We report a case of delayed acute liver injury (ALI) after treatment with delandistrogene moxeparvovec-rokl (DM), an adeno-associated virus-based gene therapy, in a 10-year-old ambulatory boy with Duchenne muscular dystrophy (DMD). The patient had a duplication of exons 8-11 in the DMD gene and was on chronic prednisone. He received prophylactic oral prednisone (1 mg/kg/d) starting 1 day before DM infusion. Baseline alanine aminotransferase (ALT), aspartate aminotransferase (AST), and creatine kinase (CK) levels were elevated, consistent with DMD. On day 34 after infusion, ALT and AST began to rise without gamma-glutamyl transpeptidase (GGT) elevation. By day 41, transaminases were more than 7× baseline with elevated GGT and bilirubin, prompting hospitalization. He was treated with intravenous methylprednisolone, increased oral prednisone, and sirolimus. Liver enzymes rapidly improved and returned to baseline by day 69. Immunologic studies revealed CD8+ T-cell activation and innate immune activation via mildly elevated IL-18 that normalized without additional immunosuppression. The patient remained clinically stable. Infectious workup, liver ultrasonogram, and coagulation studies were unremarkable. Notably, the ALT/CK ratio increased more than 10-fold 1  week before GGT elevation and before ALT exceeded the 95% CI predicted by a CK- and age-adjusted model, suggesting its potential as an early biomarker of liver injury in DMD. This case highlights the challenges of interpreting liver enzymes in DMD, the potential utility of the ALT/CK ratio for early detection of ALI, and the role of immunophenotyping in guiding immunosuppressive therapy. Early recognition and timely escalation, including use of sirolimus, may help mitigate serious liver toxicity after gene therapy.

Abstract Introduction BEACONS-NBS (Building Evidence and Collaboration for GenOmics in Nationwide Newborn Screening) is the first research study to integrate whole genome sequencing into newborn screening (NBS) across multiple U.S. states and territorial public health laboratory programs (PHLPs). We developed a list of conditions for screening. Methods We designed inclusion criteria and assembled an initial condition list from published resources. The list was revised by clinical experts, molecular geneticists, genetic counselors, PHLPs, rare disease advocacy organizations, the BEACONS-NBS Community Advisory Board, and project leadership from the National Institutes of Health. For each condition, we provided a rationale for early detection, diagnostic signs or biomarkers, and treatments or surveillance strategies. Results The BEACONS-NBS condition list includes 777 conditions associated with 743 genes, one copy number variant, and two aneuploidies and is larger than those used in other genomic NBS research studies in the U.S. and United Kingdom. Most conditions are inborn errors of immunity (37.2%), inherited metabolic disorders (18.7%), or endocrine conditions (18.1%). Nearly all conditions (93.3%) can be confirmed using a non-genetic test. Discussion BEACONS-NBS has established a condition list for implementation across multiple state and territorial PHLPs, enabling the prospective evaluation of feasibility of population-wide genomic NBS.

BACKGROUND:Newborn screening (NBS) for severe combined immunodeficiency (SCID) using T-cell receptor excision circles in dried blood spots has been implemented in the United States and many other regions and countries globally. The Clinical Immunology Society and the Association of Public Health Laboratories jointly formed the SCID Harmonization Initiative to facilitate comparison of NBS reporting practices to promote global consensus and collaboration. OBJECTIVE:To assess current NBS SCID practices using a global survey and to report the findings from the phase 1 component. METHODS:An 18-question survey was distributed to all known SCID screening programs worldwide. Only 1 response per region was analyzed. Examples of international screening algorithms were also solicited and included. RESULTS:A total of 200 responses were received, of which 80 responses were unique and used for further analysis. Of the 39 non-US countries, 15 (38%) reported national universal screening and 24 (62%) reported regional, pilot, or other screening. Additional questions pertained to methodology and reporting, with particular emphasis on communication of the clinical urgency of an abnormal T-cell receptor excision circle result. CONCLUSIONS:This global survey confirmed that the approach to NBS SCID varies widely, underscoring the need for harmonization at multiple steps, particularly for reporting and interpretation. This is the first study to capture global NBS SCID practices, and these findings provide the basis for creation of a phase 2 consensus reporting framework, which will be developed by the same SCID Harmonization Committee that created the current study.

