Dendritic cells pulsed with hepatitis C virus NS3 protein induce immune responses and protection from infection with recombinant vaccinia virus expressing NS3

Abstract

Infections with Hepatitis C virus (HCV) pose a serious health problem worldwide. In this study, the hypothesis that adoptive transfer of dendritic cells (DCs) pulsed with HCV NS3 protein and matured with an oligodeoxynucleotide (ODN) containing CpG motifs (CpG) ex vivo would initiate potent HCV-specific protective immune responses in vivo was tested. NS3 protein was efficiently transduced into DCs and treatment of DCs with CpG ODN induced phenotypic maturation and specifically increased the expression of CD40. DCs matured with CpG ODN produced higher interleukin 12 levels and a stronger allogeneic T-cell response compared with untreated DCs. Notably, there were no differences between NS3-pulsed DCs and DCs pulsed with a control protein with respect to phenotype, cytokine production or mixed lymphocyte reaction, indicating that transduction with NS3 protein did not impair DC functions. Compared with the untreated NS3-pulsed DCs, the NS3-pulsed DCs matured with CpG ODN induced stronger cellular immune responses including enhanced cytotoxicity, higher interferon-c production and stronger lymphocyte proliferation. Upon challenge with a recombinant vaccinia virus expressing NS3, all mice immunized with NS3-pulsed DCs showed a significant reduction in vaccinia virus titres when compared with mock-immunized mice. However, the NS3-pulsed DCs matured with CpG ODN induced higher levels of protection compared with the untreated NS3-pulsed DCs. These data are the first to show that NS3-pulsed DCs induce specific immune responses and provide protection from viral challenge, and also demonstrate that CpG ODNs, which have a proven safety profile, would be useful in the development of DC vaccines. INTRODUCTIONInfections with Hepatitis C virus (HCV) pose a serious health problem worldwide. An estimated 170 million people are currently infected, which amounts to 3 % of the world population. About 80 % of infected people remain chronic carriers and are at high risk of developing severe liver disease including cirrhosis and hepatocellular carcinoma (WHO, 1999). Treatment with interferon-a (IFN-a) and ribavirin is the only effective therapy against HCV infection, but the overall response rate is only 40-50 % (Fried et al., 2002;Manns et al., 2001). Currently, there is no licensed vaccine against HCV; therefore, developing strategies for vaccination as well as for treatment of HCV infection is of great importance.Dendritic cells (DCs) are the most potent type of antigen presenting cells (APCs) and are responsible for the initiation and maintenance of immune responses. Situated in peripheral tissues and in lymphoid organs, DCs are uniquely suited to detect and capture pathogens. They express members of the recently identified Toll-like receptor (TLR) family, which bind common chemical moieties associated with microbial organisms. TLR ligands include bacterial lipopolysaccharide, lipopeptides, hypomethylated CpG DNA motifs, dsRNA and flagellin (Akira et al., 2001;Iwasaki & Medzhitov, 2004). TLR signalling triggers a maturation program...

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“…Yu et al 87 employed DOTAP as a liposomal transfection reagent. Kuzushita et al 89 used Pep1, a short amphiphatic peptide carrier that delivers peptides and proteins into cells without denaturation or crosslinking, and Matsui et al 86 applied an anthrax toxinmediated antigen-delivery system.…”

“…Yu et al 87 performed immunizations with bone marrow-derived DCs that were transduced with the HCV-NS3 protein. Immunizations with the primed DCs elicited substantial NS3-specifi c cytotoxic immune responses and significant IFN-γ secretion, although the responsible cell type was not identifi ed.…”