1. Dual inhibition of HIV-1 replication by integrase-LEDGF allosteric inhibitors is predominant at the post-integration stage
- Author
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François Moreau, Richard Benarous, Jean-Michel Bruneau, Erwann Le Rouzic, Marc Ruff, Ali Saïb, Julie Brias, Stéphane Emiliani, Frédéric Le Strat, Olivier Delelis, Eric Deprez, Céline Amadori, Roxane Beauvoir, Nicolas Levy, Alessia Zamborlini, Francis Chevreuil, Juliette Nguyen, Sophie Chasset, Sylvia Eiler, Sophie Vomscheid, Benoit Ledoussal, Damien Bonnard, Les Laboratoires Biodim-Mutabilis, Biocitech, Institut de Génétique et de Biologie Moléculaire et Cellulaire ( IGBMC ), Université de Strasbourg ( UNISTRA ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratoire de Biologie et de Pharmacologie Appliquée ( LBPA ), École normale supérieure - Cachan ( ENS Cachan ) -Centre National de la Recherche Scientifique ( CNRS ), Pathologie cellulaire : aspects moléculaires et viraux / Pathologie et Virologie Moléculaire, Université Paris Diderot - Paris 7 ( UPD7 ) -Institut Universitaire d'Hématologie ( IUH ), Université Paris Diderot - Paris 7 ( UPD7 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris]-Centre National de la Recherche Scientifique ( CNRS ), Institut Cochin ( UM3 (UMR 8104 / U1016) ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), EU FP7 provided financial support under the HIVINNOV consortium, grant agreement 305137. Marc Ruff team was supported by grants from the CNRS, the INSERM, SIDACTION, the French National Agency for Research against AIDS (ANRS), the SPINE 2 European Project (FP6 Contract Nu QLG2- CT-2002-00988 and 031220), the French Infrastructure for Integrated Structural Biology (FRISBI, ANR-10-INSB-05-01) and Instruct, part of the European Strategy Forum on Research Infrastructure (ESFRI) supported by national members subscription., Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Biologie et de Pharmacologie Appliquée (LBPA), École normale supérieure - Cachan (ENS Cachan)-Centre National de la Recherche Scientifique (CNRS), Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Centre National de la Recherche Scientifique (CNRS), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and BMC, Ed.
- Subjects
Protein Conformation ,Virus Integration ,Allosteric regulation ,Integrase inhibitor ,HIV Integrase ,Plasma protein binding ,Crystallography, X-Ray ,Virus Replication ,Integrase ,Cell Line ,03 medical and health sciences ,Allosteric inhibition ,[SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases ,Virology ,Humans ,HIV Integrase Inhibitors ,LEDGF ,030304 developmental biology ,0303 health sciences ,biology ,Drug discovery ,Research ,030302 biochemistry & molecular biology ,HIV ,Antiretroviral activity ,Molecular biology ,3. Good health ,Chromatin ,Cell biology ,[ SDV.MHEP.MI ] Life Sciences [q-bio]/Human health and pathology/Infectious diseases ,Protein-protein interaction inhibitor ,Infectious Diseases ,Viral replication ,[SDV.MHEP.MI] Life Sciences [q-bio]/Human health and pathology/Infectious diseases ,HIV-1 ,biology.protein ,Intercellular Signaling Peptides and Proteins ,Co-crystallization ,Protein Binding - Abstract
Background LEDGF/p75 (LEDGF) is the main cellular cofactor of HIV-1 integrase (IN). It acts as a tethering factor for IN, and targets the integration of HIV in actively transcribed gene regions of chromatin. A recently developed class of IN allosteric inhibitors can inhibit the LEDGF-IN interaction. Results We describe a new series of IN-LEDGF allosteric inhibitors, the most active of which is Mut101. We determined the crystal structure of Mut101 in complex with IN and showed that the compound binds to the LEDGF-binding pocket, promoting conformational changes of IN which explain at the atomic level the allosteric effect of the IN/LEDGF interaction inhibitor on IN functions. In vitro, Mut101 inhibited both IN-LEDGF interaction and IN strand transfer activity while enhancing IN-IN interaction. Time of addition experiments indicated that Mut101 behaved as an integration inhibitor. Mut101 was fully active on HIV-1 mutants resistant to INSTIs and other classes of anti-HIV drugs, indicative that this compound has a new mode of action. However, we found that Mut101 also displayed a more potent antiretroviral activity at a post-integration step. Infectivity of viral particles produced in presence of Mut101 was severely decreased. This latter effect also required the binding of the compound to the LEDGF-binding pocket. Conclusion Mut101 has dual anti-HIV-1 activity, at integration and post-integration steps of the viral replication cycle, by binding to a unique target on IN (the LEDGF-binding pocket). The post-integration block of HIV-1 replication in virus-producer cells is the mechanism by which Mut101 is most active as an antiretroviral. To explain this difference between Mut101 antiretroviral activity at integration and post-integration stages, we propose the following model: LEDGF is a nuclear, chromatin-bound protein that is absent in the cytoplasm. Therefore, LEDGF can outcompete compound binding to IN in the nucleus of target cells lowering its antiretroviral activity at integration, but not in the cytoplasm where post-integration production of infectious viral particles takes place.
- Published
- 2013
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