RESUMO
OBJECTIVE: Studies of cervical cancer and its immediate precursor, cervical intraepithelial neoplasia 3 (CIN3), have identified genes that often show aberrant DNA methylation and therefore represent candidate early detection markers. We used quantitative PCR assays to evaluate methylation in five candidate genes (TNFRSF10C, DAPK1, SOCS3, HS3ST2 and CDH1) previously demonstrated as methylated in cervical cancer. METHODS: In this analysis, we performed methylation assays for the five candidate genes in 45 invasive cervical cancers, 12 histologically normal cervical specimens, and 23 liquid-based cervical cytology specimens confirmed by expert review as unequivocal demonstrating cytologic high-grade squamous intraepithelial lesions, thus representing the counterparts of histologic CIN3. RESULTS: We found hypermethylation of HS3ST2 in 93% of cancer tissues and 70% of cytology specimens interpreted as CIN3; hypermethylation of CDH1 was found in 89% of cancers and 26% of CIN3 cytology specimens. Methylation of either HS3ST2 or CDH1 was observed in 100% of cervical cancer tissues and 83% of CIN3 cytology specimens. None of the five genes showed detectable methylation in normal cervical tissues. CONCLUSION: Our data support further evaluation of HS3ST2 and CDH1 methylation as potential markers of cervical cancer and its precursor lesions.
Assuntos
Metilação de DNA , Displasia do Colo do Útero/genética , Neoplasias do Colo do Útero/genética , Adenocarcinoma/genética , Adenocarcinoma/patologia , Antígenos CD , Caderinas/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Feminino , Genes Supressores de Tumor , Marcadores Genéticos/genética , Humanos , Reação em Cadeia da Polimerase , Neoplasias do Colo do Útero/patologia , Displasia do Colo do Útero/patologiaRESUMO
The perturbations of the cytokine signaling pathway play an important role in lymphoid/hematopoietic tumors. Aberrant promoter methylation is the major mechanism of gene silencing in tumors. We examined 150 lymphoid/hematopoietic tumors or potential premalignant specimens, 55 control specimens and 12 EBV-transformed B lymphoblastoid cultures and 10 lymphoma/leukemia (L/L) or multiple myeloma (MM) cell lines for the methylation (and, in cell lines, of the expression status) of three genes involved in the cytokine signaling pathway. The genes were: SHP1, a protein tyrosine phosphatase; SYK, a protein kinase; and SOCS1, a suppressor of cytokine signaling. Our major findings were: (1) one or more of the three genes was frequently methylated in L/L and MM cell lines and there was good concordance (90-100%) between methylation and loss of gene expression; (2) treatment of L/L cell lines with a demethylating agent resulted in re-expression of SHP1 protein and downregulation of phosphorylated STAT3 in L/L cell lines; (3) all 55 control specimens and the lymphoblastoid cultures were negative for methylation of the three genes; (4) non-Hodgkin's lymphomas (100%), and leukemias (94%) had almost universal methylation of SHP1 and relatively less frequent (<30%) methylation of SOCS1 and SYK; (5) MM and monoclonal gammopathy of unknown significance (MGUS) had infrequent methylation of SHP1 (<20%), and occasional methylation of SOCS1 and SYK; and (6) comparable methylation frequencies for SOCS1 were observed in MM and MGUS, suggesting that SOCS1 methylation is an early event in MM pathogenesis. At least one gene was methylated in 119 of 130 (93%) of the malignant and 12 of 20 (60%) of the MGUS samples. Our findings demonstrate that the perturbations of cytokine signaling via silencing of these three genes are almost universal in lymphoid/hematopoietic tumors but the patterns of gene methylated for L/L and plasma cell dyscrasias are different.
Assuntos
Citocinas/metabolismo , Metilação de DNA , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/metabolismo , Transdução de Sinais , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Proteína Tirosina Fosfatase não Receptora Tipo 6 , Proteínas Tirosina Fosfatases/genética , Proteínas Tirosina Fosfatases/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/genética , Fator de Transcrição STAT3 , Transativadores/metabolismoRESUMO
Reprimo is a new candidate mediator of p53-mediated cell cycle arrest at the G2 phase. Loss of Reprimo gene expression accompanied by its promoter methylation was identified in pancreatic and lung cancers. Our aim was to examine the methylation status of Reprimo in a broad range of cancers. We examined Reprimo expression by RT-PCR and the DNA methylation status of the Reprimo promoter by MSP in 39 tumor cell lines. Loss or downregulation of Reprimo expression was frequent (62%), and we confirmed that transcriptional repression of Reprimo was caused by hypermethylation (overall concordance 92%). Treatment of expression-negative cells with 5-aza-2'-deoxycytidine restored Reprimo expression. We then examined aberrant methylation of Reprimo in 645 tumors representing 16 tumor types. Promoter methylation of Reprimo was found in 79% of gastric cancers, 62% of gallbladder cancers, 57% of lymphomas, 56% of colorectal cancers, 40% of esophageal adenocarcinomas, 37% of breast cancers and 31% of leukemias. Methylation frequencies in ovarian cancers, bladder cancers, cervical cancers, brain tumors, malignant mesotheliomas and pediatric tumors were lower (0-20%). Reprimo methylation was rarely detected in nonmalignant tissues (0-11%) except for gastric epithelia. While colorectal polyps were also frequently methylated (27%), chronic cholecystitis samples were infrequently methylated (4%). Furthermore, we failed to identify Reprimo mutation in colorectal and gastric cancer cell lines and 50 primary colorectal cancers. Aberrant methylation of Reprimo with loss of expression is a common event and may contribute to the pathogenesis of some types of human malignancy.