HBV Capsid Assembly Modulators, but not Nucleoside Analogs, Inhibit the Production of Extracellular Pregenomic RNA and Spliced RNA Variants [PublishAheadOfPrint]

The Hepatitis B virus (HBV) core protein serves multiple essential functions in the viral life cycle and is being developed as a target for antiviral agents. 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 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. Results showed that CAMs blocked extracellular HBV RNA with similar efficiencies as pgRNA encapsidation, HBV DNA replication, and Dane particles 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 cultured supernatants and patient serum showed that extracellular viral RNA consisted of pgRNA and spliced pgRNA variants with internal deletion(s) but still retained the sequences at both 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 demonstrates that HBV CAMs represent direct antiviral agents with a differentiated profile as compared to nucleos(t)ide analogs, including the inhibition of extracellular pgRNA and spliced pgRNA.

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