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OBJECTIVE: To investigate the clinical value of whole-exome sequencing (WES) in the diagnosis of foetuses with central nervous system (CNS) abnormalities but having a normal karyotyping and chromosomal microarray result. METHOD: During the period of 2016-2022, there were a total of 149 foetuses with CNS abnormalities but having negative karyotyping and chromosomal microarray analysis results; WES was performed on these foetuses and their parents. Variants were classified according to ACMG guidelines, and the association of pathogenic variants with specific types of CNS abnormalities was explored. RESULTS: Among these 149 foetuses, three categories of abnormalities, namely, single CNS abnormality, multiple CNS abnormalities, CNS abnormalities along with other organ system abnormalities were identified, for which the detection rate of P/LP variants is 17.4% (12/69), 28.6% (14/49) and 54.8% (17/31), respectively. CONCLUSION: WES brought about an increase of 28.9% in diagnostic yield in the prenatal evaluation of foetuses with CNS abnormalities but having negative karyotyping and chromosome array results. WES may also be of benefit for the diagnosis of foetuses with isolated CNS abnormalities, as well as for making more informed interpretations of imaging findings and for providing better genetic counselling.
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Sequenciamento do Exoma , Diagnóstico Pré-Natal , Humanos , Feminino , Sequenciamento do Exoma/métodos , Gravidez , Diagnóstico Pré-Natal/métodos , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/diagnóstico , Adulto , Testes Genéticos/métodos , Feto/anormalidades , Sistema Nervoso Central/anormalidades , Cariotipagem/métodosRESUMO
PURPOSE: To evaluate the relative diagnostic yield of clinical germline genomic tests in a diverse pediatric cancer population. PATIENTS AND METHODS: The KidsCanSeq study enrolled pediatric cancer patients across six sites in Texas. Germline analysis included both exome sequencing and a therapy-focused pediatric cancer gene panel. The results were categorized by participants demographics, the presence of pathogenic or likely pathogenic (P/LP) variants, and variants of uncertain significance (VUS) in cancer predisposition genes (CPGs). Pediatric actionable CPGs were defined as those with cancer surveillance recommendations during childhood. RESULTS: Cancer P/LP variants were reported by at least one platform in 103 of 578 (17.8%) participants of which 76 were dominant cancer genes (13.1%) with no significant differences by self-described race or Hispanic ethnicity. However, the proportion of participants with VUS was greater in Asian and African American participants (P = .0029). Diagnostic yield was 16.6% for exome versus 8.5% for panel (P < .0001) with 42 participants with concordant germline results. Exome-only results included P/LP variants in 30 different CPGs in 54 participants, whereas panel-only results included seven participants with a copy number or structural P/LP variants in CPGs. There was no significant difference in diagnostic yield limited to pediatric actionable CPGs (P = .6171). CONCLUSION: Approximately 18% of a diverse pediatric cancer population had germline diagnostic findings with 50% of P/LP variants reported by only one platform because of CPGs not on the targeted panel and copy number variants (CNVs)/rearrangements not reported by exome. Although diagnostic yields were similar in this diverse population, increases in VUS results were observed in Asian and African American populations. Given the clinical significance of CNVs/rearrangements in this cohort, detection is critical to optimize germline analysis of pediatric cancer populations.
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Sequenciamento do Exoma , Mutação em Linhagem Germinativa , Neoplasias , Humanos , Criança , Neoplasias/genética , Neoplasias/diagnóstico , Texas , Masculino , Feminino , Pré-Escolar , Adolescente , Sequenciamento do Exoma/métodos , Exoma/genética , Lactente , Predisposição Genética para Doença , Células GerminativasRESUMO
BACKGROUND: Whole exome sequencing (WES) has been recommended to investigate the genetic cause of fetal structural anomalies. In this retrospective study, we aimed to evaluate the diagnostic yield of WES in our cohort of families with pregnancy loss or termination of pregnancy due to structural anomalies. METHODS: As aneuploidy, triploidy and copy number variations (CNVs) could be detected by exome-based CNV analysis, only WES is performed in this study. And the results of 375 cases assessed by WES were analyzed. RESULTS: The overall detection rate was 32.3% (121/375), including aneuploidy and triploidy (7.5%, 28/375), CNVs (5.1%, 19/375) and single-nucleotide variants (SNVs) /insertions or deletions (Indels) (19.7%, 74/375). Among these, the diagnostic yield for likely pathogenic (LP) or pathogenic (P) CNVs is 4.8% (18/375), and the diagnostic yield for LP or P SNVs/Indels is 15.2% (57/375). And an additional 4.8% (18/375) of cases had CNVs or SNVs/Indels classified as variants of uncertain significance (VUS) with potential clinical significance. CONCLUSIONS: Our findings expand the known mutation spectrum of genetic variants related to fetal abnormalities, increase our understanding of prenatal phenotypes, and enable more accurate counseling of recurrence risk for future pregnancies.
