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

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.

UR - http://www.scopus.com/inward/record.url?scp=85061874310&partnerID=8YFLogxK

U2 - 10.1371/journal.pone.0212233

DO - 10.1371/journal.pone.0212233

M3 - Journal article

C2 - 30779774

AN - SCOPUS:85061874310

SN - 1932-6203

VL - 14

JO - PLOS ONE

JF - PLOS ONE

IS - 2

M1 - e0212233

ER -