Inhibitors of foot and mouth disease virus targeting a novel pocket of the RNA-dependent RNA polymerase

Authors: DURK, RYAN - University Of Missouri; SINGH, KAMALENDRA - University Of Missouri; CORNELISON, CEILI - University Of Missouri; RAI, DEVENDRA - University Of Missouri; MATZEK, KAYLA - University Of Missouri; LESLIE, MAXWELL - University Of Missouri; Schafer, Elizabeth; MARCHAND, BRUNO - University Of Missouri; ADEDEJI, ADEYEMI - University Of Missouri; MICHAILIDIS, ELEFTHERIOS - University Of Missouri; DORST, CHRISTOPHER - University Of Missouri; MORAN, JENNIFER - University Of Missouri; PAUTLER, CHRISTIE - University Of Missouri; Rodriguez, Luis; MCINTOSH, MARK - University Of Missouri; Rieder, Aida - Elizabeth; SARAFIANOS, STEFAN - University Of Missouri

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/25/2010
Publication Date: 12/21/2010
Citation: Durk, R.C., Singh, K., Cornelison, C.A., Rai, D.K., Matzek, K.B., Leslie, M.D., Schafer, E.A., Marchand, B., Adedeji, A., Michailidis, E., Dorst, C.A., Moran, J., Pautler, C., Rodriguez, L.L., Mcintosh, M.A., Rieder, A.E., Sarafianos, S.G. 2010. Inhibitors of foot and mouth disease virus targeting a novel pocket of the RNA-dependent RNA polymerase. PLoS One. DOI: 10.1371/journal.pone.0015049.

Interpretive Summary: Foot-and-mouth disease (FMD) is a highly contagious viral disease of cloven-hoofed animals that can lead to severe losses in the livestock production and export industries. Although vaccines have been extensively applied to control FMD, there is no antiviral therapy available to treat ongoing infections with FMD virus (FMDV). The purpose of this study was to identify and evaluate the efficacy of a chemical library of compounds for their inhibitory activity against the FMDV polymerase, 3Dpol. After the initial screening of the library, nine compounds were selected for further investigation based on their activity at suppressing FMDV 3Dpol-mediated RNA synthesis. Selected compounds were further characterized for their ability to inhibit FMDV replication in tissue culture. Among these compounds, C5D9 was the most active to inhibit virus yield and appeared to perform at superior levels to that of ribaviridin (a known inhibitor of Picornavirus polymerases). Considering the agricultural and socio-economic importance of FMD and need for early disease interventions, the results herein provide optimism that polymerase inhibitors be further investigated for therapeutic development against FMD.

Technical Abstract: Foot-and-Mouth Disease Virus (FMDV) is a positive stranded picornavirus which can infect cloven-hoofed animals, such as cattle, pigs and sheep, and lead to severe losses in livestock production. Unlike emergency vaccination which requires at least 7 days to trigger an effective immune response,antiviral drugs may have prophylactic and/or therapeutic effects almost immediately. However, there are currently no approved FMDV inhibitors. FMDV RNA-dependent RNA polymerase (3Dpol) is an attractive target for chemotherapeutic intervention because of its critical role in viral replication. We used a high-throughput assay to identify inhibitors of 3Dpol. RNA polymerase activity was monitored by a luciferase-based assay which measures luminescence associated with release of the pyrophosphate product during RNA synthesis. We used this assay to test compounds from the Maybridge Hitfinder chemical library in a 96-well format. Two compounds, 5-chloro-3-(thiophen-2-yl-sulfanylmethyl)-1-benzothiophene 1,1-dioxide (or C7F8) and N'1-thieno[2,3-d]pyrimidin-4-yl-4-chloro-1-benzenesulfonohydrazide (or C5D9) inhibited the RNA-dependent RNA polymerase activity of 3Dpol with IC50 values of 2.5 uM and10 µM respectively, and were chosen for further investigation. The inhibitors appeared to be specific for 3Dpol as they did not affect nucleic acid synthesis by another viral polymerase under the same conditions. Furthermore, these compounds were not cytotoxic at 20 uM as determined by the XTT assay. Compound, C5D9, suppressed virus production in FMDV-infected cells with 50% and 90% effective concentrations (EC^50 and EC^90) of 10uM and 20 uM, respectively. The results indicate that 3Dpol inhibitors merit exploration as alternative or supplementary options to contain future outbreaks of FMD.