In the last decade, undiagnosed disease programs have emerged to address the significant number of individuals with suspected but undiagnosed rare genetic diseases. In our single-center study, we have launched a pilot program for pediatric patients with undiagnosed diseases in the second-largest university hospital in the Czech Republic. This study was prospectively conducted at the Department of Pediatrics at University Hospital Brno between 2020 and 2023. A total of 58 Czech patients with undiagnosed diseases were enrolled in the study. All children underwent singleton WES with targeted phenotype-driven analysis. We identified 28 variants, including 11 pathogenic, 13 likely pathogenic, and 4 VUS according to ACMG guidelines, as diagnostic of genetic diseases in 25 patients, resulting in an overall diagnostic yield of 43%. Eleven variants were novel and had not been previously reported in any public database. The overall clinical utility (actionability) enabling at least one type of change in the medical care of the patient was 76%, whereas the average number of clinical implications to individual patient care was two. Singleton WES facilitated the diagnostic process in the Czech undiagnosed pediatric population. We believe it is an effective approach to enable appropriate counseling, surveillance, and personalized clinical management.
- MeSH
- dítě MeSH
- genetické testování metody MeSH
- kojenec MeSH
- lidé MeSH
- mladiství MeSH
- nediagnostikované nemoci * genetika diagnóza epidemiologie MeSH
- novorozenec MeSH
- předškolní dítě MeSH
- prospektivní studie MeSH
- sekvenování exomu * metody MeSH
- vzácné nemoci * genetika diagnóza MeSH
- Check Tag
- dítě MeSH
- kojenec MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- novorozenec MeSH
- předškolní dítě MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
BACKGROUND: Anaplastic Large Cell Lymphoma (ALCL) is a rare and aggressive T-cell lymphoma, classified into ALK-positive and ALK-negative subtypes, based on the presence of chromosomal translocations involving the ALK gene. The current standard of treatment for ALCL is polychemotherapy, with a high overall survival rate. However, a subset of patients does not respond to or develops resistance to these therapies, posing a serious challenge for clinicians. Recent targeted treatments such as ALK kinase inhibitors and anti-CD30 antibody-drug conjugates have shown promise but, for a fraction of patients, the prognosis is still unsatisfactory. METHODS: We investigated the genetic landscape of ALK + ALCL by whole-exome sequencing; recurring mutations were characterized in vitro and in vivo using transduced ALCL cellular models. RESULTS: Recurrent mutations in FAT family genes and the transcription factor RUNX1T1 were found. These mutations induced changes in ALCL cells morphology, growth, and migration, shedding light on potential factors contributing to treatment resistance. In particular, FAT4 silencing in ALCL cells activated the β-catenin and YAP1 pathways, which play crucial roles in tumor growth, and conferred resistance to chemotherapy. Furthermore, STAT1 and STAT3 were hyper-activated in these cells. Gene expression profiling showed global changes in pathways related to cell adhesion, cytoskeletal organization, and oncogenic signaling. Notably, FAT mutations associated with poor outcome in patients. CONCLUSIONS: These findings provide novel insights into the molecular portrait of ALCL, that could help improve treatment strategies and the prognosis for ALCL patients.
