High-throughput screening of virus-and host-targeted suppressors of HCV infection

2011 Fiscal Year Final Research Report

• Project/Area Number: 21390226
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Gastroenterology
• Research Institution: Tokyo Medical and Dental University
• Principal Investigator: SAKAMOTO Naoya 東京医科歯科大学, 大学院・医歯学総合研究科, 寄附講座教員 (10334418)
• Co-Investigator(Kenkyū-buntansha): NAKAGAWA Mina 東京医科歯科大学, 医学部附属病院, 助教 (30401342) ; AZUMA Seishin 東京医科歯科大学, 医学部附属病院, 助教 (10376783) ; WATANABE Mamoru 東京医科歯科大学, 大学院・医歯学総合研究科, 教授 (10175127)
• Co-Investigator(Renkei-kenkyūsha): FUJITA Megumi (TASAKA Megumi) 東京医科歯科大学, 大学院・医歯学総合研究科, 特任助教 (50510369)
• Project Period (FY): 2009 – 2011
• Keywords: C型肝炎 / レプリコンシステム

Research Abstract

In the present study period, we have conducted high-throughput and comprehensive screening of drugs, physiologically active compounds and small synthetic compounds that suppress HCV replication by using HCV infection and replication cell culture.(1) By using HCV Feo replicon, we conducted cell-based screening of 8, 000 diversity-oriented synthesis(DOS) compounds. We initially selected 41 compounds that suppress HCV replication and further validation identified 5 epoxide compounds that had low IC50.(2) We screened 4, 000 synthetic compounds and identified 5 compounds that suppress HCV replicon and HCV-JFH1 cell culture. We further conducted structure-activity relation(SAR) analyses and identified essential molecular structure for the antiviral activity.(3) We screened serine-rich(SR) protein kinases and identified 4 anti-HCV compounds. By integrating above results, we will further give better understanding of HCV infection life cycle and molecular targets of antiviral activity.

Research Products

• [Journal Article] Suppression of hepatitis C virus replicon by adenovirus vector-mediated expression of tough decoy RNA against miR-122a. Virus Research (2012)
  • Author(s): Sakurai F, Furukawa N, Higuchi M, Okamoto S, Ono K, Yoshida T, Kondoh M, Yagi K, Sakamoto N, Katayama K, Mizuguchi H
  • Journal Title: EPub ahead of Print
  • DOI: DOI:10.1016/j.virusres.2012.02.003.
• [Journal Article] Let-7b is a novel regulator of hepatitis C virus replication. Cell Mol Life Sci 2012 (2012)
  • Author(s): Cheng J-C, Yeh YJ, Tseng CP, Hsu SD, Chang YL, Sakamoto N, Huang HD
  • Journal Title: EPub ahead of Print
  • DOI: DOI:10.1007/s00018-012-0940-6.
• [Journal Article] Inhibition of hepatitis C virus NS5B polymerase by S-trityl-L-cysteine derivatives (2012)
  • Author(s): Nichols DB, Fournet G, Gurukumar KR, Basu A, Lee JC, Sakamoto N, Kozielski F, Musmuca I, Joseph B, Ragno R, Kaushik-Basu N
  • Journal Title: Eur J MedChem Volume: 49 Pages: 191-199
• [Journal Article] Identification of novel N-(morpholine-4-carbonyloxy) amidine compounds as potent inhibitors against hepatitis C virus replication (2012)
  • Author(s): Kusano-Kitazume A, Sakamoto N, Okuno Y, Sekine-Osajima Y, Nakagawa M, Kakinuma S, Kiyonashi K, Nitta S, Murakawa M, Azuma S, Nishimura-Sakurai Y, Hagiwara M, Watanabe M
  • Journal Title: Antimicrob Agent Chemother Volume: 56(3) Pages: 1315-1323
• [Journal Article] Use of human hepatocyte-like cells derived from induced pluripotent stem cells as a model for hepatocytes in hepatitis C virus infection (2011)
