DNA Immunization with Hepatitis C Virus (HCV) Polycistronic Genes or Immunization by HCV DNA Priming-Recombinant Canarypox Virus Boosting Induces Immune Responses and Protection from Recombinant HCV-Vaccinia Virus Infection in HLA-A2.1-Transgenic Mice

Abstract

We studied immune responses to hepatitis C virus (HCV) genes delivered as DNA encoding the entire HCV protein coding genome in two polycistronic plasmids encoding HCV capsid-E1-E2-NS2-NS3 and HCV NS3-NS4-NS5 in HLA-A2.1-transgenic mice. Immune responses to HCV DNA prime and recombinant canarypox virus boost were also studied with the above constructs. At 8 weeks after a canarypox virus boost, the DNA prime/canarypox virus boosting regimen induced potent cellular immune responses to HCV structural and nonstructural proteins on target cells expressing the HLA-A2.1 allele. High frequencies of gamma interferonsecreting cells, as detected by enzyme-linked immunospot assay, were obtained in response to several endogenously expressed HCV proteins. We also observed cytotoxic-T-lymphocyte reactivity in response to endogenously expressed HCV proteins in fresh spleen cells without in vitro expansion. Upon challenge with a recombinant vaccinia virus expressing HCV proteins at 2 months postimmunization, the HCV DNA prime/ canarypox virus-immunized mice showed a complete reduction in vaccinia virus titers compared to HCV DNA prime/boost-and mock-immunized controls. Immune responses were still detectable 4 months after canarypox virus boost in immunized mice. Interestingly, at 10 months postimmunization (8 months after canarypox virus boost), the protection in HCV DNA prime/boost-immunized mice against recombinant HCV-vaccinia virus challenge was higher than that observed in HCV DNA prime/canarypox virus boost-immunized mice.About 80% of human hepatitis C virus (HCV) infections become chronic with lifelong viremia. It is estimated that there are more than 200 million chronically HCV-infected carriers in the world, 4 million of whom are in the United States (3). Treatment of the carrier state is at present not very satisfactory. HCV is a notoriously difficult target for immunization: it exists in numerous genotypes and within an infected host in numerous quasispecies (5). It is therefore desirable to develop a strategy that will induce strong immune responses to several variable and conserved regions of the virus.Immunization against HCV will involve the generation of cell-mediated immunity to mediate killing or downregulation of infected cells (7,11). An effective way of generating cellmediated immunity is by DNA-based immunization. Numerous approaches are available for enhancing DNA-based immune responses, including the coadministration of cytokine expressing plasmids (9, 14), the use of immunostimulatory DNA sequences (15), the use of Salmonella vectors (32), and targeting of DNA to dendritic cells (33). In our previous studies with DNA-based immunization against HBV (18) and HCV (20), we found that the use of a recombinant poxvirus as a booster months after immunization with DNA produced a much stronger booster effect than that obtained when DNA was used as a booster.In the present study we evaluated plasmid DNA immunization with polycistronic gene constructs encoding the whole protein coding HCV genome in HLA-A2.1-transgen...

Recombinant Modified Vaccinia Virus Ankara Expressing a Soluble Form of Glycoprotein B Causes Durable Immunity and Neutralizing Antibodies against Multiple Strains of Human Cytomegalovirus

“…A surrogate challenge animal model using an HCMVgB-murine CMV chimera might be a valuable tool not only in the design of a gB680-MVA vaccine regime but also in optimizing gB680-MVA vaccination strategies, such as doses, routes, and timing of prime-boost administrations. In fact, the surrogate challenge animal model using recombinant VV has been used in hepatitis C vaccine development (6,45,47). However, the life cycle and tissue tropism of poxviruses are far different from those of CMV and other herpesviruses, so the utility of developing such an approach is minimal.…”

Progress on new vaccine strategies against chronic viral infections

“…Therefore, recent strategies for inducing both humoral and CD4 + and CD8 + cellular immunity could potentially provide more complete protective immunity against HCV infection. Some of these vaccines have induced CTLs in HLA transgenic or other mice sufficient to protect against a recombinant vaccinia virus expressing HCV antigens, used as a surrogate challenge virus in mice (91,93). To induce Th1-type immunity and improve T cell-mediated immunity, inclusion of cytokines and other biological adjuvants may be necessary for both prophylactic and therapeutic HCV vaccines in the future.…”

“…Thus, effort has been invested in defining HCV CTL epitopes and designing vaccine constructs (85)(86)(87)(88)(89)(90). Such approaches to HCV vaccine development include the use of DNA plasmids (90,91), recombinant viral vectors expressing HCV antigens (91)(92)(93), and HCV viruslike particles (94,95). To improve on the ability of the wild-type viral sequence to induce T cell immunity, the amino acid sequence of epitopes has been modified to increase affinity for the HLA molecule to make the epitopes more potent vaccines (known as epitope enhancement) (88).…”

Evaluating Replication-Defective Vesicular Stomatitis Virus as a Vaccine Vehicle

“…In this endeavor, many approaches to more recent HCV vaccine strategies have attempted to improve immunogenicity to antigens and protective efficacy of subsequent immune responses using chimpanzees and smaller animal models. Examples of such strategies include incorporation of HCV proteins into recombinant viruses, naked DNA vaccines with or without boosting with recombinant proteins to improve cytotoxic T-lymphocyte (CTL) responses, virosome or liposome carriers for immunogenic peptides to enhance their delivery to cells, and use of HCV-like particles to mimic virions (11,33,51,55,62). Use of the more conserved HCV core proteins among HCV genotypes as an adjuvant for envelope glycoprotein vaccine has similarly proven useful in rhesus macaque studies (60).…”