Identifying SARS-CoV-2 antiviral compounds by screening for small molecule inhibitors of nsp12/7/8 RNA-dependent RNA polymerase.
journal contributionposted on 2021-07-09, 09:42 authored by Agustina P Bertolin, Florian Weissmann, Jingkun Zeng, Viktor Posse, Jennifer C Milligan, Berta Canal, Rachel Ulferts, Mary Wu, Lucy S Drury, Michael Howell, Rupert Beale, John FX Diffley
The coronavirus disease 2019 (COVID-19) global pandemic has turned into the largest public health and economic crisis in recent history impacting virtually all sectors of society. There is a need for effective therapeutics to battle the ongoing pandemic. Repurposing existing drugs with known pharmacological safety profiles is a fast and cost-effective approach to identify novel treatments. The COVID-19 etiologic agent is the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a single-stranded positive-sense RNA virus. Coronaviruses rely on the enzymatic activity of the replication-transcription complex (RTC) to multiply inside host cells. The RTC core catalytic component is the RNA-dependent RNA polymerase (RdRp) holoenzyme. The RdRp is one of the key druggable targets for CoVs due to its essential role in viral replication, high degree of sequence and structural conservation and the lack of homologues in human cells. Here, we have expressed, purified and biochemically characterised active SARS-CoV-2 RdRp complexes. We developed a novel fluorescence resonance energy transfer-based strand displacement assay for monitoring SARS-CoV-2 RdRp activity suitable for a high-throughput format. As part of a larger research project to identify inhibitors for all the enzymatic activities encoded by SARS-CoV-2, we used this assay to screen a custom chemical library of over 5000 approved and investigational compounds for novel SARS-CoV-2 RdRp inhibitors. We identified three novel compounds (GSK-650394, C646 and BH3I-1) and confirmed suramin and suramin-like compounds as in vitro SARS-CoV-2 RdRp activity inhibitors. We also characterised the antiviral efficacy of these drugs in cell-based assays that we developed to monitor SARS-CoV-2 growth.
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-19nsp12RNA-dependent RNA polymeraseAnimalsAntiviral AgentsBenzoatesBridged Bicyclo Compounds, HeterocyclicChlorocebus aethiopsCoronavirus RNA-Dependent RNA PolymeraseDrug Evaluation, PreclinicalEnzyme AssaysFluorescence Resonance Energy TransferHigh-Throughput Screening AssaysHoloenzymesReproducibility of ResultsSARS-CoV-2Small Molecule LibrariesSuraminVero CellsViral Nonstructural ProteinsDiffley FC001066Beale FC001827HTS03 Chemical Sciences06 Biological Sciences11 Medical and Health SciencesBiochemistry & Molecular Biology