological NKG2Dhi CD8+ T cell dysregulation and accumulation

The association between cancer and autoimmune disease is unexplained, exemplified by Tcell large granular lymphocytic leukemia (T-LGL) where gain-of-function (GOF) somatic STAT3 mutations correlate with co-existing autoimmunity. To investigate whether these mutations are the cause or consequence of CD8+ Tcell clonal expansions and autoimmunity, we analyzed patients and mice with germline STAT3 GOF mutations. STAT3 GOF mutations drove the accumulation of effector CD8+ Tcell clones highly expressing NKG2D, the receptor for stress-induced MHC-class-I-related molecules. This subsetalso expressed genes for granzymes, perforin, interferon-γ, and Ccl5/Rantes and required NKG2D and the IL-15/IL-2 receptor IL2RB for maximal accumulation. Leukocyte-restricted STAT3 GOF was sufficient and CD8+ Tcells were essential for lethal pathology in mice. These results demonstrate that STAT3 GOF mutations cause effector CD8+ Tcell oligoclonal accumulation and that these rogue cells contribute to autoimmune pathology, supporting the hypothesis that somatic mutations in leukemia/lymphoma driver genes contribute to autoimmune disease.

Inborn errors of immunity with loss- and gain-of-function germline mutations in STAT1.

Signal transducer and activator of transcription 1 (STAT1) is a well-known and pivotal molecule in the regulation of the Janus activating kinases (JAK)-STAT pathway, an extensive signalling pathway downstream of several cytokine receptors that plays an indispensable role in immune defence in humans. In the JAK-STAT pathway, STAT1 homodimer-dependent IFN-γ immunity is crucial for defence against intra-macrophagic bacteria, and STAT1/STAT2/IRF9 complex-dependent IFN-α/β immunity is essential for protection against viruses. STAT1 dysfunction causes a wide range of immune dysregulation phenotypes, which have been classified into four disease types, namely, (i) autosomal recessive (AR) complete STAT1 deficiency, (ii) AR partial STAT1 deficiency, (iii) autosomal dominant (AD) STAT1 deficiency, and (iv) AD STAT1 gain-of-function (GOF), based on their mode of inheritance and function. Disease types (i, ii, and iii) are caused by STAT1 loss-of-function (LOF) mutations, whereas disease type (iv) is caused by STAT1 GOF mutations. Notably, rapid and accurate diagnosis of these four diseases can be difficult. In particular, the differential diagnosis of AD STAT1 deficiency and AD STAT1 GOF, which are caused by LOF- and GOF-STAT1 mutations, respectively, can be complicated due to their overlapping clinical symptoms. In addition, more than 150 pathological mutations have been identified in STAT1 across all domains, making functional prediction of novel mutations difficult. Thus, the functional analysis of mutations is necessary for diagnosis. The recently proposed reference database of mutant STAT1 functions based on comprehensive alanine scanning is a useful tool for evaluating unknown STAT1 mutations with high accuracy.

RECURRENT APHTHOUS STOMATITIS AND ORAL CANDIDIASIS DUE TO STAT1 MUTATION

<h3>Introduction</h3> Chronic mucocutaneous candidiasis (CMC) often presents with chronic <i>Candida</i> infections of the skin, nails or mucous membranes in the absence of invasive fungal infections. Various genetic defects are associated with CMC. Here, we describe a case of recurrent aphthous stomatitis and chronic oral candidiasis due to gain-of-function (GOF) mutation in STAT1. <h3>Case Description</h3> A 21-year-old male presented to the Allergy and Immunology clinic with long-standing history of recurrent aphthous ulcers and oral candidiasis. He had previously undergone evaluation as a child with normal immunoglobulin levels, normal antibody response to vaccines and normal delayed-type hypersensitivity skin test to <i>Candida</i> antigen, but low absolute NK cells. Targeted genetic testing was negative for APECED and IPEX. He continued to experience frequent aphthous ulcers and oral thrush that can be debilitating at times, with one episode resulting in hospital admission due to severe mucosal pain and PO intolerance. Repeat immunology evaluation was similar to prior. He had low absolute NK cells, and NK cell function study using chromium release assay was notably decreased. A comprehensive primary immunodeficiency next generation sequencing panel identified a pathogenic heterozygous c.800C>T (p.A267V) mutation in STAT1. <h3>Discussion</h3> While fungal infection of the oral mucosa is a main clinical feature of CMC, aphthous stomatitis can be found frequently in patients with STAT1 GOF mutations as well. These patients also frequently have decreased NK cells and NK cell function. It is important to be aware of these additional phenotypes in the evaluation of a patient with CMC.

