Hypermethylation and Clinicopathological Significance of RASAL1 Gene in Gastric Cancer

Background: Recent studies have suggested that expression of the RAS protein activator like-1 gene (RASAL1)is decreased in gastric carcinoma tissues and cell lines, indicated a role in tumorigenesis and development ofgastric cancer. Reduced expression of RASAL1 could result in aberrant increase of activity of RAS signalingpathways in cancer cells. However, the exact mechanism which induces down-regulation of the RASAL1 generemains unclear. This study aimed to determine the methylation status and regulation of RASAL1 in gastric cancer.Materials and
Methods: Using the methylation-specific polymerase chain reaction (MSP), the methylation statusof CpG islands in the RASAL1 promoter in gastric cancers and paired adjacent non-cancerous tissues from 40patients was assessed and its clinicopathological significance was analyzed. The methylation status of RASAL1 ingastric cancer lines MKN-28, SGC-790l, BGC-823, as well as in normal gastric epithelial cell line GES-l was alsodetermined after treatment with a DNA methyltransferase inhibitor, 5-aza-2’-doexycytidine (5-Aza-CdR). RASactivity (GAS-GTP) was assessed through a pull-down method, while protein levels of ERK1/2, a downstreammolecule of RAS signaling pathways, were determined by Western blotting.
Results: The frequencies of RASAL1promoter methylation in gastric cancer and paired adjacent non-cancerous tissues were 70% (28/40) and 30%(12/40) respectively (P<0.05). There were significantly correlations between RASAL1 promoter methylation withtumor differentiation, tumor size, invasive depth and lymph node metastasis in patients with gastric cancer(all P<0.05), but no correlation was found for age or gender. Promoter hypermethylation of the RASAL1 genewas detected in MKN-28, SGC-790l and BGC-823 cancer cells, but not in the normal gastric epithelial cell lineGES-1. Elevated expression of the RASAL1 protein, a decreased RAS-GTP and p-ERK1/2 protein were detectedin three gastric cancer cell lines after treatment with 5-Aza-CdR.
Conclusions: Aberrant hypermethylation ofthe RASAL1 gene promoter frequently occurs in gastric cancer tissues and cells. In addition, the demethylatingagent 5-Aza-CdR can reverse the hypermethylation of RASAL1 gene and up-regulate the expression of RASAL1significantly in gastric cancer cells in vivo. Our study suggests that RASAL1 promoter methylation may have acertain relationship with the reduced RASAL1 expression in gastric cancer.