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- anaplastický velkobuněčný lymfom * genetika patologie farmakoterapie MeSH
- fenotyp MeSH
- kadheriny * genetika MeSH
- lidé MeSH
- mutace * MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- prognóza MeSH
- sekvenování exomu MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- genetické testování * MeSH
- lidé MeSH
- mutace genetika MeSH
- nádory prsu * genetika prevence a kontrola MeSH
- nádory vaječníků genetika prevence a kontrola MeSH
- sekvenování exomu MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- novinové články MeSH
AIMS: Recent-onset dilated cardiomyopathy (RODCM) is characterized by heterogeneous aetiology and diverse clinical outcomes, with scarce data on genotype-phenotype correlates. Our aim was to correlate individual RODCM genotypes with left ventricular reverse remodelling (LVRR) and clinical outcomes. METHODS AND RESULTS: In this prospective study, a total of 386 Czech RODCM patients with symptom duration ≤6 months underwent genetic counselling and whole-exome sequencing (WES). The presence of pathogenic (class 5) or likely pathogenic (class 4) variants in a set of 72 cardiomyopathy-related genes was correlated with the occurrence of all-cause death, heart transplantation, or implantation of a ventricular assist device (primary outcome) and/or ventricular arrhythmia event (secondary outcome). LVRR was defined as an improvement of left ventricular ejection fraction to >50% or ≥10% absolute increase, with a left ventricular end-diastolic diameter ≤33 mm/m2 or ≥10% relative decrease. Median follow-up was 41 months. RODCM was familial in 98 (25%) individuals. Class 4-5 variants of interest (VOIs) were identified in 125 (32%) cases, with 69 (18%) having a single titin-truncating variant (TTNtv) and 56 (14%) having non-titin (non-TTN) VOIs. The presence of class 4-5 non-TTN VOIs, but not of TTNtv, heralded a lower probability of 12-month LVRR and proved to be an independent baseline predictor both of the primary and the secondary outcome. The negative result of genetic testing was a strong protective baseline variable against occurrence of life-threatening ventricular arrhythmias. Detection of class 4-5 VOIs in genes coding nuclear envelope proteins was another independent predictor of both study outcomes at baseline and also of life-threatening ventricular arrhythmias after 12 months. Class 4-5 VOIs of genes coding cytoskeleton were associated with an increased risk of life-threatening ventricular arrhythmias after baseline assessment. A positive family history of dilated cardiomyopathy alone only related to a lower probability of LVRR at 12 months and at the final follow-up. CONCLUSIONS: RODCM patients harbouring class 4-5 non-TTN VOIs are at higher risk of progressive heart failure and life-threatening ventricular arrhythmias. Genotyping may improve their early risk stratification at baseline assessment.
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- dilatační kardiomyopatie * genetika patofyziologie MeSH
- dospělí MeSH
- funkce levé komory srdeční fyziologie MeSH
- genotyp * MeSH
- lidé středního věku MeSH
- lidé MeSH
- následné studie MeSH
- prospektivní studie MeSH
- remodelace komor * genetika fyziologie MeSH
- sekvenování exomu MeSH
- tepový objem fyziologie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika MeSH
The aim of this study was to describe the clinical and molecular genetic findings in seven individuals from three unrelated families with Blau syndrome. A complex ophthalmic and general health examination including diagnostic imaging was performed. The NOD2 mutational hot spot located in exon 4 was Sanger sequenced in all three probands. Two individuals also underwent autoinflammatory disorder gene panel screening, and in one subject, exome sequencing was performed. Blau syndrome presenting as uveitis, skin rush or arthritis was diagnosed in four cases from three families. In two individuals from one family, only camptodactyly was noted, while another member had camptodactyly in combination with non-active uveitis and angioid streaks. One proband developed two attacks of meningoencephalitis attributed to presumed neurosarcoidosis, which is a rare finding in Blau syndrome. The probands from families 1 and 2 carried pathogenic variants in NOD2 (NM_022162.3): c.1001G>A p.(Arg334Gln) and c.1000C>T p.(Arg334Trp), respectively. In family 3, two variants of unknown significance in a heterozygous state were found: c.1412G>T p.(Arg471Leu) in NOD2 and c.928C>T p.(Arg310*) in NLRC4 (NM_001199139.1). In conclusion, Blau syndrome is a phenotypically highly variable, and there is a need to raise awareness about all clinical manifestations, including neurosarcoidosis. Variants of unknown significance pose a significant challenge regarding their contribution to etiopathogenesis of autoinflammatory diseases.
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- artritida * genetika diagnóza MeSH
- dědičné zánětlivé autoimunitní nemoci MeSH
- lidé MeSH
- lymfedém genetika diagnóza MeSH
- mutace * MeSH
- nemoci centrálního nervového systému MeSH
- neuropatická artropatie genetika diagnóza MeSH
- rodokmen * MeSH
- sarkoidóza * genetika diagnóza MeSH
- sekvenování exomu MeSH
- signální adaptorový protein Nod2 * genetika MeSH
- synovitida * genetika diagnóza MeSH
- uveitida * genetika diagnóza MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
Genome-wide association studies (GWAS) have identified more than 200 common genetic variants independently associated with colorectal cancer (CRC) risk, but the causal variants and target genes are mostly unknown. We sought to fine-map all known CRC risk loci using GWAS data from 100,204 cases and 154,587 controls of East Asian and European ancestry. Our stepwise conditional analyses revealed 238 independent association signals of CRC risk, each with a set of credible causal variants (CCVs), of which 28 signals had a single CCV. Our cis-eQTL/mQTL and colocalization analyses using colorectal tissue-specific transcriptome and methylome data separately from 1299 and 321 individuals, along with functional genomic investigation, uncovered 136 putative CRC susceptibility genes, including 56 genes not previously reported. Analyses of single-cell RNA-seq data from colorectal tissues revealed 17 putative CRC susceptibility genes with distinct expression patterns in specific cell types. Analyses of whole exome sequencing data provided additional support for several target genes identified in this study as CRC susceptibility genes. Enrichment analyses of the 136 genes uncover pathways not previously linked to CRC risk. Our study substantially expanded association signals for CRC and provided additional insight into the biological mechanisms underlying CRC development.
