863

Background: The aberrant methylation in regulatory regions of tumor suppressor genes is an established epigenetic mechanism for transcriptional silencing in human cancer. In addition a significant association between methylation and clinical outcome in various different cancer types has been described, including NSCLC. The molecular events for this epigenetic alteration still remain unknown. Evidence suggests that the over-expression of DNA-Methyltransferases (DNMT’s) is one potential mechanism for hypermethylation. The aim of this study was to investigate the influence of gene expression levels of the three DNMT-Isoforms (DNMT1, DNMT3a, DNMT3b) in the pathogenesis and prognosis of patients with NSCLC and describe their influence on the hypermethylation of Glutathione S-transferase pi (GST-pi), adenomatous polyposis coli (APC), the DNA repair gene O6-methylguanine DNA transferase (MGMT) and the Death-Associated Protein (DAP) Kinase. Patients and methods: Using a quantitative real time RT-PCR (Taqman®) method mRNA expression of DNMT1, DNMT3a, DNMT3b and promoter hypermethylation of GST-pi, APC, MGMT and the DAPKinase was measured in 91 matching tumor and non-malignant lung tissue samples from patients with curatively resected NSCLC, stage I to IIIa. Forty-three patients had squamous cell carcinomas, 33 patients had adenocarcinomas, and 15 patients had large cell carcinomas. Results: DNMT mRNA expression was detectable in tumor and adjacent normal tissue at following frequencies: DNMT1 normal (96.7%), tumor (98.9%); DNMT3a normal (98.9%), tumor (98.9%); DNMT3b normal (83.5%), tumor (95.6%). In tumor tissue the expression of all three 3 DNMT-isoforms was significantly higher compared with matched normal adjacent tissue (P<0.001, Wilcoxon test). Hypermethylation in tumor tissue was found in 20% for GST-pi, in 90% for APC, in 38% for MGMT and in 20% for DAPKinase. No association was found between clinical outcome or histopathologic parameters and DNMT mRNA expression or DNA-hypermethylation. Furthermore no correlation was found between the DNMT mRNA expression and DNA-hypermethylation. Multivariant analysis described DNA-hypermethylation and tumor stage as independent prognostic factors. Conclusion: The over-expression of all three DNMT-isoforms is an important mechanism in the pathogenesis of NSCLC. The deregulation of the DNMT mRNA expression seems not to be responsible for the aberrant DNA-hypermethylation in patients with NSCLC, which itself is an important prognostic marker in this malign disease.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]