Foxp3+ regulatory T cells (Treg) are critical for the maintenance of self-tolerance, and their absence or dysfunction can result in autoimmunity. To determine the critical cell type controlled by Treg and potentially the suppressor mechanism utilized by Treg in the steady state, we utilized mice expressing the diphtheria toxin receptor (DTR) exclusively on Treg cells. Complete depletion of Treg was achieved 24 h after DT treatment, but profound activation of CD4+ and CD8+ T cells as measured by induction of CD44 expression and proliferation required 3-4 days. Increased expression of CD80/CD86 was observed on dendritic cells and more prominently on macrophages after 3 days. Depletion of CD4+ T cells or macrophages resulted in ∼50% inhibition of T-cell activation. The initial steps in T-cell activation were completely independent of IFN-γ or IL-2, while upregulation of CD80/CD86 was partially dependent on IFN-γ. Complete reversal of immune activation post-Treg depletion was only achieved by blockade of CD80/CD86 interactions with CD28. We conclude that the major mechanism used by Treg in the steady state is the regulation of CD80/CD86 expression and dysregulation of this suppressor pathway results in lethal autoimmunity driven by co-stimulatory signals in concert with TCR stimulation, or even by costimulatory signals alone.

Biallelic loss-of-function mutations in the lipopolysaccharide-responsive and beige-like anchor (LRBA) gene lead to a severe syndrome of early-onset immune dysregulation called LRBA deficiency. Monoallelic CTLA4 mutations lead to a similar phenotype. In both conditions, cytotoxic T lymphocyte-associated protein 4 (CTLA-4) levels are significantly decreased. In previously reported cases of symptomatic disease associated with LRBA pathogenic variants, patients usually have severely decreased or absent LRBA protein levels.

We describe 5 patients with biallelic missense variants in the LRBA gene presenting predominantly with Evans syndrome or colitis.

LRBA and CTLA-4 levels were investigated in LRBA missense, "classic" LRBA and in CTLA-4 insufficiency samples.

Surprisingly, all 5 LRBA missense patients had normal expression of LRBA protein. However, CTLA-4 intracellular expression was reduced to similar levels as those seen in patients with CTLA-4 insufficiency at resting state. Lower levels of surface CTLA-4 are seen on cell activation, indicating that these LRBA variants lead to reduced CTLA-4 cell surface expression. Several of the missense variants are shared between unrelated patients in the cohort, suggesting a mutational hot spot or founder effect for those with shared ancestry.

Novel LRBA deficiency variants result in quantitative or qualitative LRBA defects, leading to reduced intracellular resting levels and induced surface levels of CTLA-4.

Primary immunodeficiency diseases (PIDs) are associated with multiple genetic alterations including mutations of the lipopolysaccharide responsive Beige anchor (LRBA) gene. Nonsense mutations in the LRBA gene resulting in premature termination codons cause the loss of LRBA protein expression in PID. We evaluated the impact of a translational readthrough-inducing drug (TRID) ataluren as a nonsense suppression therapy in a PID patient with a homozygous stop codon mutation in exon 30 of LRBA. A precision medicine approach allowed us to pass from "in silico" to "in vitro" to the "bedside": following the in vitro treatment of patient-derived primary fibroblasts with ataluren, we observed a restoration of the LRBA protein expression and localization. In silico predictions suggested LRBA retained function after readthrough. Based on the successful experimental and computational results we treated the patient with ataluren resulting in an improvement of his clinical symptoms and quality of life. Importantly, the clinical symptoms were associated with a recovery of LRBA expression in liver biopsies post-treatment compared with pre-treatment. Our results provide a proof of concept demonstrating that ataluren, can rescue LRBA expression in PID. This work highlights the potential for personalized precision medicine approaches to be exploited for different genetic diseases due to premature termination codons.

The aim of this study was to explore the impact of rare and ultra-rare genetic variants on the understanding and treatment of autoimmune and autoinflammatory diseases with a focus on systemic lupus erythematosus (SLE) and Behçet syndrome. This review summarizes current research on the monogenic causes of SLE and Behçet syndrome, highlighting the various pathways that can be responsible for these unique phenotypes. In monogenic SLE, the identification of complement and DNASE1L3 deficiencies has elucidated mechanisms of apoptotic body accumulation and extracellular nucleic acid sensing. Type I interferonopathies underline the specific role of DNA/RNA sensing and the interferon overexpression in the development of systemic autoimmunity. Other significant genetic defects include Toll-like receptor hypersignaling and JAK/STATopathies, which contribute to the breakdown of immune tolerance. To date, genetic defects directly affecting B and T cell biology only account for a minority of identified causes of monogenic lupus, highlighting the importance of a tight regulation of mechanistic target of rapamycin and RAS (Rat sarcoma GTPase)/MAPK (mitogen-activated protein kinase) signaling in lupus. In Behçet syndrome, rare variants in TNFAIP3, RELA, and NFKB1 genes have been identified, underscoring the importance of NF-κB overactivation. Additional monogenic diseases such as ELF4, WDR1 mutations and trisomy 8 further illustrate the genetic complexity of this condition. Observations from genetic studies in SLE and Behçet syndrome highlight the complexity of systemic inflammatory diseases in which distinct molecular defects caused by single-gene mutations can promote lupus or Behçet syndromes, often unrecognizable from their genetically complex "classical" forms. Insights gained from studying rare genetic variants enhance our understanding of immune function in health and disease, paving the way for targeted therapies and personalized medicine.

We identify BEACH domain-containing proteins (BDCPs) as novel membrane coat proteins involved in the sorting of transmembrane proteins (TMPs) on the trans-Golgi network and tubular sorting endosomes. The seven typical mammalian BDCPs share a predicted alpha-solenoid-beta propeller structure, suggesting they have a protocoatomer origin and function. We map the subcellular localization of seven BDCPs based on their dynamic colocalization with RAB and ARF small GTPases and identify five typical BDCPs on subdomains of dynamic tubular-vesicular compartments on the intersection of endocytic recycling and post-Golgi secretory pathways. We demonstrate that BDCPs interact directly with the cytosolic tails of selected TMPs and identify a subset of TMPs, whose trafficking to the plasma membrane is affected in cells lacking BDCP. We propose that the competitive binding of BDCPs and clathrin coat adaptors to the cytosolic tails of TMPs, followed by their clustering to distinct subdomains of secretory/recycling tubules function as a mechanism for sorting of TMPs in pleomorphic tubular-vesicular compartments that lack a clathrin coat.

