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1.
Cell Death Differ ; 30(1): 37-53, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-35869285

RESUMO

Despite being frequently observed in cancer cells, chromosomal instability (CIN) and its immediate consequence, aneuploidy, trigger adverse effects on cellular homeostasis that need to be overcome by anti-stress mechanisms. As such, these safeguard responses represent a tumor-specific Achilles heel, since CIN and aneuploidy are rarely observed in normal cells. Recent data have revealed that epitranscriptomic marks catalyzed by RNA-modifying enzymes change under various stress insults. However, whether aneuploidy is associated with such RNA modifying pathways remains to be determined. Through an in silico search for aneuploidy biomarkers in cancer cells, we found TRMT61B, a mitochondrial RNA methyltransferase enzyme, to be associated with high levels of aneuploidy. Accordingly, TRMT61B protein levels are increased in tumor cell lines with an imbalanced karyotype as well as in different tumor types when compared to control tissues. Interestingly, while TRMT61B depletion induces senescence in melanoma cell lines with low levels of aneuploidy, it leads to apoptosis in cells with high levels. The therapeutic potential of these results was further validated by targeting TRMT61B in transwell and xenografts assays. We show that TRM61B depletion reduces the expression of several mitochondrial encoded proteins and limits mitochondrial function. Taken together, these results identify a new biomarker of aneuploidy in cancer cells that could potentially be used to selectively target highly aneuploid tumors.


Assuntos
Metiltransferases , Neoplasias , Humanos , RNA Mitocondrial , Metiltransferases/genética , Aneuploidia , Instabilidade Cromossômica , RNA , Biomarcadores , Neoplasias/tratamento farmacológico , Neoplasias/genética
2.
Cancer Res ; 81(17): 4529-4544, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34145034

RESUMO

Cancer stem cells (CSC) are considered responsible for tumor initiation, therapeutic resistance, and metastasis. A comprehensive knowledge of the mechanisms governing the acquisition and maintenance of cancer stemness is crucial for the development of new therapeutic approaches in oncology. E2A basic helix-loop-helix (bHLH) transcription factors are associated with epithelial-mesenchymal transition (EMT) and tumor progression, but knowledge of their functional contributions to cancer biology is still limited. Using a combination of in vivo and in vitro analyses in a novel PyMT-E2A conditional knockout mouse model and derived primary tumor cell lines, we report here an essential role of E2A in stemness, metastasis, and therapeutic resistance in breast cancer. Targeted deletion of E2A in the mammary gland impaired tumor-initiating ability and dedifferentiation potential and severely compromised metastatic competence of PyMT-driven mammary tumors. Mechanistic studies in PyMT-derived cell lines indicated that E2A actions are mediated by the upregulation of Snai1 transcription. Importantly, high E2A and SNAIL1 expression occurred in aggressive human basal-like breast carcinomas, highlighting the relevance of the E2A-Snail1 axis in metastatic breast cancer. In addition, E2A factors contributed to the maintenance of genomic integrity and resistance to PARP inhibitors in PyMT and human triple-negative breast cancer cells. Collectively, these results support the potential for E2A transcription factors as novel targets worthy of translational consideration in breast cancer. SIGNIFICANCE: These findings identify key functions of E2A factors in breast cancer cell stemness, metastasis, and drug resistance, supporting a therapeutic vulnerability to targeting E2A proteins in breast cancer.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Neoplasias da Mama/metabolismo , Resistencia a Medicamentos Antineoplásicos , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Neoplasias da Mama/genética , Carcinogênese , Diferenciação Celular , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Simulação por Computador , Transição Epitelial-Mesenquimal , Feminino , Deleção de Genes , Genoma , Genótipo , Humanos , Masculino , Neoplasias Mamárias Animais , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Metástase Neoplásica , Células-Tronco Neoplásicas , Ftalazinas/farmacologia , Piperazinas/farmacologia , Fatores de Transcrição da Família Snail/metabolismo , Transgenes , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo
4.
Oncotarget ; 7(35): 56295-56308, 2016 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-27462779

