A DNA Vaccine Expressing Fusion Protein E2-NT(gp96) Induces Hepatitis C Virus Cross-Neutralizing Antibody in BALB/c Mice

摘要

Background: Development of an effective prophylactic vaccine is the optimal long-term goal for the eventual control of HCV infection. An effective HCV vaccine should be able to elicit neutralizing antibodies (NAbs). Glycoprotein E2 of HCV is the major target forNAbs.Methods: In this study, we designed and constructed a DNA vaccine (pcDNA-E2-NT(gp96)) encoding a fusion protein composed ofHCV E2 ectodomain (genotype 1a) and N-terminal domain of gp96 as a biological adjuvant. Two possible forms of a fusion protein,namely E2-NT(gp96) and NT(gp96)-E2, were made and subjected to in silico modeling and analysis. After the selection of the bestform and confirmation its expression capacity in COS-7 cells, recombinant pcDNA-E2-NT(gp96) plasmid was generated by cloning oftarget genes into pcDNA3.1(+) plasmid. Constructed DNA vaccine immunogenicity was evaluated in BALB/c mice by measurementspecific antibodies by ELISA and their neutralization capacity by neutralization assay.Results: In silico modeling and analysis showed that the E2-NT(gp96) structure was more valid than NT(gp96)-E2. Docking resultrevealed that the selected fusion protein had a high tendency for interaction with the main receptor (CD81) of HCV. GFP expressionin COS-7 cells confirmed the E2-NT(gp96) expression capacity. Restriction enzyme digestion and sequencing results confirmed theintegrity of the constructed plasmid. ELISA results showed that the pcDNA-E2-NT(gp96) induced high titers of specific antibodies inimmunized mice. The sera of immunized mice cross-neutralized JFH1/HCVcc genotype 2a by 55% relative to pre-immune sera.Conclusions: Total results showed that the generated DNA vaccine induced potent immune responses in immunized mice. Therefore, our findings are sufficiently encouraging to propose the pcDNA-E2-NT(gp96) as a promising vaccine candidate for HCV infection.