nsP2 Protease\nInhibitor Blocks the Replication of\nNew World Alphaviruses and Offer Protection in Mice

New World alphaviruses, including Venezuelan equine encephalitis\nvirus (VEEV), eastern equine encephalitis virus (EEEV), and western\nequine encephalitis virus (WEEV), are mosquito-transmitted viruses\nthat cause disease in humans. These viruses are endemic to the western\nhemisphere, and disease in humans may lead to encephalitis and long-term\nneurological sequelae. There are currently no FDA-approved vaccines\nor antiviral therapeutics available for the prevention or treatment\nof diseases caused by these viruses. The alphavirus nonstructural\nprotein 2 (nsP2) functions as a protease, which is critical for the\nestablishment of a productive viral infection by enabling accurate\nprocessing of the nsP123 polyprotein. Owing to the essential role\nplayed by nsP2 in the alphavirus infectious process, it is also a\nvaluable therapeutic target. In this article, we report the synthesis\nand evaluation of novel small molecule inhibitors that target the\nalphavirus nsP2 protease via a covalent mode of action. The two lead\ncompounds demonstrated robust inhibition of viral replication <i>in vitro</i>. These inhibitors interfered with the processing\nof the nsP123 polyprotein as determined using VEEV TC-83 as a model\npathogen and are active against EEEV and WEEV. The compounds were\nfound to be nontoxic in two different mouse strains and demonstrated\nantiviral activity in a VEEV TC-83 lethal challenge mouse model. Cumulatively,\nthe outcomes of this study provide a compelling rationale for the\npreclinical development of nsP2 protease inhibitors as direct-acting\nantiviral therapeutics against alphaviruses.