Aberrant hypermethylation of RASSF2 in tumors and peripheral blood DNA as a biomarker for malignant progression and poor prognosis of esophageal squamous cell carcinoma.

As a tumor suppressor gene, RAS-association domain family 2 (RASSF2) is inactivated by promoter hypermethylation in different tumor cell lines and primary tumors. However, the role of RASSF2 in esophageal squamous cell carcinoma (ESCC) has remained uninvestigated. The aims of this study were to determine the role and methylation status of RASSF2 in esophageal cancer cell lines, ESCC tissues and white blood cells, and to evaluate the potential prognostic role of RASSF2 in ESCC. In the present study, we found frequent silencing of RASSF2 and up-regulation of the gene by 5-Aza-dC treatment in esophageal cancer cell lines. Aberrant methylation of the CpG sites close to the transcription start site induced silencing of RASSF2 expression and in vitro methylation of RASSF2 led to a significant decrease in luciferase activity. The results were further verified in clinical specimens and aberrant methylation of the CpG sites close to the transcription start site of RASSF2 was found in ESCC tumor tissues and peripheral white blood cells. Furthermore, RASSF2 hypermethylation was associated with lower level of RASSF2 expression. ESCC patients in stage III and IV, with negative expression or hypermethylation of the CpG sites close to the transcription start of RASSF2 demonstrated poor patient survival. Taken together, our results suggest that RASSF2 may function as a tumor suppressor gene that is inactivated through hypermethylation of CpG sites close to the transcription start site in ESCC and its expression or methylation may have prognostic implications for ESCC patients.