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Variações do Número de Cópias de DNA , Sequenciamento do Exoma , Humanos , Feminino , Sequenciamento do Exoma/métodos , Gravidez , Variações do Número de Cópias de DNA/genética , Estudos Retrospectivos , Adulto , Feto , Testes Genéticos/métodos , Aborto Espontâneo/genética , AneuploidiaRESUMO
INTRODUCTION: The molecular basis and mechanisms of juvenile nasopharyngeal angiofibromas (JNA) pathogenesis are still unknown. Despite being a rare and benign neoplasm, JNA is a locally aggressive and potentially destructive head and neck neoplasm, typically found in young males. The advancement of genome technologies and analytical tools has provided an unparalleled opportunity to explore the intricacy of JNA. The present study provides the first evidence of the involvement of Y-chromosome genes in JNA. METHODS: A total of 13 JNA patients at an advanced disease stage and five age-matched male controls were registered for this study. Whole-exome sequencing (WES) analysis was conducted followed by functional analysis to understand the molecular mechanism of the JNA. RESULTS: WES analysis revealed a high prevalence of mutations in 14 genes within the protein-coding, male-specific region of the Y-chromosome of young males (mean age: 13.8 ± 2.4) with JNA. These mutations, occurring at 28 distinct positions, were characterized as moderate to high impact and were prevalent in nine JNA patients but not in the control group. The most frequently mutated genes were USP9Y and UTY, followed by KDM5D, DDX3Y, and TSPY4. The expression of USP9Y, UTY, and DDX3Y was found to be co-modulated, implying their coordinated regulation as a complex. Furthermore, somatic mutations were detected in genes previously linked to JNA. CONCLUSION: The wide array of genetic mutations in the Y-chromosome male-specific region, along with the somatic alterations identified in JNA, provides novel insights into JNA pathophysiology.
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Angiofibroma , Sequenciamento do Exoma , Mutação , Neoplasias Nasofaríngeas , Humanos , Angiofibroma/genética , Angiofibroma/patologia , Masculino , Sequenciamento do Exoma/métodos , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/patologia , Adolescente , Criança , Ubiquitina Tiolesterase/genéticaRESUMO
BACKGROUND: Hereditary Spastic Paraplegias (HSPs) and Hereditary Cerebellar Ataxias (HCAs) are progressive neurodegenerative disorders encompassing a spectrum of neurogenetic conditions with significant overlaps of clinical features. Spastic ataxias are a group of conditions that have features of both cerebellar ataxia and spasticity, and these conditions are frequently clinically challenging to distinguish. Accurate genetic diagnosis is crucial but challenging, particularly in resource-limited settings. This study aims to investigate the genetic basis of HSPs and HCAs in Pakistani families. METHODS: Families from Khyber Pakhtunkhwa with at least two members showing HSP or HCA phenotypes, and who had not previously been analyzed genetically, were included. Families were referred for genetic analysis by local neurologists based on the proband's clinical features and signs of a potential genetic neurodegenerative disorder. Whole Exome Sequencing (WES) and Sanger sequencing were then used to identify and validate genetic variants, and to analyze variant segregation within families to determine inheritance patterns. The mean age of onset and standard deviation were calculated to assess variability among affected individuals, and the success rate was compared with literature reports using differences in proportions and Cohen's h. RESULTS: Pathogenic variants associated with these conditions were identified in five of eight families, segregating according to autosomal recessive inheritance. These variants included previously reported SACS c.2182 C > T, p.(Arg728*), FA2H c.159_176del, p.(Arg53_Ile58del) and SPG11 c.2146 C > T, p.(Gln716*) variants, and two previously unreported variants in SACS c.2229del, p.(Phe743Leufs*8) and ZFYVE26 c.1926_1941del, p.(Tyr643Metfs*2). Additionally, FA2H and SPG11 variants were found to have recurrent occurrences, suggesting a potential founder effect within the Pakistani population. Onset age among affected individuals ranged from 1 to 14 years (M = 6.23, SD = 3.96). The diagnostic success rate was 62.5%, with moderate effect sizes compared to previous studies. CONCLUSIONS: The findings of this study expand the genotypic and phenotypic spectrum of HSPs and HCAs in Pakistan and emphasize the importance of utilizing exome/genome sequencing for accurate diagnosis or support accurate differential diagnosis. This approach can improve genetic counseling and clinical management, addressing the challenges of diagnosing neurodegenerative disorders in resource-limited settings.
