RESUMEN
Pathogenic variants in the JAG1 gene are a primary cause of the multi-system disorder Alagille syndrome. Although variant detection rates are high for this disease, there is uncertainty associated with the classification of missense variants that leads to reduced diagnostic yield. Consequently, up to 85% of reported JAG1 missense variants have uncertain or conflicting classifications. We generated a library of 2,832 JAG1 nucleotide variants within exons 1-7, a region with a high number of reported missense variants, and designed a high-throughput assay to measure JAG1 membrane expression, a requirement for normal function. After calibration using a set of 175 known or predicted pathogenic and benign variants included within the variant library, 486 variants were characterized as functionally abnormal (n = 277 abnormal and n = 209 likely abnormal), of which 439 (90.3%) were missense. We identified divergent membrane expression occurring at specific residues, indicating that loss of the wild-type residue itself does not drive pathogenicity, a finding supported by structural modeling data and with broad implications for clinical variant classification both for Alagille syndrome and globally across other disease genes. Of 144 uncertain variants reported in patients undergoing clinical or research testing, 27 had functionally abnormal membrane expression, and inclusion of our data resulted in the reclassification of 26 to likely pathogenic. Functional evidence augments the classification of genomic variants, reducing uncertainty and improving diagnostics. Inclusion of this repository of functional evidence during JAG1 variant reclassification will significantly affect resolution of variant pathogenicity, making a critical impact on the molecular diagnosis of Alagille syndrome.
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Síndrome de Alagille , Proteína Jagged-1 , Mutación Missense , Síndrome de Alagille/genética , Proteína Jagged-1/genética , Humanos , Exones/genéticaRESUMEN
Alagille syndrome (ALGS) is an autosomal dominant, multisystemic disorder due to haploinsufficiency in JAG1 or less frequently, mutations in NOTCH2. The disease has been difficult to diagnose and treat due to variable expression. The generation of this iPSC line (TRNDi036-A) carrying a heterozygous mutation (p.Cys693*) in the JAG1 gene provides a means of studying the disease and developing novel therapeutics towards patient treatment.
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Síndrome de Alagille , Heterocigoto , Células Madre Pluripotentes Inducidas , Proteína Jagged-1 , Mutación , Síndrome de Alagille/genética , Humanos , Proteína Jagged-1/genética , Proteína Jagged-1/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Línea Celular , Masculino , FemeninoRESUMEN
BACKGROUND AND AIMS: Alagille syndrome (ALGS) is a multisystem disorder, characterized by cholestasis. Existing outcome data are largely derived from tertiary centers, and real-world data are lacking. This study aimed to elucidate the natural history of liver disease in a contemporary, international cohort of children with ALGS. APPROACH AND RESULTS: This was a multicenter retrospective study of children with a clinically and/or genetically confirmed ALGS diagnosis, born between January 1997 and August 2019. Native liver survival (NLS) and event-free survival rates were assessed. Cox models were constructed to identify early biochemical predictors of clinically evident portal hypertension (CEPH) and NLS. In total, 1433 children (57% male) from 67 centers in 29 countries were included. The 10 and 18-year NLS rates were 54.4% and 40.3%. By 10 and 18 years, 51.5% and 66.0% of children with ALGS experienced ≥1 adverse liver-related event (CEPH, transplant, or death). Children (>6 and ≤12 months) with median total bilirubin (TB) levels between ≥5.0 and <10.0 mg/dl had a 4.1-fold (95% confidence interval [CI], 1.6-10.8), and those ≥10.0 mg/dl had an 8.0-fold (95% CI, 3.4-18.4) increased risk of developing CEPH compared with those <5.0 mg/dl. Median TB levels between ≥5.0 and <10.0 mg/dl and >10.0 mg/dl were associated with a 4.8 (95% CI, 2.4-9.7) and 15.6 (95% CI, 8.7-28.2) increased risk of transplantation relative to <5.0 mg/dl. Median TB <5.0 mg/dl were associated with higher NLS rates relative to ≥5.0 mg/dl, with 79% reaching adulthood with native liver ( p < 0.001). CONCLUSIONS: In this large international cohort of ALGS, only 40.3% of children reach adulthood with their native liver. A TB <5.0 mg/dl between 6 and 12 months of age is associated with better hepatic outcomes. These thresholds provide clinicians with an objective tool to assist with clinical decision-making and in the evaluation of therapies.
