Identifying SARS-CoV-2 antiviral compounds by screening for small molecule inhibitors of nsp15 endoribonuclease.
journal contributionposted on 2021-07-09, 09:39 authored by Berta Canal, Ryo Fujisawa, Allison W McClure, Tom D Deegan, Mary Wu, Rachel Ulferts, Florian Weissmann, Lucy S Drury, Agustina P Bertolin, Jingkun Zeng, Rupert Beale, Michael Howell, Karim Labib, John FX Diffley
SARS-CoV-2 is responsible for COVID-19, a human disease that has caused over 2 million deaths, stretched health systems to near-breaking point and endangered economies of countries and families around the world. Antiviral treatments to combat COVID-19 are currently lacking. Remdesivir, the only antiviral drug approved for the treatment of COVID-19, can affect disease severity, but better treatments are needed. SARS-CoV-2 encodes 16 non-structural proteins (nsp) that possess different enzymatic activities with important roles in viral genome replication, transcription and host immune evasion. One key aspect of host immune evasion is performed by the uridine-directed endoribonuclease activity of nsp15. Here we describe the expression and purification of nsp15 recombinant protein. We have developed biochemical assays to follow its activity, and we have found evidence for allosteric behaviour. We screened a custom chemical library of over 5000 compounds to identify nsp15 endoribonuclease inhibitors, and we identified and validated NSC95397 as an inhibitor of nsp15 endoribonuclease in vitro. Although NSC95397 did not inhibit SARS-CoV-2 growth in VERO E6 cells, further studies will be required to determine the effect of nsp15 inhibition on host immune evasion.
Crick (Grant ID: 10827, Grant title: Beale FC001827) Crick (Grant ID: 10066, Grant title: Diffley FC001066) Crick (Grant ID: 10008, Grant title: STP High Throughput Screening) Wellcome Trust (Grant ID: 106252/Z/14/Z, Grant title: WT 106252/Z/14/Z) European Commission (Grant ID: 844211 - HUMAN REPL MECH, Grant title: EC 844211 - HUMAN REPL MECH)
coronavirusCOVID-19endonucleasensp15Allosteric RegulationAnimalsAntiviral AgentsChlorocebus aethiopsDrug Evaluation, PreclinicalEndoribonucleasesEnzyme AssaysFluorescenceHigh-Throughput Screening AssaysIn Vitro TechniquesKineticsNaphthoquinonesReproducibility of ResultsSARS-CoV-2Small Molecule LibrariesSolutionsVero CellsViral Nonstructural ProteinsDiffley FC001066Beale FC001827HTS03 Chemical Sciences06 Biological Sciences11 Medical and Health SciencesBiochemistry & Molecular Biology