Dataset Information

Virtual Screening and Biological Validation of Novel Influenza Virus PA Endonuclease Inhibitors.

ABSTRACT: The influenza virus RNA-dependent RNA polymerase complex (RdRp), a heterotrimeric protein complex responsible for viral RNA transcription and replication, represents a primary target for antiviral drug development. One particularly attractive approach is interference with the endonucleolytic "cap-snatching" reaction by the RdRp subunit PA, more precisely by inhibiting its metal-dependent catalytic activity which resides in the N-terminal part of PA (PA-Nter). Almost all PA inhibitors (PAIs) thus far discovered bear pharmacophoric fragments with chelating motifs able to bind the bivalent metal ions in the catalytic core of PA-Nter. More recently, the availability of crystallographic structures of PA-Nter has enabled rational design of original PAIs with improved binding properties and antiviral potency. We here present a coupled pharmacophore/docking virtual screening approach that allowed us to identify PAIs with interesting inhibitory activity in a PA-Nter enzymatic assay. Moreover, antiviral activity in the low micromolar range was observed in cell-based influenza virus assays.

SUBMITTER: Pala N

PROVIDER: S-EPMC4538433 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature

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Virtual Screening and Biological Validation of Novel Influenza Virus PA Endonuclease Inhibitors.

Pala Nicolino N Stevaert Annelies A Dallocchio Roberto R Dessì Alessandro A Rogolino Dominga D Carcelli Mauro M Sanna Vanna V Sechi Mario M Naesens Lieve L

ACS medicinal chemistry letters 20150618 8

The influenza virus RNA-dependent RNA polymerase complex (RdRp), a heterotrimeric protein complex responsible for viral RNA transcription and replication, represents a primary target for antiviral drug development. One particularly attractive approach is interference with the endonucleolytic "cap-snatching" reaction by the RdRp subunit PA, more precisely by inhibiting its metal-dependent catalytic activity which resides in the N-terminal part of PA (PA-Nter). Almost all PA inhibitors (PAIs) thus ...[more]

PMID: 26288686

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Project description:The clinical severity and annual occurrence of influenza virus epidemics, combined with the availability of just a single class of antivirals to treat infections, underscores the urgent need to develop new anti-influenza drugs. The endonuclease activity within the viral acidic polymerase (PA) protein is an attractive target for drug discovery due to the critical role it plays in viral gene transcription. RO-7 is a next-generation PA endonuclease inhibitor of influenza A and B viruses, but its drug resistance potential is unknown. Through serial passage of influenza A(H1N1) viruses in MDCK cells under selective pressure of RO-7, we identified an I38T substitution within the PA endonuclease domain that conferred in vitro resistance to RO-7 (up to a 287-fold change in 50% effective concentration [EC50]). I38T emerged between 5 and 10 passages, and when introduced into recombinant influenza A(H1N1) viruses, alone conferred RO-7 resistance (up to an 81-fold change in EC50). Cocrystal structures of mutant and wild-type endonuclease domains with RO-7 provided the structural basis of resistance, where a key hydrophobic interaction between RO-7 and the Ile38 side chain is compromised when mutated to the polar threonine. While Ile38 does not have a crucial role in coordinating the endonuclease active site, the switch to threonine does affect the polymerase activity of some viruses and influences RO-7 affinity for the PAN target (i.e., the ≈200-residue N-terminal domain of PA). However, the change does not lead to a complete loss of replication activity in vitro Our results predict that RO-7-resistant influenza viruses carrying the I38T substitution may emerge under treatment. This should be taken into consideration for clinical surveillance and in refinement of these drugs.IMPORTANCE The effectiveness of antiviral drugs can be severely compromised by the emergence of resistant viruses. Therefore, determination of the mechanisms by which viruses become resistant is critical for drug development and clinical use. RO-7 is a compound that potently inhibits influenza virus replication and belongs to a new class of drugs in late-stage clinical trials for treatment of influenza virus infection. Here we demonstrate that a single amino acid change acquired under prolonged virus exposure to RO-7 renders influenza viruses significantly less susceptible to its inhibitory effects. We have discovered how the mutation can simultaneously interfere with drug activity and still maintain efficient virus replication. These findings have important implications for the development of more effective derivatives of RO-7-like drugs and provide guidance for how to monitor the emergence of resistance.

Dataset Information

Virtual Screening and Biological Validation of Novel Influenza Virus PA Endonuclease Inhibitors.

Publications

Virtual Screening and Biological Validation of Novel Influenza Virus PA Endonuclease Inhibitors.

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