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Ataxia Cerebelar , Linhagem , Paraplegia Espástica Hereditária , Humanos , Paraplegia Espástica Hereditária/genética , Paraplegia Espástica Hereditária/diagnóstico , Paquistão , Masculino , Feminino , Adulto , Criança , Adolescente , Ataxia Cerebelar/genética , Ataxia Cerebelar/diagnóstico , Adulto Jovem , Pessoa de Meia-Idade , Pré-Escolar , Sequenciamento do Exoma/métodos , Mutação , FenótipoRESUMO
The genetic basis of schizophrenia (SZ) remains elusive despite its characterization as a highly heritable disorder. This incomplete understanding has led to stagnation in therapeutics and treatment, leaving many suffering with insufficient relief from symptoms. However, recent large-cohort genome- and exome-wide association studies have provided insights into the underlying genetic machinery. The scale of these studies allows for the identification of ultra-rare mutations that confer substantial disease risk, guiding clinicians and researchers toward general classes of genes that are central to SZ etiology. One such large-scale collaboration effort by the Schizophrenia Exome Sequencing Meta-Analysis consortium identified ten, high-risk, ultra-rare, protein-truncating variants, providing the clearest picture to date of the dysfunctional gene products that substantially increase risk for SZ. While genetic studies of SZ provide valuable information regarding "what" genes are linked with the disorder, it is an open question as to "when" during brain development these genetic mutations impose deleterious effects. To shed light on this unresolved aspect of SZ etiology, we queried the BrainSpan developmental mRNA expression database for these ten high-risk genes and discovered three general expression trajectories throughout pre- and postnatal brain development. The elusiveness of SZ etiology, we infer, is not only borne out of the genetic heterogeneity across clinical cases, but also in our incomplete understanding of how genetic mutations perturb neurodevelopment during multiple critical periods. We contextualize this notion within the National Institute of Mental Health's Research Domain Criteria framework and emphasize the utility of considering both genetic variables and developmental context in future studies.
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Predisposição Genética para Doença , Mutação , Esquizofrenia , Esquizofrenia/genética , Humanos , Variação Genética , Estudo de Associação Genômica Ampla , Encéfalo/patologia , Encéfalo/metabolismo , Sequenciamento do Exoma/métodosRESUMO
INTRODUCTION: Whole Exome Sequencing (WES) has emerged as an efficient tool in clinical cancer diagnostics to broaden the scope from panel-based diagnostics to screening of all genes and enabling robust determination of complex biomarkers in a single analysis. METHODS: To assess concordance, six formalin-fixed paraffin-embedded (FFPE) tissue specimens and four commercial reference standards were analyzed by WES as matched tumor-normal DNA at 21 NGS centers in Germany, each employing local wet-lab and bioinformatics. Somatic and germline variants, copy-number alterations (CNAs), and complex biomarkers were investigated. Somatic variant calling was performed in 494 diagnostically relevant cancer genes. The raw data were collected and re-analyzed with a central bioinformatic pipeline to separate wet- and dry-lab variability. RESULTS: The mean positive percentage agreement (PPA) of somatic variant calling was 76 % while the positive predictive value (PPV) was 89 % in relation to a consensus list of variants found by at least five centers. Variant filtering was identified as the main cause for divergent variant calls. Adjusting filter criteria and re-analysis increased the PPA to 88 % for all and 97 % for the clinically relevant variants. CNA calls were concordant for 82 % of genomic regions. Homologous recombination deficiency (HRD), tumor mutational burden (TMB), and microsatellite instability (MSI) status were concordant for 94 %, 93 %, and 93 % of calls, respectively. Variability of CNAs and complex biomarkers did not decrease considerably after harmonization of the bioinformatic processing and was hence attributed mainly to wet-lab differences. CONCLUSION: Continuous optimization of bioinformatic workflows and participating in round robin tests are recommended.