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Síndrome de Alagille , Colestasis , Hipertensión Portal , Humanos , Niño , Masculino , Femenino , Síndrome de Alagille/epidemiología , Estudios Retrospectivos , Hipertensión Portal/etiologíaRESUMEN
This special issue of Human Mutation focuses on Innovations in Genomic Diagnostics. The increasing interest in genomic medicine, and the growing possibilities for treatment and management of genetic disease, make complete and accurate diagnosis mission critical. This issue describes leading-edge technologies with emerging utility for genomic diagnostics. Genomic testing has dramatically evolved as a result of advances in technology, data analytics, and the continuing pace of disease gene discovery. Since 2011, clinical laboratories have increasingly employed next-generation sequencing-based tests in addition to historical techniques to identify a spectrum of germline and somatic variants implicated in human disease. However, common testing platforms have known limitations, including failure to detect disease-causing variants in certain regions, inability to identify all variant types, variant phasing, measuring epigenetic changes, and ongoing challenges with variant interpretation. Innovative solutions are emerging, including increasingly rapid genome sequencing, long-read sequencing, clinical RNA sequencing, epigenomic profiling, facial phenotyping, and an array of computational tools for variant identification and interpretation.
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Genoma Humano , Genómica , Genómica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Mutación , Secuenciación del ExomaRESUMEN
Synonymous variants have been shown to alter the correct splicing of pre-mRNAs and generate disease-causing transcripts. These variants are not an uncommon etiology of genetic disease; however, they are frequently overlooked during genetic testing in the absence of functional and clinical data. Here, we describe the occurrence of a synonymous variant [NM_005422.4 (TECTA):c.327C>T, p.(Gly109=)] in seven individuals with hearing loss from six unrelated families. The variant is not located near exonic/intronic boundaries but is predicted to impact splicing by activating a cryptic splicing donor site in exon 4 of TECTA. In vitro minigene assays show that the variant disrupts the reading frame of the canonical transcript, which is predicted to cause a premature termination codon 48 amino acids downstream of the variant, leading to nonsense-mediated decay. The variant is present in population databases, predominantly in Latinos of African ancestry, but is rare in other ethnic groups. Our findings suggest that this synonymous variant is likely pathogenic for TECTA-associated autosomal recessive hearing loss and seems to have arisen as a founder variant in this specific Latino subpopulation. This study demonstrates that synonymous variants need careful splicing assessment and support from additional testing methodologies to determine their clinical impact.
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Sordera , Pérdida Auditiva , Humanos , Sitios de Empalme de ARN , Empalme del ARN/genética , Pérdida Auditiva/genética , Sordera/genética , Exones/genética , Proteínas de la Matriz Extracelular/genética , Proteínas Ligadas a GPI/genéticaRESUMEN
Mitochondrial disease diagnosis requires interrogation of both nuclear and mitochondrial (mtDNA) genomes for single-nucleotide variants (SNVs) and copy number alterations, both in the proband and often maternal relatives, together with careful phenotype correlation. We developed a comprehensive mtDNA sequencing test ('MitoGenome') using long-range PCR (LR-PCR) to amplify the full length of the mtDNA genome followed by next generation sequencing (NGS) to accurately detect SNVs and large-scale mtDNA deletions (LSMD), combined with droplet digital PCR (ddPCR) for LSMD heteroplasmy quantification. Overall, MitoGenome tests were performed on 428 samples from 394 patients with suspected or confirmed mitochondrial disease. The positive yield was 11% (43/394), including 34 patients with pathogenic or likely pathogenic SNVs (the most common being m.3243A > G in 8/34 (24%) patients), 8 patients with single LSMD, and 3 patients with multiple LSMD exceeding 10% heteroplasmy levels. Two patients with both LSMD and pathogenic SNV were detected. Overall, this LR-PCR/NGS assay provides a highly accurate and comprehensive diagnostic method for simultaneous mtDNA SNV detection at heteroplasmy levels as low as 1% and LSMD detection at heteroplasmy levels below 10%. Inclusion of maternal samples for variant classification and ddPCR to quantify LSMD heteroplasmy levels further enables accurate pathogenicity assessment and clinical correlation interpretation of mtDNA genome sequence variants and copy number alterations.
