Ubiquitin conjugation of hepatitis B virus core antigen DNA vaccine leads to enhanced cell-mediated immune response in BALB/c mice

Abstract

Background: Nearly 350 million persons worldwide are chronically infected with hepatitis B virus (HBV). Ubiquitin (Ub) is a highly conserved small regulatory protein, ubiquitous in eukaryotes, that usually serves as a signal for the target protein that is recognised and degraded in proteasomes. The Ub-mediated processing of antigens is rapid and efficient and stimulates cell-mediated immune responses. Accordingly, Ubmediated processing of antigens has been widely used in chronic-infection and cancer studies to improve immune response. Objectives: Many clinical trials have shown that DNA vaccine potency needs to be greatly enhanced. Here, we report a new strategy for designing an HBV DNA vaccine using the ubiquitin (Ub) sequence. The aim of this study was to investigate a novel DNA vaccination, based on the expression of HBV core antigen (HBcAg), fused to Ub to enhance DNA vaccine potency. Materials and Methods: Mouse ubiquitin fused to the HBcAg gene and cloned into the eukaryotic vector pcDNA3.1 (-). BALB/c mice were immunized with recombinant pUb-HBcAg or pHBcAg DNA vaccine. Lymphocyte proliferation assay, intracellular IFN-γ assay, CTL cytotoxicity assay, and antibody assay were performed to analyze the cellular and humoral immune responses to our DNA constructs. Results: HBcAg was expressed effectively in the COS-7 cells that were transiently transfected with pUb-HBcAg. Strong anti-HBc IgG responses were elicited in mice that were immunized with pUb-HBcAg. The endpoint titers of anti-HBc peaked at 1:656100 on the 42nd day after the third immunization. pUb-HBcAg stimulated greater lymphocyte proliferation and induced higher levels of IL-2 and IFN-γ and a greater percentage of HBcAg-specific CD8+ T cells in mice than pHBcAg. In the CTL assay, the specific lysis rate reached 56.5% at an effector:target ratio of 50:1 in mice that were immunized with pUb-HBcAg. Conclusions: pUb-HBcAg elicits specific anti-HBc responses and induces HBc-specific CTL responses in immunized BALB/c mice. Our results imply that Ub can be used as a molecular adjuvant that enhances the potency of DNA vaccines. © 2011 Kowsar M.P.Co. All rights reserved.