Systems Immunology Analyses of STAT1 Gain-of-Function Immune Phenotypes Reveal Heterogeneous Response to IL-6 and Broad Immunometabolic Roles for STAT1.

Patients with STAT1 gain-of-function (GOF) pathogenic variants have enhanced or prolonged STAT1 phosphorylation following cytokine stimulation and exhibit increased yet heterogeneous susceptibility to infections, autoimmunity, and cancer. Although disease phenotypes are diverse and other genetic factors contribute, how STAT1 GOF affects cytokine sensitivity and cell biology remains poorly defined. In this study, we analyzed the immune and immunometabolic profiles of two patients with known pathogenic heterozygous STAT1 GOF mutation variants. A systems immunology approach of peripheral blood cells from these patients revealed major changes in multiple immune cell compartments relative to healthy adult and pediatric donors. Although many phenotypes of STAT1 GOF donors were shared, including increased Th1 cells but decreased class-switched B cells and plasmacytoid dendritic cell populations, others were heterogeneous. Mechanistically, hypersensitivity for cytokine-induced STAT1 phosphorylation in memory T cell populations was particularly evident in response to IL-6 in one STAT1 GOF patient. Immune cell metabolism directly influences cell function, and the STAT1 GOF patients shared an immunometabolic phenotype of heightened glucose transporter 1 (GLUT1) and carnitine palmitoyl transferase 1A (CPT1a) expression across multiple immune cell lineages. Interestingly, the metabolic phenotypes of the pediatric STAT1 GOF donors more closely resembled or exceeded those of healthy adult than healthy age-similar pediatric donors, which had low expression of these metabolic markers. These results define new features of STAT1 GOF patients, including a differential hypersensitivity for IL-6 and a shared increase in markers of metabolism in many immune cell types that suggests a role for STAT1 in metabolic regulation of immunity.

Chronic demodicosis in patients with immune dysregulation: An unexpected infectious manifestation of Signal transducer and activator of transcription (STAT)1 gain-of-function.

Signal transducer and activator of transcription (STAT)1 heterozygous gain-of-function (GOF) mutations are known to induce immune dysregulation and chronic mucocutaneous candidiasis (CMCC). Previous reports suggest an association between demodicosis and STAT1 GOF. However, immune characterization of these patients is lacking. Here, we present a retrospective analysis of patients with immune dysregulation and STAT1 GOF who presented with facial and ocular demodicosis. In-depth immune phenotyping and functional studies were used to characterize the patients. We identified five patients (three males) from two non-consanguineous Jewish families. The mean age at presentation was 11.11 (range=0.58-24) years. Clinical presentation included CMCC, chronic demodicosis and immune dysregulation in all patients. Whole-exome and Sanger sequencing revealed a novel heterozygous c.1386C>A; p.S462R STAT1 GOF mutation in four of the five patients. Immunophenotyping demonstrated increased phosphorylated signal transducer and activator of transcription in response to interferon-α stimuli in all patients. The patients also exhibited decreased T cell proliferation capacity and low counts of interleukin-17-producing T cells, as well as low forkhead box protein 3+ regulatory T cells. Specific antibody deficiency was noted in one patient. Treatment for demodicosis included topical ivermectin and metronidazole. Demodicosis may indicate an underlying primary immune deficiency and can be found in patients with STAT1 GOF. Thus, the management of patients with chronic demodicosis should include an immunogenetic evaluation.

