title: Profile of methylated CpG sites of hMLH1 promoter in primary gastric carcinoma with microsatellite instability.
abstract: A recent study using colorectal cancer cell lines has identified methylation on a small region of hMLH1 promoter (-248 to -178 relative to the transcription start site) to be critical for gene silencing, but shown that methylation on a more upstream region is frequent in cell lines with hMLH1 expression. Because cultured cell lines have a higher degree of CpG methylation than primary tumors, we attempted to examine methylation profiles of CpG sites of hMLH1 promoter in primary gastric carcinomas with or without microsatellite instability (MSI). Seven cases with MSI and six cases without MSI were assessed for the methylation status of hMLH1 promoter by bisulfite-sequencing. All of the MSI-positive cases previously showed loss of hMLH1 expression and six cases displayed methylated alleles in methylation-specific PCR (MSP) for hMLH1. Sequencing analysis revealed that: (i) CpG sites were overall methylated in MSI-positive tumors with positive MSP results; (ii) a small region (-248-178) was almost invariably methylated in MSI-positive tumors; and (iii) the vast majority of CpG sites were unmethylated in MSI-negative tumors, including a more upstream region (proximal to -248). Our study suggests that methylation of a more upstream region observed in colon cancer cell lines may be an acquired change during cell line establishment and it was not identified in primary gastric carcinomas without MSI.
authors.affiliation: Department of Pathology, Seoul National University College of Medicine and Cancer Research Institute, Seoul, Korea. ghkang@snu.ac.kr
authors.forename: Gyeong Hoon
authors.initials: GH
authors.lastname: Kang
authors.forename: Sun
authors.initials: S
authors.lastname: Lee
authors.forename: Yhong Hee
authors.initials: YH
authors.lastname: Shim
authors.forename: Jin Cheon
authors.initials: JC
authors.lastname: Kim
authors.forename: Jae Y
authors.initials: JY
authors.lastname: Ro
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000230:Adenocarcinoma / Q000235:genetics* / Q000473:pathology; D000328:Adult; D000368:Aged; D002352:Carrier Proteins; D018899:CpG Islands / Q000235:genetics*; D019175:DNA Methylation; D017931:DNA Primers / Q000737:chemistry; D004273:DNA, Neoplasm / Q000032:analysis; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008297:Male; D018895:Microsatellite Repeats; D008875:Middle Aged; D000070957:MutL Protein Homolog 1; D009363:Neoplasm Proteins / Q000235:genetics*; D009687:Nuclear Proteins; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA; D013274:Stomach Neoplasms / Q000235:genetics* / Q000473:pathology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D017931:DNA Primers; D004273:DNA, Neoplasm; C501486:MLH1 protein, human; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D000070957:MutL Protein Homolog 1