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Point mutations of c-ras genes were investigated in human angiosarcomas of the liver associated with occupational exposure to vinyl chloride. DNA prepared from either frozen or paraffin-embedded tissues was amplified by the polymerase chain reaction, and putative point mutations at codons 12, 13, and 61 of c-Ha-ras, c-Ki-ras, and N-ras were analyzed by dot-blot hybridization with allele-specific oligonucleotides. A G.C----A.T transition in the second nucleotide at codon 13 of the c-Ki-ras-2 gene was detected in 5 of 6 tumors. This mutation is likely a consequence of vinyl chloride-DNA adduct formation. It leads to the substitution of glycine by aspartic acid in the resulting p21 protein, a consistent amino acid substitution found so far in all types of human cancer exhibiting a codon 13-mutated Ki-ras gene.

Previous studies have shown that a high proportion (5/6) of human liver angiosarcomas (ASL) associated with exposure to vinyl chloride (VC) contains a GC=AT mutation at the Ki-ras codon 13. This mutation, however, has not been found in 5 ASL or 2 hepatocellular carcinomas (HCC) induced in rats by VC. These 2 HCC did contain a mutation at codon 61 of the Ha-ras gene. In order to extend this study and further explore the mechanisms of tumour induction, an additional 6 ASL and 6 HCC induced in rats by VC were analysed for ras gene point mutations, as well as 10 rat and 10 murine ASL induced by vinyl fluoride (VF), and 5 ASL, 6 Kupffer cell sarcomas, 4 HCC and 2 cholangiocellular carcinomas induced by Thorotrast in rats. Tumour DNA was analysed by PCR-SSCP and direct sequencing. None of the rodent ASL contained a mutation at codon 13 of the Ki-ras gene showing that the ras gene mutational pattern is species-specific. The CAA=CTA mutation, previously found at codon 61 of the Ha-ras gene in rat HCC, was observed in 5 further VC-induced HCC but was not detected in the Thorotrast-induced HCC, suggesting carcinogen-specificity. This mutation was also absent in VC-induced ASL, which supports the cell-specificity of the ras mutational pattern in chemically induced tumours. No predominant mutation was detected in VF-and Thorotrast-induced tumours. Thus, a given mutation in a tumour may be carcinogen-specific but also depend on the species and the cell type. Int.

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