RESUMO
To explore the significance of epigenetic mechanisms in urinary bladder carcinogenesis mediated by methylation of cytosine in CpG dinucleotides at 5' promoter regions, we analysed the methylation status of a broad panel of different genes in transitional cell carcinomas (TCC) and nonurothelial cancers, among which the 14-3-3 sigma, SYK and CAGE-1 genes were recognised as promising target genes. Using methylation-specific PCR, the rate of DNA hypermethylation proved to be related to the various histopathological cancer subtypes. The higher frequency of promoter methylation of the 14-3-3 sigma (57.1%) and SYK (64.3%) genes in high-grade, high-stage TCC in association with a reduced or even lacking immunohistochemical protein expression than in low-grade, low-stage TCC (28.6% and 42.9%, respectively), indicates that aberrant methylation of these genes plays an essential role in the progression of TCC. The importance of DNA hypermethylation in the conversion of TCC from a low to a high malignant potential was strongly supported by the finding that, unlike superficial low-grade TCC, advanced muscle invasive TCC showed a concurrent promoter methylation of the 14-3-3 sigma, SYK and CAGE-1 genes. Squamous cell carcinomas revealed a peak incidence of hypermethylation of the 14-3-3 sigma gene (80%), and conversely, the lowest methylation frequency of the SYK gene (13.3%). Undifferentiated small cell carcinomas disclosed a promoter methylation of the 14-3-3 sigma, SYK and CAGE-1 genes in only a quarter each for the cases. Although a correlation between the methylation status and gene activity in squamous cell and undifferentiated small cell carcinomas was not observed, the underexpression of the SYK protein products in both cancer types and additionally of the 14-3-3 sigma protein in small cell carcinomas appeared to be related to the aggressive clinical behaviour of both these nonurothelial bladder carcinomas. The relevance of the high frequency of DNA hypermethylation of the CAGE-1 antigen in TCC and squamous cell carcinomas merits further study, particularly in relation to anticancer immunotherapy. The methylation status of the PTEN, COX-2, RUNX-3 and HIC-1 genes was found to be unaltered. In conclusion, the different patterns of aberrant methylation of the 14-3-3 sigma, SYK and CAGE-1 genes in the various histopathological cancer types of the urinary bladder point to a role in tumor cell differentiation, resulting in the phenotypical conversion of TCC into nonurothelial carcinomas and in the progression of TCC to a more malignant potential.
Assuntos
Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Carcinoma de Células de Transição/patologia , Metilação de DNA , Exonucleases/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Regiões Promotoras Genéticas/genética , Proteínas Tirosina Quinases/genética , Neoplasias da Bexiga Urinária/patologia , Proteínas 14-3-3 , Antígenos de Neoplasias/análise , Biomarcadores Tumorais/análise , Carcinoma de Células de Transição/genética , Carcinoma de Células de Transição/metabolismo , Subunidade alfa 3 de Fator de Ligação ao Core/genética , Ciclo-Oxigenase 2/genética , DNA de Neoplasias/genética , DNA de Neoplasias/metabolismo , Proteínas de Ligação a DNA/genética , Progressão da Doença , Exonucleases/análise , Exorribonucleases , Humanos , Imuno-Histoquímica , Peptídeos e Proteínas de Sinalização Intracelular/análise , Fatores de Transcrição Kruppel-Like , Proteínas de Membrana/genética , Mucosa/metabolismo , Proteínas de Neoplasias/análise , Proteínas Nucleares/análise , PTEN Fosfo-Hidrolase/genética , Fenótipo , Reação em Cadeia da Polimerase/métodos , Proteínas Tirosina Quinases/análise , Quinase Syk , Fatores de Transcrição/genética , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/metabolismoRESUMO
Tumor suppressor genes play a prominent role in the modification and progression of urinary bladder carcinogenesis as a result of classic genetic alterations. Little is known about the potential significance of epigenetic events, mediated by DNA hypermethylation. This prompted our investigation to explore the global Alu methylation and the promoter methylation of the novel putative tumor suppressor genes caveolin-1 and hDAB2IP, and of p53 in transitional cell carcinomas (TCC), squamous cell carcinomas and undifferentiated small cell carcinomas of the urinary bladder. Quantitative GeneScan analysis revealed that the various histopathological tumor entities showed considerable interindividual variations in the global methylation, but the overall rate did not significantly differ between the various cancer subtypes. With methylation-specific PCR, a high frequency of methylation of the promoter region of the caveolin-1 gene was detected in undifferentiated small cell carcinomas (50%) and in squamous cell carcinomas (25.9%), while TCC were found not to be methylated. By immunohistochemistry, all squamous cell carcinomas showed a strong diffuse overexpression of caveolin-1, whereas undifferentiated small cell cancers lacked any expression. High-grade, high-stage TCC disclosed a higher incidence (60%) and a substantially stronger expression than low-grade, low-stage TCC (42.9%). Our findings suggest that hypermethylation of the caveolin-1 gene and an abnormal protein expression play a crucial role in cell differentiation, and in the phenotypical conversion of TCC into nonurothelial carcinomas. Promoter methylation of the hDAB2IP gene occurred more frequently in advanced muscle invasive (72.7%) than in superficial noninvasive (50%) TCC. DNA hypermethylation of p53 was detected in a quarter of the low-grade, low-stage TCC and undifferentiated small cell carcinomas, but only sporadically in squamous cell carcinomas, and was absent in high-grade, high-stage TCC. In conclusion, aberrant methylation and abnormal protein expression of the caveolin-1-gene is involved in the formation of nonurothelial carcinomas of the urinary bladder and promoter methylation of the hDAB2IP gene in the progression of TCC from a low to a high malignant potential.