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Reactivation of interspersed repetitive sequences due to loss of methylation is associated with genomic instability, one of the hallmarks of cancer cells.  LINE-1  hypomethylation is a surrogate marker for global methylation loss and is potentially a new diagnostic and prognostic biomarker in tumors. However, the correlation of  LINE-1  hypomethylation with clinicopathological parameters and the CpG island methylator phenotype (CIMP) in patients with liver tumors is not yet well defined, particularly in Caucasians who show quite low rates of HCV/HBV infection and a higher incidence of liver steatosis. Therefore, quantitative DNA methylation analysis of  LINE-1 ,  RASSF1A , and  CCND2  using pyrosequencing was performed in human hepatocellular carcinomas (HCC, n = 40), hepatocellular adenoma (HCA, n = 10), focal nodular hyperplasia (FNH, n = 5), and corresponding peritumoral liver tissues as well as healthy liver tissues (n = 5) from Caucasian patients. Methylation results were correlated with histopathological findings and clinical data. We found loss of  LINE-1  DNA methylation only in HCC. It correlated significantly with poor survival (log rank test,  p  = 0.007). An inverse correlation was found for  LINE-1  and  RASSF1A  DNA methylation levels (r 2  = -0.47,  p  = 0.002).  LINE-1  hypomethylation correlated with concurrent  RASSF1 / CCND2  hypermethylation (Fisher’s exact test,  p  = 0.02). Both  LINE-1  hypomethylation and  RASSF1A / CCND2  hypermethylation were not found in benign hepatocellular tumors (HCA and FNH). Our results show that  LINE-1  hypomethylation and  RASSF1A / CCND2  hypermethylation are epigenetic aberrations specific for the process of malignant liver transformation. In addition,  LINE-1  hypomethylation might serve as a future predictive biomarker to identify HCC patients with unfavorable overall survival.

LINE-1 hypomethylation in human hepatocellular carcinomas correlates with shorter overall survival and CIMP phenotype