Although epidemiologic studies have clearly demonstrated the importance of the hepatitis B virus in the genesis of hepatocellular carcinoma, the molecular basis for this tumorigenic effect is still under debate. The finding of hepatitis B virus DNA integration into human liver DNA in many cases of hepatocellular carcinoma suggested that these integrated viral sequences may be involved in liver carcinogenesis. In an attempt to clarify this point, we studied 9 tumors which developed in non cirrhotic livers. All tumors contained viral integrations (ranging from 1 to 6 different integrants) and 4 showed abnormal hepatitis B virus mRNA (2.3 to 7.5 kilobases long). The analysis of the corresponding cDNAs revealed the existence of hybrid transcripts containing both genomic and viral sequences. In 2 cases, the viral-host junctions were mapped within the cohesive-end region of the hepatitis B virus genome leading to the production of a transcript encoding a 3' truncated X protein. In another case, the cellular sequences present in the co-transcript were located in 5' with respect to the hepatitis B virus sequences. This observation strongly suggests that, in this patient, integration took place near a cellular gene. Further analysis of this integrant should help in identifying the putative gene and its application in the development of the tumor. We conclude that the study of abnormal hepatitis B virus transcripts in liver tumors provides a positive approach to study the direct role of HBV in carcinogenesis as an insertional mutagen.