Abnormal biomarkers predict complex FAS or FADD defects missed by exome sequencing.

BACKGROUND: Elevated TCRαß+CD4-CD8- double-negative T cells (DNT) and serum biomarkers help identify FAS mutant patients with autoimmune lymphoproliferative syndrome (ALPS). However, in some patients with clinical features and biomarkers consistent with ALPS, germline or somatic FAS mutations cannot be identified on standard exon sequencing (ALPS-undetermined: ALPS-U). OBJECTIVE: We sought to explore whether complex genetic alterations in the FAS gene escaping standard sequencing or mutations in other FAS pathway-related genes could explain these cases. METHODS: Genetic analysis included whole FAS gene sequencing, copy number variation analysis, and sequencing of FAS cDNA and other FAS pathway-related genes. It was guided by FAS expression analysis on CD57+DNT, which can predict somatic loss of heterozygosity (sLOH). RESULTS: Nine of 16 patients with ALPS-U lacked FAS expression on CD57+DNT predicting heterozygous "loss-of-expression" FAS mutations plus acquired somatic second hits in the FAS gene, enriched in DNT. Indeed, 7 of 9 analyzed patients carried deep intronic mutations or large deletions in the FAS gene combined with sLOH detectable in DNT; 1 patient showed a FAS exon duplication. Three patients had reduced FAS expression, and 2 of them harbored mutations in the FAS promoter, which reduced FAS expression in reporter assays. Three of the 4 ALPS-U patients with normal FAS expression carried heterozygous FADD mutations with sLOH. CONCLUSION: A combination of serum biomarkers and DNT phenotyping is an accurate means to identify patients with ALPS who are missed by routine exome sequencing.

JAK inhibitor treatment for inborn errors of JAK/STAT signaling: An ESID/EBMT-IEWP retrospective study.

BACKGROUND: Inborn errors of immunity (IEI) with dysregulated JAK/STAT signaling present with variable manifestations of immune dysregulation and infections. Hematopoietic stem cell transplantation (HSCT) is potentially curative, but initially reported outcomes were poor. JAK inhibitors (JAKi) offer a targeted treatment option that may be an alternative or bridge to HSCT. However, data on their current use, treatment efficacy and adverse events are limited. OBJECTIVE: We evaluated the current off-label JAKi treatment experience for JAK/STAT inborn errors of immunity (IEI) among European Society for Immunodeficiencies (ESID)/European Society for Blood and Marrow Transplantation (EBMT) Inborn Errors Working Party (IEWP) centers. METHODS: We conducted a multicenter retrospective study on patients with a genetic disorder of hyperactive JAK/STAT signaling who received JAKi treatment for at least 3 months. RESULTS: Sixty-nine patients (72% children) were evaluated (45 STAT1 gain of function [GOF], 21 STAT3-GOF, 1 STAT5B-GOF, 1 suppressor of cytokine signaling 1 [aka SOCS1] loss of function, 1 JAK1-GOF). Ruxolitinib was the predominantly prescribed JAKi (80%). Overall, treatment resulted in improvement (partial or complete remission) of clinical symptoms in 87% of STAT1-GOF and in 90% of STAT3-GOF patients. We documented highly heterogeneous dosing and monitoring regimens. The response rate and time to response varied across different diseases and manifestations. Adverse events including infection and weight gain were frequent (38% of patients) but were mild (grade I-II) and transient in most patients. At last follow-up, 52 (74%) of 69 patients were still receiving JAKi treatment, and 11 patients eventually underwent HSCT after receipt of previous JAKi bridging therapy, with 91% overall survival. CONCLUSIONS: Our study suggests that JAKi may be highly effective to treat symptomatic JAK/STAT IEI patients. Prospective studies to define optimal JAKi dosing for the variable clinical presentations and age ranges should be pursued.

Interferon signature in patients with STAT1 gain-of-function mutation is epigenetically determined.

