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Address correspondence to Stefan G. Sarafianos, ssarafi@emory.edu. Present address: Karen A. Kirby, Department of Pediatrics, Emory University, Atlanta, Georgia, USA; Stefan G. Sarafianos, Department of Pediatrics, Emory University, Atlanta, Georgia, USA.

Contributor Information: Nathan Sherer, University of Wisconsin-Madison. Priscilla L. Yang, Harvard Medical School.

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Research Funding:

This work was supported in part by grants from the National Institutes of Health (NIH) (AI100890 and AI121315 to S.G.S. and Z.W.) and Trail to a Cure. A.D.H. is supported by NIH AI100890-S1, M.N.P.-C. is supported by the Fulbright Student Program, and J.J.W. is supported by a Life Sciences Fellowship from the University of Missouri.

We also acknowledge partial support from Kumamoto University and the Japan Agency for Medical Research and Development.

Keywords:

  • Science & Technology
  • Life Sciences & Biomedicine
  • Microbiology
  • antiviral agents
  • capsid
  • hepatitis B virus
  • mechanisms of action
  • virology
  • virus-host interactions
  • DNA-DAMAGE RESPONSE
  • HBV CAPSID INHIBITORS
  • IN-VITRO
  • PHENYLPROPENAMIDE DERIVATIVES
  • PRECLINICAL CHARACTERIZATION
  • MURINE CYTOMEGALOVIRUS
  • VIRAL REPLICATION
  • IMAGE-ANALYSIS
  • RESISTANT HBV
  • DELTA-ANTIGEN

The Heteroaryldihydropyrimidine Bay 38-7690 Induces Hepatitis B Virus Core Protein Aggregates Associated with Promyelocytic Leukemia Nuclear Bodies in Infected Cells

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mSphere

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Volume 3, Number 2

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Article | Final Publisher PDF

Abstract:

Heteroaryldihydropyrimidines (HAPs) are compounds that inhibit hepatitis B virus (HBV) replication by modulating viral capsid assembly. While their biophysical effects on capsid assembly in vitro have been previously studied, the effect of HAP treatment on capsid protein (Cp) in individual HBV-infected cells remains unknown. We report here that the HAP Bay 38-7690 promotes aggregation of recombinant Cp in vitro and causes a time- and dose-dependent decrease of Cp in infected cells, consistent with previously studied HAPs. Interestingly, immunofluorescence analysis showed Cp aggregating in nuclear foci of Bay 38-7690-treated infected cells in a time- and dose-dependent manner. We found these foci to be associated with promyelocytic leukemia (PML) nuclear bodies (NBs), which are structures that affect many cellular functions, including DNA damage response, transcription, apoptosis, and antiviral responses. Cp aggregation is not an artifact of the cell system used, as it is observed in HBV-expressing HepAD38 cells, in HepG2 cells transfected with an HBV-expressing plasmid, and in HepG2-NTCP cells infected with HBV. Use of a Cp overexpression vector without HBV sequences shows that aggregation is independent of viral replication, and use of an HBV-expressing plasmid harboring a HAP resistance mutation in Cp abrogated the aggregation, demonstrating that the effect is due to direct compound-Cp interactions. These studies provide novel insight into the effects of HAP-based treatment at a single-cell level.IMPORTANCE Despite the availability of effective vaccines and treatments, HBV remains a significant global health concern, with more than 240 million individuals chronically infected. Current treatments are highly effective at controlling viral replication and disease progression but rarely cure infections. Therefore, much emphasis is being placed on finding therapeutics with new drug targets, such as viral gene expression, covalently closed circular DNA formation and stability, capsid formation, and host immune modulators, with the ultimate goal of an HBV cure. Understanding the mechanisms by which novel antiviral agents act will be imperative for the development of curative HBV therapies.

Copyright information:

© 2018 Huber et al.

This is an Open Access work distributed under the terms of the Creative Commons Attribution 4.0 International License (https://creativecommons.org/licenses/by/4.0/).
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