Detection of allelic imbalance in MLH1 expression by pyrosequencing serves as a tool for the identification of germline defects in Lynch syndrome

摘要: Lynch syndrome is an autosomal dominant cancer susceptibility characterized by the early development of microsatellite unstable colorectal, endometrial and other cancers. caused germline heterozygous loss-of-function sequence mutations within mismatch repair genes MLH1, MSH2, MSH6 or PMS2. Some individuals with have constitutional epimutations, promoter methylation transcriptional inactivation a single allele in normal somatic tissues, while others lack identifiable pathogenic changes germline. We hypothesized that analysis relative levels allelic expression MLH1 would assist identification cryptic defects five presumed cases whose tumours demonstrated loss, but causative mutation remained unidentified. exploited common benign c.655A>G SNP (rs1799977) exon 8 to distinguish between two genetic alleles study their activity, using quantitative pyrosequencing assays. In one patients we detected loss deletion tumour, prompting renewed screening. A novel intronic splice was subsequently identified, which resulted entire from transcript. This assay also proved useful demonstrating gradual reversal epimutation during lymphoblastoid cell culture, suggesting this defect may not be stably maintained immortalized cells. Our findings illustrate behaviour can complement conventional molecular analyses providing new insight into epigenetic mechanisms underlying disease.