RESUMEN
The genes involved in the t(12;22)(q13;q12) translocation found recurrently in malignant melanoma of soft parts have been characterized and shown to form, in four cases studied, hybrid transcripts. The deduced chimaeric protein encoded by the der(22) chromosome consists of the N-terminal domain of EWS linked to the bZIP domain of ATF-1, a transcription factor which may normally be regulated by cAMP. ATF-1 has not previously been implicated in oncogenesis. EWS was first identified as forming a hybrid transcript in Ewing's sarcoma, which links its N-terminal domain to the DNA binding domain of the FLI-1 gene. Thus the oncogenic conversion of EWS follows a common scheme of activation, exchanging its putative RNA binding domain with different DNA binding domains that appear to be tumour-specific.
Asunto(s)
Cromosomas Humanos Par 12 , Cromosomas Humanos Par 22 , Clonación Molecular , Proteínas de Unión al ADN , Melanoma/genética , Proteínas de Fusión Oncogénica/genética , Oncogenes , Neoplasias de los Tejidos Blandos/genética , Factores de Transcripción/genética , Translocación Genética , Factor de Transcripción Activador 1 , Secuencia de Aminoácidos , Secuencia de Bases , Reordenamiento Génico , Humanos , Datos de Secuencia Molecular , Oligodesoxirribonucleótidos , Proteínas de Fusión Oncogénica/biosíntesis , Reacción en Cadena de la Polimerasa , Mapeo Restrictivo , Factores de Transcripción/biosíntesis , Transcripción Genética , Células Tumorales CultivadasRESUMEN
BACKGROUND: Using mRNA expression-derived signatures as predictors of individual patient outcome has been a goal ever since the introduction of microarrays. Here, we addressed whether analyses of tumour mRNA at the exon level can improve on the predictive power and classification accuracy of gene-based expression profiles using neuroblastoma as a model. METHODS: In a patient cohort comprising 113 primary neuroblastoma specimens expression profiling using exon-level analyses was performed to define predictive signatures using various machine-learning techniques. Alternative transcript use was calculated from relative exon expression. Validation of alternative transcripts was achieved using qPCR- and cell-based approaches. RESULTS: Both predictors derived from the gene or the exon levels resulted in prediction accuracies >80% for both event-free and overall survival and proved as independent prognostic markers in multivariate analyses. Alternative transcript use was most prominently linked to the amplification status of the MYCN oncogene, expression of the TrkA/NTRK1 neurotrophin receptor and survival. CONCLUSION: As exon level-based prediction yields comparable, but not significantly better, prediction accuracy than gene expression-based predictors, gene-based assays seem to be sufficiently precise for predicting outcome of neuroblastoma patients. However, exon-level analyses provide added knowledge by identifying alternative transcript use, which should deepen the understanding of neuroblastoma biology.
Asunto(s)
Exones/genética , Neuroblastoma/genética , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Receptor trkA/genética , Línea Celular Tumoral , Preescolar , Perfilación de la Expresión Génica , Humanos , Lactante , Proteína Proto-Oncogénica N-Myc , Neuroblastoma/mortalidad , Pronóstico , ARN Mensajero , Factores de Riesgo , Análisis de SupervivenciaRESUMEN
BACKGROUND: In the INRG dataset, the hypothesis that any segmental chromosomal alteration might be of prognostic impact in neuroblastoma without MYCN amplification (MNA) was tested. METHODS: The presence of any segmental chromosomal alteration (chromosome 1p deletion, 11q deletion and/or chromosome 17q gain) defined a segmental genomic profile. Only tumours with a confirmed unaltered status for all three chromosome arms were considered as having no segmental chromosomal alterations. RESULTS: Among the 8800 patients in the INRG database, a genomic type could be attributed for 505 patients without MNA: 397 cases had a segmental genomic type, whereas 108 cases had an absence of any segmental alteration. A segmental genomic type was more frequent in patients >18 months and in stage 4 disease (P<0.0001). In univariate analysis, 11q deletion, 17q gain and a segmental genomic type were associated with a poorer event-free survival (EFS) (P<0.0001, P=0.0002 and P<0.0001, respectively). In multivariate analysis modelling EFS, the parameters age, stage and a segmental genomic type were retained in the model, whereas the individual genetic markers were not (P<0.0001 and RR=2.56; P=0.0002 and RR=1.8; P=0.01 and RR=1.7, respectively). CONCLUSION: A segmental genomic profile, rather than the single genetic markers, adds prognostic information to the clinical markers age and stage in neuroblastoma patients without MNA, underlining the importance of pangenomic studies.
