Policresulen, a novel NS2B/NS3 protease inhibitor, effectively inhibits the replication of DENV2 virus in BHK-21 cells.

AIM: Dengue is a severe epidemic disease caused by dengue virus (DENV) infection, for which no effective treatment is available. The protease complex, consisting of nonstructural protein 3 (NS3) and its cofactor NS2B, plays a pivotal role in the replication of DENV, thus may be a potential target for anti-DENV drugs. Here, we report a novel inhibitor of DENV2 NS2B/NS3 protease and its antiviral action. METHODS: An enzymatic inhibition assay was used for screening DENV2 NS2B/NS3 inhibitors. Cytotoxicity to BHK-21 cells was assessed with MTT assay. Antiviral activity was evaluated in BHK-21 cells transfected with Rlu-DENV-Rep. The molecular mechanisms of the antiviral action was analyzed using surface plasmon resonance, ultraviolet-visible spectral analysis and differential scanning calorimetry assays, as well as molecular docking analysis combined with site-directed mutagenesis. RESULTS: In our in-house library of old drugs (~1000 compounds), a topical hemostatic and antiseptic 2-hydroxy-3,5-bis[(4-hydroxy-2-methyl-5-sulfophenyl)methyl]-4-methyl-benzene-sulfonic acid (policresulen) was found to be a potent inhibitor of DENV2 NS2B/NS3 protease with IC50 of 0.48 µg/mL. Furthermore, policresulen inhibited DENV2 replication in BHK-21 cells with IC50 of 4.99 µg/mL, whereas its IC50 for cytotoxicity to BHK-21 cells was 459.45 µg/mL. Policresulen acted as a competitive inhibitor of the protease, and slightly affected the protease stability. Using biophysical technology-based assays and molecular docking analysis combined with site-directed mutagenesis, we demonstrated that the residues Gln106 and Arg133 of DENV2 NS2B/NS3 protease directly interacted with policresulen via hydrogen bonding. CONCLUSION: Policresulen is a potent inhibitor of DENV2 NS2B/NS3 protease that inhibits DENV2 replication in BHK-21 cells. The binding mode of the protease and policresulen provides useful hints for designing new type of inhibitors against the protease.

Raddeanalin, a new flavonoid glycoside from the leaves of Salix raddeana Laksh.

A new flavonoid glycoside, diosmetin 7-O-beta-D-xylopyranosyl(1-6)-beta-D-glucopyranoside, named raddeanalin (1), was isolated from the ethanolic extract of the leaves of Salix raddeana Laksh. together with three known compounds, kaempferol 3-O-glucoside (2), quercetin 3-O-glucoside (3) and quercetin 3-O-rutinoside (4). The structure of new compound was established by the spectral data and chemical properties.