The risk of COVID-19 death is much greater and age dependent with type I IFN autoantibodies.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection fatality rate (IFR) doubles with every 5 y of age from childhood onward. Circulating autoantibodies neutralizing IFN-α, IFN-ω, and/or IFN-ß are found in ∼20% of deceased patients across age groups, and in ∼1% of individuals aged <70 y and in >4% of those >70 y old in the general population. With a sample of 1,261 unvaccinated deceased patients and 34,159 individuals of the general population sampled before the pandemic, we estimated both IFR and relative risk of death (RRD) across age groups for individuals carrying autoantibodies neutralizing type I IFNs, relative to noncarriers. The RRD associated with any combination of autoantibodies was higher in subjects under 70 y old. For autoantibodies neutralizing IFN-α2 or IFN-ω, the RRDs were 17.0 (95% CI: 11.7 to 24.7) and 5.8 (4.5 to 7.4) for individuals <70 y and ≥70 y old, respectively, whereas, for autoantibodies neutralizing both molecules, the RRDs were 188.3 (44.8 to 774.4) and 7.2 (5.0 to 10.3), respectively. In contrast, IFRs increased with age, ranging from 0.17% (0.12 to 0.31) for individuals <40 y old to 26.7% (20.3 to 35.2) for those ≥80 y old for autoantibodies neutralizing IFN-α2 or IFN-ω, and from 0.84% (0.31 to 8.28) to 40.5% (27.82 to 61.20) for autoantibodies neutralizing both. Autoantibodies against type I IFNs increase IFRs, and are associated with high RRDs, especially when neutralizing both IFN-α2 and IFN-ω. Remarkably, IFRs increase with age, whereas RRDs decrease with age. Autoimmunity to type I IFNs is a strong and common predictor of COVID-19 death.

Proceedings from the inaugural Artificial Intelligence in Primary Immune Deficiencies (AIPID) conference.

Here, we summarize the proceedings of the inaugural Artificial Intelligence in Primary Immune Deficiencies conference, during which experts and advocates gathered to advance research into the applications of artificial intelligence (AI), machine learning, and other computational tools in the diagnosis and management of inborn errors of immunity (IEIs). The conference focused on the key themes of expediting IEI diagnoses, challenges in data collection, roles of natural language processing and large language models in interpreting electronic health records, and ethical considerations in implementation. Innovative AI-based tools trained on electronic health records and claims databases have discovered new patterns of warning signs for IEIs, facilitating faster diagnoses and enhancing patient outcomes. Challenges in training AIs persist on account of data limitations, especially in cases of rare diseases, overlapping phenotypes, and biases inherent in current data sets. Furthermore, experts highlighted the significance of ethical considerations, data protection, and the necessity for open science principles. The conference delved into regulatory frameworks, equity in access, and the imperative for collaborative efforts to overcome these obstacles and harness the transformative potential of AI. Concerted efforts to successfully integrate AI into daily clinical immunology practice are still needed.

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.

Molecular Challenges in the Diagnosis of X-Linked Chronic Granulomatous Disease: CNVs, Intronic Variants, Skewed X-Chromosome Inactivation, and Gonosomal Mosaicism.

Chronic granulomatous disease (CGD) is a prototypical inborn error of immunity affecting phagocytes, in which these cells are unable to produce reactive oxygen species. CGD is caused by defects in genes encoding subunits of the NADPH oxidase enzyme complex (CYBA, CYBB, CYBC1, NCF1, NCF2, NCF4); inflammatory responses are dysregulated, and patients are highly susceptible to recurrent severe bacterial and fungal infections. X-linked CGD (XL-CGD), caused by mutations in the CYBB gene, is the most common and severe form of CGD. In this study, we describe the analytical processes undertaken in 3 families affected with XL-CGD to illustrate several molecular challenges in the genetic diagnosis of this condition: in family 1, a girl with a heterozygous deletion of CYBB exon 13 and skewed X-chromosome inactivation (XCI); in family 2, a boy with a hemizygous deletion of CYBB exon 7, defining its consequences at the mRNA level; and in family 3, 2 boys with the same novel intronic variant in CYBB (c.1151 + 6 T > A). The variant affected the splicing process, although a small fraction of wild-type mRNA was produced. Their mother was a heterozygous carrier, while their maternal grandmother was a carrier in form of gonosomal mosaicism. In summary, using a variety of techniques, including an NGS-based targeted gene panel and deep amplicon sequencing, copy number variation calling strategies, microarray-based comparative genomic hybridization, and cDNA analysis to define splicing defects and skewed XCI, we show how to face and solve some uncommon genetic mechanisms in the diagnosis of XL-CGD.