Asunto(s)
Neuroblastoma/genética , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Aberraciones Cromosómicas , Cromosomas Humanos Par 11/genética , Cromosomas Humanos Par 17/genética , Humanos , Lactante , Proteína Proto-Oncogénica N-Myc , Pronóstico , Estudios Retrospectivos , Análisis de SupervivenciaRESUMEN
T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of thymocytes and is largely driven by the NOTCH/MYC pathway. Yet, additional oncogenic drivers are required for transformation. Here, we identify protein tyrosine phosphatase type 4 A3 (PRL3) as a collaborating oncogenic driver in T-ALL. PRL3 is expressed in a large fraction of primary human T-ALLs and is commonly co-amplified with MYC. PRL3 also synergized with MYC to initiate early-onset ALL in transgenic zebrafish and was required for human T-ALL growth and maintenance. Mass-spectrometry phosphoproteomic analysis and mechanistic studies uncovered that PRL3 suppresses downstream T-cell phosphorylation signaling pathways, including those modulated by VAV1, and subsequently suppresses apoptosis in leukemia cells. Taken together, our studies have identified new roles for PRL3 as a collaborating oncogenic driver in human T-ALL and suggest that therapeutic targeting of the PRL3 phosphatase will likely be a useful treatment strategy for T-ALL.
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Biomarcadores de Tumor/metabolismo , Regulación Neoplásica de la Expresión Génica , Proteínas de Neoplasias/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patología , Proteínas Tirosina Fosfatasas/metabolismo , Linfocitos T/patología , Animales , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Femenino , Humanos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Proteínas de Neoplasias/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Pronóstico , Proteínas Tirosina Fosfatasas/genética , Linfocitos T/metabolismo , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto , Pez CebraRESUMEN
BACKGROUND: Recurrent 15q13.3 microdeletions were recently identified with identical proximal (BP4) and distal (BP5) breakpoints and associated with mild to moderate mental retardation and epilepsy. METHODS: To assess further the clinical implications of this novel 15q13.3 microdeletion syndrome, 18 new probands with a deletion were molecularly and clinically characterised. In addition, we evaluated the characteristics of a family with a more proximal deletion between BP3 and BP4. Finally, four patients with a duplication in the BP3-BP4-BP5 region were included in this study to ascertain the clinical significance of duplications in this region. RESULTS: The 15q13.3 microdeletion in our series was associated with a highly variable intra- and inter-familial phenotype. At least 11 of the 18 deletions identified were inherited. Moreover, 7 of 10 siblings from four different families also had this deletion: one had a mild developmental delay, four had only learning problems during childhood, but functioned well in daily life as adults, whereas the other two had no learning problems at all. In contrast to previous findings, seizures were not a common feature in our series (only 2 of 17 living probands). Three patients with deletions had cardiac defects and deletion of the KLF13 gene, located in the critical region, may contribute to these abnormalities. The limited data from the single family with the more proximal BP3-BP4 deletion suggest this deletion may have little clinical significance. Patients with duplications of the BP3-BP4-BP5 region did not share a recognisable phenotype, but psychiatric disease was noted in 2 of 4 patients. CONCLUSIONS: Overall, our findings broaden the phenotypic spectrum associated with 15q13.3 deletions and suggest that, in some individuals, deletion of 15q13.3 is not sufficient to cause disease. The existence of microdeletion syndromes, associated with an unpredictable and variable phenotypic outcome, will pose the clinician with diagnostic difficulties and challenge the commonly used paradigm in the diagnostic setting that aberrations inherited from a phenotypically normal parent are usually without clinical consequences.