  • Author(s): Yoshida T, Takayama K, Kondoh M, Sakurai F, Tani H, Sakamoto N, Matsuura Y, Mizuguchi H, Yagi K
  • Journal Title: Biochem Biophys Res Commun Volume: 416(1-2) Pages: 119-124
• [Journal Article] A new method for induced fit docking(GENIUS) and its application to virtual screening of novel HCV NS3-4A protease inhibitors (2011)
  • Author(s): Takaya D, Yamashita A, Kamijo K, Gomi J, Ito M, Maekawa S, Enomoto N, SakamotoN, Watanabe Y, Arai R, Umeyama H, Honma T, Matsumoto T, Yokoyama S
  • Journal Title: Bioorganic & Medicinal Chemistry Volume: 19(22) Pages: 6892-6905
• [Journal Article] Genome-wide association study identified ITPA/DDRGK1 variants reflecting thrombocytopenia in pegylatedinterferon and ribavirin therapy for chronic hepatitis C (2011)
  • Author(s): Tanaka Y, Kurosaki M, Nishida N, Sugiyama M, Matsuura K, Sakamoto N, Enomoto N, Yatsuhashi H, Nishiguchi S, Hino K, Hige S, Itoh Y, Tanaka E, Mochida S, Honda M, Hiasa Y, Koike A, Sugauchi F, Kaneko S, Izumi N, Tokunaga K, Mizokami M
  • Journal Title: Human Molecular Genetics Volume: 20(17) Pages: 3507-3516
• [Journal Article] Analysis of the complete open reading frame of genotype 2b hepatitis C virus inassociation with the response to peginterferon and ribavirin therapy (2011)
  • Author(s): Kadokura M, Maekawa S, Sueki R, Miura M, Komase K, Shindo H, Amemiya F, Uetake T, Inoue T, Sakamoto M, Nakagawa M, Sakamoto N, Watanabe M, Enomoto N
  • Journal Title: PLoSOne Volume: 6(9) Pages: e24514
• [Journal Article] Adenovirus vector-mediated assay system for hepatitis C virus replication (2011)
  • Author(s): Yoshida T, Kondoh M, Ojima M, Mizuguchi H, Yamagishi Y, Sakamoto N, Yagi K
  • Journal Title: Nucleic Acids Res Volume: 39(10) Pages: e64
• [Journal Article] Analysis of interferon signaling by infectious hepatitis C virus clones with substitutions of core amino acids 70 and 91 (2011)
  • Author(s): Funaoka Y, Sakamoto N, Nakagawa M, Kakinuma S, Suda S, Watanabe T, Nitta S, Kitazume A, Kiyohashi K, Murakawa M, Azuma S, Tsuchiya K, and Watanabe M
  • Journal Title: J Virol Volume: 85(12) Pages: 5986-5994
• [Journal Article] The inhibitory effect on hepatitis C virus infection of a triterpenoid compound, with or without interferon-alpha (2011)
  • Author(s): Watanabe T, Sakamoto N, Nakagawa M, Kakinuma S, Itsui Y, Nishimura-Sakurai Y, Ueyama M, Funaoka Y, Kitazume A, Nitta S, Kiyohashi K, Murakawa M, Azuma S, Tsuchiya K, Oooka S, Watanabe M
  • Journal Title: Antimicrob Agents Chemother Volume: 55(6) Pages: 2537-2545
• [Journal Article] Studies on virus kinetics using infectious fluorescence-tagged hepatitis C virus cell culture (2011)
  • Author(s): Yamamoto M, Sakamoto N, Nakamura T, Itsui Y, Nakagawa M, Nishimura-Sakurai Y, Kakinuma S, Azuma S, Tsuchiya K, Kato T, Wakita T, Watanabe M
  • Journal Title: Hepatol Res Volume: 41 Pages: 258-269
• [Journal Article] Association between lipid accumulation and the cannabinoid system in Huh7 cells expressing HCV genes (2011)
  • Author(s): Toyoda M, Kitaoka A, Machida K, Nishinakagawa T, Yada R, Kohjima M, Kato M, Kotoh K, Sakamoto N, Shiota G, Nakamuta M, Nakashima M, Enjoji M
  • Journal Title: Int J Mol Med Volume: 27 Pages: 619-624
