Silencing-specific methylation and single nucleotide polymorphism of hMLH1 promoter in gastric carcinomas

AIM:To investigate CpG methylation and single nucleotide polymorphism(SNP)of a specific promoter region of hMLH1 in primary gastric carcinoma. METHODS:Primary gastric carcinomas(n=80),their corresponding normal mucosal samples,and gastric mucosal biopsies from normal/gastritis control patients(n=54)were used.Hypermethylation at-253 nt and-251 nt in relation with the translational start site and SNP of a silencing specific region(-339 nt-46 nt)in the hMLH1 promoter were analyzed by BstUI-combined bisulfite assay(COBRA),denaturing high performance liquid chromatogram(DHPLC),and sequencing. RESULTS:(A)The specific methylation at-253 nt and-251 nt was observed in 2 of 60 primary gastric carcinomas,but neither in all of the corresponding mucosa nor in normal/ gastritis samples,by Bst UI-COBRA and DHPLC.(B)The hMLH1 promoter was methylated homogeneously in the xenograft of the primary gastric carcinoma with the methylated and unmethylated hMLH1.(C)The pattern of SNP at-93 nt of the hMLH1 promoter in 54 Chinese patients with gastric carcinoma was the same as that in the control patients:51% was A/G heteroalleles,34% and 15% were A/A and G/G homoalleles,respectively. CONCLUSION:Biallelic inactivation of hMLH1 by epigenetic silencing existed in human primary gastric carcinoma homogeneously.Hypermethylation of hMLH1 may play a role in the early stage of development of a few gastric carcinomas.The SNP at-93 nt is not related to the susceptibility of gastric carcinomas.

Antitumor effect of Gefitinib, an epidermal growth factor receptor tyrosine kinase inhibitor, combined with cytotoxic agent on murine hepatocellular carcinoma

AIM: To investigate the inhibitory effect of gefitinib combined with cytotoxic agent cisplatin (CDDP) on hepatocellular carcinoma (HCC). METHODS: Female Kunming mice and H22 hepatocarcinoma cells were used. Gefitinib at daily dose of 100 mg/kg body weight (BW) or lecithin liquid was given by gastrogavage once a day for 5 or 10 successive days. CDDP or normal saline (NS) was administered intraperitoneally (i.p.) once a day for 5 successive days. Mice were randomly divided into control group (lecithin, or NS, i.p.), CDDP group (daily dose, 1.2 mg/kg BW; d1-5, or d6-10), Gefitinib (d1-5, or d6-10, or d1-10), and Gefitinib combined with CDDP groups. The inhibitory rate (IR) of tumor, net BW, spleen index (SI), thymus index (TI) and the amount of peripheral blood cells of mice were detected on the 12th experiment day. RESULTS: The growth of HCC in mice was inhibited by Gefitinib alone (IR: 41% in d1-10 group and 30% in d1-5 group, respectively) or CDDP alone (IR: 32-54% in d1-5 group or d6-10 group). The highest inhibitory effect (IR: 56%) on HCC growth was observed in Gefitinib (d1-10) combined with CDDP (d1-5) group. Higher inhibition was also observed in CDDP (d1-5) followed by Gefitinib (d6-10) group than that in Gefitinib (d1-5) followed by CDDP (d6-10) group (IR: 61% vs 36%, P<0.01) in the independent study. Net BW, SI, TI and the amount of blood cells of mice in Gefitinib alone group were not significantly different from those in control groups. CONCLUSION: Gefitinib can significantly inhibit the growth of murine H22 hepatocellular carcinoma. If Gefitinib is used after CDDP treatment in animal experiments, the inhibitory effect could be enhanced.