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Aberraciones Cromosómicas , Deleción Cromosómica , Trastornos de los Cromosomas/genética , Cromosomas Humanos Par 15/genética , Duplicación de Gen , Adolescente , Adulto , Niño , Preescolar , Trastornos de los Cromosomas/patología , Femenino , Humanos , Lactante , Recién Nacido , Discapacidad Intelectual/genética , Discapacidad Intelectual/patología , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Linaje , Embarazo , SíndromeRESUMEN
Neuroblastoma serves as a paradigm for utilising tumour genomic data for determining patient prognosis and treatment allocation. However, before the establishment of the International Neuroblastoma Risk Group (INRG) Task Force in 2004, international consensus on markers, methodology, and data interpretation did not exist, compromising the reliability of decisive genetic markers and inhibiting translational research efforts. The objectives of the INRG Biology Committee were to identify highly prognostic genetic aberrations to be included in the new INRG risk classification schema and to develop precise definitions, decisive biomarkers, and technique standardisation. The review of the INRG database (n=8800 patients) by the INRG Task Force finally enabled the identification of the most significant neuroblastoma biomarkers. In addition, the Biology Committee compared the standard operating procedures of different cooperative groups to arrive at international consensus for methodology, nomenclature, and future directions. Consensus was reached to include MYCN status, 11q23 allelic status, and ploidy in the INRG classification system on the basis of an evidence-based review of the INRG database. Standardised operating procedures for analysing these genetic factors were adopted, and criteria for proper nomenclature were developed. Neuroblastoma treatment planning is highly dependant on tumour cell genomic features, and it is likely that a comprehensive panel of DNA-based biomarkers will be used in future risk assignment algorithms applying genome-wide techniques. Consensus on methodology and interpretation is essential for uniform INRG classification and will greatly facilitate international and cooperative clinical and translational research studies.
Asunto(s)
Neuroblastoma/diagnóstico , Neuroblastoma/genética , Aberraciones Cromosómicas , Cromosomas Humanos Par 11 , Cromosomas Humanos Par 17 , Consenso , Amplificación de Genes , Marcadores Genéticos , Humanos , Cooperación Internacional , Proteína Proto-Oncogénica N-Myc , Neuroblastoma/epidemiología , Neuroblastoma/psicología , Neuroblastoma/terapia , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Planificación de Atención al Paciente , Ploidias , Pronóstico , Biosíntesis de Proteínas , Medición de Riesgo , Factores de Riesgo , Análisis de SupervivenciaRESUMEN
BACKGROUND: Recent molecular studies of breakpoints of recurrent chromosome rearrangements revealed the role of genomic architecture in their formation. In particular, segmental duplications representing blocks of >1 kb with >90% sequence homology were shown to mediate non-allelic homologous recombination (NAHR). However, the occurrence of the majority of newly detected submicroscopic imbalances cannot be explained by the presence of segmental duplications. Therefore, further studies are needed to investigate whether architectural features other than segmental duplications mediate these rearrangements. METHODS: We analysed a series of patients with breakpoints clustering within chromosome band 5q35. Using high density arrays and subsequent quantitative polymerase chain reaction (qPCR), we characterised the breakpoints of four interstitial deletions (including one associated with an unbalanced paracentric inversion), a duplication and a familial reciprocal t(5;18)(q35;q22) translocation. RESULTS AND CONCLUSION: Five of the breakpoints were located within an interval of approximately 265 kb encompassing the RANBP17 and TLX3 genes. This region is also targeted by the recurrent cryptic t(5;14)(q35;q32) translocation, which occurs in approximately 20% of childhood T cell acute lymphoblastic leukaemia (T-ALL). In silico analysis indicated the architectural features most likely to contribute to the genomic instability of this region, which was supported by our molecular data. Of further interest, in two patients and the familial translocation, the delineated breakpoint regions encompassed highly homologous LINEs (long interspersed nuclear elements), suggesting that NAHR between these LINEs may have mediated these rearrangements.