Deleterious mutations in the lipopolysaccharide responsive beige-like anchor protein (LRBA) gene cause severe childhood immune dysregulation. The complexity of the symptoms involving multiple organs and the broad range of unpredictable clinical manifestations of LRBA deficiency complicate the choice of therapeutic interventions. Although LRBA has been linked to Rab11-dependent trafficking of the immune checkpoint protein CTLA-4, its precise cellular role remains elusive. We show that LRBA, however, only slightly colocalizes with Rab11. Instead, LRBA is recruited by members of the small GTPase Arf protein family to the TGN and to Rab4+ endosomes, where it controls intracellular traffic. In patient-derived fibroblasts, loss of LRBA led to defects in the endosomal pathway promoting the accumulation of enlarged endolysosomes and lysosome secretion. Thus, LRBA appears to regulate flow through the endosomal system on Rab4+ endosomes. Our data strongly suggest functions of LRBA beyond CTLA-4 trafficking and provide a conceptual framework to develop new therapies for LRBA deficiency.

Anophthalmia, microphthalmia and coloboma (AMC) comprise a spectrum of developmental eye disorders, accounting for approximately 20% of childhood visual impairment. While non-coding regulatory sequences are increasingly recognised as contributing to disease burden, characterising their impact on gene function and phenotype remains challenging. Furthermore, little is known of the nature and extent of their contribution to AMC phenotypes. We report two families with variants in or near MAB21L2, a gene where genetic variants are known to cause AMC in humans and animal models. The first proband, presenting with microphthalmia and coloboma, has a likely pathogenic missense variant (c.338 G > C; p.[Trp113Ser]), segregating within the family. The second individual, presenting with microphthalmia, carries an ~ 113.5 kb homozygous deletion 19.38 kb upstream of MAB21L2. Modelling of the deletion results in transient small lens and coloboma as well as midbrain anomalies in zebrafish, and microphthalmia and coloboma in Xenopus tropicalis. Using conservation analysis, we identify 15 non-coding conserved elements (CEs) within the deleted region, while ChIP-seq data from mouse embryonic stem cells demonstrates that two of these (CE13 and 14) bind Otx2, a protein with an established role in eye development. Targeted disruption of CE14 in Xenopus tropicalis recapitulates an ocular coloboma phenotype, supporting its role in eye development. Together, our data provides insights into regulatory mechanisms underlying eye development and highlights the importance of non-coding sequences as a source of genetic diagnoses in AMC.

Common Variable Immunodeficiency (CVID) is a primary immunodeficiency characterized by reduced levels of specific immunoglobulins, resulting in frequent infections, autoimmune disorders, increased cancer risk, and diminished antibody production despite an adequate B cell count. With its clinical manifestations being highly variable, the classification of CVID, including the widely recognized Freiburg classification, is primarily based on clinical symptoms and genetic variations. Our study aims to refine the classification of CVID by analyzing transcriptomics data to identify distinct disease subtypes. We utilized the GSE51405 dataset, examining transcriptomic profiles from 30 CVID patients without complications. Employing a combination of clustering techniques-KMeans, hierarchical agglomerative clustering, spectral clustering, and Gaussian Mixture models-and differential gene expression analysis with R's limma package, we integrated molecular findings with demographic data (age and gender) through correlation analysis and identified common genes among clusters. Three distinct clusters of CVID patients were identified using KMeans, Agglomerative Clustering, and Gaussian Mixture Models, highlighting the disease's heterogeneity. Differential expression analysis unveiled 31 genes with variable expression levels across these clusters. Notably, nine genes (EIF5A, RPL21, ANP32A, DTX3L, NCF2, CDC42EP3, CHP1, FOLR3, and DEFA4) exhibited consistent differential expression across all clusters, independent of demographic factors. The study recommends categorizing patients based on the four genes, NCF2, CHP1, FOLR3, and DEFA4-as they may assist in prognostic prediction. Transcriptomic analysis of common variable immunodeficiency (CVID) patients identified three distinct clusters based on gene expression, independent of age and gender. Nine differentially expressed genes were identified across these clusters, suggesting potential biomarkers for CVID subtype classification. These findings highlight the genetic heterogeneity of CVID and provide novel insights into disease classification and potential personalized treatment approaches.

LRBA deficiency is an inborn error of immunity defined by autoimmunity, lymphoproliferation, recurrent infections, cytopenia, and inflammatory bowel disease. Despite recent advances in managing this disease with targeted biologic therapy, haematopoietic stem cell transplant (HSCT) remains the only cure. However, great variability exists between protocols used to transplant patients with LRBA deficiency. We describe a cohort of seven patients with LRBA deficiency who underwent HSCT using a myeloablative, reduced toxicity regime of fludarabine, treosulfan, and thiotepa at two transplantation centres from 2016 to 2019. Data were collected both retrospectively and prospectively, measuring time to engraftment, infectious complications, incidence of graft versus host disease, and post-transplantation chimerism. Six of seven patients survived transplantation, and four of six surviving patients achieving treatment-free survival. We thus recommend that HSCT with fludarabine, treosulfan, and thiotepa-based conditioning be considered in patients with LRBA deficiency.

Newly developing genomic technologies are an increasingly important part of clinical care and thus, it is not only important to understand the technologies and their limitations, but to also interpret the findings in an actionable fashion. Clinical geneticists and genetic counselors are now an integral part of the clinical team and are able to bridge the complexities of this rapidly changing science between the bedside clinicians and patients. This manuscript reviews the terminology, the current technology, some of the known genetic disorders that result in lung disease, and indications for genetic testing with associated caveats. Because this field is evolving quickly, we also provide links to websites that provide continuously updated information important for integrating genomic technology results into clinical decision-making.

