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A time-of–drug addition approach to target identification of antiviral compounds

Nature Protocols volume 6, pages 925–933 (2011)Cite this article

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Abstract

Insight into the mode of action of newly discovered antiviral agents is now almost a prerequisite for clinical development. This protocol describes a method that provides information on the target of inhibitors of the human immunodeficiency virus (HIV); it can also be adapted to other viruses. The results from this experiment are available within 2 d. This time-based approach determines how long the addition of a compound can be postponed before losing its antiviral activity in cell culture. The target of an antiviral compound can be identified by comparing its relative position in the time scale to that of reference drugs. Therefore, it is more precise than, for example, in the case of HIV, a determination of pre- or postintegrational mode of action, and combines in one routine different assays for studying mechanisms of action.

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Figure 1: Determination of the time necessary for one round of HIV-1 replication.
Figure 2: Flow diagram outlining the time-of-addition procedure.
Figure 3: Schematic illustration of the HIV replication cycle and a typical result from a time-of-addition experiment.
Figure 4: A time-of-addition experiment reveals the last occurring target in the viral replication cycle.
Figure 5: Result obtained with a polyanionic dendrimer targeting two steps in the viral replication depending on the concentration tested.

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Acknowledgements

We thank K. Erven, C. Heens and L. Bral for fine technical and editorial assistance.

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Author notes

  1. Rudi Pauwels

    Present address: Present address: Biocartis SA, Parc Scientifique EPFL, Lausanne, Switzerland.,

Authors and Affiliations

  1. Rega Institute for Medical Research, Katholieke Universiteit Leuven, Leuven, Belgium

    Dirk Daelemans, Rudi Pauwels, Erik De Clercq & Christophe Pannecouque

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  1. Dirk Daelemans
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  2. Rudi Pauwels
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  4. Christophe Pannecouque
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Contributions

R.P. and E.D.C. conceived the original protocol and D.D. and C.P. adjusted it to its current form and co-wrote the manuscript with E.D.C.

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Correspondence to Dirk Daelemans.

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Daelemans, D., Pauwels, R., De Clercq, E. et al. A time-of–drug addition approach to target identification of antiviral compounds. Nat Protoc 6, 925–933 (2011). https://doi.org/10.1038/nprot.2011.330

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