Asunto(s)
Rotura Cromosómica , Cromosomas Humanos Par 5 , Inestabilidad Genómica , Mapeo Cromosómico , Eliminación de Gen , Duplicación de Gen , Humanos , Translocación GenéticaRESUMEN
Recent studies have unveiled copy number variants (CNVs) as an important source of genetic variation. Many of these CNVs contain coding sequences, which have been shown to be dosage sensitive. Evidence is accumulating that certain CNVs have impact on susceptibility to human diseases such as HIV infection and autoimmune diseases, as well as on adaptability to environmental conditions or nutrition. The possible role and impact of CNVs on cancer development and progression is only now emerging. In this review we look into the role of CNVs and their associated genomic structural features in relation to the formation of chromosome alterations in cancer cells and evolutionary genomic plasticity, as well as the de novo occurrence of known or putative CNVs as somatic events during oncogenesis. The role of germline CNVs in cancer predisposition is still largely unexplored. A number of observations seem to warrant the importance of further studies to elucidate the impact of these variants in the early steps of carcinogenesis.
Asunto(s)
Dosificación de Gen/genética , Neoplasias/genética , Animales , Evolución Molecular , Duplicación de Gen , Predisposición Genética a la Enfermedad/genética , Genoma/genética , HumanosRESUMEN
Recently, we and others described a new chromosomal rearrangement, that is, inv(7)(p15q34) and t(7;7)(p15;q34) involving the T-cell receptor beta (TCRbeta) (7q34) and the HOXA gene locus (7p15) in 5% of T-cell acute lymphoblastic leukemia (T-ALL) patients leading to transcriptional activation of especially HOXA10. To further address the clinical, immunophenotypical and molecular genetic findings of this chromosomal aberration, we studied 330 additional T-ALLs. This revealed TCRbeta-HOXA rearrangements in five additional patients, which brings the total to 14 cases in 424 patients (3.3%). Real-time quantitative PCR analysis for HOXA10 gene expression was performed in 170 T-ALL patients and detected HOXA10 overexpression in 25.2% of cases including all the cases with a TCRbeta-HOXA rearrangement (8.2%). In contrast, expression of the short HOXA10 transcript, HOXA10b, was almost exclusively found in the TCRbeta-HOXA rearranged cases, suggesting a specific role for the HOXA10b short transcript in TCRbeta-HOXA-mediated oncogenesis. Other molecular and/or cytogenetic aberrations frequently found in subtypes of T-ALL (SIL-TAL1, CALM-AF10, HOX11, HOX11L2) were not detected in the TCRbeta-HOXA rearranged cases except for deletion 9p21 and NOTCH1 activating mutations, which were present in 64 and 67%, respectively. In conclusion, this study defines TCRbeta-HOXA rearranged T-ALLs as a distinct cytogenetic subgroup by clinical, immunophenotypical and molecular genetic characteristics.
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Proteínas de Homeodominio/genética , Leucemia-Linfoma de Células T del Adulto/genética , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Adolescente , Adulto , Niño , Deleción Cromosómica , Inversión Cromosómica , Femenino , Reordenamiento Génico de Linfocito T , Proteínas Homeobox A10 , Humanos , Inmunofenotipificación , Leucemia-Linfoma de Células T del Adulto/patología , Leucemia-Linfoma de Células T del Adulto/fisiopatología , Masculino , Persona de Mediana Edad , Receptor Notch1/genética , Activación Transcripcional , Translocación GenéticaRESUMEN
Chromosomal aberrations of T-cell receptor (TCR) gene loci often involve the TCRalphadelta (14q11) locus and affect various known T-cell oncogenes. A systematic fluorescent in situ hybridization (FISH) screening for the detection of chromosomal aberrations involving the TCR loci, TCRalphadelta (14q11), TCRbeta (7q34) and TCRgamma (7p14), has not been conducted so far. Therefore, we initiated a screening of 126 T-cell acute lymphoblastic leukemia (T-ALL) and T-cell lymphoblastic lymphoma cases and 19 T-ALL cell lines using FISH break-apart assays for the different TCR loci. Genomic rearrangements of the TCRbeta locus were detected in 24/126 cases (19%), most of which (58.3%) were not detected upon banding analysis. Breakpoints in the TCRalphadelta locus were detected in 22/126 cases (17.4%), whereas standard cytogenetics only detected 14 of these 22 cases. Cryptic TCRalphadelta/TCRbeta chromosome aberrations were thus observed in 22 of 126 cases (17.4%). Some of these chromosome aberrations target new putative T-cell oncogenes at chromosome 11q24, 20p12 and 6q22. Five patients and one cell line carried chromosomal rearrangements affecting both TCRbeta and TCRalphadelta loci. In conclusion, this study presents the first inventory of chromosomal rearrangements of TCR loci in T-ALL, revealing an unexpected high number of cryptic chromosomal rearrangements of the TCRbeta locus and further broadening the spectrum of genes putatively implicated in T-cell oncogenesis.
