Featured Article p16 Hypermethylation Is Associated with Hepatitis Virus Infection, Age, and Gender in Hepatocellular Carcinoma

Purpose: The tumor suppressor gene p16 is mainly inactivated by an epigenetic change involving promoter hypermethylation in hepatocarcinogenesis. The possible clinical impact of p16 methylation and the potential risk factors for this epigenetic alteration have not been thoroughly investigated. Experimental Design: We studied the methylation status and mRNA and protein expression of p16 in 50 hepatocellular carcinomas and corresponding nonneoplastic liver lesions using methylation-specific PCR, reverse transcription-PCR, and immunohistochemical techniques. Results: p16 hypermethylation was observed in 58% (29 of 50) of the hepatocellular carcinomas and 16% (6 of 38) of the corresponding chronic hepatitis and cirrhosis tissue samples. p16 methylation was significantly associated with mRNA and protein expression (P < 0.001 and P 0.003, respectively). All of the p16-methylated tumors were positive for hepatitis B virus or hepatitis C virus markers, but none of the virus-negative tumors exhibited p16 methylation (P 0.006). The frequency of p16 hypermethylation tended to be higher in hepatitis C virus-related tumors (23 of 32, 72%) than in hepatitis B virus-related tumors (6 of 13, 46%; P 0.1). Aberrant methylation of p16 was also related significantly to increasing age, female gender, and normal levels of serum PIVKA-II (P 0.02, 0.04, and 0.04, respectively). No statistically significant difference in survival was observed between patients with p16 hypermethylation and those without. Conclusions: Our observations suggest that p16 hypermethylation may contribute to hepatocarcinogenesis from an early stage and that multiple risk factors, such as viral infections, age, and gender, may be associated with p16 hypermethylation in hepatocarcinogenesis.