The Role of the Woodchuck Model in the Treatment of Hepatitis B Virus Infection

Kulkarni, Ketan; Jacobson, Ira M.; Tennant, Bud C.

doi:10.1016/j.cld.2007.08.012

Cited by 16 publications

(14 citation statements)

References 76 publications

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“…This observation indicates a wide spectrum for these compounds in hepadnavirus cccDNA intervention and also provides an opportunity to evaluate their antiviral activity (alone or in combination with nucleos[t]ide analogues) in the duck model. Other hepadnavirus animal models, including humanized uPA/ SCID mice and woodchucks (4,21), are also available for in vivo testing of DSS compounds. First, however, the potency of the hit compounds needs to be improved.…”

Section: Discussionmentioning

confidence: 99%

Identification of Disubstituted Sulfonamide Compounds as Specific Inhibitors of Hepatitis B Virus Covalently Closed Circular DNA Formation

Cai

Mills

et al. 2012

Antimicrob Agents Chemother

203

170

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cHepatitis B virus (HBV) covalently closed circular DNA (cccDNA) plays a central role in viral infection and persistence and is the basis for viral rebound after the cessation of therapy, as well as the elusiveness of a cure even after extended treatment. Therefore, there is an urgent need for the development of novel therapeutic agents that directly target cccDNA formation and maintenance. By employing an innovative cell-based cccDNA assay in which secreted HBV e antigen is a cccDNA-dependent surrogate, we screened an in-house small-molecule library consisting of 85,000 drug-like compounds. Two structurally related disubstituted sulfonamides (DSS), termed CCC-0975 and CCC-0346, emerged and were confirmed as inhibitors of cccDNA production, with low micromolar 50% effective concentrations (EC 50 s) in cell culture. Further mechanistic studies demonstrated that DSS compound treatment neither directly inhibited HBV DNA replication in cell culture nor reduced viral polymerase activity in the in vitro endogenous polymerase assay but synchronously reduced the levels of HBV cccDNA and its putative precursor, deproteinized relaxed circular DNA (DP-rcDNA). However, DSS compounds did not promote the intracellular decay of HBV DPrcDNA and cccDNA, suggesting that the compounds interfere primarily with rcDNA conversion into cccDNA. In addition, we demonstrated that CCC-0975 was able to reduce cccDNA biosynthesis in duck HBV-infected primary duck hepatocytes. This is the first attempt, to our knowledge, to identify small molecules that target cccDNA formation, and DSS compounds thus potentially serve as proof-of-concept drug candidates for development into therapeutics to eliminate cccDNA from chronic HBV infection. It is estimated that 2 billion people worldwide have been infected with hepatitis B virus (HBV). Although most adulthood infections are transient, approximately 5 to 10% of infected adults and over 90% of infected neonates fail to mount a sufficient immune response to clear the virus and develop a life-long chronic infection (23,27). Chronic hepatitis B is currently a substantial public health burden, affecting approximately 350 million individuals worldwide. These patients have an elevated risk of liver cirrhosis, hepatocellular carcinoma, and other severe clinical sequelae (1, 23). It is therefore a global health priority to cure chronic HBV infection and prevent its dire consequences.HBV is a noncytopathic, liver-tropic DNA virus belonging to the Hepadnaviridae family. Upon infection, the viral genomic relaxed circular DNA (rcDNA) is transported into the cell nucleus and converted into episomal covalently closed circular DNA (cccDNA), which serves as the transcription template for the viral mRNAs. After transcription and nuclear export, cytoplasmic viral pregenomic RNA (pgRNA) is assembled with HBV polymerase and capsid proteins to form the nucleocapsid, inside which polymerase-catalyzed reverse transcription yields minus-strand DNA, which is subsequently copied into plus-strand DNA to form the progeny rcDNA g...

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Section: Discussionmentioning

confidence: 99%

Identification of Disubstituted Sulfonamide Compounds as Specific Inhibitors of Hepatitis B Virus Covalently Closed Circular DNA Formation

Cai

Mills

et al. 2012

Antimicrob Agents Chemother

203

170

View full text Add to dashboard Cite

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“…Efforts to produce better treatments remain an active goal of HBV research. Woodchuck hepatitis virus (WHV) and the woodchuck model system have been key to understanding natural viral infection and to developing antiviral therapies (9)(10)(11). In this regard, a better understanding of the WHV model system is desirable.…”

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confidence: 99%

Structurally Similar Woodchuck and Human Hepadnavirus Core Proteins Have Distinctly Different Temperature Dependences of Assembly