RESUMO

Around, 30-40% of HER2-positive breast cancers do not show substantial clinical benefit from the targeted therapy and, thus, the mechanisms underlying resistance remain partially unknown. Interestingly, ERBB2 is frequently co-amplified and co-expressed with neighbour genes that may play a relevant role in this cancer subtype. Here, using an in silico analysis of data from 2,096 breast tumours, we reveal a significant correlation between Gasdermin B (GSDMB) gene (located 175 kilo bases distal from ERBB2) expression and the pathological and clinical parameters of poor prognosis in HER2-positive breast cancer. Next, the analysis of three independent cohorts (totalizing 286 tumours) showed that approximately 65% of the HER2-positive cases have GSDMB gene amplification and protein over-expression. Moreover, GSDMB expression was also linked to poor therapeutic responses in terms of lower relapse free survival and pathologic complete response as well as positive lymph node status and the development of distant metastasis under neoadjuvant and adjuvant treatment settings, respectively. Importantly, GSDMB expression promotes survival to trastuzumab in different HER2-positive breast carcinoma cells, and is associated with trastuzumab resistance phenotype in vivo in Patient Derived Xenografts. In summary, our data identifies the ERBB2 co-amplified and co-expressed gene GSDMB as a critical determinant of poor prognosis and therapeutic response in HER2-positive breast cancer.


Assuntos
Neoplasias da Mama/genética , Carcinoma Ductal de Mama/genética , Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/genética , Recidiva Local de Neoplasia/genética , Receptor ErbB-2/antagonistas & inibidores , Adulto , Idoso , Animais , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Neoplasias da Mama/terapia , Carcinoma Ductal de Mama/mortalidade , Carcinoma Ductal de Mama/patologia , Carcinoma Ductal de Mama/terapia , Quimioterapia Adjuvante , Intervalo Livre de Doença , Resistencia a Medicamentos Antineoplásicos , Feminino , Seguimentos , Amplificação de Genes , Humanos , Metástase Linfática , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Terapia Neoadjuvante , Proteínas de Neoplasias/metabolismo , Recidiva Local de Neoplasia/mortalidade , Recidiva Local de Neoplasia/patologia , Recidiva Local de Neoplasia/terapia , Prognóstico , Receptor ErbB-2/metabolismo , Trastuzumab/farmacologia , Trastuzumab/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
5.
BMC Cancer ; 15: 940, 2015 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-26620706

RESUMO

BACKGROUND: High grade serous ovarian cancer is characterised by high initial response to chemotherapy but poor outcome in the long term due to acquired resistance. One of the main genetic features of this disease is TP53 mutation. The majority of TP53 mutated tumors harbor missense mutations in this gene, correlated with p53 accumulation. TP53 null tumors constitute a specific subgroup characterised by nonsense, frameshift or splice-site mutations associated to complete absence of p53 expression. Different studies show that this kind of tumors may have a worse prognosis than other TP53 mutated HGSC. METHODS: In this study, we sought to characterise the intra-tumor heterogeneity of a TP53 null HGSC consisting of six primary tumor samples, two intra-pelvic and four extra-pelvic recurrences using exome sequencing and comparative genome hybridisation. RESULTS: Significant heterogeneity was found among the different tumor samples, both at the mutational and copy number levels. Exome sequencing identified 102 variants, of which only 42 were common to all three samples; whereas 7 of the 18 copy number changes found by CGH analysis were presented in all samples. Sanger validation of 20 variants found by exome sequencing in additional regions of the primary tumor and the recurrence allowed us to establish a sequence of the tumor clonal evolution, identifying those populations that most likely gave rise to recurrences and genes potentially involved in this process, like GPNMB and TFDP1. Using functional annotation and network analysis, we identified those biological functions most significantly altered in this tumor. Remarkably, unexpected functions such as microtubule-based movement and lipid metabolism emerged as important for tumor development and progression, suggesting its potential interest as therapeutic targets. CONCLUSIONS: Altogether, our results shed light on the clonal evolution of the distinct tumor regions identifying the most aggressive subpopulations and at least some of the genes that may be implicated in its progression and recurrence, and highlights the importance of considering intra-tumor heterogeneity when carrying out genetic and genomic studies, especially when these are aimed to diagnostic procedures or to uncover possible therapeutic strategies.


