Frequent k-ras-2 mutations and p16(INK4A) methylation in hepatocellular carcinomas in workers exposed to vinyl chloride

Vinyl chloride (VC) is a know animal and human carcinogen associated with l iver angiosarcomas (LAS) and hepatocellular carcinomas (HCC). In VC-associa ted LAS mutations of the K-ras-2 gene have been reported; however, no data about the prevalence of such mutations in VC associated HCCs are available. Recent data indicate K-ras-2 mutations induce P16 methylation accompanied by inactivation of the p16 gene. The presence of K-ras-2 mutations was anal ysed in tissue from 18 patients with VC associated HCCs, As a control group , 20 patients with hepatocellular carcinoma due to hepatitis B (n = 7), hep atitis C (n = 5) and alcoholic liver cirrhosis (n = 8) was used. The specif ic mutations were determined by direct sequencing of codon 12 and 13 of the K-ras-2 gene in carcinomatous and adjacent nonneoplastic liver tissue afte r microdissection. The status of p16 was evaluated by methylation-specific PCR (MSP), microsatellite analysis, DNA sequencing and immunohistochemical staining. All patients had a documented chronic quantitated exposure to VC (average 8883 ppmy, average duration: 245 months). K-ras-2 mutations were f ound in 6 of 18 (33%) examined VC-associated HCCs and in 3 cases of adjacen t non-neoplastic liver tissue. There were 3 G -->A point mutations in the t umour tissue. Air 3 mutations found in non-neoplastic liver from VC-exposed patients were also G --> A point mutations (codon 12- and codon 13-asparta te mutations). Hypermethylation of the 5' CpG island of the p16 gene was fo und in 13 of 18 examined carcinomas (72%). Of 6 cancers with K-ras-2 mutati ons, 5 specimens also showed methylated p16. Within the control group, K-ra s-2 mutation were found in 3 of 20 (15%) examined HCC, p16 methylation occu rred in 11 out of 20 (55%) patients. K-ras-2 mutations and p16 methylation are frequent events in VC associated HCCs. We observed a K-ras-2 mutation p attern characteristic of chloroethylene oxide, a carcinogenic metabolite of VC. Our results strongly suggest that K-ras-2 mutations play an important role in the pathogenesis of VG-associated HCC. (C) 2001 Cancer Research Cam paign.