Combination of DNA prime--adenovirus boost immunization with entecavir elicits sustained control of chronic hepatitis B in the woodchuck model.

A potent therapeutic T-cell vaccine may be an alternative treatment of chronic hepatitis B virus (HBV) infection. Previously, we developed a DNA prime-adenovirus (AdV) boost vaccination protocol that could elicit strong and specific CD8+ T-cell responses to woodchuck hepatitis virus (WHV) core antigen (WHcAg) in mice. In the present study, we first examined whether this new prime-boost immunization could induce WHcAg-specific T-cell responses and effectively control WHV replication in the WHV-transgenic mouse model. Secondly, we evaluated the therapeutic effect of this new vaccination strategy in chronically WHV-infected woodchucks in combination with a potent antiviral treatment. Immunization of WHV-transgenic mice by DNA prime-AdV boost regimen elicited potent and functional WHcAg-specific CD8+ T-cell response that consequently resulted in the reduction of the WHV load below the detection limit in more than 70% of animals. The combination therapy of entecavir (ETV) treatment and DNA prime-AdV boost immunization in chronic WHV carriers resulted in WHsAg- and WHcAg-specific CD4+ and CD8+ T-cell responses, which were not detectable in ETV-only treated controls. Woodchucks receiving the combination therapy showed a prolonged suppression of WHV replication and lower WHsAg levels compared to controls. Moreover, two of four immunized carriers remained WHV negative after the end of ETV treatment and developed anti-WHs antibodies. These results demonstrate that the combined antiviral and vaccination approach efficiently elicited sustained immunological control of chronic hepadnaviral infection in woodchucks and may be a new promising therapeutic strategy in patients.

Construction and comparative study of monovalent and multivalent DNA vaccines against Streptococcus iniae.

Streptococcus iniae is an important fish pathogen with a broad host range that includes both marine and freshwater fish species. With an aim to develop effective vaccines against S. iniae, we in this study constructed three monovalent DNA vaccines, i.e., pSagF, pSagG, and pSagI, based on sagF, G, and I, which are components of the streptolysin S cluster. The immunoprotective potentials of these vaccines were examined in a model of Japanese flounder (Paralichthys olivaceus). The results showed that following intramuscular administration, the vaccine plasmids were transported to spleen, kidney, and liver, where the vaccine-encoding transgenes were expressed. Immunocolloidal gold electron microscopy detected production of the vaccine protein in fish vaccinated with each of the vaccine plasmids. Following lethal-dose S. iniae challenge, pSagF-, pSagG-, and pSagI-vaccinated fish exhibited relative percent of survival (RPS) rates of 78%, 65%, and 76% respectively. To examine whether multivalent vaccines composed of different combinations of monovalent vaccines would produce better protections, flounder were vaccinated with FG (pSagF plus pSagG), FI (pSagF plus pSagG), or FGI (pSagF plus pSagG and pSagI). Subsequent challenging study showed that the RPS rates of the fish vaccinated with the divalent and trivalent vaccines were 4%-17% and 13%-26% respectively higher than those of the fish vaccinated with the component monovalent vaccines. Furthermore, FGI exhibited a strong cross protection against both serotype I and serotype II S. iniae, apparently due to, as revealed by sequence analysis, the existence of highly conserved SagF, SagG, and SagI homologs in these serotypes. Immunological analysis showed that all vaccines induced (i) specific serum antibody production, (ii) enhanced complement-mediated bactericidal activity, and (iii) significant induction of a wide range of immune genes. However, the levels of gene expression and serum bactericidal activity induced by FGI were in general more potent than those induced by monovalent vaccines. Taken together, these results indicate that the DNA vaccines based on sagF, G, and I, especially when they are formulated as multivalent vaccines, are highly efficacious against S. iniae infection.

Therapeutic DNA vaccine induces broad T cell responses in the gut and sustained protection from viral rebound and AIDS in SIV-infected rhesus macaques.