Hematopoietic stem cells (HSCs) can generate all blood cells. This ability is exploited in HSC transplantation (HSCT) to treat hematologic disease. A clear understanding of the molecular mechanisms that regulate HSCT is necessary to continue improving transplant protocols. We identified the Beige and Chediak-Higashi domain-containing protein (BDCP), Neurobeachin (NBEA), as a putative regulator of HSCT. Here, we demonstrated that NBEA and related BDCPs, including LPS Responsive Beige-Like Anchor Protein (LRBA), Neurobeachin Like 1 (NBEAL1) and Lysosomal Trafficking Regulator (LYST), are required during HSCT to efficiently reconstitute the hematopoietic system of lethally irradiated mice. Nbea knockdown in mouse HSCs induced apoptosis and a differentiation block after transplantation. Nbea deficiency in hematopoietic progenitor cells perturbed the expression of genes implicated in vesicle trafficking and led to changes in NOTCH receptor localization. This resulted in perturbation of the NOTCH transcriptional program, which is required for efficient HSC engraftment. In summary, our findings reveal a novel role for NBEA in the control of NOTCH receptor turnover in hematopoietic cells and supports a model in which BDCP-regulated vesicle trafficking is required for efficient HSCT.

Lipopolysaccharide-responsive and beige-like anchor (LRBA) is a scaffolding protein that interacts with proteins such as CTLA-4 and PKA, the importance of which has been determined in various cell types, including T regulatory cells, B cells, and renal cells. LRBA deficiency is associated with an inborn error in immunity characterized by immunodeficiency and autoimmunity. In addition to defects in T regulatory cells, patients with LRBA deficiency also exhibit B cell defects, such as reduced cell number, low memory B cells, hypogammaglobulinemia, impaired B cell proliferation, and increased autophagy. Although Lrba-/- mice do not exhibit the immunodeficiency observed in humans, responses to B cell receptors (BCR) in B cells have not been explored. Therefore, a murine model is for elucidating the mechanism of Lrba mechanism in B cells.

To compare and evaluate spleen-derived B cell responses to BCR crosslinking in C57BL6 Lrba-/- and Lrba+/+ mice.

Spleen-derived B cells were obtained from 8 to 12-week-old mice. Subpopulations were determined by immunostaining and flow cytometry. BCR crosslinking was assessed by the F(ab')2 anti-μ chain. Activation, proliferation and viability assays were performed using flow cytometry and protein phosphorylation was evaluated by immunoblotting. The nuclear localization of p65 was determined using confocal microscopy. Nur77 expression was evaluated by Western blot.

Lrba-/- B cells showed an activated phenotype and a decreased proportion of transitional 1 B cells, and both proliferation and survival were affected after BCR crosslinking in the Lrba-/- mice. The NF-κB pathway exhibited a basal activation status of several components, resulting in increased activation of p50, p65, and IκBα, basal p50 activation was reduced by the Plcγ2 inhibitor U73122. BCR crosslinking in Lrba-/ - B cells resulted in poor p50 phosphorylation and p65 nuclear localization. Increased levels of Nur77 were detected.

These results indicate the importance of Lrba in controlling NF-κB activation driven by BCR. Basal activation of NF-κB could impact cellular processes, such as, activation, differentiation, proliferation, and maintenance of B cells after antigen encounter.

Lrba is a cytoplasmic protein involved in vesicular trafficking. Lrba-deficient (Lrba-/-) mice exhibit substantially higher levels of IgA in both serum and feces than wild-type (WT) mice. Transforming growth factor β1 (TGFβ1) and its receptors (TGFβR I and II) is essential for differentiating IgA+ B cells. Furthermore, increased IgA production suggests a potential connection between Lrba and the TGFβR signaling pathway in IgA production. However, the specific function of Lrba in B cell biology remains unknown.

Given the increased IgA levels in Lrba-/- mice, the goal in this work was to explore the lymph organs where the switch to IgA occurs, and if TGFβR function is affected.

Non-immunized Lrba-/- mice were compared with Lrba+/+ mice. IgA levels in the serum and feces, as well as during peripheral B cell development, were determined. IgA+ B cells and plasma cells were assessed in the small intestine and secondary lymphoid organs, such as the spleen, mesenteric lymph nodes, and Peyer's patches. The TGFβR signaling pathway was evaluated by determining the expression of TGFβR on B cells. Additionally, SMAD2 phosphorylation was measured under basal conditions and in response to recombinant TGFβ. Finally, confocal microscopy was performed to investigate a possible interaction between Lrba and TGFβR in B cells.

Lrba-/- mice exhibited significantly higher levels of circulating IgA, IgA+ B, and plasma cells than in peripheral lymphoid organs those in WT mice. TGFβR expression on the membrane of B cells was similar in both Lrba-/- and Lrba+/+ mice. However, intracellular TGFβR expression was reduced in Lrba-/- mice. SMAD2 phosphorylation showed increased levels under basal conditions; stimulation with recombinant TGFβ elicited a poorer response than in that in Lrba+/+ B cells. Finally, we found that Lrba colocalizes with TGFβR in B cells.

Lrba is essential in controlling TGFβR signaling, subsequently regulating SMAD2 phosphorylation on B cells. This mechanism may explain the increased differentiation of IgA+ B cells and production of IgA-producing plasma cells.

