RESUMO
Epithelial ovarian cancer (EOC) has an innate susceptibility to become chemoresistant. Up to 30% of patients do not respond to conventional chemotherapy [paclitaxel (Taxol®) in combination with carboplatin] and, of those who have an initial response, many patients relapse. Therefore, an understanding of the molecular mechanisms that regulate cellular chemotherapeutic responses in EOC cells has the potential to impact significantly on patient outcome. The mitotic arrest deficiency protein 2 (MAD2), is a centrally important mediator of the cellular response to paclitaxel. MAD2 immunohistochemical analysis was performed on 82 high-grade serous EOC samples. A multivariate Cox regression analysis of nuclear MAD2 IHC intensity adjusting for stage, tumour grade and optimum surgical debulking revealed that low MAD2 IHC staining intensity was significantly associated with reduced progression-free survival (PFS) (p = 0.0003), with a hazard ratio of 4.689. The in vitro analyses of five ovarian cancer cell lines demonstrated that cells with low MAD2 expression were less sensitive to paclitaxel. Furthermore, paclitaxel-induced activation of the spindle assembly checkpoint (SAC) and apoptotic cell death was abrogated in cells transfected with MAD2 siRNA. In silico analysis identified a miR-433 binding domain in the MAD2 3' UTR, which was verified in a series of experiments. Firstly, MAD2 protein expression levels were down-regulated in pre-miR-433 transfected A2780 cells. Secondly, pre-miR-433 suppressed the activity of a reporter construct containing the 3'-UTR of MAD2. Thirdly, blocking miR-433 binding to the MAD2 3' UTR protected MAD2 from miR-433 induced protein down-regulation. Importantly, reduced MAD2 protein expression in pre-miR-433-transfected A2780 cells rendered these cells less sensitive to paclitaxel. In conclusion, loss of MAD2 protein expression results in increased resistance to paclitaxel in EOC cells. Measuring MAD2 IHC staining intensity may predict paclitaxel responses in women presenting with high-grade serous EOC.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biomarcadores Tumorais/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ciclo Celular/metabolismo , Resistencia a Medicamentos Antineoplásicos , Neoplasias Císticas, Mucinosas e Serosas/metabolismo , Neoplasias Epiteliais e Glandulares/tratamento farmacológico , Neoplasias Epiteliais e Glandulares/metabolismo , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/metabolismo , Proteínas Repressoras/metabolismo , Regiões 3' não Traduzidas , Biomarcadores Tumorais/genética , Proteínas de Ligação ao Cálcio/genética , Carboplatina/administração & dosagem , Carcinoma Epitelial do Ovário , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Quimioterapia Adjuvante , Intervalo Livre de Doença , Relação Dose-Resposta a Droga , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Proteínas Mad2 , MicroRNAs/metabolismo , Análise Multivariada , Gradação de Tumores , Estadiamento de Neoplasias , Neoplasias Císticas, Mucinosas e Serosas/genética , Neoplasias Císticas, Mucinosas e Serosas/mortalidade , Neoplasias Císticas, Mucinosas e Serosas/patologia , Neoplasias Císticas, Mucinosas e Serosas/terapia , Neoplasias Epiteliais e Glandulares/genética , Neoplasias Epiteliais e Glandulares/mortalidade , Neoplasias Epiteliais e Glandulares/patologia , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/mortalidade , Neoplasias Ovarianas/patologia , Paclitaxel/administração & dosagem , Inclusão em Parafina , Modelos de Riscos Proporcionais , Interferência de RNA , Proteínas Repressoras/genética , Estudos Retrospectivos , Medição de Risco , Fatores de Risco , Fatores de Tempo , Transfecção , Resultado do TratamentoRESUMO
The aim of this study was to identify genomic aberrations in endometrial cancer cells treated with the phyto-estrogenic compounds tectorigenin, irigenin and apigenin and to compare with those treated with beta-estradiol using array-based comparative genomic hybridisation (array CGH). The microarray contains 287 targets and includes telomeres, microdeletions, oncogenes and tumour suppressor genes and has increased mapping resolution compared to conventional CGH. An endometrial cancer cell line (Ishikawa) was cultured and treated with the phyto-estrogens. Treated cells were examined using the CGH microarray. Over 20 % of the array genes were aberrated in the cells treated with beta-estradiol, tectorigenin and irigenin compared to 3 % in those treated with the same concentration of apigenin. Protein kinase c zeta form, insulin, insulin receptor and protein-tyrosine phosphatase non-receptor-type 1 which are involved in insulin metabolism were aberrated by tectorigenin and irigenin. Apigenin may play a role in the treatment of endometrial cancer and in the treatment of postmenopausal women. Further studies in normal endometrium and primary endometrial cancer cells are needed to elucidate the role of the phyto-estrogens.