Phase 3 randomized trial of mavorixafor, CXCR4 antagonist, in WHIM syndrome.

We investigated efficacy and safety of mavorixafor, an oral CXCR4 antagonist for participants with Warts, Hypogammaglobulinemia, Infections, and Myelokathexis (WHIM) syndrome, a rare immunodeficiency caused by CXCR4 gain-of-function variants. This randomized (1:1), double-blind, placebo-controlled, phase 3 trial enrolled participants aged ≥12 years with WHIM syndrome and absolute neutrophil count (ANC) ≤400/µL. Participants received once-daily mavorixafor or placebo for 52 weeks. Primary endpoint was time (hours) above ANC threshold ≥500/µL (TATANC; over 24 hours). Secondary endpoints included TAT absolute lymphocyte count ≥1000/µL (TATALC; defined similar to TATANC); absolute changes in white blood cell (WBC), ANC, and ALC from baseline; annualized infection rate; infection duration and total infection score (combined infection number/severity). In 31 participants (mavorixafor, n=14; placebo, n=17), mavorixafor least squares (LS) mean TATANC was 15.0 hours, placebo 2.8 hours (P<0.001). Mavorixafor LS mean TATALC was 15.8 hours, placebo 4.6 hours (P<0.001). Higher absolute WBC, ANC, and ALC levels were seen with mavorixafor than placebo at each timepoint assessed. Annualized infection rates were 60% lower with mavorixafor versus placebo (LS mean 1.7 versus 4.2; nominal P=0.007) and total infection scores were 40% lower (7.4 [95% CI, 1.6-13.2] versus 12.3 [95% CI, 7.2-17.3]). Treatment with mavorixafor reduced infection frequency, severity, duration, and antibiotic use. No discontinuations occurred due to treatment-emergent adverse events (TEAEs); no related serious TEAEs were observed. Overall, mavorixafor-treated participants showed significant increases in LS mean TATANC and TATALC, reduced infection frequency, severity/duration. Mavorixafor was well tolerated in participants with WHIM syndrome. Trial was registered at ClinicalTrials.gov NCT03995108.

CVID-Associated Intestinal Disorders in the USIDNET Registry: An Analysis of Disease Manifestations, Functional Status, Comorbidities, and Treatment.

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.

Comprehensive phenotypic analysis of diverse FOXN1 variants.

BACKGROUND: Thymus hypoplasia due to stromal cell problems has been linked to mutations in several transcription factors, including Forkhead box N1 (FOXN1). FOXN1 supports T-cell development by regulating the formation and expansion of thymic epithelial cells (TECs). While autosomal recessive FOXN1 mutations result in a nude and severe combined immunodeficiency phenotype, the impact of single-allelic or compound heterozygous FOXN1 mutations is less well-defined. OBJECTIVE: With more than 400 FOXN1 mutations reported, their impact on protein function and thymopoiesis remains unclear for most variants. We developed a systematic approach to delineate the functional impact of diverse FOXN1 variants. METHODS: Selected FOXN1 variants were tested with transcriptional reporter assays and imaging studies. Thymopoiesis was assessed in mouse lines genocopying several human FOXN1 variants. Reaggregate thymus organ cultures were used to compare the thymopoietic potential of the FOXN1 variants. RESULTS: FOXN1 variants were categorized into benign, loss- or gain-of-function, and/or dominant-negatives. Dominant negative activities mapped to frameshift variants impacting the transactivation domain. A nuclear localization signal was mapped within the DNA binding domain. Thymopoiesis analyses with mouse models and reaggregate thymus organ cultures revealed distinct consequences of particular Foxn1 variants on T-cell development. CONCLUSIONS: The potential effect of a FOXN1 variant on T-cell output from the thymus may relate to its effects on transcriptional activity, nuclear localization, and/or dominant negative functions. A combination of functional assays and thymopoiesis comparisons enabled a categorization of diverse FOXN1 variants and their potential impact on T-cell output from the thymus.

CVID-associated intestinal disorders in the USIDNET registry: An analysis of disease manifestations, functional status, comorbidities, and treatment.

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.

Chromosomal Numerical Aberrations and Rare Copy Number Variation in Patients with Inflammatory Bowel Disease.