LRBA is a cytoplasmic protein that is ubiquitously distributed. Almost all LRBA domains have a scaffolding function. In 2012, it was reported that homozygous variants in LRBA are associated with early-onset hypogammaglobulinemia. Since its discovery, more than 100 pathogenic variants have been reported. This review focuses on the variants reported in LRBA and their possible associations with clinical phenotypes. In this work LRBA deficiency cases reported more than 11 years ago have been revised. A database was constructed to analyze the type of variants, age at onset, clinical diagnosis, infections, autoimmune diseases, and cellular and immunoglobulin levels. The review of cases from 2012 to 2023 showed that LRBA deficiency was commonly diagnosed in patients with a clinical diagnosis of Common Variable Immunodeficiency, followed by enteropathy, neonatal diabetes mellitus, ALPS, and X-linked-like syndrome. Most cases show early onset of presentation at <6 years of age. Most cases lack protein expression, whereas hypogammaglobulinemia is observed in half of the cases, and IgG and IgA levels are isotypes reported at low levels. Patients with elevated IgG levels exhibited more than one autoimmune manifestation. Patients carrying pathogenic variants leading to a premature stop codon show a severe phenotype as they have an earlier onset of disease presentation, severe autoimmune manifestations, premature death, and low B cells and regulatory T cell levels. Missense variants were more common in patients with low IgG levels and cytopenia. This work lead to the conclusion that the type of variant in LRBA has association with disease severity, which leads to a premature stop codon being the ones that correlates with severe disease.

Mutations in LRBA, a BEACH domain protein, cause severe immune deficiency in humans. LRBA is expressed in many tissues and organs according to biochemical analysis, but little is known about its cellular and subcellular localization, and its deficiency phenotype outside the immune system. By LacZ histochemistry of Lrba gene-trap mice, we performed a comprehensive survey of LRBA expression in numerous tissues, detecting it in many if not all epithelia, in exocrine and endocrine cells, and in subpopulations of neurons. Immunofluorescence microscopy of the exocrine and endocrine pancreas, salivary glands, and intestinal segments, confirmed these patterns of cellular expression and provided information on the subcellular localizations of the LRBA protein. Immuno-electron microscopy demonstrated that in neurons and endocrine cells, which co-express LRBA and its closest relative, neurobeachin, both proteins display partial association with endomembranes in complementary, rather than overlapping, subcellular distributions. Prominent manifestations of human LRBA deficiency, such as inflammatory bowel disease or endocrinopathies, are believed to be primarily due to immune dysregulation. However, as essentially all affected tissues also express LRBA, it is possible that LRBA deficiency enhances their vulnerability and contributes to the pathogenesis.

Regulatory T cells (Tregs) are a specialized subgroup of T-cell lymphocytes that is crucial for maintaining immune homeostasis and preventing excessive immune responses. Depending on their differentiation route, Tregs can be subdivided into thymically derived Tregs (tTregs) and peripherally induced Tregs (pTregs), which originate from conventional T cells after extrathymic differentiation at peripheral sites. Although the regulatory attributes of tTregs and pTregs partially overlap, their modes of action, protein expression profiles, and functional stability exhibit specific characteristics unique to each subset. Over the last few years, our knowledge of Treg differentiation, maturation, plasticity, and correlations between their phenotypes and functions has increased. Genetic and functional studies in patients with numeric and functional Treg deficiencies have contributed to our mechanistic understanding of immune dysregulation and autoimmune pathologies. This review provides an overview of our current knowledge of Treg biology, discusses monogenetic Treg pathologies and explores the role of Tregs in various other autoimmune disorders. Additionally, we discuss novel approaches that explore Tregs as targets or agents of innovative treatment options.

Primary immune regulatory disorders (PIRDs) constitute a spectrum of inborn errors of immunity (IEIs) that are primarily characterized by autoimmunity, lymphoproliferation, atopy, and malignancy. In PIRDs, infections are infrequent compared to other IEIs. While susceptibility to infection primarily stems from antibody deficiency, it is sometimes associated with additional innate immune and T or NK cell defects. The use of immunotherapy and chemotherapy further complicates the immune landscape, increasing the risk of diverse infections. Recurrent sinopulmonary infections, particularly bacterial infections such as those associated with staphylococcal and streptococcal organisms, are the most reported infectious manifestations. Predisposition to viral infections, especially Epstein-Barr virus (EBV)-inducing lymphoproliferation and malignancy, is also seen. Notably, mycobacterial and invasive fungal infections are rarely documented in these disorders. Knowledge about the spectrum of infections in these disorders would prevent diagnostic delays and prevent organ damage. This review delves into the infection profile specific to autoimmune lymphoproliferative syndrome (ALPS), Tregopathies, and syndromes with autoimmunity within the broader context of PIRD. Despite the critical importance of understanding the infectious aspects of these disorders, there remains a scarcity of comprehensive reports on this subject.

The study aims to report a rare case of a novel homozygous variant in the LRBA gene, originating from uniparental disomy of paternal origin. This case contributes new clinical data to the LRBA gene variant database.

The study details the case of a 2-year-old child diagnosed in May 2023 at our center with a homozygous LRBA gene variant. Detailed clinical data of the patient were collected, including whole-exome sequencing of peripheral blood mononuclear cells, with parental genetic verification.

The child presented with recurrent respiratory infections and chronic neutropenia, progressing to pancytopenia. Imaging showed splenomegaly and enlarged lymph nodes in the axillary and abdominal regions. Peripheral blood lymphocyte count revealed reduced B cells and NK cells. Elevated cytokine levels of IFN-α and IFN-r were observed. Whole-exome sequencing revealed a nonsense homozygous variant in the LRBA gene, specifically c.2584C>T (p.Gln862Ter). The father exhibited a heterozygous variant at this locus, while no variant was found in the mother. Sample analysis indicated characteristics of uniparental disomy. According to the guidelines of the American College of Medical Genetics and Genomics (ACMG), this variant is preliminarily classified as "Likely pathogenic". Currently, there are no reports in academic literature regarding this specific variant site.

LRBA gene variants can lead to a rare inborn error of immunity disease. The c.2584C>T (p.Gln862Ter) variant in exon 22 of the LRBA gene is a newly identified pathogenic variant, and the homozygous variant caused by uniparental disomy is exceedingly rare. This case represents the second global report of an LRBA gene function loss due to uniparental disomy abnormalities.

