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Khaya grandifoliola C.DC: a potential source of active ingredients against hepatitis C virus in vitro

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Abstract

In this study, we examined the antiviral properties of Khaya grandifoliola C.DC (Meliaceae) on the hepatitis C virus (HCV) life cycle in vitro and identified some of the chemical constituents contained in the fraction with the most antiviral activity. Dried bark powder was extracted by maceration in a methylene chloride/methanol (MCM) system (50:50; v/v) and separated on silica gel by flash chromatography. Infection and replication rates in Huh-7 cells were investigated by luciferase reporter assay and indirect immunofluorescence assay using subgenomic replicons, HCV pseudotyped particles, and cell-culture-derived HCV (HCVcc), respectively. Cell viability was assessed by MTT assay, and cellular gene expression was analysed by qRT-PCR. The chemical composition of the fraction with the most antiviral activity was analysed by coupled gas chromatography and mass spectrometry (GC-MS). Five fractions of different polarities (F0-F100) were obtained from the MCM extract. One fraction (KgF25) showed the strongest antiviral effect on LucUbiNeoET replicons at nontoxic concentrations. Tested at 100 µg/mL, KgF25 had a high inhibitory effect on HCV replication, comparable to that of 0.01 µM daclatasvir or 1 µM telaprevir. This fraction also inhibited HCVcc infection by mostly targeting the entry step. KgF25 inhibited HCV entry in a pan-genotypic manner by directly inactivating free viral particles. Its antiviral effects were mediated by the transcriptional upregulation of the haem oxygenase-1 gene and interferon antiviral response. Three constituents, namely, benzene, 1,1′-(oxydiethylidene)bis (1), carbamic acid, (4-methylphenyl)-, 1-phenyl (2), and 6-phenyl, 4-(1′-oxyethylphenyl) hexene (3), were identified from the active fraction KgF25 by GC-MS. Khaya grandifoliola contains ingredients capable of acting on different steps of the HCV life cycle.

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Abbreviations

BCP:

Boceprevir

CC50 :

Cytotoxic concentration 50

CCLR:

Cell culture lysis reagent

EGCG:

Epigallocatechin-3-gallate

GC/MS:

Gas chromatography/mass spectrometry

HCV:

Hepatitis C virus

HCVcc:

Cell-culture-derived hepatitis C virions

HCVpp:

Pseudotyped particles of hepatitis C virus

IC50 :

50 % inhibitory concentration

JFH-1:

Japanese fulminant hepatitis 1

Kg:

Khaya grandifoliola

KgCE:

Khaya grandifoliola crude extract

KgF25:

25 % methanol fraction of Khaya grandifoliola

LAR:

Luciferase assay reagent

MCM:

Methylene chloride/methanol

MOI:

Multiplicity of infection

Nrf-2:

Nuclear factor erythoid 2-related factor-2

NS:

Nonstructural

PEG-IFN:

Pegylated interferon

qRT-PCR:

Quantitative real-time-polymerase chain reaction

SVR:

Sustained virological response

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Acknowledgments

We thank the DAAD (German Academic Exchange Service) who Granted a six-month scholarship to B.R.T.G. at the University of Hamburg in Germany (Grant Number A/12/90499). We also thank Professor Hartmut Schlüter from the Department of Clinical Chemistry of the Hamburg University Medical Center for the GC and mass spectrometry analysis. This work was partly funded by the ‘Agence Nationale de la Recherche sur le Sida et les Hépatites virales’ (ANRS) in France through the “‘Centre d’Infection et d’Immunité de Lille”‘ and by the International Foundation for Sciences (IFS), Stockholm, Sweden, who supported the collection and fractionation of plant material through Grant No. F/4223-2, awarded to Dr. F.N. Njayou. Finally, we are grateful to Dr. Manfo Faustin Pascal for the critical review of this manuscript.

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Authors and Affiliations

  1. Laboratory of Pharmacology and Toxicology, Department of Biochemistry, Faculty of Science, University of Yaounde I, PO Box 812, Yaounde, Cameroon

    Borris Rosnay Tietcheu Galani, Frédéric Nico Njayou & Paul Fewou Moundipa

  2. Laboratory of Applied Biochemistry, Department of Biological Sciences, Faculty of Science, University of Ngaoundéré, Ngaoundéré, Cameroon

    Borris Rosnay Tietcheu Galani

  3. CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL, Centre d’Infection et d’Immunité de Lille, Univ. Lille, 59000, Lille, France

    Marie-Emmanuelle Sahuc, Gaspard Deloison, Priscille Brodin, Yves Rouillé & Karin Séron

  4. Institute of Experimental Immunology and Hepatology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

    Gabriele Sass, Christine Loscher & Gisa Tiegs

  5. Department of Organic Chemistry, Faculty of science, University of Yaounde I, Yaounde, Cameroon

    Pierre Mkounga

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  1. Borris Rosnay Tietcheu Galani
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  2. Marie-Emmanuelle Sahuc
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Galani, B.R.T., Sahuc, ME., Sass, G. et al. Khaya grandifoliola C.DC: a potential source of active ingredients against hepatitis C virus in vitro . Arch Virol 161, 1169–1181 (2016). https://doi.org/10.1007/s00705-016-2771-5

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