STAT1 gain of function mutation impairs immune response to viral infections

Abstract Regulation of T cells by interferons and other cytokines, which act via STAT family transcription factors, is critical for host defense. Patients with STAT1 gain-of-function (GOF) mutations exhibit heightened STAT1 activation after cytokine stimulation and develop lifelong ailments, including chronic infections, autoimmunity, and cancer. A characteristic feature of this disorder is chronic mucocutaneous candidiasis (CMC), related to exagerated type 1/IFNγ bias that antagonizes a type 3/Th17 response required for fungal clearance. However, this type 1/IFNγ bias does not explain why almost 40% of reported patients exhibit chronic and sometimes lethal viral infections. This observation is unexpected, as cells with STAT1-GOF mutations exhibit increased expression of interferon stimulated genes following IFNα/β and IFNγ stimulation and patients show more IFNγ-producing CD4 T cells. This paradox emphasizes the critical need to understand basic principles of cytokine signaling, specifically how a STAT1-GOF mutation alters cytokine output. Using a novel conditional knock-in mouse expressing a common GOF STAT1 mutation, T385M, we found that this model recapitulates observations in patients (i.e. elevated STAT1 activation, increased total STAT1 protein, susceptibility to CMC), as well as a strong type 1/interferon bias at steady state. Despite this type 1 bias, STAT1-GOF exhibit impaired NK and CD8 T cell response to systemic viral infections. These results reveal that STAT1 activity must be tuned for proper immune response to viral infections.

STAT Immunodeficiency Disorders and Fungal Infection Susceptibility

Invasive fungal diseases (IFD) or complicated mucocutaneous fungal infections can occur secondary to both primary and acquired immunodeficiency disorders. The signal transducer and activator of transcription (STAT) proteins are critical transcription factors involved in a wide range of processes including the immune response. This review highlights the immune pathways, clinical phenotypes and current management options of fungal infections in patient with mutations in STAT1 and STAT3, the most relevant STAT molecules involved in the host response to fungal pathogens. STAT1 gain of function (GOF) and STAT3 loss of function (LOF) (cause of autosomal dominant Hyper IgE syndrome) mutations have been associated with both chronic mucocutaneous candidiasis (CMC) and invasive fungal infections, not only inhaled moulds, mostly Aspergillus spp., but also Pneumocystis, Cryptococcus and other endemic dimorphic fungi. Significant progress has been made in the insights into the immunopathogenesis of fungal infections in these patients; nevertheless, the concrete mechanisms involved are still not fully elucidated. Early initiation of antifungal therapy followed by long-term secondary prophylaxis is normally required. Some adjunctive therapies have also been employed, although their use has not been completely established. Besides a developing knowledge in STAT1 GOF and STAT3 LOF mutations, additional research is needed. It should aim for a better understanding of the pathogenesis, diagnosis, and management of fungal infections in these patients. The identification of new therapeutic targets should help to improve patient quality of life and outcomes.

