RESUMO
BACKGROUND: Cellular angiofibroma, a rare benign mesenchymal neoplasm, is classified within the 13q/RB1 family of tumors due to morphological, immunohistochemical, and genetic similarities with spindle cell lipoma. Here, genetic data reveal pathogenetic heterogeneity in cellular angiofibroma. METHODS: Three cellular angiofibromas were studied using G-banding/Karyotyping, array comparative genomic hybridization, RNA sequencing, and direct cycling sequencing. RESULTS: The first tumor carried a del(13)(q12) together with heterozygous loss and minimal expression of the RB1 gene. Tumors two and three displayed chromosome 8 abnormalities associated with chimeras of the pleomorphic adenoma gene 1 (PLAG1). In tumor 2, the cathepsin B (CTSB) fused to PLAG1 (CTSB::PLAG1) while in tumor 3, the mir-99a-let-7c cluster host gene (MIR99AHG) fused to PLAG1 (MIR99AHG::PLAG1), both leading to elevated expression of PLAG1 and insulin growth factor 2. CONCLUSION: This study uncovers two genetic pathways contributing to the pathogenetic heterogeneity within cellular angiofibromas. The first aligns with the 13q/RB1 family of tumors and the second involves PLAG1-chimeras. These findings highlight the diverse genetic landscape of cellular angiofibromas, providing insights into potential diagnostic strategies.
Assuntos
Angiofibroma , Cromossomos Humanos Par 13 , Heterogeneidade Genética , Humanos , Angiofibroma/genética , Angiofibroma/patologia , Masculino , Cromossomos Humanos Par 13/genética , Proteínas de Ligação a DNA/genética , Adulto , Feminino , Proteínas de Ligação a Retinoblastoma/genética , MicroRNAs/genética , Ubiquitina-Proteína Ligases/genética , Pessoa de Meia-Idade , Hibridização Genômica Comparativa , Cromossomos Humanos Par 8/genética , Catepsina BRESUMO
AIMS: The methylation status of the O6-methylguanine-DNA methyltransferase (MGMT) promoter region is essential in evaluating the prognosis and predicting the drug response in patients with glioblastoma. In this study, we evaluated the utility of using nanopore long-read sequencing as a method for assessing methylation levels throughout the MGMT CpG-island, compared its performance to established techniques and demonstrated its clinical applicability. METHODS: We analysed 165 samples from CNS tumours, focusing on the MGMT CpG-island using nanopore sequencing. Oxford Nanopore Technologies (ONT) MinION and PromethION flow cells were employed for single sample or barcoded assays, guided by a CRISPR/Cas9 protocol, adaptive sampling or as part of a whole genome sequencing assay. Methylation data obtained through nanopore sequencing were compared to results obtained via pyrosequencing and methylation bead arrays. Hierarchical clustering was applied to nanopore sequencing data for patient stratification. RESULTS: Nanopore sequencing displayed a strong correlation (R2 = 0.91) with pyrosequencing results for the four CpGs of MGMT analysed by both methods. The MGMT-STP27 algorithm's classification was effectively reproduced using nanopore data. Unsupervised hierarchical clustering revealed distinct patterns in methylated and unmethylated samples, providing comparable survival prediction capabilities. Nanopore sequencing yielded high-confidence results in a rapid timeframe, typically within hours of sequencing, and extended the analysis to all 98 CpGs of the MGMT CpG-island. CONCLUSIONS: This study presents nanopore sequencing as a valid and efficient method for determining MGMT promotor methylation status. It offers a comprehensive view of the MGMT promoter methylation landscape, which enables the identification of potentially clinically relevant subgroups of patients. Further exploration of the clinical implications of patient stratification using nanopore sequencing of MGMT is warranted.
Assuntos
Metilação de DNA , Sequenciamento por Nanoporos , Regiões Promotoras Genéticas , Humanos , Sequenciamento por Nanoporos/métodos , Regiões Promotoras Genéticas/genética , Ilhas de CpG/genética , Proteínas Supressoras de Tumor/genética , Metilases de Modificação do DNA/genética , Enzimas Reparadoras do DNA/genética , Neoplasias Encefálicas/genética , Feminino , Masculino , Glioblastoma/genética , IdosoRESUMO
Endometrial stromal sarcomas (ESS) are a heterogeneous group of rare mesenchymal cancers. Considerable knowledge has been gained in recent years about the molecular characteristics of these cancers, which helps to classify them in a more meaningful manner leading to improved diagnosis, prognostication, and treatment. According to this classification, ESS is now grouped as low- or high-grade. ESS may have overlapping clinical presentation, morphology, and immunohistochemical profile. Their genetic characteristics allow subdivision of many of them depending on which pathogenetically important fusion genes they carry, but clearly much more needs to be unraveled in this regard. We here provide an overview of the molecular pathogenetic knowledge gained so far on low- and high-grade ESS.
