Abstract 180: Potential utility of Reprimo methylation as a sensitizer to anticancer treatment in gastric cancer

Abstract Introduction: Reprimo is a highly glycosylated protein, inducing cell cycle G2 arrest. Reprimo promoter methylation has been reported as the mechanism regulating its expression in gastric cancer, but the biological roles remain unclear. The aim of this study is to examine Reprimo from a functional point of view. Methods: The methylation status of Reprimo promoter was analyzed using quantitative-methylation specific polymerase chain reaction (Q-MSP), and the effects of expression were examined in gastric cancer cell lines (AZ521 and SH10) with no endogenous expression of Reprimo using stable transfection and phenotype analysis (WST assay and apoptosis assay). Results: Reprimo methylation was detected in 6 of 8 gastric cancer cell lines; the silenced or reduced expression could be restored by treatment with demethylating agents such as 5-aza-2’-deoxycitidine and/or trichostatin A. On Q-MSP, the optimal cutoff value (0.42) determined by the receiver operating characteristic (ROC) curve could significantly discriminate primary tumor (69%, 57/83) from the corresponding normal tissues (18%, 15/83, P<0.0001). AZ521 cells with enforced expression significantly enhanced sensitivity to 5-FU therapy, and augmented apoptosis without affecting cell cycle. On the other hand, SH10 cells without methylation exhibited the undetectable level of endogenous expression, and Reprimo expression was robustly induced under 5-fluorouracil (5-FU), one of the standard anticancer agents for gastric cancer. As a result, SH10 cells stably transfected with Reprimo exhibited similar level apoptosis with mock cells. Conclusion: Reprimo expression is inducible under 5-FU agent, that may lead to apoptosis in gastric cancer. However, its expression is frequently suppressed by promoter methylation. Thus, Reprimo methylation status is considered to have a great potential as a predictive biomarker to select patients who may be effective for 5-FU therapy in gastric cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 180.