PULMONARY HYPERTENSION IN A CHILD WITH STAT3 GAIN OF FUNCTION MUTATION-ASSOCIATED INTERSITIAL LUNG DISEASE

SESSION TITLE: Fellows Pediatrics Posters SESSION TYPE: Fellow Case Report Posters PRESENTED ON: October 18-21, 2020 INTRODUCTION: STAT3 gain of function mutations are a recently described cause of autoimmunity that affect multiple organ systems, including the lungs. CASE PRESENTATION: The patient was first diagnosed with autoimmune lymphoproliferative syndrome (ALPS) at age 2 in the setting of lymphadenopathy, hepatosplenomegaly, & autoimmune cytopenias. Treatment was initially steroids & IVIG; he later started sirolimus as a steroid-sparing agent. No genetic defect consistent with ALPS was identified though he was found to have short telomeres, of unclear significance as he did not fit a dyskeratosis congenita phenotype. Further evaluation revealed a gain of function (GOF) mutation in STAT3. At age 9, he developed dyspnea & syncope; evaluation revealed mild pulmonary hypertension (PH) with positive vasoreactivity testing and CT chest with upper-lobe predominant subpleural cystic lung disease & scattered pulmonary nodules. Immunosuppression was switched from sirolimus to mycophenolate. For the PH, he was prescribed dual oral therapy, with symptomatic improvement. He developed hypoxemia at age 11. VQ scan was notable for markedly heterogeneous perfusion despite homogenous ventilation & PFTs revealed a reduced DLCO with mild restriction. Given concern for worsening pulmonary disease in the setting of the STAT3 mutation, he was started on tocilizumab (in place of mycophenolate) with partial improvement in hypoxemia. At age 12, given persistent oxygen requirement & CT showing new upper lobe predominant mosaic attenuation with groundglass (& overall stable cystic lung disease), he started ruloxitinib with some improvement in his dyspnea & hypoxemia. Since tocilizumab & ruloxitinib were initiated, his TLC & DLCO have normalized. DISCUSSION: In the largest series of patients with STAT3 GOF autoimmunity, 36% were reported to have interstitial lung disease (ILD), though none had cystic lung disease1. Upper-lobe predominant subpleural cystic disease, as seen in our patient, is most reminiscent of that often incidentally found in Trisomy 21, and has not been described in other pediatric populations2. In addition to autoimmunity related to STAT3 GOF involving the lungs, there is evidence that inappropriate activation of STAT3 is involved in the development of fibrosis in patients with UIP/IPF, suggestive of a potential alternative pathway to ILD in these patients3. Pulmonary hypertension has not previously been described as a consequence of STAT3 GOF mutations. Our patient developed PH years before he became hypoxemic, suggestive that his PH is not simply due to chronic hypoxemia. CONCLUSIONS: In this patient, STAT3 GOF mutation was associated with cystic lung disease and pulmonary hypertension, manifestations that have not previously been described. Reference #1: Fabre A, Marchal S, Barlogis V, et al. Clinical aspects of STAT3 gain-of-function germline mutations: a systematic review. J Allergy Clin Immunol Pract. 2019;7(6):1958-69. Reference #2: George M, Amodio J, Lee H. Cystic lung disease in Down syndrome: a case report and literature review. Case Rep Pediatr. 2016;2016:4048501. Reference #3: Pechkovsky DV, Prele CM, Wong J, et al. STAT3-mediated signaling dysregulates lung fibroblast-myofibroblast activation and differentiation in UIP/IPF. AJP. 2012;180(4):1398-1412. DISCLOSURES: No relevant relationships by Sarah Cohen, source=Web Response No relevant relationships by Stephen Kirkby, source=Web Response

Three Copies of Four Interferon Receptor Genes Underlie a Mild Type I Interferonopathy in Down Syndrome.

Down syndrome (DS) is characterized by the occurrence of three copies of human chromosome 21 (HSA21). HSA21 contains a cluster of four interferon receptor (IFN-R) genes: IFNAR1, IFNAR2, IFNGR2, and IL10RB. DS patients often develop mucocutaneous infections and autoimmune diseases, mimicking patients with heterozygous gain-of-function (GOF) STAT1 mutations, which enhance cellular responses to three types of interferon (IFN). A gene dosage effect at these four loci may contribute to the infectious and autoimmune manifestations observed in individuals with DS. We report high levels of IFN-αR1, IFN-αR2, and IFN-γR2 expression on the surface of monocytes and EBV-transformed-B (EBV-B) cells from studying 45 DS patients. Total and phosphorylated STAT1 (STAT1 and pSTAT1) levels were constitutively high in unstimulated and IFN-α- and IFN-γ-stimulated monocytes from DS patients but lower than those in patients with GOF STAT1 mutations. Following stimulation with IFN-α or -γ, but not with IL-6 or IL-21, pSTAT1 and IFN-γ activation factor (GAF) DNA-binding activities were significantly higher in the EBV-B cells of DS patients than in controls. These responses resemble the dysregulated responses observed in patients with STAT1 GOF mutations. Concentrations of plasma type I IFNs were high in 12% of the DS patients tested (1.8% in the healthy controls). Levels of type I IFNs, IFN-Rs, and STAT1 were similar in DS patients with and without recurrent skin infections. We performed a genome-wide transcriptomic analysis based on principal component analysis and interferon modules on circulating monocytes. We found that DS monocytes had levels of both IFN-α- and IFN-γ-inducible ISGs intermediate to those of monocytes from healthy controls and from patients with GOF STAT1 mutations. Unlike patients with GOF STAT1 mutations, patients with DS had normal circulating Th17 counts and a high proportion of terminally differentiated CD8+ T cells with low levels of STAT1 expression. We conclude a mild interferonopathy in Down syndrome leads to an incomplete penetrance at both cellular and clinical level, which is not correlate with recurrent skin bacterial or fungal infections. The constitutive upregulation of type I and type II IFN-R, at least in monocytes of DS patients, may contribute to the autoimmune diseases observed in these individuals.

