Case report and review of the literature: immune dysregulation in a large familial cohort due to a novel pathogenic RELA variant.

OBJECTIVE: To explore and define the molecular cause(s) of a multi-generational kindred affected by Bechet's-like mucocutaneous ulcerations and immune dysregulation. METHODS: Whole genome sequencing and confirmatory Sanger sequencing were performed. Components of the NFκB pathway were quantified by immunoblotting, and function was assessed by cytokine production (IL-6, TNF-α, IL-1ß) after lipopolysaccharide (LPS) stimulation. Detailed immunophenotyping of T-cell and B-cell subsets was performed in four patients from this cohort. RESULTS: A novel variant in the RELA gene, p. Tyr349LeufsTer13, was identified. This variant results in premature truncation of the protein before the serine (S) 536 residue, a key phosphorylation site, resulting in enhanced degradation of the p65 protein. Immunoblotting revealed significantly decreased phosphorylated [p]p65 and pIκBα. The decrease in [p]p65 may suggest reduced heterodimer formation between p50/p65 (NFκB1/RelA). Immunophenotyping revealed decreased naïve T cells, increased memory T cells, and expanded senescent T-cell populations in one patient (P1). P1 also had substantially higher IL-6 and TNF-α levels post-stimulation compared with the other three patients. CONCLUSION: Family members with this novel RELA variant have a clinical phenotype similar to other reported RELA cases with predominant chronic mucocutaneous ulceration; however, the clinical phenotype broadens to include Behçet's syndrome and IBD. Here we describe the clinical, immunological and genetic evaluation of a large kindred to further expand identification of patients with autosomal dominant RELA deficiency, facilitating earlier diagnosis and intervention. The functional impairment of the canonical NFκB pathway suggests that this variant is causal for the clinical phenotype in these patients.

Infectious Complications Predict Premature CD8+ T-cell Senescence in CD40 Ligand-Deficient Patients.

PURPOSE: CD40 ligand (CD40L)-deficient patients display increased susceptibilities to infections that can be mitigated with effective prophylactic strategies including immunoglobulin G (IgG) replacement and prophylactic antibiotics. CD8+ T-cell senescence has been described in CD40L deficiency, but it is unclear if this is an intrinsic feature of the disease or secondary to infectious exposures. To address this question, we assessed CD8+ T-cell senescence and its relationship to clinical histories, including prophylaxis adherence and infections, in CD40L-deficient patients. METHODS: Peripheral CD8+ T-cells from seven CD40L-deficient patients and healthy controls (HCs) were assessed for senescent features using T-cell receptor excision circle (TREC) analysis, flow cytometry, cytometry by time of flight (CyTOF) and in vitro functional determinations including CMV-specific proliferation and cytokine release assays. RESULTS: Three patients (5, 28, and 34 years old) who were poorly adherent to immunoglobulin G replacement and Pneumocystis jirovecii pneumonia (PJP) prophylaxis and/or experienced multiple childhood pneumonias (patient group 1) had an expansion of effector memory CD8+ T-cells with the senescent phenotype when compared to HCs. Such changes were not observed in the patient group 2 (four patients, 16, 22, 24, and 33 years old) who were life-long adherents to prophylaxis and experienced few infectious complications. CyTOF analysis of CD8+ T-cells from the 5-year-old patient and older adult HCs showed similar expression patterns of senescence-associated molecules. CONCLUSIONS: Our findings support that recurrent infections and non-adherence to prophylaxis promote early CD8+ T-cell senescence in CD40L deficiency. Premature senescence may increase malignant susceptibilities and further exacerbate infectious risk in CD40L-deficient patients.

Differing Performance of the Warning Signs for Immunodeficiency in the Diagnosis of Pediatric Versus Adult Patients in a Two-Center Tertiary Referral Population.

