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

Abstract

Sequence-dependent formation and lack of repair of polycyclic aromatic hydrocarbon-induced DNA adducts correlates well with the positions of p53 mutational hotspots in smoking-related lung cancers (Denissenko et al, 1996(Denissenko et al, , 1998. The mycotoxin a¯atoxin B 1 (AFB 1 ) is considered to be a major causative agent in hepatocellular carcinoma (HCC) in regions with presumed high food contamination by AFB 1 . A unique mutational hotspot, a G to T transversion at the third base of codon 249 of the p53 gene is observed in these tumors. To test whether a selectivity of AFB 1 adduct formation is related to this peculiar mutational spectrum, we have mapped AFB 1 -DNA adducts at nucleotide resolution using ligation-mediated PCR and terminal transferasedependent PCR. Human HepG2 cells were exposed to AFB 1 metabolically activated in the presence of rat liver microsomes. Signi®cant adduct formation was seen at the third base of codon 249. However, this was not the major site of AFB 1 adducts and strong adduction was also observed at codons 226, 243, 244, 245 and 248 in exon 7 of the p53 gene and at several codons in exon 8. The damage at codon 249 does not consist of a unique abasic site or ring-opened a¯atoxin B 1 adduct but rather is consistent with the principal N7-guanine adduct of AFB 1 . Time course experiments indicate that, under the conditions used, AFB 1 adducts are not removed in a strand-selective manner and adduct removal from the third base of codon 249 proceeds at a relatively fast rate (50% in 7 h). The incomplete correspondence between sites of persistent AFB 1 damage and the speci®c codon 249 mutation suggests that AFB 1 may not be involved in mutation of this site or that additional mechanisms such as parallel infection with hepatitis B virus may be required for selection of codon 249 mutants in HCC.

Potential role ofp53 mutation in chemical hepatocarcinogenesis of rats

“…In fact, a certain oncogenic agent could act on only one or more spots of the p53 gene DNA sequence. The effect of the hotspots is yet uncertain, because a certain oncogenic agent has made different mutational hotspots of p53 gene in different species [37][38][39][40] and tissues [28,39,40] . Amino acid residues 278 and 279 of the p53 protein have been found to be located in helix H 2 of loop-sheet-helix fitting in the major groove of DNA, allowing them to contact edges of the bases, and to play a central role in DNA recognition [41] , so mutation occurring in this area may cause p53 to lose its growth regulation.…”

DNA Repair Capacity-Related to Genetic Polymorphisms of DNA Repair Genes and Aflatoxin B1-Related Hepatocellular Carcinoma Among Chinese Population

“…Using the human p53 gene in an in vitro assay, codon 249 has been exhibited to be a preferential site for formation of AFB1-N 7 -Gua adducts, evidence consistent with a role for AFB1 in the mutations observed in HCC (50,53). Therefore, the codon 249 mutation of p53 gene has been defined as the hotspot mutation of p53 gene resulting from AFB1 and has become the molecular symbol of HCC induced by AFB1 exposure (54)(55)(56). A wide diversity of DNA damage produced by AFB1 exposure, if not repaired, may cause chromosomal aberrations, micronuclei, sister chromatid exchange, unscheduled DNA synthesis, and chromosomal strand breaks, and can be converted into gene mutations and genomic instability, which in turn results in cellular malignant transformation (19).…”

Genetic Single Nucleotide Polymorphisms (GSNPs) in the DNA Repair Genes and Hepatocellular Carcinoma Related to Aflatoxin B1 among Guangxiese Population

“…This mutation is tested in more than 40% of HCC from high-AFB1-exposure areas, but in either very rare or absent for those from low or null AFB1 exposure areas [36][37][38][39]. Therefore, G:C > T:A mutation in codon 249 of TP53 gene is regarded as the hotspot mutation caused by AFB1 and the molecular symbol of AFB1-related HCC [40][41][42].…”