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Benchmarking , Variações do Número de Cópias de DNA , Sequenciamento do Exoma , Neoplasias , Medicina de Precisão , Humanos , Sequenciamento do Exoma/métodos , Alemanha , Medicina de Precisão/métodos , Medicina de Precisão/normas , Neoplasias/genética , Biomarcadores Tumorais/genética , Biologia Computacional/métodosRESUMO
Background: Wiedemann-Steiner syndrome (WSS), a rare autosomal-dominant disorder caused by haploinsufficiency of the KMT2A gene product, is part of a group of disorders called chromatinopathies. Chromatinopathies are neurodevelopmental disorders caused by mutations affecting the proteins responsible for chromatin remodeling and transcriptional regulation. The resulting gene expression dysregulation mediates the onset of a series of clinical features such as developmental delay, intellectual disability, facial dysmorphism, and behavioral disorders. Aim of the Study: The aim of this study was to investigate a 10-year-old girl who presented with clinical features suggestive of WSS. Methods: Clinical and genetic investigations were performed. Whole exome sequencing (WES) was used for genetic testing, performed using Illumina technology. The bidirectional capillary Sanger resequencing technique was used in accordance with standard methodology to validate a mutation discovered by WES in all family members who were available. Utilizing computational protein modeling for structural and functional studies as well as in silico pathogenicity prediction models, the effect of the mutation was examined. Results: WES identified a de novo heterozygous missense variant in the KMT2A gene KMT2A(NM_001197104.2): c.3451C>G, p.(Arg1151Gly), absent in the gnomAD database. The variant was classified as Likely Pathogenetic (LP) according to the ACMG criteria and was predicted to affect the CXXC-type zinc finger domain functionality of the protein. Modeling of the resulting protein structure suggested that this variant changes the protein flexibility due to a variation in the Gibbs free energy and in the vibrational entropy energy difference between the wild-type and mutated domain, resulting in an alteration of the DNA binding affinity. Conclusions: A novel and de novo mutation discovered by the NGS approach, enhancing the mutation spectrum in the KMT2A gene, was characterized and associated with WSS. This novel KMT2A gene variant is suggested to modify the CXXC-type zinc finger domain functionality by affecting protein flexibility and DNA binding.
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Sequenciamento do Exoma , Histona-Lisina N-Metiltransferase , Proteína de Leucina Linfoide-Mieloide , Humanos , Feminino , Sequenciamento do Exoma/métodos , Proteína de Leucina Linfoide-Mieloide/genética , Criança , Histona-Lisina N-Metiltransferase/genética , Deficiência Intelectual/genética , Deficiência Intelectual/patologia , Deficiência Intelectual/diagnóstico , Mutação de Sentido Incorreto , Anormalidades Múltiplas/genéticaRESUMO
Advances in genetics led to the identification of hundreds of epilepsy-related genes, some of which are treatable with etiology-specific interventions. However, the diagnostic yield of next-generation sequencing (NGS) in unexplained epilepsy is highly variable (10-50%). We sought to determine the diagnostic yield and clinical utility of NGS in children with unexplained epilepsy that is accompanied by neurodevelopmental delays and/or is medically intractable. A 5-year retrospective review was conducted at the American University of Beirut Medical Center to identify children who underwent whole exome sequencing (WES) or whole genome sequencing (WGS). Data on patient demographics, neurodevelopment, seizures, and treatments were collected. Forty-nine children underwent NGS with an overall diagnostic rate of 68.9% (27/38 for WES, and 4/7 for WGS). Most children (42) had neurodevelopmental delays with (18) or without (24) refractory epilepsy, and only three had refractory epilepsy without delays. The diagnostic yield was 77.8% in consanguineous families (18), and 61.5% in non-consanguineous families (26); consanguinity information was not available for one family. Genetic test results led to anti-seizure medication optimization or dietary therapies in six children, with subsequent improvements in seizure control and neurodevelopmental trajectories. Not only is the diagnostic rate of NGS high in children with unexplained epilepsy and neurodevelopmental delays, but also genetic testing in this population may often lead to potentially life-altering interventions.
