Screening for inhibitor of episomal DNA identified dicumarol as a hepatitis B virus inhibitor

Fumihiko Takeuchi; Sotaro Ikeda; Yuta Tsukamoto; Yoshikazu Iwasawa; Chen Qihao; Yukie Otakaki; Ouda Ryota; Wan Ling Yao; Ryo Narita; Hijikata Makoto; Koichi Watashi; Takaji Wakita; Koh Takeuchi; Kazuaki Chayama; Amane Kogure; Hiroki Kato; Takashi Fujita

doi:10.1371/journal.pone.0212233

Screening for inhibitor of episomal DNA identified dicumarol as a hepatitis B virus inhibitor

Fumihiko Takeuchi, Sotaro Ikeda, Yuta Tsukamoto, Yoshikazu Iwasawa, Chen Qihao, Yukie Otakaki, Ouda Ryota, Wan Ling Yao, Ryo Narita, Hijikata Makoto, Koichi Watashi, Takaji Wakita, Koh Takeuchi, Kazuaki Chayama, Amane Kogure, Hiroki Kato, Takashi Fujita^*

^*Corresponding author for this work

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaper › Journal article › Research › peer-review

Abstract

Currently, there is no available therapy to eradicate hepatitis B virus (HBV) in chronically infected individuals. This is due to the difficulty in eliminating viral covalently closed circular (ccc) DNA, which is central to the gene expression and replication of HBV. We developed an assay system for nuclear circular DNA using an integration-deficient lentiviral vector. This vector produced non-integrated circular DNA in nuclei of infected cells. We engineered this vector to encode firefly luciferase to monitor the lentiviral episome DNA. We screened 3,840 chemicals by this assay for luciferase-reducing activity and identified dicumarol, which is known to have anticoagulation activity. We confirmed that dicumarol reduced lentiviral episome DNA. Furthermore, dicumarol inhibited HBV replication in cell culture using NTCP-expressing HepG2 and primary human hepatocytes. Dicumarol reduced intracellular HBV RNA, DNA, supernatant HBV antigens and DNA. We also found that dicumarol reduced the cccDNA level in HBV infected cells, but did not affect HBV adsorption/entry. This is a novel assay system for screening inhibitors targeting nuclear cccDNA and is useful for finding new antiviral substances for HBV.

Original language	English
Article number	e0212233
Journal	PLOS ONE
Volume	14
Issue	2
ISSN	1932-6203
DOIs	https://doi.org/10.1371/journal.pone.0212233
Publication status	Published - Feb 2019

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Takeuchi, F., Ikeda, S., Tsukamoto, Y., Iwasawa, Y., Qihao, C., Otakaki, Y., Ryota, O., Yao, W. L., Narita, R., Makoto, H., Watashi, K., Wakita, T., Takeuchi, K., Chayama, K., Kogure, A., Kato, H., & Fujita, T. (2019). Screening for inhibitor of episomal DNA identified dicumarol as a hepatitis B virus inhibitor. PLOS ONE, 14(2), Article e0212233. https://doi.org/10.1371/journal.pone.0212233

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title = "Screening for inhibitor of episomal DNA identified dicumarol as a hepatitis B virus inhibitor",

abstract = "Currently, there is no available therapy to eradicate hepatitis B virus (HBV) in chronically infected individuals. This is due to the difficulty in eliminating viral covalently closed circular (ccc) DNA, which is central to the gene expression and replication of HBV. We developed an assay system for nuclear circular DNA using an integration-deficient lentiviral vector. This vector produced non-integrated circular DNA in nuclei of infected cells. We engineered this vector to encode firefly luciferase to monitor the lentiviral episome DNA. We screened 3,840 chemicals by this assay for luciferase-reducing activity and identified dicumarol, which is known to have anticoagulation activity. We confirmed that dicumarol reduced lentiviral episome DNA. Furthermore, dicumarol inhibited HBV replication in cell culture using NTCP-expressing HepG2 and primary human hepatocytes. Dicumarol reduced intracellular HBV RNA, DNA, supernatant HBV antigens and DNA. We also found that dicumarol reduced the cccDNA level in HBV infected cells, but did not affect HBV adsorption/entry. This is a novel assay system for screening inhibitors targeting nuclear cccDNA and is useful for finding new antiviral substances for HBV.",