PURPOSE: Primary immunodeficiency (PID) represents disorders with a spectrum of clinical presentations. The medical community seeks clinical features to prompt evaluation for immunodeficiency given improved prognosis with early identification. We hoped to identify clinical characteristics that would improve the diagnostic accuracy of the widely disseminated Jeffrey Modell Foundation warning signs for immunodeficiency. METHODS: We performed a retrospective chart review in a two-center North American cohort of patients with PID. Charts of 137 pediatric and 400 adult patients with PID were evaluated for the presence of these warning signs and compared to controls with normal preliminary biochemical immune evaluation. RESULTS: Fewer than 45% of adults with PID presented with ≥ 2 warning signs, while diagnostic utility was improved in the pediatric population where the warning signs were found to be 64% sensitive. The warning signs found in a significantly increased proportion compared to controls differed for pediatric PID patients (recurrent pneumonia (OR 2.9, p < 0.001), failure to thrive (OR 2.1, p < 0.001), need for IV antibiotics (OR 2.1, p < 0.001), serious bacterial infection (OR 4.8, p < 0.001), recurrent otitis media (OR 1.5, p = 0.027)), versus adult PID patients (recurrent otitis media (OR 2.9, p < 0.001), recurrent sinusitis (OR 2.1, p < 0.001), diarrhea with weight loss (OR 2.2, p < 0.001), recurrent viral infection (OR 3.3 p < 0.001)). In evaluation for additional criteria to promote identification of immunodeficiency, linear regression models showed slightly improved diagnostic accuracy of the warning signs with the addition of autoimmunity in our pediatric PID cohort (8.7% v 2.8%, p < 0.001, ROC 0.58). Adult PID patients demonstrated atopy more frequently than controls (48.0% vs 40.3%, p = 0.011), while atopy was found to have a negative association with the presence of PID in the pediatric age group (OR 0.3, p < 0.01). No improvement in diagnostic accuracy of the warning signs was found with the addition of allergic disease, autoimmunity, or malignant and benign proliferative disease in the adult cohort. CONCLUSIONS: We demonstrate poor diagnostic performance of warning signs for immunodeficiency in patients with PID in a retrospective chart review. Divergent warning signs of statistically significant diagnostic utility were found in pediatric versus adult patients. We suggest education of physicians on differing presentations of possible immunodeficiency between age groups, and expansion of the warning signs to include non-infectious comorbidities such as autoimmunity in pediatric patients.

Role of Allogeneic Hematopoietic Stem Cell Transplant for Chronic Granulomatous Disease (CGD): a Report of the United States Immunodeficiency Network.

PURPOSE: Chronic granulomatous disease (CGD) is a primary immunodeficiency for which allogeneic hematopoietic stem cell transplant (HSCT) offers potential cure. Direct comparison of HSCT to non-HSCT management in the North American population was performed to identify clinical factors associated with overall survival (OS) and transplant-related survival (TRS). METHODS: Retrospective review of CGD subjects enrolled in the United States Immunodeficiency Network. Survival was estimated by the Kaplan-Meier method and modeled by proportional hazards regression. RESULTS: We identified 507 patients (66% CYBB mutants) diagnosed in 1953-2016. Fifty underwent allogeneic HSCT. Median follow-up was 9.1 years after diagnosis (0-45.8 years). OS was negatively associated with CYBB mutation (HR = 6.25; p = 0.034) and not associated with HSCT (88% v. 85% ± HCT) (HR = 1.26; p = 0.65). Transplant at ≤ 14 years old was associated with improved TRS (93% v. 82% at T + 60 months) (HR = - 4.51; p = 0.035). Patients transplanted before 15 years old had fewer severe infections pre-HSCT (mean 0.95 v. 2.13; p = 0.047). No mortality was reported in patients receiving stem cells from matched siblings. Infection incidence declined post-HSCT in subjects with greater than or equal to four infections pre-HSCT (p = 0.0010). Compared to non-HSCT patients ≥ 15 years old, post-transplant survivors had higher mean performance score (93.2 v. 85.9; p = 0.0039) and lower frequency of disability (11% v. 52%; p = 0.014). CONCLUSION: Allogeneic HSCT was associated with reduced infection incidence and improved functional performance, but not with a change in overall survival. Transplant-related survival was elevated in patients undergoing HSCT before 15 years old. Consider HSCT prior to late adolescence in patients with severely diminished reactive oxygen intermediate synthesis, particularly if a matched sibling is available.

Delayed and Severe Reactions to Baked Egg and Baked Milk Challenges.

