ABSTRACT FINAL ID: 922
TITLE: Preclinical Profile of Potent Second Generation CpAMs Capable of Inhibiting the Generation of HBsAg, HBeAg, pgRNA and cccDNA in HBV Infected Cells
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ABSTRACT BODY:
Background: Clinical cure remains elusive in chronic HBV (CHB) patients, despite prolonged treatment periods with current available therapies. Core Protein Allosteric Modulators (CpAMs) are a novel class of direct acting antivirals with the potential to help improve cure rates by targeting multiple aspects of the viral life cycle. Here we characterize potent 2nd generation CpAMs that exhibit favorable drug properties and inhibit cccDNA establishment and replenishment via multiple mechanisms.
Methods: CpAM antiviral activities were determined using the HepAD38 cell line (Gt D), and infected HepG2-NTCP cells and Primary Human Hepatocytes (PHH). Pan-genotypic activity was established using transient transfection assays or HBV stable cell lines. Combination studies were performed using a range of inhibitory concentrations in HepAD38 cells. HBV DNA levels were quantified by Taqman PCR (qPCR) using primers and a probe specific to the HBV core gene. HBeAg and HBsAg were quantified by ELISA. HBV total RNA/encapsidated pgRNA, Cp, capsids, and capsid-associated core DNA were detected by Northern Blot or b-DNA, Western Blot, Enzyme Immunoassay (EIA), and Southern Blot, respectively, as previously described.
Results: Representative 2nd generation CpAMs exhibited potent inhibition of viral DNA replication in HepAD38 cells (EC50s <30 nM), and reductions in HBeAg, HBsAg and pgRNA in infected HepG2-NTCP cells and PHH (EC50s <300 nM). Antiviral activity extended to other major HBV genotypes in stably or transiently transfected HepG2 cells. No significant activity was exhibited against a panel of other viruses (EC50s >10 µM) or cell lines in cytotoxicity assays (CC50s >10 µM), indicating that the compounds are selective inhibitors of HBV. Lead molecules possess promising physical properties, low drug-drug interaction potential and favorable PK profiles in multiple species. Mechanism of action studies suggest that the enhanced potency of these new CpAMs is likely due to inhibition of multiple steps of the HBV life cycle, including acceleration of aberrant capsid assembly, disruption of pre-formed capsids and premature disassembly during trafficking of incoming mature capsids and rcDNA containing capsids to the nucleus during infection.
Conclusions: Novel 2nd generation pan-genotypic CpAMs demonstrate potent HBV antiviral activity, particularly against cccDNA generation and its surrogate markers. They appear to work via newly identified mechanisms and possess promising drugable properties. The potency and favorable preclinical profile support advancement of this next generation of CpAMs into clinical development.作者: antiHBVren 时间: 2018-2-24 15:23
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