Objective: Hepatitis B virus (HBV) is a major etiological agent of hepatocellular carcinoma (HCC). The x gene of HBV genome (HBVx) is considered as its oncogene. In order to assess the extent of its involvement in hepatocarcinogenesis, both HBsAg sero-positive and mainly sero-negative HCC were searched for the presence, expression and mutation status of the x gene of HBV, as well as its association with 249 codon hot spot mutation of p53 gene.
Methods: Using PCR, RT-PCR, PCR-RFLP and DNA sequencing, studies were more focused on 25 HBsAg sero-negative, pathologically diagnosed HCC patients operated during 1991 to 1996 mainly in Qidong and also in Beijing.
Results: The x gene sequence of HBV was found by PCR without exception in all 25 seronegative HCC DNA (100% 25/25) and also in all 19 seropositive counterparts. The RNA messages of HBVx gene were found in all 8 HCC patients randomly selected from the seronegative group. Sequence analysis of the HBVx gene showed the presence of missense mutation in the 130 and 131 codon in 4 of 6 samples studied. Using PCR-RFLP, missense mutation of the 249 codon was identified in 57% (12/21) of all HBsAg negative cases from Qidong. No such mutations were found in the 4 Beijing counterparts.
Conclusion: HBVx gene sequence was universally present in HBsAg negative HCC samples of Qidog studied, indicating the important role of HBVx gene in hepatocarcinogenesis of the high incidence area. The close association of the hotspot mutation of p53 gene in Qidong HCC with the presence of HBVx gene sequence suggests that such mutation is the molecular footprint of the combined effect of aflatoxin B1 exposure and HBVx gene product.