Assuntos
Cistadenocarcinoma Seroso/patologia , Heterogeneidade Genética , Neoplasias Ovarianas/patologia , Proteína Supressora de Tumor p53/genética , Evolução Clonal , Hibridização Genômica Comparativa , Cistadenocarcinoma Seroso/genética , Feminino , Variação Genética , Humanos , Glicoproteínas de Membrana/genética , Pessoa de Meia-Idade , Mutação , Gradação de Tumores , Neoplasias Ovarianas/genética , Prognóstico , Análise de Sequência de DNA , Fator de Transcrição DP1/genética
6.
Mod Pathol ; 28(11): 1492-503, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26381823

RESUMO

Although TAZ, the final effector of the Hippo pathway that modulates epithelial to mesenchymal transition and stemness, has been implicated in the development of different types of cancer, its role in endometrial cancer has not yet been studied. Thus, we evaluated the expression of TAZ in different types of endometrial cancer by immunohistochemistry. TAZ expression was detected in 76% of undifferentiated endometrial carcinomas, 54% of endometrial carcinosarcomas, 46% of endometrial serous carcinomas, 36% of grade 3 endometrioid carcinomas, and 18% of grade 1-2 endometrioid carcinomas, with statistically significant differences. We analyzed the WWTR1 gene that encodes TAZ by FISH and MassARRAY spectrometry, ruling out gene amplification and differential promoter methylation as the main mechanisms that modulate TAZ expression in endometrial tumors. However, we did detect a significant association between Scribble hypoexpression and delocalization with TAZ expression. Moreover, we demonstrated that TAZ promoted invasiveness, and it favored cell motility and tumor growth, in endometrial cancer cell lines. In addition, TAZ expression was associated with the transition from an epithelial to mesenchymal phenotype, both in vitro and in human tumors. Together, these data reveal a previously unknown role for TAZ and the Hippo pathway in the progression of aggressive subtypes of endometrial cancer.


Assuntos
Carcinoma/patologia , Neoplasias do Endométrio/patologia , Transição Epitelial-Mesenquimal/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Animais , Western Blotting , Carcinoma/metabolismo , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Neoplasias do Endométrio/metabolismo , Feminino , Imunofluorescência , Xenoenxertos , Via de Sinalização Hippo , Humanos , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Espectrometria de Massas , Camundongos , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase , Proteínas Serina-Treonina Quinases/metabolismo , Transativadores , Fatores de Transcrição , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional
7.
Mol Cell Biol ; 35(20): 3566-78, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26240282

RESUMO

Aurora kinase B, one of the three members of the mammalian Aurora kinase family, is the catalytic component of the chromosomal passenger complex, an essential regulator of chromosome segregation in mitosis. Aurora B is overexpressed in human tumors although whether this kinase may function as an oncogene in vivo is not established. Here, we report a new mouse model in which expression of the endogenous Aurkb locus can be induced in vitro and in vivo. Overexpression of Aurora B in cultured cells induces defective chromosome segregation and aneuploidy. Long-term overexpression of Aurora B in vivo results in aneuploidy and the development of multiple spontaneous tumors in adult mice, including a high incidence of lymphomas. Overexpression of Aurora B also results in a reduced DNA damage response and decreased levels of the p53 target p21(Cip1) in vitro and in vivo, in line with an inverse correlation between Aurora B and p21(Cip1) expression in human leukemias. Thus, overexpression of Aurora B may contribute to tumor formation not only by inducing chromosomal instability but also by suppressing the function of the cell cycle inhibitor p21(Cip1).


Assuntos
Aneuploidia , Aurora Quinase B/fisiologia , Inibidor de Quinase Dependente de Ciclina p21/genética , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Fibroblastos/metabolismo , Expressão Gênica , Inativação Gênica , Camundongos Endogâmicos C57BL , Leucemia-Linfoma Linfoblástico de Células Precursoras/enzimologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética
8.
Methods Mol Biol ; 1094: 109-35, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24162984

RESUMO

This chapter describes the various methods derived from the protocol of standard fluorescent in situ hybridization (FISH) that are used in human, animal, plant, and microbial studies. These powerful techniques allow us to detect and physically map on interphase nuclei, chromatin fibers, or metaphase chromosomes probes derived from single-copy genes to repetitive DNA sequences. Other variants of the technique enable the co-localization of genes and the overall comparison of the genome among individuals of the same species or of different taxa. A further variant detects and localizes bacteria on tissues and cells. Overall, this offers a remarkable multiplicity of possible applications ranging from strict physical mapping, to clinical and evolutionary studies, making it a powerful and informative complement to other molecular, functional, or genomic approaches.