STAT1 gain-of-function (GOF) variants lead to defective Th17 cell development and chronic mucocutaneous candidiasis (CMC), but frequently also to autoimmunity. Stimulation of cells with STAT1 inducing cytokines like interferons (IFN) result in hyperphosphorylation and delayed dephosphorylation of GOF STAT1. However, the mechanism how the delayed dephosphorylation exactly causes the increased expression of STAT1-dependent genes, and how the intracellular signal transduction from cytokine receptors is affected, remains unknown. In this study we show that the circulating levels of IFN-α were not persistently elevated in STAT1 GOF patients. Nevertheless, the expression of interferon signature genes was evident even in the patient with low or undetectable serum IFN-α levels. Chromatin immunoprecipitation (ChIP) experiments revealed that the active chromatin mark trimethylation of lysine 4 of histone 3 (H3K4me3), was significantly enriched in areas associated with interferon-stimulated genes in STAT1 GOF cells in comparison to cells from healthy donors. This suggests that the chromatin binding of GOF STAT1 variant promotes epigenetic changes compatible with higher gene expression and elevated reactivity to type I interferons, and possibly predisposes for interferon-related autoimmunity. The results also suggest that epigenetic rewiring may be responsible for treatment failure of Janus kinase 1/2 (JAK1/2) inhibitors in certain patients.

Multiorgan involvement and management in children with Down syndrome.

AIM: To review multiorgan involvement and management in children with Down syndrome (DS). METHODS: A literature review of articles from 1980 to 2019 using the MEDLINE interface of PubMed was performed using the following search terms- [Down syndrome] or [Trisomy 21] AND [Cardiology] or [Respiratory] or [neurodevelopment] or [epilepsy] or [musculoskeletal] or [immune system] or [haematological] or [endocrine] or [gastrointestinal] or [ophthalmological] or [Ear Nose Throat] or [dermatology] or [renal]. RESULTS: Congenital heart disease particularly septal defects occur in over 60% of infants with DS and 5%-34% of infants develop persistent pulmonary hypertension of the newborn irrespective of a diagnosis of congenital heart disease. Early recognition and management of aspiration, obstructive sleep apnoea and recurrent lower respiratory tract infections (LRTI) could reduce risk of developing pulmonary hypertension in later childhood. Children with DS have an increased risk of autistic spectrum disorder, attention deficit disorder and epilepsy particularly infantile spasms, which are associated with poor neurodevelopmental outcomes. Congenital anomalies of the gastrointestinal and renal system as well as autoimmune diseases, coeliac disease, arthropathy, thyroid dysfunction fold diabetes mellitus and dermatological conditions are more common. Hearing and visual anomalies are also well recognised association with DS (Table 1). CONCLUSION: Children with DS are at an increased risk of multiorgan comorbidities. Organ-specific health surveillance may provide holistic care for the children and families with DS throughout childhood.

Topical cidofovir for the treatment of recalcitrant viral warts and molluscum contagiosum in Jacobsen syndrome.

Jacobsen syndrome is caused by a terminal deletion on the long arm of chromosome 11 and can be associated with immunodeficiency. Patients with Jacobsen syndrome can be predisposed to cutaneous viral infections that are difficult to treat. We report successful use of topical 1% cidofovir as treatment of recalcitrant verruca vulgaris in one patient and molluscum contagiosum in another patient with Jacobsen syndrome. Topical cidofovir appears to be a good treatment option in this cohort and should be considered early for treatment-resistant cutaneous viral infections.

Altered endotoxin responsiveness in healthy children with Down syndrome.

BACKGROUND: Down syndrome (DS) is the most common syndromic immunodeficiency with an increased risk of infection, mortality from sepsis, and autoinflammation. Innate immune function is altered in DS and therefore we examined responses in CD11b and Toll like receptor 4 (TLR-4), which are important immune cell surface markers upregulated in response to Lipopolysaccharide (LPS) endotoxin, and the immunomodulator melatonin. Neutrophil and monocyte responses to LPS and melatonin in children with Down syndrome (DS) who were clinically stable were compared to age-matched controls. Whole blood was incubated with LPS and melatonin and the relative expression of CD11b and TLR-4 evaluated by flow cytometry. RESULTS: Children with DS had an increased response to LPS in neutrophils and intermediate monocytes, while also having elevated TLR-4 expression on non-classical monocytes compared to controls at baseline. Melatonin reduced CD11b expression on neutrophils, total monocytes, both classical and intermediate sub-types, in children with DS and controls. CONCLUSION: Melatonin could represent a useful clinical adjunct in the treatment of sepsis as an immunomodulator. Children with DS had increased LPS responses which may contribute to the more adverse outcomes seen in sepsis.

