Expanding the PRAAS spectrum: De novo mutations of immunoproteasome subunit ß-type 10 in six infants with SCID-Omenn syndrome.

Mutations in proteasome ß-subunits or their chaperone and regulatory proteins are associated with proteasome-associated autoinflammatory disorders (PRAAS). We studied six unrelated infants with three de novo heterozygous missense variants in PSMB10, encoding the proteasome ß2i-subunit. Individuals presented with T-B-NK± severe combined immunodeficiency (SCID) and clinical features suggestive of Omenn syndrome, including diarrhea, alopecia, and desquamating erythematous rash. Remaining T cells had limited T cell receptor repertoires, a skewed memory phenotype, and an elevated CD4/CD8 ratio. Bone marrow examination indicated severely impaired B cell maturation with limited V(D)J recombination. All infants received an allogeneic stem cell transplant and exhibited a variety of severe inflammatory complications thereafter, with 2 peri-transplant and 2 delayed deaths. The single long-term transplant survivor showed evidence for genetic rescue through revertant mosaicism overlapping the affected PSMB10 locus. The identified variants (c.166G>C [p.Asp56His] and c.601G>A/c.601G>C [p.Gly201Arg]) were predicted in silico to profoundly disrupt 20S immunoproteasome structure through impaired ß-ring/ß-ring interaction. Our identification of PSMB10 mutations as a cause of SCID-Omenn syndrome reinforces the connection between PRAAS-related diseases and SCID.

Investigating Concomitant RAG-2 and LRBA Mutations in SCID and Autoimmunity.

Inborn errors of immunity (IEI) are a large heterogenous group of diseases characterized by immunodeficiency, immune dysregulation, allergy, auto-inflammation and predisposition for malignancies. Most are inherited in an autosomal recessive trait. We studied a patient with severe combined immunodeficiency (SCID) and immune dysregulation who harbored two distinct bi-allelic IEI-associated genetic mutations. Clinical, immunological and genetic data were collected. Genetic investigation included whole exome sequencing on DNA extracted from skin fibroblasts. Family segregation was performed by Sanger sequencing. Immunological evaluation included absolute and functional evaluation of lymphocytes and chimerism analysis post hematopoietic stem cell transplantation (HSCT). Treg subsets, LRBA and CTLA4 expression levels were measured by flow-cytometric analysis. A nineteen-year-old female patient from a consanguine background underwent unconditioned matched sibling related HSCT during infancy due to clinical presentation of SCID with an Omenn phenotype. At that time her underlying genetic defect was not defined. Years after HSCT, severe auto-immune phenomena were noted, including a systemic lupus erythematosus-like syndrome and ophthalmic manifestations. Genetic evaluation revealed bi-allelic homozygous mutations in RAG-2 (c.685C>T, p.Arg229Trp) and a previously undescribed mutation in LRBA (c.3325G>T, p.Asp1109Tyr). LRBA and CTLA4 expression levels were normal, suggesting that the LRBA variant identified in these kindred is unlikely to be pathogenic. Multiple genetic defects causing complex IEIs may be identified in the same individual in highly consanguineous populations. Functional immunological testing is essential for evaluation of novel genetic variants.

Inadequate Activation of γδT- and B-cells in Patient with Wiskott-Aldrich Syndrome (WAS) Portrayed by TRG and IGH Repertoire Analyses.

