A specific CpG methylation pattern of theMGMT promoter region associated with reduced MGMT expression in primary colorectal cancers

Abstract

The enzyme O6-methylguanine-DNA methyltransferase (MGMT) protects cells from the cytotoxic and mutagenic effects of alkylating agents. Approximately 20% of tumor cell lines lack MGMT activity and are highly sensitive to alkylating agents. In established cancer cell lines, MGMT expression appears to be correlated with methylation of residues in both the promoter and the body of the gene. The effect of methylation of the MGMT promoter on gene expression and carcinogenesis in primary tumors is unknown. We investigated methylation of the MGMT promoter region in primary colorectal cancers and normal colonic mucosa. We used five methylation-sensitive restriction enzymes (BssHII, SacII, Eagl, Nael, and Smal) and Southern blot analysis to assess methylation in 46 cancers and 22 controls. Methylation of Eagl and Nael sites was seen in 12 tumors but in none of the 22 normal colorectal mucosa specimens. This difference was statistically significant (P<0.01). Methylation-sensitive single-nucleotide primer extension analysis of four additional cytosine residues confirmed methylation of the promoter region in the tumors identified by Eagl and Nael digestions and served to further quantitate the extent of methylation. Western blot analysis of 21 tumors revealed statistically significant lower MGMT expression in the eight tumors with methylation of the Eagl and Nael sites and nt -128 than in the 13 tumors lacking the methylation pattern (P<0.05). MGMT activity was lower in tumors with methylation than in tumors that were not methylated. The difference was not, however, statistically significant. We conclude that a subset of colorectal tumors is characterized by a specific methylation pattern in the MGMT promoter associated with reduced MGMT expression.

Generating mutations but providing chemosensitivity: the role of O6-methylguanine DNA methyltransferase in human cancer

“…This profile, thanks to the popularization of bisulfite-PCR techniques (Fraga and Esteller, 2002), has been expanded in additional reports of MGMT inactivation in other tumor types (Toyooka et al, 2001;Virmani et al, 2001;Choy et al, 2002;Lee et al, 2002;Maruyama et al, 2002;Smith-Sorensen et al, 2002), summarized in Figure 3. This aberrant MGMT methylation has been correlated with the loss of MGMT protein (Esteller et al, 1999a(Esteller et al, , 2002cHerfarth et al, 1999. ), lack of mRNA expression (Esteller et al, 2000a;Yin et al, 2003) and loss of enzymatic activity (Herfarth et al, 1999).…”

Promoter hypermethylation of MGMT is associated with protein loss in gastric carcinoma

“…Promoter hypermethylation and reduced MGMT expression has been found in some primary human carcinomas. [7][8][9] Data have also indicated that wild-type p53 may negatively regulate basal MGMT promoter activity. in vitro and in vivo studies have shown that induction of MGMT in the presence of DNAdamaging agents involves p53.…”

Methylation of O6-Methyl Guanine Methyltransferase Gene Promoter in Meningiomas - Comparison between Tumor Grades I, II, and III

“…Aberrant DNA hypermethylation at the promoter region DNA repair gene contributes to the progression of some cancers (Herfarth et al, 1999;Wu et al, 2008;Hibi et al, 2009;Sharma et al, 2009). We used MSP (Methylation-Specific PCR) to evaluate the status of the promoter methylation of MGMT gene in the different grades of meningioma tumors.…”

“…Previous reports have shown that MGMT hypermethylation occurs in many tumor types, including gliomas, large B-cell lymphomas, retinoblastomas, and cancers of the breast, lung, prostate, stomach, and colon (Herfarth et al, 1999;Wu et al, 2008;Hibi et al, 2009;Sharma et al, 2009). Many studies have observed a high frequency of MGMT methylation in tumors, but there are only a few studies on the status of MGMT methylation in meningiomas.…”