Treatments for multi-system inflammatory syndrome in children - discharge, fever, and second-line therapies.

Scarce evidence exists about the best treatment for multi-system inflammatory syndrome (MIS-C). We analyzed the effects of steroids, intravenous immunoglobulin (IVIG), and their combination on the probability of discharge over time, the probability of switching to second-line treatment over time, and the persistence of fever 2 days after treatment. We did a retrospective study to investigate the effect of different treatments on children with MIS-C from 1 March 2020 to 1 June 2021. We estimated the time-to-event probability using a Cox model weighted by propensity score to balance the baseline characteristics. Thirty of 132 (22.7%) patients were initially treated with steroids alone, 29/132 (21.9%) with IVIG alone, and 73/132 (55%) with IVIG plus steroids. The probability of early discharge was higher with IVIG than with IVIG plus steroids (hazard ratio [HR] 1.65, 95% CI 1.11-2.45, p = 0.013), but with a higher probability of needing second-line therapy compared to IVIG plus steroids (HR 3.05, 95% CI 1.12-8.25, p = 0.028). Patients on IVIG had a higher likelihood of persistent fever than patients on steroids (odds ratio [OR] 4.23, 95% CI 1.43-13.5, p = 0.011) or on IVIG plus steroids (OR 4.4, 95% CI 2.05-9.82, p < 0.001). No differences were found for this endpoint between steroids or steroids plus IVIG.    Conclusions: The benefits of each approach may vary depending on the outcome assessed. IVIG seemed to increase the probability of earlier discharge over time but also of needing second-line treatment over time. Steroids seemed to reduce persistent fever, and combination therapy reduced the need for escalating treatment. What is Known: • Steroids plus intravenous immunoglobulin, compared with intravenous immunoglobulin alone for multi-system inflammatory syndrome (MIS-C) might reduce the need for hemodynamic support and the duration of fever, but the certainty of the evidence is low. What is New: • Intravenous immunoglobulin, steroids, and their combination for MIS-C may have different outcomes. • In this study, intravenous immunoglobulin increased the probability of discharge over time, steroids reduced persistent fever, while combination therapy reduced the need for second-line treatments.

Multisystem inflammatory syndrome in children and SARS-CoV-2 variants: a two-year ambispective multicentric cohort study in Catalonia, Spain.

Multisystem inflammatory syndrome in children (MIS-C) is a rare but severe disease temporarily related to SARS-CoV-2. We aimed to describe the epidemiological, clinical, and laboratory findings of all MIS-C cases diagnosed in children < 18 years old in Catalonia (Spain) to study their trend throughout the pandemic. This was a multicenter ambispective observational cohort study (April 2020-April 2022). Data were obtained from the COVID-19 Catalan surveillance system and from all hospitals in Catalonia. We analyzed MIS-C cases regarding SARS-CoV-2 variants for demographics, symptoms, severity, monthly MIS-C incidence, ratio between MIS-C and accumulated COVID-19 cases, and associated rate ratios (RR). Among 555,848 SARS-CoV-2 infections, 152 children were diagnosed with MIS-C. The monthly MIS-C incidence was 4.1 (95% CI: 3.4-4.8) per 1,000,000 people, and 273 (95% CI: 230-316) per 1,000,000 SARS-CoV-2 infections (i.e., one case per 3,700 SARS-CoV-2 infections). During the Omicron period, the MIS-C RR was 8.2 (95% CI: 5.7-11.7) per 1,000,000 SARS-CoV-2 infections, which was significantly lower (p < 0.001) than that for previous variant periods in all age groups. The median [IQR] age of MIS-C was 8 [4-11] years, 62.5% male, and 80.2% without comorbidities. Common symptoms were gastrointestinal findings (88.2%) and fever > 39 °C (81.6%); nearly 40% had an abnormal echocardiography, and 7% had coronary aneurysm. Clinical manifestations and laboratory data were not different throughout the variant periods (p > 0.05).  Conclusion: The RR between MIS-C cases and SARS-CoV-2 infections was significantly lower in the Omicron period for all age groups, including those not vaccinated, suggesting that the variant could be the main factor for this shift in the MISC trend. Regardless of variant type, the patients had similar phenotypes and severity throughout the pandemic. What is Known: • Before our study, only two publications investigated the incidence of MIS-C regarding SARS-CoV-2 variants in Europe, one from Southeast England and another from Denmark. What is New: • To our knowledge, this is the first study investigating MIS-C incidence in Southern Europe, with the ability to recruit all MIS-C cases in a determined area and analyze the rate ratio for MIS-C among SARS-CoV-2 infections throughout variant periods. • We found a lower rate ratio of MISC/infections with SARS-CoV-2 in the Omicron period for all age groups, including those not eligible for vaccination, suggesting that the variant could be the main factor for this shift in the MISC trend.