Patients with Wiskott-Aldrich syndrome (WAS) harbor mutations in the WAS gene and suffer from immunodeficiency, microthrombocytopenia, and eczema. T-cells play an important role in immune response in the skin and the γδT-cells have an important role in skin homeostasis. Since WAS patients often present with eczema, we wanted to examine whether the T-cell receptor gamma (TRG) repertoire of the γδT-cells is affected in these patients. In addition, the immunoglobulin heavy chain (IGH) repertoire from genomic DNA of WAS patients was not yet studied. Thus, we sought to determine the effects that specific WAS mutations from our patients have in shaping the TRG and IGH immune repertoires. We collected clinical and genetic data on four WAS patients, each harboring a different mutation in the WAS gene. Using next-generation sequencing (NGS), we analyzed their TRG and IGH repertoires using genomic DNA isolated from their peripheral blood. We analyzed the TRG and IGH repertoire sequences to show repertoire restriction, clonal expansions, preferential utilization of specific V genes, and unique characteristics of the antigen binding region in WAS patients with eczema compared to healthy controls. Both the TRG and IGH repertoire showed diverse repertoire comparable to healthy controls on one the hand, and on the other hand, the IGH repertoire showed increased diversity, more evenly distributed repertoire and immaturity of the antigen binding region. Thus, we demonstrate by analyzing the repertoire based on genomic DNA, the various effect that WAS mutations have in shaping the TRG and IGH adaptive immune repertoires.

SLP76 Mutation Associated with Combined Immunodeficiency and EBV-Related Lymphoma.

Increased susceptibility to develop severe forms of Epstein-Barr virus (EBV) infection in early age is a significant hallmark of an underlying primary immunodeficiency (PID). Here, we present immunologic and genetic evaluations of a 3-year-old child who was born to first-cousins parents and presented with recurrent infections, failure to thrive, and severe EBV-related infection and proliferation. A diagnosis of diffuse large B cell lymphoma was made and the immunological workup was suggestive of T cell immunodeficiency. Unfortunately, the patient succumbed to EBV-related lymphoma. Whole-exome sequencing revealed a novel homozygous mutation, c.991del.C; p. Q331Sfs*6 in the SLP76 gene. The SLP76 protein, a TCR signaling molecule, was recently linked to a human disease of the immune system. In order to examine the effect of this new SLP76 mutation on T cell signaling, a SLP76-deficient Jurkat-derived T cell line was transduced either with wild-type (WT), or with the specific SLP76 mutant, or with a mock vector. Downstream TCR signaling events, including ERK1/2 phosphorylation, CD69 expression, and Ca2 + mobilization, were reduced in cells harboring the reported mutation, linking this novel mutation to the expected immunological outcome. SLP76 deficiency should be added to the growing list of monogenetic diseases that predispose affected individuals to acquire severe and uncontrolled EBV infections and to develop substantial complications. This case further links mutations in the SLP76 gene to a significant human immunodeficiency and extends its clinical phenotype.

Combined Immunodeficiency Caused by a Novel Nonsense Mutation in LCK.

Mutations affecting T-cell receptor (TCR) signaling typically cause combined immunodeficiency (CID) due to varying degrees of disturbed T-cell homeostasis and differentiation. Here, we describe two cousins with CID due to a novel nonsense mutation in LCK and investigate the effect of this novel nonsense mutation on TCR signaling, T-cell function, and differentiation. Patients underwent clinical, genetic, and immunological investigations. The effect was addressed in primary cells and LCK-deficient T-cell lines after expression of mutated LCK. RESULTS: Both patients primarily presented with infections in early infancy. The LCK mutation led to reduced expression of a truncated LCK protein lacking a substantial part of the kinase domain and two critical regulatory tyrosine residues. T cells were oligoclonal, and especially naïve CD4 and CD8 T-cell counts were reduced, but regulatory and memory including circulating follicular helper T cells were less severely affected. A diagnostic hallmark of this immunodeficiency is the reduced surface expression of CD4. Despite severely impaired TCR signaling mTOR activation was partially preserved in patients' T cells. LCK-deficient T-cell lines reconstituted with mutant LCK corroborated partially preserved signaling. Despite detectable differentiation of memory and effector T cells, their function was severely disturbed. NK cell cytotoxicity was unaffected. Residual TCR signaling in LCK deficiency allows for reduced, but detectable T-cell differentiation, while T-cell function is severely disturbed. Our findings expand the previous report on one single patient on the central role of LCK in human T-cell development and function.

Can T-cell and B-cell excision circles predict development of inhibitors in pediatric hemophilia A?

