RESUMO
BACKGROUND: Neurofibromatosis type 1 is one of the most common familial diseases, the hallmark of which is the development of multiple neurofibromas. These are benign nerve sheath tumours, which can transform into malignant peripheral nerve sheath tumours (MPNST). METHODS: The aim of this study was to identify differentially expressed microRNA (miRNA) in neurofibromas and MPNST obtained from patients with neurofibromatosis type 1 using microarray analysis. Differential expression was validated by reverse transcription quantitative-PCR, and functional studies were performed after transfection of miRNA oligonucleotide mimics into MPNST cells. RESULTS: Sixteen miRNA were significantly differentially expressed in MPNST compared with NF, and of these fourteen were downregulated in MPNST: these included miR-30e*, miR-29c*, miR-29c, miR-340*, miR-30c, miR-139-5p, miR-195, miR-151-5p, miR-342-5p, miR-146a, miR-150, miR-223, let-7 a and let-7 g with a false discovery rate of q=8.48E-03 for the least significant miRNA. In contrast, miR-210 and miR-339-5p were upregulated in MPNST compared with neurofibromas. Prediction softwares/algorithms identified a list of genes targeted by miR-29c including extracellular matrix genes and matrix metalloproteinase (MMP)-2, all of which are reported to be involved in cell migration and invasion. Functional studies in a MPNST cell line, sNF96.2, using a mimic of the mature miR-29c showed reduced invasion, whereas there was no change in proliferation. Zymography of the manipulated cells showed that MMP2 activity was also reduced when miR-29c expression was forced in sNF96.2. CONCLUSION: We provide evidence that reduction of miR-29c has a pivotal role in the progression of nerve sheath tumours and results by increasing the invasive/migratory properties of nerve sheath tumours.
Assuntos
Genes Supressores , MicroRNAs , Neoplasias de Bainha Neural/genética , Linhagem Celular Tumoral , Progressão da Doença , Regulação para Baixo , Matriz Extracelular/genética , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Neurofibromatose 1/genéticaRESUMO
Chordomas are radio- and chemo-resistant tumours and metastasise in as many as 40% of patients. The aim of this study was to identify potential molecular targets for the treatment of chordoma. In view of the reported association of chordoma and tuberous sclerosis complex syndrome, and the available therapeutic agents against molecules in the PI3K/AKT/TSC1/TSC2/mTOR pathway, a tissue microarray of 50 chordoma cases was analysed for expression of active molecules involved in this signalling pathway by immunohistochemistry and a selected number by western blot analysis. Chordomas were positive for p-AKT (92%), p-TSC2 (96%), p-mTOR (27%), total mTOR (75%), p-p70S6K (62%), p-RPS6 (22%), p-4E-BP1 (96%) and eIF-4E (98%). Phosphatase and tensin homologue deleted on chromosome 10 expression was lost in 16% of cases. Mutations failed to be identified in PI3KCA and RHEB1 in the 23 cases for which genomic DNA was available. Fluorescence in situ hybridisation analysis for mTOR and RPS6 loci showed that 11 of 33 and 21 of 44 tumours had loss of one copy of the respective genes, results which correlated with the loss of the relevant total proteins. Fluorescence in situ hybridisation analysis for loci containing TSC1 and TSC2 revealed that all cases analysed harboured two copies of the respective genes. On the basis of p-mTOR and or p-p70S6K expression there is evidence indicating that 65% of the chordomas studied may be responsive to mTOR inhibitors, rapamycin or its analogues, and that patients may benefit from combined therapy including drugs that inhibit AKT.
Assuntos
Cordoma/genética , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Western Blotting , Cordoma/patologia , Feminino , Humanos , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina , Pessoa de Meia-Idade , Complexos Multiproteicos , Análise de Sequência com Séries de Oligonucleotídeos , Fosfatidilinositol 3-Quinases/metabolismo , Análise Serial de Proteínas , Proteínas , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR , Fatores de Transcrição/metabolismo , Esclerose Tuberosa/tratamento farmacológico , Esclerose Tuberosa/genética , Esclerose Tuberosa/patologia , Proteína 1 do Complexo Esclerose Tuberosa , Proteína 2 do Complexo Esclerose Tuberosa , Proteínas Supressoras de Tumor/metabolismo , Adulto JovemRESUMO
Chordomas are malignant tumours that occur along the spine and are thought to derive from notochordal remnants. There is significant morphological variability between and within chordomas, with some showing prominent areas of chondroid differentiation. Our microarray data from a broad range of connective tissue neoplasms indicate that, at the transcriptional level, chordomas resemble cartilaginous neoplasms. Here we show that chordomas express many genes known to be involved in cartilage development, but they also uniquely express genes distinguishing them from chondroid neoplasms. The brachyury transcription factor, known to be involved in notochordal development, is only expressed by chordomas. Using a polyclonal antibody, we show that brachyury is expressed in the embryonic notochord and in all 53 chordomas analysed, labelling both chondroid and chordoid areas of these tumours. In contrast, the protein was not detected in over 300 neoplasms, including 163 chondroid tumours. Brachyury was not detected in the nucleus pulposus, arguing against the hypothesis that this tissue derives directly from the notochord. These data provide compelling evidence that chordomas derive from notochord and demonstrate that brachyury is a specific marker for the notochord and notochord-derived tumours.