• [Journal Article] IL-6-mediated intersubgenotypic variation of interferon sensitivity in hepatitis C virus genotype 2a/2b chimeric clones (2010)
  • Author(s): Suda G, Sakamoto N, Itsui Y, Nakagawa M, Tasaka-Fujita M, Funaoka Y, Watanabe T, Nitta S, Kiyohashi K, Azuma S, Kakinuma S, Tsuchiya K, Imamura M, Hiraga N, Chayama K, Watanabe M
  • Journal Title: Virology Volume: 407 Pages: 80-90
• [Journal Article] ITPA gene variant protects against anemia induced by pegylated interferon-alpha and ribavirin therapy for Japanese patients with chronic hepatitis C (2010)
  • Author(s): Sakamoto N, Tanaka Y, Nakagawa M, Yatsuhashi H, Nishiguchi S, Enomoto N, Azuma S, Nishimura-Sakurai Y, Kakinuma S, Nishida N, Tokunaga K, Honda M, Ito K, Mizokami M, Watanabe M
  • Journal Title: Hepatol Res Volume: 40 Pages: 1063-1071
• [Journal Article] Cell culture and in vivo analyses of cytopathic hepatitis C virus mutants (2010)
  • Author(s): Mishima K, Sakamoto N, Sekine-Osajima Y, Nakagawa M, Itsui Y, Azuma S, Kakinuma S, Kiyohashi K, Kitazume A, Tsuchiya K, Imamura M, Hiraga N, Chayama K, Wakita T, Watanabe M
  • Journal Title: Virology Volume: 405 Pages: 361-369
• [Journal Article] Curcumin inhibits hepatitis C virus replication via suppressing the Akt-SREBP-1 pathway (2010)
  • Author(s): Kim K, Kim KH, Kim HY, Cho HK, Sakamoto N, Cheong J
  • Journal Title: FEBS Lett Volume: 584 Pages: 707-712
• [Journal Article] Inhibition of hepatitis C virus replication by a specific inhibitor of serine-arginine-rich protein kinase (2010)
  • Author(s): Karakama Y, Sakamoto N, Itsui Y, Nakagawa M, Tasaka-Fujita M, Nishimura-Sakurai Y, Kakinuma S, Oooka M, Azuma S, Tsuchiya K, Onogi H, Hagiwara M, Watanabe M
  • Journal Title: Antimicrob Agents Chemother Volume: 54 Pages: 3179-3186
• [Journal Article] Bioactivity-guided screening identifies pheophytin a as a potent anti-hepatitis C virus compound from Lonicera hypoglauca Miq (2009)
  • Author(s): Wang SY, Tseng CP, Tsai KC, Lin CF, Wen CY, Tsay HS, Sakamoto N, Tseng CH, Cheng JC
  • Journal Title: Biochem BiophysRes Commun Volume: 385 Pages: 230-235
• [Journal Article] Genome-wide association of IL28B with response to pegylated interferon-alpha and ribavirin therapy for chronic hepatitis C (2009)
  • Author(s): Tanaka Y, Nishida N, Sugiyama M, Kurosaki M, Matsuura K, Sakamoto N, Nakagawa M, Korenaga M, Hino K, Hige S, Ito Y, Mita E, Tanaka E, Mochida S, Murawaki Y, Honda M, Sakai A, Hiasa Y, Nishiguchi S, Koike A, Sakaida I, Imamura M, Ito K, Yano K, Masaki N, Sugauchi F, Izumi N, Tokunaga K, Mizokami M
  • Journal Title: NatGenet Volume: 41 Pages: 1105-1109
• [Journal Article] A functional genomic screen identifies cellular cofactors of hepatitis C virus replication (2009)
  • Author(s): Tai AW, Benita Y, Peng LF, Kim SS, Sakamoto N, Xavier RJ, Chung RT
  • Journal Title: Cell Host Microbe Volume: 5 Pages: 298-307
• [Journal Article] Two flavonoids extracts from Glycyrrhizae radix inhibit in vitro hepatitis C virus replication (2009)
  • Author(s): Sekine-Osajima Y, Sakamoto N, Nakagawa M, Itsui Y, Tasaka M, Nishimura-Sakurai Y, Chen CH, Suda G, Mishima K, Onuki Y, Yamamoto M, Maekawa S, Enomoto N, Kanai T, Tsuchiya K, Watanabe M