BACKGROUND: Hemophilia A (HA) therapy requires intravenous replacement infusions of factor (F) VIII concentrate. Inhibitors are high-affinity immunoglobulin G that are directed against FVIII and thereby render replacement therapy ineffective. This complication has significant prognostic implications. We aimed to examine the immune system involvement in inhibitor formation specifically T-cell excision circles (TRECs) and B-cell excision circles (KRECs), markers of new T and B cells, respectively, and examine them as surrogate markers for inhibitor formation. METHODS: Blood samples were collected from 35 children with severe HA. Children were divided into two groups: with FVIII inhibitors and without FVIII inhibitors. TRECs and KRECs were measured in peripheral blood. RESULTS: A total of 11 patients with inhibitors and 24 without were evaluated. Children with inhibitors had higher levels of TRECs however not statistically significant (p = 0.085). CjKREC levels were higher in the inhibitor patients (p = 0.003). Moreover, the sj/cjKREC ratio was lower in the inhibitor patients (p = 0.015). CONCLUSIONS: Our findings may add to the notion that inhibitor formation is attributed to humoral immunity due to peripheral B-cell expansion and loss of peripheral tolerance. Improved knowledge regarding the involvement of the immune system in the formation of FVIII inhibitors will enable better therapy tailoring in the era of non-replacement therapies. IMPACT: The etiology of FVIII inhibitor formation is multifactorial, in which the immune system plays a pivotal role. Our findings may add to the notion that inhibitor formation is attributed to humoral immunity due to peripheral B-cell expansion and production of antibodies against FVIII. Improved knowledge regarding the involvement of the immune system in the development of FVIII inhibitors will enable the identification of patients prone to inhibitor development and better therapy tailoring in the new era of non-replacement therapies.

Newborn screening for severe combined immunodeficiency and inborn errors of immunity.

PURPOSE OF REVIEW: Severe combined immune deficiency (SCID) is the most devastating genetic disease of the immune system with an unfavorable outcome unless diagnosed early in life. Newborn screening (NBS) programs play a crucial role in facilitating early diagnoses and timely interventions for affected infants. RECENT FINDINGS: SCID marked the pioneering inborn error of immunity (IEI) to undergo NBS, a milestone achieved 15 years ago through the enumeration of T-cell receptor excision circles (TRECs) extracted from Guthrie cards. This breakthrough has revolutionized our approach to SCID, enabling not only presymptomatic identification and prompt treatments (including hematopoietic stem cell transplantation), but also enhancing our comprehension of the global epidemiology of SCID. SUMMARY: NBS is continuing to evolve with the advent of novel diagnostic technologies and treatments. Following the successful implementation of SCID-NBS programs, a call for the early identification of additional IEIs is the next step, encompassing a broader spectrum of IEIs, facilitating early diagnoses, and preventing morbidity and mortality.

Genetic workup as a complementary tool for the diagnosis of primary complement component deficiencies: a multicenter experience.

Diagnosis of primary complement deficiencies requires a high index of suspicion. Thus, susceptible patients are often underdiagnosed and untreated. Here, we present a multicenter experience with two novel inborn errors of the classical complement system. This is a retrospective multicenter analysis of computerized medical records of children (<18 years) admitted in the period between 2012 and 2018 at Shaare Zedek Medical Center in Jerusalem and Edmond and Lily Safra Children's Hospital, Tel-Hashomer Medical Center, in Ramat Gan, Israel. Patients were genetically diagnosed by a complementary immune workup. We identified 5 patients (3 males) from four different families harboring two novel mutations in the complement components C6-C8. Genetic mutations were identified by whole-exome sequencing or by sequencing of the coding exons of a single gene based on the findings in the immune workup. Clinical manifestations consisted of meningitis with or without meningococcemia. The immune workup demonstrated nearly absent levels of CH50, compatible with a complement pathway defect. Diagnosis delay ranged between 0 and 30 years. CONCLUSION: Awareness of risk factors for primary complement deficiencies, even at the first infectious episode, should facilitate prompt immune and genetic workup, commencing diagnosis and proper treatment for the patient and family. WHAT IS KNOWN: • Deficiencies in the classical terminal complement components increase susceptibility to invasive meningococcal infections. • Recurrent meningococcal infections mandate a diagnostic workup of the complement system. WHAT IS NEW: • Genetic workup can be utilized for prompt diagnosis of complement deficiencies. • High rates of consanguinity, even in the presence of a single meningococcal infection, should promote immune and genetic workups.