Common variable immunodeficiency (CVID) is a heterogenous disease category created to distinguish late-onset antibody deficiencies from early-onset diseases like agammaglobulinemia or more expansively dysfunctional combined immunodeficiencies. Opinions vary on which affected patients should receive a CVID diagnosis which confuses clinicians and erects reproducibility barriers for researchers. Most experts agree that CVID's most indeliable feature is defective germinal center (GC) production of isotype-switched, affinity-maturated antibodies. Here, we review the biological factors contributing to CVID-associated GC dysfunction including genetic, epigenetic, tolerogenic, microbiome, and regulatory abnormalities. We also discuss the consequences of these biological phenomena to the development of non-infectious disease complications. Finally, we opine on topics and lines of investigation we think hold promise for expanding our mechanistic understanding of this protean condition and for improving the lives of affected patients.

Genome-wide association studies have successfully identified genetic variants associated with human disease. Various statistical approaches based on penalized and machine learning methods have recently been proposed for disease prediction. In this study, we evaluated the performance of several such methods for predicting asthma using the Korean Chip (KORV1.1) from the Korean Genome and Epidemiology Study (KoGES).

First, single-nucleotide polymorphisms were selected via single-variant tests using logistic regression with the adjustment of several epidemiological factors. Next, we evaluated the following methods for disease prediction: ridge, least absolute shrinkage and selection operator, elastic net, smoothly clipped absolute deviation, support vector machine, random forest, boosting, bagging, naïve Bayes, and k-nearest neighbor. Finally, we compared their predictive performance based on the area under the curve of the receiver operating characteristic curves, precision, recall, F1-score, Cohen's Kappa, balanced accuracy, error rate, Matthews correlation coefficient, and area under the precision-recall curve. Additionally, three oversampling algorithms are used to deal with imbalance problems.

Our results show that penalized methods exhibit better predictive performance for asthma than that achieved via machine learning methods. On the other hand, in the oversampling study, randomforest and boosting methods overall showed better prediction performance than penalized methods.

Chronic nonmalignant lymphoproliferation and autoimmune cytopenia are relevant manifestations of immunohematologic diseases of childhood. Their diagnostic classification is challenging but important for therapy. Autoimmune lymphoproliferative syndrome (ALPS) is a genetically defined inborn error of immunity combining these manifestations, but it can explain only a small proportion of cases. Diagnostic categories such as ALPS-like disease, common variable immunodeficiency, or Evans syndrome have therefore been used. Advances in genetics and increasing availablity of targeted therapies call for more therapy-oriented disease classification. Moreover, recent discoveries in the (re)analysis of genetic conditions affecting FAS signaling ask for a more precise definition of ALPS. In this review, we propose the term autoimmune lymphoproliferative immunodeficiencies for a disease phenotype that is enriched for patients with genetic diseases for which targeted therapies are available. For patients without a current molecular diagnosis, this term defines a subgroup of immune dysregulatory disorders for further studies. Within the concept of autoimmune lymphoproliferative immunodeficiencies, we propose a revision of the ALPS classification, restricting use of this term to conditions with clear evidence of perturbation of FAS signaling and resulting specific biologic and clinical consequences. This proposed approach to redefining ALPS and other lymphoproliferative conditions provides a framework for disease classification and diagnosis that is relevant for the many specialists confronted with these diseases.

Adult stem cells are long-lived and quiescent with unique metabolic requirements. Macroautophagy/autophagy is a fundamental survival mechanism that allows cells to adapt to metabolic changes by degrading and recycling intracellular components. Here we address why autophagy depletion leads to a drastic loss of the stem cell compartment. Using inducible deletion of autophagy specifically in adult hematopoietic stem cells (HSCs) and in mice chimeric for autophagy-deficient and normal HSCs, we demonstrate that the stem cell loss is cell-intrinsic. Mechanistically, autophagy-deficient HSCs showed higher expression of several amino acid transporters (AAT) when compared to autophagy-competent cells, resulting in increased amino acid (AA) uptake. This was followed by sustained MTOR (mechanistic target of rapamycin) activation, with enlarged cell size, glucose uptake and translation, which is detrimental to the quiescent HSCs. MTOR inhibition by rapamycin treatment in vivo was able to rescue autophagy-deficient HSC loss and bone marrow failure and resulted in better reconstitution after transplantation. Our results suggest that targeting MTOR may improve aged stem cell function, promote reprogramming and stem cell transplantation.List of abbreviations: 5FU: fluoracil; AA: amino acids; AKT/PKB: thymoma viral proto-oncogene 1; ATF4: activating transcription factor 4; BafA: bafilomycin A1; BM: bone marrow; EIF2: eukaryotic initiation factor 2; EIF4EBP1/4EBP1: eukaryotic translation initiation factor 4E binding protein 1; KIT/CD117/c-Kit: KIT proto-oncogene receptor tyrosine kinase; HSCs: hematopoietic stem cells; HSPCs: hematopoietic stem and progenitor cells; Kyn: kynurenine; LSK: lineage- (Lin-), LY6A/Sca-1+, KIT/c-Kit/CD117+; LY6A/Sca-1: lymphocyte antigen 6 family member A; MTOR: mechanistic target of rapamycin kinase; MTORC1: MTOR complex 1; MTORC2: MTOR complex 2; OPP: O-propargyl-puromycin; PI3K: phosphoinositide 3-kinase; poly(I:C): polyinosinic:polycytidylic acid; RPS6/S6: ribosomal protein S6; tam: tamoxifen; TCA: tricarboxylic acid; TFEB: transcription factor EB; PTPRC/CD45: Protein Tyrosine Phosphatase Receptor Type C, CD45 antigen.