BACKGROUND:Type I interferons (IFN-I) and type II interferon (IFN-γ) contribute to the pathogenesis of inflammatory diseases, but measurement of these cytokines remains challenging in the clinical setting. OBJECTIVE:We aimed to develop a clinical test that rapidly captures the activity of IFN-I and IFN-γ. METHODS:We compared RNA sequencing data from healthy controls and patients with macrophage activation syndrome, multisystem inflammatory syndrome in children, and systemic lupus erythematosus to evaluate biomarkers of IFN-I and IFN-γ signaling. We utilized a flow cytometry assay to measure interferon-inducible markers. The findings were verified by a clinical laboratory that independently developed and validated this assay. RESULTS:We identified CD274 (programmed death ligand 1, or PD-L1) as a biomarker of IFN-γ signaling and confirmed CD169 (SIGLEC-1) as a readout of IFN-I activity. We utilized a flow cytometry assay to quantify CD169 and CD274 expression on monocytes and demonstrated the validity of this method for rapid screening of interferon dysregulation in patients with various inflammatory diseases including, among others, macrophage activation syndrome, hemophagocytic lymphohistiocytosis, multisystem inflammatory syndrome in children, systemic lupus erythematosus, juvenile idiopathic arthritis, and juvenile dermatomyositis. We further demonstrated the utility of this test for assessment of interferon dysregulation in monogenic inflammatory diseases and for efficacy monitoring of medications that target the interferon pathways including Janus kinase inhibitors, emapalumab, and anifrolumab. Finally, we outlined the steps of assay validation and clinical implementation, and we showed reproducible findings in a clinical laboratory. CONCLUSION:Flow cytometry analysis of CD169 and CD274 is an effective method to rapidly quantify IFN-I and IFN-γ activity in the clinical setting.

Biallelic loss-of-function variants in capping protein regulator and myosin 1 linker 2 (CARMIL2) cause a complex disorder of immune dysregulation hallmarked by susceptibility to infections, inflammatory bowel and skin disease, and Epstein-Barr virus-positive smooth muscle tumors (EBV+ SMTs). We report a multicenter retrospective study to evaluate hematopoietic cell transplantation (HCT) outcomes in CARMIL2-deficient patients. 17 patients underwent 19 HCTs, with a total follow-up of 768 mo and a median follow-up of 37 mo (range 1-195). Three patients died during the early posttransplant period (overall survival, 82.4%), and two required a second HCT for graft failure. Despite limitations due to cohort size and high pre-transplant morbidity in individual patients, HCT improved all major disease manifestations, including infections, inflammatory disease, and previously treatment-refractory EBV+ SMTs. No patient required immunoglobulin replacement after HCT, and no detrimental effects of mixed chimerism were noted. Allogeneic HCT is therefore a curative option for patients with CARMIL2 deficiency, which should be offered upon diagnosis.

Allogeneic HCT substantially addresses immunodeficiency, immune dysregulation, and Epstein-Barr virus–positive smooth muscle tumors in patients with CARMIL2 deficiency and should be offered shortly following diagnosis.

Background and Objectives: Early identification of cardiac dysfunction in multi-system inflammatory syndrome in children (MIS-C) is crucial for effective management. Our primary objective was to predict left ventricular systolic dysfunction (LVSD) through a multicenter collaborative assessing admission laboratory data and echocardiogram findings. Methods: Laboratory and clinical data were collected by retrospective chart review from a cohort of pediatric patients admitted and treated for MIS-C in our institutions. Laboratory data including absolute lymphocyte count, albumin, sedimentation rate, C-reactive protein, procalcitonin, d-dimer, fibrinogen, ferritin, interleukin-6 level, and lymphocyte subsets (T, B and NK quantitation, TBNK) were collected. We built a LASSO logistic regression model to predict which MIS-C patients would have left ventricular systolic dysfunction LVSD using only laboratory data obtained within the first 24 h of admission. Results: Of the 1474 MIS-C patients evaluated, 297 had LVSD. The linear kinetic analysis found differences in albumin, lymphocyte count, C-reactive proteins and fibrinogen for systolic dysfunction patients, and of these C-reactive proteins, fibrinogen and procalcitonin were more predictive earlier. The best model for coronary artery abnormalities (CAAs) performed poorly, with a mean cross-validated AUC of 0.57. The model performed well with a cross-validated AUC of 0.845. Conclusions: This model identified widely available biomarkers to successfully predict systolic dysfunction in MIS-C patients. Those at high risk of systolic dysfunction had higher peak laboratory values for C-reactive protein, fibrinogen, and procalcitonin early on. A regularized logistic regression model was validated to provide excellent discrimination for LVSD.

