THE VIRUS OF HEPATITIS B

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Hepatitis B virus (HBV) is an important cause of liver disease including liver cancer in man. It is a member of a family of viruses which have interesting structure, mode of replication and biological properties. We propose to study the role of HBV in liver cancer by analyzing integrated viral DNA sequences with flanking cellular DNA cloned from two hepatocellular carcinomas (HCC), each with single integration sites. We will also further investigate NIH 3T3 cell transformation observed with a cloned HBV DNA dimer, an 1850 bp BamHI fragment of HBV DNA and DNA from HCC. We will also investigate the effect of liver regeneration on viral integration and HCC formation. We will further characterize high molecular weight HBV DNA forms (5 and 9 kd) found in virions from plasma. A final goal is to investigate a viral vaccine strategy by which relevant viral antigens are expressed by enteric bacterial strains which result in immune responses but not disease when live bacteria are ingested by mouth. Although this approach might offer a particular advantage for enteric pathogens such as rotavirus for which local enteric immunity is important, the approach could be equally effective for other agents such as HBV for which only systemic immunity is important. We have introduced genes for hepatitis B surface (HBsAg) and core (HBcAg) antigens and rotavirus VP7 into plasmid expression vectors and have achieved expression of HBsAg and rotavirus VP7 as fusion proteins and HBcAg as a discrete polypeptide in E. coli. We propose to optimize expression of these antigens in E. coli; introduce the hybrid plasmids into Salmonella typhi strain Ty 21a and S. typhimurine 1479 (mutants with attenuated virulence and successfully used as live, oral Salmonella vaccines); and test the transformed Salmonella for viral antigen expression and ability to induce immune responses in mice. The long-term goal is live oral vaccines for rotavirus and HBV in man.