Aberrant methylation of Reprimo in lung cancer

Deregulation of cell cycle inhibition contributes to human carcinogenesis. Reprimo (for stop/repress) is a newly identified mediator of the p53-mediated cell cycle arrest at the G₂ phase. Loss of Reprimo expression due to promoter methylation was recently identified in pancreatic cancer. We examined Reprimo expression by reverse transcription PCR (RT-PCR) and aberrant methylation of Reprimo by methylation specific PCR (MSP) in lung cancer cell lines (n = 35) and primary tumors (n = 167). We also correlated the p53 gene status with Reprimo methylation in cell lines. Aberrant methylation of Reprimo was present in 32% (six of 19) of non-small cell lung cancer (NSCLC) cell lines, 6% (one of 16) of small cell lung cancer (SCLC) cell lines, and 31% (51 of 167) of primary tumors. Methylation was absent in normal lymphocytes and was rare in corresponding nonmalignant lung tissues (7%; four of 57). Overall concordance between loss of expression and aberrant methylation of Reprimo was 94% (33 of 35) in cell lines. Reprimo expression was restored after treatment with the demethylating agent 5-aza-2′-deoxycytidine in all five-cell lines tested that lacked Reprimo expression. There was no significant correlation between p53 gene status and Reprimo methylation in cell lines. These data indicate that Reprimo methylation is frequent in lung cancers and occurs independently of p53 status. Methylation of Reprimo may play a role in the pathogenesis of lung cancers.