needed to reduce the risk of developing chronic liver disease Chronic hepatitis B virus (HBV) infection is a major and HCC in chronic HBV carriers. Current established thercause of acute and chronic liver diseases. We have reapy involves treatment with interferon alfa but results in cently described HBV and woodchuck hepatitis virus limited clinical success. The use of inhibitors of the viral poly-(WHV) dominant negative (DN) core mutants that were merase such as nucleoside analogues so far have been unsatcapable of inhibiting wild-type viral replication by 95%.
isfactory because of transient effects on HBV replication, These mutants may represent a potent class of antiviral significant toxicity in clinical trials, or emergence of drug agents that act as ''intracellular immunogens.'' To faciliresistant strains. 4 tate their potential use in animal model systems, we now Characterization of the HBV genome 1 and elucidation of have studied the duck HBV (DHBV) and placed the DN viral protein structure, as well as understanding steps of the mutant constructs in recombinant retroviral and adenoviral expression vectors. Transient expression of the viral life cycle, have led to potential new antiviral strategies. DHBV molecular equivalent of the WHV and HBV DN For example, antisense oligodeoxynucleotides (ODNs) and riconstructs inhibited wild-type DHBV replication by 98%. bozymes have been developed to target various regions of Recombinant retroviral and adenoviral vectors conthe HBV or DHBV genome, and antiviral effects have been taining the HBV and DHBV DN complementary DNAs observed both in vitro and in vivo. [5][6][7][8] The use of ODNs is (cDNAs) were used to transiently and stably transduce complicated by their instability in complex protein mixtures hepatoma-derived cell lines constitutively expressing and by the lack of a cell-specific delivery system to reach the replicating wild-type virus. These investigations show intracellular concentrations required to achieve an antiviral that the DN core mutants were powerful inhibitors of effect. Furthermore, the use of ribozymes has been limited HBV and DHBV replication when delivered intracelluby their inability to efficiently cleave their RNA viral targets, larly and appear as promising antiviral agents for gene sequences within the cell.
therapy of persistent viral infection of the liver. (HEPA-
Another attractive antiviral strategy is based on the design TOLOGY 1996;24:1010-1017.)
and use of dominant negative (DN) polypeptides that are able to interact and/or disrupt the function of their native counterparts. For example, cells overexpressing mutated Hepatitis B virus (HBV) is the prototypic member of the forms of the human immunodeficiency virus (HIV) gag and hepadnavirus family, a group of small enveloped DNA vi-Rev proteins have been able to inhibit ''wild-type'' HIV repliruses. Other members of the family include the woodchuck cation. 9,10 The expression of such mutants has led to the conhepatitis virus (WHV), the ground squirrel hepatitis virus, cept of ''intracellular immunization'' as a potential gene therthe duck hepatitis B virus (DHBV), and the heron hepatitis apeutic approach for chronic HIV infection. 11 virus. Each viral agent will cause acute and chronic infection
In this regard, we have recently identified and functionally of the liver in their respective hosts. 1 In this respect, HBV characterized mutants of the HBV and WHV core protein infection is a significant cause of liver disease in the world that strikingly inhibit wild-type viral replication. Transient today, ranging from acute liver failure to chronic active hepatransfection experiments in hepatoma cell lines show that titis, liver cirrhosis, and hepatocellular carcinoma (HCC). 2,3 Although HBV infection may be prevented by vaccination truncated core proteins fused in frame to the carboxy-termiand passive immunization, no effective therapy is yet availnus of the small s envelope(s) protein are capable of inhibiting able for the more than 300 million individuals worldwide wild-type viral replication by as much as 95%. The antiviral with chronic HBV infection. New antiviral approaches are effect was principally attributable to interference by the mutant core protein with wild-type nucleocapsid assembly. As a consequence of this phenomenon, the process of pregenomic RNA encapsidation was inhibited and viral DNA synthesis Abbreviations: HBV, hepatitis B virus; WHV, woodchuck hepatitis virus; DHBV, duck was terminated. 12 Such mutant core proteins may be among hepatitis B virus; HCC, hepatocellular carcinoma; ODNs, oligodeoxynucleotides; DN, domithe most potent inhibitors of HBV replication and prove usenant negative; HIV, human immunodeficiency virus; LMH, ; aa, amino acid; DMEM, ful as antiviral agents within infected hepatocytes. Moreover, Dulbecco minimal essential medium; Ig, immunoglobulin; PCR, polymerase chain reaction; cDNA, complementary DNA; MOI, multiplicity of infection.