Cancer cells display complex genomic aberrations that include large-scale genetic rearrangements and epigenetic modulation that are not easily captured by short-read sequencing. This study presents a novel approach for simultaneous profiling of long-range genetic and epigenetic changes in matched cancer samples, focusing on clear cell renal cell carcinoma (ccRCC). ccRCC is a common kidney cancer subtype frequently characterized by a 3p deletion and the inactivation of the von Hippel-Lindau (VHL) gene. We performed integrated genetic, cytogenetic, and epigenetic analyses on paired tumor and adjacent nontumorous tissue samples. Optical genome mapping identified genomic aberrations as structural and copy number variations, complementing exome-sequencing findings. Single-molecule methylome and hydroxymethylome mapping revealed a significant global reduction in 5hmC level in both sample pairs, and a correlation between both epigenetic signals and gene expression was observed. The single-molecule epigenetic analysis identified numerous differentially modified regions, some implicated in ccRCC pathogenesis, including the genes VHL, PRCC, and PBRM1. Notably, pathways related to metabolism and cancer development were significantly enriched among these differential regions. This study demonstrates the feasibility of integrating optical genome and epigenome mapping for comprehensive characterization of matched tumor and adjacent tissue, uncovering both established and novel somatic aberrations.

• MeSH
• DNA vazebné proteiny MeSH
• epigeneze genetická * genetika   MeSH
• epigenom * genetika   MeSH
• karcinom z renálních buněk * genetika   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mapování chromozomů metody   MeSH
• metylace DNA * genetika   MeSH
• nádorový supresorový protein VHL genetika   MeSH
• nádory ledvin * genetika   patologie   MeSH
• regulace genové exprese u nádorů MeSH
• transkripční faktory MeSH
• variabilita počtu kopií segmentů DNA * genetika   MeSH
• Check Tag
• lidé středního věku MeSH
• 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: Paediatric tumours are often characterised by the presence of recurrent DNA copy number alterations (CNAs). These DNA copy number profiles, obtained from a tissue biopsy, can aid in the correct prognostic classification and therapeutic stratification of several paediatric cancer entities (e.g. MYCN amplification in neuroblastoma) and are part of the routine diagnostic practice. Liquid biopsies (LQBs) offer a potentially safer alternative for such invasive tumour tissue biopsies and can provide deeper insight into tumour heterogeneity. PROCEDURE: The robustness and reliability of LQB CNA analyses was evaluated. We performed retrospective CNA profiling using shallow whole-genome sequencing (sWGS) on paired plasma circulating cell-free DNA (cfDNA) and tissue DNA samples from routinely collected samples from paediatric patients (n = 128) representing different tumour entities, including osteosarcoma, Ewing sarcoma, rhabdomyosarcoma, Wilms tumour, brain tumours and neuroblastoma. RESULTS: Overall, we observed a good concordance between CNAs in tissue DNA and cfDNA. The main cause of CNA discordance was found to be low cfDNA sample quality (i.e. the ratio of cfDNA (<700 bp) and high molecular weight DNA (>700 bp)). Furthermore, CNAs were observed that were present in cfDNA and not in tissue DNA, or vice-versa. In neuroblastoma samples, no false-positives or false-negatives were identified for the detection of the prognostic marker MYCN amplification. CONCLUSION: In future prospective studies, CNA analysis on LQBs that are of sufficient quality can serve as a complementary assay for CNA analysis on tissue biopsies, as either cfDNA or tissue DNA can contain CNAs that cannot be identified in the other biomaterial.

• MeSH
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• nádorové biomarkery genetika   MeSH
• předškolní dítě MeSH
• prospektivní studie MeSH
• retrospektivní studie MeSH
• studie proveditelnosti MeSH
• tekutá biopsie metody   MeSH
• variabilita počtu kopií segmentů DNA genetika   MeSH
• volné cirkulující nukleové kyseliny genetika   MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

AIMS: The aim of this retrospective study was to determine the detection rate of the pathogenic copy number variants (CNVs) in a cohort of 33 foetuses - 32 with CHD (congenital heart defects) and 1 with kidney defect, after exclusion of common aneuploidies (trisomy 13, 18, 21, and monosomy X) by karyotyping, Multiplex ligation - dependent probe amplification (MLPA) and chromosomal microarray analysis (CMA). We also assess the effectivity of MLPA as a method of the first tier for quick and inexpensive detection of mutations, causing congenital malformations in foetuses. METHODS: MLPA with probe mixes P070, P036 - Telomere 3 and 5, P245 - microdeletions, P250 - DiGeorge syndrome, and P311 - CHD (Congenital heart defects) was performed in 33 samples of amniotic fluid and chorionic villi. CMA was performed in 10 relevant cases. RESULTS: Pathogenic CNVs were found in 5 samples: microdeletions in region 22q11.2 (≈2 Mb) in two foetuses, one distal microdeletion of the 22q11.2 region containing genes LZTR1, CRKL, AIFM3 and SNAP29 (≈416 kb) in the foetus with bilateral renal agenesis, 8p23.1 (3.8 Mb) microdeletion syndrome and microdeletion in area 9q34.3 (1.7 Mb, Kleefstra syndrome). MLPA as an initial screening method revealed unambiguously pathogenic CNVs in 15.2 % of samples. CONCLUSION: Our study suggests that MLPA and CMA are a reliable and high-resolution technology and should be used as the first-tier test for prenatal diagnosis of congenital heart disease. Determination of the cause of the abnormality is crucial for genetic counselling and further management of the pregnancy.