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

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

CD137 deficiency causes immune dysregulation with predisposition to lymphomagenesis.

Dysregulated immune responses are essential underlying causes of a plethora of pathologies including cancer, autoimmunity, and immunodeficiency. We here investigated 4 patients from unrelated families presenting with immunodeficiency, autoimmunity, and malignancy. We identified 4 distinct homozygous mutations in TNFRSF9 encoding the tumor necrosis factor receptor superfamily member CD137/4-1BB, leading to reduced, or loss of, protein expression. Lymphocytic responses crucial for immune surveillance, including activation, proliferation, and differentiation, were impaired. Genetic reconstitution of CD137 reversed these defects. CD137 deficiency is a novel inborn error of human immunity characterized by lymphocytic defects with early-onset Epstein-Barr virus (EBV)-associated lymphoma. Our findings elucidate a functional role and relevance of CD137 in human immune homeostasis and antitumor responses.

Immune and TRG repertoire signature of the thymus in Down syndrome patients.

BACKGROUND: Patients with Down syndrome (DS) are at increased risk for infections and autoimmune disorders. Although several immunological abnormalities were previously found, differences in T cell receptor repertoire have never been shown. Thus we compared the T cell receptor gamma (TRG) repertoire in DS and non-syndromic pediatric patients by next-generation sequencing, in addition to other immunological markers. METHODS: Genomic DNA was extracted from thymuses of pediatric patients who underwent heart surgery, where six were with DS and six were non-syndromic patients. Peripheral blood counts, T cell subpopulations, thymus TCR excision circles (TRECs), spectratyping, and next-generation sequencing for TRG were analyzed. RESULTS: The mean age of the patients was 7 months and the mean lymphocyte count was slightly lower in patients with DS, whereas thymus TREC results were similar to non-syndromic patients (p = 0.197). The TRG repertoire analysis showed that patients with DS had a significantly larger number of unique TRG sequences, together with decreased clonal expansion. Lastly, the V and J gene usages in the thymus were similar in DS and non-syndromic patients. CONCLUSIONS: Patients with DS showed increased TRG repertoire diversity with decreased clonal expansion compared to non-syndromic patients. IMPACT: Alterations in T cell receptor gamma repertoire were found in patients with Down syndrome using next-generation sequencing (NGS) technique. Patients showed increased repertoire diversity and decreased clonal expansion compared to controls. These findings add to previous reports on abnormalities of other immune system components in patients with Down syndrome. NGS technique may point out differences not seen by previous methods. Repertoire abnormalities may contribute to those patients' predisposition to infections and autoimmune diseases.

Atypical immune phenotype in severe combined immunodeficiency patients with novel mutations in IL2RG and JAK3.

Mutations in the common gamma chain of the interleukin 2 receptor (IL2RG) or the associated downstream signaling enzyme Janus kinase 3 (JAK3) genes are typically characterized by a T cell-negative, B cell-positive, natural killer (NK) cell-negative (T-B+NK-) severe combined immunodeficiency (SCID) immune phenotype. We report clinical course, immunological, genetic and proteomic work-up of two patients with different novel mutations in the IL-2-JAK3 pathway with a rare atypical presentation of T-B+NK- SCID. Lymphocyte subpopulation revealed significant T cells lymphopenia, normal B cells, and NK cells counts (T-B+NK+SCID). Despite the presence of B cells, IgG levels were low and IgA and IgM levels were undetectable. T-cell proliferation in response to mitogens in patient 1 was very low and T-cell receptor V-beta chain repertoire in patient 2 was polyclonal. Whole-exome sequencing revealed novel mutations in both patients (patient 1-c.923delC frame-shift mutation in the IL2RG gene, patient 2-c.G172A a homozygous missense mutation in the JAK3 gene). Bioinformatic analysis of the JAK3 mutation indicated deleterious effect and 3D protein modeling located the mutation to a surface exposed alpha-helix structure. Our findings help to link between genotype and phenotype, which is a key factor for the diagnosis and treatment of SCID patients.