Assuntos
Neoplasias do Endométrio/patologia , Sarcoma do Estroma Endometrial/patologia , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/metabolismo , Feminino , Humanos , Estadiamento de Neoplasias , Prognóstico , Sarcoma do Estroma Endometrial/genética , Sarcoma do Estroma Endometrial/metabolismoRESUMO
OBJECTIVE: The aim of this study was to analyse the promoter methylation status of the gene O6-methylguanine-DNA methyltransferase (MGMT) in malignant effusions, with focus on serous carcinoma. METHODS: Fresh-frozen cell pellets from 81 effusions (42 peritoneal, 38 pleural, one pericardial), consisting of 71 carcinomas of different origin (33 ovarian, 23 breast, six lung, five uterine corpus and four cervical carcinomas) and 10 malignant mesotheliomas, were analysed for MGMT methylation using pyrosequencing analysis. RESULTS: MGMT methylation at all four cytosine-guanine dinucleotide sites examined was detected in only 2/81 (2%) specimens, consisting of a high-grade serous carcinoma with high frequency of methylation, and a breast carcinoma with low methylation frequency. CONCLUSION: The findings in the present study suggest that MGMT methylation is a rare epigenetic change in malignant effusions of different origin.
Assuntos
Carcinoma/genética , Metilação de DNA/genética , Metilases de Modificação do DNA/genética , Enzimas Reparadoras do DNA/genética , Epigênese Genética/genética , Regiões Promotoras Genéticas/genética , Proteínas Supressoras de Tumor/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Sarcomas account for 3% of all uterine malignancies and many of them are characterized by acquired, specific fusion genes whose detection has increased pathogenetic knowledge and diagnostic precision. We describe a novel fusion gene, GREB1-NCOA2, detected by transcriptome sequencing and validated by reverse transcriptase polymerase chain reaction and Sanger sequencing in an undifferentiated uterine sarcoma. The chimeric transcript was an in-frame fusion between exon 3 of GREB1 and exon 15 of NCOA2. The fusion is reported here for the first time, but it involves the GREB1 gene, an important promoter of tumor growth and progression, and NCOA2 which is known to be involved in transcriptional regulation. The alteration and recombination of these genes played a role in the tumorigenesis and/or progression of this sarcoma.
Assuntos
Proteínas de Neoplasias/genética , Coativador 2 de Receptor Nuclear/genética , Proteínas de Fusão Oncogênica/genética , Sarcoma/genética , Neoplasias Uterinas/genética , Sequência de Bases , Cromossomos Humanos Par 2 , Cromossomos Humanos Par 8 , Feminino , Humanos , Hibridização in Situ Fluorescente , Pessoa de Meia-Idade , Proteínas de Neoplasias/metabolismo , Coativador 2 de Receptor Nuclear/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Translocação GenéticaRESUMO
Formation of fusion genes is pathogenetically crucial in many solid tumors. They are particularly characteristic of several mesenchymal tumors, but may also be found in epithelial neoplasms. Ovarian carcinomas, too, may harbor fusion genes but only few of these were found to be recurrent with a rate ranging from 0.5 to 5%. Because most attempts to find specific and recurrent fusion transcripts in ovarian carcinomas focused exclusively on high-grade serous carcinomas, the situation in the other carcinoma subgroups remains largely uninvestigated as far as fusion genes are concerned. We performed transcriptome sequencing on a series of 34 samples from ovarian tumors that included borderline, clear cell, mucinous, endometrioid, low-grade and high-grade serous carcinomas in search of fusion genes typical of these subtypes. We found a total of 24 novel fusion transcripts. The PCMTDI-CCNL2 fusion transcript, which involves a member of the cyclin family, was found recurrently involved but only in endometrioid carcinomas (4 of 18 tumors; 22%). We also found three additional fusion transcripts involving genes belonging to the cyclin family: ANXA5-CCNA2 and PDE4D-CCNB1 were detected in two endometrioid carcinomas, whereas CCNY-NRG4 was identified in a clear cell carcinoma. The recurrent involvement of CCNL2 in four fusions and of three other genes of the cyclin family in three additional transcripts hints that deregulation of cyclin genes is important in the pathogenesis of ovarian carcinomas in general but of endometrioid carcinomas particularly.