Assuntos
Análise Citogenética/métodos , Hibridização in Situ Fluorescente/métodos , Animais , Bactérias/metabolismo , Cromossomos Humanos/metabolismo , Cromossomos de Plantas/metabolismo , Hibridização Genômica Comparativa , DNA/metabolismo , Humanos , Sondas de Oligonucleotídeos/metabolismo , Plantas/metabolismo , Cariotipagem Espectral
9.
Clin Transl Oncol ; 8(8): 560-5, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16952844

RESUMO

Childhood leukemia is a common pediatric cancer in the developed world, the disease is biologically diverse and there is much discussion as to its causal mechanisms. Acute lymphoblastic leukemia (ALL) is the most common subtype and infants with ALL have a greatly increased risk of treatment failure. There are molecular and biological properties of leukemic cells that determine treatment outcome; these can usually be attributed to distinct genetic abnormalities that alter the normal proliferative and survival signals of hematopoietic cells. Experimental evidence for the existence of leukemic stem cells (LSC) has been obtained, and it is presumed that these cells arise from mutations in normal hematopoetic stem cells or progenitor cells, and they are difficult to eradicate. LSC seem to be surprisingly different from their normal counterparts and therefore are obvious new targets for drug therapy. Therapeutic concepts using monoclonal antibodies have substantially improved response rates in patients with malignant lymphomas and are currently being evaluated in other types of cancer.


Assuntos
Leucemia/genética , Leucemia/terapia , Adolescente , Anticorpos Monoclonais/uso terapêutico , Criança , Pré-Escolar , Humanos , Imunização Passiva , Imunoterapia Ativa , Lactente , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Células-Tronco
10.
Genes Chromosomes Cancer ; 42(3): 260-8, 2005 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-15609347

RESUMO

Although the histologic distinction between pheochromocytomas and head and neck paragangliomas is clear, little is known about the genetic differences between them. To date, various sets of genes have been found to be involved in inherited susceptibility to developing both tumor types, but the genes involved in sporadic pathogenesis are still unknown. To define new candidate regions, we performed CGH analysis on 29 pheochromocytomas and on 24 paragangliomas mainly of head and neck origin (20 of 24), which allowed us to differentiate between the two tumor types. Loss of 3q was significantly more frequent in pheochromocytomas, and loss of 1q appeared only in paragangliomas. We also found gain of 11q13 to be a significantly frequent alteration in malignant cases of both types. In addition, recurrent loss of 8p22-23 was found in 62% of pheochromocytomas (including all malignant cases) versus in 33% of paragangliomas, suggesting that this region contains candidate genes involved in the pathogenesis of this abnormality. Using FISH analysis on tissue microarrays, we confirmed genomic deletion of this region in 55% of pheochromocytomas compared to 12% of paragangliomas. Loss of 8p22-23 appears to be an important event in the sporadic development of these tumors, and additional molecular studies are necessary to identify candidate genes in this chromosomal region.


Assuntos
Neoplasias das Glândulas Suprarrenais/genética , Biomarcadores Tumorais/genética , Neoplasias de Cabeça e Pescoço/genética , Paraganglioma/genética , Feocromocitoma/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 11/genética , Cromossomos Humanos Par 3/genética , Cromossomos Humanos Par 8/genética , Feminino , Deleção de Genes , Humanos , Masculino , Pessoa de Meia-Idade , Hibridização de Ácido Nucleico
11.
Genes Chromosomes Cancer ; 42(3): 287-98, 2005 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-15611930