Activation-induced deaminase is critical for the establishment of DNA methylation patterns prior to the germinal center reaction.

Activation-induced deaminase (AID) initiates antibody diversification in germinal center B cells by deaminating cytosines, leading to somatic hypermutation and class-switch recombination. Loss-of-function mutations in AID lead to hyper-IgM syndrome type 2 (HIGM2), a rare human primary antibody deficiency. AID-mediated deamination has been proposed as leading to active demethylation of 5-methycytosines in the DNA, although evidence both supports and casts doubt on such a role. In this study, using whole-genome bisulfite sequencing of HIGM2 B cells, we investigated direct AID involvement in active DNA demethylation. HIGM2 naïve and memory B cells both display widespread DNA methylation alterations, of which ∼25% are attributable to active DNA demethylation. For genes that undergo active demethylation that is impaired in HIGM2 individuals, our analysis indicates that AID is not directly involved. We demonstrate that the widespread alterations in the DNA methylation and expression profiles of HIGM2 naïve B cells result from premature overstimulation of the B-cell receptor prior to the germinal center reaction. Our data support a role for AID in B cell central tolerance in preventing the expansion of autoreactive cell clones, affecting the correct establishment of DNA methylation patterns.

Curation and expansion of Human Phenotype Ontology for defined groups of inborn errors of immunity.

BACKGROUND: Accurate, detailed, and standardized phenotypic descriptions are essential to support diagnostic interpretation of genetic variants and to discover new diseases. The Human Phenotype Ontology (HPO), extensively used in rare disease research, provides a rich collection of vocabulary with standardized phenotypic descriptions in a hierarchical structure. However, to date, the use of HPO has not yet been widely implemented in the field of inborn errors of immunity (IEIs), mainly due to a lack of comprehensive IEI-related terms. OBJECTIVES: We sought to systematically review available terms in HPO for the depiction of IEIs, to expand HPO, yielding more comprehensive sets of terms, and to reannotate IEIs with HPO terms to provide accurate, standardized phenotypic descriptions. METHODS: We initiated a collaboration involving expert clinicians, geneticists, researchers working on IEIs, and bioinformaticians. Multiple branches of the HPO tree were restructured and extended on the basis of expert review. Our ontology-guided machine learning coupled with a 2-tier expert review was applied to reannotate defined subgroups of IEIs. RESULTS: We revised and expanded 4 main branches of the HPO tree. Here, we reannotated 73 diseases from 4 International Union of Immunological Societies-defined IEI disease subgroups with HPO terms. We achieved a 4.7-fold increase in the number of phenotypic terms per disease. Given the new HPO annotations, we demonstrated improved ability to computationally match selected IEI cases to their known diagnosis, and improved phenotype-driven disease classification. CONCLUSIONS: Our targeted expansion and reannotation presents enhanced precision of disease annotation, will enable superior HPO-based IEI characterization, and hence benefit both IEI diagnostic and research activities.

Multi-inflammatory Syndrome in Children Related to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in Spain.

Some clusters of children with a multisystem inflammatory syndrome (MIS-C) associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection have been reported. We describe the epidemiological and clinical features of children with MIS-C in Spain. MIS-C is a potentially severe condition that presents in children with recent SARS-CoV-2 infection.

Atypical Inflammatory Syndrome Triggered by SARS-CoV-2 in Infants with Down Syndrome.

