STATE OF THE P53 GENE IN HEPATOCELLULAR CARCINOMAS OF GROUND-SQUIRRELS AND WOODCHUCKS WITH PAST AND ONGOING INFECTION WITH HEPADNAVIRUSES

Infection with hepadnaviruses and exposure to dietary aflatoxin are co nsidered major risk factors in the development of hepatocellular carci noma (RCC) both in humans and in animals. Recently, a broad range of m utations in the p53 tumor suppressor gene has been reported in human H CCs, predominantly from hepatitis B virus carriers in areas with eithe r high or low levels of exposure to dietary aflatoxin. To determine wh ether p53 mutations are common to HCCs of hosts infected with related hepadnaviruses with and without treatment with anatoxin, we studied th e occurrence of mutations in the p53 gene in HCCs of ground squirrels and woodchucks with history of infection with ground squirrel hepatiti s virus (GSHV) and woodchuck hepatitis virus, respectively. Sequencing of wild type p53 genes from ground squirrels and woodchucks revealed remarkable homology between the two species with only a few amino acid differences in exons 4, 8, and 9. Using direct polymerase chain react ion sequencing, we analyzed the state of the p53 gene (exons 4-9) in 2 0 HCCs from ground squirrels (2 uninfected, 7 with past, and 11 with o ngoing infection with GSHV) and in 11 HCCs from woodchucks persistentl y infected with woodchuck hepatitis virus. Five GSHV carrier and two u ninfected ground squirrels received i.p. administration of aflatoxin B ,. We detected only one mutation in the p53 gene, of the tested animal s. This mutation was located in codon 176 of exon 5 in the HCC of a GS HV-positive ground squirrel treated with aflatoxin. Mutation was cause d by a G to T transversion in the second position of the codon, result ing in the replacement of cysteine with phenylalanine, and was accompa nied by a tumor-specific loss of heterozygosity. p53 allelic amino aci d variation with sequences coding for aspartic acid or asparagine was present in codon 61 in the variable region of exon 4 in both HCCs and nonneoplastic tissues of ground squirrels. In view of the considerably lower apparent rate of mutations in comparison to human HCCs, we sugg est a less important role for aflatoxin in the induction of p53 mutati ons in HCCs of ground squirrels. Alternatively, etiological factors ot her than p53 mutations may be of greater significance in the developme nt of HCC in ground squirrels and woodchucks.