Assuntos
Biomarcadores Tumorais/análise , Cordoma/genética , Proteínas Fetais/análise , Notocorda/embriologia , Neoplasias da Coluna Vertebral/genética , Proteínas com Domínio T/análise , Biomarcadores Tumorais/genética , Doenças das Cartilagens/diagnóstico , Doenças das Cartilagens/genética , Condrossarcoma/diagnóstico , Condrossarcoma/embriologia , Condrossarcoma/genética , Cordoma/diagnóstico , Cordoma/embriologia , Proteínas Fetais/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Imuno-Histoquímica/métodos , Queratinas/análise , Queratinas/genética , Notocorda/química , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Neoplasias da Coluna Vertebral/diagnóstico , Neoplasias da Coluna Vertebral/embriologia , Proteínas com Domínio T/genética , Distribuição TecidualAssuntos
Neoplasias da Mama/genética , Frequência do Gene/genética , Genes BRCA1/genética , Repetições de Microssatélites/genética , Mutação/genética , Expansão das Repetições de Trinucleotídeos/genética , Alelos , Proteína BRCA1/química , Proteína BRCA1/genética , Sítios de Ligação , Análise Mutacional de DNA , Feminino , França , Mutação em Linhagem Germinativa/genética , Humanos , Perda de Heterozigosidade/genética , Neoplasias Ovarianas/genética , Estrutura Terciária de Proteína , Dedos de Zinco/genéticaRESUMO
BRCA1 is a breast cancer susceptibility gene. Germline mutations in BRCA1 gene are found in 5 to 10% of breast cancer. The aim of this study is to screen the tunisian women with familial or sporadic breast cancer for BRCA1 gene mutations. The authors used the Protein Truncation Test (PTT) and DNA sequencing to detect BRCA1 gene mutations in 12 tunisian families with breast cancer and the Allele Specific Oligonucleotide-PCR (ASO-PCR) to detect the 185delAG and 1294del40 mutations in 150 tunisian women with sporadic breast cancer. A nonsens mutation was found, by PTT, in exon 11 of BRCA1 gene in one case of familial breast cancer. No mutation in the rest of exons was found by the DNA sequencing. The BRCA1 1294del40 mutation was found only in a patient with non familial breast cancer. The 185delAG mutation was absent in all cases of breast cancer. These data suggest that the germline mutation of BRCA1 is implicated in breast cancer in Tunisia and that the 185delAG mutation is absent in arab tunisian women.
Assuntos
Neoplasias da Mama/genética , Genes BRCA1/fisiologia , Predisposição Genética para Doença/genética , Mutação em Linhagem Germinativa/genética , Neoplasias Ovarianas/genética , Árabes/genética , Neoplasias da Mama/epidemiologia , Estudos de Casos e Controles , Códon sem Sentido/genética , Análise Mutacional de DNA , Éxons/genética , Feminino , Predisposição Genética para Doença/epidemiologia , Testes Genéticos , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Neoplasias Ovarianas/epidemiologia , Linhagem , Reação em Cadeia da Polimerase , Tunísia/epidemiologiaRESUMO
ATM mutations predispose cells to malignancy by promoting chromosomal instability. We have identified a family with multiple cancers that segregates a mutant allele of ATM, IVS61+2insTA, which causes skipping of exon 61 in the mRNA, as well as a previously undescribed polymorphism, IVS61+104C(54):T(46). The mutation was inherited by two sisters, one who developed breast cancer at age 39 and the second at age 44, from their mother, who developed kidney cancer at age 67. Molecular studies were undertaken to determine the role of the ATM gene in the development of cancer in this family. Studies of irradiated lymphocytes from both sisters revealed elevated numbers of chromatid breaks, typical of A-T heterozygotes. Studies on lymphoblastoid cell lines established from these individuals revealed abnormal p53 induction and apoptosis after DNA damage. Loss of heterozygosity (LOH) in the ATM region of chromosome 11q23.1 showed that the normal ATM allele was lost in the breast tumor of the older sister. LOH was not seen at the BRCA1 or BRCA2 loci. BRCA2 is not likely to be a cancer-predisposing gene in this family because each sister inherited different chromosomes 13 from each parent. The sisters share their maternal BRCA1 allele, although no mutation in this gene was detected in the family. Our findings suggest that haploinsufficiency at ATM may promote tumorigenesis, even though LOH at the locus supports a more classic two-hit tumor suppressor gene model.
