Activation of ras oncogene in aflatoxin-induced rat liver carcinogenesis.

Abstract

The presence of activated transforming genes was investigated in four primary aflatoxin-induced rat liver tumors in male Fischer rats, in two cell lines generated from such tumors, in an epithelial liver-derived nontransformed cell line, and in the latter cell line after transformation by aflatoxin B1 in vitro. When DNA extracted from these sources was transfected into NIH 3T3 cells, negative results were obtained from focus assays. Cotransfection of these DNA samples with a gene for resistance to G418, followed by selection for resistance to that antibiotic, and tumorigenicity testing in nude mice demonstrated DNA-mediated transfer of the neoplastic phenotype in all cases except for DNA from the nontransformed cell line. DNA extracted from these primary nude mouse tumors used in a secondary round of transfection with NIH 3T3 cells gave positive results in focus assays, which were conserved through succeeding rounds of transfection. By use of appropriate radiolabeled probes, activated ras oncogenes were detected in all samples. N-ras activation was detected in three of the primary rat liver tumors and both hepatoma cell lines. Ki-ras activation was detected in one primary rat liver tumor, and Ha-ras activation was detected in the cell line transformed in vitro with activated aflatoxin B1. The activated Ki-ras oncogene was further characterized by use of synthetic oligonucleotide probes and was shown to contain a G -* A transition at the second nucleotide in codon 12.There is strong epidemiological evidence for involvement of aflatoxins in the high incidence of hepatoma in certain third world areas (1), especially in synergism with hepatitis B virus (2) and alcohol (3). Within the same country, highly significant correlations with aflatoxin ingestion have been obtained for incidences of primary hepatoma in different areas (1, 4). Aflatoxin B1 (AFB,)-induced rat liver cancer has been extensively studied to gain insight into the mode of toxin action and the process of neoplastic induction (5). AFB1 is metabolized into its active form, AFB1-8,9-epoxide (6), which forms bulky adducts with DNA. In vivo, binding of AFB1 appears to be exclusively to guanine residues (7). On a molar basis, activated AFB1 is one of the most potent mutagens (8) so far studied, a property that correlates well with carcinogenicity (9). In foodstuffs naturally contaminated with aflatoxins (as in the peanut meal used in the present study) smaller amounts of aflatoxin G1 (AFG1) are frequently found, but the predominant aflatoxin present is AFB1. AFG, is hepatocarcinogenic in the rat but is less potent than AFB1 (10). The mechanism of activation of AFG1 and its binding to guanine bases in the DNA are similar to that of AFB1 (11).Activation of cellular oncogenes has been shown in many human and experimental tumors. Although 10-15% of all human tumors show activated oncogenes, mainly ras genes (12), some well-defined and controlled experimental carcinogenesis systems show a higher incidence of detectable transforming genes (13-18). In this...

Mutational activation of the c-Ha-ras gene in liver tumors of different rodent strains: correlation with susceptibility to hepatocarcinogenesis.

“…In contrast to the B6C3F1 mouse, liver tumors from rats analyzed so far did not contain any mutations within the Ha-ras gene (5). There are, however, reports demonstrating mutated Ki-ras or N-ras genes in some aflatoxin B1-induced liver tumors of the Fischer 344 rat (7)(8)(9). The reason for the differences in the activation of ras genes between B6C3F1 mouse and rat liver tumors is not known at present.…”

Frequent genomic imbalances suggest commonly altered tumour genes in human hepatocarcinogenesis

“…Growth factors, like IGF-II and transforming growth factor (TGF)-α, are known to be frequently overexpressed and support growth in an autocrine manner in HCCs (Derynck et al, 1987;Schirmacher et al, 1992;Nong et al, 1997), while the tumorigenic role of proto-oncogenes is poorly characterized. So far MYC overexpression and ras-gene mutations appear to be rather infrequent in human HCCs (Geissler and Gesien, 1997), in contrast to some well-characterized animal models (Sinha et al, 1988;Fourel et al, 1990;McMahon et al, 1990). Recently oncogenic mutations of the β-catenin gene have been found in 26% of human HCCs (de la Coste et al, 1998).…”

Do Gastroenterologists Consider Aflatoxins as Origin of Digestive System Cancers?

“…Transversions G→T or transitions G→A are produced in the third base of codon 249 of the p53 gene and in the first or second base of codon 12 of the H-ras gene [80][81][82][83][84][85]. [86]. When 12 tumors induced by AFB 1 were examined in rats, the genomic DNA of the 10 tumors was transformed and 2/8 had activated Ki-ras [87].…”