Beyond Basicity: Discovery of Nonbasic DENV‑2 Protease Inhibitors with Potent Activity in Cell Culture

The viral serine protease NS2B-NS3
is one of the promising targets
for drug discovery against dengue virus and other flaviviruses. The
molecular recognition preferences of the protease favor basic, positively
charged moieties as substrates and inhibitors, which leads to pharmacokinetic
liabilities and off-target interactions with host proteases such as
thrombin. We here present the results of efforts that were aimed specifically
at the discovery and development of noncharged, small-molecular inhibitors
of the flaviviral proteases. A key factor in the discovery of these
compounds was a cellular reporter gene assay for the dengue protease,
the DENV2proHeLa system. Extensive structure–activity relationship
explorations resulted in novel benzamide derivatives with submicromolar
activities in viral replication assays (EC50 0.24 μM),
selectivity against off-target proteases, and negligible cytotoxicity.
This structural class has increased drug-likeness compared to most
of the previously published active-site-directed flaviviral protease
inhibitors and includes promising candidates for further preclinical
development.