  • Journal Title: Hepatol Res Volume: 39 Pages: 60-69
• [Journal Article] Prospects for future therapy of hepatitis Cvirus infection (2009)
  • Author(s): Sakamoto N, Wu GY
  • Journal Title: Future Virology Volume: 4 Pages: 453-462
• [Journal Article] New therapeutic approaches to hepatitis Cvirus (2009)
  • Author(s): Sakamoto N, Watanabe M
  • Journal Title: JGastroenterol Volume: 44 Pages: 643-649
• [Journal Article] Antiviral effects of the interferon-induced protein guanylate binding protein 1 and its interaction with the hepatitis C virus NS5B protein (2009)
  • Author(s): Itsui Y, Sakamoto N, Kakinuma S, Nakagawa M, Sekine-Osajima Y, Tasaka-Fujita M, Nishimura-Sakurai Y, Suda G, Karakama Y, Mishima K, Yamamoto M, Watanabe T, Ueyama M, Funaoka Y, Azuma S, Watanabe M
  • Journal Title: Hepatology Volume: 50 Pages: 1727-1737
• [Journal Article] The isomerase active site of cyclophilin A is critical for hepatitis C virus replication (2009)
  • Author(s): Chatterji U, Bobardt M, Selvarajah S, Yang F, Tang H, Sakamoto N, Vuagniaux G, Parkinson T, Gallay P
  • Journal Title: J Biol Chem Volume: 284 Pages: 16998-17005
• [Journal Article] Inhibition of hepatitis C virus replication and viral helicase by ethyl acetate extract of the marine feather star Alloeocomatella polycladia
  • Author(s): Yamashita A, Abdus K, Furuta A, Matsuda Y, Fujita O, Tani H, Fujita Y, Fujimoto Y, Ikeda M, Kato N, Sakamoto N, Maekawa S, Enomoto N, Nakakoshi M, Tsubuki M, Sekiguchi Y, Thuneda S, Akimitsu N, Noda N, Tanaka J, Moriishi K
  • Journal Title: Marine Drugs Volume: (in press)
• [Presentation] A high-content screening assay using infectious fluorescence-tagged hepatitis C virus reveals candidates fo rsmall molecule inhibitors of viral entry (2011)
  • Author(s): Kusano-Kitazume A, Sakamoto N, Okuno Y, Yamamoto M, Sekine-Osajima Y, Nakagawa M, Kakinuma S, Kiyohashi K, Nitta S, Murakawa M, Hagiwara M, Watanabe M
  • Organizer: 62th. Annual Meeting of American Association for the Study of Liver Diseases
  • Place of Presentation: San Francisco, CA.(Poster# 383)
  • Year and Date: 2011-11-04
• [Presentation] Development of an RNA polymerase I-driven adenoviral vector and its application in an HCV replication assay (2011)
  • Author(s): Takeshi Yoshida, Fumi Satoh, Watari Akihiro, Masuo Kondoh, Hiroyuk iMizuguchi, Naoya Sakamoto, Kiyohito Yagi
  • Organizer: 18th. International Meeting on Hepatitis C Virus & Related Viruses
  • Place of Presentation: Seattle, WA
  • Year and Date: 2011-09-08
• [Presentation] Analysis of interferon signaling by infectious hepatitis C virus clones with substitutions of core amino acids 70 and 91 (2011)
  • Author(s): Naoya Sakamoto, Yusuke Funaoka, Goki Suda, Mina Nakagawa, Sei Kakinuma, Mamoru Watanabe
  • Organizer: 18th. International Meeting on Hepatitis C Virus & RelatedViruses
  • Place of Presentation: Seattle, WA
  • Year and Date: 2011-09-08
• [Presentation] HCV-NS4B targets STING anda brogates RIG-I-mediated type-Iinterferon-dependent innate immune response (2011)
  • Author(s): Sayuri Nitta, Naoya Sakamoto, Megumi Tasaka-Fujita, Kei Kiyohashi, Akiko Kusano-Kitazume, Miyako Murakawa, Kouhei Yoshino, Kako Mishima, SeiKakinuma, Mina Nakagawa, Mamoru Watanabe