CD57 identifies T cells with functional senescence before terminal differentiation and relative telomere shortening in patients with activated PI3 kinase delta syndrome.

Premature T-cell immunosenescence with CD57+ CD8+ T-cell accumulation has been linked to immunodeficiency and autoimmunity in primary immunodeficiencies including activated PI3 kinase delta syndrome (APDS). To address whether CD57 marks the typical senescent T-cell population seen in adult individuals or identifies a distinct population in APDS, we compared CD57+ CD8+ T cells from mostly pediatric APDS patients to those of healthy adults with similarly prominent senescent T cells. CD57+ CD8+ T cells from APDS patients were less differentiated with more CD27+ CD28+ effector memory T cells showing increased PD1 and Eomesodermin expression. In addition, transition of naïve to CD57+ CD8+ T cells was not associated with the characteristic telomere shortening. Nevertheless, they showed the increased interferon-gamma secretion, enhanced degranulation and reduced in vitro proliferation typical of senescent CD57+ CD8+ T cells. Thus, hyperactive PI3 kinase signaling favors premature accumulation of a CD57+ CD8+ T-cell population, which shows most functional features of typical senescent T cells, but is different in terms of differentiation and relative telomere shortening. Initial observations indicate that this specific differentiation state may offer the opportunity to revert premature T-cell immunosenescence and its potential contribution to inflammation and immunodeficiency in APDS.

Clinical spectrum and features of activated phosphoinositide 3-kinase δ syndrome: A large patient cohort study.

BACKGROUND: Activated phosphoinositide 3-kinase δ syndrome (APDS) is a recently described combined immunodeficiency resulting from gain-of-function mutations in PIK3CD, the gene encoding the catalytic subunit of phosphoinositide 3-kinase δ (PI3Kδ). OBJECTIVE: We sought to review the clinical, immunologic, histopathologic, and radiologic features of APDS in a large genetically defined international cohort. METHODS: We applied a clinical questionnaire and performed review of medical notes, radiology, histopathology, and laboratory investigations of 53 patients with APDS. RESULTS: Recurrent sinopulmonary infections (98%) and nonneoplastic lymphoproliferation (75%) were common, often from childhood. Other significant complications included herpesvirus infections (49%), autoinflammatory disease (34%), and lymphoma (13%). Unexpectedly, neurodevelopmental delay occurred in 19% of the cohort, suggesting a role for PI3Kδ in the central nervous system; consistent with this, PI3Kδ is broadly expressed in the developing murine central nervous system. Thoracic imaging revealed high rates of mosaic attenuation (90%) and bronchiectasis (60%). Increased IgM levels (78%), IgG deficiency (43%), and CD4 lymphopenia (84%) were significant immunologic features. No immunologic marker reliably predicted clinical severity, which ranged from asymptomatic to death in early childhood. The majority of patients received immunoglobulin replacement and antibiotic prophylaxis, and 5 patients underwent hematopoietic stem cell transplantation. Five patients died from complications of APDS. CONCLUSION: APDS is a combined immunodeficiency with multiple clinical manifestations, many with incomplete penetrance and others with variable expressivity. The severity of complications in some patients supports consideration of hematopoietic stem cell transplantation for severe childhood disease. Clinical trials of selective PI3Kδ inhibitors offer new prospects for APDS treatment.

Acute Genital Swelling Heralding C1 Esterase Inhibitor Deficiency in a Child.

A healthy 5-year-old boy presented to the emergency department with an acute genital swelling. He had no relevant family history. His presentation and blood investigations were consistent with C1 esterase inhibitor deficiency, mostly likely arising de novo. A rare cause of acute genital swelling and its management are discussed.

Changes in airway inflammation during pulmonary exacerbations in patients with cystic fibrosis and primary ciliary dyskinesia.