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Deficiências do Desenvolvimento , Epilepsia , Sequenciamento do Exoma , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Feminino , Criança , Epilepsia/genética , Epilepsia/diagnóstico , Pré-Escolar , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Estudos Retrospectivos , Deficiências do Desenvolvimento/genética , Deficiências do Desenvolvimento/diagnóstico , Lactente , Sequenciamento do Exoma/métodos , Transtornos do Neurodesenvolvimento/genética , Transtornos do Neurodesenvolvimento/diagnóstico , Testes Genéticos/métodos , Adolescente , Sequenciamento Completo do Genoma/métodosRESUMO
Single-cell genomic analysis enables researchers to gain novel insights across diverse research areas, including developmental biology, tumor heterogeneity, and disease pathogenesis. Conducting single-cell genomic analysis using next-generation sequencing (NGS) methods has traditionally been challenging as the amount of genomic DNA present in a single cell is limited. Advancements in multiple displacement amplification (MDA) technologies allow the unbiased amplification of limited quantities of DNA under conditions that maintain its integrity. This method of amplification results in high yield and facilitates the generation of high-complexity NGS libraries that ensure the highest coverage to effectively allow variant calling. With the introduction of new sequencing platforms and chemistry, whole genome sequencing became a more cost-effective application, but enrichment of specific regions of interest further reduces the amount of required sequencing output and associated costs. There are two enrichment methods, polymerase chain reaction (PCR)-based and hybrid-capture-based methods. PCR-based methods are very flexible and highly effective but focus on specific loci, typically known to be associated with disease. Inherited diseases of unknown genetic origin require a more comprehensive approach to capture the genetic variation that is not yet associated with a specific disease. Hybrid capture enrichment methods require considerable amounts of DNA such that exome enrichment from single cells is only possible after preamplification of this limited material. This article describes the complete workflow from single cells and small quantities of DNA to exome-NGS libraries for Illumina sequencing instruments and includes the following protocols: © 2024 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Whole genome amplification from single cells or small amounts of gDNA Basic Protocol 2: NGS library generation of MDA-amplified material Basic Protocol 3: Exome enrichment.
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Análise de Célula Única , Análise de Célula Única/métodos , Humanos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequenciamento do Exoma/métodos , Reação em Cadeia da Polimerase/métodos , Exoma/genéticaRESUMO
Rationale: Extrachromosomal circular DNA is a hallmark of cancer, but its role in shaping the genome heterogeneity of urothelial bladder carcinoma (UBC) remains poorly understood. Here, we comprehensively analyzed the features of extrachromosomal circular DNA in 80 UBC patients. Methods: We performed whole-genome/exome sequencing (WGS/WES), Circle-Seq, single-molecule real-time (SMRT) long-read sequencing of circular DNA, and RNA sequencing (RNA-Seq) on 80 pairs of tumor and AT samples. We used our newly developed circular DNA analysis software, Circle-Map++ to detect small extrachromosomal circular DNA from Circle-Seq data. Results: We observed a high load and significant heterogeneity of extrachromosomal circular DNAs in UBC, including numerous single-locus and complex chimeric circular DNAs originating from different chromosomes. This includes highly chimeric circular DNAs carrying seven oncogenes and circles from nine chromosomes. We also found that large tumor-specific extrachromosomal circular DNAs could influence genome-wide gene expression, and are detectable in time-matched urinary sediments. Additionally, we found that the extrachromosomal circular DNA correlates with hypermutation, copy number variation, oncogene amplification, and clinical outcome. Conclusions: Overall, our study provides a comprehensive extrachromosomal circular DNA map of UBC, along with valuable data resources and bioinformatics tools for future cancer and extrachromosomal circular DNA research.
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Variações do Número de Cópias de DNA , DNA Circular , Neoplasias da Bexiga Urinária , Neoplasias da Bexiga Urinária/genética , Humanos , DNA Circular/genética , Variações do Número de Cópias de DNA/genética , Sequenciamento Completo do Genoma/métodos , Heterogeneidade Genética , Masculino , Feminino , Sequenciamento do Exoma/métodos , Idoso , Mutação/genéticaRESUMO
BACKGROUND: Diagnosis of hereditary myopathy is often challenging owing to overlapping clinical phenotypes and muscle histopathological findings. This retrospective study aimed to identify the phenotypic and genotypic spectra of hereditary myopathies at a tertiary hospital in Riyadh, Saudi Arabia. METHODS: We reviewed the medical records of patients with hereditary myopathy who were evaluated between January 2018 and December 2022. RESULTS: Eighty-seven patients (78 families) were included, two-thirds were men with a mean age of 35 (SD 14.2) years. Limb-girdle muscular dystrophy (LGMD) was the most prevalent clinical diagnosis (25 cases; 29%), of whom, a genetic diagnosis was achieved in 15 of 22 patients tested (68%). In genetically confirmed LGMD, the most prevalent disorders were dysferlinopathy (27%) followed by fukutin-related protein (FKRP) - related limb girdle muscular dystrophy (20%), sarcoglycanopathy (20%), lamin A/C related myopathy (13%), and calpain-3 myopathy (13%). In 26 patients with pathogenic/likely pathogenic variants, the genetic testing method was whole exome sequencing (WES) (42%), Next generation sequencing (NGS) (31%), and targeted single gene analysis (27%). The sensitivity of each genetic testing method was as follows: 100% for targeted single-gene analysis, 100% for targeted analysis of D4Z4 repeat array units, 88% for myotonic dystrophy protein kinase (DMPK) repeat expansion analysis, 42% for NGS-neuromuscular panel, and 46% for WES. CONCLUSION: The prevalent types of hereditary myopathies were consistent with those reported locally and internationally. This study highlights the diagnostic yield of various molecular genetic tests for the diagnosis of hereditary myopathy in an adult cohort and the need for improved access to advanced molecular testing in cases suspected to have facioscapulohumeral muscular dystrophy (FSHD) or mitochondrial myopathies.
