Exploring the Pharmacological Mechanism of the Widely Used Huashi Baidu Decoction against Severe Acute Respiratory Syndrome Coronavirus 2 by in silico Simulations and in vitro Assays

Abstract Huashi Baidu decoction (HSBD) is an approved traditional Chinese medicine formula that has demonstrated clinical efficacy against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but its therapeutic mechanisms remain incompletely elucidated. The purpose of this study is to explore the pharmacological mechanism of HSBD against SARS-CoV-2. In this work, potential target proteins of HSBD were predicted by network pharmacology, deep learning, and ligand-based approaches. Molecular docking was performed to screen natural products with high inhibitory potential against predicted viral targets. The antiviral activities of the screened candidates were validated by enzymatic inhibition assays, followed by molecular dynamics (MD) simulations to explore the interaction mechanism between active compounds and the viral targets. Our results revealed that HSBD could potentially interact with the host proteins, including TP53, HSP90AA1, STAT3, MCL1, EGFR, and GSK3B, and suppress viral replication and inflammation by modulating the PI3K-Akt and MAPK signaling pathways. Furthermore, SARS-CoV-2 main protease (Mpro) and papain-like protease (PLpro) were predicted to be key antiviral targets of HSBD by deep learning and ligand-based approaches, followed by in vitro enzymatic inhibition assays, which confirmed that glabrol inhibits Mpro, licoflavone B inhibits PLpro, and isobavachromene acts as a dual inhibitor. MD simulations identified Q110 in Mpro and Y268 in PLpro as critical residues contributing to ligand binding. In summary, HSBD exerts its therapeutic effects against SARS-CoV-2 through direct antiviral action and the regulation of multiple host targets and signaling pathways. Those bioactive HSBD compounds with Mpro or PLpro inhibitory activities could serve as candidates for anti-SARS-CoV-2 drug development.