Heterozygous FOXN1 Variants Cause Low TRECs and Severe T Cell Lymphopenia, Revealing a Crucial Role of FOXN1 in Supporting Early Thymopoiesis.

FOXN1 is the master regulatory gene of thymic epithelium development. FOXN1 deficiency leads to thymic aplasia, alopecia, and nail dystrophy, accounting for the nude/severe combined immunodeficiency (nu/SCID) phenotype in humans and mice. We identified several newborns with low levels of T cell receptor excision circles (TRECs) and T cell lymphopenia at birth, who carried heterozygous loss-of-function FOXN1 variants. Longitudinal analysis showed persistent T cell lymphopenia during infancy, often associated with nail dystrophy. Adult individuals with heterozygous FOXN1 variants had in most cases normal CD4+ but lower than normal CD8+ cell counts. We hypothesized a FOXN1 gene dosage effect on the function of thymic epithelial cells (TECs) and thymopoiesis and postulated that these effects would be more prominent early in life. To test this hypothesis, we analyzed TEC subset frequency and phenotype, early thymic progenitor (ETP) cell count, and expression of FOXN1 target genes (Ccl25, Cxcl12, Dll4, Scf, Psmb11, Prss16, and Cd83) in Foxn1nu/+ (nu/+) mice and age-matched wild-type (+/+) littermate controls. Both the frequency and the absolute count of ETP were significantly reduced in nu/+ mice up to 3 weeks of age. Analysis of the TEC compartment showed reduced expression of FOXN1 target genes and delayed maturation of the medullary TEC compartment in nu/+ mice. These observations establish a FOXN1 gene dosage effect on thymic function and identify FOXN1 haploinsufficiency as an important genetic determinant of T cell lymphopenia at birth.

Dominant-negative loss of function arises from a second, more frequent variant within the SAND domain of autoimmune regulator (AIRE).

A genetic variant in the SAND domain of autoimmune regulator (AIRE), R247C, was identified in a patient with type I diabetes mellitus (T1DM) and his mother with rheumatoid arthritis. In vitro, the variant dominantly inhibited AIRE; however, typical features of Autoimmune Polyendocrinopathy Candidiasis and Ectodermal Dysplasia (APECED) were not seen in the subjects. Rather, early manifestation of autoimmunity appeared to be dependent on additional genetic factors. On a population level, diverse variants were identified in this region. Surprisingly, many likely pathogenic variants were seen disproportionately in Africans when compared to Europeans, reinforcing the importance of these variants in altering the immune repertoire. In light of these findings, we propose that R247C and other variants within the SAND-domain alter protein function in a dominant fashion and hold potential as drivers of autoimmunity.

Long-term outcomes of 176 patients with X-linked hyper-IgM syndrome treated with or without hematopoietic cell transplantation.

BACKGROUND: X-linked hyper-IgM syndrome (XHIGM) is a primary immunodeficiency with high morbidity and mortality compared with those seen in healthy subjects. Hematopoietic cell transplantation (HCT) has been considered a curative therapy, but the procedure has inherent complications and might not be available for all patients. OBJECTIVES: We sought to collect data on the clinical presentation, treatment, and follow-up of a large sample of patients with XHIGM to (1) compare long-term overall survival and general well-being of patients treated with or without HCT along with clinical factors associated with mortality and (2) summarize clinical practice and risk factors in the subgroup of patients treated with HCT. METHODS: Physicians caring for patients with primary immunodeficiency diseases were identified through the Jeffrey Modell Foundation, United States Immunodeficiency Network, Latin American Society for Immunodeficiency, and Primary Immune Deficiency Treatment Consortium. Data were collected with a Research Electronic Data Capture Web application. Survival from time of diagnosis or transplantation was estimated by using the Kaplan-Meier method compared with log-rank tests and modeled by using proportional hazards regression. RESULTS: Twenty-eight clinical sites provided data on 189 patients given a diagnosis of XHIGM between 1964 and 2013; 176 had valid follow-up and vital status information. Sixty-seven (38%) patients received HCT. The average follow-up time was 8.5 ± 7.2 years (range, 0.1-36.2 years). No difference in overall survival was observed between patients treated with or without HCT (P = .671). However, risk associated with HCT decreased for diagnosis years 1987-1995; the hazard ratio was significantly less than 1 for diagnosis years 1995-1999. Liver disease was a significant predictor of overall survival (hazard ratio, 4.9; 95% confidence limits, 2.2-10.8; P < .001). Among survivors, those treated with HCT had higher median Karnofsky/Lansky scores than those treated without HCT (P < .001). Among patients receiving HCT, 27 (40%) had graft-versus-host disease, and most deaths occurred within 1 year of transplantation. CONCLUSION: No difference in survival was observed between patients treated with or without HCT across all diagnosis years (1964-2013). However, survivors treated with HCT experienced somewhat greater well-being, and hazards associated with HCT decreased, reaching levels of significantly less risk in the late 1990s. Among patients treated with HCT, treatment at an early age is associated with improved survival. Optimism remains guarded as additional evidence accumulates.