Assuntos
Ciclinas/genética , Neoplasias do Endométrio/genética , Fusão Gênica , Recidiva Local de Neoplasia/genética , Proteínas de Fusão Oncogênica/genética , Neoplasias Ovarianas/genética , Adenocarcinoma de Células Claras/genética , Adenocarcinoma de Células Claras/patologia , Adenocarcinoma Mucinoso/genética , Adenocarcinoma Mucinoso/patologia , Biomarcadores Tumorais/genética , Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/patologia , Neoplasias do Endométrio/patologia , Feminino , Seguimentos , Humanos , Recidiva Local de Neoplasia/patologia , Neoplasias Ovarianas/patologia , PrognósticoRESUMO
High-grade endometrial stromal sarcoma likely encompasses underrecognized tumors harboring genetic abnormalities besides YWHAE-NUTM2 fusion. Triggered by three initial endometrial stromal sarcomas with ZC3H7B-BCOR fusion characterized by high-grade morphology and aggressive clinical behavior, we herein investigate the clinicopathologic features of this genetic subset by expanding the analysis to 17 such tumors. All of them occurred in adult women with a median age of 54 (range, 28-71) years. They were predominantly based in the endomyometrium and demonstrated tongue-like and/or pushing myometrial invasion. Most were uniformly cellular and displayed haphazard fascicles of spindle cells with mild to moderate nuclear atypia. Myxoid matrix was seen in 14 of 17 (82%) tumors, and collagen plaques were seen in 8 (47%). The mitotic index was ≥10 mitotic figures/10 high-power fields (HPFs) in 14 of 17 (82%) tumors with a median of 14.5 mitotic figures/10 HPFs. No foci of conventional or variant low-grade endometrial stromal sarcoma were seen. All tumors expressed CD10 with only limited or absent desmin, SMA and/or h-caldesmon staining. ER and PR expression in >5% of cells was seen in 4 of 12 (33%) tumors. Diffuse cyclin D1 and BCOR immunoreactivity was present in 7 of 8 (88%) and 7 of 14 (50%) tumors, respectively. Fluorescence in situ hybridization or targeted RNA sequencing confirmed ZC3H7B-BCOR fusion in all tumors, including four and two previously diagnosed as myxoid leiomyosarcoma and undifferentiated uterine sarcoma, respectively. Limited clinical data suggest that patients present at higher stage and have worse prognosis compared with published outcomes in low-grade endometrial stromal sarcoma. Tumors with ZC3H7B-BCOR fusion constitute a distinct group of endometrial stromal sarcomas with high-grade morphology that should be distinguished from other uterine mesenchymal neoplasms that may demonstrate myxoid morphology.
Assuntos
Neoplasias do Endométrio/patologia , Proteínas de Fusão Oncogênica/genética , Proteínas Proto-Oncogênicas/genética , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Sarcoma do Estroma Endometrial/patologia , Adulto , Idoso , Neoplasias do Endométrio/genética , Feminino , Humanos , Pessoa de Meia-Idade , Sarcoma do Estroma Endometrial/genéticaRESUMO
We present a new endometrial stromal sarcoma (ESS)-associated genomic rearrangement involving chromosome arms 5p and 6p and leading to the formation of a BRD8-PHF1 fusion gene. The PHF1 (PHD finger protein 1) gene, from 6p21, is known to be rearranged in ESS in a promiscuous way inasmuch as it has been shown to recombine with JAZF1, EPC1, MEAF6, and now also with BRD8, in tumors of this type. In all rearrangements of PHF1, including the present one, a recurrent theme is that the entire coding part of PHF1 constitutes the 3' end of the fusion. BRD8 (bromodomain containing 8) encodes a protein which is involved in regulation of protein acetylation and/or histone acetyl transferase activity. All the genetic fusions identified so far in ESS appear to recombine genes involved in transcriptional regulation, that is, polycomb group complex-mediated and aberrant methylation/acetylation genes. This adds to the likelihood that the new BRD8-PHF1 shares the same pathogenetic mechanism as the other ESS-specific rearrangements.