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Genoma Mitocondrial , Enfermedades Mitocondriales , ADN Mitocondrial/genética , Genoma Mitocondrial/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Mitocondrias/genética , Enfermedades Mitocondriales/diagnóstico , Enfermedades Mitocondriales/genéticaRESUMEN
PURPOSE: Detection of all major classes of genomic variants in a single test would decrease cost and increase the efficiency of genomic diagnostics. Genome sequencing (GS) has the potential to provide this level of comprehensive detection. We sought to demonstrate the utility of GS in the molecular diagnosis of 18 patients with clinically defined Alagille syndrome (ALGS), who had a negative or inconclusive result by standard-of-care testing. METHODS: We performed GS on 16 pathogenic variant-negative probands and two probands with inconclusive results (of 406 ALGS probands) and analyzed the data for sequence, copy-number, and structural variants in JAG1 and NOTCH2. RESULTS: GS identified four novel pathogenic alterations including a copy-neutral inversion, a partial deletion, and a promoter variant in JAG1, and a partial NOTCH2 deletion, for an additional diagnostic yield of 0.9%. Furthermore, GS resolved two complex rearrangements, resulting in identification of a pathogenic variant in 97.5% (n = 396/406) of patients after GS. CONCLUSION: GS provided an increased diagnostic yield for individuals with clinically defined ALGS who had prior negative or incomplete genetic testing by other methods. Our results show that GS can detect all major classes of variants and has potential to become a single first-tier diagnostic test for Mendelian disorders.
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Síndrome de Alagille , Síndrome de Alagille/diagnóstico , Síndrome de Alagille/genética , Secuencia de Bases , Mapeo Cromosómico , Pruebas Genéticas , Humanos , Proteína Jagged-1/genéticaRESUMEN
Alagille syndrome (ALGS) is a multisystem autosomal dominant developmental disorder caused predominantly by pathogenic variants in JAGGED1 (JAG1), and also by pathogenic variants in NOTCH2 in a much smaller number of individuals. Clinical presentation is highly variable and includes liver, heart, eye, skeleton, and facial abnormalities, with a subset of individuals also presenting with kidney, vascular, and central nervous system phenotypes. Hepatocellular carcinoma (HCC) is a rare complication of ALGS, though little is known about its incidence or etiology among affected individuals. Previous reports have identified HCC occurrence in both pediatric and adult cases of ALGS. We present a case report of HCC in a 58-year-old woman with a pathogenic JAG1 variant and no overt hepatic features of ALGS. Through a comprehensive literature review, we compile all reported pediatric and adult cases, and further highlight one previously reported case of HCC onset in an adult ALGS patient without any hepatic disease features, similar to our own described patient. Our case report and literature review suggest that ALGS-causing variants could confer risk for developing HCC regardless of phenotypic severity and highlight a need for a cancer screening protocol that would enable early detection and treatment in this at-risk population.
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Síndrome de Alagille/complicaciones , Carcinoma Hepatocelular/etiología , Proteína Jagged-1/genética , Neoplasias Hepáticas/etiología , Mutación , Receptor Notch2/genética , Síndrome de Alagille/genética , Carcinoma Hepatocelular/patología , Femenino , Humanos , Neoplasias Hepáticas/patología , Persona de Mediana Edad , Pronóstico , Literatura de Revisión como AsuntoRESUMEN
Alagille syndrome (ALGS) is an autosomal dominant multisystem disorder with cholestasis as a defining clinical feature. We sought to characterize hepatic outcomes in a molecularly defined cohort of children with ALGS-related cholestasis. Two hundred and ninety-three participants with ALGS with native liver were enrolled. Participants entered the study at different ages and data were collected retrospectively prior to enrollment, and prospectively during the study course. Genetic analysis in 206 revealed JAGGED1 mutations in 91% and NOTCH2 mutations in 4%. Growth was impaired with mean height and weight z-scores of <-1.0 at all ages. Regression analysis revealed that every 10 mg/dL increase in total bilirubin was associated with a decrease in height z-score by 0.10 (P = 0.03) and weight z-score by 0.15 (P = 0.007). Total bilirubin was higher for younger participants (P = 0.03) with a median of 6.9 mg/dL for those less than 1 year old compared with a median of 1.3 mg/dL for participants 13 years or older. The median gamma glutamyl transferase also dropped from 612 to 268 in the same age groups. After adjusting for age, there was substantial within-individual variation of alanine aminotransferase. By 20 years of age, 40% of participants had developed definite portal hypertension. Estimated liver transplant-free survival at the age of 18.5 years was 24%. Conclusions: This is the largest multicenter natural history study of cholestasis in ALGS, demonstrating a previously underappreciated burden of liver disease with early profound cholestasis, a second wave of portal hypertension later in childhood, and less than 25% of patients reaching young adulthood with their native liver. These findings will promote optimization of ALGS management and development of clinically relevant endpoints for future therapeutic trials.