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Testes Genéticos , Distrofia Muscular do Cíngulo dos Membros , Humanos , Masculino , Arábia Saudita/epidemiologia , Adulto , Feminino , Pessoa de Meia-Idade , Distrofia Muscular do Cíngulo dos Membros/genética , Distrofia Muscular do Cíngulo dos Membros/diagnóstico , Distrofia Muscular do Cíngulo dos Membros/epidemiologia , Estudos Retrospectivos , Testes Genéticos/métodos , Adulto Jovem , Estudos de Coortes , Adolescente , Doenças Musculares/genética , Doenças Musculares/diagnóstico , Sequenciamento do Exoma/métodosRESUMO
BACKGROUND: Ectodermal dysplasia (ED) is a rare genetic disorder that affects structures derived from the ectodermal germ layer. RESULTS: In this study, we analyzed the genetic profiles of 27 Korean patients with ED. Whole exome sequencing (WES) was performed on 23 patients, and targeted panel sequencing was conducted on the remaining 4 patients. Among the patients in the cohort, 74.1% (20/27) tested positive for ED. Of these positive cases, EDA and EDAR mutations were found in 80% (16/20). Notably, 23.1% (3/13) of EDA-positive cases exhibited copy number variations. Among the 23 patients who underwent WES, we conducted a virtual panel analysis of eight well-known genes, resulting in diagnoses for 56.5% (13/23) of the cases. Additionally, further analysis of approximately 5,000 OMIM genes identified four more cases, increasing the overall positivity rate by approximately 17%. These findings underscore the potential of WES for improving the diagnostic yield of ED. Remarkably, 94.1% of the patients manifesting the complete triad of ED symptoms (hair/skin/dental) displayed detectable EDA/EDAR mutations. In contrast, none of the 7 patients without these three symptoms exhibited EDA/EDAR mutations. CONCLUSIONS: When conducting molecular diagnostics for ED, opting for targeted sequencing of EDA/EDAR mutations is advisable for cases with classical symptoms, while WES is deemed an effective strategy for cases in which these symptoms are absent.
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Displasia Ectodérmica , Sequenciamento do Exoma , Mutação , Humanos , Displasia Ectodérmica/genética , Displasia Ectodérmica/diagnóstico , República da Coreia , Masculino , Feminino , Sequenciamento do Exoma/métodos , Mutação/genética , Criança , Variações do Número de Cópias de DNA/genética , Perfil Genético , Pré-Escolar , Adulto , Adolescente , Receptor Edar/genética , Ectodisplasinas/genética , Lactente , Adulto JovemRESUMO
BACKGROUND: Hereditary spastic paraplegia (HSP) represents a group of monogenic neurodegenerative disorders characterized by high clinical and genetic heterogeneity. HSP is characterized by slowly progressing hypertonia of both lower extremities, spastic gait, and myasthenia. The most prevalent autosomal dominant form of HSP, known as spastic paraplegia 4 (SPG4), is attributed to variants in the spastin (SPAST) gene. METHODS AND RESULTS: Here, a Chinese family presenting with spasticity in both legs and a shuffling gait participated in our investigation. Whole exome sequencing of the proband was utilized to identify the genetic lesion in the family. Through data filtering, Sanger sequencing validation, and co-separation analysis, a novel variant (NM_014946.3: c.1669G > C:p.A557P) of SPAST was identified as the genetic lesion of this family. Furthermore, bioinformatic analysis revealed that this variant was deleterious and located in a highly evolutionarily conserved site. CONCLUSION: Our study confirmed the diagnosis of SPG4 in this family, contributing to genetic counseling for families affected by SPG4. Additionally, our study broadened the spectrum of SPAST variants and highlighted the importance of ATPases associated with various cellular activity domains of SPAST.