Newborn screening for severe combined immunodeficiency in 11 screening programs in the United States.

IMPORTANCE: Newborn screening for severe combined immunodeficiency (SCID) using assays to detect T-cell receptor excision circles (TRECs) began in Wisconsin in 2008, and SCID was added to the national recommended uniform panel for newborn screened disorders in 2010. Currently 23 states, the District of Columbia, and the Navajo Nation conduct population-wide newborn screening for SCID. The incidence of SCID is estimated at 1 in 100,000 births. OBJECTIVES: To present data from a spectrum of SCID newborn screening programs, establish population-based incidence for SCID and other conditions with T-cell lymphopenia, and document early institution of effective treatments. DESIGN: Epidemiological and retrospective observational study. SETTING: Representatives in states conducting SCID newborn screening were invited to submit their SCID screening algorithms, test performance data, and deidentified clinical and laboratory information regarding infants screened and cases with nonnormal results. Infants born from the start of each participating program from January 2008 through the most recent evaluable date prior to July 2013 were included. Representatives from 10 states plus the Navajo Area Indian Health Service contributed data from 3,030,083 newborns screened with a TREC test. MAIN OUTCOMES AND MEASURES: Infants with SCID and other diagnoses of T-cell lymphopenia were classified. Incidence and, where possible, etiologies were determined. Interventions and survival were tracked. RESULTS: Screening detected 52 cases of typical SCID, leaky SCID, and Omenn syndrome, affecting 1 in 58,000 infants (95% CI, 1/46,000-1/80,000). Survival of SCID-affected infants through their diagnosis and immune reconstitution was 87% (45/52), 92% (45/49) for infants who received transplantation, enzyme replacement, and/or gene therapy. Additional interventions for SCID and non-SCID T-cell lymphopenia included immunoglobulin infusions, preventive antibiotics, and avoidance of live vaccines. Variations in definitions and follow-up practices influenced the rates of detection of non-SCID T-cell lymphopenia. CONCLUSIONS AND RELEVANCE: Newborn screening in 11 programs in the United States identified SCID in 1 in 58,000 infants, with high survival. The usefulness of detection of non-SCID T-cell lymphopenias by the same screening remains to be determined.

Composition of the CD27+ Memory-B-Cell Compartment Delineates Immunoglobulin Deficiency Endotypes.

Abstract Purpose: The finding of reduced numbers of class-switched memory B cells (CSM) in peripheral blood is widely used to assist the diagnosis and subclassification of CVID. Limited data exists on this finding in relation to the entire class of PADs. In this study, consecutive 8-marker comprehensive B-cell panel results were analyzed to determine how reduced CSM quantities might inform the pathophysiology of CVID and other humoral immunodeficiencies. Methods: Subpopulations of CD27+ memory B cells from 64 consecutive subjects with or without humoral immunodeficiency were examined to identify associations with diagnosis and serum immunoglobulin level. Results: CD27+IgM-IgD- percentage (CSM%) was correlated with IgG level in a discontinuous manner with an estimated change point of 9.7% (95% CI: 4.7, 12.4). All subjects with a CSM% below 9.7% had substantially lower serum IgG and IgA levels compared with those above 9.7. CSM% below 9.7% is not associated with serum IgM level. Rather, the proportion of CD27+IgMonly B cells (IgMonly or IgMonly%) is correlated with serum IgM. Conclusion: Low CSM% may mark an endotype of humoral immune dysfunction defined by either loss of class switching or critical failure of the coordinated production of both memory cells and long-lived plasma cells responsible for adequate immunoglobulin levels in humans. In patients with low CSM%, maintenance or expansion of IgMonly cells and IgM production suggests the former explanation, while concomitant loss of IgMonly cells suggests the latter. These findings provide a simple endotypic stratification method for future studies on the failed coordinated B cell response in humans with PAD.

Meeting Report: 2023 Muscular Dystrophy Association Summit on 'Safety and Challenges in Gene Therapy of Neuromuscular Diseases'.