Assuntos
Proteínas de Ligação a DNA/genética , Neoplasias do Endométrio/genética , Proteínas de Fusão Oncogênica/genética , Proteínas do Grupo Polycomb/genética , Receptores dos Hormônios Tireóideos/genética , Sarcoma do Estroma Endometrial/genética , Células Cultivadas , Cromossomos Humanos Par 5/genética , Proteínas de Ligação a DNA/metabolismo , Neoplasias do Endométrio/patologia , Feminino , Humanos , Pessoa de Meia-Idade , Fusão Oncogênica , Proteínas de Fusão Oncogênica/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Receptores dos Hormônios Tireóideos/metabolismo , Sarcoma do Estroma Endometrial/patologia , Fatores de TranscriçãoRESUMO
The identification of recurrent gene fusions in common epithelial cancers--for example, TMPRSS2/ERG in prostate cancer and EML4/ALK in nonsmall cell lung carcinomas--has raised the question of whether fusion genes are pathogenetically important also in ovarian carcinomas. The first recurrent fusion transcript in serous ovarian carcinomas was reported by Salzman et al. in 2011, who used deep paired-end sequencing to detect the fusion gene ESRRA-C11orf20 in 10 out of 67 (15%) serous ovarian carcinomas examined, a finding that holds great promise for our understanding of ovarian tumorigenesis as well as, potentially, for new treatment strategies. We wanted to test how frequent the ESRRA/C11orf20 fusion is in ovarian carcinomas of all subtypes, and therefore examined a series of 230 ovarian carcinomas of which 197 were of the serous subtype and 163 of the 197 were of stages III and IV--that is, the very same carcinoma subset where the fusion transcript had been found. We performed PCR and high-throughput sequencing analyses in search of the fusion transcript. We used the same primers described previously for the detection of the fusion and the same primer combination, but found no ESRRA/C11orf20 fusion in our series. A synthetic DNA plasmid containing the reported ESRRA/C11orf20 fusion was included as a positive control for our PCR experiments. Data from high-throughput sequencing of 23 ovarian carcinomas were screened in search of alternative partner(s) for the ESRRA and/or C11orf20 gene, but none was found. We conclude that the frequency of the ESRRA/C11orf20 gene fusion in serous ovarian carcinomas of stages III and IV must be considerable less than that reported previously (0/163 in our experience compared with 10/67 in the previous study). At the very least, it seems clear that the said fusion cannot be a common pathogenetic event in this tumor type.
Assuntos
Adenocarcinoma/genética , Proteínas Nucleares/genética , Proteínas de Fusão Oncogênica/genética , Neoplasias Ovarianas/genética , Receptores de Estrogênio/genética , Adenocarcinoma/patologia , Feminino , Frequência do Gene , Humanos , Estadiamento de Neoplasias , Neoplasias Ovarianas/patologia , Receptor ERRalfa Relacionado ao EstrogênioRESUMO
Recent cytogenetic and molecular investigations have improved our understanding of endometrial stromal tumors, including sarcomas (ESS), and helped redefine their classification into more pathogenetically meaningful categories. Because much more can be gained through such studies, we add information on another 22 ESS examined by karyotyping, PCR analysis, expression array analysis, and transcriptome sequencing. In spite of the known preference for certain pathogenetic pathways, we found considerable genetic heterogeneity in high-grade (HG) as well as in low-grade (LG) ESS. Not all HG tumors showed a YWHAE-NUTM chimeric transcript and as many as six LGESS showed no hitherto known ESS-related fusions. Among the transcripts identified by transcriptome sequencing and verified by Sanger sequencing, new variants of ZC3H7-BCOR and its reciprocal BCOR-ZC3H7 were identified as was involvement of the CREBBP and MLLT4 genes (both well known leukemia-related genes) in two new fusions. FISH analysis identified a known EPC1-PHF1 fusion which led to the identification of a new variant at the molecular level. The fact that around 70 genes were found differentially expressed, by microarray analysis, when comparing LGESS showing ESS-related fusions with LGESS without such transcripts, underscores the biochemical importance of the observed genetic heterogeneity and hints that new subgroups/entities in LGESS still remain undiscovered. © 2016 The Authors. Genes, Chromosomes & Cancer Published by Wiley Periodicals, Inc.
Assuntos
Citogenética , Heterogeneidade Genética , Sarcoma do Estroma Endometrial/patologia , Transcriptoma/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Gradação de Tumores , Sarcoma do Estroma Endometrial/classificação , Sarcoma do Estroma Endometrial/genéticaRESUMO
We have previously identified two ALK rearrangements in a subset of ependymal tumors using a combination of cytogenetic data and RNA sequencing. The aim of this study was to perform an unbiased search for fusion transcripts in our entire series of ependymal tumors. Fusion analysis was performed using the FusionCatcher algorithm on 12 RNA-sequenced ependymal tumors. Candidate transcripts were prioritized based on the software's filtering and manual visualization using the BLAST (Basic Local Alignment Search Tool) and BLAT (BLAST-like alignment tool) tools. Genomic and reverse transcriptase PCR with subsequent Sanger sequencing was used to validate the potential fusions. Fluorescent in situ hybridization (FISH) using locus-specific probes was also performed. A total of 841 candidate chimeric transcripts were identified in the 12 tumors, with an average of 49 unique candidate fusions per tumor. After algorithmic and manual filtering, the final list consisted of 24 potential fusion events. Raw RNA-seq read sequences and PCR validation supports two novel fusion genes: a reciprocal fusion gene involving UQCR10 and C1orf194 in an adult spinal ependymoma and a TSPAN4-CD151 fusion gene in a pediatric infratentorial anaplastic ependymoma. Our previously reported ALK rearrangements and the RELA and YAP1 fusions found in supratentorial ependymomas were until now the only known fusion genes present in ependymal tumors. The chimeric transcripts presented here are the first to be reported in infratentorial or spinal ependymomas. Further studies are required to characterize the genomic rearrangements causing these fusion genes, as well as the frequency and functional importance of the fusions. © 2016 Wiley Periodicals, Inc.
