A Combination of Structure-based Virtual Screening and Experimental Strategies to Identify the Potency of Caffeic Acid Ester Derivatives as SARS-CoV-2 3CLpro Inhibitor from an In-house Database (preprint)

Drug development requires significant time and resources, and computer-aided drug discovery techniques that integrate chemical and biological spaces offer valuable tools for the process. This study focused on the field of COVID-19 therapeutics and aimed to identify new active non-covalent inhibitors for 3CLpro, a key protein target. By combining in silico and in vitro approaches, an in-house database was utilized to identify potential inhibitors. The drug-likeness criteria was considered to pre-filter 553 compounds from 12 groups of natural products. Using structure-based virtual screening, 296 compounds were identified that matched the chemical features of SARS-CoV-2 3CLpro peptidomimetic inhibitor pharmacophore models. Subsequent molecular docking resulted in 43 hits with high binding affinities. Among the hits, caffeic acid analogs showed significant interactions with the 3CLpro active site, indicating their potential as promising candidates. To further evaluate their efficacy, enzyme-based assays were conducted, revealing that two ester derivatives of caffeic acid (4k and 4l) exhibited more than a 30% reduction in 3CLpro activity. Overall, these findings suggest that the screening approach employed in this study holds promise for the discovery of novel anti-SARS-CoV-2 therapeutics. Furthermore, the methodology could be extended for optimization or retrospective evaluation to enhance molecular targeting and antiviral efficacy of potential drug candidates.

Virtual flavonoids screening identifies potent compounds against enterovirus A71 and coxsackievirus A16 (preprint)

Worldwide outbreaks of hand, foot, and mouth disease (HFMD) are caused by enterovirus A71 (EV-A71) and coxsackievirus A16 (CV-A16). Since no anti-HFMD drugs are currently available, it is interesting to study potential viral inhibitors. Rupintrivir is a rhinovirus 3C protease (3Cpro) inhibitor with apparent inhibitory activity against HFMD. This study constructs pharmacophore models of the EV-A71 and CV-A16 3Cpro complexed with rupintrivir using molecular dynamics (MD) simulations. Both models had similar pharmacophore features, including hydrogen bond donors and acceptors and hydrophobic interactions. These pharmacophore models were then used as a template to screen 39 flavonoid compounds as potential novel inhibitors. Diosmin, epigallocatechin gallate (EGCG), and RTH-011 showed high binding affinities for EV-A71 and CV-A16 3Cpro. They formed hydrogen bonds with important surrounding residues in both proteins, including H40, L127, T142, A144, T145, H161, I162, G163, and G164. In addition, their effective concentrations against rhabdomyosarcoma (RD) cell infection by EV-A71 and CV-A16 were determined. EGCG had the highest half maximal effective concentration (EC50) of 12.86 ± 1.30 µM for EV-A71 and 15.54 ± 1.50 µM for CV-A16, while diosmin had EC50 of 21.01 ± 1.57 µM for EV-A71 and 30.68 ± 3.25 µM for CV-A16. Both compounds were non-toxic in RD cells, with 50% cytotoxic concentrations of > 100 µM for EGCG and > 500 µM for diosmin. In addition, MD simulation analysis showed that EGCG had a higher binding affinity than diosmin, supported by its significantly lower solvated binding free energies and greater numbers of contact atoms and key binding residues. Moreover, previous studies reported EGCG’s inhibitory effect on other viruses, such as severe acute respiratory syndrome coronavirus 2. Therefore, our findings suggest that EGCG can effectively inhibit the EV-A71 and CV-A16 3Cpro.