BACKGROUND: Although many egg- and milk-allergic children tolerate baked egg (BE) and baked milk (BM), reactions elicited by these oral food challenges (OFCs) can be severe. Previous studies comparing BE and BM reactions to other OFC reactions are limited. OBJECTIVE: To compare the clinical features of reactions to BE and BM with other OFCs to promote challenge safety. METHODS: A retrospective review of OFCs eliciting objective reactions to BE, BM, lightly cooked egg (CE), fresh cow's milk (CM), peanut (PN), and tree nuts (TN) which were performed at a tertiary referral center from June 1, 2017, to June 1, 2019. RESULTS: We identified 174 OFCs conducted in 158 subjects, age 6 to 187 months (34 BE, 19 BM, 14 CE, 25 CM, 52 PN, and 30 TN). TN reactors were older than BE (P = .049) and PN (P < .01) reactors and had a higher frequency of persistent asthma than PN-reactive subjects (P = .02). Mucocutaneous symptoms occurred less frequently during BE (56%) and BM (42%) OFCs compared with other OFCs (P < .05). Lower respiratory tract reactions were increased during BM (37%) versus BE (12%), CM (8%), and PN (12%) OFCs (P < .05). Epinephrine was administered to more BE (44%) and TN (50%) than PN (17%) OFCs (P < .01). New reaction manifestations developed 60 minutes or later after OFC termination during 29% BE and 21% BM versus 0% PN OFCs (P < .05). One-third of anaphylactic reactions to BE began more than 60 minutes after OFC termination. CONCLUSIONS: BE and BM challenge reactions differed phenotypically from other OFC reactions, including significantly increased frequencies of new symptoms beginning 60 minutes or later after challenge termination. Consider amended dose-escalation and prolonged observation after BE and BM OFCs.

Delayed-Onset ADA1 (ADA) Deficiency Not Detected by TREC Screen.

A 9-month-old boy presented to a community pediatrician with a recent history of failure to thrive. Workup revealed neutropenia and lymphopenia. Subsequent admission for fever and pneumonia revealed an absolute neutrophil count of 860 and absolute lymphocyte count of 214. Lymphopenia affected all lymphocyte subsets and his naïve and memory CD4+ T-cell ratio was inverted for age. Immunoglobulin levels were normal for age, and tetanus and diphtheria antibody titers were protective. The profound lymphopenia raised suspicion for severe combined immunodeficiency (SCID), despite a normal newborn screening by T-cell receptor excision circle analysis. He did not have a previous history of recurrent fevers or infections, had attended day care, and had received all age-appropriate vaccines. He subsequently was diagnosed with Pneumocystis jirovecii pneumonia, adenovirus upper respiratory infection, and rotaviral diarrhea. An enzyme assay revealed absent adenosine deaminase (ADA) activity and elevated erythrocyte deoxyadenosine nucleotides. With genetic sequencing, 2 pathogenic variants in the ADA gene were confirmed. Acute management of ADA-SCID is aimed at restoration of enzyme activity, followed by curative therapy. The patient is currently on immunoglobulin therapy and recombinant ADA (Revcovi), with an excellent immune response, while awaiting sibling hematopoietic cell transplant from a matched sibling. Hypomorphic ADA variants can present with delayed-onset SCID, and some of these patients are missed by SCID newborn screening. A careful review of a complete blood cell count might offer clues and promote confirmatory diagnostic investigation.

A Novel Pathogenic Variant in CARMIL2 (RLTPR) Causing CARMIL2 Deficiency and EBV-Associated Smooth Muscle Tumors.

CARMIL2 deficiency is a rare combined immunodeficiency (CID) characterized by defective CD28-mediated T cell co-stimulation, altered cytoskeletal dynamics, and susceptibility to Epstein Barr Virus smooth muscle tumors (EBV-SMTs). Case reports associated with EBV-SMTs are limited. We describe herein a novel homozygous CARMIL2 variant (c.1364_1393del) in two Saudi Arabian male siblings born to consanguineous parents who developed EBV-SMTs. CARMIL2 protein expression was significantly reduced in CD4+ T cells and CD8+ T cells. T cell proliferation on stimulation with soluble (s) anti-CD3 or (s) anti-CD3 plus anti-CD28 antibodies was close to absent in the proband, confirming altered CD28-mediated co-signaling. CD28 expression was substantially reduced in the proband's T cells, and was diminished to a lesser degree in the T cells of the younger sibling, who has a milder clinical phenotype. Defects in both T and B cell compartments were observed, including absent central memory CD8+ T cells, and decreased frequencies of total and class-switched memory B cells. FOXP3+ regulatory T cells (Treg) were also quantitatively decreased, and furthermore CD25 expression within the Treg subset was substantially reduced. These data confirm the pathogenicity of this novel loss-of-function (LOF) variant in CARMIL2 and expand the genotypic and phenotypic spectrum of CIDs associated with EBV-SMTs.