Abstract
ABSTRACTThe hepatitis B virus (HBV) core protein serves multiple essential functions in the viral life cycle, and antiviral agents that target the core protein are being developed. Capsid assembly modulators (CAMs) are compounds that target core and misdirect capsid assembly, resulting in the suppression of HBV replication and virion production. Besides HBV DNA, circulating HBV RNA has been detected in patient serum and can be associated with the treatment response. Here we studied the effect of HBV CAMs on the production of extracellular HBV RNA using infected HepaRG cells and primary human hepatocytes. Representative compounds from the sulfonamide carboxamide and heteroaryldihydropyrimidine series of CAMs were evaluated and compared to nucleos(t)ide analogs as inhibitors of the viral polymerase. The results showed that CAMs blocked extracellular HBV RNA with efficiencies similar to those with which they blocked pregenomic RNA (pgRNA) encapsidation, HBV DNA replication, and Dane particle production. Nucleos(t)ide analogs inhibited viral replication and virion production but not encapsidation or production of extracellular HBV RNA. Profiling of HBV RNA from both culture supernatants and patient serum showed that extracellular viral RNA consisted of pgRNA and spliced pgRNA variants with an internal deletion(s) but still retained the sequences at both the 5′ and 3′ ends. Similar variants were detected in the supernatants of infected cells with and without nucleos(t)ide analog treatment. Overall, our data demonstrate that HBV CAMs represent direct antiviral agents with a profile differentiated from that of nucleos(t)ide analogs, including the inhibition of extracellular pgRNA and spliced pgRNA.
Highlights
The hepatitis B virus (HBV) core protein serves multiple essential functions in the viral life cycle, and antiviral agents that target the core protein are being developed
If the extracellular RNA particles were produced from assembled capsids, the assembly modulator mechanism of action would prevent the generation of such RNA particles through inhibition of RNA encapsidation
The HBV core protein is a promising antiviral target for the treatment of hepatitis B, and small-molecule compounds targeting core are currently being developed as a new class of direct-acting antiviral agents
Summary
The hepatitis B virus (HBV) core protein serves multiple essential functions in the viral life cycle, and antiviral agents that target the core protein are being developed. We studied the effect of HBV CAMs on the production of extracellular HBV RNA using infected HepaRG cells and primary human hepatocytes. The results showed that CAMs blocked extracellular HBV RNA with efficiencies similar to those with which they blocked pregenomic RNA (pgRNA) encapsidation, HBV DNA replication, and Dane particle production. As different chemical classes of CAMs are currently being developed as new antiviral therapies for the treatment of chronic hepatitis B, it is important to profile their activities against HBV markers during different stages of the viral life cycle. We investigated the effect of CAMs from the sulfonamide carboxamide series (represented by NVR 3-1983) and the HAP series (represented by BAY 41-4109) on the production of extracellular HBV RNA using naturally infected HepaRG cells and primary human hepatocytes. Our study further profiled the nature of extracellular HBV RNA from the supernatants of infected cells treated or not treated with nucleoside analogs and from hepatitis B virus genotype A to D-infected patient serum
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