Individuals with JAK1 variants are affected by syndromic features encompassing autoimmunity, atopy, colitis, and dermatitis.

Inborn errors of immunity lead to autoimmunity, inflammation, allergy, infection, and/or malignancy. Disease-causing JAK1 gain-of-function (GoF) mutations are considered exceedingly rare and have been identified in only four families. Here, we use forward and reverse genetics to identify 59 individuals harboring one of four heterozygous JAK1 variants. In vitro and ex vivo analysis of these variants revealed hyperactive baseline and cytokine-induced STAT phosphorylation and interferon-stimulated gene (ISG) levels compared with wild-type JAK1. A systematic review of electronic health records from the BioME Biobank revealed increased likelihood of clinical presentation with autoimmunity, atopy, colitis, and/or dermatitis in JAK1 variant-positive individuals. Finally, treatment of one affected patient with severe atopic dermatitis using the JAK1/JAK2-selective inhibitor, baricitinib, resulted in clinically significant improvement. These findings suggest that individually rare JAK1 GoF variants may underlie an emerging syndrome with more common presentations of autoimmune and inflammatory disease (JAACD syndrome). More broadly, individuals who present with such conditions may benefit from genetic testing for the presence of JAK1 GoF variants.

Development of a human genetics-guided priority score for 19,365 genes and 399 drug indications.

Studies have shown that drug targets with human genetic support are more likely to succeed in clinical trials. Hence, a tool integrating genetic evidence to prioritize drug target genes is beneficial for drug discovery. We built a genetic priority score (GPS) by integrating eight genetic features with drug indications from the Open Targets and SIDER databases. The top 0.83%, 0.28% and 0.19% of the GPS conferred a 5.3-, 9.9- and 11.0-fold increased effect of having an indication, respectively. In addition, we observed that targets in the top 0.28% of the score were 1.7-, 3.7- and 8.8-fold more likely to advance from phase I to phases II, III and IV, respectively. Complementary to the GPS, we incorporated the direction of genetic effect and drug mechanism into a directional version of the score called the GPS with direction of effect. We applied our method to 19,365 protein-coding genes and 399 drug indications and made all results available through a web portal.

Genome-wide prediction of pathogenic gain- and loss-of-function variants from ensemble learning of a diverse feature set.

Gain-of-function (GOF) variants give rise to increased/novel protein functions whereas loss-of-function (LOF) variants lead to diminished protein function. Experimental approaches for identifying GOF and LOF are generally slow and costly, whilst available computational methods have not been optimized to discriminate between GOF and LOF variants. We have developed LoGoFunc, a machine learning method for predicting pathogenic GOF, pathogenic LOF, and neutral genetic variants, trained on a broad range of gene-, protein-, and variant-level features describing diverse biological characteristics. LoGoFunc outperforms other tools trained solely to predict pathogenicity for identifying pathogenic GOF and LOF variants and is available at https://itanlab.shinyapps.io/goflof/ .

Genetic susceptibility to diabetic kidney disease is linked to promoter variants of XOR.

The lifetime risk of kidney disease in people with diabetes is 10-30%, implicating genetic predisposition in the cause of diabetic kidney disease (DKD). Here we identify an expression quantitative trait loci (QTLs) in the cis-acting regulatory region of the xanthine dehydrogenase, or xanthine oxidoreductase (Xor), a binding site for C/EBPß, to be associated with diabetes-induced podocyte loss in DKD in male mice. We examine mouse inbred strains that are susceptible (DBA/2J) and resistant (C57BL/6J) to DKD, as well as a panel of recombinant inbred BXD mice, to map QTLs. We also uncover promoter XOR orthologue variants in humans associated with high risk of DKD. We introduced the risk variant into the 5'-regulatory region of XOR in DKD-resistant mice, which resulted in increased Xor activity associated with podocyte depletion, albuminuria, oxidative stress and damage restricted to the glomerular endothelium, which increase further with type 1 diabetes, high-fat diet and ageing. Therefore, differential regulation of Xor contributes to phenotypic consequences with diabetes and ageing.

Identifying high-impact variants and genes in exomes of Ashkenazi Jewish inflammatory bowel disease patients.