Precision medicine aims to enhance diagnosis, treatment, and prognosis by integrating multimodal data at the point of care. However, challenges arise due to the vast number of diseases, differing methods of classification, and conflicting terminological coding systems and practices used to represent molecular definitions of disease. This lack of interoperability artificially constrains the potential for diagnosis, clinical decision support, care outcome analysis, as well as data linkage across research domains to support the development or repurposing of therapeutics. There is a clear and pressing need for a unified system for managing disease entities⁠-including identifiers, synonyms, and definitions. To address these issues, we created the Mondo disease ontology-a community-driven, open-source, unified disease classification system that harmonizes diverse terminologies into a consistent, computable framework. Mondo integrates key medical and biomedical terminologies, including Online Mendelian Inheritance in Man (OMIM), Orphanet, Medical Subject Headings (MeSH), National Cancer Institute Thesaurus (NCIt), and more, to provide a comprehensive and accurate representation of disease concepts with fully provenanced and attributed links back to the sources. Mondo can be used as the handle for curation of gene-disease associations utilized in diagnostic applications, research applications such as computational phenotyping, and in clinical coding systems in clinical decision support by pointing the clinician to the numerous knowledge resources linked to the Mondo identifier. Mondo's community-centric approach, stewarded by the Monarch Initiative's expertise in ontologies, ensures that the ontology remains adaptable to the evolving needs of biomedical research and clinical communities, as well as the knowledge providers.

Background: Autoantibodies against IFN-α (anti–IFN-α) have been reported in recombinase activating gene (RAG) deficiency, attributed to impaired central and peripheral T-cell/B-cell tolerance. However, the clinical features, especially viral infections, associated with these autoantibodies at baseline, their kinetics over time, and their response to hematopoietic cell transplantation are not well defined. Objective: We described the clinical and immunologic findings linked to anti–IFN-α IgG in RAG deficiency and tracked its kinetics longitudinally, including in those who underwent hematopoietic cell transplantation. Methods: We measured anti–IFN-α IgG by enzyme-linked immunosorbent assay in 80 RAG-deficient patients with curated clinical and immunologic data from a multinational collaboration. Results: Forty-eight patients (60.0%) had positive anti–IFN-α at baseline; these patients were typically older at time of testing, fulfilled the phenotype of delayed-onset combined immunodeficiency with granuloma and/or autoimmunity (70.8% vs 31.3%, P = .001), and had a history of more frequent viral infections, mainly from the Herpesviridae family (62.5% vs 21.9%, P < .001). These patients also showed higher levels of serum immunoglobulins and expanded populations of peripheral blood autoreactive-prone (CD19hiCD21lo) (14.3 vs 5.2%, P = .016) and double-negative (IgD−CD27−) B cells (12.8 vs 5.8%, P = .001). In cases with longitudinal evaluation, anti–IFN-α titers were largely stable, although an increase was observed with concurrent active cytomegalovirus infections. Despite some decline after transplantation, these autoantibodies persisted during follow-up. Conclusions: Anti–IFN-α autoantibodies reflect immune dysregulation in partial RAG deficiency. Their production is likely aggravated by environmental factors, especially frequent viral infections. Further studies are needed to define their pathogenic role in RAG deficiency.

We report the development of REMEDY (REpair of heterozygous Mutations independent of Exogenous Donor template with high efficiencY), a genome editing strategy that allows efficient repair of heterozygous mutations in human and mouse cells without necessitating an exogenous donor DNA template. Here, we used in-PAM or near-PAM CRISPR strategies to induce a double-strand break (DSB) in mutant alleles. Following the DSB, the wild-type homologous chromosome itself serves as an endogenous DNA donor template and initiates the correction of the mutant allele. Concurrently treating the cells with HDR enhancers, such as AZD7648, further improved the efficiency of the correction. We demonstrated the utility of REMEDY in the context of six different diseases with heterozygous mutations such as IBMPFD, cystic fibrosis, progeria, ITPR3-associated combined immunodeficiency, ACTA1, and TBCD in human patient derived primary cells and complementary mouse model cell lines. ![Figure][1] Graphical Abstract ### Competing Interest Statement MNK, YS, ZS, BO, JM, NYS have provisional patents related to the technology described here. MNK, ZS and NYS have received funding related to this technology. [1]: pending:yes

