Rosmarinic acid is a novel inhibitor for Hepatitis B virus replication targeting viral epsilon RNA-polymerase interaction

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  • Yuta Tsukamoto, Kyoto University, University Hospital Bonn
  • ,
  • Sotaro Ikeda, Kyoto University
  • ,
  • Koji Uwai, Muroran Institute of Technology
  • ,
  • Riho Taguchi, Muroran Institute of Technology
  • ,
  • Kazuaki Chayama, Hiroshima University
  • ,
  • Takemasa Sakaguchi, Hiroshima University
  • ,
  • Ryo Narita
  • Wan Ling Yao, Kyoto University
  • ,
  • Fumihiko Takeuchi, Kyoto University
  • ,
  • Yukie Otakaki, Kyoto University
  • ,
  • Koichi Watashi, National Institute of Infectious Diseases, Tokyo University of Science, Japan Science and Technology Agency
  • ,
  • Takaji Wakita, National Institute of Infectious Diseases
  • ,
  • Hiroki Kato, Kyoto University, University Hospital Bonn
  • ,
  • Takashi Fujita, Kyoto University

Current therapeutics for hepatitis B virus (HBV) patients such as nucleoside analogs (NAs) are effective; however, new antiviral drugs against HBV are still desired. Since the interaction between the epsilon (ε) sequence of HBV pregenomic RNA and viral polymerase (Pol) is a key step in the HBV replication cycle, we aimed to identify small compounds for its inhibition, and established a pull-down assay system for the detection of ε-RNA-binding-Pol. Screening showed that 5 out of 3,965 compounds inhibited ε-Pol binding, and we identified rosmarinic acid, which exhibited specificity, as a potential antiviral agent. In order to examine the anti-HBV effects of rosmarinic acid, HBV-infected primary human hepatocytes from a humanized mouse liver were treated with rosmarinic acid. The rosmarinic acid treatment decreased HBV components including the amounts of extracellular HBV DNA with negligible cytotoxicity. We also investigated the combined effects of rosmarinic acid and the NA, lamivudine. rosmarinic acid slightly enhanced the anti-HBV activity of lamivudine, suggesting that the HBV replication step targeted by rosmarinic acid is distinct from that of NA. We analyzed an additional 25 rosmarinic acid derivatives, and found that 5 also inhibited ε-Pol. Structural comparisons between these derivatives implied that the “two phenolic hydroxyl groups at both ends” and the “caffeic acid-like structure” of rosmarinic acid are critical for the inhibition of ε-Pol binding. Collectively, our results demonstrate that rosmarinic acid inhibits HBV replication in HBV-infected cells by specifically targeting ε-Pol binding.

Original languageEnglish
Article numbere0197664
JournalPLOS ONE
Volume13
Issue5
Number of pages16
ISSN1932-6203
DOIs
Publication statusPublished - 21 May 2018

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