Virtual Screening of Hepatitis B Virus Pre-Genomic RNA as a Novel Therapeutic Target

Abstract

The global burden imposed by hepatitis B virus (HBV) infection necessitates the discovery and design of novel antiviral drugs to complement existing treatments. One attractive and underexploited therapeutic target is ε, an ~85-nucleotide (nt) cis-acting regulatory stem-loop RNA located at the 3′- and 5′-ends of the pre-genomic RNA (pgRNA). Binding of the 5′-end ε to the viral polymerase protein (P) triggers two early events in HBV replication: pgRNA and P packaging and reverse transcription. Our recent solution nuclear magnetic resonance spectroscopy structure of ε permits structure-informed drug discovery efforts that are currently lacking for P. Here, we employ a virtual screen against ε using a Food and Drug Administration (FDA)-approved compound library, followed by in vitro binding assays. This approach revealed that the anti-hepatitis C virus drug Daclatasvir is a selective ε-targeting ligand. Additional molecular dynamics simulations demonstrated that Daclatasvir targets ε at its flexible 6-nt priming loop (PL) bulge and modulates its dynamics. Given the functional importance of the PL, our work supports the notion that targeting ε dynamics may be an effective anti-HBV therapeutic strategy.

Hepatitis B Virus Epsilon (ε) RNA Element: Dynamic Regulator of Viral Replication and Attractive Therapeutic Target

“…As a second method to test our structure-informed hypothesis, we employed a structure-based virtual screening (VS) approach [ 219 ]. Computational docking can provide complementary data and corroborating evidence to experimental binding assays.…”

“…We then selected a 1604-compound FDA-approved library curated on the ZINC15 database [ 229 , 230 , 231 ]. With our receptor and library set, we executed VS to identify FDA-approved drugs that could be repurposed as anti-HBV therapeutics [ 219 ]. We implemented selection criteria based on affinity, commercial availability, drug-like properties, and docking site [ 219 ].…”

“…With our receptor and library set, we executed VS to identify FDA-approved drugs that could be repurposed as anti-HBV therapeutics [ 219 ]. We implemented selection criteria based on affinity, commercial availability, drug-like properties, and docking site [ 219 ]. Ultimately, we selected 12 potential lead compounds ( Figure 12 A) with diverse chemotypes and uses that could now be experimentally verified [ 219 ].…”

“…Similar to Raloxifene, compound insolubility precluded the use of NMR to acquire a Daclatasvir-bound full-length ε structure, so we once again employed computational docking and MD simulations. Our docking analysis revealed that Daclatasvir selectively targets the ε PL, with its core wedged between nucleotides U15 and U17–C19, and also contacting the adjacent A20–G22 and U47–G51 and C5 and A6 in the LH ( Figure 12 C) [ 219 ]. Interestingly, akin to Raloxifene, MD simulations reveal that Daclatasvir binding modulates ε dynamics, albeit in a different manner.…”

“…Interestingly, akin to Raloxifene, MD simulations reveal that Daclatasvir binding modulates ε dynamics, albeit in a different manner. Daclatasvir binding rigidifies the dynamics of nucleotides U17–C19 but increases the conformational variety of U15 [ 219 ].…”