Primary immunodeficiency (PID) is characterised by recurrent and often life-threatening infections, autoimmunity and cancer, and it presents major diagnostic and therapeutic challenges. Although the most severe forms present in early childhood, the majority of patients present in adulthood, typically with no apparent family history and a variable clinical phenotype of widespread immune dysregulation: about 25% of patients have autoimmune disease, allergy is prevalent, and up to 10% develop lymphoid malignancies 1 – 3 . Consequently, in sporadic PID genetic diagnosis is difficult and the role of genetics is not well defined. We addressed these challenges by performing whole genome sequencing (WGS) of a large PID cohort of 1,318 participants. Analysis of coding regions of 886 index cases found disease-causing mutations in known monogenic PID genes in 10.3%, while a Bayesian approach (BeviMed 4 ) identified multiple potential new candidate genes, including IVNS1ABP . Exploration of the non-coding genome revealed deletions in regulatory regions which contribute to disease causation. Finally, a genome-wide association study (GWAS) identified PID-associated loci and uncovered evidence for co-localisation of, and interplay between, novel high penetrance monogenic variants and common variants (at the PTPN2 and SOCS1 loci). This begins to explain the contribution of common variants to variable penetrance and phenotypic complexity in PID. Thus, a cohort-based WGS approach to PID diagnosis can increase diagnostic yield while deepening our understanding of the key pathways influencing human immune responsiveness.

VarFish is a user-friendly web application for the quality control, filtering, prioritization, analysis, and user-based annotation of DNA variant data with a focus on rare disease genetics. It is capable of processing variant call files with single or multiple samples. The variants are automatically annotated with population frequencies, molecular impact, and presence in databases such as ClinVar. Further, it provides support for pathogenicity scores including CADD, MutationTaster, and phenotypic similarity scores. Users can filter variants based on these annotations and presumed inheritance pattern and sort the results by these scores. Variants passing the filter are listed with their annotations and many useful link-outs to genome browsers, other gene/variant data portals, and external tools for variant assessment. VarFish allows users to create their own annotations including support for variant assessment following ACMG-AMP guidelines. In close collaboration with medical practitioners, VarFish was designed for variant analysis and prioritization in diagnostic and research settings as described in the software's extensive manual. The user interface has been optimized for supporting these protocols. Users can install VarFish on their own in-house servers where it provides additional lab notebook features for collaborative analysis and allows re-analysis of cases, e.g. after update of genotype or phenotype databases.

Background: Kynurenine, a metabolite of the L-tryptophan pathway, plays a pivotal role in neuro-inflammation, cancer immunology, and cardiovascular inflammation, and has been shown to predict cardiovascular events. Objectives: It was our objective to increase the body of data regarding the value of kynurenine as a biomarker in chronic heart failure (CHF). Methods: We investigated the predictive value of plasma kynurenine in a CHF cohort (CHF, n = 114); in a second cohort of defibrillator carriers with CHF (AICD, n = 156), we determined clinical and biochemical determinants of the marker which was measured by enzyme immunoassay. Results: In the CHF cohort, both kynurenine and NT-proBNP increased with NYHA class. Univariate binary logistic regression showed kynurenine to predict death within a 6-month follow-up (OR 1.43, 95% CI 1.03–2.00, p = 0.033) whereas NT-proBNP did not contribute significantly. Kynurenine, like NT-proBNP, was able to discriminate at a 30% threshold of left ventricular ejection fraction (LVEF; AUC-ROC, both 0.74). Kynurenine correlated inversely with LVEF (ϱ = –0.394), glomerular filtration fraction (GFR; ϱ = –0.615), and peak VO2 (ϱ = –0.626). Moreover, there was a strong correlation of kynurenine with NT-proBNP (ϱ = 0.615). In the AICD cohort, multiple linear regression analysis demonstrated highly significant associations of kynurenine with GFR, hsCRP, and tryptophan, as well as a significant impact of age. Conclusions: This work speaks in favor of kynurenine being a new and valuable biomarker of CHF, with particular attention placed on its ability to predict mortality and reflect exercise capacity.

