No consistent pattern of mutations in p53 and ras genes in liver tumors ofrat treated with the drinking water mutagen 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX)

The frequency of point mutations in p53 (exons 4-7) and in Ki-ras, Ha-ras, and N-ras (exons 1 and 2) and the expression of p53 protein were evaluated in the liver tumors of Wistar rats of a 104-week carcinogenicity study on 3 -chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), a chlorine disinf ection by-product in drinking water. Mutations were analyzed in 16 hepatoce llular adenomas, 7 hepatocellular carcinomas, 23 cholangiomas, and 2 cholan giocarcinomas of the MX-treated animals and one hepatocellular carcinoma an d cholangiocarcinoma in control animals using PCR-SSCP (polymerase chain re action-single-strand conformation polymorphism) or PCR-TGGE (temperature gr adient gel electrophoresis) and direct sequencing. The expression of the p5 3 protein (wild-type and mutated protein) was detected by immunohistochemis try (CM5 antibody). The expression of p53 and that of the proliferating cel l nuclear antigen (PCNA, 19 A2) were also evaluated in livers of female ani mals exposed to MX for week, 3 weeks, or 18 weeks. Altogether, Four mutatio ns were found in p53 in three tumors, in two hepatocellular adenomas, and o ne cholangiocarcinoma, all in Females receiving the highest MX dose (6.6 mg /kg/day) of the study. Three of the mutations were G:C --> A:T transitions and one was an A:T --> T:A transversion. The mutations were scattered at di fferent codons and positions of the codon. One hepatocellular adenoma conta ined two p53 mutations. All cholangiomas and cholangiocarcinomas, but no he patocellular adenomas and carcinomas, overexpressed the p53 protein. My tre atment did not induce p53 expression at any age in the liver or alter the e xpression of the PCNA in the liver of younger animals. The p53 protein was overexpressed in hyperplastic ducts in aged rats but not in bile ducts of y ounger rats (up to 24 weeks). No mutations were observed in either Ki-ras, Ha-ras, or N-ras of the liver tumors. These data suggest that point mutatio ns in p53 Ki-ras, Ha-ras, and N-ras are not involved in the MX-induced live r carcinogenesis in rats. Environ. Mel. Mutagen. 36:292-300, 2000. (C) 2000 Wiley-Liss, Inc.