Hepatitis B virus (HBV) is an enveloped dsDNA virus with a ssRNA intermediate form. Correct assembly of the icosahedral core of HBV is required for replication; reverse transcription of the RNA pre-genome takes place within the completed capsid, the protein shell of the core. Current anti-HBV therapeutics are interferon and nucleoside analog inhibitors of HBV reverse transcriptase (Pol). Both approaches have limited efficacy and can be expensive. Like any monotherapy, Pol inhibitors select for resistant mutants; some of these mutations escape the vaccine because Pol and surface protein genes overlap. We advocate capsid assembly as a complementary target for antiviral therapy. Assembly is critical to HBV replication and is unique to the virus (there are no cellular homologs to capsid protein). The HBV capsid is constructed from 120 protein dimers. In vitro, assembly can be induced by a number of small molecules and salts. We have found that HBV assembly is allosterically regulated: capsid protein has assembly-active and -inactive conformations. Consequently, small molecules that favor the inactive form will inhibit assembly; molecules that favor the active form will accelerate the process. We have found that some molecules misdirect capsid assembly, presumably by distorting the dimer geometry, giving rise to the rapid production of misshapen and nonfunctional structures. At this time, we have a small selection of molecules in each category. In this proposal we describe a strategy for developing assembly-directed antivirals. Starting with the pre-existing leads, we will investigate the physical and structural basis for altered assembly by the most active molecules. Using this information, we will then take advantage of recent advances in the quantification of assembly to screen candidate small-molecule libraries for enhanced activity in assembly acceleration, inhibition, and misdirection. The effects of these molecules will also be tested in cultured cells that express HBV. More than 350 million individuals suffer from chronic infection with hepatitis B virus (HBV), including more than 1.25 million Americans. Worldwide, HBV will contribute to 1 million deaths this year. Current antiviral strategies focus on virus enzymes. We propose a new strategy for developing antiviral molecules that target virus assembly.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
7R01AI067417-04
Application #
7739772
Study Section
Special Emphasis Panel (ZRG1-DDR (01))
Program Officer
Berard, Diana S
Project Start
2005-12-06
Project End
2010-11-30
Budget Start
2008-08-01
Budget End
2008-11-30
Support Year
4
Fiscal Year
2008
Total Cost
$233,403
Indirect Cost
Name
Indiana University Bloomington
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
006046700
City
Bloomington
State
IN
Country
United States
Zip Code
47401
Lomonosova, Elena; Zlotnick, Adam; Tavis, John E (2017) Synergistic Interactions between Hepatitis B Virus RNase H Antagonists and Other Inhibitors. Antimicrob Agents Chemother 61:
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Venkatakrishnan, Balasubramanian; Katen, Sarah P; Francis, Samson et al. (2016) Hepatitis B Virus Capsids Have Diverse Structural Responses to Small-Molecule Ligands Bound to the Heteroaryldihydropyrimidine Pocket. J Virol 90:3994-4004
Singharoy, Abhishek; Venkatakrishnan, Balasubramanian; Liu, Yun et al. (2015) Macromolecular Crystallography for Synthetic Abiological Molecules: Combining xMDFF and PHENIX for Structure Determination of Cyanostar Macrocycles. J Am Chem Soc 137:8810-8
Zlotnick, Adam; Venkatakrishnan, Balasubramanian; Tan, Zhenning et al. (2015) Core protein: A pleiotropic keystone in the HBV lifecycle. Antiviral Res 121:82-93
Tan, Zhenning; Pionek, Karolyn; Unchwaniwala, Nuruddin et al. (2015) The interface between hepatitis B virus capsid proteins affects self-assembly, pregenomic RNA packaging, and reverse transcription. J Virol 89:3275-84
Wang, Joseph Che-Yen; Nickens, David G; Lentz, Thomas B et al. (2014) Encapsidated hepatitis B virus reverse transcriptase is poised on an ordered RNA lattice. Proc Natl Acad Sci U S A 111:11329-34
Zlotnick, Adam; Tan, Zhenning; Selzer, Lisa (2013) One protein, at least three structures, and many functions. Structure 21:6-8
Katen, Sarah P; Tan, Zhenning; Chirapu, Srinivas Reddy et al. (2013) Assembly-directed antivirals differentially bind quasiequivalent pockets to modify hepatitis B virus capsid tertiary and quaternary structure. Structure 21:1406-16
Li, Lichun; Chirapu, Srinivas Reddy; Finn, M G et al. (2013) Phase diagrams map the properties of antiviral agents directed against hepatitis B virus core assembly. Antimicrob Agents Chemother 57:1505-8

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