BACKGROUND AND AIMS: Inflammatory bowel diseases [IBD] have a complex polygenic aetiology. Rare genetic variants can cause monogenic intestinal inflammation. The impact of chromosomal aberrations and large structural abnormalities on IBD susceptibility is not clear. We aimed to comprehensively characterise the phenotype and prevalence of patients with IBD who possess rare numerical and structural chromosomal abnormalities. METHODS: We performed a systematic literature search of databases PubMed and Embase; and analysed gnomAD, Clinvar, the 100 000 Genomes Project, and DECIPHER databases. Further, we analysed international paediatric IBD cohorts to investigate the role of IL2RA duplications in IBD susceptibility. RESULTS: A meta-analysis suggests that monosomy X [Turner syndrome] is associated with increased expressivity of IBD that exceeds the population baseline (1.86%, 95% confidence interval [CI] 1.48 to 2.34%) and causes a younger age of IBD onset. There is little evidence that Klinefelter syndrome, Trisomy 21, Trisomy 18, mosaic Trisomy 9 and 16, or partial trisomies contribute to IBD susceptibility. Copy number analysis studies suggest inconsistent results. Monoallelic loss of X-linked or haploinsufficient genes is associated with IBD by hemizygous or heterozygous deletions, respectively. However, haploinsufficient gene deletions are detected in healthy reference populations, suggesting that the expressivity of IBD might be overestimated. One duplication that has previously been identified as potentially contributing to IBD risk involves the IL2RA/IL15R loci. Here we provide additional evidence that a microduplication of this locus may predispose to very-early-onset IBD by identifying a second case in a distinct kindred. However, the penetrance of intestinal inflammation in this genetic aberration is low [<2.6%]. CONCLUSIONS: Turner syndrome is associated with increased susceptibility to intestinal inflammation. Duplication of the IL2RA/IL15R loci may contribute to disease risk.

Epigenetic and immunological indicators of IPEX disease in subjects with FOXP3 gene mutation.

BACKGROUND: Forkhead box protein 3 (FOXP3) is the master transcription factor in CD4+CD25hiCD127lo regulatory T (Treg) cells. Mutations in FOXP3 result in IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked) syndrome. Clinical presentation of IPEX syndrome is broader than initially described, challenging the understanding of the disease, its evolution, and treatment choice. OBJECTIVE: We sought to study the type and extent of immunologic abnormalities that remain ill-defined in IPEX, across genetic and clinical heterogeneity. METHODS: We performed Treg-cell-specific epigenetic quantification and immunologic characterization of severe "typical" (n = 6) and "atypical" or asymptomatic (n = 9) patients with IPEX. RESULTS: Increased number of cells with Treg-cell-Specific Demethylated Region demethylation in FOXP3 is a consistent feature in patients with IPEX, with (1) highest values in those with typical IPEX, (2) increased values in subjects with pathogenic FOXP3 but still no symptoms, and (3) gradual increase over the course of disease progression. Large-scale profiling using Luminex identified plasma inflammatory signature of macrophage activation and TH2 polarization, with cytokines previously not associated with IPEX pathology, including CCL22, CCL17, CCL15, and IL-13, and the inflammatory markers TNF-α, IL-1A, IL-8, sFasL, and CXCL9. Similarly, both Treg-cell and Teff compartments, studied by Mass Cytometry by Time-Of-Flight, were skewed toward the TH2 compartment, especially in typical IPEX. CONCLUSIONS: Elevated TSDR-demethylated cells, combined with elevation of plasmatic and cellular markers of a polarized type 2 inflammatory immune response, extends our understanding of IPEX diagnosis and heterogeneity.

Mesenchymal cell replacement corrects thymic hypoplasia in murine models of 22q11.2 deletion syndrome.

22q11.2 deletion syndrome (22q11.2DS) is the most common human chromosomal microdeletion, causing developmentally linked congenital malformations, thymic hypoplasia, hypoparathyroidism, and/or cardiac defects. Thymic hypoplasia leads to T cell lymphopenia, which most often results in mild SCID. Despite decades of research, the molecular underpinnings leading to thymic hypoplasia in 22q11.2DS remain unknown. Comparison of embryonic thymuses from mouse models of 22q11.2DS (Tbx1neo2/neo2) revealed proportions of mesenchymal, epithelial, and hematopoietic cell types similar to those of control thymuses. Yet, the small thymuses were growth restricted in fetal organ cultures. Replacement of Tbx1neo2/neo2 thymic mesenchymal cells with normal ones restored tissue growth. Comparative single-cell RNA-Seq of embryonic thymuses uncovered 17 distinct cell subsets, with transcriptome differences predominant in the 5 mesenchymal subsets from the Tbx1neo2/neo2 cell line. The transcripts affected included those for extracellular matrix proteins, consistent with the increased collagen deposition we observed in the small thymuses. Attenuating collagen cross-links with minoxidil restored thymic tissue expansion for hypoplastic lobes. In colony-forming assays, the Tbx1neo2/neo2-derived mesenchymal cells had reduced expansion potential, in contrast to the normal growth of thymic epithelial cells. These findings suggest that mesenchymal cells were causal to the small embryonic thymuses in the 22q11.2DS mouse models, which was correctable by substitution with normal mesenchyme.

