Genome-wide DNA Methylation Profiles in Liver Tissue at the Precancerous Stage and in Hepatocellular Carcinoma

To clarify genome-wide DNA methylation profiles during hepatocarcinogenesis, bacterial artificial chromosome (BAC) array-based methylated CpG island amplification was performed on 126 tissue samples. The average numbers of BAC clones showing DNA hypo- or hypermethylation increased from noncancerous liver tissue obtained from patients with hepatocellular carcinomas (HCCs) (N) to HCCs. N appeared to be at the precancerous stage, showing DNA methylation alterations that were correlated with the future development of HCC. Using Wilcoxon test, 25 BAC clones, whose DNA methylation status was inherited by HCCs from N and were able to discriminate 15 N samples from 10 samples of normal liver tissue obtained from patients without HCCs (C) with 100% sensitivity and specificity, were identified. The criteria using the 25 BAC clones were able to discriminate 24 additional N samples from 26 C samples in the validation set with 95.8% sensitivity and 96.2% specificity. Using Wilcoxon test, 41 BAC clones, whose DNA methylation status was able to discriminate patients who survived more than 4 years after hepatectomy from patients who suffered recurrence within 6 months and died within a year after hepatectomy, were identified. The DNA methylation status of the 41 BAC clones was correlated with the cancer-free and overall survival rates of patients with HCC. Multivariate analysis revealed that satisfying the criteria using the 41 BAC clones was an independent predictor of overall outcome. Genome-wide alterations of DNA methylation may participate in hepatocarcinogenesis from the precancerous stage, and DNA methylation profiling may provide optimal indicators for carcinogenetic risk estimation and prognostication.