Common variable immunodeficiency (CVID) has been subdivided into five phenotypes, including one marked by non-infectious enteropathies that lead to significant morbidity and mortality. We examined a large national registry of patients with CVID to better characterize this population and understand how the presence of enteropathy influences nutritional status, patient function, and the risk of additional non-infectious disorders in CVID patients. We also sought to illustrate the range of treatment strategies for CVID-associated enteropathies. We extracted patient data from the United States Immunodeficiency Network (USIDNET) database, which included 1415 patients with CVID, and compared those with and without intestinal disorders. Demographic and genetic profiles, functional status, and treatments targeting intestinal disorders are reported. Intestinal disorders were present in 20% of patients with CVID, including chronic diarrhea, inflammatory bowel disease, malabsorption, and others. Compared to those without enteropathies, this patient subset exhibited significantly lower Karnofsky-Lansky functional scores, greater reliance on nutritional support, higher rates of vitamin deficiencies, and increased prevalence of hematologic disorders, liver disease, pulmonary disease, granulomatous disease, and lymphoma. Genetic data were reported for only 5% of the cohort. No mutations segregated significantly to patients with or without intestinal disease. Corticosteroids were most frequently used for treatment. Patients with CVID-associated intestinal disorders exhibit higher rates of autoimmune and inflammatory comorbidities, lymphoma, malnutrition, and debility. We review recent studies implicating specific pathways underlying this immune dysregulation. Further studies are needed to evaluate the role of targeted immunomodulatory therapies for CVID-associated intestinal disorders.

Common variable immunodeficiency (CVID) is the most common symptomatic inborn error of immunity. The disorder is characterized by variable clinical and immunological manifestations, and, in a small minority of patients, a monogenic cause may be identified. In this review, we focalized on three different monogenic forms of CVID-like disease.

Activated phosphoinositide 3-kinase delta syndrome (APDS) is a rare disorder characterized by hyperactivated class I phosphatidylinositol-3 kinase (PI3K) pathway. Affected patients present with respiratory infectious episodes, impaired viral clearance and lymphoproliferation. Recently, a direct PI3K inhibitor has been approved and it showed encouraging results both in controlling clinical and immunological manifestations of the disease. On the other hand, patients with defects in CTLA-4 or LRBA gene present with life-threatening immune dysregulation, autoimmunity and lymphocytic infiltration of multiple organs. Abatacept, a soluble cytotoxic T lymphocyte antigen 4 (CTLA-4) fusion protein that acts as a costimulation modulator, has been widely implemented for affected patients with good results as bridge treatment.

Understanding the biological basis of CVID is important not only for enriching our knowledge of the human immune system, but also for setting the basis for potential targeted treatments in this disorder.

Whole-exome sequencing (WES) provides a powerful diagnostic tool for identifying primary immunodeficiency diseases (PIDs). This study explores the utility of this approach in uncovering previously undiagnosed PIDs in children with community-acquired sepsis (CAS), with a medical history of recurrent infections or a family history of PIDs.

We performed WES on DNA samples extracted from the blood of the 34 enrolled patients, followed by bioinformatic analysis for variant calling, annotation, and prioritization. We also performed a segregation analysis in available family members to confirm the inheritance patterns and assessed the potential impact of the identified variants on protein function.

From 34 patients enrolled in the study, 29 patients (85%) with previously undiagnosed genetic diseases, including 28 patients with PIDs and one patient with interstitial lung and liver disease, were identified. We identified two patients with severe combined immunodeficiency (SCID), patients with combined immunodeficiency (CID), six patients with combined immunodeficiency with syndromic features (CID-SF), four patients with defects in intrinsic and innate immunity, four patients with congenital defects of phagocyte function (CPDF), and six patients with the disease of immune dysregulation. Autoinflammatory disorders and predominantly antibody deficiency were diagnosed in one patient each.

Our findings demonstrate the potential of WES in identifying undiagnosed PIDs in children with CAS. Implementing WES in the clinical evaluation of CAS patients with a warning sign for PIDs can aid in their timely diagnosis and potentially lead to improved patient care.

LPS-responsive beige-like anchor (LRBA) deficiency (LRBA-/-) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA4) insufficiency (CTLA4+/-) are mechanistically overlapped diseases presenting with recurrent infections and autoimmunity. The effectiveness of different treatment regimens remains unknown.

Our aim was to determine the comparative efficacy and long-term outcome of therapy with immunosuppressants, CTLA4-immunoglobulin (abatacept), and hematopoietic stem cell transplantation (HSCT) in a single-country multicenter cohort of 98 patients with a 5-year median follow-up.

The 98 patients (63 LRBA-/- and 35 CTLA4+/-) were followed and evaluated at baseline and every 6 months for clinical manifestations and response to the respective therapies.

The LRBA-/- patients exhibited a more severe disease course than did the CTLA4+/- patients, requiring more immunosuppressants, abatacept, and HSCT to control their symptoms. Among the 58 patients who received abatacept as either a primary or rescue therapy, sustained complete control was achieved in 46 (79.3%) without severe side effects. In contrast, most patients who received immunosuppressants as primary therapy (n = 61) showed either partial or no disease control (72.1%), necessitating additional immunosuppressants, abatacept, or transplantation. Patients with partial or no response to abatacept (n = 12) had longer disease activity before abatacept therapy, with higher organ involvement and poorer disease outcomes than those with a complete response. HSCT was performed in 14 LRBA-/- patients; 9 patients (64.2%) showed complete remission, and 3 (21.3%) continued to receive immunosuppressants after transplantation. HSCT and abatacept therapy gave rise to similar probabilities of survival.

Abatacept is superior to immunosuppressants in controlling disease manifestations over the long term, especially when started early, and it may provide a safe and effective therapeutic alternative to transplantation.

Our goal is to review current understanding of interstitial lung disease (ILD) affecting patients with inborn errors of immunity (IEI). This includes understanding how IEI might predispose to and promote development or progression of ILD as well as how our growing understanding of IEI can help shape treatment of ILD in these patients. Additionally, by examining current knowledge of ILD in IEI, we hope to identify key knowledge gaps that can become focus of future investigative efforts.

Recent identification of novel IEI associated with ILD and the latest reports examining treatment of ILD in IEI are included. Of noted interest, are recent clinical studies of immunomodulatory therapy for ILD in common variable immunodeficiency.

ILD is a frequent complication found in many IEI. This article provides a guide to identifying manifestations of ILD in IEI. We review a broad spectrum of IEI that develop ILD, including antibody deficiency and immune dysregulation disorders that promote autoimmunity and autoinflammation. This work integrates clinical information with molecular mechanisms of disease and diagnostic assessments to provide an expedient overview of a clinically relevant and expanding topic.

