Germline epimutation of MLH1 in early-onset colorectal cancer

Abstract

Germline mutation of the DNA mismatch repair (MMR) genes, particularly the MSH2 and MLH1 genes, causes the most common hereditary cancer, known as hereditary nonpolyposis colorectal cancer (HNPCC) syndrome. However, there remains a proportion of HNPCC families that have no detectable mutation in the DNA MMR genes despite of protein loss. This has prompted us to hypothesize that there might be other mechanisms apart from DNA mutation that causes inactivation of the MMR genes in certain HNPCC families. We have recently reported the germline epimutation of MSH2 gene in a putative HNPCC family and have confirmed its inheritance across three successive generations. Similarly, germline epimutation of MLH1 has been reported in a small number of individuals from several studies, yet it has never been documented in Chinese population. In this study, we identified germline MLH1 methylation in an early onset CRC patient with two metachronous cancers that lost MLH1 protein, where there is no detectable MLH1 gene mutation. Promoter hypermethylation of MLH1 were detected in both carcinomas and the adjacent normal mucosae by methylation specific PCR (MS-PCR). DNAs extracted from the peripheral blood leukocyte, buccal swab and hair follicles from that patient were all methylation positive by MS-PCR. We have further confirmed that the methylation event is specifically linked to a unique allele through SNP analysis. Unlike the case of MSH2 germline methylation, however, hypermethylation of MLH1 was not detected in the parents and two siblings in any tissues screened. Subsequent SNP and linkage analysis of this family has further revealed that the epimutation is not observed in the mother who carrying the same haplotype that is methylated in her daughter. These findings are consistent with previous reports of infrequent inheritance of germline MLH1 epimutation across generations and suggest that more than one epigenetic regulatory element may be involved in germline epimutation silencing.