Asunto(s)
Reordenamiento Génico de Linfocito T/genética , Genes Codificadores de la Cadena beta de los Receptores de Linfocito T/genética , Leucemia-Linfoma de Células T del Adulto/epidemiología , Leucemia-Linfoma de Células T del Adulto/genética , Adolescente , Adulto , Niño , Preescolar , Femenino , Genes Codificadores de la Cadena alfa de los Receptores de Linfocito T/genética , Genes Codificadores de la Cadena delta de los Receptores de Linfocito T/genética , Humanos , Hibridación Fluorescente in Situ , Incidencia , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Translocación GenéticaRESUMEN
The assignment with chromosome banding techniques of the breakpoints of the recurrent translocation t(3;5) which leads to NPM1/MLF1 gene fusion in myeloid malignancies has not been unequivocal. In order to assess whether this is due to uncertainty in interpretation of the observed banding pattern or whether it reflects true genomic heterogeneity, we decided to analyze the breakpoint positions using fluorescence in situ (FISH) techniques in eight patients with myeloid malignancies and rearrangements of chromosomes 3 and 5. In three patients, colocalization of the NPM1 and MLF1 spanning BACs was demonstrated and NPM1/MLF1 fusion shown by PCR in one while in the remaining cases breakpoints were located outside the NPM1 and MLF1 loci. Interestingly, loss of a copy of the NPM1 gene was found in three of these latter patients. This findings suggest that haploinsufficiency of NPM1 may play a role in subtypes of myelodysplasias and leukemias.
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Cromosomas Humanos Par 5/genética , Leucemia Mieloide/genética , Síndromes Mielodisplásicos/genética , Proteínas Nucleares/genética , Translocación Genética , Adulto , Anciano , Niño , Preescolar , Bandeo Cromosómico , Cromosomas Humanos Par 3/genética , Femenino , Hematopoyesis/genética , Humanos , Hibridación Fluorescente in Situ/métodos , Masculino , Nucleofosmina , Proteínas de Fusión Oncogénica/genética , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
BACKGROUND: Chromosomal abnormalities are a major cause of mental retardation and multiple congenital anomalies (MCA/MR). Screening for these chromosomal imbalances has mainly been done by standard karyotyping. Previous array CGH studies on selected patients with chromosomal phenotypes and normal karyotypes suggested an incidence of 10-15% of previously unnoticed de novo chromosomal imbalances. OBJECTIVE: To report array CGH screening of a series of 140 patients (the largest published so far) with idiopathic MCA/MR but normal karyotype. RESULTS: Submicroscopic chromosomal imbalances were detected in 28 of the 140 patients (20%) and included 18 deletions, seven duplications, and three unbalanced translocations. Seventeen of 24 imbalances were confirmed de novo and 19 were assumed to be causal. Excluding subtelomeric imbalances, our study identified 11 clinically relevant interstitial submicroscopic imbalances (8%). Taking this and previously reported studies into consideration, array CGH screening with a resolution of at least 1 Mb has been undertaken on 432 patients with MCA/MR. Most imbalances are non-recurrent and spread across the genome. In at least 8.8% of these patients (38 of 432) de novo intrachromosomal alterations have been identified. CONCLUSIONS: Array CGH should be considered an essential aspect of the genetic analysis of patients with MCA/MR. In addition, in the present study three patients were mosaic for a structural chromosome rearrangement. One of these patients had monosomy 7 in as few as 8% of the cells, showing that array CGH allows detection of low grade mosaicisims.