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- Asijci * genetika MeSH
- běloši * genetika MeSH
- celogenomová asociační studie * MeSH
- genetická predispozice k nemoci * MeSH
- jednonukleotidový polymorfismus * MeSH
- kolorektální nádory * genetika MeSH
- lidé MeSH
- lokus kvantitativního znaku * MeSH
- mapování chromozomů MeSH
- sekvenování exomu MeSH
- studie případů a kontrol MeSH
- transkriptom MeSH
- východní Asiaté MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
BACKGROUND: Tobacco is the main risk factor for developing lung cancer. Yet, while some heavy smokers develop lung cancer at a young age, other heavy smokers never develop it, even at an advanced age, suggesting a remarkable variability in the individual susceptibility to the carcinogenic effects of tobacco. We characterized the germline profile of subjects presenting these extreme phenotypes with Whole Exome Sequencing (WES) and Machine Learning (ML). METHODS: We sequenced germline DNA from heavy smokers who either developed lung adenocarcinoma at an early age (extreme cases) or who did not develop lung cancer at an advanced age (extreme controls), selected from databases including over 6600 subjects. We selected individual coding genetic variants and variant-rich genes showing a significantly different distribution between extreme cases and controls. We validated the results from our discovery cohort, in which we analysed by WES extreme cases and controls presenting similar phenotypes. We developed ML models using both cohorts. FINDINGS: Mean age for extreme cases and controls was 50.7 and 79.1 years respectively, and mean tobacco consumption was 34.6 and 62.3 pack-years. We validated 16 individual variants and 33 variant-rich genes. The gene harbouring the most validated variants was HLA-A in extreme controls (4 variants in the discovery cohort, p = 3.46E-07; and 4 in the validation cohort, p = 1.67E-06). We trained ML models using as input the 16 individual variants in the discovery cohort and tested them on the validation cohort, obtaining an accuracy of 76.5% and an AUC-ROC of 83.6%. Functions of validated genes included candidate oncogenes, tumour-suppressors, DNA repair, HLA-mediated antigen presentation and regulation of proliferation, apoptosis, inflammation and immune response. INTERPRETATION: Individuals presenting extreme phenotypes of high and low risk of developing tobacco-associated lung adenocarcinoma show different germline profiles. Our strategy may allow the identification of high-risk subjects and the development of new therapeutic approaches. FUNDING: See a detailed list of funding bodies in the Acknowledgements section at the end of the manuscript.
- MeSH
- adenokarcinom plic * genetika MeSH
- fenotyp MeSH
- genetická predispozice k nemoci MeSH
- lidé středního věku MeSH
- lidé MeSH
- nádory plic * genetika patologie MeSH
- sekvenování exomu MeSH
- senioři MeSH
- zárodečné buňky patologie MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- senioři MeSH
- Publikační typ
- časopisecké články MeSH
High-grade serous ovarian carcinoma (HGSC) is the most common subtype of ovarian cancer and is among the most fatal gynecological malignancies worldwide, due to late diagnosis at advanced stages and frequent therapy resistance. In 47 HGSC patients, we assessed somatic and germline genetic variability of a custom panel of 144 known or suspected HGSC-related genes by high-coverage targeted DNA sequencing to identify the genetic determinants associated with resistance to platinum-based therapy. In the germline, the most mutated genes were DNAH14 (17%), RAD51B (17%), CFTR (13%), BRCA1 (11%), and RAD51 (11%). Somatically, the most mutated gene was TP53 (98%), followed by CSMD1/2/3 (19/19/36%), and CFTR (23%). Results were compared with those from whole exome sequencing of a similar set of 35 HGSC patients. Somatic variants in TP53 were also validated using GENIE data of 1287 HGSC samples. Our approach showed increased prevalence of high impact somatic and germline mutations, especially those affecting splice sites of TP53, compared to validation datasets. Furthermore, nonsense TP53 somatic mutations were negatively associated with patient survival. Elevated TP53 transcript levels were associated with platinum resistance and presence of TP53 missense mutations, while decreased TP53 levels were found in tumors carrying mutations with predicted high impact, which was confirmed in The Cancer Genome Atlas data (n = 260). Targeted DNA sequencing of TP53 combined with transcript quantification may contribute to the concept of precision oncology of HGSC. Future studies should explore targeting the p53 pathway based on specific mutation types and co-analyze the expression and mutational profiles of other key cancer genes.