• MeSH
• lidé MeSH
• pilotní projekty MeSH
• plod MeSH
• retrospektivní studie MeSH
• těhotenství MeSH
• transkripční faktory genetika   MeSH
• variabilita počtu kopií segmentů DNA * genetika   MeSH
• vrozené srdeční vady * diagnóza   genetika   MeSH
• Check Tag
• lidé MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Transcriptome sequencing (RNA-seq) is widely used to detect gene rearrangements and quantitate gene expression in acute lymphoblastic leukemia (ALL), but its utility and accuracy in identifying copy number variations (CNVs) has not been well described. CNV information inferred from RNA-seq can be highly informative to guide disease classification and risk stratification in ALL due to the high incidence of aneuploid subtypes within this disease. Here we describe RNAseqCNV, a method to detect large scale CNVs from RNA-seq data. We used models based on normalized gene expression and minor allele frequency to classify arm level CNVs with high accuracy in ALL (99.1% overall and 98.3% for non-diploid chromosome arms, respectively), and the models were further validated with excellent performance in acute myeloid leukemia (accuracy 99.8% overall and 99.4% for non-diploid chromosome arms). RNAseqCNV outperforms alternative RNA-seq based algorithms in calling CNVs in the ALL dataset, especially in samples with a high proportion of CNVs. The CNV calls were highly concordant with DNA-based CNV results and more reliable than conventional cytogenetic-based karyotypes. RNAseqCNV provides a method to robustly identify copy number alterations in the absence of DNA-based analyses, further enhancing the utility of RNA-seq to classify ALL subtype.

• MeSH
• algoritmy MeSH
• karyotypizace MeSH
• lidé MeSH
• sekvenování transkriptomu MeSH
• variabilita počtu kopií segmentů DNA * genetika   MeSH
• vysoce účinné nukleotidové sekvenování * metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

So-called oncocytic papillary renal cell carcinoma (OPRCC) is a poorly defined variant of papillary renal cell carcinoma. Since its first description, several studies were published with conflicting results, and thus precise definition is lacking. A cohort of 39 PRCCs composed of oncocytic cells were analyzed. Cases were divided into 3 groups based on copy number variation (CNV) pattern. The first group consisted of 23 cases with CNV equal to renal oncocytoma. The second group consisted of 7 cases with polysomy of chromosomes 7 and 17 and the last group of 9 cases included those with variable CNV. Epidemiologic, morphologic and immunohistochemical features varied among the groups. There were not any particular histomorphologic features correlating with any of the genetic subgroups. Further, a combination of morphologic, immunohistochemical, and molecular-genetic features did not allow to precisely predict biologic behavior. Owing to variable CNV pattern in OPRCC, strict adherence to morphology and immunohistochemical profile is recommended, particularly in limited samples (i.e., core biopsy). Applying CNV pattern as a part of a diagnostic algorithm can be potentially misleading. OPRCC is a highly variable group of tumors, which might be misdiagnosed as renal oncocytoma. Using the term OPRCC as a distinct diagnostic entity is, thanks to its high heterogeneity, questionable.

• MeSH
• biopsie dutou jehlou normy   MeSH
• chromozomální aberace MeSH
• chybná diagnóza MeSH
• diferenciální diagnóza MeSH
• dospělí MeSH
• hybridizace in situ fluorescenční metody   MeSH
• imunohistochemie metody   MeSH
• karcinom z renálních buněk epidemiologie   genetika   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• nádorové biomarkery metabolismus   MeSH
• nádory ledvin diagnóza   genetika   patologie   MeSH
• oxyfilní adenom diagnóza   genetika   patologie   MeSH
• oxyfilní buňky metabolismus   patologie   MeSH
• překrývající se geny genetika   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• staging nádorů metody   MeSH
• variabilita počtu kopií segmentů DNA genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

