Characterization of the dynamics of hepatitis B virus resistance to adefovir by ultra-deep pyrosequencing

Authors

  • Christophe Rodriguez,

    1. National Reference Center for Viral Hepatitis B, C and Delta, Department of Virology, Hôpital Henri Mondor, Université Paris-Est, Créteil, France
    2. INSERM U955, Créteil, France
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  • Stéphane Chevaliez,

    1. National Reference Center for Viral Hepatitis B, C and Delta, Department of Virology, Hôpital Henri Mondor, Université Paris-Est, Créteil, France
    2. INSERM U955, Créteil, France
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  • Paul Bensadoun,

    1. INSERM U955, Créteil, France
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  • Jean-Michel Pawlotsky

    Corresponding author
    1. INSERM U955, Créteil, France
    • National Reference Center for Viral Hepatitis B, C and Delta, Department of Virology, Hôpital Henri Mondor, Université Paris-Est, Créteil, France
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  • Potential conflicts of interest: C.R., S.C., and J.M.P. have received research grants from Gilead Sciences. S.C. and J.M.P. have served as advisors for Gilead Sciences.

  • This work was funded by the French National Agency for Research on AIDS and Viral Hepatitis (ANRS). The authors are particularly grateful to Prof. Jean-François Delfraissy, the Director of ANRS, for his continuous support to our UDPS projects. Christophe Rodriguez is the recipient of a doctoral fellowship from ANRS. The hosting laboratory is funded by the Fondation pour la Recherche Médicale (FRM) as an “FRM Team.“ The authors thank Katyna Borroto-Esoda and Gilead Sciences for providing the samples from patients treated with adefovir used in this study and Prof. Manh-Tong Dao for providing the samples from unselected patients newly observed in the Department of Hepatology of the University Hospital of Caen, France.

Address reprint requests to: Jean-Michel Pawlotsky, M.D., Ph.D., Department of Virology, Hôpital Henri Mondor, 51 avenue du Maréchal de Lattre de Tassigny, 94010 Créteil, France. E-mail: jean-michel.pawlotsky@hmn.aphp.fr; fax: +33-1-4981-4831.

Abstract

Hepatitis B virus (HBV) resistance to nucleoside/nucleotide analogs is frequent. Ultra-deep pyrosequencing (UDPS) is a powerful new tool that can detect minor viral variants and characterize complex quasispecies mixtures. We used UDPS to analyze the dynamics of adefovir-resistant HBV variants in patients with chronic HBV infection in whom adefovir resistance occurred during treatment. Amino acid substitutions known to confer resistance to adefovir were detected at baseline in most patients. The dynamics of adefovir-resistant variants were complex and differed among patients as a result of evolving differences in variant fitness. UDPS analysis revealed successive waves of selection of HBV populations with single and multiple amino acid substitutions. Adefovir-resistant variants were partially inhibited by lamivudine, but remained fit in its presence. Conclusion: Substitutions conferring HBV resistance to nucleoside/nucleotide analogs exist before treatment, and that the dynamics of adefovir-resistant populations are much more complex and heterogeneous than previously thought and involve thus far unknown amino acid substitutions. The UDPS-based approach described here is likely to have important implications for the assessment of antiviral drug resistance in research and clinical practice. (Hepatology 2013;53:890–901)

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