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- chemorezistence * genetika MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mutace MeSH
- nádorový supresorový protein p53 * genetika MeSH
- nádory vaječníků * genetika farmakoterapie patologie MeSH
- platina terapeutické užití farmakologie MeSH
- regulace genové exprese u nádorů MeSH
- sekvenování exomu metody MeSH
- senioři MeSH
- serózní cystadenokarcinom * genetika farmakoterapie patologie MeSH
- zárodečné mutace MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND: Whole exome sequencing (WES) and whole genome sequencing (WGS) have become standard methods in human clinical diagnostics as well as in population genomics (POPGEN). Blood-derived genomic DNA (gDNA) is routinely used in the clinical environment. Conversely, many POPGEN studies and commercial tests benefit from easy saliva sampling. Here, we evaluated the quality of variant call sets and the level of genotype concordance of single nucleotide variants (SNVs) and small insertions and deletions (indels) for WES and WGS using paired blood- and saliva-derived gDNA isolates employing genomic reference-based validated protocols. METHODS: The genomic reference standard Coriell NA12878 was repeatedly analyzed using optimized WES and WGS protocols, and data calls were compared with the truth dataset published by the Genome in a Bottle Consortium. gDNA was extracted from the paired blood and saliva samples of 10 participants and processed using the same protocols. A comparison of paired blood-saliva call sets was performed in the context of WGS and WES genomic reference-based technical validation results. RESULTS: The quality pattern of called variants obtained from genomic-reference-based technical replicates correlates with data calls of paired blood-saliva-derived samples in all levels of tested examinations despite a higher rate of non-human contamination found in the saliva samples. The F1 score of 10 blood-to-saliva-derived comparisons ranged between 0.8030-0.9998 for SNVs and between 0.8883-0.9991 for small-indels in the case of the WGS protocol, and between 0.8643-0.999 for SNVs and between 0.7781-1.000 for small-indels in the case of the WES protocol. CONCLUSION: Saliva may be considered an equivalent material to blood for genetic analysis for both WGS and WES under strict protocol conditions. The accuracy of sequencing metrics and variant-detection accuracy is not affected by choosing saliva as the gDNA source instead of blood but much more significantly by the genomic context, variant types, and the sequencing technology used.
- MeSH
- DNA genetika MeSH
- exom MeSH
- genom lidský MeSH
- genomika MeSH
- lidé MeSH
- metagenomika * MeSH
- sekvenování celého genomu MeSH
- sekvenování exomu MeSH
- sliny * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Neurodevelopmental disorders (NDDs) and/or associated multiple congenital abnormalities (MCAs) represent a genetically heterogeneous group of conditions with an adverse prognosis for the quality of intellectual and social abilities and common daily functioning. The rapid development of exome sequencing (ES) techniques, together with trio-based analysis, nowadays leads to up to 50% diagnostic yield. Therefore, it is considered as the state-of-the-art approach in these diagnoses. RESULTS: In our study, we present the results of ES in a cohort of 85 families with 90 children with severe NDDs and MCAs. The interconnection of the in-house bioinformatic pipeline and a unique algorithm for variant prioritization resulted in a diagnostic yield of up to 48.9% (44/90), including rare and novel causative variants (41/90) and intragenic copy-number variations (CNVs) (3/90). Of the total number of 47 causative variants, 53.2% (25/47) were novel, highlighting the clinical benefit of ES for unexplained NDDs. Moreover, trio-based ES was verified as a reliable tool for the detection of rare CNVs, ranging from intragenic exon deletions (GRIN2A, ZC4H2 genes) to a 6-Mb duplication. The functional analysis using PANTHER Gene Ontology confirmed the involvement of genes with causative variants in a wide spectrum of developmental processes and molecular pathways, which form essential structural and functional components of the central nervous system. CONCLUSION: Taken together, we present one of the first ES studies of this scale from the central European region. Based on the high diagnostic yield for paediatric NDDs in this study, 48.9%, we confirm trio-based ES as an effective and reliable first-tier diagnostic test in the genetic evaluation of children with NDDs.
