DNA Prime-Adenovirus Boost Immunization Induces a Vigorous and Multifunctional T-Cell Response against Hepadnaviral Proteins in the Mouse and Woodchuck Model

Abstract

Induction of hepatitis B virus (HBV)-specific cytotoxic T cells by therapeutic immunization may be a strategy to treat chronic hepatitis B. In the HBV animal model, woodchucks, the application of DNA vaccine expressing woodchuck hepatitis virus (WHV) core antigen (WHcAg) in combination with antivirals led to the prolonged control of viral replication. However, it became clear that the use of more potent vaccines is required to overcome WHV persistence. Therefore, we asked whether stronger and more functional T-cell responses could be achieved using the modified vaccines and an optimized prime-boost vaccination regimen. We developed a new DNA plasmid (pCGWHc) and recombinant adenoviruses (AdVs) showing high expression levels of WHcAg. Mice vaccinated with the improved plasmid pCGWHc elicited a stronger WHcAg-specific CD8 + T-cell response than with the previously used vaccines. Using multicolor flow cytometry and an in vivo cytotoxicity assay, we showed that immunization in a DNA prime-AdV boost regimen resulted in an even more vigorous and functional T-cell response than immunization with the new plasmid alone. Immunization of naïve woodchucks with pCGWHc plasmid or AdVs induced a significant WHcAg-specific degranulation response prior to the challenge, this response had not been previously detected. Consistently, this response led to a rapid control of infection after the challenge. Our results demonstrate that high antigen expression levels and the DNA prime-AdV boost immunization improved the T-cell response in mice and induced significant T-cell responses in woodchucks. Therefore, this new vaccination strategy may be a candidate for a therapeutic vaccine against chronic HBV infection.

Novel Woodchuck Hepatitis Virus (WHV) Transgene Mouse Models Show Sex-Dependent WHV Replicative Activity and Development of Spontaneous Immune Responses to WHV Proteins

“…This finding is consistent with the low level of expression of the WHV x gene in the woodchuck liver during natural infection (27). The detection of WHV proteins has failed so far, as no suitable antibodies were available (15,25,28,29).…”

“…The possibility of breaking immune tolerance to WHcAg in Tg mice allows the testing of different therapeutic vaccines and vaccination regimens. Recently, an immunization protocol including priming with DNA vaccines and boosts with adenoviral vectors was shown to elicit robust and effective humoral and cellular immune responses against WHV in woodchucks (25,38). This protocol was also evaluated in Tg mice and was superior to DNA vaccination alone (A. D. Kosinska, M. Lu, and M.…”

“…Plasmids pWHsIm and pWHcIm containing the WHV preS2-S or core gene, respectively, under the control of the cytomegalovirus (CMV) promoter, were described previously (24). Plasmid pCGWHc containing the WHV core gene downstream of the ␤-globin intron sequence, under the control of the cytomegalovirus (CMV) promoter, was described previously by Kosinska et al (25). Immunization of mice was performed according to a procedure described previously by Schirmbeck et al (26).…”

Prime/Boost Immunization with DNA and Adenoviral Vectors Protects from Hepatitis D Virus (HDV) Infection after Simultaneous Infection with HDV and Woodchuck Hepatitis Virus

“…BHK-21 cells (baby hamster kidney cells; ATCC CCL-10) were handled as described previously (7), and 10 4 cells were transfected with 1 g of pcDNA3p24 or pcDNA3p27 using the Lipofectamine reagent (Invitrogen, Karlsruhe, Germany) according to the manufacturers' instructions. For Western blot analysis, cultured cells were handled as usual (7). Expression of HDAgp27 was verified using a polyclonal human anti-HDV serum (Fig.…”

“…In this study, we intended to improve the strength of the CD8 ϩ T cell response by a DNA prime and adenoviral boost immunization protocol. Recombinant adenoviruses have been shown to induce vigorous T cell responses in many viral infections such as Ebola virus, simian immunodeficiency virus, and WHV (7,11,12). Recently, the adenoviral vector approach was shown to induce sustained T cell responses against hepatitis C virus in healthy human volunteers (13).…”