Distinct molecular response patterns of activating STAT3 mutations associate with penetrance of lymphoproliferation and autoimmunity

Germline STAT3 gain-of-function (GOF) mutations have been linked to poly-autoimmunity and lymphoproliferation with variable expressivity and incomplete penetrance. Here we studied the impact of 17 different STAT3 GOF mutations on the canonical STAT3 signaling pathway and correlated the molecular results with clinical manifestations. The mutations clustered in three groups. Group 1 mutants showed altered STAT3 phosphorylation kinetics and strong basal transcriptional activity. They were associated with the highest penetrance of lymphoproliferation and autoimmunity. Group 2 mutants showed a strongly inducible transcriptional reporter activity and were clinically less penetrant. Group 3 mutants were mostly located in the DNA binding domain and showed the strongest DNA binding affinity despite a poor transcriptional reporter response. Thus, the GOF effect of STAT3 mutations is determined by a heterogeneous response pattern at the molecular level. The correlation of response pattern and clinical penetrance indicates a significant contribution of mutation-determined effects on disease manifestations.

STAT1 Gain-of-Function Mutations Cause High Total STAT1 Levels With Normal Dephosphorylation.

Signal transducer and activator of transcription (STAT1)1 gain of function (GOF) pathogenic variants have been associated with increased levels of phosphorylated STAT1 and STAT1-dependent cellular responses. Delayed dephosphorylation was proposed as the underlying mechanism leading to the characteristically raised pSTAT1 levels. We examined the levels of STAT1 protein and message as well as rates of STAT1 phosphorylation, dephosphorylation, and degradation associated with STAT1 GOF pathogenic variants. Fresh peripheral blood mononuclear cells (PBMC) from 14 STAT1 GOF patients carrying 10 different pathogenic variants in the coiled-coil, DNA binding, and SH2 domains and healthy donors were used to study STAT1 levels and phosphorylation (pSTAT1) following IFNγ and IFNα stimulation. STAT1 protein levels were measured by flow cytometry and immunoblot. STAT1 mRNA levels were measured using quantitative reverse transcription PCR. STAT1 protein degradation was studied using cycloheximide. Patient IFNγ and IFNα induced peak pSTAT1 was higher than in healthy controls. The velocity of pSTAT1 dephosphorylation after treatment of IFNγ stimulated CD14+ monocytes with the Janus Kinase (JAK)-inhibitor ruxolitinib was significantly faster in patient cells. STAT1 protein levels in patient CD14+ monocytes and CD3+ T cells were higher than in healthy donors. There was a strong and positive correlation between CD14+ STAT1 protein levels and peak pSTAT1 levels. Patient fresh PBMC STAT1 mRNA levels were increased at rest and after 16 h of incubation. STAT1 protein degradation was similar in patient and healthy volunteer cells. Patient IFNγ receptors 1 and 2 and JAK2 levels were normal. One patient in our cohort was treated with the oral JAK inhibitor ruxolitinib. Treatment was associated with normalization of both STAT1 protein and peak pSTAT1 levels. After JAK inhibitor treatment was stopped the patient's CD14+ monocyte STAT1 protein and peak phosphorylation levels increased proportionally. These findings suggest that patients with STAT1 GOF mutations have higher levels of total STAT1 protein, leading to high levels of pSTAT1 after stimulation, despite rapid STAT1 dephosphorylation and normal degradation.

Type 1 Diabetes in STAT Protein Family Mutations: Regulating the Th17/Treg Equilibrium and Beyond.

