Key result
High-throughput screening identified MPPI and 2-MPO as 2C helicase inhibitors, with MPPI achieving effective foot-and-mouth disease virus suppression at 160 μM.
Why the study?
The foot-and-mouth disease virus 2C protein drives viral genome replication and serves as a critical target for antiviral drug development.
A FRET-based high-throughput screening platform identified MPPI as a potent inhibitor of FMDV 2C helicase, offering a potential therapeutic strategy against picornaviruses.
Background: The 2C protein of foot-and-mouth disease virus (FMDV), a member of helicase superfamily 3 (SF3), drives viral genome replication and serves as a critical target for antiviral drug development. Methods: A fluorescence resonance energy transfer (FRET)-based high-throughput screening (HTS) platform was developed to identify 2C helicase inhibitors. Primary screening evaluated 4424 compounds for helicase inhibition. Molecular docking analyzed inhibitor interactions with the N207 residue within the catalytic core and helicase inhibition assays classified the inhibitor type (mixed, competitive, noncompetitive). Differential scanning fluorimetry (nanoDSF) quantified 2C thermal destabilization. Antiviral activity was assessed via indirect immunofluorescence, RT-qPCR, and plaque reduction assays. Results: Six compounds inhibited 2C helicase activity at >620 μM. Molecular docking revealed hydrogen bonding, hydrophobic interactions, and π-cation stabilization at the catalytic core. 2-MPO and MPPI were classified as mixed-type inhibitors, 5-TzS− and 2-PyOH as competitive, and DCMQ/Spiro-BD-CHD-dione as noncompetitive. NanoDSF showed a ΔTm ≥ 1.5 °C (2.5 mM compounds), with reduced destabilization in N207A mutants. Antiviral assays identified 2-MPO and MPPI as optimal inhibitors. MPPI achieved effective FMDV suppression at 160 μM, exhibiting two orders of magnitude higher potency than 2-MPO (400 μM). Conclusions: The established FRET-based HTS platform targeting 2C helicase facilitates anti-FMDV lead discovery, while 2C inhibitors may serve as an effective therapeutic strategy against other picornaviruses.
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Zhou et al. (2025) studied Foot-and-mouth disease virus (FMDV). 2C helicase inhibitors (2-MPO and MPPI) was evaluated on 2C helicase inhibition and FMDV suppression. High-throughput screening identified MPPI and 2-MPO as 2C helicase inhibitors, with MPPI achieving effective foot-and-mouth disease virus suppression at 160 μM.
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