Inflammatory bowel disease (IBD) is a group of chronic digestive tract inflammatory conditions whose genetic etiology is still poorly understood. The incidence of IBD is particularly high among Ashkenazi Jews. Here, we identify 8 novel and plausible IBD-causing genes from the exomes of 4453 genetically identified Ashkenazi Jewish IBD cases (1734) and controls (2719). Various biological pathway analyses are performed, along with bulk and single-cell RNA sequencing, to demonstrate the likely physiological relatedness of the novel genes to IBD. Importantly, we demonstrate that the rare and high impact genetic architecture of Ashkenazi Jewish adult IBD displays significant overlap with very early onset-IBD genetics. Moreover, by performing biobank phenome-wide analyses, we find that IBD genes have pleiotropic effects that involve other immune responses. Finally, we show that polygenic risk score analyses based on genome-wide high impact variants have high power to predict IBD susceptibility.

Identifying shared genetic factors underlying epilepsy and congenital heart disease in Europeans.

Epilepsy (EP) and congenital heart disease (CHD) are two apparently unrelated diseases that nevertheless display substantial mutual comorbidity. Thus, while congenital heart defects are associated with an elevated risk of developing epilepsy, the incidence of epilepsy in CHD patients correlates with CHD severity. Although genetic determinants have been postulated to underlie the comorbidity of EP and CHD, the precise genetic etiology is unknown. We performed variant and gene association analyses on EP and CHD patients separately, using whole exomes of genetically identified Europeans from the UK Biobank and Mount Sinai BioMe Biobank. We prioritized biologically plausible candidate genes and investigated the enriched pathways and other identified comorbidities by biological proximity calculation, pathway analyses, and gene-level phenome-wide association studies. Our variant- and gene-level results point to the Voltage-Gated Calcium Channels (VGCC) pathway as being a unifying framework for EP and CHD comorbidity. Additionally, pathway-level analyses indicated that the functions of disease-associated genes partially overlap between the two disease entities. Finally, phenome-wide association analyses of prioritized candidate genes revealed that cerebral blood flow and ulcerative colitis constitute the two main traits associated with both EP and CHD.

Novel brown adipose tissue candidate genes predicted by the human gene connectome.

Brown adipose tissue (BAT) is a promising therapeutic target against obesity. Therefore, research on the genetic architecture of BAT could be key for the development of successful therapies against this complex phenotype. Hypothesis-driven candidate gene association studies are useful for studying genetic determinants of complex traits, but they are dependent upon the previous knowledge to select candidate genes. Here, we predicted 107 novel-BAT candidate genes in silico using the uncoupling protein one (UCP1) as the hallmark of BAT activity. We first identified the top 1% of human genes predicted by the human gene connectome to be biologically closest to the UCP1, estimating 167 additional pathway genes (BAT connectome). We validated this prediction by showing that 60 genes already associated with BAT were included in the connectome and they were biologically closer to each other than expected by chance (p < 2.2 × 10-16). The rest of genes (107) are potential candidates for BAT, being also closer to known BAT genes and more expressed in BAT biopsies than expected by chance (p < 2.2 × 10-16; p = 4.39 × 10-02). The resulting new list of predicted human BAT genes should be useful for the discovery of novel BAT genes and metabolic pathways.

VIPPID: a gene-specific single nucleotide variant pathogenicity prediction tool for primary immunodeficiency diseases.

Distinguishing pathogenic variants from non-pathogenic ones remains a major challenge in clinical genetic testing of primary immunodeficiency (PID) patients. Most of the existing mutation pathogenicity prediction tools treat all mutations as homogeneous entities, ignoring the differences in characteristics of different genes, and use the same model for genes in different diseases. In this study, we developed a single nucleotide variant (SNV) pathogenicity prediction tool, Variant Impact Predictor for PIDs (VIPPID; https://mylab.shinyapps.io/VIPPID/), which was tailored for PIDs genes and used a specific model for each of the most prevalent PID known genes. It employed a Conditional Inference Forest model and utilized information of 85 features of SNVs and scores from 20 existing prediction tools. Evaluation of VIPPID showed that it had superior performance (area under the curve = 0.91) over non-specific conventional tools. In addition, we also showed that the gene-specific model outperformed the non-gene-specific models. Our study demonstrated that disease-specific and gene-specific models can improve SNV pathogenicity prediction performance. This observation supports the notion that each feature of mutations in the model can be potentially used, in a new algorithm, to investigate the characteristics and function of the encoded proteins.

