Promoter hypermethylation of RASSF1A and RUNX3 genes as an independent prognostic prediction marker in surgically resected non-small cell lung cancers

Abstract

Aberrant methylation of promoter CpG islands is known to be a major inactivation mechanism of the tumor suppressor and tumor-related genes. Some published studies suggest a relationship to exist between the methylation status of several genes and the prognosis in non-small cell lung cancer (NSCLC); hypermethylation of the specific genes may be expected to serve as a biomarker for the prognosis, after a curative resection of NSCLC. To determine the relationship between the methylation status of the tumor suppressor and the tumor-related genes, and the clinicopathologic characteristics, including the survival rate, in patients with NSCLC after a surgical resection, we studied methylation in 10 genes (DAPK, FHIT, H-cadherin, MGMT, p14, p16, RAR-beta, RASSF1A, RUNX3, and TIMP-3) in 101 NSCLC cases by methylation-specific PCR (MSP). The methylation frequencies of the 10 genes examined in NSCLC were 26% for DAPK, 34% for FHIT, 26% for H-cadherin, 14% for MGMT, 8% for p14, 27% for p16, 38% for RAR-beta, 42% for RASSF1A, 25% for RUNX3, and 12% for TIMP-3. Clinicopathologically, the patients with all stages of disease who had positive RASSF1A, RUNX3, or H-cadherin methylation status were found to have a significantly shorter duration of survival, as compared with the patients with a negative methylation status for those genes (RASSF1A:P=0.023, RUNX3:P=0.035, H-cadherin:P=0.039) in univariate analysis. Thereafter, while limiting our examination to patients with stage I disease, the patients who had a positive RASSF1A or RUNX3 methylation status were found to have a significantly shorter duration of survival, in comparison to the patients with negative methyaltion status for each of those genes (RASSF1A:P=0.022, RUNX3:P<0.01) in univariate analysis. Next, the histological differences were recognized that the patients with RUNX3 methylation had a shorter duration of survival in adenocarcinomas (ACs) (P=0.045), in contrast to those with RASSF1A methylation who had a shorter duration of survival in squamous cell carcinomas (SCCs) (P=0.021). In multivariate analysis, both positive RASSF1A methylation status, and positive RUNX3 methylation status were found to be independent prognostic factors (RASSF1A:P=0.031, RUNX3:P=0.028), as was TNM stage (P=0.004) and pleural involvement (P=0.037). In conclusion, the hypermethylation of RASSF1A or RUNX3 gene is therefore a useful biomarker to predict the prognosis in NSCLC, particularly RASSF1A due to SCCs and RUNX3 due to ACs.