Anti-SARS-CoV-2 activities in vitro of Shuanghuanglian preparations and bioactive ingredients

Abstract

Human infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) and there is no cure currently. The 3CL protease (3CLpro) is a highly conserved protease which is indispensable for CoVs replication, and is a promising target for development of broad-spectrum antiviral drugs. In this study we investigated the anti-SARS-CoV-2 potential of Shuanghuanglian preparation, a Chinese traditional patent medicine with a long history for treating respiratory tract infection in China. We showed that either the oral liquid of Shuanghuanglian, the lyophilized powder of Shuanghuanglian for injection or their bioactive components dose-dependently inhibited SARS-CoV-2 3CLpro as well as the replication of SARS-CoV-2 in Vero E6 cells. Baicalin and baicalein, two ingredients of Shuanghuanglian, were characterized as the first noncovalent, nonpeptidomimetic inhibitors of SARS-CoV-2 3CLpro and exhibited potent antiviral activities in a cell-based system. Remarkably, the binding mode of baicalein with SARS-CoV-2 3CLpro determined by X-ray protein crystallography was distinctly different from those of known 3CLpro inhibitors. Baicalein was productively ensconced in the core of the substrate-binding pocket by interacting with two catalytic residues, the crucial S1/S2 subsites and the oxyanion loop, acting as a “shield” in front of the catalytic dyad to effectively prevent substrate access to the catalytic dyad within the active site. Overall, this study provides an example for exploring the in vitro potency of Chinese traditional patent medicines and effectively identifying bioactive ingredients toward a specific target, and gains evidence supporting the in vivo studies of Shuanghuanglian oral liquid as well as two natural products for COVID-19 treatment.

Scutellaria baicalensis Flavones as Potent Drugs against Acute Respiratory Injury during SARS-CoV-2 Infection: Structural Biology Approaches

“…Baicalein presented a high affinity for the viral 3CLpro, with the K d measured by isothermal titration calorimetry being 4.03 µM and the calculated ligand efficiency being 0.37 kcal/mol [40]. The crystal structure determined by Su et al [40] comprises four 3CLpro protomers that assemble as two dimers, with each protomer being bound by a baicalein molecule. Structurally, a 3CLpro protomer comprises domains I (residues 10 to 99), II (residues 100 to 182), and III (residues 198 to 303); Glu166 residues are important in shaping S1 pockets of substrate binding sites from the catalytically active 3CLpro dimer [65].…”

“…Several studies have pointed out the anti-SARS-CoV-2 effects of S. baicalensis compounds, with an emphasis on baicalein and baicalin [36][37][38][39][40], wogonin [39], and oroxylin A [39].…”

“…The interaction of flavones with 3CLpro was investigated based on the crystal structure of 3CLpro bound to baicalein-PDB:6M2N [40,52].…”

“…The structure of SARS-CoV-2 3CLpro bound to baicalein was solved by Su et al [40]. The authors identified baicalein as a novel nonpeptidomimetic compound that inhibits SARS-CoV-2 3CLpro with micromolar potency, with the IC 50 being 0.94 µM.…”

“…The authors identified baicalein as a novel nonpeptidomimetic compound that inhibits SARS-CoV-2 3CLpro with micromolar potency, with the IC 50 being 0.94 µM. Baicalein presented a high affinity for the viral 3CLpro, with the K d measured by isothermal titration calorimetry being 4.03 µM and the calculated ligand efficiency being 0.37 kcal/mol [40]. The crystal structure determined by Su et al [40] comprises four 3CLpro protomers that assemble as two dimers, with each protomer being bound by a baicalein molecule.…”