RESUMO

Molecular cytogenetic techniques enabled us to clarify numerical and structural alterations previously detected by conventional cytogenetic techniques in 37 patients who had myelodysplastic syndromes with complex karyotypes. Using high-resolution comparative genomic hybridization (HR-CGH), we found the most recurrent alterations to be deletion of 5q (70%), 18q (35%), 7q (32%), 11q (30%), and 20q (24%), gain of 11q (35%) and 8q (24%), and trisomy of chromosome 8 (19%). Furthermore, in 35% of the patients, 20 amplifications were identified. These amplifications were shown by FISH to involve some genes previously described as amplified in hematological malignancies, such as ERBB2, MLL, and RUNX1. In addition, two other genes, BCL6 and BCL2, which are classically related to apoptosis and non-Hodgkin lymphoma, were shown for the first time to be involved in amplification. Genomic alterations involving different subtelomeric regions with losses in 4p16, 5p15.3, 6q27, 18p11.3, and 18q23 and gains in 1p36.3 and 19p13.3 were detected by HR-CGH. Array CGH analysis of the subtelomeric regions in some samples was able to confirm a number of these alterations and found some additional alterations not detected by conventional CGH.


Assuntos
Aberrações Cromossômicas , Cromossomos Humanos/genética , Cariotipagem , Síndromes Mielodisplásicas/genética , Hibridização de Ácido Nucleico , Telômero/genética , Idoso , Idoso de 80 Anos ou mais , Cromossomos Artificiais Bacterianos/genética , Subunidade alfa 2 de Fator de Ligação ao Core , Proteínas de Ligação a DNA/genética , Feminino , Genes erbB-2/genética , Histona-Lisina N-Metiltransferase , Humanos , Masculino , Pessoa de Meia-Idade , Proteína de Leucina Linfoide-Mieloide , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-bcl-2 , Proteínas Proto-Oncogênicas c-bcl-6 , Proto-Oncogenes/genética , Fatores de Transcrição/genética
12.
Genes Chromosomes Cancer ; 41(4): 353-65, 2004 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15382261

RESUMO

Comparative genomic hybridization (CGH) has been widely used to detect copy number alterations in cancer and to identify regions containing candidate tumor-responsible genes; however, gene expression changes have been described only in highly amplified regions (amplicons). To study the overall impact of slight copy number changes on gene expression, we analyzed 16 T-cell lymphomas by using CGH and a custom-designed cDNA microarray containing 7,657 genes and expressed sequence tags related to tumorigenesis. We evaluated mean gene expression and variability within CGH-altered regions and explored the relationship between the effects of the gene and its position within these regions. Minimally overlapping CGH candidate areas (6q25, 13q21-q22, and 19q13.1) revealed a weak relationship between altered genomic content and gene expression. However, some candidate genes showed modified expression within these regions in the majority of tumors; these candidate genes were evaluated and confirmed in another independent series of 23 T-cell lymphomas by use of the same cDNA microarray and by FISH on a tissue microarray. When all the CGH regions detected for each tumor were considered, we found a significant increase or decrease in the mean expression of the genes contained in gained or lost regions, respectively. In addition, we found that the expression of a gene was dependent not only on its position within an altered region but also on its own mechanism of regulation: genes in the same altered region responded very differently to the gain or loss of genetic material. Supplementary material for this article can be found on the Genes, Chromosomes, and Cancer website at http://www.interscience.wiley.com/jpages/1045-2257/suppmat/index.html.


Assuntos
Cromossomos Humanos Par 13/genética , Cromossomos Humanos Par 19/genética , Cromossomos Humanos Par 6/genética , Dosagem de Genes , Linfoma de Células T/genética , Etiquetas de Sequências Expressas , Perfilação da Expressão Gênica , Humanos , Hibridização in Situ Fluorescente , Hibridização de Ácido Nucleico , Análise de Sequência com Séries de Oligonucleotídeos
13.
Genes Chromosomes Cancer ; 41(3): 191-202, 2004 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-15334542