Whole exome sequencing (WES) approach for diagnosing primary immunodeficiencies (PIDs) in a highly consanguineous community.

Primary immunodeficiencies (PIDs) are a heterogeneous group of monogenic inborn errors of immunity. The genetic causes of these diseases can be identified using whole exome sequencing (WES). Here, DNA samples from 106 patients with a clinical suspicion of PID were subjected to WES in order to test the diagnostic yield of this test in a highly consanguineous community. A likely genetic diagnosis was achieved in 70% of patients. Several factors were considered to possibly influence the diagnostic rate of WES among our cohort including early age, presence of consanguinity, family history suggestive of PID, the number of family members who underwent WES and the clinical phenotype of the patient. The highest diagnostic rate was in patients with combined immunodeficiency or with a syndrome. Notably, WES findings altered the clinical management in 39% (41/106) of patients in our cohort. Our findings support the use of WES as an important diagnostic tool in patients with suspected PID, especially in highly consanguineous communities.

A Large Cohort of RAG1/2-Deficient SCID Patients-Clinical, Immunological, and Prognostic Analysis.

INTRODUCTION: Severe combined immunodeficiency (SCID) is a fatal disorder resulting from various genetic defects. In the Middle East, where consanguineous marriage is prevalent, autosomal recessive mutations in recombination-activating genes (RAG) are a leading cause of SCID. We present a large cohort of SCID patients due to RAG1 or RAG2 mutations. METHODS: Twenty-six patients with RAG1 or RAG2 deficiency, diagnosed at Sheba Medical Center, were retrospectively investigated. Clinical presentation, immunologic phenotype, genetic analysis, treatment, and outcome were analyzed. RESULTS: Majority of patients were referred from the Palestinian Authority. Most patients were males of Muslim Arab descent, 77% were born to consanguineous parents, and 65% had family history of immunodeficiency. Nearly all patients suffered from various infections before turning 2 months old, eight patients (31%) presented with Omenn and Omenn-like syndrome, and three patients (11%) had maternal engraftment. Notably, seven patients (27%) suffered from vaccine-derived infections, including a rare case of measles encephalitis. Nineteen patients underwent hematopoietic stem cell transplantation (HSCT) at a median age of 6 months, with a successful outcome for 72% of them. Genetic analysis revealed 11 different mutations (7 RAG2, 4 RAG1), two of them novel. CONCLUSIONS: Consanguineous marriages account for a genetic "founder effect." SCID is a pediatric emergency that dictates immediate precautions and curative treatment with HSCT. Due to lack of newborn screening for SCID within the Palestinian population, most patients in this cohort were diagnosed upon clinical symptoms, which led to a delayed diagnosis, harmful administration of contra-indicated live vaccines, delay in HSCT, and poor outcome.

Congenital neutropenia with variable clinical presentation in novel mutation of the SRP54 gene.

