Novel NS2B/NS3-protease dengue virus inhibitors

Abstract

Dengue disease, caused by the virus of the same name, is expanding globally with an annual estimate of 390 million people infected. However, available antivirals have been ineffective. Therefore, the search of small-molecule therapeutic candidates is a recurrent practice. The dengue virus (DENV) NS2B/NS3 protease complex is a recognized target for the design of specific antivirals, due to its importance in the replication and high degree of sequence conservation. The aim of this research was to find molecules capable of inhibiting this protein complex. A total of 210,903 molecules, from the PubChem database, were docked in silico to the NS2B/NS3 crystallographic structure (PDB: 2FOM). The recombinant protease was expressed in E. coli and purified by affinity chromatography. Five of the best 500 leading compounds, according to the virtual screening affinity values (-11.6 and -13.5 kcal/mol), were purchased. The inhibitory activity of these molecules by the protease was evaluated in vitro, using the action of the recombinant enzyme on the Boc-Gly-Arg-Arg-AMC substrate (IC50), and antiviral assays for the detection of NS1 viral titles in infected human hepatocytes. Compounds CID54681617, CID54692801, and CID54715399 showed to be strong inhibitors of NS2B/NS3, with IC50 values (μM) and percentages of NS1 viral title reductions corresponding to 19.9, 79.9%; 17.5, 69.8%; 9.1, 73.9 %, respectively. Multivariate methods on molecular descriptors showed that two of these compounds are structurally different from DENV inhibitors reported by other authors. This discovery opens new possibilities to obtain drug candidates against Dengue.