RESUMO

The functional consequences of a high-hyperdiploid karyotype, found in up to one-third of cases of acute lymphoblastic leukemia (ALL), are unknown. Using the technique of comparative expressed sequence hybridization (CESH), we sought to address the question of whether increased chromosome copies in hyperdiploid ALL lead to increased gene expression. Relative expression of hyperdiploid ALL blasts versus peripheral blood mononuclear cells was analyzed in 18 patients. Common regions of overexpression corresponding to the presence of tri-/tetrasomies included: Xp22.1-22.2, 4q28, 6q14-15, 6q24, 10p13, 14q23-24, 17q21, 18q12, and 21q21, identified in 28-89% of cases. However, increased expression without underlying trisomy occurred at 3p21.3, 7q11.2, 8p21, and 8q24.1 in 39-90% of cases. High expression at 7q11.2, the most consistent change detected, was confirmed by quantitative PCR. Poor correlation between the presence of tri-/tetrasomy and overexpression was observed for chromosomes 14 and 17. Two cases were reanalyzed versus (i) B cells, (ii) transformed B cells, and (iii) CD34+19+ cells (the putative counterpart of the leukemic cell). A reduction in the number of relatively overexpressed regions was observed with CD34+19+ cells. In particular, the peak at 7q11.2 disappeared, suggesting up-regulation of genes from this region in the early ontology of normal B-cell development. In conclusion, we have shown that tri-/tetrasomies in hyperdiploid ALL lead to an increase in the expression of associated sequences. The choice of a biologically relevant reference is crucial for data interpretation.


Assuntos
Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Antígenos CD19/biossíntese , Antígenos CD34/biossíntese , Linfócitos B/metabolismo , Pré-Escolar , Cromossomos Humanos Par 7 , Primers do DNA/química , Diploide , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Lactente , Cariotipagem , Masculino , Hibridização de Ácido Nucleico , Reação em Cadeia da Polimerase , RNA/metabolismo , Análise de Sequência de DNA
14.
Cancer Genet Cytogenet ; 153(1): 39-47, 2004 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-15325092

RESUMO

We have performed a cytogenetic analysis of 23 myelodysplastic syndromes (MDS) with complex karyotypes (CK) using GTG-banding and spectral karyotyping techniques. Fifty-five percent of cases were hypodiploid, 34% were hyperdiploid, and 11% were pseudodiploid. The most recurrent alterations were monosomy of chromosomes 18, 5, and 7; trisomy of chromosome 8; and deletion of 5q, 11q, and 12p. Ninety-two structural alterations were mostly identified as unbalanced. The chromosomes and regions more frequently affected were 16q12, 17p11, and 20q11. Eight of 92 structural alterations were reciprocal translocations. Two translocations were recurrent, t(X;20)(p11.4;q11.2) and der(17)t(5;17)(?;p11.2); each one was present in about 10% of cases (2 cases, t[X:20] and 3 cases, t[5:17]). Mutations of TP53 were observed in five cases (22%), all with rearrangements affecting 17p. Total or partial inactivation of TP53 was detected in six cases (26%) as a result of loss of either both copies (four cases) or just one copy (two cases). Fluorescence in situ hybridization analysis showed amplification of genes previously identified in myeloid and/or hematological processes, such as HER2neu, MLL, and AML1, which could represent frequent events in MDS with CK.


Assuntos
Síndromes Mielodisplásicas/genética , Cariotipagem Espectral , Idoso , Idoso de 80 Anos ou mais , Anemia Refratária com Excesso de Blastos/genética , Aneuploidia , Aberrações Cromossômicas , Bandeamento Cromossômico , Feminino , Amplificação de Genes , Genes p53 , Humanos , Perda de Heterozigosidade , Masculino , Pessoa de Meia-Idade , Oncogenes , Translocação Genética/genética
15.
Cancer Genet Cytogenet ; 148(1): 35-43, 2004 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-14697639

RESUMO

Alveolar rhabdomyosarcomas (ARMS) are soft-tissue tumors that are genetically characterized by the presence of reciprocal translocations that generate the fusion gene PAX3-FOXO1A or PAX7-FOXO1A. For the study of the biologic consequences of such rearrangements, several cell lines have been generated. However, established cell lines accumulate chromosome and genetic aberrations that make it difficult to draw significant conclusions. We have applied a set of techniques that includes spectral karyotyping, fluorescence in situ hybridization (FISH), comparative genomic hybridization (CGH), and microarray CGH, to the most commonly used cell lines carrying the two fusion genes that are present in ARMS. We have identified the bacterial artificial chromosomes that cover the breakpoints at genes PAX3, PAX7, and FOXO1A, which can be used as FISH probes for the translocations. The RH30 cell line, positive for the PAX3-FOXO1A fusion gene, was found to be highly complex: wide range of chromosome number, more than 50 chromosome rearrangements, amplification of the hybrid gene, 24 DNA changes detected by conventional CGH, and 21 gene copy changes detected by microarray CGH (including several high-level amplifications). RMZ-RC2 cell line, positive for the PAX7-FOXO1A, was in the near-tetraploid range with only nonclonal structural rearrangements, amplification of the hybrid gene, 24 DNA changes by CGH, and 8 gene copy changes, confirming the previously reported high-level amplification of MYCN.