BACKGROUND: The SRP54 (signal recognition protein 54) is a conserved component of the ribonucleoprotein complex that mediates cotranslational targeting and translocation of proteins to the endoplasmic reticulum. In 2017, mutations in the gene have been described as a cause of congenital neutropenia with or without pancreatic insufficiency, and since then, only limited cases were added to the literature. METHODS: Two patients with neutropenia underwent hematological, immunological, and genetic work-up, including lymphocyte phenotyping, immunoglobulins, and complement levels, antineutrophil and antinuclear antibodies, bone marrow FISH panel for myelodysplastic syndrome, whole-exome sequencing, and in silico proteomic analysis. RESULTS: Clinical findings in the two families revealed a wide spectrum of immunological and clinical manifestations, ranging from mild asymptomatic neutropenia during febrile illnesses to severe neutropenia and life-threatening infection requiring leg amputation. Immunological and hematological work-up showed isolated neutropenia with normal lymphocyte subpopulations, immunoglobulin and complement levels, and negative autoimmune tests. Bone marrow aspirations showed variability ranging from normal myelopoiesis to myeloid maturation arrest at the promyelocytic stage, with normal FISH panel for myelodysplastic syndrome. Genetic analysis identified a novel, de novo, in-frame deletion in the SRP54 gene, c.342-344delAAC, p.T115del. In silico proteomic analysis suggested impaired SRP54 protein function due to reduced GTP activity and stability. CONCLUSIONS: We describe congenital neutropenia with variable clinical presentation in novel mutation of the SRP54 gene.

Novel MALT1 Mutation Linked to Immunodeficiency, Immune Dysregulation, and an Abnormal T Cell Receptor Repertoire.

MALT1 (mucosa-associated lymphoid tissue lymphoma-translocation gene 1) is an intracellular signaling protein that activates NFκB and is crucial for both the adaptive and innate immune responses. Only 6 patients with immune deficiencies secondary to inherited mutations in the MALT1 gene have been described. PURPOSE: To provide clinical and immunological insights from 2 patients diagnosed with MALT1 immunodeficiency syndrome due to a novel MALT1 mutation. METHODS: Two cousins with suspected combined immunodeficiency underwent immunological and genetic work-up, including lymphocyte phenotyping, lymphocyte activation by mitogen stimulation, and next-generation sequencing (NGS) of T cell receptor gamma chain (TRG) repertoire. Whole exome sequencing was performed to identify the underlying genetic defect. RESULTS: Clinical findings included recurrent infections, failure to thrive, lymphadenopathy, dermatitis, and autoimmunity. Immune work-up revealed lymphocytosis, low to normal levels of immunoglobulins, absence of regulatory T cells, and low Th17 cells. A normal proliferative response was induced by phytohemagglutinin and IL-2 but was diminished with anti-CD3. TRG repertoire was diverse with a clonal expansion pattern. Genetic analysis identified a novel autosomal recessive homozygous c.1799T>A; p. I600N missense mutation in MALT1. MALT1 protein expression was markedly reduced, and in vitro IL-2 production and NFκB signaling pathway were significantly impaired. CONCLUSIONS: Two patients harboring a novel MALT1 mutation presented with signs of immune deficiency and dysregulation and were found to have an abnormal T cell receptor repertoire. These findings reinforce the link between MALT1 deficiency and combined immunodeficiency. Early diagnosis is crucial, and curative treatment by hematopoietic stem cell transplantation may be warranted.

Fetuin-A deficiency is associated with infantile cortical hyperostosis (Caffey disease).

BACKGROUND: Infantile cortical hyperostosis (ICH)/Caffey disease is an inflammatory collagenopathy of infancy, manifested by subperiosteal bone hyperplasia. Genetically, ICH was linked with heterozygosity for an R836C mutation in the COL1A1 gene. Although an autosomal-recessive trait is also suspected, it has not been proven thus far. METHODS: A case of an infant male born to consanguineous parents is reported, presenting with classical findings, course, and clinical outcome of ICH. Whole-exome sequencing (WES) was performed in order to identify a possible underlying genetic defect. RESULTS: WES analysis revealed a novel homozygous nonsense mutation in lysine 2 of fetuin-A, encoded by the ALPHA-2-HS-GLYCOPROTEIN (AHSG) gene (c.A4T; p.K2X). Fetuin-A is an important regulator of bone remodeling and an inhibitor of ectopic mineralization. By enzyme-linked immunosorbent assay (ELISA), we show a complete deficiency of this protein in the patient's serum, compared to controls. CONCLUSION: A novel homozygous nonsense mutation in AHSG gene has been found in ICH patient with a typical phenotype, resulting in fetuin-A deficiency. This finding postulates an autosomal-recessive mode of inheritance in ICH, which, unlike the autosomal-dominant inheritance associated with COL1A1, is associated with AHSG and fetuin-A deficiency.