Identification of discriminative gene-level and protein-level features associated with pathogenic gain-of-function and loss-of-function variants.

Identifying whether a given genetic mutation results in a gene product with increased (gain-of-function; GOF) or diminished (loss-of-function; LOF) activity is an important step toward understanding disease mechanisms because they may result in markedly different clinical phenotypes. Here, we generated an extensive database of documented germline GOF and LOF pathogenic variants by employing natural language processing (NLP) on the available abstracts in the Human Gene Mutation Database. We then investigated various gene- and protein-level features of GOF and LOF variants and applied machine learning and statistical analyses to identify discriminative features. We found that GOF variants were enriched in essential genes, for autosomal-dominant inheritance, and in protein binding and interaction domains, whereas LOF variants were enriched in singleton genes, for protein-truncating variants, and in protein core regions. We developed a user-friendly web-based interface that enables the extraction of selected subsets from the GOF/LOF database by a broad set of annotated features and downloading of up-to-date versions. These results improve our understanding of how variants affect gene/protein function and may ultimately guide future treatment options.

The genetic structure of the Turkish population reveals high levels of variation and admixture.

The construction of population-based variomes has contributed substantially to our understanding of the genetic basis of human inherited disease. Here, we investigated the genetic structure of Turkey from 3,362 unrelated subjects whose whole exomes (n = 2,589) or whole genomes (n = 773) were sequenced to generate a Turkish (TR) Variome that should serve to facilitate disease gene discovery in Turkey. Consistent with the history of present-day Turkey as a crossroads between Europe and Asia, we found extensive admixture between Balkan, Caucasus, Middle Eastern, and European populations with a closer genetic relationship of the TR population to Europeans than hitherto appreciated. We determined that 50% of TR individuals had high inbreeding coefficients (≥0.0156) with runs of homozygosity longer than 4 Mb being found exclusively in the TR population when compared to 1000 Genomes Project populations. We also found that 28% of exome and 49% of genome variants in the very rare range (allele frequency < 0.005) are unique to the modern TR population. We annotated these variants based on their functional consequences to establish a TR Variome containing alleles of potential medical relevance, a repository of homozygous loss-of-function variants and a TR reference panel for genotype imputation using high-quality haplotypes, to facilitate genome-wide association studies. In addition to providing information on the genetic structure of the modern TR population, these data provide an invaluable resource for future studies to identify variants that are associated with specific phenotypes as well as establishing the phenotypic consequences of mutations in specific genes.

Sudden Cardiac Arrest in a Patient With Mitral Valve Prolapse and LMNA and SCN5A Mutations.

Bileaflet mitral valve prolapse (Bi-MVP) is associated with increased risk for cardiac arrest. We describe a patient who presented after a cardiac arrest with Bi-MVP and variants in Lamin A/C (LMNA) and the sodium channel alpha-subunit 5a (SCN5A). Genetic variants may be the culprit for arrhythmogenesis in Bi-MVP patients. (Level of Difficulty: Intermediate.).

Burden of Cardiomyopathic Genetic Variation in Lethal Pediatric Myocarditis.

