Hepatocellular carcinoma (HCC) is the third and fourth commonest cancer in Chinese men and women, respectively. Hepatitis B virus (HBV) has been identified as the risk factor with more significance in populations in which infection by this virus is endemic. Among the environmental factors that putatively contribute to the etiology of HCC is aflatoxin contamination of the diet, in particular aflatoxin B1 (AFB1). Aflatoxins represent a group of mycotoxins produced by the fungi Aspergillus flavus and Aspergillus parasiticus. In vitro studies have shown that AFB1 preferentially induces the transversion of G to T in the third position of codon 249 of p53 gene. The extent of AFB1 contamination varies significantly among different countries or regions of the world. The arbitrary classification of a certain region as a high or low aflatoxin exposure area has led to contradictory results in terms of the correlation between mutations at codon 249 in the p53 gene and exposure to AFB1. One of the major limitations in these studies is the number of HCC subjects from high AFB1 exposure areas compared to moderate and low areas. The only studies from high exposure areas are from Qi-Dong and southern Africa, there are no reports from Guangxi, China, an area where aflatoxin is well confirmed to play a significant role in the etiology of HCC. Our working hypothesis is that by analyzing the mutational spectra for p53 and the HBV status in an area well-known for its high aflatoxin exposure we will be able to better define the role of these elements in aflatoxin hepatocarcinogenesis. We have obtained DNA from 68 paraffin-embbeded HCC from patients from southern Guangxi. We analyzed for the presence of 249 mutation by using an RFLP assay and direct sequencing. We have completed the analysis for 16 samples, and we have found an incidence of 249 mutation of 31% (5/16). The analysis of the rest of the samples is currently being done and should be completed in the next two months. All samples are currently being immunostained for p53 in order to correlate presence of mutation and accumulation of protein. We have also investigated the status of infection by HBV of the samples using an immunohistochemistry assay and we found that of 53 samples analyzed so far, 51% (27/53) are negative, 40% (21/53) are positive and 9% (5/53) could not be analyzed. All samples have being microdissected using a laser capture microdissection equipment in order to retrieve samples of normal tissue from each patient. These samples could be later used for genotyping of polymorphims of several enzymes involved in the metabolism of AFB1.