Common variable immunodeficiency (CVID) is an inborn error of immunity characterized by low levels of antibodies. In addition to infections, many patients also suffer from T-helper 1-driven immune dysregulation, which is associated with increased mortality. The aim of this study was to perform in-depth characterization of the T and the B cell compartments in a well-defined cohort of patients affected by CVID and correlate the findings to the level of clinical immune dysregulation. We used mass cytometry, targeted proteomics, flow cytometry and functional assays to delineate the immunological phenotype of 15 CVID-affected patients with different levels of immune dysregulation. Unbiased clustering of T cell mass cytometry data correlated with CVID-related immune dysregulation and plasma protein profiles. Expanded CXCR3+ T-bet-expressing B cells correlated with effector memory CD4+ T cell clusters, and increased plasma levels of CXCR3-ligands. Our findings indicate an interplay between B cells and T cells in CVID-related immune dysregulation and provide a better understanding of the underlying pathological mechanisms.

Primary immune regulatory disorders (PIRDs) are inborn errors of immunity caused by a loss in the regulatory mechanism of the inflammatory or immune response, leading to impaired immunological tolerance or an exuberant inflammatory response to various stimuli due to loss or gain of function mutations. Whilst PIRDs may feature susceptibility to recurrent, severe, or opportunistic infection in their phenotype, this group of syndromes has broadened the spectrum of disease caused by defects in immunity-related genes to include autoimmunity, autoinflammation, lymphoproliferation, malignancy, and allergy; increasing focus on PIRDs has thus redefined the classical 'primary immunodeficiency' as one aspect of an overarching group of inborn errors of immunity. The growing number of genetic defects associated with PIRDs has expanded our understanding of immune tolerance mechanisms and prompted identification of molecular targets for therapy. However, PIRDs remain difficult to recognize due to incomplete penetrance of their diverse phenotype, which may cross organ systems and present to multiple clinical specialists prior to review by an immunologist. Control of immune dysregulation with immunosuppressive therapies must be balanced against the enhanced infective risk posed by the underlying defect and accumulated end-organ damage, posing a challenge to clinicians. Whilst allogeneic hematopoietic stem cell transplantation may correct the underlying immune defect, identification of appropriate patients and timing of transplant is difficult. The relatively recent description of many PIRDs and rarity of individual genetic entities that comprise this group means data on natural history, clinical progression, and treatment are limited, and so international collaboration will be needed to better delineate phenotypes and the impact of existing and potential therapies. This review explores pathophysiology, clinical features, current therapeutic strategies for PIRDs including cellular platforms, and future directions for research.

Nonsense mutations cause several genetic diseases such as cystic fibrosis, Duchenne muscular dystrophy, β-thalassemia, and Shwachman-Diamond syndrome. These mutations induce the formation of a premature termination codon (PTC) inside the mRNA sequence, resulting in the synthesis of truncated polypeptides. Nonsense suppression therapy mediated by translational readthrough-inducing drugs (TRIDs) is a promising approach to correct these genetic defects. TRIDs generate a ribosome miscoding of the PTC named "translational readthrough" and restore the synthesis of full-length and potentially functional proteins. The new oxadiazole-core TRIDs NV848, NV914, and NV930 (NV) showed translational readthrough activity in nonsense-related in vitro systems. In this work, the possible off-target effect of NV molecules on natural termination codons (NTCs) was investigated. Two different in vitro approaches were used to assess if the NV molecule treatment induces NTC readthrough: (1) a study of the translational-induced p53 molecular weight and functionality; (2) the evaluation of two housekeeping proteins' (Cys-C and β2M) molecular weights. Our results showed that the treatment with NV848, NV914, or NV930 did not induce any translation alterations in both experimental systems. The data suggested that NV molecules have a specific action for the PTCs and an undetectable effect on the NTCs.

Many inborn errors of immunity (IEI) manifest with hallmarks of both immunodeficiency and immune dysregulation due to uncontrolled immune responses and impaired immune homeostasis. A subgroup of these disorders frequently presents with autoimmunity and lymphoproliferation (ALPID phenotype). After the initial description of the genetic basis of autoimmune lymphoproliferative syndrome (ALPS) more than 20 years ago, progress in genetics has helped to identify many more genetic conditions underlying this ALPID phenotype. Among these, the majority is caused by a group of autosomal-dominant conditions including CTLA-4 haploinsufficiency, STAT3 gain-of-function disease, activated PI3 kinase syndrome, and NF-κB1 haploinsufficiency. Even within a defined genetic condition, ALPID patients may present with staggering clinical heterogeneity, which makes diagnosis and management a challenge. In this review, we discuss the pathophysiology, clinical presentation, approaches to diagnosis, and conventional as well as targeted therapy of the most common ALPID conditions.

Regulatory T (Treg) cells that express the transcription factor forkhead box protein P3 (FOXP3) are naturally present in the immune system and have roles in the maintenance of immunological self-tolerance and immune system and tissue homeostasis. Treg cells suppress T cell activation, expansion and effector functions by various mechanisms, particularly by controlling the functions of antigen-presenting cells. They can also contribute to tissue repair by suppressing inflammation and facilitating tissue regeneration, for example, via the production of growth factors and the promotion of stem cell differentiation and proliferation. Monogenic anomalies of Treg cells and genetic variations of Treg cell functional molecules can cause or predispose patients to the development of autoimmune diseases and other inflammatory disorders, including kidney diseases. Treg cells can potentially be utilized or targeted to treat immunological diseases and establish transplantation tolerance, for example, by expanding natural Treg cells in vivo using IL-2 or small molecules or by expanding them in vitro for adoptive Treg cell therapy. Efforts are also being made to convert antigen-specific conventional T cells into Treg cells and to generate chimeric antigen receptor Treg cells from natural Treg cells for adoptive Treg cell therapies with the aim of achieving antigen-specific immune suppression and tolerance in the clinic.