TRPM3 is a temperature- and neurosteroid-sensitive plasma membrane cation channel expressed in a variety of neuronal and non-neuronal cells. Recently, rare de novo variants in TRPM3 were identified in individuals with developmental and epileptic encephalopathy (DEE), but the link between TRPM3 activity and neuronal disease remains poorly understood. We previously reported that two disease-associated variants in TRPM3 lead to a gain of channel function (Van Hoeymissen et al., 2020; Zhao et al., 2020). Here, we report a further ten patients carrying one of seven additional heterozygous TRPM3 missense variants. These patients present with a broad spectrum of neurodevelopmental symptoms, including global developmental delay, intellectual disability, epilepsy, musculo-skeletal anomalies, and altered pain perception. We describe a cerebellar phenotype with ataxia or severe hypotonia, nystagmus, and cerebellar atrophy in more than half of the patients. All disease-associated variants exhibited a robust gain-of-function phenotype, characterized by increased basal activity leading to cellular calcium overload and by enhanced responses to the neurosteroid ligand pregnenolone sulphate, when co-expressed with wild-type TRPM3 in mammalian cells. The antiseizure medication primidone, a known TRPM3 antagonist, reduced the increased basal activity of all mutant channels. These findings establish gain-of-function of TRPM3 as the cause of a spectrum of autosomal dominant neurodevelopmental disorders with frequent cerebellar involvement in humans, and provide support for the evaluation of TRPM3 antagonists as a potential therapy.

BackgroundInborn errors of immunity (IEI) present with a large phenotypic spectrum of disease, which can pose diagnostic and therapeutic challenges. Suppressor of cytokine signaling 1 (SOCS1) is a key negative regulator of cytokine signaling, and has recently been associated with a novel IEI. Of patients described to date, it is apparent that SOCS1 haploinsufficiency has a pleiotropic effect in humans.ObjectiveWe sought to investigate whether dysregulation of immune pathways, in addition to STAT1, play a role in the broad clinical manifestations of SOCS1 haploinsufficiency.MethodsWe assessed impacts of reduced SOCS1 expression across multiple immune cell pathways utilizing patient cells and CRISPR/Cas9 edited primary human T cells.ResultsSOCS1 haploinsufficiency phenotypes straddled across the International Union of Immunological Societies classifications of IEI. We found that reduced SOCS1 expression led to dysregulation of multiple intracellular pathways in immune cells. STAT1 phosphorylation is enhanced, comparably with STAT1 gain-of-function mutations, and STAT3 phosphorylation is similarly reduced with concurrent reduction of Th17 cells. Furthermore, reduced SOCS1 E3 ligase function was associated with increased FAK1 in immune cells, and increased AKT and p70 ribosomal protein S6 kinase phosphorylation. We also found Toll-like receptor responses are increased in SOCS1 haploinsufficiency patients.ConclusionsSOCS1 haploinsufficiency is a pleiotropic monogenic IEI. Dysregulation of multiple immune cell pathways may explain the variable clinical phenotype associated with this new condition. Knowledge of these additional dysregulated immune pathways is important when considering the optimum management for SOCS1 haploinsufficient patients.

Biallelic variants in PUS3 have recently been recognized as a rare cause of neurodevelopmental disorders. Pseudouridine synthase-3 encoded by PUS3 is an enzyme important for modification of various RNAs, including transfer RNA (tRNA). Here we present the clinical and genetic features of 21 individuals with biallelic PUS3 variants: seven new and 14 previously reported individuals, where clinical features of Miriam Nøstvik, Sarah M. Kateta and Bitten Schönewolf-Greulich shared first authorship. Rikke S. Møller and Zeynep Tümer shared senior authorship.

The inverse associations of zonulin with creatinine and markers of cardio-vascular risk (high CMLcalprotectin and kynurenine, low homoarginine) are novel findings that need further experimental and clinical clarification. Our study indicates zonulin involvement in metabolic inflammation in T2D, but no association with disease status in CHF.

Neurodevelopmental disorder with dysmorphic facies and distal limb anomalies (NEDDFL), defined primarily by developmental delay/intellectual disability, speech delay, postnatal microcephaly, and dysmorphic features, is a syndrome resulting from heterozygous variants in the dosage‐sensitive bromodomain PHD finger chromatin remodeler transcription factor BPTF gene. To date, only 11 individuals with NEDDFL due to de novo BPTF variants have been described. To expand the NEDDFL phenotypic spectrum, we describe the clinical features in 25 novel individuals with 20 distinct, clinically relevant variants in BPTF, including four individuals with inherited changes in BPTF. In addition to the previously described features, individuals in this cohort exhibited mild brain abnormalities, seizures, scoliosis, and a variety of ophthalmologic complications. These results further support the broad and multi‐faceted complications due to haploinsufficiency of BPTF.