Immunotherapies that induce durable immune control of chronic HIV infection may eliminate the need for life-long dependence on drugs. We investigated a DNA vaccine formulated with a novel genetic adjuvant that stimulates immune responses in the blood and gut for the ability to improve therapy in rhesus macaques chronically infected with SIV. Using the SIV-macaque model for AIDS, we show that epidermal co-delivery of plasmids expressing SIV Gag, RT, Nef and Env, and the mucosal adjuvant, heat-labile E. coli enterotoxin (LT), during antiretroviral therapy (ART) induced a substantial 2-4-log fold reduction in mean virus burden in both the gut and blood when compared to unvaccinated controls and provided durable protection from viral rebound and disease progression after the drug was discontinued. This effect was associated with significant increases in IFN-γ T cell responses in both the blood and gut and SIV-specific CD8+ T cells with dual TNF-α and cytolytic effector functions in the blood. Importantly, a broader specificity in the T cell response seen in the gut, but not the blood, significantly correlated with a reduction in virus production in mucosal tissues and a lower virus burden in plasma. We conclude that immunizing with vaccines that induce immune responses in mucosal gut tissue could reduce residual viral reservoirs during drug therapy and improve long-term treatment of HIV infection in humans.

The treatment and prevention of mouse melanoma with an oral DNA vaccine carried by attenuated Salmonella typhimurium.

Therapeutic vaccines of cancer are attractive for their capacity of breaking the immune tolerance and invoking long-term immune response targeting cancer cells without autoimmunity. An efficient antigen delivery system is the key issue of developing an effective cancer vaccine. Attenuated Salmonella typhimurium as the carrier of cancer vaccine are able to transfer DNA from the prokaryote to the eukaryote and preferentially replicate within the tumor tissue. Heat shock protein 70 delivers the tumor-associated antigens to antigen presenting cells through its polypeptide-binding domain and breaks immune tolerance of the cancer cells. Here we described a novel low-copy-number DNA vaccine based on the Hsp70-TAA complex and carried by the attenuated S. typhimurium strain SL3261. Oral administration of this vaccine elicited specific CTL-mediated lysis of the melanoma tumor cells and marked activation of the T-cells. The therapeutic vaccine effectively protected 57.1% C57BL/6J mice from lethal challenge with B16F10 melanoma tumor cells in prophylactic settings and eraicated 62.5% tumor growth in therapeutic settings. This approach may provide a new strategy for the prevention and treatment of cancer.

Bacteria as DNA vaccine carriers for genetic immunization.

Genetic immunization with plasmid DNA vaccines has proven to be a promising tool in conferring protective immunity in various experimental animal models of infectious diseases or tumors. Recent research focuses on the use of bacteria, in particular enteroinvasive species, as effective carriers for DNA vaccines. Attenuated strains of Shigella flexneri, Salmonella spp., Yersinia enterocolitica or Listeria monocytogenes have shown to be attractive candidates to target DNA vaccines to immunological inductive sites at mucosal surfaces. This review summarizes recent progress in bacteria-mediated delivery of plasmid DNA vaccines in the field of infectious diseases and cancer.

Protective effect of vaccination with DNA of the H. Pylori genomic library in experimentally infected mice.

Immunologically mediated protection against H. pylori infection is an attractive alternative to antibiotic treatment. We compared the efficacy of conventional protein vaccination with that of genetic vaccination against experimental infection with H. pylori in mice. For oral immunization, we used the recombinant peptide of an antigenic fragment of UreB (rUreB) or H. pylori-whole cell lysate antigens, and for genetic immunization, we used recombinant pcDNA and pSec plasmids inserted with the fragment of ureB or DNA of the H. pylori genomic library. Mice were challenged with the mouse stomach-adapted H. pylori Sidney Strain. The detection of gastric bacterial colonization was performed by real-time PCR of a 26-kDa Helicobacter-specific gene, and the presence of serum H. pylori-specific antibodies was determined using direct ELISA assay. The most effective treatment appeared to be oral vaccination with rUreB and either intramuscular or intradermal vaccination with DNA of the H. pylori genomic library. Intradermal genetic vaccination with genomic library DNA significantly increased the IgG antibody response. Our study revealed acceptable efficacies of genetic vaccination with DNA of the H. pylori genomic library.