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Powerful, recent advances in technologies to analyze the genome have had a profound impact on the practice of medical genetics, both in the laboratory and in the clinic. Increasing utilization of genome-wide testing such as chromosomal microarray analysis and exome sequencing have lead a shift toward a "genotype-first" approach. Numerous techniques are now available to diagnose a particular syndrome or phenotype, and while traditional techniques remain efficient tools in certain situations, higher-throughput technologies have become the de facto laboratory tool for diagnosis of most conditions. However, selecting the right assay or technology is challenging, and the wrong choice may lead to prolonged time to diagnosis, or even a missed diagnosis. In this review, we will discuss current core technologies for the diagnosis of classic genetic disorders to shed light on the benefits and disadvantages of these strategies, including diagnostic efficiency, variant interpretation, and secondary findings. Finally, we review upcoming technologies posed to impart further changes in the field of genetic diagnostics as we move toward "genome-first" practice.
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Mosaicism denotes an individual who has at least two populations of cells with distinct genotypes that are derived from a single fertilized egg. Genetic variation among the cell lines can involve whole chromosomes, structural or copy-number variants, small or single-nucleotide variants, or epigenetic variants. The mutational events that underlie mosaic variants occur during mitotic cell divisions after fertilization and zygote formation. The initiating mutational event can occur in any types of cell at any time in development, leading to enormous variation in the distribution and phenotypic effect of mosaicism. A number of classification proposals have been put forward to classify genetic mosaicism into categories based on the location, pattern, and mechanisms of the disease. We here propose a new classification of genetic mosaicism that considers the affected tissue, the pattern and distribution of the mosaicism, the pathogenicity of the variant, the direction of the change (benign to pathogenic vs. pathogenic to benign), and the postzygotic mutational mechanism. The accurate and comprehensive categorization and subtyping of mosaicisms is important and has potential clinical utility to define the natural history of these disorders, tailor follow-up frequency and interventions, estimate recurrence risks, and guide therapeutic decisions.
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Variaciones en el Número de Copia de ADN , Mosaicismo , Análisis Mutacional de ADN , Humanos , Mutación , Programas InformáticosRESUMEN
BACKGROUND & AIMS: Extrahepatic biliary atresia (BA) is a pediatric liver disease with no approved medical therapy. Recent studies using human samples and experimental modeling suggest that glutathione redox metabolism and heterogeneity play a role in disease pathogenesis. We sought to dissect the mechanistic basis of liver redox variation and explore how other stress responses affect cholangiocyte injury in BA. METHODS: We performed quantitative in situ hepatic glutathione redox mapping in zebrafish larvae carrying targeted mutations in glutathione metabolism genes and correlated these findings with sensitivity to the plant-derived BA-linked toxin biliatresone. We also determined whether genetic disruption of HSP90 protein quality control pathway genes implicated in human BA altered biliatresone toxicity in zebrafish and human cholangiocytes. An in vivo screening of a known drug library was performed to identify novel modifiers of cholangiocyte injury in the zebrafish experimental BA model, with subsequent validation. RESULTS: Glutathione metabolism gene mutations caused regionally distinct changes in the redox potential of cholangiocytes that differentially sensitized them to biliatresone. Disruption of human BA-implicated HSP90 pathway genes sensitized zebrafish and human cholangiocytes to biliatresone-induced injury independent of glutathione. Phosphodiesterase-5 inhibitors and other cyclic guanosine monophosphate signaling activators worked synergistically with the glutathione precursor N-acetylcysteine in preventing biliatresone-induced injury in zebrafish and human cholangiocytes. Phosphodiesterase-5 inhibitors enhanced proteasomal degradation and required intact HSP90 chaperone. CONCLUSION: Regional variation in glutathione metabolism underlies sensitivity to the biliary toxin biliatresone and may account for the reported association between BA transplant-free survival and glutathione metabolism gene expression. Human BA can be causatively linked to genetic modulation of protein quality control. Combined treatment with N-acetylcysteine and cyclic guanosine monophosphate signaling enhancers warrants further investigation as therapy for BA.
