Céline Lacroix1, Jordi Querol-Audí2, Manon Roche3, David Franco1, Mathy Froeyen4, Pablo Guerra2, Thierry Terme3, Patrice Vanelle3, Núria Verdaguer2, Johan Neyts5, Pieter Leyssen1. 1. Laboratory of Virology and Experimental Chemotherapy, Rega Institute for Medical Research, KU Leuven, Leuven, Belgium. 2. Molecular Biology Institute of Barcelona (IBMB), Consejo Superior de Investigaciones Científicas, Barcelona, Spain. 3. Aix-Marseille Université, Institut de Chimie Radicalaire UMR 7273 CNRS 27, Boulevard Jean Moulin, Marseille, France. 4. Laboratory of Medicinal Chemistry, Rega Institute for Medical Research, KU Leuven, Leuven, Belgium. 5. Laboratory of Virology and Experimental Chemotherapy, Rega Institute for Medical Research, KU Leuven, Leuven, Belgium johan.neyts@rega.kuleuven.be.
Abstract
OBJECTIVES: To study the characteristics and the mode of action of the anti-rhinovirus compound 4-[1-hydroxy-2-(4,5-dimethoxy-2-nitrophenyl)ethyl]benzonitrile (LPCRW_0005). METHODS: The antiviral activity of LPCRW_0005 was evaluated in a cytopathic effect reduction assay against a panel of human rhinovirus (HRV) strains. To unravel its precise molecular mechanism of action, a time-of-drug-addition study, resistance selection and thermostability assays were performed. The crystal structure of the HRV14/LPCRW_0005 complex was elucidated as well. RESULTS: LPCRW_0005 proved to be a selective inhibitor of the replication of HRV14 (EC(50) of 2 ± 1 μM). Time-of-drug-addition studies revealed that LPCRW_0005 interferes with the earliest stages of virus replication. Phenotypic drug-resistant virus variants were obtained (≥30-fold decrease in susceptibility to the inhibitory effect of LPCRW_0005), which carried either an A150T or A150V amino acid substitution in the VP1 capsid protein. The link between the mutant genotype and drug-resistant phenotype was confirmed by reverse genetics. Cross-resistance studies and thermostability assays revealed that LPCRW_0005 has a similar mechanism of action to the capsid binder pleconaril. Elucidation of the crystal structure of the HRV14/LPCRW_0005 complex revealed the existence of multiple hydrophobic and polar interactions between the VP1 pocket and LPCRW_0005. CONCLUSIONS: LPCRW_0005 is a novel inhibitor of HRV14 replication that acts as a capsid binder. The compound has a chemical structure that is markedly smaller than that of other capsid binders. Structural studies show that LPCRW_0005, in contrast to pleconaril, leaves the toe end of the pocket in VP1 empty. This suggests that extended analogues of LPCRW_0005 that fill the full cavity could be more potent inhibitors of rhinovirus replication.
OBJECTIVES: To study the characteristics and the mode of action of the anti-rhinovirus compound 4-[1-hydroxy-2-(4,5-dimethoxy-2-nitrophenyl)ethyl]benzonitrile (LPCRW_0005). METHODS: The antiviral activity of LPCRW_0005 was evaluated in a cytopathic effect reduction assay against a panel of human rhinovirus (HRV) strains. To unravel its precise molecular mechanism of action, a time-of-drug-addition study, resistance selection and thermostability assays were performed. The crystal structure of the HRV14/LPCRW_0005 complex was elucidated as well. RESULTS:LPCRW_0005 proved to be a selective inhibitor of the replication of HRV14 (EC(50) of 2 ± 1 μM). Time-of-drug-addition studies revealed that LPCRW_0005 interferes with the earliest stages of virus replication. Phenotypic drug-resistant virus variants were obtained (≥30-fold decrease in susceptibility to the inhibitory effect of LPCRW_0005), which carried either an A150T or A150V amino acid substitution in the VP1 capsid protein. The link between the mutant genotype and drug-resistant phenotype was confirmed by reverse genetics. Cross-resistance studies and thermostability assays revealed that LPCRW_0005 has a similar mechanism of action to the capsid binder pleconaril. Elucidation of the crystal structure of the HRV14/LPCRW_0005 complex revealed the existence of multiple hydrophobic and polar interactions between the VP1 pocket and LPCRW_0005. CONCLUSIONS:LPCRW_0005 is a novel inhibitor of HRV14 replication that acts as a capsid binder. The compound has a chemical structure that is markedly smaller than that of other capsid binders. Structural studies show that LPCRW_0005, in contrast to pleconaril, leaves the toe end of the pocket in VP1 empty. This suggests that extended analogues of LPCRW_0005 that fill the full cavity could be more potent inhibitors of rhinovirus replication.
Authors: Liang Sun; Hyunwook Lee; Hendrik Jan Thibaut; Kristina Lanko; Eva Rivero-Buceta; Carol Bator; Belen Martinez-Gualda; Kai Dallmeier; Leen Delang; Pieter Leyssen; Federico Gago; Ana San-Félix; Susan Hafenstein; Carmen Mirabelli; Johan Neyts Journal: PLoS Pathog Date: 2019-05-09 Impact factor: 6.823
Authors: James T Kelly; Luigi De Colibus; Lauren Elliott; Elizabeth E Fry; David I Stuart; David J Rowlands; Nicola J Stonehouse Journal: Antiviral Res Date: 2015-10-30 Impact factor: 5.970