Lung disease in patients with both primary ciliary dyskinesia (PCD) or cystic fibrosis (CF) is associated with impaired mucociliary clearance; however, clinical outcomes are typically worse in CF patients. We assessed whether CF and PCD patients differ in inflammatory response in the airways during pulmonary exacerbation.We first studied clinically stable PCD patients with a spectrum of bacterial pathogens to assess inflammatory response to different pathogens. Subsequently, PCD and CF patients with similar bacterial pathogens were studied at the time of a pulmonary exacerbation and after 21 days of antibiotics treatment. Qualitative and quantitative microbiology, cell counts, interleukin-8 concentrations, and neutrophil elastase activity were assessed in sputum samples obtained before and after treatment.In stable PCD patients, no significant differences were found in sputum inflammatory markers between individuals colonised with different bacterial pathogens. Pulmonary exacerbation severity assessed by a pulmonary exacerbation score and lung function decline from their previous baseline did not differ between CF and PCD patients. Bacterial density for Staphylococcus aureus and Haemophilus influenzae was higher in CF versus PCD (p<0.05), but absolute neutrophil counts were higher in PCD patients (p=0.02). While sputum elastase activity was similar in PCD and CF at the time of exacerbation, it decreased with antibiotic therapy in PCD (p<0.05) but not CF patients.PCD patients differ from those with CF in their responses to treatment of pulmonary exacerbations, with higher neutrophil elastase activity persisting in the CF airways at the end of treatment.

Functional characterization of the human dendritic cell immunodeficiency associated with the IRF8(K108E) mutation.

We have previously reported on a unique patient in whom homozygosity for a mutation at IRF8 (IRF8(K108E)) causes a severe immunodeficiency. Laboratory evaluation revealed a highly unusual myeloid compartment, remarkable for the complete absence of CD141 and CD161 monocytes, absence of CD11c1 conventional dendritic cells (DCs) and CD11c1/CD1231 plasmacytoid DCs, and striking granulocytic hyperplasia. The patient initially presented with severe disseminated mycobacterial and mucocutaneous fungal infections and was ultimately cured by cord blood transplant. Sequencing RNA from the IRF8(K108E) patient's primary blood cells prior to transplant shows not only depletion of IRF8-bound and IRF8-regulated transcriptional targets, in keeping with the distorted composition of the myeloid compartment, but also a paucity of transcripts associated with activated CD41 and CD81 T lymphocytes. This suggests that T cells reared in the absence of a functional antigen-presenting compartment in IRF8(K108E) are anergic. Biochemical characterization of the IRF8(K108E) mutant in vitro shows that loss of the positively charged side chain at K108 causes loss of nuclear localization and loss of transcriptional activity, which is concomitant with decreased protein stability, increased ubiquitination, increased small ubiquitin-like modification, and enhanced proteasomal degradation. These findings provide functional insight into the molecular basis of immunodeficiency associated with loss of IRF8.

Variant ataxia telangiectasia identified during evaluation for short stature.

Ataxia telangiectasia (A-T) (OMIM 208900) is an autosomal recessive multisystem disorder characterised by progressive cerebellar ataxia, telangiectasias, immunodeficiency and a predisposition to malignancy. 'Variant' A-T has later onset of neurological symptoms and slower progression compared with the 'classic' form. A woman presented with short stature in late childhood. Karyotype revealed rearrangements involving chromosomes 7 and 14. A chromosomal breakage disorder gene panel demonstrated compound heterozygote mutations in her ATM gene including one mutation c.7271T>G with residual ATM function, confirming the diagnosis of variant A-T. Since diagnosis, she has developed progressive cerebellar ataxia and telangiectasias. Long-standing restrictive and aversive feeding behaviours presented challenges for her management and necessitated gastrostomy.

Experience of pediatric to adult transition in immunology services: patient experience questionnaire and micro-costing analysis.