Asunto(s)
Anomalías Múltiples/genética , Aberraciones Cromosómicas , Discapacidad Intelectual/genética , Adolescente , Adulto , Niño , Preescolar , Cromosomas Humanos Par 7/genética , Femenino , Dosificación de Gen/genética , Genoma Humano/genética , Humanos , Lactante , Masculino , Persona de Mediana Edad , Hibridación de Ácido NucleicoRESUMEN
In leukemias chromosomal aberrations, balanced translocations in particular, play a critical role in the oncogenic process. The characterization of these chromosomal alterations was crucial to the discovery of the genes implicated in leukemogenesis, as the chromosomal breakpoints indicated their genomic localization. In addition, these molecular defects may serve as targets for diagnostic essays and can have a major prognostic value. Finally, the characterization of the deregulated cellular pathways potentially identifies targets for therapeutic intervention. In this paper we summarize our efforts to expand the current knowledge of the diagnostic, prognostic or biological significance of selected chromosomal aberrations identified in M-FISH studies. First, we illustrated the power of M-FISH in dissecting complex chromosomal aberrations in myeloid neoplasms. MLL amplification was defined as a clinical entity characterized by adverse prognosis and within the multitude and variety of chromosomal rearrangements a pattern of a limited number of cytogenetic subclasses was discerned. In leukemias characterized by 11q23 amplification, we described the amplicon and confirmed MLL, in addition to DDX6, as a principal amplification target. Molecular characterization of a large series of unselected sporadic and recurrent 3q26 rearranged leukemias confirmed the decisive role of ectopic EVI1 expression in these malignancies. We contributed to an extensive analysis of the phenotypical and prognostic features of T-ALL characterized by HOX11L2 expression and identified HOX11L2 overexpression as one of the most frequent genetic defects in childhood T-ALL, associated with intermediate prognosis. Finally, we designed and validated diagnostic tools for the detection of the t(9;14) (p13;q34) resulting in PAX5 overexpression and convincingly associated the presence of this rearrangement to high-grade morphology and karyotype complexity. In conclusion, the series of investigations presented here clearly illustrate the benefits of M-FISH as molecular tool for the dissection and characterization of complex and cryptic rearrangements. The subsequent reports demonstrate the utility of molecular cytogenetics and expression analyses to the clinical management of patients diagnosed with hematological malignancies.
Asunto(s)
Aberraciones Cromosómicas , Leucemia Mieloide/clasificación , Leucemia Mieloide/genética , Translocación Genética , Aberraciones Cromosómicas/clasificación , Rotura Cromosómica , Análisis Citogenético , Marcadores Genéticos , Humanos , Hibridación Fluorescente in Situ/métodos , Leucemia Mieloide/diagnóstico , PronósticoRESUMEN
Neuroblastoma (NB) is, next to acute lymphoblastic leukaemia, brain tumours and lymphoma the most frequent paediatric tumour (8-10%). Our research group aims to contribute to the unravelling of the genetic basis of NB. Insight into the genes and signalling pathways involved in tumour formation and development can represent an essential step towards the development of more efficient molecular targeted therapies. A first part of our research work was devoted to the analysis of genomic alterations in NB. By means of a new highly sensitive method for detecting gains and losses of chromosomal segments, we recognised three major prognostic relevant genomic subtypes of NB. In addition smaller subgroups with deviating genomic patterns were detected. In addition, this work yielded important information regarding delineation of critical regions of gain and loss in NB which should facilitate further selection of candidate oncogenes or tumour suppressor genes. A second important part of our work focussed on the gene expression profiling of NB precursor cells. We were able as the first to isolate these cells and determine their transcriptome, a finding of fundamental importance for future expression studies in NB. Another study focussed on the identification of MYCN transcriptional target genes. Gene expression analyses of model systems developed in our lab and of a large panel of cell lines and tumours allowed us to subtract a list of candidate genes which are now under further study. Finally, we initiated research towards the understanding of the role of methylation in NB oncogenesis. From this, we were able to create a list of potentially relevant methylated genes in NB. From the above it is clear that our team has made important contributions to the understanding of the complex biology and clinical behaviour of NB. Also, a broad technically innovative research platform has been developed which will allow us to dissect NB genetics with greater speed and accuracy.
