β-Thujaplicinol inhibits hepatitis B virus replication by blocking the viral ribonuclease H activity

• β-Thujaplicinol inhibits HBV RNAseH activity with low micromolar IC50 values.
• β-Thujaplicinol inhibits HBV replication in culture by blocking the viral RNAseH.
• RNAseH activity of at least 3 of HBV’s 8 genotypes is sensitive to β-thujaplicinol.
• Derivatization of β-thujaplicinol to increase its therapeutic index is warranted.

Hepatitis B virus (HBV) is a hepatotropic DNA virus that replicates by reverse transcription. It chronically infects >350 million people and kills about 1 million patients annually. Therapy primarily employs nucleos(t)ide analogs that suppress viral DNA synthesis by the viral reverse transcriptase very well but that rarely cure the infection, so additional therapies are needed. Reverse transcription requires the viral ribonuclease H (RNAseH) to destroy the viral RNA after it has been copied into DNA. We recently produced active recombinant HBV RNAseH and demonstrated that Human Immunodeficiency Virus (HIV) RNAseH antagonists could inhibit the HBV enzyme at a high frequency. Here, we extended these results to β-thujaplicinol, a hydroxylated tropolone which inhibits the HIV RNAseH. β-Thujaplicinol inhibited RNAseHs from HBV genotype D and H in biochemical assays with IC50 values of 5.9 ± 0.7 and 2.3 ± 1.7 μM, respectively. It blocked replication of HBV genotypes A and D in culture by inhibiting the RNAseH activity with an estimated EC50 of ∼5 μM and a CC50 of 10.1 ± 1.7 μM. Activity of β-thujaplicinol against RNAseH sequences from multiple HBV genotypes implies that if chemical derivatives of β-thujaplicinol with improved efficacy and reduced toxicity can be identified, they would have promise as anti-HBV agents.