The mammalian SKIV2L RNA exosome is essential for early B cell development.

The SKIV2L RNA exosome is an evolutionarily conserved RNA degradation complex in the eukaryotes. Mutations in the SKIV2L gene are associated with a severe inherited disorder, trichohepatoenteric syndrome (THES), with multisystem involvement but unknown disease mechanism. Here, we reported a THES patient with SKIV2L mutations showing severe primary B cell immunodeficiency, hypogammaglobulinemia, and kappa-restricted plasma cell dyscrasia but normal T cell and NK cell function. To corroborate these findings, we made B cell-specific Skiv2l knockout mice (Skiv2lfl/flCd79a-Cre), which lacked both conventional B-2 and innate-like B-1 B cells in the periphery and secondary lymphoid organs. This was linked to a requirement of SKIV2L RNA exosome activity in the bone marrow during early B cell development at the pro-B cell to large pre-B cell transition. Mechanistically, Skiv2l-deficient pro-B cells exhibited cell cycle arrest and DNA damage. Furthermore, loss of Skiv2l led to substantial out-of-frame V(D)J rearrangement of immunoglobulin heavy chain and severely reduced surface expression of µH, both of which are crucial for pre-BCR signaling and proliferative burst during early B cell development. Together, our data demonstrated a crucial role for SKIV2L RNA exosome in early B cell development in both human and mice by ensuring proper V(D)J recombination and Igh expression, which serves as the molecular basis for immunodeficiency associated with THES.

Cytoplasmic RNA quality control failure engages mTORC1-mediated autoinflammatory disease.

Inborn errors of nucleic acid metabolism often cause aberrant activation of nucleic acid sensing pathways, leading to autoimmune or autoinflammatory diseases. The SKIV2L RNA exosome is cytoplasmic RNA degradation machinery that was thought to be essential for preventing the self-RNA-mediated interferon (IFN) response. Here, we demonstrate the physiological function of SKIV2L in mammals. We found that Skiv2l deficiency in mice disrupted epidermal and T cell homeostasis in a cell-intrinsic manner independently of IFN. Skiv2l-deficient mice developed skin inflammation and hair abnormality, which were also observed in a SKIV2L-deficient patient. Epidermis-specific deletion of Skiv2l caused hyperproliferation of keratinocytes and disrupted epidermal stratification, leading to impaired skin barrier with no appreciable IFN activation. Moreover, Skiv2l-deficient T cells were chronically hyperactivated and these T cells attacked lesional skin as well as hair follicles. Mechanistically, SKIV2L loss activated the mTORC1 pathway in both keratinocytes and T cells. Both systemic and topical rapamycin treatment of Skiv2l-deficient mice ameliorated epidermal hyperplasia and skin inflammation. Together, we demonstrate that mTORC1, a classical nutrient sensor, also senses cytoplasmic RNA quality control failure and drives autoinflammatory disease. We also propose SKIV2L-associated trichohepatoenteric syndrome (THES) as a new mTORopathy for which sirolimus may be a promising therapy.

Obstructive hydrocephalus and intracerebral mass secondary to Epicoccum nigrum.

Here we report a case of a 14-week-old girl with a history of intrauterine drug exposure and hypoxic ischemic encephalopathy secondary to cardiac arrest requiring prolonged resuscitation at birth presented with irritability and a bulging anterior fontanelle. After neurosurgical resection, pathologic examination showed fungal hyphae, and Epicoccum nigrum was detected by fungal PCR and sequencing. To our knowledge, this is the first reported case of a central nervous system infection due to Epicoccum nigrum.

Lymphomatoid granulomatosis of the central nervous system (CNS-LYG) posing a management challenge.

Isolated central nervous system lymphomatoid granulomatosis (CNS-LYG) can mimic aggressive glioblastomas. We describe a complex presentation of CNS-LYG coexisting with immune thrombocytopenia successfully managed with rituximab and ultra-low-dose radiation therapy.

SARS-CoV-2 infection associated with hepatitis in an infant with X-linked severe combined immunodeficiency.