Asunto(s)
Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Neuroblastoma/genética , Genes Relacionados con las Neoplasias , Humanos , Neuroblastoma/patologíaRESUMEN
In the past few years high throughput methods for assessment of DNA copy number alterations have witnessed rapid progress. Both 'in house' developed BAC, cDNA, oligonucleotide and commercial arrays are now available and widely applied in the study of the human genome, particularly in the context of disease. Cancer cells are known to exhibit DNA losses, gains and amplifications affecting tumor suppressor genes and proto-oncogenes. Moreover, these patterns of genomic imbalances may be associated with particular tumor types or subtypes and may have prognostic value. Here we summarize recent array CGH findings in neuroblastoma, a pediatric tumor of the sympathetic nervous system. A total of 176 primary tumors and 53 cell lines have been analyzed on different platforms. Through these studies the genomic content and boundaries of deletions, gains and amplifications were characterized with unprecedented accuracy. Furthermore, in conjunction with cytogenetic findings, array CGH allows the mapping of breakpoints of unbalanced translocations at a very high resolution.
Asunto(s)
Biología Computacional/métodos , ADN/análisis , Genoma Humano , Neuroblastoma/genética , Línea Celular Tumoral , ADN de Neoplasias , Humanos , Hibridación Fluorescente in Situ , Hibridación de Ácido Nucleico , Análisis de Secuencia por Matrices de Oligonucleótidos , Oligonucleótidos/química , Oligonucleótidos/genética , Translocación GenéticaRESUMEN
Chromosomal translocations with breakpoints in T-cell receptor (TCR) genes are recurrent in T-cell malignancies. These translocations involve the TCRalphadelta gene (14q11), the TCRbeta gene (7q34) and to a lesser extent the TCRgamma gene at chromosomal band 7p14 and juxtapose T-cell oncogenes next to TCR regulatory sequences leading to deregulated expression of those oncogenes. Here, we describe a new recurrent chromosomal inversion of chromosome 7, inv(7)(p15q34), in a subset of patients with T-cell acute lymphoblastic leukemia characterized by CD2 negative and CD4 positive, CD8 negative blasts. This rearrangement juxtaposes the distal part of the HOXA gene cluster on 7p15 to the TCRbeta locus on 7q34. Real time quantitative PCR analysis for all HOXA genes revealed high levels of HOXA10 and HOXA11 expression in all inv(7) positive cases. This is the first report of a recurrent chromosome rearrangement targeting the HOXA gene cluster in T-cell malignancies resulting in deregulated HOXA gene expression (particularly HOXA10 and HOXA11) and is in keeping with a previous report suggesting HOXA deregulation in MLL-rearranged T- and B cell lymphoblastic leukemia as the key factor in leukaemic transformation. Finally, our observation also supports the previous suggested role of HOXA10 and HOXA11 in normal thymocyte development.
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Inversión Cromosómica , Cromosomas Humanos Par 7/genética , Proteínas de Unión al ADN/genética , Proteínas de Homeodominio/genética , Leucemia-Linfoma de Células T del Adulto/genética , Activación Transcripcional/genética , Adolescente , Adulto , Niño , Preescolar , Análisis Citogenético , Proteínas de Unión al ADN/fisiología , Femenino , Regulación Neoplásica de la Expresión Génica , Reordenamiento Génico de Linfocito T/genética , Proteínas Homeobox A10 , Proteínas de Homeodominio/fisiología , Humanos , Inmunofenotipificación , Masculino , Persona de Mediana Edad , Translocación Genética/genéticaRESUMEN
Merkel cell carcinoma or small cell carcinoma of the skin is a rare skin cancer seen in increasing numbers in Queensland, Australia. In its clinical course and histopathology, it resembles small cell lung carcinoma (SCLC). Little is known of the genetic basis of this disease except for a number of cytogenetic studies and three loss of heterozygosity studies. Therefore, comparative genomic hybridization was performed to determine the characteristic DNA gains and losses that occur in this tumor. Comparative genomic hybridization analysis of 34 specimens from 24 patients revealed a pattern of gains and losses that closely resembles that seen in SCLC. Overall frequent loss was seen for chromosomes 3p (46%), 5q (21%), 8p (21%), 10 (33%), 11q (17%), 13q (33%), and 17p (25%). Significant gains were seen for chromosomes 1 (63%), 3q (33%), 5p (38%), 8q (38%), 19 (63%), and X (41%), with smaller numbers having gains for chromosomes 6, 7, 20, and 21. In contrast to SCLC, amplification in Merkel cell carcinoma is a rare event.
