Frequent k- ras -2 mutations and p16INK4Amethylation in hepatocellular carcinomas in workers exposed to vinyl chloride

Abstract

Summary Vinyl chloride (VC) is a know animal and human carcinogen associated with liver angiosarcomas (LAS) and hepatocellular carcinomas (HCC). In VC-associated 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 analysed in tissue from 18 patients with VC associated HCCs. As a control group, 20 patients with hepatocellular carcinoma due to hepatitis B (n = 7), hepatitis C (n = 5) and alcoholic liver cirrhosis (n = 8) was used. The specific mutations were determined by direct sequencing of codon 12 and 13 of the K-ras-2 gene in carcinomatous and adjacent nonneoplastic liver tissue after 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 found in 6 of 18 (33%) examined VC-associated HCCs and in 3 cases of adjacent non-neoplastic liver tissue. There were 3 G → A point mutations in the tumour tissue. All 3 mutations found in non-neoplastic liver from VC-exposed patients were also G → A point mutations (codon 12-and codon 13-aspartate mutations). Hypermethylation of the 5′ CpG island of the p16 gene was found in 13 of 18 examined carcinomas (72%). Of 6 cancers with K-ras-2 mutations, 5 specimens also showed methylated p16. Within the control group, K-ras-2 mutation were found in 3 of 20 (15%) examined HCC. p16 methylation occurred 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 pattern 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 VC-associated HCC. prove the association of their cancer with VC exposure. As a control group, hepatocellular carcinoma tissue from 20 patients with chronic hepatitis B and C virus infection and alcohol-induced cirrhosis with HCC was used. The presence of K-ras-2 mutations was examined by direct sequencing after microdissection of the tumour nodules. MATERIALS AND METHODS Patients and tissue samplesData were extracted from the German industrial professional association for the chemical industry which is responsible for statutory accident insurance and provides medical and social rehabilitation and financial compensation to persons who incur an industrial injury or occupational disease. According to the German social legislation code VII, compensation for work-related impairment and disability is only possible in cases of a proven association of occupational factor and disease. The criteria for the patients included in this study were a documented chronic quantitated exposure to VC, which leads ...

Hepatocellular carcinoma and the risk of occupational exposure

“…induce specific gene mutations in the liver [105][106][107][108][109][110][111][112][113][114][115][116][117] . Various European and Italian studies have reported the apparent association between the amount and timing of exposure to VCM and development of HCC in those exposed [118][119][120] .…”

Genetics of hepatocellular tumors

“…Recently, the presence of KRAS2 mutations was observed in 33% of 18 vinyl chloride-associated HCCs and three mutations were found in adjacent nonneoplastic liver tissue (Weihrauch et al, 2001). The KRAS mutations observed were probably caused by chloroethylene oxide, a carcinogenic metabolite of vinyl chloride.…”

“…Loss of heterozygosity at the short arm of chromosome 9, in the region of p16 INK4 , was found in about 20% of HCC cases (Boige et al, 1997;Nagai et al, 1997;Laurent-Puig et al, 2001). Methylation of the p16 INK4 promoter leading to the absence of protein expression was found in 30-70% of the tumors (Liew et al, 1999;Matsuda et al, 1999;Jin et al, 2000;Weihrauch et al, 2001). Somatic inactivating mutations and homozygous deletions of the gene are rarely observed in HCC (Biden et al, 1997;Jin et al, 2000).…”

Prevalence of K-Ras mutations in hepatocellular carcinoma: A Turkish Oncology Group pilot study

“…This is similar to the findings of previous studies, which reported that the K-Ras mutation is an uncommon or even rare event in HCC. This is in contrast to the frequent presence of K-Ras mutations in chemical models of HCC in animals (1,(19)(20)(21)(22)(23)(24). Considering the small sample size of the previous and present studies, the true prevalence of K-Ras mutations in HCC has yet to be determined.…”