Evaluation of Zika virus DNA vaccines based on NS1 and domain III of E

Background: A large-scale outbreak of Zika virus (ZIKV) has occurred in Brazil and other South American countries, and has rapidly spread to 60 countries and regions worldwide since 2015, but no approved anti-ZIKV vaccines are available as of 2021.Methods: We developed four types of anti-ZIKV DNA vaccine candidates: VPC-NS1, VPC-prME, VPC-prME-NS1, and VPC-EIII-NS1. They were developed against the structural proteins prM and E, and non-structural protein 1 (NS1) of ZIKV using the mammalian cell expression vector pcDNA3.1(+) as the backbone. For immunization, we intramuscularly injected mice with each vaccine candidate (n = 12 to 15 per group) on day 0 and day 14, with mice injected with phosphate-buffered saline (PBS) and pcDNA3.1(+) backbone vector as controls. On day 7, 21, and 35 after initial immunization, the effect of DNA vaccines was evaluated by ZIKV-specific humoral immunity determined by enzyme-linked immunosorbent assay (ELISA), ZIKV-specific T cell immunity determined by intracellular cytokine staining by flow cytometry and serum neutralization capacity determined by plaque reduction neutralization test (PRNT50) assay.Results: The sequencing results showed that DNA vaccine vectors were successfully constructed. Western blotting and immunofluorescence results demonstrated the successful expression of immunogens carried by the DNA vaccines. On day 21 and 35 after the initial immunization, the levels of serum total immunoglobulin (Ig)G in all vaccine-given groups were slightly higher (approximately 1.5- to 2-fold) than those in the control groups. By contrast, ZIKV-specific IgG levels of all vaccine-given groups were significantly higher (approximately 10- to 1000- fold) than those of the control groups. The PRNT50 assay showed that the average serum dilution factors for neutralizing half ZIKV virions from vaccine-given groups were at least 32-fold (highest, 93-fold), while the sera from control group showed no protection. For cellular immunity, the proportions of CD11b+ myeloid cells, CD19+ B lymphocytes and CD3+ T lymphocytes in the mouse spleens as well as the percentages of CD4+ and CD8+ subsets of T cell were not changed 35 days after initial immunization. By contrast, the proportions of ZIKVspecific CD4+T cell and CD8+T cell in all vaccine-given groups were 2- to 10-folds and 2- to 30-fold than those in the control groups, respectively.Conclusion: All four DNA vaccines designed for the ZIKV induced neutralizing IgGs and cellular immune responses against ZIKV. Particularly, VPC-EIII-NS1 induced high level of humoral response comparable to the vaccine candidate containing prM, E and NS1 polyprotein, suggesting a potent reduced ADE effect and reserved neutralizing activity. Our findings may provide guidance for improving safety of anti-ZIKV vaccines in the future.