Assuntos
Aberrações Cromossômicas , Rabdomiossarcoma/genética , Linhagem Celular Tumoral , Bandeamento Cromossômico , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Hibridização de Ácido Nucleico , Translocação Genética
16.
Genes Chromosomes Cancer ; 36(4): 406-12, 2003 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-12619165

RESUMO

Erythroid leukemia (ERL or AML-M6) is an uncommon subtype of acute myeloid leukemia, the clinical, morphological, and genetic behavior of which needs further characterization. We analyzed a homogeneous group of 23 de novo AML-M6 patients whose bone marrow cells showed complex karyotypes. We also analyzed eight leukemia cell lines with erythroid phenotype, performing detailed molecular cytogenetic analyses, including spectral karyotyping (SKY) in all samples. The main features are: (1) A majority of patients (56%) had hypodiploidy. Loss of genetic material was the most common genetic change, especially monosomies of chromosome 7 or 18, and deletions of chromosome arm 5q. Taken together, 87% of the cases displayed aberrations involving chromosome 5 or 8. (2) We describe a novel, cryptic, and recurrent translocation, t(11;19)(p11.2;q13.1). Another translocation, t(12;21)(p11.2;q11.2), was found to be recurrent in a patient with ERL and in the K562 cell line. (3) MLL gene rearrangements were detected in 20% of cases (three translocations and three amplifications) and, overall, we defined 52 rearrangements (excluding deletions) with a mean of 2.3 translocations per patient. (4) Of the structural aberrations, 21% involved chromosomes 11 and 19. Most of the rearrangements were unbalanced; only 13 reciprocal translocations were observed. The general picture of chromosomal aberrations in cell lines did not reflect what occurred in patient samples. However, both primary samples and cell lines shared three common breakpoints at 19q13.1, 20q11.2, and 21q11.2. This is the first molecular cytogenetic description of the karyotype abnormalities present in patients with ERL. It should assist in the identification of genes involved in erythroleukemogenesis.


Assuntos
Aberrações Cromossômicas , Cromossomos Humanos Par 11/genética , Cromossomos Humanos Par 19/genética , Rearranjo Gênico/genética , Leucemia Eritroblástica Aguda/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Coloração Cromossômica , Feminino , Humanos , Lactente , Cariotipagem , Masculino , Pessoa de Meia-Idade , Taxa de Sobrevida
17.
Hum Genet ; 112(2): 178-85, 2003 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-12522559

RESUMO

An alternative model has been proposed for the development of clear-cell renal cell carcinoma (RCC) in families where chromosome 3 translocations segregate with the disease. In this model, the existence of a translocation involving chromosome 3 would favour the non-disjunctional loss of the derivative chromosome carrying the 3p segment. Additionally, subsequent somatic mutations in the VLH gene, located in 3p25-26, would inactivate this tumour suppressor gene. In the present work, we describe a new family with two clear-cell RCC affected members and a t(3;8)(p13;q24.1) translocation in two consecutive generations. We observed loss of the derivative chromosome carrying the 3p segment (der(8)) and somatic mutation of the VHL gene in the left-kidney tumoral tissue of the proband. His right-kidney tumour carried a different VHL mutation and loss of heterozygosity (LOH) was not detected. The mother of the proband was also clear-cell RCC-affected but the tumoral tissue analysed did not carry any VHL gene mutations. Another member of the family, a maternal aunt, had a papillary RCC and did not carry this translocation, the LOH on 3p or the VHL somatic mutations. Haplotype analysis of the three affected members revealed that they did not inherit a common region on 3p, confirming the different genetic origin of both tumour types. Finally, the presence of RCC in other non-available members of the family highlights the overall risk for RCC in families with chromosome 3 translocations. In the present work, we have confirmed the proposed mechanism for the development of clear-cell RCC in this family, although we cannot discard the existence of other genes, in addition to VHL, being involved in hereditary RCC.


