P16(INK4-ALPHA) PROMOTER IS HYPERMETHYLATED AT A HIGH-FREQUENCY IN ESOPHAGEAL ADENOCARCINOMAS

Loss of heterozygosity (LOH) of 9p21, which contains the p16(INK4a) tu mor suppressor gene locus, is one of the most frequent genetic abnorma lities in human neoplasia, including esophageal adenocarcinomas. Only a minority of Barrett's adenocarcinomas with 9p21 LOH have a somatic m utation in the remaining p16 allele, and none have been found to have homozygous deletions. To determine whether p16 promoter hypermethylati on may be an alternative mechanism for p16 inactivation in esophageal adenocarcinomas, we examined the methylation status of the p16 promote r in flow-sorted aneuploid cell populations from 21 patients with prem alignant Barrett's epithelium or esophageal adenocarcinoma. Using bisu lfite modification, primer-extension preamplification, and methylation -specific PCR, we demonstrate that the methylation assay can be perfor med on 2 ng of DNA (similar to 275 cells). Eight of 21 patients (38%) had p16 promoter hypermethylation and 9p21 LOH, including 3 patients w ho had only premalignant Barrett's epithelium. Our data suggest that p romoter hypermethylation with LOH is a common mechanism for inactivati on of p16 in the pathogenesis of esophageal adenocarcinomas.