Production and purification of virus-like particles of different enterovirus subtypes as vaccines

Abstract Enterovirus A71 (EV-A71) and coxsackievirus A16 (CV-A16), both belonging to enterovirus species A, are the major etiological agents for hand-foot-and-mouth-disease that causes severe neurological complications and death in young children. Since prevalent EV-A71 subgenotypes change, developing bivalent vaccines composed of the epidemic EV-A71 subgenotype and CV-A16 is desirable. Virus-like particles (VLPs) of enteroviruses can be produced by co-expressing P1 polyprotein and 3CD protease and are promising vaccine platforms, but VLP production and purification issues are rarely addressed. To produce VLPs of different enterovirus subtypes as vaccines, we exploited the flashBAC GOLD™ system to generate 4 baculoviruses expressing P1 and 3CD derived from different prevalent enterovirus subtypes (EV-A71 C2, C4 and B5 and CV-A16). By fine-tuning the strategies to infect High Five™ cells, we achieved high level production of VLPs derived from EV-A71 C2, C4, B5 and CV-A16, reaching 195, 85, 157 and 173 mg/L, respectively. We further purified and characterized these 4 subtypes of VLPs. Immunization of mice with VLPs of subtypes that were predominantly prevalent over the last decade (C4 and CV-A16), alone or in combination, successfully induced humoral and cellular immune responses. These data demonstrated the potential of such baculovirus/insect cell platform for timely, swift and mass production of VLP vaccines against epidemic enterovirus strains.