Assuntos
Biomarcadores Tumorais/genética , Neoplasias Encefálicas/genética , Ependimoma/genética , Proteínas de Fusão Oncogênica/genética , Adulto , Idoso , Neoplasias Encefálicas/classificação , Neoplasias Encefálicas/patologia , Estudos de Casos e Controles , Ependimoma/classificação , Ependimoma/patologia , Feminino , Seguimentos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Hibridização in Situ Fluorescente , Lactente , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Software , Taxa de Sobrevida , Tetraspanina 24/genética , Tetraspaninas/genéticaRESUMO
Leiomyomas of the gastrointestinal tract are mostly found in the esophagus, stomach, and colon. Genetic information about them is very limited and no fusion genes have been described. We present herein cytogenetic and molecular genetic analyses of two gastrointestinal leiomyomas found in the esophagus and small intestine. The esophageal leiomyoma had the karyotype 45,Y,der(X)t(X;6)(p22;p21),inv(2)(p23q35),add(6)(p21),-11[cp6]/46,XY[7]. The intestinal leiomyoma karyotype was 46,X,add(X)(q2?),der(2)add(2)(p23)add(2)(q33),add(4)(p14),add(14)(q22)[10]/47,XX,+12[2]/46,XX[1]. RNA-sequencing detected FN1-ALK fusion transcripts in both tumors. RT-PCR together with Sanger sequencing verified the presence of the FN1-ALK fusion transcripts. Fluorescence in situ hybridization using an ALK breakapart probe further confirmed the rearrangement of the ALK gene. Immunohistochemical investigation of ALK in the leiomyoma of the small intestine revealed positivity with strong granular cytoplasmatic staining in the tumor cells. This is the first ever ALK fusion reported in gastrointestinal leiomyomas. Our results are of potential clinical importance because crizotinib, a selective ALK inhibitor, has demonstrated effect in patients whose tumors harbor ALK rearrangements. Thus, ALK emerges as a possible therapeutic target in patients whose tumors, including gastrointestinal leiomyomas, carry ALK fusions.
Assuntos
Fibronectinas/genética , Neoplasias Gastrointestinais/genética , Leiomioma/genética , Receptores Proteína Tirosina Quinases/genética , Adulto , Quinase do Linfoma Anaplásico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas de Fusão Oncogênica/genética , RecidivaRESUMO
Endometrial stromal sarcomas (ESS) are genetically heterogeneous uterine tumors in which a JAZF1-SUZ12 chimeric gene resulting from the chromosomal translocation t(7;17)(p15;q21) as well as PHF1 rearrangements (in chromosomal band 6p21) with formation of JAZF1-PHF1, EPC1-PHF1, and MEAF6-PHF1 chimeras have been described. Here, we investigated two ESS characterized cytogenetically by the presence of a der(22)t(X;22)(p11;q13). Whole transcriptome sequencing one of the tumors identified a ZC3H7-BCOR chimeric transcript. Reverse transciptase-PCR with the ZC3H7B forward and BCOR reverse primer combinations confirmed the presence of a ZC3H7-BCOR chimeric transcript in both ESS carrying a der(22)t(X;22) but not in a control ESS with t(1;6) and the MEAF6-PHF1 fusion. Sequencing of the amplified cDNA fragments showed that in both cases ESS exon 10 of ZC3H7B (from 22q13; accession number NM_017590 version 4) was fused to exon 8 of BCOR (from Xp11; accession number NM_001123385 version 1). Reciprocal multiple BCOR-ZC3H7B cDNA fragments were amplified in only one case suggesting that ZC3H7B-BCOR, on the der(22)t(X;22), is the pathogenetically important fusion gene. The putative ZC3H7B-BCOR protein would contain the tetratricopeptide repeats and LD motif from ZC3H7B and the AF9 binding site (1093-1233aa), the 3 ankyrin repeats (1410-1509 aa), and the NSPC1 binding site of BCOR. Although the presence of these motifs suggests various functions of the chimeric protein, it is possible that its most important role may be in epigenetic regulation. Whether or not the (patho)genetic subsets JAZF1-SUZ12, PHF1 rearrangements, and ZC3H7B-BCOR correspond to any phenotypic, let alone clinically important, differences in ESS remain unknown.