BACKGROUND: Acute myocarditis (AM) is a well-known cause of sudden death and heart failure, often caused by prevalent viruses. We previously showed that some pediatric AM correlates with putatively damaging variants in genes related to cardiomyocyte structure and function. We sought to evaluate whether deleterious cardiomyopathic variants were enriched among fatal pediatric AM cases in New York City compared with ancestry-matched controls. METHODS: Twenty-four children (aged 3 weeks to 20 years) with death due to AM were identified through autopsy records; histologies were reviewed to confirm that all cases met Dallas criteria for AM and targeted panel sequencing of 57 cardiomyopathic genes was performed. Controls without cardiovascular disease were identified from a pediatric database and matched by genetic ancestry to cases using principal components from exome sequencing. Rates of putative deleterious variations (DV) were compared between cases and controls. Where available, AM tissues underwent viral analysis by polymerase chain reaction. RESULTS: DV were identified in 4 of 24 AM cases (16.7%), compared with 2 of 96 age and ancestry-matched controls (2.1%, P=0.014). Viral causes were proven for 6 of 8 AM cases (75%), including the one DV+ case where tissue was available for testing. DV+ cases were more likely to be female, have no evidence of chronic inflammation, and associate with sudden cardiac death than DV- cases. CONCLUSIONS: Deleterious variants in genes related to cardiomyocyte integrity are more common in children with fatal AM than controls, likely conferring susceptibility. Additionally, genetically mediated AM may progress more rapidly and be more severe.

A computational approach for detecting physiological homogeneity in the midst of genetic heterogeneity.

The human genetic dissection of clinical phenotypes is complicated by genetic heterogeneity. Gene burden approaches that detect genetic signals in case-control studies are underpowered in genetically heterogeneous cohorts. We therefore developed a genome-wide computational method, network-based heterogeneity clustering (NHC), to detect physiological homogeneity in the midst of genetic heterogeneity. Simulation studies showed our method to be capable of systematically converging genes in biological proximity on the background biological interaction network, and capturing gene clusters harboring presumably deleterious variants, in an efficient and unbiased manner. We applied NHC to whole-exome sequencing data from a cohort of 122 individuals with herpes simplex encephalitis (HSE), including 13 individuals with previously published monogenic inborn errors of TLR3-dependent IFN-α/ß immunity. The top gene cluster identified by our approach successfully detected and prioritized all causal variants of five TLR3 pathway genes in the 13 previously reported individuals. This approach also suggested candidate variants of three reported genes and four candidate genes from the same pathway in another ten previously unstudied individuals. TLR3 responsiveness was impaired in dermal fibroblasts from four of the five individuals tested, suggesting that the variants detected were causal for HSE. NHC is, therefore, an effective and unbiased approach for unraveling genetic heterogeneity by detecting physiological homogeneity.

Genome-wide discovery of genetic loci that uncouple excess adiposity from its comorbidities.

Obesity is a major risk factor for cardiometabolic diseases. Nevertheless, a substantial proportion of individuals with obesity do not suffer cardiometabolic comorbidities. The mechanisms that uncouple adiposity from its cardiometabolic complications are not fully understood. Here, we identify 62 loci of which the same allele is significantly associated with both higher adiposity and lower cardiometabolic risk. Functional analyses show that the 62 loci are enriched for genes expressed in adipose tissue, and for regulatory variants that influence nearby genes that affect adipocyte differentiation. Genes prioritized in each locus support a key role of fat distribution (FAM13A, IRS1 and PPARG) and adipocyte function (ALDH2, CCDC92, DNAH10, ESR1, FAM13A, MTOR, PIK3R1 and VEGFB). Several additional mechanisms are involved as well, such as insulin-glucose signalling (ADCY5, ARAP1, CREBBP, FAM13A, MTOR, PEPD, RAC1 and SH2B3), energy expenditure and fatty acid oxidation (IGF2BP2), browning of white adipose tissue (CSK, VEGFA, VEGFB and SLC22A3) and inflammation (SH2B3, DAGLB and ADCY9). Some of these genes may represent therapeutic targets to reduce cardiometabolic risk linked to excess adiposity.

Inflamed Ulcerative Colitis Regions Associated With MRGPRX2-Mediated Mast Cell Degranulation and Cell Activation Modules, Defining a New Therapeutic Target.

