DNA Damage-Inducible Gene, Reprimo Functions as a Tumor Suppressor and Is Suppressed by Promoter Methylation in Gastric Cancer

Abstract

In several types of human cancer, the gene expression of Reprimo, a highly glycosylated protein, is frequently silenced via methylation of its promoter. The aim of this study was to characterize the epigenetic inactivation of Reprimo and its biologic function and clinical relevance in gastric cancer. The correlation between Reprimo methylation and clinical relevance was assessed in 83 primary human gastric cancer tissues. The effects of Reprimo expression were also examined using in vitro and in vivo assays. Reprimo methylation was cancer specific and frequently observed. In two gastric cancer cell lines without Reprimo methylation, we observed faint or weak Reprimo expression under normal conditions and high expression under DNAdamaging conditions. In four gastric cancer cell lines with Reprimo methylation, however, Reprimo expression remained faint even under DNA-damaging conditions, with expression being restored in combination with agents that induce demethylation. Enforced Reprimo expression robustly inhibited cell proliferation and anchorage-independent colony formation and enhanced DNA damage-induced apoptosis. Inverse effects were observed via siRNA-mediated knockdown of endogenous Reprimo. Reprimo expression inhibited tumorigenesis in vivo. Reprimo methylation was also associated with a poor response in patients with gastric cancer treated with chemotherapy (P

Methylation-Sensitive Melt Curve Analysis of the Reprimo Gene Methylation in Gastric Cancer

“…The aberrant RPRM gene methylation has been associated with dysfunction of the cell cycle and carcinogenesis [5, 19]. Our data revealed that in healthy population or normal functional cell types (such as PBMCs), the RPRM gene was less frequently methylated.…”

Casticin inhibits the activity of transcription factor Sp1 and the methylation of RECK in MGC803 gastric cancer cells

“…Following treatment with casticin, MGC803 cells (1x10 7 ) underwent genomic DNA extraction and purification using the DNeasy Blood & Tissue kit (Qiagen GmbH, Hilden, Germany). A total of 200 ng genomic DNA was denatured at 100˚C for 3 min and placed on ice for cooling.…”

“…Therefore, DNA methylation serves an important role in maintaining chromatin structures, DNA conformation, DNA stability and DNA-protein interactions (5). However, abnormal methylation of DNA in certain pathological conditions may deactivate tumor suppressor genes and activate oncogenes (6,7). The deactivation of tumor suppressor genes, including RECK and Ras association domain family 1 isoform A (RASSF1A), is closely associated with the occurrence of gastric cancer (8)(9)(10).…”

Reprimo tissue-specific expression pattern is conserved between zebrafish and human

“…RPRM is a potential class II (inactivation occurring by epigenetic mechanisms) tumor suppressor gene (TSG). Inactivation is driven principally by aberrant methylation of its promoter region [4, 7, 8] and correlates with genetic instability in human cancers, such as gastrointestinal tumors [9, 10]. In addition, aberrant methylation of its promoter region has been proposed as a biomarker for non-invasive detection of gastric cancer [7].…”