The effective transition from pediatric to adult care for individuals with chronic medical conditions should address the medical, psychosocial and educational needs of the cohort. The views and experiences of service users and their families are an integral component of service development. This study sought to evaluate the current provision of transition services from pediatric immunology services to adult immunology services for patients with a diagnosis of an inborn error of immunity at St. James's Hospital, Dublin. We gathered patient perspectives on the experience of the transition process using a structured survey. In addition, we adopted a micro-costing technique to estimate the cost of implementing the current standard of care for these patients. Results of a micro-costing analysis suggest that the most significant component of cost in assessing these patients is on laboratory investigation, an area where there is likely significant duplication between pediatric and adult care. Perspectives from patients suggested that the transition period went well for the majority of the cohort and that they felt ready to move to adult services, but the transition was not without complications in areas such as self-advocacy and medication management. The transition process may benefit from enhanced communication and collaboration between pediatric and adult services.

Diagnostic evaluation of paediatric autoimmune lymphoproliferative immunodeficiencies (ALPID): a prospective cohort study.

BACKGROUND: Lymphoproliferation and autoimmune cytopenias characterise autoimmune lymphoproliferative syndrome. Other conditions sharing these manifestations have been termed autoimmune lymphoproliferative syndrome-like diseases, although they are frequently more severe. The aim of this study was to define the genetic, clinical, and immunological features of these disorders to improve their diagnostic classification. METHODS: In this prospective cohort study, patients were referred to the Center for Chronic Immunodeficiency in Freiburg, Germany, between Jan 1, 2008 and March 5, 2022. We enrolled patients younger than 18 years with lymphoproliferation and autoimmune cytopenia, lymphoproliferation and at least one additional sign of an inborn error of immunity (SoIEI), bilineage autoimmune cytopenia, or autoimmune cytopenia and at least one additional SoIEI. Autoimmune lymphoproliferative syndrome biomarkers were determined in all patients. Sanger sequencing followed by in-depth genetic studies were recommended for patients with biomarkers indicative of autoimmune lymphoproliferative syndrome, while IEI panels, exome sequencing, or genome sequencing were recommended for patients without such biomarkers. Genetic analyses were done as decided by the treating physician. The study was registered on the German Clinical Trials Register, DRKS00011383, and is ongoing. FINDINGS: We recruited 431 children referred for autoimmune lymphoproliferative syndrome evaluation, of whom 236 (55%) were included on the basis of lymphoproliferation and autoimmune cytopenia, 148 (34%) on the basis of lymphoproliferation and another SoIEI, 33 (8%) on the basis of autoimmune bicytopenia, and 14 (3%) on the basis of autoimmune cytopenia and another SoIEI. Median age at diagnostic evaluation was 9·8 years (IQR 5·5-13·8), and the cohort comprised 279 (65%) boys and 152 (35%) girls. After biomarker and genetic assessments, autoimmune lymphoproliferative syndrome was diagnosed in 71 (16%) patients. Among the remaining 360 patients, 54 (15%) had mostly autosomal-dominant autoimmune lymphoproliferative immunodeficiencies (AD-ALPID), most commonly affecting JAK-STAT (26 patients), CTLA4-LRBA (14), PI3K (six), RAS (five), or NFκB (three) signalling. 19 (5%) patients had other IEIs, 17 (5%) had non-IEI diagnoses, 79 (22%) were unresolved despite extended genetics (ALPID-U), and 191 (53%) had insufficient genetic workup for diagnosis. 16 (10%) of 161 patients with a final diagnosis had somatic mutations. Alternative classification of patients fulfilling common variable immunodeficiency or Evans syndrome criteria did not increase the proportion of genetic diagnoses. INTERPRETATION: The ALPID phenotype defined in this study is enriched for patients with genetic diseases treatable with targeted therapies. The term ALPID might be useful to focus diagnostic and therapeutic efforts by triggering extended genetic analysis and consideration of targeted therapies, including in some children currently classified as having common variable immunodeficiency or Evans syndrome. FUNDING: Deutsche Forschungsgemeinschaft under Germany's Excellence Strategy. TRANSLATION: For the German translation of the abstract see Supplementary Materials section.

Innate immune dysregulation in multisystem inflammatory syndrome in children (MIS-C).

