High-throughput screening of the ReFRAME, Pandemic Box, and COVID Box drug repurposing libraries against SARS-CoV-2 nsp15 endoribonuclease to identify small-molecule inhibitors of viral activity

Abstract

SARS-CoV-2 has caused a global pandemic, and has taken over 1.7 million lives as of mid-December, 2020. Although great progress has been made in the development of effective countermeasures, with several pharmaceutical companies approved or poised to deliver vaccines to market, there is still an unmet need of essential antiviral drugs with therapeutic impact for the treatment of moderate-to-severe COVID-19. Towards this goal, a high-throughput assay was used to screen SARS-CoV-2 nsp15 uracil-dependent endonuclease (endoU) function against 13 thousand compounds from drug and lead repurposing compound libraries. While over 80% of initial hit compounds were pan-assay inhibitory compounds, three hits were confirmed as nsp15 endoU inhibitors in the 1–20 μM range in vitro. Furthermore, Exebryl-1, a ß-amyloid anti-aggregation molecule for Alzheimer’s therapy, was shown to have antiviral activity between 10 to 66 μM, in Vero 76, Caco-2, and Calu-3 cells. Although the inhibitory concentrations determined for Exebryl-1 exceed those recommended for therapeutic intervention, our findings show great promise for further optimization of Exebryl-1 as an nsp15 endoU inhibitor and as a SARS-CoV-2 antiviral.

Molecular Docking and Dynamics Investigations for Identifying Potential Inhibitors of the 3-Chymotrypsin-like Protease of SARS-CoV-2: Repurposing of Approved Pyrimidonic Pharmaceuticals for COVID-19 Treatment

“…Gemcitabine and cidofovir were reported to inhibit SARS-CoV and SARS-CoV-2 proteins with IC 50 values of 4.95 µM and 36 µM [54,55]. Further, telbivudine, tipiracil, cytarabine, and citicoline were recommended as 3CL pro inhibitors [56][57][58][59][60][61]. This transitory literature scanning confirms these pharmaceuticals' activity against 3CL pro of SARS-CoV-2, as demonstrated in the present study.…”

Inhibition of the hexamerization of SARS-CoV-2 endoribonuclease and modeling of RNA structures bound to the hexamer

“…Of these, functional groups of Compound IV share analogous interactions with Nsp15 in Pocket A/B with those of the compound of pose 4 that shows the lowest . Choi et al used high-throughput assay to screen compounds against Nsp15, and inhibition was confirmed for three hits in vitro and showed that Exebryl-1 (ß-amyloid anti-aggregation molecule for Alzheimer’s therapy) has antiviral activity 38 . They also showed that the most plausible binding residues with Exebryl-1 significantly overlap with those of Pocket B. Interestingly, the compounds that we found by AutoDock Vina shares some similarity in functional groups with Exebryl-1.…”

The Role of Coronavirus RNA-Processing Enzymes in Innate Immune Evasion

“…In this direction, Benzopurpurin B and Congo red were shown to display the higher inhibition on Nsp15 activity. In addition, these drugs reduced the infectivity of the SARS-CoV in Vero cells [ 123 ]. Based on this evidence, a more comprehensive investigation is needed for the exploitation of RNAse A inhibitors as drugs that can target Nsp15 activity.…”

“…Using the available crystallographic data for NSP15 from MERS, SARS-CoV, and SARS-CoV-2, several groups have employed virtual screening, molecular docking, and molecular dynamic simulation techniques to identify putative antivirals against Nsp15 [ 123 , 124 ]. Nsp15 is a uridine-specific endoribonuclease and amino acid residue Ser294 has been proposed to play a crucial role in uracil recognition in the catalytic pocket [ 75 ].…”