Conjugation of Hemoglobin and Mannan Markedly Improves the Immunogenicity of Domain III of the Zika Virus E Protein: Structural and Immunological Study.

Zika virus (ZIKV) leads to congenital microcephaly and anomalies and severe neurological diseases such as Guillain-Barre syndrome. Safe and effective vaccines are necessitated to deal with these severe health threats. As an ideal antigen, the domain III of the envelope protein (EDIII) of ZIKV can evoke potent neutralizing antibodies without any antibody-dependent enhancement (ADE) effect. However, EDIII necessitates to be formulated with an antigen delivery system or adjuvants to improve its immunogenicity. Hemoglobin (Hb) regulates inflammation, cytokine levels, and activate macrophage. Mannan is a polysaccharide of the fungal cell wall with an immunomodulatory activity. In this study, EDIII was conjugated with Hb and mannan, using the disulfide bond as the linker. Hb and mannan both functioned as the adjuvants. Conjugation of Hb and mannan acted as the delivery system for EDIII. The structure of EDIII was essentially maintained upon conjugation of Hb and mannan. The intracellular release of EDIII from the conjugate (HM-EDIII-2) was achieved by reduction of the glutathione-sensitive disulfide bond. As compared with EDIII, HM-EDIII-2 elicited high EDIII-specific IgG titers and high levels of Th1-type cytokines (IFN-γ and IL-2) and Th2-type cytokines (IL-5 and IL-10), along with no apparent toxicity to the organs. Moreover, the pharmacokinetic study revealed a prolonged serum exposure of HM-EDIII-2 to the immune cells. Thus, HM-EDIII-2 could boost a strong humoral and cellular immune response to EDIII. Our study was expected to provide the feasibility necessary to develop a robust and potentially safe ZIKV vaccine.