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Conductos Biliares/patología , Atresia Biliar/tratamiento farmacológico , Depuradores de Radicales Libres/farmacología , Oxidación-Reducción/efectos de los fármacos , Proteostasis/efectos de los fármacos , Acetilcisteína/farmacología , Acetilcisteína/uso terapéutico , Animales , Animales Modificados Genéticamente , Benzodioxoles/toxicidad , Conductos Biliares/citología , Conductos Biliares/efectos de los fármacos , Atresia Biliar/inducido químicamente , Atresia Biliar/genética , Atresia Biliar/patología , Línea Celular , GMP Cíclico/agonistas , GMP Cíclico/metabolismo , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Quimioterapia Combinada , Depuradores de Radicales Libres/uso terapéutico , Glutatión/metabolismo , Humanos , Proteostasis/genética , Transducción de Señal/efectos de los fármacos , Pez CebraRESUMEN
Next-generation sequencing has revolutionized the diagnostic process, making broadscale testing affordable and applicable to almost all specialties; however, there remain several challenges in its widespread implementation. Barriers such as lack of infrastructure or expertise within local health systems and complex result interpretation or counseling make it harder for frontline clinicians to incorporate genomic testing in their existing workflow. The general population is more informed and interested in pursuing genetic testing, and this has been coupled with the increasing accessibility of direct-to-consumer testing. As a result of these changes, primary care physicians and nongenetics specialty providers find themselves seeing patients for whom genetic testing would be beneficial but managing genetic test results that are out of their scope of practice. In this report, we present a practical and centralized approach to providing genomic services through an independent, enterprise-wide clinical service model. We present 4 years of clinical experience, with >3400 referrals, toward designing and implementing the clinical service, maximizing resources, identifying barriers, and improving patient care. We provide a framework that can be implemented at other institutions to support and integrate genomic services across the enterprise.
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Atención a la Salud/organización & administración , Pruebas Genéticas , Pediatría , Niño , Genómica , HumanosRESUMEN
Biliary atresia (BA) is a severe pediatric liver disease resulting in necroinflammatory obliteration of the extrahepatic biliary tree. BA presents within the first few months of life as either an isolated finding or with additional syndromic features. The etiology of isolated BA is unknown, with evidence for infectious, environmental, and genetic risk factors described. However, to date, there are no definitive causal genes identified for isolated BA in humans, and the question of whether single gene defects play a major role remains open. We performed exome-sequencing in 101 North American patients of European descent with isolated BA (including 30 parent-child trios) and considered several experimental designs to identify potentially deleterious protein-altering variants that may be involved in the disease. In a case-only analysis, we did not identify genes with variants shared among more than two probands, and burden tests of rare variants using a case-case control design did not yield significant results. In the trio analysis of 30 simplex families (patient and parent trios), we identified 66 de novo variants in 66 genes including potentially deleterious variants in STIP1 and REV1. STIP1 is a co-chaperone for the heat-shock protein, HSP90, and has been shown to have diverse functions in yeast, flies and mammals, including stress-responses. REV1 is known to be a key player in DNA repair pathway and to interact with HSP90. In conclusion, our results do not support the hypothesis that a simple genetic model is responsible for the majority of cases of isolated BA. Our finding of de novo variants in genes linked to evolutionarily conserved stress responses (STIP1 and REV1) suggests that exploration of how genetic susceptibility and environmental exposure may interact to cause BA is warranted.