  • Organizer: 18th. International Meeting on Hepatitis C Virus & Related Viruses
  • Place of Presentation: Seattle, WA
  • Year and Date: 2011-09-08
• [Presentation] Development of RNA pol-driven adenovirus vector expressing hepatitis C virus replicon (2011)
  • Author(s): Yoshiaki Yamagishi, Takeshi Yoshida, Masuo Kondoh, Hiroyuki Mizuguchi, Naoya Sakamoto, Akihiro Watari, Kiyohito Yagi
  • Organizer: Experimental Biology 2011
  • Place of Presentation: Washington, DC
  • Year and Date: 2011-04-13
• [Presentation] Identification of Novel Small Molecules Inhibitors of Hepatitis C Virus Replication Using a Cell-Based High-Throughput Screen (2010)
  • Author(s): Akiko Kusano-Kitazume, Naoya Sakamoto, Yuko Sekine-Osajima, Seishin Azuma, Mina Nakagawa, Sei Kakinuma, Yuki Nishimura-Sakurai, Yukiko Okuno, Masatoshi Hagiwara, and Mamoru Watanabe
  • Organizer: 17th. International Meeting on Hepatitis C Virus & Related Viruses
  • Place of Presentation: Yokohama, Japan
  • Year and Date: 2010-09-09
• [Presentation] The inhibitory effect of triterpenoid compound with or without interferonαon Hepatitis C virus infection (2010)
  • Author(s): Takako Watanabe, Naoya Sakamoto, Mina Nakagawa, Yasuhiro Itsui, Yuki Nishimura-Sakurai, Megumi Tasaka-Fujita, Yuko Sekine-Osajima, and Mamoru Watanabe
  • Organizer: 17th. International Meeting on Hepatitis C Virus & Related Viruses
  • Place of Presentation: Yokohama, Japan
  • Year and Date: 2010-09-09
• [Presentation] In-vitro replication and interferon sensitivity of core aa 70 and 91 mutant HCV cell culture (2009)
  • Author(s): Yusuke Funaoka, Naoya Sakamoto, Goki Suda, Yasuhiro Itsui, Mina Nakagawa, Kako Mishima, Yuko Karakama, Machi Yamamoto, Cheng-Hsin Chen-Azuma, Sei Kakinuma and Mamoru Watanabe
  • Organizer: 60th. Annual Meeting of American Association for the Study of Liver Diseases
  • Place of Presentation: Boston, MA
  • Year and Date: 2009-10-30
• [Presentation] In-vitro and in-vivo Characterization of a new genotype 2b HCV clone and 2b/JFH1 intergenotypic chimera and analyses of the factor that regulate interferon sensitivity (2009)
  • Author(s): Goki Suda, Naoya Sakamoto, Yasuhiro Itsui, Mina Nakagawa, Kako Mishima, Yuko Karakama, Machi Yamamoto, Cheng-Hsin Chen, Sei Kaki numa, KazuakiChayama, Michio Imamura and Mamoru Watanabe
  • Organizer: 60th. Annual Meeting of American Association for the Study of Liver Diseases
  • Place of Presentation: Boston, MA
  • Year and Date: 2009-10-30
• [Presentation] Establishment and genetic analyses of cytopathogenic HCV-JFH1 mutants by plaque-forming assay (2009)
  • Author(s): Kako Mishima, Naoya Sakamoto, Yuko Sekine-Osajima, Mina Nakagawa, Megumi Tasaka, Yuki Nishimura-Sakurai, Yasuhiro Itsui, Takaji Wakita, and Mamoru Watanabe
  • Organizer: 16th. International Meeting on Hepatitis C Virus & Related Viruses
  • Place of Presentation: Nice, France
  • Year and Date: 2009-10-03
• [Book] siRNAによるC型肝炎遺伝子治療.バイオ医薬品の開発技術とシーズ (2009)
  • Author(s): 坂本直哉、横田隆〓
  • Total Pages: 246-251
  • Publisher: シーエムシー出版
• [Patent(Industrial Property Rights)] C型肝炎ウイルスの増殖を抑制する医薬組成物 (2011)
  • Inventor(s): 坂本直哉、渡辺守、北詰晶子、萩原正敏、奥野友紀子
  • Industrial Property Rights Holder: 東京医科歯科大学
  • Industrial Property Number: 特許、特願2011-194082
  • Filing Date: 2011-09-06