Assuntos
Ataxia Telangiectasia/genética , Mutação , Neoplasias/genética , Proteínas Serina-Treonina Quinases/genética , Adulto , Idoso , Proteínas Mutadas de Ataxia Telangiectasia , Sequência de Bases , Neoplasias da Mama/genética , Proteínas de Ciclo Celular , DNA/genética , Dano ao DNA , Proteínas de Ligação a DNA , Éxons , Feminino , Genes BRCA1 , Heterozigoto , Humanos , Íntrons , Neoplasias Renais/genética , Perda de Heterozigosidade , Linfócitos/efeitos da radiação , Masculino , Dados de Sequência Molecular , Linhagem , Proteínas Supressoras de TumorRESUMO
A new type of mutation by deletion-insertion in BRCA-1 gene is found in three unrelated French breast/ovarian cancer families. Surprisingly, deletion and insertion occurred at the same nucleotide position at the end of exon 11 (3958del5ins4), thus generating a truncated protein. This original mutation consists in a deletion of 5 bp (CTCAG) and in an insertion of 4 different bp (AGGC). Here, we proposed two hypothesis to explain this phenomenom. The first hypothesis is the formation of a hairpin stem-loop structure comprising the mutational site and the sequence corresponding to the duplication insertion 2 nucleotides before the mutation. The second hypothesis, more speculative, consists in an abortive integration of a human mobile element as a human transposon (tigger 1) which involved a deletion of 5 bp during its excision and an insertion of 4 bases corresponding to the 5' extremity of the transposon.
Assuntos
Neoplasias da Mama/genética , Deleção de Genes , Genes BRCA1/genética , Mutação em Linhagem Germinativa/genética , Sequências Repetitivas Dispersas/genética , Mutagênese Insercional/genética , Neoplasias Ovarianas/genética , Adulto , Feminino , Humanos , Pessoa de Meia-IdadeRESUMO
Seventy French high-risk families were screened for germ-line BRCA-1 mutations. The BRCA-1 coding region and the exon-intron boundaries were sequenced, except when pre-screening by PTT revealed a truncated protein in exon 11. Germ-line BRCA-1 mutations were detected in 24% of families. The number of breast and ovarian cancers per family, a relative young age at ovarian cancer diagnosis, and the occurrence of breast and ovarian cancer in the same patient significantly predicted the presence of a BRCA-1 mutation. The low BRCA-1 mutation frequency suggested that some alterations were not detected and some families were probably BRCA-2 or BRCAx carriers.
Assuntos
Proteína BRCA1/genética , Neoplasias da Mama/genética , Mutação em Linhagem Germinativa , Neoplasias Ovarianas/genética , Adulto , Alelos , Neoplasias da Mama/epidemiologia , Feminino , França/epidemiologia , Deleção de Genes , Humanos , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Neoplasias Ovarianas/epidemiologia , Polimorfismo Genético , Fatores de RiscoRESUMO
A novel complex mutation consisting of a small deletion/insertion (3958del5ins4) was found in the breast cancer-1 gene (BRCA-1) in three unrelated French breast and/or ovarian cancer families. These mutations occurred at the same nucleotide position of the 3' end of exon 11. The wild-type sequence, CTCAG, was deleted and replaced by AGGC in the three families. The consequence is the generation of a stop codon, TAG, resulting in a truncated protein. We propose two different mechanisms to explain the generation of this complex mutation: (i) the simultaneous occurrence of a deletion and an insertion in a stem-loop structure and (ii) the abortive integration of a human transposable element (Tigger 1) that deleted 5 nucleotides and inserted a 4-nucleotide "scar", corresponding to the 5' extremity of the transposon.
Assuntos
Neoplasias da Mama/genética , Genes BRCA1/genética , Neoplasias Ovarianas/genética , Proteína BRCA1/genética , Códon de Terminação/genética , Feminino , França , Marcadores Genéticos , Haplótipos , Humanos , Pessoa de Meia-Idade , Mutação/genética , Reação em Cadeia da Polimerase , Deleção de Sequência/genéticaRESUMO
Ataxia-Telangiectasia (A-T) is a rare autosomal recessive disease characterised by cutaneous telangiectasia, cerebellar ataxia, immunodeficiency, high sensitivity to ionising radiation, chromosomal instability and an increased risk of cancer. The gene mutated in A-T patients, ATM, is located on chromosome 11q22-23. ATM heterozygotes are thought to have a high tendency to develop malignancies, such as breast cancer. In order to determine the contribution of heterozygous ATM mutation to cancer, studies of cancer-affected patients have been undertaken in non site-specific cancer families and sporadic breast cancer cases. No evidence of an important role of ATM heterozygous mutations has been shown. In order to give another contribution to these results, we tried to define a specific family phenotype according to the most common cancers observed in ATM heterozygotes. Breast and gastric cancers appear to be the most frequent malignancies in A-T carriers and one ATM germ-line mutation has been described in a breast/gastric cancer family. Therefore we further investigated the role of ATM mutation in additional breast/gastric cancer families. In eighteen families associating these two malignancies, we used the protein transcription/translation test to detect ATM mutations in the index case from each family. We found one case of ATM mutation which did not cosegregate with the gastric cancer in the family.