Asunto(s)
Carcinoma de Células de Merkel/genética , Aberraciones Cromosómicas , Neoplasias Cutáneas/genética , Deleción Cromosómica , ADN de Neoplasias/genética , Femenino , Amplificación de Genes , Humanos , Masculino , Hibridación de Ácido Nucleico , Pronóstico , Células Tumorales CultivadasRESUMEN
Esophageal cancer (EC) is the leading cause of cancer death in the Black male population in South Africa. Although several oncogenes and tumor suppressor genes have previously been found altered in this cancer, many novel genes remain to be identified. To identify the chromosomal location of these unknown genes, we have analyzed DNA of 29 South African EC patients by comparative genomic hybridization. Frequent loss occurred at chromosome 1p (52%), 4p (52%), 18q (48%), 19p (52%), 19q (55%), and 22q (41%). The most common gains were detected at 1q (41%), 2q (52%), 3q (72%), 5p (31%), 7p (48%), 7q (45%), 8q (55%), and Xq (69%). High level amplification was detected at 2q24-33, 6p21.1-q14, 7p12-q21, 7q11.2-31, 8q22-24, 8q13-qter, 13q21-34, and at 13q32-34. The present comparative genomic hybridization study opens the way for additional targeted studies on these particular chromosomal regions to identify the specific genes involved in the higher susceptibility to specific subtypes of esophageal carcinoma in different geographical regions. The loss of 8p (28%) and Xp (17%) in tumors of male individuals may provide clues to the basis of the sex-biased frequency of occurrence of EC favoring men.
Asunto(s)
ADN de Neoplasias/análisis , Neoplasias Esofágicas/genética , Población Negra/genética , Deleción Cromosómica , Mapeo Cromosómico , Neoplasias Esofágicas/etnología , Femenino , Dosificación de Gen , Humanos , Masculino , Hibridación de Ácido Nucleico , SudáfricaRESUMEN
Gene expression analysis is increasingly important in many fields of biological research. Understanding patterns of expressed genes is assumed to provide insight into complex regulatory networks and can lead to the identification of genes relevant to specific biological processes, including disease. Among different techniques, reverse transcription quantitative polymerase chain reaction (RT-qPCR) is currently regarded as the gold standard for targeted quantification of RNA gene expression, especially because of its high sensitivity, specificity, accuracy, and precision, and also because of its practical simplicity and processing speed. However, different critical factors can influence the outcome of RT-qPCR studies, including isolation of RNA, reverse transcription to cDNA, and data analysis. These factors need to be addressed in order to obtain biologically meaningful results. In this chapter, we describe how RT-qPCR can be used in a reliable way to successfully study differential gene expression in zebrafish. Hereby, we especially focus on how expressed repetitive elements can be employed as reference targets in zebrafish RT-qPCR studies and how they can further improve the quality of the data.
Asunto(s)
Perfilación de la Expresión Génica/métodos , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Secuencias Repetitivas de Ácidos Nucleicos/genética , Pez Cebra/genética , Animales , ADN Complementario/genética , Regulación de la Expresión Génica/genética , ARN/biosíntesis , ARN/genéticaRESUMEN
Neuroblastoma is characterized by deletions of the short arm of chromosome 1 (1p) and amplification of the N-myc oncogene. We have made somatic cell hybrids of two human neuroblastoma cell lines, one with and one without N-myc expression and amplification. The expression of the amplified N-myc gene is completely switched off in the hybrids. This suggests that N-myc expression results from loss of a repressor function. As N-myc amplification is associated with loss of heterozygosity (LOH) of 1p36, we analysed 1p deletions in 16 neuroblastoma cell lines. The seven cell lines without N-myc amplification have no deletions or relatively small deletions, with an SRO on 1p36.23-33. This suggests that a tumor suppressor gene maps in this region. All nine cell lines with N-myc amplification have larger deletions, with an SRO from 1p35-36.1 to the telomere. This suggests that a second tumor suppressor gene which is associated with N-myc amplification maps more proximally. Fine mapping of 1p36 deletions in the two cell lines of the fusion experiment suggests that the distal locus is not a repressor of N-myc expression, but the more proximal locus could be a candidate for this function.