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Atresia Biliar/diagnóstico , Exoma , Proteínas HSP90 de Choque Térmico/genética , Proteínas de Choque Térmico/genética , Mutación Missense , Nucleotidiltransferasas/genética , Atresia Biliar/genética , Atresia Biliar/metabolismo , Atresia Biliar/patología , Estudios de Casos y Controles , Niño , Preescolar , Femenino , Expresión Génica , Interacción Gen-Ambiente , Predisposición Genética a la Enfermedad , Proteínas HSP90 de Choque Térmico/metabolismo , Proteínas de Choque Térmico/metabolismo , Humanos , Masculino , Nucleotidiltransferasas/metabolismo , Polimorfismo de Nucleótido Simple , Secuenciación del ExomaRESUMEN
Gastrointestinal motility disorders include a spectrum of mild to severe clinical phenotypes that are caused by smooth muscle dysfunction. We investigated the genetic etiology of severe esophageal, gastric, and colonic dysmotility in two unrelated families with autosomal dominant disease presentation. Using exome sequencing, we identified a 2 base pair insertion at the end of the myosin heavy chain 11 (MYH11) gene in all affected members of Family 1 [NM_001040113:c.5819_5820insCA(p.Gln1941Asnfs*91)] and a 1 base pair deletion at the same genetic locus in Proband 2 [NM_001040113:c.5819del(p.Pro1940Hisfs*91)]. Both variants are predicted to result in a similarly elongated protein product. Heterozygous dominant negative MYH11 pathogenic variants have been associated with thoracic aortic aneurysm and dissection while biallelic null alleles have been associated with megacystis microcolon intestinal hypoperistalsis syndrome. This report highlights heterozygous protein-elongating MYH11 variants affecting the SM2 isoforms of MYH11 as a cause for severe gastrointestinal dysmotility, and we hypothesize that the mechanistic pathogenesis of this disease, dominant hypercontractile loss-of-function, is distinct from those implicated in other diseases involving MYH11 dysfunction.
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Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Músculo Liso/metabolismo , Músculo Liso/fisiopatología , Mutación , Cadenas Pesadas de Miosina/genética , Fenotipo , Adulto , Niño , Análisis Mutacional de ADN , Electromiografía , Endoscopía del Sistema Digestivo , Trastornos de la Motilidad Esofágica/diagnóstico , Trastornos de la Motilidad Esofágica/genética , Femenino , Gastroparesia/diagnóstico , Gastroparesia/genética , Estudios de Asociación Genética/métodos , Estudio de Asociación del Genoma Completo , Humanos , Lactante , Enfermedades Intestinales/diagnóstico , Enfermedades Intestinales/genética , Masculino , Persona de Mediana Edad , Linaje , Polimorfismo de Nucleótido Simple , Radiografía , Síndrome , Adulto JovenRESUMEN
Alagille syndrome is an autosomal dominant disease with a known molecular etiology of dysfunctional Notch signaling caused primarily by pathogenic variants in JAGGED1 (JAG1), but also by variants in NOTCH2. The majority of JAG1 variants result in loss of function, however disease has also been attributed to lesser understood missense variants. Conversely, the majority of NOTCH2 variants are missense, though fewer of these variants have been described. In addition, there is a small group of patients with a clear clinical phenotype in the absence of a pathogenic variant. Here, we catalog our single-center study, which includes 401 probands and 111 affected family members amassed over a 27-year period, to provide updated mutation frequencies in JAG1 and NOTCH2 as well as functional validation of nine missense variants. Combining our cohort of 86 novel JAG1 and three novel NOTCH2 variants with previously published data (totaling 713 variants), we present the most comprehensive pathogenic variant overview for Alagille syndrome. Using this data set, we developed new guidance to help with the classification of JAG1 missense variants. Finally, we report clinically consistent cases for which a molecular etiology has not been identified and discuss the potential for next generation sequencing methodologies in novel variant discovery.
