Development Of The Novel Vaccine Against H5N1 Virus By ASP-1 Adjuvant

Objective:This study aimed to perform study on novel vaccines against H5N1 virus, combing with the adjuvanticity of the recombinant ASP-1 protein serving as a novel biological adjuvant, to develop a novel vaccine with good immunogenicity and immunoprotection against H5N1 virus.Methods:First of all, the truncated ASP-1 designed by analysis of bioinformatics was expressed in prokaryotic expression system, obtaining the recombinant ASP-1 protein. And subsequent studies on two recombinant vaccine candidates against H5N1 virus were carried out as follows:(1) Recombinant HA protein was expressed and purified in baculovirus expression system, based on HA sequence with coding optimization on HA sequence of A/Vietnam/1194/2004(H5N1) isolate. BALB/c mice were vaccinated with the recombinant HA vaccine adjuvanted with recombinant ASP-1 or conventional aluminum hydroxide adjuvant, followed by detection and evaluation of immunogenicity and immunoprotection. (2) Acquire the consensus sequence of extracellular domain of M2 protein (M2e) by analysising the conservation of H5N1 virus isolate-derived M2 protein. The M2e consensus sequence was selected as the target antigen to design the vaccine with pET32a(+) vector protein, trxA, fused with 3 tandem copies of M2e, designated as TrxA-M2e3 which was expressed and purified in E.coli expression system. BALB/c mice were vaccinated with the recombinant TrxA-M2e3 adjuvanted with recombinant ASP-1. The immunogenicity was detected and the cross protection against different clades of H5N1 viruses was evaluated by virus challenge experiment.Results and Conclusion:The recombinant antigens, HA and TrxA-M2e3, and recombinant ASP-1 adjuvant protein were expressed sucessfully in this study. Low dose of recombinant HA vaccine adjuvanted with ASP-1 can induce high titers of antigen-specific antibody and provide vaccinated mice full protection against lethal virus challenge. The recombinant TrxA-M2e3 adjuvanted with ASP-1 can also induce high titers of antibody and provide effective cross-protection against different clades of H5N1 viruses. Results from ASP-1-adjuvanted vaccination show the good adjuvanticity of recombinant ASP-1 producing Thl-excursive immune response. In conclusion, the truncated recombinant ASP-1, as a novel adjuvant, will be helpful for developing the novel vaccine against H5N1 virus. There is a potential to develop novel vaccine candidates with good immunogenicity and protection against H5N1 virus targeting to HA inducing neutralizing antibody and highly conserved M2e, combined with the adjuvanticity of ASP-1. This study laid a foundation to develop a novel vaccine against H5N1 virus.