Identification of Natural Products Inhibiting SARS-CoV-2 by Targeting Viral Proteases: A Combined in Silico and in Vitro Approach

Abstract

In this study, an integrated in silico−in vitro approach was employed to discover natural products (NPs) active against SARS-CoV-2. The two SARS-CoV-2 viral proteases, i.e., main protease (M pro ) and papain-like protease (PL pro ), were selected as targets for the in silico study. Virtual hits were obtained by docking more than 140,000 NPs and NP derivatives available in-house and from commercial sources, and 38 virtual hits were experimentally validated in vitro using two enzyme-based assays. Five inhibited the enzyme activity of SARS-CoV-2 M pro by more than 60% at a concentration of 20 μM, and four of them with high potency (IC 50 < 10 μM). These hit compounds were further evaluated for their antiviral activity against SARS-CoV-2 in Calu-3 cells. The results from the cell-based assay revealed three mulberry Diels−Alder-type adducts (MDAAs) from Morus alba with pronounced anti-SARS-CoV-2 activities. Sanggenons C (12), O (13), and G (15) showed IC 50 values of 4.6, 8.0, and 7.6 μM and selectivity index values of 5.1, 3.1 and 6.5, respectively. The docking poses of MDAAs in SARS-CoV-2 M pro proposed a butterfly-shaped binding conformation, which was supported by the results of saturation transfer difference NMR experiments and competitive 1 H relaxation dispersion NMR spectroscopy.

Synthesis and investigation of new indole‐containing vinyl sulfide derivatives: In silico and in vitro studies for potential therapeutic applications

“…These inhibitors find applications as therapeutic agents in various fields, such as blood coagulation, retroviral diseases (HIV), and cancer (mastoparanos) [23,24] . Given the diverse applications of protease inhibitors, the discovery of new ones has generated substantial interest for commercial purposes [25,26] . Additionally, compounds 3 b , 3 e , and 3 c exhibit a moderate inhibitory capacity in the context of protease inhibition.…”

“…Notably, extensive literature studies have emphasized the antioxidant properties of both synthetic and natural indole compounds [48] . This heightened activity can be primarily attributed to the presence of the highly electroreactive aromatic ring system [8–48] …”

“…[23,24] Given the diverse applications of protease inhibitors, the discovery of new ones has generated substantial interest for commercial purposes. [25,26] Additionally, compounds 3 b, 3 e, and 3 c exhibit a moderate inhibitory capacity in the context of protease inhibition.…”

Discovery of anti-SARS-CoV-2 secondary metabolites from the heartwood of Pterocarpus santalinus using multi-informative molecular networking

“…To evaluate the potential molecular target involved in the phenotypic activity, compound 5 with the strongest anti-SARS-CoV-2 activity was tested for SARS-CoV-2 main protease (M pro ) inhibition using an enzyme-based FRET assay ( Wasilewicz et al, 2023 ). However, compound 5 did not show any inhibitory activity against M pro at 50 µM when compared to the DMSO control (data not shown, method in Supplementary Material).…”

Protease inhibitors from Theobroma cacao impair SARS-CoV-2 replication in vitro

“…A biflavonoid from Anacardium occidentale and the flavonoids isorhoifolin and rutin from Theobroma cacao were studied as candidate anti -SARS-CoV-2 compounds, preferentially targeting the major viral protease (Mpro) [ [ 9 , 10 ]]. More recently, mulberry Diels–Alder-type adducts (MDAAs) from Morus alba have also been described as a new class of Mpro inhibitors of the SARS-CoV-2 virus [ 11 ].…”