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Síndrome de Alagille/genética , Proteína Jagged-1/genética , Mutación con Pérdida de Función , Mutación Missense , Receptor Notch2/genética , Síndrome de Alagille/metabolismo , Femenino , Predisposición Genética a la Enfermedad , Humanos , Proteína Jagged-1/metabolismo , Masculino , Tasa de Mutación , Linaje , Receptor Notch2/metabolismoRESUMEN
Importance: Although genetic testing is important for bringing precision medicine to children with epilepsy, it is unclear what genetic testing strategy is best in maximizing diagnostic yield. Objectives: To evaluate the diagnostic yield of an exome-based gene panel for childhood epilepsy and discuss the value of follow-up testing. Design, Setting, and Participants: A case series study was conducted on data from clinical genetic testing at Children's Hospital of Philadelphia was conducted from September 26, 2016, to January 8, 2018. Initial testing targeted 100 curated epilepsy genes for sequence and copy number analysis in 151 children with idiopathic epilepsy referred consecutively by neurologists. Additional genetic testing options were offered afterward. Exposures: Clinical genetic testing. Main Outcomes and Measures: Molecular diagnostic findings. Results: Of 151 patients (84 boys [55.6%]; median age, 4.2 years [interquartile range, 1.4-8.7 years]), 16 children (10.6%; 95% CI, 6%-16%) received a diagnosis after initial panel analysis. Parental testing for 15 probands with inconclusive results revealed de novo variants in 7 individuals (46.7%), resulting in an overall diagnostic yield of 15.3% (23 of 151; 95% CI, 9%-21%). Twelve probands with nondiagnostic panel findings were reflexed to exome sequencing, and 4 were diagnostic (33.3%; 95% CI, 6%-61%), raising the overall diagnostic yield to 17.9% (27 of 151; 95% CI, 12%-24%). The yield was highest (17 of 44 [38.6%; 95% CI, 24%-53%]) among probands with epilepsy onset in infancy (age, 1-12 months). Panel diagnostic findings involved 16 genes: SCN1A (n = 4), PRRT2 (n = 3), STXBP1 (n = 2), IQSEC2 (n = 2), ATP1A2, ATP1A3, CACNA1A, GABRA1, KCNQ2, KCNT1, SCN2A, SCN8A, DEPDC5, TPP1, PCDH19, and UBE3A (all n = 1). Exome sequencing analysis identified 4 genes: SMC1A, SETBP1, NR2F1, and TRIT1. For the remaining 124 patients, analysis of 13 additional genes implicated in epilepsy since the panel was launched in 2016 revealed promising findings in 6 patients. Conclusions and Relevance: Exome-based targeted panels appear to enable rapid analysis of a preselected set of genes while retaining flexibility in gene content. Successive genetic workup should include parental testing of select probands with inconclusive results and reflex to whole-exome trio analysis for the remaining nondiagnostic cases. Periodic reanalysis is needed to capture information in newly identified disease genes.
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Epilepsia/diagnóstico , Epilepsia/genética , Secuenciación del Exoma/métodos , Predisposición Genética a la Enfermedad/genética , Pruebas Genéticas/métodos , Niño , Preescolar , Femenino , Humanos , Lactante , Masculino , Tripeptidil Peptidasa 1Asunto(s)
Ácidos Nucleicos Libres de Células , Diagnóstico Prenatal , ADN , Femenino , Feto , Humanos , Embarazo , Atención PrenatalRESUMEN
The originally published version of this Article contained errors in Fig. 2. The numbers below the black arrowheads were incorrect; please see incorrect Figure in associated Correction. These errors have now been corrected in the PDF and HTML versions of the Article.
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Clinical exome sequencing (CES) has become the preferred diagnostic platform for complex pediatric disorders with suspected monogenic etiologies. Despite rapid advancements, the major challenge still resides in identifying the casual variants among the thousands of variants detected during CES testing, and thus establishing a molecular diagnosis. To improve the clinical exome diagnostic efficiency, we developed Phenoxome, a robust phenotype-driven model that adopts a network-based approach to facilitate automated variant prioritization. Phenoxome dissects the phenotypic manifestation of a patient in concert with their genomic profile to filter and then prioritize variants that are likely to affect the function of the gene (potentially pathogenic variants). To validate our method, we have compiled a clinical cohort of 105 positive patient samples that represent a wide range of genetic heterogeneity. Phenoxome identifies the causative variants within the top 5, 10, or 25 candidates in more than 50%, 71%, or 88% of these exomes, respectively. Furthermore, we show that our method is optimized for clinical testing by outperforming the current state-of-art method. We have demonstrated the performance of Phenoxome using a clinical cohort and showed that it enables rapid and accurate interpretation of clinical exomes. Phenoxome is available at https://phenoxome.chop.edu/ .