Improvements in the immunological, molecular, and genetic technologies such as next-generation sequencing have led to an exponential increase in the number of monogenic immune dysregulatory syndromes diagnosed, where type 1 diabetes (T1D) forms part of the autoimmune manifestations. Here, we reviewed the mutations in the signal transducer and activator of transcription (STAT) protein family, namely gain-of-function (GOF) mutations in STAT1 and STAT3 as well as STAT5b deficiency, that show strong association to T1D susceptibility. The equilibrium of T-helper 17 (Th17) and regulatory T cells (Tregs) is often found altered in patients affected by STAT GOF mutations. While the increased number of Th17 cells and the concomitant decrease in Treg cells may explain T1D in STAT3 GOF patients, the reduced number of Th17 cells found in those carrying STAT1 GOF mutations added a new level of complexity on the exact role of Th17 in the pathogenesis of T1D. Here, we describe the possible mechanisms through which STAT3 and STAT1 GOF mutations may perturb the fate and function of Th17 and Tregs and explore how this may lead to the development of T1D. We propose that the study of monogenic diseases, and in particular STAT mutations, may not only improve our understanding of the function of the human immune system but also shed light onto the pathogenic mechanisms of T1D and the genetic variants that confer predisposition to the disease.

Dysregulatory syndromes: the role of signal transducers and activators of transcription.

A comparative description of dysregulatory syndromes with mutations in signal transducer and activator of transcription (STAT) genes. STAT 1, 3 and 5b loss of function (LOF) and gain of function (GOF) mutations are a heterogeneous group of genetic disorders that range from immunodeficiency (ID) to autoimmune disease (AID), depending on the underlying signalling pathway defect. Between them, there are clear overlapping and differences in clinical presentation and laboratory findings. Dysregulatory syndromes due to LOF and GOF mutations in STAT1, 3 and 5b are a particular group of primary immunodeficiencies (PIDs) in which AID may be the predominant finding in addition to infections susceptibility. STAT1 GOF mutations were described as the major cause of chronic mucocutaneous candidiasis, while activating STAT3 mutations result in early-onset multiorgan autoimmunity and ID. Human STAT5b deficiency is a rare disease that also involves ID and severe growth failure. In recent years, the identification of the genes involved in these disorders allowed to differentiate these overlapping syndromes in order to choose the most effective therapeutic options.

Human TH9 differentiation is dependent on signal transducer and activator of transcription (STAT) 3 to restrain STAT1-mediated inhibition

Patients with loss-of-function (LOF) signal transducer and activator of transcription 3 (STAT3) mutations have dermatitis, enhanced IgE production despite a relative lack of immediate hypersensitivity, recurrent infection, and an increased rate of lymphoma in addition to a number of skeletal and connective tissue abnormalities. Patients with STAT1 gain-of-function (GOF) mutations also have susceptibility to candidiasis and sinopulmonary infection, as well as autoimmunity and squamous cell carcinoma, in addition to even more broad phenotypes. Because of the link between TH9 cells and allergic inflammation, autoimmunity, and antitumor surveillance and because evidence shows a role for either STAT3 or STAT1 in TH9 differentiation conflicts, we sought to determine the status on this lineage of STAT1 GOF and STAT3 LOF mutations in human subjects. We detected IL-9 levels and TH9 differentiation in patients with STAT3 LOF and STAT1 GOF mutations, together with TH9 transcript factors, and partially rescued their deficiency invitro by adding cytokines they lacked or transfecting key molecules. We found that PBMCs or sorted naive CD4+ T cells from patients with STAT3 LOF and STAT1 GOF mutations had impaired TH9 generation/differentiation. STAT3 inhibition in normal TH9 cultures diminished early IL-21 induction and late IL-9 production, whereas exogenous IL-21 enhanced TH9 differentiation, even with STAT3 inhibition, by restoring suppressor of cytokine signaling 3 expression and thus inhibiting excessive phosphorylated signal transducer and activator of transcription (p-STAT) 1 activation. Furthermore, exogenous expression of suppressor of cytokine signaling 3 or either T-bet or STAT1 RNA interference in STAT3 LOF cells partially rescued IL-9 differentiation. Collectively, these results suggest that human TH9 differentiation depends on normal p-STAT3 and IL-21 production to suppress p-STAT1 activation and T-bet transcription.

