An alphavirus vaccine development utilizing RNA replication-defective strategy.

Alphaviruses are arthropod-borne viruses that cause widespread disease. However, many pathogenic alphaviruses are classified as risk group 3 human pathogens, which hampers the development of vaccines and therapeutic agents targeting alphavirus infections. In this study, we developed an RNA replication-defective Venezuelan equine encephalitis virus that has a complete nsP4 gene deletion (VEEV-ΔnsP4). A BHK cell line expressing nsP4 (BHKnsP4) was selected for trans-complementation to support the replication cycle of VEEV-ΔnsP4. The VEEV-ΔnsP4 replicates only in BHKnsP4 cells and is immunologically similar to its parental wild type. Significantly, VEEV-ΔnsP4 is highly attenuated and a single dose immunization can protect mice from a lethal challenge. Furthermore, the RNA replication-defective vaccine strategy has been successfully employed for Chikungunya virus (CHIKV), Western equine encephalitis virus (WEEV) and Eastern equine encephalitis virus (EEEV). Overall, our study highlights the potential of the nsP4 trans-complementation system as a safe and effective platform for alphavirus vaccines development.