Absence of mutations in the p53 tumor suppressor gene in woodchuck hepatocellular carcinomas associated with hepadnavirus infection and intake of aflatoxin B₁

Abstract

Infection with hepadnaviruses and exposure to aflatoxin B1 (AFB1) are considered major risk factors in the development of hepatocellular carcinoma (HCC) in humans and in animals. A high rate of mutations in the p53 tumor suppressor gene in hepatocellular carcinomas of predominantly hepatitis B virus (HBV) carrier patients has been recently related to dietary aflatoxin. Another member of the hepadnavirus family, the woodchuck hepatitis virus (WHV), infects woodchucks in a manner similar to that of HBV in humans. Therefore, it was of particular interest to determine whether the p53 gene in woodchuck HCCs associated with hepadnavirus infection and with exposure to AFB1 is affected in the same manner as in human HCCs. By direct PCR-sequencing, we analyzed exons 4-9 of the p53 gene in 13 HCCs from 12 woodchucks (two uninfected, ten WHV carriers). Six WHV carrier and two uninfected woodchucks were treated with AFB1. None of the analyzed HCC samples exhibited mutations, either in p53 gene exons 4-9, or in splicing donor-acceptor sites. The present data are consistent with our previous study that indicated a low rate of p53 mutations in HCCs of AFB1-treated ground squirrels, either infected or not infected with ground squirrel hepatitis virus, and in WHV carrier woodchucks not exposed to AFB1. Overall, our findings indicate that in woodchucks and in ground squirrels exposure to aflatoxin may affect the development of p53 mutations less than in humans.

The p53 codon 249 mutational hotspot in hepatocellular carcinoma is not related to selective formation or persistence of aflatoxin B1 adducts

“…Such a supposition is supported by the lack of codon 249 equivalent mutations in HCCs and preneoplastic hepatic lesions induced by a¯atoxin B 1 in nonhuman primates and other animals such as ducks, rats, and squirrels (Fujimoto et al, 1992;Du¯ot et al, 1994;Rivkina et al, 1994;Shen and Ong, 1996). The p53 gene in ducks lacks the equivalent codon 249 guanine base which is the preferred mutation site in humans (Du¯ot et al, 1994).…”

HBV integrants of hepatocellular carcinoma cell lines contain an active enhancer

“…A number of studies suggested that pX functionally inactivates p53 via several di erent mechanisms (Doitsh and Shaul, 1999 and the references therein). Interestingly, in woodchucks although no role has been attributed to pX in HCC development, the p53 gene is by large wild-type (Rivkina et al, 1996). It remained to be clari®ed how p53 is inactivated in these tumors.…”

Altered DNA Mutation Spectrum in Aflatoxin B1-Treated Transgenic Mice That Express the Hepatitis B Virus X Protein

“…Although immunohistochemical staining of liver tissue sections isolated from the mice used in this study (both AFB1 treated and untreated) did not reveal WT or mutant p53 accumulation (data not shown), this may reflect a difference between species for AFB1-induced HCC. Inactivating mutations in p53 are rare to nonexistent in liver tumor samples isolated from woodchucks infected with woodchuck hepatitis virus (42). In rats, AFB1 treatment leads to a high frequency of liver tumors containing mutations in codon 12 of the ras gene (35).…”