Signal transducer and activator of transcription gain-of-function primary immunodeficiency/immunodysregulation disorders.

To describe primary immunodeficiencies caused by gain-of-function (GOF) mutations of signal transducer and activator of transcription (STAT) genes, a group of genetically determined disorders characterized by susceptibility to infections and, in many cases, autoimmune manifestations. GOF mutations affecting STAT1 result in increased STAT tyrosine phosphorylation and secondarily increased response to STAT1-signaling cytokines, such as interferons. In contrast, STAT3 hyperactivity is not usually related to hyperphosphorylation but rather to increased STAT3-mediated transcriptional activity. In both cases, heterozygous STAT1 and STAT3 GOF mutations trigger a distinct set of genes in target cells that lead to abnormal functioning of antimicrobial response and/or autoimmunity and result in autosomal dominant diseases. Clinical manifestations of patients with STAT1 GOF are characterized by mucocutaneous candidiasis and recurrent lower tract respiratory infections. In addition, many patients have thyroiditis, type 1 diabetes mellitus, autoimmune cytopenias, cancer or aneurysms. Patients with germline STAT3 GOF mutations have an increased frequency of early-onset multiorgan autoimmunity (i.e. autoimmune enteropathy, type 1 diabetes mellitus, autoimmune interstitial lung disease and autoimmune cytopenias), lymphoproliferation, short stature and, less frequently, severe recurrent infections. Treatment options range from antimicrobial therapy, intravenous or subcutaneous immunoglobulin and immunosuppressive drugs. Some patients with STAT1 GOF disorder have undergone hematopoietic stem cell transplantation, although these have been difficult because of the underlying proinflammatory milieu from the mutation.

Autoimmune Regulator Deficiency Results in a Decrease in STAT1 Levels in Human Monocytes.

Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a rare primary immunodeficiency disorder typically caused by biallelic autoimmune regulator (AIRE) mutations that manifests with chronic mucocutaneous candidiasis (CMC) and autoimmunity. Patients with STAT1 gain-of-function (GOF) mutations also develop CMC and autoimmunity; they exhibit increased STAT1 protein levels at baseline and STAT1 phosphorylation (pSTAT1) upon interferon (IFN)-γ stimulation relative to healthy controls. AIRE interacts functionally with a protein that directly regulates STAT1, namely protein inhibitor of activated STAT1, which inhibits STAT1 activation. Given the common clinical features between patients with AIRE and STAT1 GOF mutations, we sought to determine whether APECED patients also exhibit increased levels of STAT1 protein and phosphorylation in CD14+ monocytes. We obtained peripheral blood mononuclear cells from 8 APECED patients and 13 healthy controls and assessed the levels of STAT1 protein and STAT1 tyrosine phosphorylation at rest and following IFN-γ stimulation, as well as the levels of STAT1 mRNA. The mean STAT1 protein levels in CD14+ monocytes exhibited a ~20% significant decrease in APECED patients both at rest and after IFN-γ stimulation relative to that of healthy donors. Similarly, the mean peak value of IFN-γ-induced pSTAT1 level was ~20% significantly lower in APECED patients compared to that in healthy controls. The decrease in STAT1 and peak pSTAT1 in APECED patients was not accompanied by decreased STAT1 mRNA or anti-IFN-γ autoantibodies; instead, it correlated with the presence of autoantibodies to type I IFN and decreased AIRE−/− monocyte surface expression of IFN-γ receptor 2. Our data show that, in contrast to patients with STAT1 GOF mutations, APECED patients show a moderate but consistent and significant decrease in total STAT1 protein levels, associated with lower peak levels of pSTAT1 molecules after IFN-γ stimulation.

STAT mutations as program switchers: turning primary immunodeficiencies into autoimmune diseases.