Kukreja

Wang

Pierson

et al. 2014

J Virol

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Woodchuck hepatitis virus (WHV), a close relative of human hepatitis B virus (HBV), has been a key model for disease progression and clinical studies. Sequences of the assembly domain of WHV and HBV core proteins (wCp149 and hCp149, respectively) have 65% identity, suggesting similar assembly behaviors. We report a cryo-electron microscopy (cryo-EM) structure of the WHV capsid at nanometer resolution and characterization of wCp149 assembly. At this resolution, the T‫4؍‬ capsid structures of WHV and HBV are practically identical. In contrast to their structural similarity, wCp149 demonstrates enhanced assembly kinetics and stronger dimer-dimer interactions than hCp149: at 23°C and at 100 mM ionic strength, the pseudocritical concentrations of assembly of wCp149 and hCp149 are 1.8 M and 43.3 M, respectively. Transmission electron microscopy reveals that wCp149 assembles into predominantly T‫4؍‬ capsids with a sizeable population of larger, nonicosahedral structures. Charge detection mass spectrometry indicates that T‫3؍‬ particles are extremely rare compared to the ϳ5% observed in hCp149 reactions. Unlike hCp149, wCp149 capsid assembly is favorable over a temperature range of 4°C to 37°C; van't Hoff analyses relate the differences in temperature dependence to the high positive values for heat capacity, enthalpy, and entropy of wCp149 assembly. Because the final capsids are so similar, these findings suggest that free wCp149 and hCp149 undergo different structural transitions leading to assembly. The difference in the temperature dependence of wCp149 assembly may be related to the temperature range of its hibernating host. IMPORTANCEIn this paper, we present a cryo-EM structure of a WHV capsid showing its similarity to HBV. We then observe that the assembly properties of the two homologous proteins are very different. Unlike human HBV, the capsid protein of WHV has evolved to function in a nonhomeostatic environment. These studies yield insight into the interplay between core protein self-assembly and the host environment, which may be particularly relevant to plant viruses and viruses with zoonotic cycles involving insect vectors.

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“…This finding is consistent with the low level of expression of the WHV x gene in the woodchuck liver during natural infection (27). The detection of WHV proteins has failed so far, as no suitable antibodies were available (15,25,28,29).…”

Section: Resultsmentioning

confidence: 99%

Novel Woodchuck Hepatitis Virus (WHV) Transgene Mouse Models Show Sex-Dependent WHV Replicative Activity and Development of Spontaneous Immune Responses to WHV Proteins

Zhao

Ma²,

Zhang³

et al. 2014

J Virol

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HBV has a very narrow host range and can infect only humans and higher primates such as chimpanzees (3, 4). However, experiments using chimpanzees are costly and require both scientific and ethical justification. HBV transgenic (Tg) mice have allowed examination of the influence of viral and host factors on HBV pathogenesis and replication and assessment of the antiviral potential of pharmacological agents (5). For example, the transfer of HBsAg-specific CD8 ϩ T cells into these mice led to the inhibition of HBV replication in the liver by a noncytolytic mechanism (6), leading to the important hypothesis that the antiviral cytokines IFN-␥ and tumor necrosis factor alpha are major mediators of noncytolytic inhibition of HBV replication (7). Furthermore, activation of innate immune responses in HBV Tg mice by Toll-like receptor (TLR) ligands also inhibits HBV replication (8). Thus, the HBV Tg mouse model was and remains a useful research tool to study HBV infection. However, as HBV Tg mice are immunologically tolerant to HBV proteins (5, 9), obvious disease-related phenotypes have not been observed. On the other hand, HBsAg is supposed to play an important role in HBV persistence and hepatocarcinogenesis (10). The presence of HBsAg may inhibit host immune responses and facilitate HBV persistence (11, 12). Therefore, transgenic mice replicating HBV but lacking small HBs expression represent an interesting model for studies of the role of HBsAg in HBV persistence, hepatocarcinogenesis, and antiviral immune responses (13).Woodchuck hepatitis virus (WHV) is a member of the Hepadnaviridae family and was discovered in 1978 (14). WHV causes acute and chronic infections in woodchucks (Marmota monax) similar to HBV infections in humans. The woodchuck model has been proven to be an informative model for studies of hepadnaviral infection and pathogenesis and for the screening and development of antiviral drugs (15). Despite the establishment of immunological assays and tools for the woodchuck model during the last years, immunological studies in outbred and immunogenetically uncharacterized woodchucks are still difficult (16,17). Therefore, it was desirable to have a WHV Tg mouse model that would allow the determination of immunological parameters and complement infection experiments in woodchucks.In the present study, we report the establishment of two WHV Tg mouse strains, one with a wild-type WHV genome and the other with a WHV genome harboring artificially introduced stop codons in the coding region of WHV surface anti-

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The Role of the Woodchuck Model in the Treatment of Hepatitis B Virus Infection

Cited by 16 publications

References 76 publications

Identification of Disubstituted Sulfonamide Compounds as Specific Inhibitors of Hepatitis B Virus Covalently Closed Circular DNA Formation

Identification of Disubstituted Sulfonamide Compounds as Specific Inhibitors of Hepatitis B Virus Covalently Closed Circular DNA Formation

Structurally Similar Woodchuck and Human Hepadnavirus Core Proteins Have Distinctly Different Temperature Dependences of Assembly

Novel Woodchuck Hepatitis Virus (WHV) Transgene Mouse Models Show Sex-Dependent WHV Replicative Activity and Development of Spontaneous Immune Responses to WHV Proteins

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