Neutrophil Functions in Immunodeficiency Due to DOCK8 Deficiency.

Neutrophil chemotactic defects have been reported previously in patients with hyper-IgE syndrome. Bi-allelic mutations in dedicator of cytokinesis 8 (DOCK8) gene usually cause an autosomal recessive hyper-IgE syndrome phenotype. Data are lacking about expression of DOCK8 protein in neutrophils or the possible role of DOCK8 in neutrophil function. We sought to determine if DOCK8 protein is expressed in neutrophils and if DOCK8 plays a role in neutrophil function. The expression of DOCK8 protein was assessed in neutrophils from healthy volunteers with and without activators. Neutrophil chemotaxis, phagocytosis and superoxide generation were studied in neutrophils from DOCK8-deficient patients compared to neutrophils from healthy controls before and after stimulation with activators: phorbol 12-myristate 13-acetate (PMA) or N-Formylmethionyl-leucyl-phenylalanine (fMLP). DOCK8 protein is expressed in resting neutrophils from healthy controls, with a significant increase in DOCK8 expression after stimulation. Neutrophil functions were assessed in 6 DOCK8-deficient patients. All patients had the same non-sense mutation (c.C5134A, p.S1711X). Normal chemotaxis was recorded in 4/6 patients while a mild to moderate chemotaxis defect was recorded in 2/6. Superoxide generation was mainly normal in neutrophils from all six patients and phagocytosis was normal in five patients tested. We conclude that DOCK8 protein is expressed in resting human neutrophils and DOCK8 expression is increased after stimulation with either PMA or fMLP. Most patients with a disease-causing mutation in DOCK8 have normal neutrophil functions, while a minority showed a mild to moderate chemotactic defect.

Disruption of Thrombocyte and T Lymphocyte Development by a Mutation in ARPC1B.

Regulation of the actin cytoskeleton is crucial for normal development and function of the immune system, as evidenced by the severe immune abnormalities exhibited by patients bearing inactivating mutations in the Wiskott-Aldrich syndrome protein (WASP), a key regulator of actin dynamics. WASP exerts its effects on actin dynamics through a multisubunit complex termed Arp2/3. Despite the critical role played by Arp2/3 as an effector of WASP-mediated control over actin polymerization, mutations in protein components of the Arp2/3 complex had not previously been identified as a cause of immunodeficiency. Here, we describe two brothers with hematopoietic and immunologic symptoms reminiscent of Wiskott-Aldrich syndrome (WAS). However, these patients lacked mutations in any of the genes previously associated with WAS. Whole-exome sequencing revealed a homozygous 2 bp deletion, n.c.G623DEL-TC (p.V208VfsX20), in Arp2/3 complex component ARPC1B that causes a frame shift resulting in premature termination. Modeling of the disease in zebrafish revealed that ARPC1B plays a critical role in supporting T cell and thrombocyte development. Moreover, the defects in development caused by ARPC1B loss could be rescued by the intact human ARPC1B ortholog, but not by the p.V208VfsX20 variant identified in the patients. Moreover, we found that the expression of ARPC1B is restricted to hematopoietic cells, potentially explaining why a mutation in ARPC1B has now been observed as a cause of WAS, whereas mutations in other, more widely expressed, components of the Arp2/3 complex have not been observed.

Correction to: Novel Mutations in RASGRP1 Are Associated with Immunodeficiency, Immune Dysregulation, and EBV-Induced Lymphoma.

The original version of this article unfortunately contained mistakes in Author's name, in Table 1 and in result section.