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Newcastle disease (ND) is still one of the major plagues of birds worldwide. Combat actions are limited to vaccines, highlighting the urgent need for new and amply available antiviral drugs. Previous results have shown that Newcastle disease virus (NDV) downregulates the intracellular Raf kinase inhibitor protein (RKIP) expression for efficient replication, suggesting that this molecular may be a suitable target for antiviral intervention. In the present work, we investigated whether or not the Raf kinase inhibitor V (RKIV), which functions in the same way as RKIP by targeting the intracellular Raf kinase, is able to suppress the propagation of enzootic virulent NDV in vitro and in vivo. In vitro antiviral activity of RKIV was assessed by cell-based assay, and in vivo activity was determined in the chicken model. Our results clearly showed that RKIV treatment protected the cells from NDV-induced CPE with the effective concentrations on nM level, and inhibited virus replication in the lungs of infected chickens in a dose-dependent manner and protected chickens from the lethal infection by NDV. Thus, we conclude that the Raf kinase inhibitor compound RKIV, by inhibiting the host cellular target Raf kinase, might be very promising as a new class of antivirals against the enzootic virulent NDV infection.RKIV protects cells from NDV-induced CPE at effective concentrations in the nM range.RKIV treatment reduces NDV titers isolated from the lungs of infected chickens in a dose-dependent manner.RKIV treatment protects chickens from lethal NDV inoculation.No toxic side effects of RKIV are found in vitro and in vivo with suitable doses and treatment period.