Synergistic Interactions between Hepatitis B Virus RNase H Antagonists and Other Inhibitors

Abstract

Combination therapies are standard for management of human immunodeficiency virus (HIV) and hepatitis C virus (HCV) infections; however, no such therapies are established for human hepatitis B virus (HBV). Recently, we identified several promising inhibitors of HBV RNase H (here simply RNase H) activity that have significant activity against viral replication in vitro. Here, we investigated the in vitro antiviral efficacy of combinations of two RNase H inhibitors with the current anti-HBV drug nucleoside analog lamivudine, with HAP12, an experimental core protein allosteric modulator, and with each other. Anti-HBV activities of the compounds were tested in a HepG2-derived cell line by monitoring intracellular core particle DNA levels, and cytotoxicity was assessed by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay. The antiviral efficiencies of the drug combinations were evaluated using the median-effect equation derived from the mass-action law principle and combination index theorem of Chou and Talalay. We found that combinations of two RNase H inhibitors from different chemical classes were synergistic with lamivudine against HBV DNA synthesis. Significant synergism was also observed for the combination of the two RNase H inhibitors. Combinations of RNase H inhibitors with HAP12 had additive antiviral effects. Enhanced cytotoxicity was not observed in the combination experiments. Because of these synergistic and additive effects, the antiviral activity of combinations of RNase H inhibitors with drugs that act by two different mechanisms and with each other can be achieved by administering the compounds in combination at doses below the respective single drug doses.

3,7-Dihydroxytropolones Inhibit Initiation of Hepatitis B Virus Minus-Strand DNA Synthesis

“…Figiel et al have demonstrated that, under conditions where the DNA polymerase active site of HIV-1 RT is covalently liked to its substrate, nucleic acid can be simultaneously accessed by the RNA polymerase and RNase H active centers, indicating an important degree of coordination between its synthetic and hydrolytic activities [ 24 ]. Using this precedent, we speculated that ligand binding at the HBV RNase H domain might allosterically modulate P protein DNA polymerase activity, since we and others have demonstrated that α-HTs inhibit HBV RNase H activity in vitro, and virus replication in culture [ 25 , 26 , 27 ]. In addition, Didierjean et al have reported that DNA polymerase activity of HIV-1 RT can be inhibited by structurally related 3,7-dHTs [ 28 ].…”

Recent Advances in Understanding, Diagnosing, and Treating Hepatitis B Virus Infection

“…Several other studies demonstrated the potential of inhibiting the viral ribonuclease H strategy as a promising alternative for developing new anti-HBV drugs. Combinations of RNase H inhibitors with other antiviral agents had additive antiviral effects without enhanced cytotoxicity [ 159 ].…”

Shedding light on RNaseH: a promising target for hepatitis B virus (HBV)

“…The RNaseH inhibitors repress HBV RNaseHs from three genotypes, indicating that HBV's high genetic diversity is unlikely to complicate drug development [13]. The inhibitors are synergistic with the nucleoside analog drug Lamivudine and additive with the experimental HBV capsid protein assembly modifier Hap12 without enhancing cytotoxicity [10,14]. Two RNaseH inhibitors, an HPD (208) and an αHT (110), can significantly suppress HBV replication in chimeric mice carrying humanized livers [15].…”

“…2). This is because HBV RNaseH inhibitors work synergistically with nucleoside analogs and additively with capsid assembly modifiers, are insensitive to HBV's genetic variation, and different chemotypes of RNaseH inhibitors can be synergistic with each other [10,14]. Developing such a battery of anti-HBV drugs may finally achieve the long-sought goal of achieving a functional cure for chronic HBV infections.…”