author = "Fumihiko Takeuchi and Sotaro Ikeda and Yuta Tsukamoto and Yoshikazu Iwasawa and Chen Qihao and Yukie Otakaki and Ouda Ryota and Yao, {Wan Ling} and Ryo Narita and Hijikata Makoto and Koichi Watashi and Takaji Wakita and Koh Takeuchi and Kazuaki Chayama and Amane Kogure and Hiroki Kato and Takashi Fujita",

year = "2019",

month = feb,

doi = "10.1371/journal.pone.0212233",

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volume = "14",

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Takeuchi, F, Ikeda, S, Tsukamoto, Y, Iwasawa, Y, Qihao, C, Otakaki, Y, Ryota, O, Yao, WL, Narita, R, Makoto, H, Watashi, K, Wakita, T, Takeuchi, K, Chayama, K, Kogure, A, Kato, H & Fujita, T 2019, 'Screening for inhibitor of episomal DNA identified dicumarol as a hepatitis B virus inhibitor', PLOS ONE, vol. 14, no. 2, e0212233. https://doi.org/10.1371/journal.pone.0212233

TY - JOUR

T1 - Screening for inhibitor of episomal DNA identified dicumarol as a hepatitis B virus inhibitor

AU - Takeuchi, Fumihiko

AU - Ikeda, Sotaro

AU - Tsukamoto, Yuta

AU - Iwasawa, Yoshikazu

AU - Qihao, Chen

AU - Otakaki, Yukie

AU - Ryota, Ouda

AU - Yao, Wan Ling

AU - Narita, Ryo

AU - Makoto, Hijikata

AU - Watashi, Koichi

AU - Wakita, Takaji

AU - Takeuchi, Koh

AU - Chayama, Kazuaki

AU - Kogure, Amane

AU - Kato, Hiroki

AU - Fujita, Takashi

PY - 2019/2

Y1 - 2019/2

N2 - Currently, there is no available therapy to eradicate hepatitis B virus (HBV) in chronically infected individuals. This is due to the difficulty in eliminating viral covalently closed circular (ccc) DNA, which is central to the gene expression and replication of HBV. We developed an assay system for nuclear circular DNA using an integration-deficient lentiviral vector. This vector produced non-integrated circular DNA in nuclei of infected cells. We engineered this vector to encode firefly luciferase to monitor the lentiviral episome DNA. We screened 3,840 chemicals by this assay for luciferase-reducing activity and identified dicumarol, which is known to have anticoagulation activity. We confirmed that dicumarol reduced lentiviral episome DNA. Furthermore, dicumarol inhibited HBV replication in cell culture using NTCP-expressing HepG2 and primary human hepatocytes. Dicumarol reduced intracellular HBV RNA, DNA, supernatant HBV antigens and DNA. We also found that dicumarol reduced the cccDNA level in HBV infected cells, but did not affect HBV adsorption/entry. This is a novel assay system for screening inhibitors targeting nuclear cccDNA and is useful for finding new antiviral substances for HBV.

AB - Currently, there is no available therapy to eradicate hepatitis B virus (HBV) in chronically infected individuals. This is due to the difficulty in eliminating viral covalently closed circular (ccc) DNA, which is central to the gene expression and replication of HBV. We developed an assay system for nuclear circular DNA using an integration-deficient lentiviral vector. This vector produced non-integrated circular DNA in nuclei of infected cells. We engineered this vector to encode firefly luciferase to monitor the lentiviral episome DNA. We screened 3,840 chemicals by this assay for luciferase-reducing activity and identified dicumarol, which is known to have anticoagulation activity. We confirmed that dicumarol reduced lentiviral episome DNA. Furthermore, dicumarol inhibited HBV replication in cell culture using NTCP-expressing HepG2 and primary human hepatocytes. Dicumarol reduced intracellular HBV RNA, DNA, supernatant HBV antigens and DNA. We also found that dicumarol reduced the cccDNA level in HBV infected cells, but did not affect HBV adsorption/entry. This is a novel assay system for screening inhibitors targeting nuclear cccDNA and is useful for finding new antiviral substances for HBV.

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U2 - 10.1371/journal.pone.0212233

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M3 - Journal article

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AN - SCOPUS:85061874310

SN - 1932-6203

VL - 14

JO - PLOS ONE

JF - PLOS ONE

IS - 2

M1 - e0212233

ER -

Screening for inhibitor of episomal DNA identified dicumarol as a hepatitis B virus inhibitor

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