While adults with Down syndrome (DS) are at increased risk of severe COVID-19 pneumonia, little is known about COVID-19 in children with DS. In children without DS, SARS-CoV-2 can rarely cause severe COVID-19 pneumonia, or an even rarer and more typically pediatric condition, multisystem inflammatory syndrome in children (MIS-C). Although the underlying mechanisms are still unknown, MIS-C is thought to be primarily immune-mediated. Here, we describe an atypical, severe form of MIS-C in two infant girls with DS who were hospitalized for over 4 months. Immunological evaluation revealed pronounced neutrophilia, B cell depletion, increased circulating IL-6 and IL-8, and elevated markers of immune activation ICAM1 and FcÉ£RI. Importantly, uninfected children with DS presented with similar but less stark immune features at steady state, possibly explaining risk of further uncontrolled inflammation following SARS-CoV-2 infection. Overall, a severe, atypical form of MIS-C may occur in children with DS.

Long-term outcome of LRBA deficiency in 76 patients after various treatment modalities as evaluated by the immune deficiency and dysregulation activity (IDDA) score.

BACKGROUND: Recent findings strongly support hematopoietic stem cell transplantation (HSCT) in patients with severe presentation of LPS-responsive beige-like anchor protein (LRBA) deficiency, but long-term follow-up and survival data beyond previous patient reports or meta-reviews are scarce for those patients who do not receive a transplant. OBJECTIVE: This international retrospective study was conducted to elucidate the longitudinal clinical course of patients with LRBA deficiency who do and do not receive a transplant. METHOD: We assessed disease burden and treatment responses with a specially developed immune deficiency and dysregulation activity score, reflecting the sum and severity of organ involvement and infections, days of hospitalization, supportive care requirements, and performance indices. RESULTS: Of 76 patients with LRBA deficiency from 29 centers (median follow-up, 10 years; range, 1-52), 24 underwent HSCT from 2005 to 2019. The overall survival rate after HSCT (median follow-up, 20 months) was 70.8% (17 of 24 patients); all deaths were due to nonspecific, early, transplant-related mortality. Currently, 82.7% of patients who did not receive a transplant (43 of 52; age range, 3-69 years) are alive. Of 17 HSCT survivors, 7 are in complete remission and 5 are in good partial remission without treatment (together, 12 of 17 [70.6%]). In contrast, only 5 of 43 patients who did not receive a transplant (11.6%) are without immunosuppression. Immune deficiency and dysregulation activity scores were significantly lower in patients who survived HSCT than in those receiving conventional treatment (P = .005) or in patients who received abatacept or sirolimus as compared with other therapies, and in patients with residual LRBA expression. Higher disease burden, longer duration before HSCT, and lung involvement were associated with poor outcome. CONCLUSION: The lifelong disease activity, implying a need for immunosuppression and risk of malignancy, must be weighed against the risks of HSCT.

Health-Related Quality of Life and Multidimensional Fatigue Scale in Children with Primary Immunodeficiencies.

PURPOSE: Patients with primary immunodeficiency disease (PID) have an increased risk of experiencing physical activity limitations, social difficulties, and psychological problems due to their chronic condition. Evaluation of their health-related quality of life (HRQOL) and fatigue is crucial in these patients to help understand their complex disease and provide adequate medical care. METHODS: In this study, we evaluated HRQOL and fatigue in pediatric and young adult patients with PID attending our center. Participants completed the Pediatric Quality of Life Inventory (PedsQL), version 4.0, and the PedsQL multidimensional fatigue module, standard version. RESULTS: Fifty-three PID patients were recruited (age range: 2-23 years). The mean HRQOL score obtained was 66.61 (SD: 18.73) out of 100, and the emotional and work/school dimensions were the ones most highly affected. There were no significant differences in reported quality of life between patients and their caregivers. The mean patient-reported fatigue value was 68.81 (SD: 17.80) out of 100, and the rest-related dimension was the one most highly affected. In the caregivers' assessment, general fatigue was the most highly affected dimension. CONCLUSIONS: The results of this study show that quality of life is poor and fatigue measures are considerably increased in our young adult and pediatric patients with PIDs. These findings can indicate areas requiring more intensive interventions, and they will serve as a basis for comparison of future results.

Neuroinflammatory Disease as an Isolated Manifestation of Hemophagocytic Lymphohistiocytosis.