MIS-C is a systemic inflammation disorder with poorly characterised immunopathological mechanisms. We compared changes in the systemic immune response in children with MIS-C (n = 12, 5-13 years) to healthy controls (n = 14, 5-15 years). Analysis was done in whole blood treated with LPS. Expression of CD11b and Toll-like receptor-4 (TLR4) in neutrophils and monocytes were analysed by flow cytometry. Serum cytokines (IL-1ß, IL-2, IL-6, IL-8, IL-10, IL-Ira, TNF-α, TNF-ß, IFN-Υ, VEGF, EPO and GM-CSF) and mRNA levels of inflammasome molecules (NLRP3, ASC and IL-1ß) were evaluated. Subpopulations of lymphocytes (CD3+, CD19+, CD56+, CD4+, CD8+, TCR Vδ1+, TCR Vδ2+) were assessed at basal levels. Absolute counts of neutrophils and NLR were high in children with MIS-C while absolute counts of lymphocytes were low. Children with MIS-C had increased levels of IL-6, IL-10, TNF-ß and VEGF serum cytokines at the basal level, and significantly increased TNF-ß post-LPS, compared to controls. IL-1RA and EPO decreased at baseline and post-LPS in MIS-C patients compared to controls. The percentage of CD3+ cells, NK cells and Vδ1 was lower while B cells were higher in children with MIS-C than in controls. Dysregulated immune response in children with MIS-C was evident and may be amenable to immunomodulation.

Incomplete Kawasaki disease associated with complicated Streptococcus pyogenes pneumonia: A case report.

A three-year-old boy presented with community-acquired pneumonia complicated by empyema. Streptococcus pyogenes (group A streptococcus) was identified on culture of the pleural fluid. The patient improved with antibiotic therapy and drainage of the empyema. During his convalescence, the patient developed persistent fever, lethargy and anorexia. His inflammatory markers were elevated, and repeat cultures were negative. Although the patient had none of the classical mucocutaneous features of Kawasaki disease, an echocardiogram was performed, which revealed coronary artery dilation. The patient was diagnosed with incomplete Kawasaki disease and treated with intravenous immunoglobulin and high-dose acetylsalicylic acid. The fever subsided within 48 h. To the authors' knowledge, the present report is the first report of Kawasaki disease associated with complicated S pyogenes pneumonia. It emphasizes the importance of considering incomplete Kawasaki disease among children with persistent fever, the role of echocardiography in diagnosis, and the potential link between Kawasaki disease and superantigen-producing organisms such as S pyogenes.

Un garçon de trois ans a consulté à cause d'une pneumonie non nosocomiale compliquée par un empyème. On a décelé un Streptococcus pyogenes (streptocoque du groupe A) à la culture du liquide pleural. L'état du patient s'est amélioré au moyen d'une antibiothérapie et d'un drainage de l'empyème.Pendant sa convalescence, le patient a présenté une fièvre persistante, une léthargie et une anorexie. Ses marqueurs inflammatoires étaient élevés, et les nouvelles cultures étaient négatives. Même s'il ne présentait aucune des caractéristiques mucocutanées classiques de la maladie de Kawasaki, on a effectué un échocardiogramme, qui a révélé une dilatation des artères coronaires.Le patient a reçu un diagnostic de maladie de Kawasaki incomplète et a été traité au moyen d'immunoglobuline intraveineuse et d'acide acétylsalicylique à forte dose. La fièvre a disparu en 48 heures. En autant que le sache les auteurs, c'est le premier rapport de cas de maladie Kawasaki incomplète compliquée par une pneumonie à S pyogenes, ce qui fait ressortir l'importance d'envisager une maladie de Kawasaki chez les enfants atteints de fièvre persistante, le rôle de l'échocardiographie dans le diagnostic et le lien potentiel entre la maladie de Kawasaki et les organismes producteurs de superantigène comme le S pyogenes.

Roifman syndrome: a description of further immunological and radiological features.

Roifman syndrome is a rare autosomal recessive inherited syndromic immunodeficiency. We wish to add to the available literature by reporting two brothers with clinical, radiological and immunological features of Roifman syndrome, confirmed on whole exome sequencing. We report an excellent response to subcutaneous immunoglobulin therapy in both brothers, reducing infection burden and hospital admissions. New radiological features are also described here which may assist in the diagnosis of other patients.