Atraumatic oral spray immunization with replication-deficient viral vector vaccines.

The development of needle-free vaccines is one of the recently defined "grand challenges in global health" (H. Varmus, R. Klausner, R. Klausner, R. Zerhouni, T. Acharya, A. S. Daar, and P. A. Singer, Science 302:398-399, 2003). To explore whether a natural pathway to the inductive site of the mucosa-associated lymphatic tissue could be exploited for atraumatic immunization purposes, replication-deficient viral vector vaccines were sprayed directly onto the tonsils of rhesus macaques. Tonsillar immunization with viral vector vaccines encoding simian immunodeficiency virus (SIV) antigens induced cellular and humoral immune responses. Viral RNA levels after a stringent SIV challenge were reduced, providing a level of protection similar to that observed after systemic immunization with the same vaccines. Thus, atraumatic oral spray immunization with replication-deficient vectors can overcome the epithelial barrier, deliver the vaccine antigen to the mucosa-associated lymphatic tissue, and avoid induction of tolerance, providing a novel approach to circumvent acceptability problems of syringe and needle vaccines for children and in developing countries.

Engineering N-glycosylation mutations in IL-12 enhances sustained cytotoxic T lymphocyte responses for DNA immunization.

Interleukin-12 (IL-12), consisting of p40 and p35 subunits, produces both p70 heterodimer and free p40. p70 is essential for the induction of T-helper 1 (Th1) and cytotoxic T-cell (CTL) immunity, whereas p40 inhibits p70-mediated function. Here, we found that mutations introduced into N-glycosylation sites (N220 of murine p40 and N222 of human p40) reduced secretion of p40 but not p70. Co-immunization of N220 mutant mIL-12 gene with hepatitis C virus (HCV) E2 DNA significantly enhanced long-term E2-specific CD8+ T-cell response and protection against tumor challenge compared with that of wild type. Our results indicate that the ratio of p70 to p40 is important for generating sustained long-term cell-mediated immunity. Thus, the mutant IL-12 could be utilized for the development of DNA vaccines as an adjuvant for the generation of long-term memory T-cell responses.

Prolonged survival of vaccinated macaques after oral SIVmac239 challenge regardless of viremia control in the chronic phase.

To evaluate the efficacy of a multigenic vaccine and its protective immunity in the SIVmac239 challenge model, 12 rhesus macaques were divided into two groups. The vaccine group was intramuscularly immunized with multigenic DNA and recombinant adenovirus vaccine, while the control group received buffers. At 16 weeks after the last immunization, all macaques were challenged orally with pathogenic SIVmac239. The mean plasma SIV RNA loads of the vaccine group were significantly lower than those of the placebo control group up to 16 weeks post-challenge. The vaccine-induced Gag-specific IFN-gamma ELISPOT T cell responses inversely correlated with the viral loads before the chronic phase. Two out of six vaccinated macaques with strong and sustained Gag-specific T cell responses showed viremia control and maintained CD4+ T cell percentage. However, the other four vaccinated macaques showed high viral loads and reduced level of CD4+ T cell percentages during the chronic phase, comparable to those in control macaques. Five out of six vaccinated macaques survived for more than 72 weeks, while five out of six controls died of an AIDS-related disease. Therefore, the vaccination conferred not only reduction of viral loads in a portion of vaccinated macaques (2/6), but also prolonged survival of all vaccinated macaques regardless of viremia control. Our results further suggest that new experimental approaches may be needed to assess protective effects from AIDS-associated disease in the immunized macaques after oral SIV challenge.

Reduction of viral loads by multigenic DNA priming and adenovirus boosting in the SIVmac-macaque model.

In this study, we investigated the ability of a multigenic SIV DNA prime/replication-defective adenovirus serotype 5 (rAd/SIV) boost regimen to induce SIV-specific immune responses and protection against intrarectal challenge with SIVmac251 in rhesus macaques. Four rhesus macaques were immunized intramuscularly three times at 8-week intervals with SIV DNA vaccine and boosted once with rAd/SIV vaccine Four control macaques received the same amount of mock plasmid DNA and mock adenovirus vector. While the SIV DNA vaccine included plasmids expressing a mutated human IL-12 gene (IL-12N222L) as well as SIVmac239 structural and regulatory genes, the rAd/SIV vaccine contained rAd vectors expressing SIVmac239 genes only. Immunization with SIV DNA vaccine alone induced SIV-specific IFN-gamma ELISPOT responses in only two of four vaccinated macaques, whereas all animals developed SIV-specific T-cell responses and Env- and Tat-specific antibody responses following the rAd/SIV vaccine boost. Upon intrarectal challenge with pathogenic SIVmac251, strong anamnestic Env-specific binding and neutralizing antibody responses were detected in the vaccinated macaques. Overall, the immunized macaques had lower peak and set-point viral loads than control macaques, suggesting that the induced immune responses play a role in the control of viremia. In addition, the loss of CD4+ T cells was delayed in the vaccinated macaques after challenge. These results indicate that the multigenic DNA prime-adenovirus boost immunization may be a promising approach in developing an effective AIDS vaccine.