In the last 20 years, discoveries about the autoimmune regulator (AIRE) protein and its critical role in immune tolerance have provided fundamental insights into understanding the molecular basis of autoimmunity. This review provides a comprehensive overview of the effect of AIRE on immunological tolerance and the characteristics of autoimmune diseases in Autoimmune Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy (APECED), which is caused by biallelic AIRE mutations. A better understanding of the immunological mechanisms of AIRE deficiency may enlighten immune tolerance mechanisms and new diagnostic and treatment strategies for autoimmune diseases. Considering that not all clinical features of APECED are present in a given follow-up period, the diagnosis is not easy in a patient at the first visit. Longer follow-up and a multidisciplinary approach are essential for diagnosis. It is challenging to prevent endocrine and other organ damage compared with other diseases associated with multiple autoimmunities, such as FOXP3, LRBA, and CTLA4 deficiencies. Unfortunately, no curative therapy like haematopoietic stem cell transplantation or specific immunomodulation is present that is successful in the treatment.

Type 1 classical dendritic cells (cDC1s) have emerged as the major antigen-presenting cell performing cross-presentation (XP) in vivo, but the antigen-processing pathway in this cell remains obscure. Two competing models for in vivo XP of cell-associated antigens by cDC1 include a vacuolar pathway and cytosolic pathway. A vacuolar pathway relies on directing antigens captured in vesicles toward a class I major histocompatibility complex loading compartment independently of cytosolic entry. Alternate proposals invoke phagosomal rupture, either constitutive or triggered by spleen tyrosine kinase (SYK) signaling in response to C-type lectin domain family 9 member A (CLEC9A) engagement, that releases antigens into the cytosol for proteasomal degradation. The Beige and Chediak-Higashi (BEACH) protein WD repeat- and FYVE domain-containing protein 4 (WDFY4) is strictly required for XP of cell-associated antigens in vivo. However, the cellular mechanism for WDFY4 activity remains unknown and its requirement in XP in vivo is currently indifferent regarding the vacuolar versus cytosolic pathways. Here, we review the current status of these models and discuss the need for future investigation.

LPS-responsive beige-like anchor protein (LRBA) deficiency is a primary immunodeficiency disease (PID) characterized by a regulatory T cell defect resulting in immune dysregulation and autoimmunity. We present two siblings born to consanguineous parents of North African descent with LRBA deficiency and central nervous system (CNS) manifestations. As no concise overview of these manifestations is available in literature, we compared our patient's presentation with a reviewed synthesis of the available literature.

The younger brother presented with enteropathy at age 1.5 years, and subsequently developed Evans syndrome and diabetes mellitus. These autoimmune manifestations led to the genetic diagnosis of LRBA deficiency through whole exome sequencing with PID gene panel. At 11 years old, he had two tonic-clonic seizures. Brain MRI showed multiple FLAIR-hyperintense lesions and a T2-hyperintense lesion of the cervical medulla.  His sister presented with immune cytopenia at age 9 years, and developed diffuse lymphadenopathy and interstitial lung disease. Genetic testing confirmed the same mutation as her brother. At age 13 years, a brain MRI showed multiple T2-FLAIR-hyperintense lesions. She received an allogeneic hematopoietic stem cell transplantation (allo-HSCT) 3 months later. Follow-up MRI showed regression of these lesions.

Neurological disease is documented in up to 25% of patients with LRBA deficiency. Manifestations range from cerebral granulomas to acute disseminating encephalomyelitis, but detailed descriptions of neurological and imaging phenotypes are lacking. LRBA deficiency amongst other PIDs should be part of the differential diagnosis in patients with inflammatory brain lesions. We strongly advocate for a more detailed description of CNS manifestations in patients with LRBA deficiency, when possible with MR imaging. This will aid clinical decision concerning both anti-infectious and anti-inflammatory therapy and in considering the indication for allo-HSCT.

Type 1 diabetes (T1D) is a polygenic disease and does not follow a mendelian pattern. Inborn errors of immunity (IEIs), on the other hand, are caused by damaging germline variants, suggesting that T1D and IEIs have nothing in common. Some IEIs, resulting from mutations in genes regulating regulatory T-cell homeostasis, are associated with elevated incidence of T1D. The genetic spectrum of IEIs is gradually being unraveled; consequently, molecular pathways underlying human monogenic autoimmunity are being identified. There is an appreciable overlap between some of these pathways and the genetic variants that determine T1D susceptibility, suggesting that after all, IEI and T1D are 2 sides of the same coin. The study of monogenic IEIs with a variable incidence of T1D has the potential to provide crucial insights into the mechanisms leading to T1D. These insights contribute to the definition of T1D endotypes and explain disease heterogeneity. In this review, we discuss the interconnected pathogenic pathways of autoimmunity, β-cell function, and primary immunodeficiency. We also examine the role of environmental factors in disease penetrance as well as the circumstantial evidence of IEI drugs in preventing and curing T1D in individuals with IEIs, suggesting the repositioning of these drugs also for T1D therapy.

The understanding of common variable immunodeficiency disorders (CVID) is in evolution. CVID was previously a diagnosis of exclusion. New diagnostic criteria have allowed the disorder to be identified with greater precision. With the advent of next-generation sequencing (NGS), it has become apparent that an increasing number of patients with a CVID phenotype have a causative genetic variant. If a pathogenic variant is identified, these patients are removed from the overarching diagnosis of CVID and are deemed to have a CVID-like disorder. In populations where consanguinity is more prevalent, the majority of patients with severe primary hypogammaglobulinemia will have an underlying inborn error of immunity, usually an early-onset autosomal recessive disorder. In nonconsanguineous societies, pathogenic variants are identified in approximately 20% to 30% of patients. These are often autosomal dominant mutations with variable penetrance and expressivity. To add to the complexity of CVID and CVID-like disorders, some genetic variants such as those in TNFSF13B (transmembrane activator calcium modulator cyclophilin ligand interactor) predispose to, or enhance, disease severity. These variants are not causative but can have epistatic (synergistic) interactions with more deleterious mutations to worsen disease severity. This review is a description of the current understanding of genes associated with CVID and CVID-like disorders. This information will assist clinicians in interpreting NGS reports when investigating the genetic basis of disease in patients with a CVID phenotype.