Selenium binding protein 1 (SELENBP1) has been known to be reduced in various types cancer, and epigenetic change is shown to be likely to account for the reduction of expression. With cDNA microarray comparative analysis, we found that SELENBP1 is markedly decreased in hepatitis B virus-X (HBx)-expressing cells. To clarify the effect of HBx on expression, we compared the expression levels of mRNA and protein by semi-quantitative RT-PCR, Northern blot, and Western blot. As expected, expression was shown to be reduced in cells expressing HBx, and reporter gene analysis showed that the promoter is repressed by HBx. In addition, the stepwise deletion of 5' flanking promoter sequences resulted in a gradual decrease in basal promoter activity and inhibition of expression by HBx. Moreover, immunohistochemistry on tissue microarrays containing 60 pairs of human liver tissue showed decreased intensity of SELENBP1 in tumor tissues as compared with their matched non-tumor liver tissues. Taken together, our findings suggest that inhibition of expression by HBx might act as one of the causes in the development of hepatocellular carcinoma caused by HBV infection.