Assuntos
Proteínas de Fusão Oncogênica/genética , Proteínas Proto-Oncogênicas/genética , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Sarcoma do Estroma Endometrial/genética , Translocação Genética , Adulto , Cromossomos Humanos Par 22/genética , Cromossomos Humanos X/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Rearranjo Gênico , Humanos , Hibridização in Situ Fluorescente , Pessoa de Meia-Idade , Proteínas de Fusão Oncogênica/isolamento & purificação , Sarcoma do Estroma Endometrial/etiologia , Sarcoma do Estroma Endometrial/patologia , Translocação Genética/genéticaRESUMO
Mesothelioma is a rare but very aggressive tumor derived from mesothelial cells. A number of often complex but nonrandom cytogenetic abnormalities have been found in these tumors, resulting in loss of chromosome bands 14q32 and 22q12 in more than 35% of the cases. In this study, we used RNA sequencing to search for fusion transcripts in a mesothelioma carrying a t(14;22)(q32;q12) as the sole chromosomal aberration and found an EWSR1-YY1 and its reciprocal YY1-EWSR1 fusion transcript. Screening 15 additional cases of mesothelioma from which we had RNA but no cytogenetic information, we identified one more tumor carrying an EWSR1-YY1 fusion gene but not the reciprocal YY1-EWSR1 transcript. RT-polymerase chain reaction and sequencing showed that in both cases exon 8 of EWSR1 (nucleotide 1,139, accession number NM_013986 version 3, former exon 7 in sequence with accession number X66899) was fused to exon 2 of YY1 (nucleotide 1,160, accession number NM_003403 version 3). The EWSR1 breakpoint in exon 8 in the EWSR1-YY1 chimeric transcript is similar to what is found in other fusions involving EWSR1 such as EWSR1-FLI1, EWSR1-DDIT3, and EWSR1-ATF1. The EWSR1-YY1-encoded protein is an abnormal transcription factor with the transactivation domain of EWSR1 and the DNA-binding domain of YY1. This is the first study to detect a specific fusion gene in mesothelioma (the reason how frequent the EWSR1-YY1 fusion is remains uncertain) and also the first time that direct involvement of YY1 in oncogenesis has been demonstrated.
Assuntos
Proteínas de Ligação a Calmodulina/genética , Mesotelioma/genética , Proteínas de Fusão Oncogênica/genética , Proteínas de Ligação a RNA/genética , Fator de Transcrição YY1/genética , Adulto , Idoso , Proteínas de Ligação a Calmodulina/isolamento & purificação , Cromossomos Humanos Par 14/genética , Cromossomos Humanos Par 22/genética , Humanos , Hibridização in Situ Fluorescente , Masculino , Mesotelioma/patologia , Pessoa de Meia-Idade , Proteínas de Fusão Oncogênica/isolamento & purificação , Proteína EWS de Ligação a RNA , Proteínas de Ligação a RNA/isolamento & purificação , Análise de Sequência de RNA , Translocação Genética , Fator de Transcrição YY1/isolamento & purificaçãoRESUMO
Little is known about the genomic abnormalities of squamous cell carcinomas (SCC) of the vulva and how they correlate with gene expression. We determined the genomic and expression profiles of 15 such SCC using karyotyping, DNA ploidy analysis, arrayCGH, and expression arrays. Four of the five cases with clonal chromosomal aberrations found by G-banding showed highly abnormal karyotypes with multiple rearrangements. The imbalances scored by arrayCGH mapped to different chromosomes with losses being more common than gains. Frequent losses were scored from 3p and 8p whereas gains were frequent from 3q and 8q (loss of 8p with concomitant gain of 8q mostly occurred via 8q isochromosome formation). This is the first study of vulvar tumors using arrayCGH, and some frequent imbalances could be defined precisely. Of particular note were the sometimes large, sometimes small deletions of 3p and 9p which had minute areas in 3p14 and 9p23 as minimal commonly deleted regions. FHIT (3p14) and PTPRD (9p23) are the only genes here. They were both lost in seven cases, including homozygous losses of PTPRD in four tumors. Using qPCR we could demonstrate deregulation of the FHIT gene in tumor cells. Hence, this gene is likely to play a pathogenetic role in vulvar SCC tumorigenesis. Expression array analyses also identified a number of other genes whose expression profile was altered. Notable among the downregulated genes were MAL (in 2q11), KRT4 (in 12q13), and OLFM4 (in 13q14), whereas upregulated genes included SPRR2G (in 1q21.3) and S100A7A (in 1q21.3).
