Discovery of Immunologically Inspired Small Molecules That Target the Viral Envelope Protein

Dengue virus is a major human pathogen that infects over 390 million people annually leading to approximately 500 000 hospitalizations due to severe dengue. Since the only marketed vaccine, Dengvaxia, has recently been shown to increase disease severity in those lacking natural immunity, antivirals to prevent or treat dengue  infection represent a large, unmet medical need. Small molecules that target the dengue virus envelope protein, E, on the surface of the virion could act analogously to antibodies by engaging E extracellularly to block infection; however, a shortage of target-based assays suitable for screening and medicinal chemistry studies has limited efforts in this area. Here we demonstrate that the dengue E protein offers a tractable drug target for preventing dengue infection by developing a target-based assay using a recombinantly expressed dengue serotype 2 E protein. We performed a high-throughput screen of ∼20 000 compounds followed by secondary assays to confirm target-binding and antiviral activity and counter-screens to exclude compounds with nonspecific activities. These efforts yielded eight distinct chemical leads that inhibit dengue infection by binding to E and preventing E-mediated membrane fusion with potencies equal to or greater than previously described small molecule inhibitors of E. We show that a subset of these compounds inhibit viruses representative of the other three dengue serotypes and Zika virus. This work provides tools for discovery and optimization of direct-acting antivirals against dengue E and shows that this approach may be useful in developing antivirals with broad-spectrum activity against other flavivirus pathogens.