Isolated neuroinflammatory disease has been described in case reports of familial hemophagocytic lymphohistiocytosis (FHL), but the clinical spectrum of disease manifestations, response to therapy and prognosis remain poorly defined. We combined an international survey with a literature search to identify FHL patients with (i) initial presentation with isolated neurological symptoms; (ii) absence of cytopenia and splenomegaly at presentation; and (iii) systemic HLH features no earlier than 3 months after neurological presentation. Thirty-eight (20 unreported) patients were identified with initial diagnoses including acute demyelinating encephalopathy, leukoencephalopathy, CNS vasculitis, multiple sclerosis, and encephalitis. Median age at presentation was 6.5 years, most commonly with ataxia/gait disturbance (75%) and seizures (53%). Diffuse multifocal white matter changes (79%) and cerebellar involvement (61%) were common MRI findings. CSF cell count and protein were increased in 22/29 and 15/29 patients, respectively. Fourteen patients progressed to systemic inflammatory disease fulfilling HLH-2004 criteria at a mean of 36.9 months after initial neurological presentation. Mutations were detected in PRF1 in 23 patients (61%), RAB27A in 10 (26%), UNC13D in 3 (8%), LYST in 1 (3%), and STXBP2 in 1 (3%) with a mean interval to diagnosis of 28.3 months. Among 19 patients who underwent HSCT, 11 neurologically improved, 4 were stable, one relapsed, and 3 died. Among 14 non-transplanted patients, only 3 improved or had stable disease, one relapsed, and 10 died. Isolated CNS-HLH is a rare and often overlooked cause of inflammatory brain disease. HLH-directed therapy followed by HSCT seems to improve survival and outcome.

Determining value in the treatment of activated PI3Kδ syndrome in Spain: a multicriteria decision analysis from the perspective of key stakeholders.

Introduction: Activated phosphoinositide 3-kinase (PI3K)δ syndrome (APDS) is an ultra-rare inborn error of immunity (IEI) combining immunodeficiency and immune dysregulation. This study determined what represents value in APDS in Spain from a multidisciplinary perspective applying multicriteria decision analysis (MCDA) methodology. Methods: A multidisciplinary committee of nine experts scored the evidence matrix. A specific framework for orphan drug evaluation in Spain and the weights assigned by a panel of 98 evaluators and decision-makers was used. Re-evaluation of scores was performed. Results: APDS is considered a very severe disease with important unmet needs, including misdiagnosis and diagnostic delay. Current management is limited to treatment of symptoms with off-label use of therapies supported by limited evidence. Therapeutic benefit is partial, resulting in limited disease control. Haematopoietic stem cell transplantation (HSCT), the only potential curative alternative, is restricted to a reduced patient population and without evidence of long-term efficacy or safety. All options present a limited safety profile. Data on patients' quality of life are lacking. APDS is associated with high pharmacological, medical and indirect costs. Conclusions: APDS is considered a severe disease, with limited understanding by key stakeholders of how treatment success is assessed in clinical practice, the serious impact that has on patients and the associated high economic burden. This study brings to light how MCDA methodology could represent a useful tool to complement current clinical and decision-making methods used by APDS experts and evaluators.

Higher COVID-19 pneumonia risk associated with anti-IFN-α than with anti-IFN-ω auto-Abs in children.

We found that 19 (10.4%) of 183 unvaccinated children hospitalized for COVID-19 pneumonia had autoantibodies (auto-Abs) neutralizing type I IFNs (IFN-α2 in 10 patients: IFN-α2 only in three, IFN-α2 plus IFN-ω in five, and IFN-α2, IFN-ω plus IFN-ß in two; IFN-ω only in nine patients). Seven children (3.8%) had Abs neutralizing at least 10 ng/ml of one IFN, whereas the other 12 (6.6%) had Abs neutralizing only 100 pg/ml. The auto-Abs neutralized both unglycosylated and glycosylated IFNs. We also detected auto-Abs neutralizing 100 pg/ml IFN-α2 in 4 of 2,267 uninfected children (0.2%) and auto-Abs neutralizing IFN-ω in 45 children (2%). The odds ratios (ORs) for life-threatening COVID-19 pneumonia were, therefore, higher for auto-Abs neutralizing IFN-α2 only (OR [95% CI] = 67.6 [5.7-9,196.6]) than for auto-Abs neutralizing IFN-ω only (OR [95% CI] = 2.6 [1.2-5.3]). ORs were also higher for auto-Abs neutralizing high concentrations (OR [95% CI] = 12.9 [4.6-35.9]) than for those neutralizing low concentrations (OR [95% CI] = 5.5 [3.1-9.6]) of IFN-ω and/or IFN-α2.

