title: Synthesis and anti-hepatitis B virus activity of C4 amide-substituted isosteviol derivatives.
abstract: A series of novel isosteviol derivatives having C4-amide substituents were synthesized in order to test for antiviral effects against the hepatitis B virus (HBV) in vitro. Among them, IN-4 [N-(propylcarbonyl)-4α-amino-19-nor-ent-16-ketobeyeran] (5) exhibited inhibitory activity against secretion of HBsAg and HBeAg as well as inhibition of HBV DNA replication. Therefore, the mechanism of its antiviral activity was further analyzed using HBV-transfected Huh7 cells. Exposure to IN-4 produced minimal inhibitory effects on viral precore/pregenomic RNA expression. However, expression levels of the 2.4/2.1-kb preS/major S RNA of the viral surface gene significantly decreased, along with intracellular levels of HBV DNA. A promoter activity analysis demonstrated that IN-4 significantly inhibited viral X, S, and preS expression levels but not viral core promoter activities. In particular, IN-4 was observed to significantly inhibit HBV gene regulation by disrupting nuclear factor (NF)-κB-associated promoter activity. In addition, the nuclear expression of p65/p50 NF-κB member proteins was attenuated following IN-4 treatment, while cytoplasmic IκBα protein levels were enhanced. Meanwhile, IN-4 was observed to inhibit the binding activity of NF-κB to putative DNA elements. Furthermore, transfection of a p65 expression plasmid into Huh7 cells significantly reversed the inhibitory effect of IN-4 on HBV DNA levels, providing further evidence of the central role of NF-κB in its antiviral mechanism. It is therefore suggested that IN-4 inhibits HBV by interfering with the NF-κB signaling pathway, resulting in downregulation of viral gene expression and DNA replication.
authors.affiliation: School of Medicine, China Medical University, Taichung 404, Taiwan.
authors.forename: Tsurng-Juhn
authors.initials: TJ
authors.lastname: Huang
authors.affiliation: Graduate Institute of Biomedical Sciences, National Chung Hsing University, Taichung 402, Taiwan.
authors.forename: Cheng-Lin
authors.initials: CL
authors.lastname: Yang
authors.affiliation: Department of Radiation Oncology, China Medical University Hospital, Taichung 404, Taiwan; Department of Biomedical Imaging and Radiological Science, China Medical University, Taichung 404, Taiwan.
authors.forename: Yu-Cheng
authors.initials: YC
authors.lastname: Kuo
authors.affiliation: Department of Medical Laboratory Science and Biotechnology, China Medical University, Taichung 404, Taiwan.
authors.forename: Yi-Chih
authors.initials: YC
authors.lastname: Chang
authors.affiliation: Division of Chinese Medicinal Chemistry, National Research Institute of Chinese Medicine, Ministry of Health and Welfare, Taipei 112, Taiwan; Department of Pharmaceutical Sciences, School of Pharmacy, Taipei Medical University, Taipei 110, Taiwan.
authors.forename: Li-Ming
authors.initials: LM
authors.lastname: Yang
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, Taipei Medical University, Taipei 110, Taiwan.
authors.forename: Bo-Hon
authors.initials: BH
authors.lastname: Chou
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, Taipei Medical University, Taipei 110, Taiwan; Ph.D. Program for the Clinical Drug Discovery from Botanical Herbs, College of Pharmacy, Taipei Medical University, Taipei 110, Taiwan. Electronic address: shwu-lin@tmu.edu.tw.
authors.forename: Shwu-Jiuan
authors.initials: SJ
authors.lastname: Lin
mesh_terms: D000577:Amides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D000998:Antiviral Agents / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D002460:Cell Line; D004261:DNA Replication / Q000187:drug effects; D004279:DNA, Viral / Q000235:genetics; D045786:Diterpenes, Kaurane / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D006509:Hepatitis B / Q000188:drug therapy / Q000378:metabolism / Q000821:virology; D006514:Hepatitis B Surface Antigens / Q000378:metabolism; D006513:Hepatitis B e Antigens / Q000378:metabolism; D006515:Hepatitis B virus / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism
chemical_list: D000577:Amides; D000998:Antiviral Agents; D004279:DNA, Viral; D045786:Diterpenes, Kaurane; D006514:Hepatitis B Surface Antigens; D006513:Hepatitis B e Antigens; D016328:NF-kappa B; C515747:isosteviol
keywords: Antiviral effect; Hepatitis B virus; Isosteviol derivative; NF-κB