BACKGROUND & AIMS: Recent literature has implicated a key role for mast cells in murine models of colonic inflammation, but their role in human ulcerative colitis (UC) is not well established. A major advance has been the identification of mrgprb2 (human orthologue, MRGPX2) as mediating IgE-independent mast cell activation. We sought to define mechanisms of mast cell activation and MRGPRX2 in human UC. METHODS: Colon tissues were collected from patients with UC for bulk RNA sequencing and lamina propria cells were isolated for MRGPRX2 activation studies and single-cell RNA sequencing. Genetic association of all protein-altering G-protein coupled receptor single-nucleotide polymorphism was performed in an Ashkenazi Jewish UC case-control cohort. Variants of MRGPRX2 were transfected into Chinese hamster ovary (CHO) and human mast cell (HMC) 1.1 cells to detect genotype-dependent effects on ß-arrestin recruitment, IP-1 accumulation, and phosphorylated extracellular signal-regulated kinase. RESULTS: Mast cell-specific mediators and adrenomedullin (proteolytic precursor of PAMP-12, an MRGPRX2 agonist) are up-regulated in inflamed compared to uninflamed UC. MRGPRX2 stimulation induces carboxypeptidase secretion from inflamed UC. Of all protein-altering GPCR alleles, a unique variant of MRGPRX2, Asn62Ser, was most associated with and was bioinformatically predicted to alter arrestin recruitment. We validated that the UC protective serine allele enhances ß-arrestin recruitment, decreases IP-1, and increases phosphorylated extracellular signal-regulated kinase with MRGPRX2 agonists. Single-cell RNA sequencing defines that adrenomedullin is expressed by activated fibroblasts and epithelial cells and that interferon gamma is a key upstream regulator of mast cell gene expression. CONCLUSION: Inflamed UC regions are distinguished by MRGPRX2-mediated activation of mast cells, with decreased activation observed with a UC-protective genetic variant. These results define cell modules of UC activation and a new therapeutic target.

TLR3 controls constitutive IFN-ß antiviral immunity in human fibroblasts and cortical neurons.

Human herpes simplex virus 1 (HSV-1) encephalitis can be caused by inborn errors of the TLR3 pathway, resulting in impairment of CNS cell-intrinsic antiviral immunity. Deficiencies of the TLR3 pathway impair cell-intrinsic immunity to vesicular stomatitis virus (VSV) and HSV-1 in fibroblasts, and to HSV-1 in cortical but not trigeminal neurons. The underlying molecular mechanism is thought to involve impaired IFN-α/ß induction by the TLR3 recognition of dsRNA viral intermediates or by-products. However, we show here that human TLR3 controls constitutive levels of IFNB mRNA and secreted bioactive IFN-ß protein, and thereby also controls constitutive mRNA levels for IFN-stimulated genes (ISGs) in fibroblasts. Tlr3-/- mouse embryonic fibroblasts also have lower basal ISG levels. Moreover, human TLR3 controls basal levels of IFN-ß secretion and ISG mRNA in induced pluripotent stem cell-derived cortical neurons. Consistently, TLR3-deficient human fibroblasts and cortical neurons are vulnerable not only to both VSV and HSV-1, but also to several other families of viruses. The mechanism by which TLR3 restricts viral growth in human fibroblasts and cortical neurons in vitro and, by inference, by which the human CNS prevents infection by HSV-1 in vivo, is therefore based on the control of early viral infection by basal IFN-ß immunity.

Common and Rare Variant Prediction and Penetrance of IBD in a Large, Multi-ethnic, Health System-based Biobank Cohort.

BACKGROUND AND AIMS: Polygenic risk scores (PRS) may soon be used to predict inflammatory bowel disease (IBD) risk in prevention efforts. We leveraged exome-sequence and single nucleotide polymorphism (SNP) array data from 29,358 individuals in the multiethnic, randomly ascertained health system-based BioMe biobank to define effects of common and rare IBD variants on disease prediction and pathophysiology. METHODS: PRS were calculated from European, African American, and Ashkenazi Jewish (AJ) reference case-control studies, and a meta-GWAS run using all three association datasets. PRS were then combined using regression to assess which combination of scores best predicted IBD status in European, AJ, Hispanic, and African American cohorts in BioMe. Additionally, rare variants were assessed in genes associated with very early-onset IBD (VEO-IBD), by estimating genetic penetrance in each BioMe population. RESULTS: Combining risk scores based on association data from distinct ancestral populations improved IBD prediction for every population in BioMe and significantly improved prediction among European ancestry UK Biobank individuals. Lower predictive power for non-Europeans was observed, reflecting in part substantially lower African IBD case-control reference sizes. We replicated associations for two VEO-IBD genes, ADAM17 and LRBA, with high dominant model penetrance in BioMe. Autosomal recessive LRBA risk alleles are associated with severe, early-onset autoimmunity; we show that heterozygous carriage of an African-predominant LRBA protein-altering allele is associated with significantly decreased LRBA and CTLA-4 expression with T-cell activation. CONCLUSIONS: Greater genetic diversity in African populations improves prediction across populations, and generalizes some VEO-IBD genes. Increasing African American IBD case-collections should be prioritized to reduce health disparities and enhance pathophysiological insight.