Common Variable Immunodeficiency and Neurodevelopmental Delay Due to a 13Mb Deletion on Chromosome 4 Including the NFKB1 Gene: A Case Report.

Syndromic immunodeficiencies are a heterogeneous group of inborn errors of immunity that can affect the development of non-immune organs and systems. The genetic basis of these immunodeficiencies is highly diverse, ranging from monogenic defects to large chromosomal aberrations. Antibody deficiency is the most prevalent immunological abnormality in patients with syndromic immunodeficiencies caused by chromosomal rearrangements, and usually manifests as a common variable immunodeficiency (CVID)-like phenotype. Here we describe a patient with a complex phenotype, including neurodevelopmental delay, dysmorphic features, malformations, and CVID (hypogammaglobulinemia, reduced pre-switch and switch memory B cells, and impaired vaccine response). Microarray-based comparative genomic hybridization (aCGH) revealed a 13-Mb deletion on chromosome 4q22.2-q24 involving 53 genes, some of which were related to the developmental manifestations in our patient. Although initially none of the affected genes could be linked to his CVID phenotype, subsequent reanalysis identified NFKB1 haploinsufficiency as the cause. This study underscores the value of periodic reanalysis of unsolved genetic studies performed with high-throughput technologies (eg, next-generation sequencing and aCGH). This is important because of the ongoing incorporation of new data establishing the relationship between genes and diseases. In the present case, NFKB1 had not been associated with human disease at the time aCGH was performed. Eight years later, reanalysis of the genes included in the chromosome 4 deletion enabled us to identify NFKB1 haploinsufficiency as the genetic cause of our patient's CVID. In the future, other genes included in the deletion may be linked to human disease, allowing us to better define the molecular basis of our patient's complex clinical phenotype.

Detection and evolutionary dynamics of somatic FAS variants in autoimmune lymphoproliferative syndrome: Diagnostic implications.

Autoimmune lymphoproliferative syndrome (ALPS) is a rare primary immune disorder characterized by impaired apoptotic homeostasis. The clinical characteristics include lymphoproliferation, autoimmunity (mainly cytopenia), and an increased risk of lymphoma. A distinctive biological feature is accumulation (>2.5%) of an abnormal cell subset composed of TCRαß+ CD4-CD8- T cells (DNTs). The most common genetic causes of ALPS are monoallelic pathogenic variants in the FAS gene followed by somatic FAS variants, mainly restricted to DNTs. Identification of somatic FAS variants has been typically addressed by Sanger sequencing in isolated DNTs. However, this approach can be costly and technically challenging, and may not be successful in patients with normal DNT counts receiving immunosuppressive treatment. In this study, we identified a novel somatic mutation in FAS (c.718_719insGTCG) by Sanger sequencing on purified CD3+ cells. We then followed the evolutionary dynamics of the variant along time with an NGS-based approach involving deep amplicon sequencing (DAS) at high coverage (20,000-30,000x). Over five years of clinical follow-up, we obtained six blood samples for molecular study from the pre-treatment (DNTs>7%) and treatment (DNTs<2%) periods. DAS enabled detection of the somatic variant in all samples, even the one obtained after five years of immunosuppressive treatment (DNTs: 0.89%). The variant allele frequency (VAF) range was 4%-5% in pre-treatment samples and <1.5% in treatment samples, and there was a strong positive correlation between DNT counts and VAF (Pearson's R: 0.98, p=0.0003). We then explored whether the same approach could be used in a discovery setting. In the last follow-up sample (DNT: 0.89%) we performed somatic variant calling on the FAS exon 9 DAS data from whole blood and purified CD3+ cells using VarScan 2. The c.718_719insGTCG variant was identified in both samples and showed the highest VAF (0.67% blood, 1.58% CD3+ cells) among >400 variants called. In summary, our study illustrates the evolutionary dynamics of a somatic FAS mutation before and during immunosuppressive treatment. The results show that pathogenic somatic FAS variants can be identified with the use of DAS in whole blood of ALPS patients regardless of their DNT counts.