X-linked severe combined immunodeficiency (X-SCID) is a disorder of adaptive immunity caused by mutations in the IL-2 receptor common gamma chain gene resulting in deficiencies of T and natural killer cells, coupled with severe dysfunction in B cells. X-SCID is lethal without allogeneic stem cell transplant or gene therapy due to opportunistic infections. An infant with X-SCID became infected with SARS-CoV-2 while awaiting transplant. The patient developed severe hepatitis without the respiratory symptoms typical of COVID-19. He was treated with convalescent plasma, and thereafter was confirmed to have SARS-CoV-2 specific antibodies, as detected with a microfluidic antigen array. After resolution of the hepatitis, he received a haploidentical CD34 selected stem cell transplant, without conditioning, from his father who had recovered from COVID-19. SARS CoV-2 was detected via RT-PCR on nasopharyngeal swabs until 61 days post transplantation. He successfully engrafted donor T and NK cells, and continues to do well clinically.

Molecular Insights Into the Causes of Human Thymic Hypoplasia With Animal Models.

22q11.2 deletion syndrome (DiGeorge), CHARGE syndrome, Nude/SCID and otofaciocervical syndrome type 2 (OTFCS2) are distinct clinical conditions in humans that can result in hypoplasia and occasionally, aplasia of the thymus. Thymic hypoplasia/aplasia is first suggested by absence or significantly reduced numbers of recent thymic emigrants, revealed in standard-of-care newborn screens for T cell receptor excision circles (TRECs). Subsequent clinical assessments will often indicate whether genetic mutations are causal to the low T cell output from the thymus. However, the molecular mechanisms leading to the thymic hypoplasia/aplasia in diverse human syndromes are not fully understood, partly because the problems of the thymus originate during embryogenesis. Rodent and Zebrafish models of these clinical syndromes have been used to better define the underlying basis of the clinical presentations. Results from these animal models are uncovering contributions of different cell types in the specification, differentiation, and expansion of the thymus. Cell populations such as epithelial cells, mesenchymal cells, endothelial cells, and thymocytes are variably affected depending on the human syndrome responsible for the thymic hypoplasia. In the current review, findings from the diverse animal models will be described in relation to the clinical phenotypes. Importantly, these results are suggesting new strategies for regenerating thymic tissue in patients with distinct congenital disorders.

A late preterm infant with lymphopenia.

The newborn screen for severe combined immunodeficiency (SCID) uses real-time quantitative polymerase chain reaction for T-cell receptor excision circles and is highly sensitive for SCID. However, T-cell lymphopenia from other primary and secondary causes, such as DiGeorge syndrome, prematurity, thymic involution from stress, and thymectomy during cardiac surgery, is also detected. We present a newborn girl with T-cell lymphopenia of unknown etiology detected via abnormal newborn screen.

Connecting the Dots From Fever of Unknown Origin to Myelodysplastic Syndrome: GATA2 Haploinsufficiency.

Leukemia-predisposing conditions, such as GATA2 haploinsufficiency, are known for their high penetrance and expressivity profiles. These disorders pose a difficult diagnostic challenge to even the most experienced clinician when they first present. We describe the case of a 17-year-old male presenting with features of nontuberculous mycobacterial infection, pulmonary fibrinoid granulomatous vasculitis, and myelodysplasia in the setting of a pathogenic GATA2 frameshift mutation confirmed by next-generation sequencing. The broad differential for GATA2 haploinsufficiency requires prompt recognition of key clinical features and laboratory abnormalities towards directing diagnosis and guiding appropriate and perhaps life-saving therapy.

FOXN1 compound heterozygous mutations cause selective thymic hypoplasia in humans.

We report on 2 patients with compound heterozygous mutations in forkhead box N1 (FOXN1), a transcription factor essential for thymic epithelial cell (TEC) differentiation. TECs are critical for T cell development. Both patients had a presentation consistent with T-/loB+NK+ SCID, with normal hair and nails, distinct from the classic nude/SCID phenotype in individuals with autosomal-recessive FOXN1 mutations. To understand the basis of this phenotype and the effects of the mutations on FOXN1, we generated mice using CRISPR-Cas9 technology to genocopy mutations in 1 of the patients. The mice with the Foxn1 compound heterozygous mutations had thymic hypoplasia, causing a T-B+NK+ SCID phenotype, whereas the hair and nails of these mice were normal. Characterization of the functional changes due to the Foxn1 mutations revealed a 5-amino acid segment at the end of the DNA-binding domain essential for the development of TECs but not keratinocytes. The transcriptional activity of this Foxn1 mutant was partly retained, indicating a region that specifies TEC functions. Analysis of an additional 9 FOXN1 mutations identified in multiple unrelated patients revealed distinct functional consequences contingent on the impact of the mutation on the DNA-binding and transactivation domains of FOXN1.