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Paraplegia Espástica Hereditária , Espastina , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , China , População do Leste Asiático/genética , Sequenciamento do Exoma/métodos , Mutação/genética , Paraplegia , Linhagem , Paraplegia Espástica Hereditária/genética , Espastina/genéticaRESUMO
Moebius syndrome (MBS) is a rare congenital disorder characterized by non-progressive facial palsy and ocular abduction paralysis. Most cases are sporadic, but also rare familial cases with autosomal dominant transmission and incomplete penetrance/variable expressivity have been described. The genetic etiology of MBS is still unclear: de novo pathogenic variants in REV3L and PLXND1 are reported in only a minority of cases, suggesting the involvement of additional causative genes. With the aim to uncover the molecular causative defect and identify a potential genetic basis of this condition, we performed trio-WES on a cohort of 37 MBS and MBS-like patients. No de novo variants emerged in REV3L and PLXND1. We then proceeded with a cohort analysis to identify possible common causative genes among all patients and a trio-based analysis using an in silico panel of candidate genes. However, identified variants emerging from both approaches were considered unlikely to be causative of MBS, mainly due to the lack of clinical overlap. In conclusion, despite this large cohort, WES failed to identify mutations possibly associated with MBS, further supporting the heterogeneity of this syndrome, and suggesting the need for integrated omics approaches to identify the molecular causes underlying MBS development.
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Sequenciamento do Exoma , Síndrome de Möbius , Humanos , Sequenciamento do Exoma/métodos , Masculino , Feminino , Síndrome de Möbius/genética , Mutação , Criança , Pré-Escolar , Estudos de Coortes , Lactente , Adolescente , Predisposição Genética para DoençaRESUMO
We present the results of the first study of a large cohort of patients with inherited retinal dystrophies (IRD) performed for the Polish population using whole-exome sequencing (WES) in the years 2016-2019. Moreover, to facilitate such diagnostic analyses and enable future application of gene therapy and genome editing for IRD patients, a Polish genomic reference database (POLGENOM) was created based on whole-genome sequences of healthy Polish Caucasian nonagenarians and centenarians. The newly constructed database served as a control, providing a comparison for variant frequencies in the Polish population. The diagnostic yield for the selected group of IRD patients reached 64.9%. The study uncovered the most common pathogenic variants in ABCA4 and USH2A in the European population, along with several novel causative variants. A significant frequency of the ABCA4 complex haplotype p.(Leu541Pro; Ala1038Val) was observed, as well as that of the p.Gly1961Glu variant. The first VCAN causative variant NM_004385.5:c.4004-2A>G in Poland was found and described. Moreover, one of the first patients with the RPE65 causative variants was identified, and, in consequence, could receive the dedicated gene therapy. The availability of the reference POLGENOM database enabled comprehensive variant characterisation during the NGS data analysis, confirming the utility of a population-specific genomic database for enhancing diagnostic accuracy. Study findings suggest the significance of genetic testing in elder patients with unclear aetiology of eye diseases. The combined approach of NGS and the reference genomic database can improve the diagnosis, management, and future treatment of IRDs.
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Sequenciamento do Exoma , Distrofias Retinianas , População Branca , Humanos , Distrofias Retinianas/genética , Polônia , Masculino , População Branca/genética , Feminino , Sequenciamento do Exoma/métodos , Proteínas da Matriz Extracelular/genética , Bases de Dados Genéticas , Transportadores de Cassetes de Ligação de ATP/genética , Idoso de 80 Anos ou mais , cis-trans-Isomerases/genética , Idoso , Pessoa de Meia-Idade , Adulto , MutaçãoRESUMO
Male infertility affects approximately 7% of the male population, and about 15% of these cases are predicted to have a genetic etiology. One gene implicated in autosomal dominant male infertility, SYCP2, encodes a protein critical for the synapsis of homologous chromosomes during meiosis I, resulting in impaired spermatogenesis. However, the clinical validity of the gene-disease pair was previously categorized as on the border of limited and moderate due to few reported cases. This study investigates the genetic cause of infertility for three unrelated Chinese patients with oligoasthenozoospermia. Whole exome sequencing (WES) and subsequent Sanger sequencing revealed novel heterozygous loss-of-function (LOF) variants in SYCP2 (c.89dup, c.946_947del, and c.4378_4379del). These cases, combined with the previously reported cases, provide strong genetic evidence supporting an autosomal dominant inheritance pattern. The experimental evidence also demonstrates a critical role for SYCP2 in spermatogenesis. Collectively, this updated assessment of the genetic and experimental evidence upgrades the gene-disease association strength of SYCP2 and autosomal dominant male infertility from on the border of limited and moderate to strong. The reclassification improves SYCP2 variant interpretation and qualifies it for the inclusion on diagnostic male infertility gene panels and prioritization in whole exome or genome studies for related phenotypes. These findings therefore improve the clinical interpretation of SYCP2 LOF variants.