Safety and Acceptance of COVID-19 Vaccination After Multisystem Inflammatory Syndrome in Children (MIS-C) in Spain.

In this cohort of 42 adolescents with a previous multisystem inflammatory syndrome (MIS-C) diagnosis, 32 (76.2%) were vaccinated with COVID-19 vaccines, with a low incidence of relevant adverse events. More importantly, no new MIS-C or myocarditis occurred after a median of 10 weeks (range 5.3-19.7) post-vaccination.

Epigenetic profiling linked to multisystem inflammatory syndrome in children (MIS-C): A multicenter, retrospective study.

Background: Most children and adolescents infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remain asymptomatic or develop a mild coronavirus disease 2019 (COVID-19) that usually does not require medical intervention. However, a small proportion of pediatric patients develop a severe clinical condition, multisystem inflammatory syndrome in children (MIS-C). The involvement of epigenetics in the control of the immune response and viral activity prompted us to carry out an epigenomic study to uncover target loci regulated by DNA methylation that could be altered upon the appearance of MIS-C. Methods: Peripheral blood samples were recruited from 43 confirmed MIS-C patients. 69 non-COVID-19 pediatric samples and 15 COVID-19 pediatric samples without MIS-C were used as controls. The cases in the two groups were mixed and divided into discovery (MIS-C = 29 and non-MIS-C = 56) and validation (MIS-C = 14 and non-MIS-C = 28) cohorts, and balanced for age, gender and ethnic background. We interrogated 850,000 CpG sites of the human genome for DNA methylation variants. Findings: The DNA methylation content of 33 CpG loci was linked with the presence of MIS-C. Of these sites, 18 (54.5%) were located in described genes. The top candidate gene was the immune T-cell mediator ZEB2; and others highly ranked candidates included the regulator of natural killer cell functional competence SH2D1B; VWA8, which contains a domain of the Von Willebrand factor A involved in the pediatric hemostasis disease; and human leukocyte antigen complex member HLA-DRB1; in addition to pro-inflammatory genes such as CUL2 and AIM2. The identified loci were used to construct a DNA methylation profile (EPIMISC) that was associated with MIS-C in both cohorts. The EPIMISC signature was also overrepresented in Kawasaki disease patients, a childhood pathology with a possible viral trigger, that shares many of the clinical features of MIS-C. Interpretation: We have characterized DNA methylation loci that are associated with MIS-C diagnosis. The identified genes are likely contributors to the characteristic exaggerated host inflammatory response observed in these patients. The described epigenetic signature could also provide new targets for more specific therapies for the disorder. Funding: Unstoppable campaign of Josep Carreras Leukaemia Foundation, Fundació La Marató de TV3, Cellex Foundation and CERCA Programme/Generalitat de Catalunya.

Multisystem Inflammatory Syndrome in Children in Western Countries? Decreasing Incidence as the Pandemic Progresses?: An Observational Multicenter International Cross-sectional Study.

BACKGROUND: SARS-CoV-2 variations as well as immune protection after previous infections and/or vaccination may have altered the incidence of multisystemic inflammatory syndrome in children (MIS-C). We aimed to report an international time-series analysis of the incidence of MIS-C to determine if there was a shift in the regions or countries included into the study. METHODS: This is a multicenter, international, cross-sectional study. We collected the MIS-C incidence from the participant regions and countries for the period July 2020 to November 2021. We assessed the ratio between MIS-C cases and COVID-19 pediatric cases in children <18 years diagnosed 4 weeks earlier (average time for the temporal association observed in this disease) for the study period. We performed a binomial regression analysis for 8 participating sites [Bogotá (Colombia), Chile, Costa Rica, Lazio (Italy), Mexico DF, Panama, The Netherlands and Catalonia (Spain)]. RESULTS: We included 904 cases of MIS-C, among a reference population of 17,906,432 children. We estimated a global significant decrease trend ratio in MIS-C cases/COVID-19 diagnosed cases in the previous month ( P < 0.001). When analyzing separately each of the sites, Chile and The Netherlands maintained a significant decrease trend ( P < 0.001), but this ratio was not statistically significant for the rest of sites. CONCLUSIONS: To our knowledge, this is the first international study describing a global reduction in the trend of the MIS-C incidence during the pandemic. COVID-19 vaccination and other factors possibly linked to the virus itself and/or community transmission may have played a role in preventing new MIS-C cases.