Human CRY1 variants associate with attention deficit/hyperactivity disorder.

Attention deficit/hyperactivity disorder (ADHD) is a common and heritable phenotype frequently accompanied by insomnia, anxiety, and depression. Here, using a reverse phenotyping approach, we report heterozygous coding variations in the core circadian clock gene cryptochrome 1 in 15 unrelated multigenerational families with combined ADHD and insomnia. The variants led to functional alterations in the circadian molecular rhythms, providing a mechanistic link to the behavioral symptoms. One variant, CRY1Δ11 c.1657+3A>C, is present in approximately 1% of Europeans, therefore standing out as a diagnostic and therapeutic marker. We showed by exome sequencing in an independent cohort of patients with combined ADHD and insomnia that 8 of 62 patients and 0 of 369 controls carried CRY1Δ11. Also, we identified a variant, CRY1Δ6 c.825+1G>A, that shows reduced affinity for BMAL1/CLOCK and causes an arrhythmic phenotype. Genotype-phenotype correlation analysis revealed that this variant segregated with ADHD and delayed sleep phase disorder (DSPD) in the affected family. Finally, we found in a phenome-wide association study involving 9438 unrelated adult Europeans that CRY1Δ11 was associated with major depressive disorder, insomnia, and anxiety. These results defined a distinctive group of circadian psychiatric phenotypes that we propose to designate as "circiatric" disorders.

Non-syndromic Oculocutaneous Albinism: Novel Genetic Variants and Clinical Follow Up of a Brazilian Pediatric Cohort.

Oculocutaneous albinism (OCA) is a genetic disorder characterized by skin, hair, and eye hypopigmentation due to a reduction or absence of melanin. Clinical manifestations include vision problems and a high susceptibility to skin cancer. In its non-syndromic form, OCA is associated with six genes and one chromosomal region. Because OCA subtypes are not always clinically distinguishable, molecular analysis has become an important tool for classifying types of OCA, which facilitates genetic counseling and can guide the development of new therapies. We studied eight Brazilian individuals aged 1.5-18 years old with clinical diagnosis of OCA. Assessment of ophthalmologic characteristics showed results consistent with albinism, including reduced visual acuity, nystagmus, and loss of stereoscopic vision. We also observed the appearance of the strabismus and changes in static refraction over a 2-year period. Dermatologic evaluation showed that no participants had preneoplastic skin lesions, despite half of the participants reporting insufficient knowledge about skin care in albinism. Whole-exome and Sanger sequencing revealed eight different mutations: six in the TYR gene and two in the SLC45A2 gene, of which one was novel and two were described in a population study but were not previously associated with the OCA phenotype. We performed two ophthalmological evaluations, 2 years apart; and one dermatological evaluation. To the best of our knowledge, this is the first study to perform clinical follow-up and genetic analysis of a Brazilian cohort with albinism. Here, we report three new OCA causing mutations.

Identifying disease-causing mutations in genomes of single patients by computational approaches.

Over the last decade next generation sequencing (NGS) has been extensively used to identify new pathogenic mutations and genes causing rare genetic diseases. The efficient analyses of NGS data is not trivial and requires a technically and biologically rigorous pipeline that addresses data quality control, accurate variant filtration to minimize false positives and false negatives, and prioritization of the remaining genes based on disease genomics and physiological knowledge. This review provides a pipeline including all these steps, describes popular software for each step of the analysis, and proposes a general framework for the identification of causal mutations and genes in individual patients of rare genetic diseases.