Extensive methylation of hMLH1 promoter region predominates in proximal colon cancer with microsatellite instability

Background & Aims: Methylation of the hMLH1 promoter region has been sugges ted to cause microsatellite instability (MSI) in sporadic colorectal carcin oma (CRC). We studied the methylation profile in a wide region of the hMLH1 promoter and compared with the hMLH1 protein expression and MSI status in 88 cases of sporadic CRC. Methods: Na-bisulfite treatment and polymerase ch ain reaction single-strand conformation polymorphism analysis was performed using 5 sets of polymerase chain reaction primers spanning the promoter re gion of the hMLH1 to examine methylation status. Results were compared with immunostaining using anti-MLH1 monoclonal antibody and MSI status of the t umor samples. Results: Methylation status was classified as full or partial methylation. Full methylation indicates the methylation of all CpG sites i n the examined regions. Methylation of the hMLH1 promoter was observed in 8 8.9% (16 of 18) of CRCs showing high frequency MSI (MSI-H), among which 89% (14 of 16) had full methylation with reduced hMLH1 protein expression. All cases showing full methylation were proximal colon tumors with MSI-H. In c ases with partial methylation, only the upstream region of the hMLH1 promot er was methylated. Partial methylation was also shown in 33.3% (6 of 18) of the normal mucosa of MSI-H cases. Frequencies of methylation were signific antly correlated with female gender (P = 0.0009) and aging (P = 0.007). Con clusions: Full methylation of the hMLH1 promoter region and subsequent gene inactivation may play a crucial role in the carcinogenesis of MSI-H CRCs i n the proximal colon. Methylation upstream of the hMLH1 promoter appears to be an early event in the carcinogenesis of MSI-H tumors.