Assuntos
Adenocarcinoma de Células Claras/genética , Carcinoma de Células Renais/genética , Cromossomos Humanos Par 3/genética , Cromossomos Humanos Par 8/genética , Neoplasias Renais/genética , Perda de Heterozigosidade , Translocação Genética , Proteínas Supressoras de Tumor , Ubiquitina-Proteína Ligases , Adenocarcinoma de Células Claras/patologia , Adulto , Idoso , Carcinoma de Células Renais/patologia , Aberrações Cromossômicas , Mapeamento Cromossômico , Segregação de Cromossomos , DNA de Neoplasias , Feminino , Genes Supressores de Tumor , Doenças Genéticas Inatas , Heterozigoto , Humanos , Hibridização in Situ Fluorescente , Neoplasias Renais/patologia , Ligases/genética , Masculino , Repetições de Microssatélites , Pessoa de Meia-Idade , Mutação , Hibridização de Ácido Nucleico , Linhagem , Reação em Cadeia da Polimerase , Proteína Supressora de Tumor Von Hippel-Lindau , Doença de von Hippel-Lindau/genética
19.
Int J Cancer ; 99(2): 305-9, 2002 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-11979449

RESUMO

The BRCA1 and BRCA2 genes are responsible for a high proportion of familial breast cancer; germline mutations in these genes confer a lifetime risk of about 70% for developing breast cancer. Most of the described deleterious mutations are small deletions or insertions that originate a truncated protein; however, in many cases, they are amino acid changes whose significance is unknown. In these cases, there are some tests that can analyze the meaning of these variants, but most remain unclassified. The BRCA genes are tumor suppressors and it is believed that complete loss of the wild-type allele is a common mechanism of inactivation in tumors from patients carrying a germline deleterious mutation in these genes; if this is true, loss of heterozygosity (LOH) analysis in the tumor sample could help to distinguish if a rare variant is either a deleterious mutation or a common polymorphism. In the present study, we performed LOH analysis at the BRCA loci in 47 tumors from patients who belonged to high-risk breast cancer families and were carriers of any type of alteration in these genes. Our results suggest that (i) loss of the wild-type allele is the most common mechanism of inactivation in tumors from patients who carry a deleterious mutation in any of the genes, (ii) this loss is not common when we analyze familial tumors not associated with mutations in BRCA and (iii) LOH can be used to clarify variants of unknown significance in the BRCA genes.


Assuntos
Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias da Mama/genética , Perda de Heterozigosidade , Alelos , Eletroforese , Feminino , Heterozigoto , Humanos , Mutação , Neoplasias Ovarianas/genética , Reação em Cadeia da Polimerase , Polimorfismo Genético , Polimorfismo Conformacional de Fita Simples , Sensibilidade e Especificidade
20.
Micron ; 33(4): 399-402, 2002.
Artigo em Inglês | MEDLINE | ID: mdl-11814878

RESUMO

N-Bromosuccinimide (NBS) is a known protein reagent able to modify amino acids and proteins, resulting in oxidation of tryptophan, tyrosine and histidine residues, as well as sulfhydryl, alcohol and phenol groups. These properties make NBS a suitable reagent to selectively block certain amino acid residues in biochemistry, and also permit the histochemical detection of proteins by oxidative deamination followed by the Schiff reaction. In this paper we show that, under ultraviolet excitation, NBS selectively reveals the cytoplasmic granules of mammalian eosinophils and chicken heterophils, rendering considerable white--blue fluorescence, in a remarkable fluorogenic reaction which rapidly increases at the beginning of the observation. This emission slightly decays afterwards and then remains almost stable still yielding a high level of emission after 10min of continuous excitation. Possible mechanisms underlying these results are discussed and we propose NBS as a very suitable fluorogenic reagent for the microscopical detection and analysis of proteins.


Assuntos
Bromosuccinimida/farmacologia , Eritrócitos/citologia , Animais , Galinhas , Eritrócitos/efeitos dos fármacos , Humanos , Microscopia de Fluorescência
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