Efficacy and cytotoxicity in cell culture of novel α-hydroxytropolone inhibitors of hepatitis B virus ribonuclease H

Abstract

Chronic Hepatitis B virus (HBV) infection is a major worldwide public health problem. Current direct-acting anti-HBV drugs target the HBV DNA polymerase activity, but the equally essential viral ribonuclease H (RNaseH) activity is unexploited as a drug target. Previously, we reported that α–hydroxytropolone compounds can inhibit the HBV RNaseH and block viral replication. Subsequently, we found that our biochemical RNaseH assay underreports efficacy of the α-hydroxytropolones against HBV replication. Therefore, we conducted a structure-activity analysis of 59 troponoids against HBV replication in cell culture. These studies revealed that antiviral efficacy is diminished by larger substitutions on the tropolone ring, identified key components in the substitutions needed for high efficacy, and revealed that cytotoxicity correlates with increased lipophilicity of the α-hydroxytropolones. These data provide key guidance for further optimization of the α-hydroxytropolone scaffold as novel HBV RNaseH inhibitors.

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 ].…”

Meeting report: 31st International Conference on Antiviral Research

“…Tiffany Edwards of Saint Louis University, MO, USA, reported the development of HBV RNaseH inhibitors belonging to either the N-hydroxy-isoquinolinedione (HID) and N-hydroxy-pyridinedione (HPD) families. This work follows published efforts of this team to improve the efficacy and characterize the anti-RNAse activity of these compounds in vitro (Edwards et al, 2017;Lomonosova et al, 2017). As a recall, RNAse activity in HBV polymerase is crucial for the reverse transcription of pregenomic RNA into rcDNA within nucleocapsids.…”

Endonuclease Activity Inhibition of the NS1 Protein of Parvovirus B19 as a Novel Target for Antiviral Drug Development

“…To identify small-molecule compounds that can inhibit the NS1N-mediated Ori cleavage, we used 96 compounds (see Table S1 in the supplemental material), which we have used previously to screen inhibitors for the RNase H nuclease activity of the human hepatitis B virus (HBV) polymerase (43)(44)(45)(46)(47), to test them for inhibitory effects on the NS1N-mediated Ori cleavage using a 32 P-labeled Ori30 template. We first tested them at 100 M and found that 25 compounds inhibited the nicking of NS1N by greater than 80%, as shown in Fig.…”

“…In the current study, we identified 8 compounds that exhibit inhibition of NS1 nicking of the B19V Ori from the screening of 96 small-molecule compounds, which have been used to screen inhibitors of the RNase H nuclease activity of the HBV polymerase (43)(44)(45)(46)(47), using an in vitro nicking assay. We chose three compounds, compounds 7, 135, and 201, that share a similar flavonoid chemical structure for detailed study.…”