Design and Validation of Novel Chikungunya Virus Protease Inhibitors

Abstract

dChikungunya virus (CHIKV; genus Alphavirus) is the causative agent of chikungunya fever. CHIKV replication can be inhibited by some broad-spectrum antiviral compounds; in contrast, there is very little information about compounds specifically inhibiting the enzymatic activities of CHIKV replication proteins. These proteins are translated in the form of a nonstructural (ns) P1234 polyprotein precursor from the CHIKV positive-strand RNA genome. Active forms of replicase enzymes are generated using the autoproteolytic activity of nsP2. The available three-dimensional (3D) structure of nsP2 protease has made it a target for in silico drug design; however, there is thus far little evidence that the designed compounds indeed inhibit the protease activity of nsP2 and/or suppress CHIKV replication. In this study, a set of 12 compounds, predicted to interact with the active center of nsP2 protease, was designed using target-based modeling. The majority of these compounds were shown to inhibit the ability of nsP2 to process recombinant protein and synthetic peptide substrates. Furthermore, all compounds found to be active in these cell-free assays also suppressed CHIKV replication in cell culture, the 50% effective concentration (EC 50 ) of the most potent inhibitor being ϳ1.5 M. Analysis of stereoisomers of one compound revealed that inhibition of both the nsP2 protease activity and CHIKV replication depended on the conformation of the inhibitor. Combining the data obtained from different assays also indicates that some of the analyzed compounds may suppress CHIKV replication using more than one mechanism. C hikungunya virus (CHIKV; genus Alphavirus, family Togaviridae) is the causative agent of chikungunya fever, a disease that has affected millions in the last decade. It is characterized by high fever, arthralgia, myalgia, headache, and rash. The disease is usually self-limiting; however long-lasting chronic symptoms are observed in nearly 50% of CHIKV-infected patients (1). As there is no approved vaccine or specific licensed antiviral compounds (2), the treatment of CHIKV infection is largely based on relief of symptoms.CHIKV infection can be inhibited by targeting host factors essential for virus infection. Targeting of several metabolic pathways has revealed anti-CHIKV effects of several licensed drugs (3). Compounds most likely targeting virus entry or host cell-specific components required for virus infection have been described (4-8). Several nucleosides or nucleotides, acting as pseudosubstrates for CHIKV RNA-dependent RNA polymerase and/or using another, more general mechanism of action, have shown anti-CHIKV activity (9). In addition, anti-CHIKV effects have been described for several groups of novel synthetic compounds (10-12).Computer-aided design based on molecular docking and molecular dynamics simulations and pharmacophore approach allows identifying potential in silico hits as active inhibitors for different CHIKV replicase proteins. This approach, however, requires the three-dimensional (3D) structure...

Chikungunya virus: epidemiology, replication, disease mechanisms, and prospective intervention strategies

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Characterization of β- d - N 4 -Hydroxycytidine as a Novel Inhibitor of Chikungunya Virus

“…To date, several studies have identified small-molecule inhibitors that inhibit CHIKV replication in cell culture (9)(10)(11)(12)(13)(14). Some nucleoside analogs, such as ribavirin, have also been shown to interfere with CHIKV replication.…”

Functions of Chikungunya Virus Nonstructural Proteins

“…In this study, the most active compound had an IC 50 below 1 μM and molecular docking simulation suggested protease inhibition as a possible mode of action (Jadav et al 2015). Recently, for the first time, a new panel of predicted nsP2 inhibitors was shown to inhibit both the protease activity of nsP2 as well as CHIKV replication in cell culture (Das et al 2016). …”