INTRODUCTION: Next-generation sequencing is now used on a routine basis for molecular testing but studies on copy-number variant (CNV) detection from next-generation sequencing data are underrepresented. Utilizing an existing whole-exome sequencing (WES) dataset, we sought to investigate the contribution of rare CNVs to the genetic causality of dystonia. METHODS: The CNV read-depth analysis tool ExomeDepth was applied to the exome sequences of 953 unrelated patients with dystonia (600 with isolated dystonia and 353 with combined dystonia; 33% with additional neurological involvement). We prioritized rare CNVs that affected known disease genes and/or were known to be associated with defined microdeletion/microduplication syndromes. Pathogenicity assessment of CNVs was based on recently published standards of the American College of Medical Genetics and Genomics and the Clinical Genome Resource. RESULTS: We identified pathogenic or likely pathogenic CNVs in 14 of 953 patients (1.5%). Of the 14 different CNVs, 12 were deletions and 2 were duplications, ranging in predicted size from 124bp to 17 Mb. Within the deletion intervals, BRPF1, CHD8, DJ1, EFTUD2, FGF14, GCH1, PANK2, SGCE, UBE3A, VPS16, WARS2, and WDR45 were determined as the most clinically relevant genes. The duplications involved chromosomal regions 6q21-q22 and 15q11-q13. CNV analysis increased the diagnostic yield in the total cohort from 18.4% to 19.8%, as compared to the assessment of single-nucleotide variants and small insertions and deletions alone. CONCLUSIONS: WES-based CNV analysis in dystonia is feasible, increases the diagnostic yield, and should be combined with the assessment of single-nucleotide variants and small insertions and deletions.

• MeSH
• dospělí MeSH
• dystonické poruchy diagnóza   genetika   MeSH
• dystonie diagnóza   genetika   MeSH
• kohortové studie MeSH
• lidé MeSH
• sekvenování exomu * MeSH
• variabilita počtu kopií segmentů DNA * genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Cullin 4B (CUL4B), lysosomal-associated membrane protein Type 2 (LAMP2), ATP1B4, TMEM255A, and ZBTB33 are neighboring genes on Xq24. Mutations in CUL4B result in Cabezas syndrome (CS). Male CS patients present with dysmorphic, neuropsychiatric, genitourinary, and endocrine abnormalities. Heterozygous CS females are clinically asymptomatic. LAMP2 mutations cause Danon disease (DD). Cardiomyopathy is a dominant feature of DD present in both males and heterozygous females. No monogenic phenotypes have been associated with mutations in ATP1B4, TMEM255A, and ZBTB33 genes. To facilitate diagnostics and counseling in CS and DD families, we present a female DD patient with a de novo Alu-mediated Xq24 rearrangement causing a deletion encompassing CUL4B, LAMP2, and also the other three neighboring genes. Typical to females heterozygous for CUL4B mutations, the patient was CS asymptomatic, however, presented with extremely skewed X-chromosome inactivation (XCI) ratios in peripheral white blood cells. As a result of the likely selection against CUL4B deficient clones, only minimal populations (~3%) of LAMP2 deficient leukocytes were identified by flow cytometry. On the contrary, myocardial LAMP2 protein expression suggested random XCI. We demonstrate that contiguous CUL4B and LAMP2 loss-of-function copy number variations occur and speculate that male patients carrying similar defects could present with features of both CS and DD.

• MeSH
• chromozomální delece MeSH
• dospělí MeSH
• elementy Alu genetika   MeSH
• exony genetika   MeSH
• glykogenóza typu IIb diagnóza   genetika   patofyziologie   MeSH
• inaktivace chromozomu X genetika   MeSH
• kardiomyopatie genetika   patofyziologie   MeSH
• kulinové proteiny genetika   MeSH
• lidé MeSH
• membránový protein 2 asociovaný s lyzozomy genetika   MeSH
• mentální retardace vázaná na chromozom X genetika   patofyziologie   MeSH
• mutace ztráty funkce genetika   MeSH
• myokard metabolismus   MeSH
• sodíko-draslíková ATPasa genetika   MeSH
• transkripční faktory genetika   MeSH
• variabilita počtu kopií segmentů DNA genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH

Mutations in the FMS-like tyrosine kinase 3 (FLT3) gene in 13q12.2 are among the most common driver events in acute leukemia, leading to increased cell proliferation and survival through activation of the phosphatidylinositol 3-kinase/AKT-, RAS/MAPK-, and STAT5-signaling pathways. In this study, we examine the pathogenetic impact of somatic hemizygous 13q12.2 microdeletions in B-cell precursor (BCP) acute lymphoblastic leukemia (ALL) using 5 different patient cohorts (in total including 1418 cases). The 13q12.2 deletions occur immediately 5' of FLT3 and involve the PAN3 locus. By detailed analysis of the 13q12.2 segment, we show that the deletions lead to loss of a topologically associating domain border and an enhancer of FLT3. This results in increased cis interactions between the FLT3 promoter and another enhancer located distally to the deletion breakpoints, with subsequent allele-specific upregulation of FLT3 expression, expected to lead to ligand-independent activation of the receptor and downstream signaling. The 13q12.2 deletions are highly enriched in the high-hyperdiploid BCP ALL subtype (frequency 3.9% vs 0.5% in other BCP ALL) and in cases that subsequently relapsed. Taken together, our study describes a novel mechanism of FLT3 involvement in leukemogenesis by upregulation via chromatin remodeling and enhancer hijacking. These data further emphasize the role of FLT3 as a driver gene in BCP ALL.