Objectives Proteasome-associated autoinflammatory syndromes (PRAAS) include a group of autoinflammatory interferonopathies caused by 20S proteasome dysfunction. We characterised pathomechanisms and treatment responses of patients with a de novo, dominant-negative (DN)-proteasome subunit beta type-8 (PSMB8) variant. Methods Patients with the DN-PSMB8 p.G209R variant encoding a mutant β5i subunit of the 20S immunoproteasome were evaluated. Interferon biomarkers, proteasome activity, structural modelling, and proteotoxic stress responses were assessed. Patients’ T cells underwent integrated transcriptomic and proteomic profiling to characterise immune dysregulation, proteotoxic stress responses, mitochondrial function, and type I interferon (IFN-I) associated stress signalling. Results Patients with DN-PRAAS presented with early-onset systemic inflammation, panniculitis, cytopenias, infections, and porto-sinusoidal vascular liver disease (PSVD), indicating broader immune dysfunction that partially responds to Janus kinase inhibition and/or interferon-α/β receptor blockade (anifrolumab). Mechanistically, the PSMB8 p.G209R variant caused steric hindrance that impaired β5i propeptide processing and final 20S proteasome formation, resulting in intracellular protein aggregation, impaired mitochondrial metabolism, and altered neutral lipid processing. The IFN-I signature of patients’ T cells was reduced by blockade of 2 integrated stress response (ISR)-regulating kinases, protein kinase R (PKR) and general control nonderepressible 2 (GCN2), and by Janus kinase signalling. Conclusions The DN-PSMB8 p.G209R variant broadens the clinical and mechanistic PRAAS spectrum by causing 20S proteasome maturation arrest and uncovering a convergence between mitochondrial dysfunction and the ISR. Our findings implicate cytopenia in a pattern of vascular pathology, including PSVD of the liver. We further identify PKR and GCN2 as key mediators of maladaptive IFN-I responses and potential therapeutic targets.

Human recombination-activating gene (RAG) deficiency can manifest with distinct clinical and immunological phenotypes. By applying a multiomics approach to a large group of RAG-mutated patients, we aimed at characterizing the immunopathology associated with each phenotype. Although defective T and B cell development is common to all phenotypes, patients with hypomorphic RAG variants can generate T and B cells with signatures of immune dysregulation and produce autoantibodies to a broad range of self-antigens, including type I interferons. T helper 2 (TH2) cell skewing and a prominent inflammatory signature characterize Omenn syndrome, whereas more hypomorphic forms of RAG deficiency are associated with a type 1 immune profile both in blood and tissues. We used cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) analysis to define the cell lineage-specific contribution to the immunopathology of the distinct RAG phenotypes. These insights may help improve the diagnosis and clinical management of the various forms of the disease.

Cutaneous squamous-cell carcinoma (SCC) is primarily caused by oncogenesis mediated by ultraviolet radiation, and β-human papillomavirus (β-HPV) is believed to be a mere facilitator that is dispensable for the maintenance of cutaneous SCC. Here, we describe a woman with benign and malignant HPV-related diseases that include a recurrent, unresectable, invasive cutaneous SCC with β-HPV19 genomic integration in the context of germline pathogenic mutations in ZAP70, an adapter required for T-cell receptor (TCR) signal transduction. Restoration of the integrity of TCR signaling by allogeneic hematopoietic-cell transplantation led to the resolution of all HPV-related diseases, thereby revealing a direct role of β-HPV in skin carcinogenesis in hosts with defective adaptive T-cell responses. (Funded by the National Institutes of Health.).

The United States Immunodeficiency Network (USIDNET) is an NIH-funded research consortium that advances scientific investigation in the field of inborn errors of immunity (IEI). Capturing demographic diversity within the patient registry is important for improving research on IEI and supporting health equity in clinical immunology. Formerly, registry data were collected via an opt-in system with patient informed consent. Now, the registry uses semi-automated de-identified data extraction from EPIC with a consent waiver. This study was performed to assess whether the new ascertainment method improved diversity using data from one site. De-identified data contributed from CHOP was reviewed from both the old and the new USIDNET registries. Race, ethnicity, sex, and age within the new (n = 1,145) and old (n = 551) registries were defined. Patients without data for a given demographic (i.e. race) were excluded from the analysis for that demographic. Chi-squared and Fisher’s exact test were completed using GraphPad Prism to determine statistical significance. The racial breakdown of the new registry was: 0.18% American Indian or Alaska Native, 2.55% Asian, 6.94% Black, 72.93% White, and 17.40% other or more than one race. This was a significant (p < 0.0001) difference in racial distribution compared to the old registry: 0.78% Asian or Pacific Islander, 10.85% Black, 86.05% White, and 2.33% other or more than one race. Relative to the old registry, the new registry had a significantly higher proportion of patients from racial minority backgrounds for the diagnosis of agammaglobulinemia (p = 0.0194), but not for severe combined immunodeficiency (p = 0.3937) or chronic granulomatous disease (p = 0.7305). The new registry trended toward a younger median age: 17 years old (0-54) versus 28 years old (9-72). The design of the new USIDNET registry may better capture a greater representation of patients from racial minority backgrounds compared to the old registry. However, the proportion of patients from racial minority backgrounds within the registry continues to be lower than the populations seen in either total or immunology department CHOP patient visits. While there are many contributing factors, our data may indicate disparities in accessing tertiary care or receiving an IEI diagnosis.