STAT proteins are a family of transcription factors that mediate cellular response to cytokines and growth factors. Study of patients with familial susceptibility to pathogens and/or autoimmune diseases has led to the identification of 7 inherited disorders that are caused by mutations of 4 STAT family genes. Homozygous or compound heterozygous mutations of STAT1 lead to complete or partial forms of STAT1 deficiency that are associated with susceptibility to intracellular pathogens and herpetic infections. Patients with heterozygous STAT1 gain-of-function (GOF) mutations usually present with chronic mucocutaneous candidiasis (CMC) but may also experience bacterial and viral infections, autoimmune manifestations, lymphopenia, cerebral aneurysms, and increased risk to develop tumors. STAT2 deficiency has been described in 5 family members and is characterized by selective susceptibility to viral infections, whereas STAT3 loss-of-function (LOF) mutations are causative of the autosomal-dominant hyper-IgE syndrome (HIES), a condition that is characterized by cutaneous and respiratory infections in association with mucocutaneous candidiasis, eczema, skeletal and connective tissue abnormalities, eosinophilia, and high levels IgE. STAT5B LOF and STAT3 GOF mutations are both associated with disorders characterized by autoimmune or allergic manifestations, together with increased risk of infections. Particularly, STAT5b deficiency results in growth hormone (GH) insensitivity, immunodeficiency, diarrhea, and generalized eczema, whereas STAT3 GOF mutations result in autoimmune cytopenia, lymphadenopathy, short stature, infections, enteropathy, and multiorgan autoimmunity, including early-onset type I diabetes, thyroiditis, hepatitis, arthritis, and interstitial lung disease.

Heterozygous STAT1 gain-of-function mutations underlie an unexpectedly broad clinical phenotype

AbstractSince their discovery in patients with autosomal dominant (AD) chronic mucocutaneous candidiasis (CMC) in 2011, heterozygous STAT1 gain-of-function (GOF) mutations have increasingly been identified worldwide. The clinical spectrum associated with them needed to be delineated. We enrolled 274 patients from 167 kindreds originating from 40 countries from 5 continents. Demographic data, clinical features, immunological parameters, treatment, and outcome were recorded. The median age of the 274 patients was 22 years (range, 1-71 years); 98% of them had CMC, with a median age at onset of 1 year (range, 0-24 years). Patients often displayed bacterial (74%) infections, mostly because of Staphylococcus aureus (36%), including the respiratory tract and the skin in 47% and 28% of patients, respectively, and viral (38%) infections, mostly because of Herpesviridae (83%) and affecting the skin in 32% of patients. Invasive fungal infections (10%), mostly caused by Candida spp. (29%), and mycobacterial disease (6%) caused by Mycobacterium tuberculosis, environmental mycobacteria, or Bacille Calmette-Guérin vaccines were less common. Many patients had autoimmune manifestations (37%), including hypothyroidism (22%), type 1 diabetes (4%), blood cytopenia (4%), and systemic lupus erythematosus (2%). Invasive infections (25%), cerebral aneurysms (6%), and cancers (6%) were the strongest predictors of poor outcome. CMC persisted in 39% of the 202 patients receiving prolonged antifungal treatment. Circulating interleukin-17A–producing T-cell count was low for most (82%) but not all of the patients tested. STAT1 GOF mutations underlie AD CMC, as well as an unexpectedly wide range of other clinical features, including not only a variety of infectious and autoimmune diseases, but also cerebral aneurysms and carcinomas that confer a poor prognosis.

Treatment options for chronic mucocutaneous candidiasis

Autosomal dominant chronic mucocutaneous candidiasis (AD-CMC) is a rare and severe primary immunodeficiency that is characterized by mucocutaneous fungal infection, autoimmunity, cerebral aneurysms, and oropharyngeal and esophageal cancer. Recently, it was discovered that STAT1 mutations are responsible for AD-CMC. These mutations lead to the inability of STAT1 to be dephosphorylated, resulting in hyperphosphorylation, increased binding to the DNA, and gain of function (GOF) effects on STAT1 signaling. Furthermore, a characteristic feature of AD-CMC patients is deficiency in the T-helper 17 (Th17) responses, which is believed to be the immunological cause of the mucocutaneous fungal infection. No targeted treatment other than lifelong antifungal prophylaxis exists for AD-CMC. However, the discovery of the genetic and immunological defects makes it now possible to explore new treatment strategies. This review will discuss immunomodulatory treatment options that can be explored in patients with STAT1 GOF mutations.