Genetic and epigenetic markers to identify high risk patients for multiple early gastric cancers after treatment with endoscopic mucosal resection

Abstract

The development of multiple gastric cancer is a major problem after the endoscopic resection of the first early gastric cancer. To find out markers to identify high risk patients, we analyzed the microsatellite instability (MSI) status and hypermethylation of tumor-related genes in multiple gastric cancers. Sixty-four adenocarcinomas resected by endoscopy, including 32 early solitary gastric cancers (SGCs) from 32 patients and 32 multiple gastric cancers (MGCs) from 14 patients, were employed. We analyzed MSI and the methylation status of promoter regions of the hMLH1, MGMT, p16 and E-cadherin using methylation-specific Polymerase Chain Reaction. Expression levels of hMLH1 were examined by immunohistochemistry. MSI (+) was detected in 5 of the 14 (35.7%) patients with MGCs, and in only 3 of the 32 patients (9.3%) with SGCs. Significant differences were observed between the 2 groups (p<0.001). Hypermethylation of hMLH1 was more frequently detected in MGCs than in SGCs (p<0.01), whereas significant difference was not observed in the frequency of MGMT, p16 or E-cadherin promoter methylation between the 2 groups. In conclusion, our results indicate that inactivation of hMLH1 through promoter hypermethylation may be involved in the development of multiple gastric cancers following the MSI pathway.

Keywords: CpG island methylation; microsatellite instability; multiple gastric cancer.

Fig. 1

Representative results of microsatellite analyses in early gastric carcinomas. In case 2 (upper panel), each microsatellite demonstrates identical pattern between normal gastric mucosa (N) and cancer (T). On the other hand, BAT-26, BAT-25 and BAT 40 show extra-peaks (arrows) in cancer DNA from case 16.

Fig. 2

Representative results of methylation-specific PCR of hMLH1, p16, MGMT and E-cadherin by using both methylated (M) and unmethylated (U) specific primers in gastric cancer. Methylated PCR products were detected in case 57 for hMLH1, in case 17 and 18 for p16, in case 47, 49 and 50 for MGMT and in case 31 for E-cadherin. P, positive control.

Fig. 3

Comparison of the frequency of the promoter hypermethylation of hMLH1, MGMT, p16 and E-cadherin in MSI (+) and MSI (−) gastric cancers. The incidences of promoter hypermethylation of hMLH1, MGMT and p16 are significantly higher in the MSI (+) tumors than in the MSI (−) tumors. On the other hand, significant difference is not observed in the frequency of E-cadherin promoter methylation between the 2 groups.