 This meeting report summarizes the presentations and discussions held at the summit on Challenges in Gene Therapy hosted by the Muscular Dystrophy Association (MDA) in 2023. Topics covered include safety issues, mitigation strategies and practical considerations pertaining to the clinical translation of gene therapies for neuromuscular disease. The listing of actionable recommendations will assist in overall efforts in the field to achieve safe and efficacious translation of gene therapies for neuromuscular disease patients.

Autophagy-associated immune dysregulation and hyperplasia in a patient with compound heterozygous mutations in ATG9A.

ABBREVIATIONS: BCL2: BCL2 apoptosis regulator; BCL10: BCL10 immune signaling adaptor; CARD11: caspase recruitment domain family member 11; CBM: CARD11-BCL10-MALT1; CR2: complement C3d receptor 2; EBNA: Epstein Barr nuclear antigen; EBV: Epstein-Barr virus; FCGR3A; Fc gamma receptor IIIa; GLILD: granulomatous-lymphocytic interstitial lung disease; HV: healthy volunteer; IKBKB/IKB kinase: inhibitor of nuclear factor kappa B kinase subunit beta; IL2RA: interleukin 2 receptor subunit alpha; MALT1: MALT1 paracaspase; MS4A1: membrane spanning 4-domain A1; MTOR: mechanistic target of rapamycin kinase; MYC: MYC proto-oncogene, bHLH: transcription factor; NCAM1: neural cell adhesion molecule 1; NFKB: nuclear factor kappa B; NIAID: National Institute of Allergy and Infectious Diseases; NK: natural killer; PTPRC: protein tyrosine phosphatase receptor type C; SELL: selectin L; PBMCs: peripheral blood mononuclear cells; TR: T cell receptor; Tregs: regulatory T cells; WT: wild-type.

"Common variable immunodeficiency: Challenges for diagnosis".

Common variable immunodeficiency is a heterogeneous condition characterized by B cell dysfunction with reduced serum immunoglobulin levels and a highly variable spectrum of clinical manifestations ranging from recurrent infections to autoimmune disease. The diagnosis of CVID is often challenging due to the diverse clinical presentation of patients and the existence of multiple diagnostic criteria without a universally adopted consensus. Laboratory evaluation to assist with diagnosis currently includes serum immunoglobulin testing, immunophenotyping, assessment of vaccine response, and genetic testing. Additional emerging techniques include investigation of the B cell repertoire and the use of machine learning algorithms. Advances in our understanding of common variable immunodeficiency will ultimately contribute to earlier diagnosis and novel interventions with the goal of improving prognosis for these patients.

Fluctuations in quality of life and immune responses during intravenous immunoglobulin infusion cycles.

Despite adequate infection prophylaxis, variation in self-reported quality of life (QOL) throughout the intravenous immunoglobulin (IVIG) infusion cycle is a widely reported but infrequently studied phenomenon. To better understand this phenomenon, subjects with humoral immunodeficiency receiving replacement doses of IVIG were studied over 3 infusion cycles. Questionnaire data from 6 time points spread over 3 IVIG infusions cycles (infusion day and 7 days after each infusion) were collected in conjunction with monitoring the blood for number of regulatory T-cells (Treg) and levels of 40 secreted analytes: primarily cytokines, chemokines, and growth factors. At day 7, self-reported well-being increased, and self-reported fatigue decreased, reflecting an overall improvement in QOL 7 days after infusion. Over the same period, percentage of Treg cells in the blood increased (p<0.01). Multiple inflammatory chemokine and cytokine levels increased in the blood by 1 hour after infusion (CCL4 (MIP-1b), CCL3 (MIP-1a), CCL2 (MCP-1), TNF-α, granzyme B, IL-10, IL-1RA, IL-8, IL-6, GM-CSF, and IFN- γ). The largest changes in analytes occurred in subjects initiated on IVIG during the study. A significant decrease in IL-25 (IL-17E) following infusion was seen in most intervals among subjects already receiving regular infusions prior to study entry. These findings reveal several short-term effects of IVIG given in replacement doses to patients with humoral immunodeficiency: QOL consistently improves in the first week of infusion, levels of a collection of monocyte-associated cytokines increase immediately after infusion whereas IL-25 levels decrease, and Treg levels increase. Moreover, patients that are new to IVIG experience more significant fluctuations in cytokine levels than those receiving it regularly.

Expression and activation of the steroidogenic enzyme CYP11A1 is associated with IL-13 production in T cells from peanut allergic children.

