Design and synthesis of naturally-inspired SARS-CoV-2 inhibitors.

A naturally inspired chemical library of 25 molecules was synthesised guided by 3-D dimensionality and natural product likeness factors to explore a new chemical space. The synthesised chemical library, consisting of fused-bridged dodecahydro-2a,6-epoxyazepino[3,4,5-c,d]indole skeletons, followed lead likeness factors in terms of molecular weight, C-sp3 fraction and Clog P. Screening of the 25 compounds against lung cells infected with SARS-CoV-2 led to the identification of 2 hits. Although the chemical library showed cytotoxicity, the two hits (3b, 9e) showed the highest antiviral activity (EC50 values of 3.7 and 1.4 µM, respectively) with an acceptable cytotoxicity difference. Computational analysis based on docking and molecular dynamics simulations against main protein targets in SARS-CoV-2 (main protease Mpro, nucleocapsid phosphoprotein, non-structural protein nsp10-nsp16 complex and RBD/ACE2 complex) were performed. The computational analysis proposed the possible binding targets to be either Mpro or the nsp10-nsp16 complex. Biological assays were performed to confirm this proposition. A cell-based assay for Mpro protease activity using a reverse-nanoluciferase (Rev-Nluc) reporter confirmed that 3b targets Mpro. These results open the way towards further hit-to-lead optimisations.

SS148 and WZ16 inhibit the activities of nsp10-nsp16 complexes from all seven human pathogenic coronaviruses.

Seven coronaviruses have infected humans (HCoVs) to-date. SARS-CoV-2 caused the current COVID-19 pandemic with the well-known high mortality and severe socioeconomic consequences. MERS-CoV and SARS-CoV caused epidemic of MERS and SARS, respectively, with severe respiratory symptoms and significant fatality. However, HCoV-229E, HCoV-NL63, HCoV-HKU1, and HCoV-OC43 cause respiratory illnesses with less severe symptoms in most cases. All coronaviruses use RNA capping to evade the immune systems of humans. Two viral methyltransferases, nsp14 and nsp16, play key roles in RNA capping and are considered valuable targets for development of anti-coronavirus therapeutics. But little is known about the kinetics of nsp10-nsp16 methyltransferase activities of most HCoVs, and reliable assays for screening are not available. Here, we report the expression, purification, and kinetic characterization of nsp10-nsp16 complexes from six HCoVs in parallel with previously characterized SARS-CoV-2. Probing the active sites of all seven by SS148 and WZ16, the two recently reported dual nsp14 / nsp10-nsp16 inhibitors, revealed pan-inhibition. Overall, our study show feasibility of developing broad-spectrum dual nsp14 / nsp10-nsp16-inhibitor therapeutics.