Assuntos
Biomarcadores Tumorais/genética , Carcinoma de Células Escamosas/genética , Aberrações Cromossômicas , Perfilação da Expressão Gênica , Genoma Humano , Neoplasias Vulvares/genética , Cromossomos Humanos/genética , Hibridização Genômica Comparativa , Feminino , Humanos , Cariotipagem , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais CultivadasRESUMO
BACKGROUND/AIM: The term "calcified chondroid mesenchymal neoplasm" was introduced in 2021 to describe a group of tumors characterized by various morphological features, including the formation of cartilage or chondroid matrix. These tumors frequently carry chimeric genes where the 5'-end partner gene is fibronectin 1 and the 3'-end partner gene codes for receptor tyrosine kinase. Our study explores fusion of the genes platelet-derived growth factor receptor alpha (PDGFRA) and ubiquitin-specific peptidase 8 (USP8) in calcified chondroid mesenchymal neoplasm. CASE REPORT: Genetic investigations were conducted on a tumor located in the leg of a 71-year-old woman. G-banding analysis of short-term cultured tumor cells revealed the karyotype 46,XX,t(4;15)(q12;q21)[6]/46,XX[4]. RNA sequencing detected in-frame PDGFRA::USP8 and USP8::PDGFRA chimeric transcripts, which were validated by RT-PCR/Sanger sequencing. The PDGFRA::USP8 chimeric protein is predicted to have cell membrane location and functions as a chimeric ubiquitinyl hydrolase. The USP8::PDGFRA protein was predicted to be nuclear and function as a positive regulator of cellular metabolic process. CONCLUSION: We report, for the first time, a calcified chondroid mesenchymal neoplasm carrying a balanced t(4;15)(q12;q21) chromosomal translocation, resulting in the generation of both PDGFRA::USP8 and USP8::PDGFRA chimeras. The PDGFRA::USP8 protein is located on the cell membrane and functions as a chimeric ubiquitinyl hydrolase, activated by PDGFs. Conversely, USP8::PDGFRA is a nuclear protein regulating metabolic processes.
Assuntos
Receptor alfa de Fator de Crescimento Derivado de Plaquetas , Humanos , Feminino , Idoso , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Translocação Genética , Ubiquitina Tiolesterase/genética , Ubiquitina Tiolesterase/metabolismo , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Calcinose/genética , Calcinose/patologia , Cromossomos Humanos Par 4/genéticaRESUMO
BACKGROUND/AIM: In precursor B-cell lineage acute lymphoblastic leukemia (BCP-ALL), leukemic cells harbor genetic abnormalities that play an important role in the diagnosis, prognosis, and treatment. A subgroup of BCP-ALL is characterized by the presence of a Philadelphia (Ph) chromosome and a chimeric BCR::ABL1 gene, whereas in another subgroup, leukemic cells exhibit near-haploidy with chromosome number 24-30. This study presents the third documented case of BCP-ALL in which a near haploid clone concurrently displayed a Ph chromosome/BCR::ABL1. CASE REPORT: Bone marrow cells obtained at diagnosis from a 25-year-old man with BCP-ALL were genetically investigated using G-banding, fluorescence in situ hybridization, and array comparative genomic hybridization. Leukemic cells had an abnormal karyotype 28
Assuntos
Cromossomo Filadélfia , Leucemia-Linfoma Linfoblástico de Células Precursoras , Masculino , Humanos , Adulto , Haploidia , Hibridização in Situ Fluorescente , Hibridização Genômica Comparativa , Aberrações Cromossômicas , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Cariótipo , Proteínas de Fusão bcr-abl/genética , Translocação GenéticaRESUMO
BACKGROUND/AIM: Constitutional chromosomal aberrations are rare in hematologic malignancies and their pathogenetic role is mostly poorly understood. We present a comprehensive molecular characterization of a novel constitutional chromosomal translocation found in two siblings - sisters - diagnosed with myelodysplastic syndrome (MDS). MATERIALS AND METHODS: Bone marrow and blood cells from the two patients were examined using G-banding, RNA sequencing, PCR, and Sanger sequencing. RESULTS: We identified a balanced t(17;19)(q21;p13) translocation in both siblings' bone marrow, blood cells, and phytohemagglutinin-stimulated lymphocytes. The translocation generated a MYO1F::WNK4 chimera on the der(19)t(17;19), encoding a chimeric serine/threonine kinase, and a VPS25::MYO1F on the der(17), potentially resulting in an aberrant VPS25 protein. CONCLUSION: The t(17;19)(q21;p13) translocation found in the two sisters probably predisposed them to myelodysplasia. How the MYO1F::WNK4 and/or VPS25::MYO1F chimeras, perhaps especially MYO1F::WNK4 that encodes a chimeric serine/threonine kinase, played a role in MDS pathogenesis, remains incompletely understood.