Activation of the steroidogenic enzyme CYP11A1 was shown to be necessary for the development of peanut-induced intestinal anaphylaxis and IL-13 production in allergic mice. We determined if levels of CYP11A1 in peripheral blood T cells from peanut-allergic (PA) children compared to non-allergic controls were increased and if levels correlated to IL-13 production and oral challenge outcomes to peanut. CYP11A1 mRNA and protein levels were significantly increased in activated CD4+ T cells from PA patients. In parallel, IL-13 production was significantly increased; IFNγ levels were not different between groups. There were significant correlations between expression levels of CYP11A1 mRNA and levels of IL13 mRNA and protein, levels of serum IgE anti-Ara h 2 and to outcomes of peanut challenge. The importance of CYP11A1 on cytokine production was tested using a CYP11A1 CRISPR/Cas9 KO plasmid or an inhibitor of enzymatic CYP11A1 activity. Inhibition of CYP11A1 activation in patient cells treated with the inhibitor, aminoglutethimide, or CD4+ T cell line transfected with the CYP11A1 KO plasmid resulted in reduced IL-13 production. These data suggest that the CYP11A1-CD4+Tcell-IL-13 axis in activated CD4+ T cells from PA children is associated with development of PA reactions. CYP11A1 may represent a novel target for therapeutic intervention in PA children.

Vasculitis in a Child With the Hyper-IgM Variant of Ataxia-Telangiectasia.

A subset of patients with Ataxia-Telangiectasia (A-T) have dramatically reduced levels of IgG, IgA, and IgE with retained or elevated IgM levels. Several reports suggest that these A-T patients with a "hyper-IgM phenotype" (HIgM) suffer more clinical immunologic consequences than other A-T patients. The immunopathologic mechanism driving this phenomenon is unknown, making it difficult to predict response to immunomodulatory therapy. We describe an A-T patient with HIgM who underwent tumor necrosis factor (TNF) receptor blockade for cutaneous granuloma and after several months of successful therapy developed non-malignant lymphoproliferation, cytopenia, and increased serum immunoglobulin levels. This process was subsequently followed by an immune-complex-mediated intrarenal small vessel vasculitis that led to renal failure. The vasculitis was successfully treated with rituximab and corticosteroids. This case underscores the importance of HIgM as an unfavorable prognostic indicator in A-T and highlights the complexity of immunomodulatory treatment in this population, and the potential for a successful approach tailored to the immune defect.

Biallelic mutations in DNA ligase 1 underlie a spectrum of immune deficiencies.

We report the molecular, cellular, and clinical features of 5 patients from 3 kindreds with biallelic mutations in the autosomal LIG1 gene encoding DNA ligase 1. The patients exhibited hypogammaglobulinemia, lymphopenia, increased proportions of circulating γδT cells, and erythrocyte macrocytosis. Clinical severity ranged from a mild antibody deficiency to a combined immunodeficiency requiring hematopoietic stem cell transplantation. Using engineered LIG1-deficient cell lines, we demonstrated chemical and radiation defects associated with the mutant alleles, which variably impaired the DNA repair pathway. We further showed that these LIG1 mutant alleles are amorphic or hypomorphic, and exhibited variably decreased enzymatic activities, which lead to premature release of unligated adenylated DNA. The variability of the LIG1 genotypes in the patients was consistent with that of their immunological and clinical phenotypes. These data suggest that different forms of autosomal recessive, partial DNA ligase 1 deficiency underlie an immunodeficiency of variable severity.

Recurrent rhinovirus infections in a child with inherited MDA5 deficiency.

MDA5 is a cytosolic sensor of double-stranded RNA (ds)RNA including viral byproducts and intermediates. We studied a child with life-threatening, recurrent respiratory tract infections, caused by viruses including human rhinovirus (HRV), influenza virus, and respiratory syncytial virus (RSV). We identified in her a homozygous missense mutation in IFIH1 that encodes MDA5. Mutant MDA5 was expressed but did not recognize the synthetic MDA5 agonist/(ds)RNA mimic polyinosinic-polycytidylic acid. When overexpressed, mutant MDA5 failed to drive luciferase activity from the IFNB1 promoter or promoters containing ISRE or NF-κB sequence motifs. In respiratory epithelial cells or fibroblasts, wild-type but not knockdown of MDA5 restricted HRV infection while increasing IFN-stimulated gene expression and IFN-ß/λ. However, wild-type MDA5 did not restrict influenza virus or RSV replication. Moreover, nasal epithelial cells from the patient, or fibroblasts gene-edited to express mutant MDA5, showed increased replication of HRV but not influenza or RSV. Thus, human MDA5 deficiency is a novel inborn error of innate and/or intrinsic immunity that causes impaired (ds)RNA sensing, reduced IFN induction, and susceptibility to the common cold virus.