Assuntos
Sequenciamento do Exoma , Infertilidade Masculina , Masculino , Humanos , Infertilidade Masculina/genética , Adulto , Sequenciamento do Exoma/métodos , Espermatogênese/genética , Mutação com Perda de Função , Proteínas de Ligação a DNA/genética , Proteínas de Ciclo Celular/genética , LinhagemRESUMO
We built a reference panel with 342 million autosomal variants using 78,195 individuals from the Genomics England (GEL) dataset, achieving a phasing switch error rate of 0.18% for European samples and imputation quality of r2 = 0.75 for variants with minor allele frequencies as low as 2 × 10-4 in white British samples. The GEL-imputed UK Biobank genome-wide association analysis identified 70% of associations found by direct exome sequencing (P < 2.18 × 10-11), while extending testing of rare variants to the entire genome. Coding variants dominated the rare-variant genome-wide association results, implying less disruptive effects of rare non-coding variants.
Assuntos
Frequência do Gene , Estudo de Associação Genômica Ampla , Haplótipos , Polimorfismo de Nucleotídeo Único , Humanos , Inglaterra , Sequenciamento do Exoma/métodos , Genoma Humano , Estudo de Associação Genômica Ampla/métodos , Genômica/métodos , Biobanco do Reino Unido , Reino Unido , População Branca/genéticaRESUMO
BACKGROUND: Global developmental delay with speech and behavioral abnormalities (OMIM: 619243) is an autosomal dominant disease caused by variants in TNRC6B gene. METHOD: We reviewed and summarized clinical manifestations and genotypes in patients previously reported with TNRC6B gene variants. We used several prediction tools to predict pathogenicity and performed minigene assays to verify the function of the synonymous variant affecting RNA splicing. RESULT: The patient presented with convulsive seizures and developmental delay. WES combined with functional studies diagnosed a child with a synonymous variant in TNRC6B gene. Through minigene assay and Sanger sequencing, we demonstrated that c.3141G > A variant induced exon 7 skipping and the synonymous variant was pathogenic. CONCLUSION: Synonymous variants do not change the amino acids encoded by the codon, so we usually consider synonymous variants to be benign and ignore their pathogenicity. Minigene assay is a valuable tool to identify the effect of variation on RNA splicing and identify synonymous variants' benign or pathogenic. We showed that the synonymous variant was pathogenic by minigene assay. WES combined with minigene assay establishes a robust basis for genetic counseling and diagnosing diseases.
Assuntos
Splicing de RNA , Humanos , Splicing de RNA/genética , Deficiências do Desenvolvimento/genética , Éxons/genética , Masculino , Mutação Silenciosa , Sequenciamento do Exoma/métodos , Feminino , Genótipo , Criança , Pré-EscolarRESUMO
BACKGROUND: Cerebral venous sinus thrombosis (CVST) is a rare cause of stroke. Acquired and inherited prothrombotic conditions are the most common risk factors for CVST. Sometimes, an etiology is not found. Wide utilization of next generation sequencing technologies in clinical practice may lead to identification of risk factors other than those classically associated with CVST. METHOD AND RESULTS: This retrospective clinical-laboratory observational study has a reference patient who presented with CVST as an adolescent. Work up for prothrombotic conditions showed high homocysteine level secondary to homozygosity for a common polymorphism, c.677 C > T in the methylenetetrahydrofolate reductase (MTHFR) gene. His older unaffected brother has a similar MTHFR genotype and high homocysteine. The whole exome sequencing revealed a likely pathogenic variant in the sodium voltage gated channel, alpha subunit 1(SCN1A) gene. CONCLUSION: CVST is a multifactorial disease. Prothrombotic conditions are the most common risk factors for CVST. High homocysteine due to the common MTHFR polymorphisms was previously attributed to various thrombotic conditions including CVST. Although high homocysteine due to MTHFR polymorphism may be a contributing factor, additional risk factors such as blood flow abnormalities during SCN1A related seizures may be needed for thrombosis.