Assuntos
Síndromes Mielodisplásicas , Irmãos , Translocação Genética , Humanos , Síndromes Mielodisplásicas/genética , Síndromes Mielodisplásicas/patologia , Feminino , Cromossomos Humanos Par 17/genética , Cromossomos Humanos Par 19/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas de Transporte Vesicular/genética , Proteínas de Fusão Oncogênica/genética , Pessoa de Meia-IdadeRESUMO
BACKGROUND/AIM: Mixed phenotype acute leukemia (MPAL) is a rare hematologic malignancy in which the leukemic cells cannot be assigned to any specific lineage. The lack of well-defined, pathogenetically relevant diagnostic criteria makes the clinical handling of MPAL patients challenging. We herein report the genetic findings in bone marrow cells from two pediatric MPAL patients. PATIENTS AND METHODS: Bone marrow cells were examined using G-banding, array comparative genomic hybridization, RNA sequencing, reverse transcription polymerase chain reaction, Sanger sequencing, and fluorescence in situ hybridization. RESULTS: In the first patient, the genetic analyses revealed structural aberrations of chromosomal bands 8p11, 10p11, 11q21, and 17p11, the chimeras MLLT10::PICALM and PICALM::MLLT10, and imbalances (gains/losses) on chromosomes 2, 4, 8, 13, and 21. A submicroscopic deletion in 21q was also found including the RUNX1 locus. In the second patient, there were structural aberrations of chromosome bands 1p32, 8p11, 12p13, 20p13, and 20q11, the chimeras ETV6::LEXM and NCOA6::ETV6, and imbalances on chromosomes 2, 8, 11, 12, 16, 19, X, and Y. CONCLUSION: The leukemic cells from both MPAL patients carried chromosome aberrations resulting in fusion genes as well as genomic imbalances resulting in gain and losses of many gene loci. The detected fusion genes probably represent the main leukemogenic events, although the gains and losses are also likely to play a role in leukemogenesis.
Assuntos
Proteínas de Fusão Oncogênica , Leucemia-Linfoma Linfoblástico de Células Precursoras , Humanos , Criança , Hibridização in Situ Fluorescente , Hibridização Genômica Comparativa , Proteínas de Fusão Oncogênica/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Fenótipo , Translocação GenéticaRESUMO
Introduction: Intramuscular myxomas are benign tumors that are challenging to diagnose, especially on core needle biopsies. Acquired chromosomal aberrations and pathogenic variants in codon 201 or codon 227 in GNAS complex locus gene (GNAS) have been reported in these tumors. Here we present our genetic findings in a series of 22 intramuscular myxomas. Materials and methods: The tumors were investigated for the presence of acquired chromosomal aberrations using G-banding and karyotyping. Pathogenic variants in codon 201 or codon 227 of GNAS were assessed using direct cycle Sanger sequencing and Ion AmpliSeq Cancer Hotspot Panel v2 methodologies. Results: Eleven tumors carried chromosomal abnormalities. Six tumors had numerical, four had structural, and one had both numerical and structural chromosomal aberrations. Gains of chromosomes 7 and 8 were the most common abnormalities being found in five and four tumors respectively. Pathogenic variants in GNAS were detected in 19 myxomas (86%) with both methodologies. The detected pathogenic variants were p.R201H in nine cases (seven with abnormal and two with normal karyotypes), p.R201C in five cases, all with normal karyotypes, p.R201S in three cases (two with abnormal and one with normal karyotype), p.R201G in one case with a normal karyotype, and p.Q227E in one case with a normal karyotype. Conclusion: Firstly, our data indicate a possible association between chromosomal abnormalities and GNAS pathogenic variants in intramuscular myxomas. Secondly, the presence of the rare pathogenic variants R201S, p.R201G and p.Q227E in 26% (5 out of 19) of myxomas with GNAS pathogenic variants shows that methodologies designed to detect only the common "hotspot" of p.R201C and p.R201H will give false negative results. Finally, a comparison between Ion AmpliSeq Cancer Hotspot Panel v2 and direct cycle Sanger sequencing showed that direct cycle Sanger sequencing provides a quick, reliable, and relatively cheap method to detect GNAS pathogenic variants, matching even the most cutting-edge sequencing methods.