• MeSH
• akutní lymfatická leukemie genetika   MeSH
• buněčné linie MeSH
• chromozomální delece MeSH
• chromozomální poruchy komplikace   genetika   MeSH
• jednonukleotidový polymorfismus MeSH
• kohortové studie MeSH
• lidé MeSH
• lidské chromozomy, pár 13 genetika   MeSH
• mikročipová analýza MeSH
• regulace genové exprese u leukemie MeSH
• restrukturace chromatinu genetika   fyziologie   MeSH
• sekvenování celého genomu MeSH
• sekvenování transkriptomu MeSH
• tyrosinkinasa 3 podobná fms genetika   MeSH
• upregulace genetika   MeSH
• variabilita počtu kopií segmentů DNA genetika   MeSH
• zesilovače transkripce genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Tumor protein p53 (TP53) is the most frequently mutated gene in cancer1,2. In patients with myelodysplastic syndromes (MDS), TP53 mutations are associated with high-risk disease3,4, rapid transformation to acute myeloid leukemia (AML)5, resistance to conventional therapies6-8 and dismal outcomes9. Consistent with the tumor-suppressive role of TP53, patients harbor both mono- and biallelic mutations10. However, the biological and clinical implications of TP53 allelic state have not been fully investigated in MDS or any other cancer type. We analyzed 3,324 patients with MDS for TP53 mutations and allelic imbalances and delineated two subsets of patients with distinct phenotypes and outcomes. One-third of TP53-mutated patients had monoallelic mutations whereas two-thirds had multiple hits (multi-hit) consistent with biallelic targeting. Established associations with complex karyotype, few co-occurring mutations, high-risk presentation and poor outcomes were specific to multi-hit patients only. TP53 multi-hit state predicted risk of death and leukemic transformation independently of the Revised International Prognostic Scoring System (IPSS-R)11. Surprisingly, monoallelic patients did not differ from TP53 wild-type patients in outcomes and response to therapy. This study shows that consideration of TP53 allelic state is critical for diagnostic and prognostic precision in MDS as well as in future correlative studies of treatment response.

• MeSH
• alely MeSH
• analýza přežití MeSH
• fenotyp MeSH
• frekvence genu MeSH
• kohortové studie MeSH
• lidé MeSH
• mutace MeSH
• mutační analýza DNA MeSH
• myelodysplastické syndromy diagnóza   genetika   mortalita   terapie   MeSH
• nádorový supresorový protein p53 genetika   MeSH
• nestabilita genomu genetika   MeSH
• prognóza MeSH
• variabilita počtu kopií segmentů DNA genetika   MeSH
• výsledek terapie MeSH
• ztráta heterozygozity genetika   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Centromeric and pericentromeric chromosome regions are occupied by satellite DNA. Satellite DNAs play essential roles in chromosome segregation, and, thanks to their extensive sequence variability, to some extent, they can also be used as phylogenetic markers. In this paper, we isolated and sequenced satellite DNA I-IV in 11 species of Cervidae. The obtained satellite DNA sequences and their chromosomal distribution were compared among the analysed representatives of cervid subfamilies Cervinae and Capreolinae. Only satI and satII sequences are probably present in all analysed species with high abundance. On the other hand, fluorescence in situ hybridisation (FISH) with satIII and satIV probes showed signals only in a part of the analysed species, indicating interspecies copy number variations. Several indices, including FISH patterns, the high guanine and cytosine (GC) content, and the presence of centromere protein B (CENP-B) binding motif, suggest that the satII DNA may represent the most important satellite DNA family that might be involved in the centromeric function in Cervidae. The absence or low intensity of satellite DNA FISH signals on biarmed chromosomes probably reflects the evolutionary reduction of heterochromatin following the formation of chromosome fusions. The phylogenetic trees constructed on the basis of the satellite I-IV DNA relationships generally support the present cervid taxonomy.

• MeSH
• centromera genetika   MeSH
• heterochromatin genetika   MeSH
• hybridizace in situ fluorescenční metody   MeSH
• lidé MeSH
• přežvýkavci genetika   MeSH
• repetitivní sekvence nukleových kyselin genetika   MeSH
• satelitní DNA genetika   MeSH
• variabilita počtu kopií segmentů DNA genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH