These replicon DNA or particle pentavalent vaccines could simultaneously and effectively induce antibody responses and protect effects against the 5 agents. the binding of THc to the ganglioside GT1b as the sera of individual replicon DNA or particle-immunized mice. These results indicated these pentavalent replicon vaccines could protect against the 4 BoNT serotypes and effectively neutralize and protect the TeNT. Therefore, our studies demonstrate the utility of combining replicon DNA or particle vaccines into multi-agent formulations as potent pentavalent vaccines for eliciting protective responses against BoNTs and TeNT. and were shown to elicit protective immune responses in mice and other animals.9-15 DNA Rabbit polyclonal to AFG3L1 or viral replicon vaccines encoding the same Hc antigens were described as the next generation of candidate vaccines.16-21 Recently, we developed individual and multivalent candidate replicon vaccines against BoNTs using DNA-based Semliki Forest virus (SFV) replicon vectors.22-24 In this current study, we further developed several pentavalent replicon vaccines for BoNTs (serotypes A, B, E, and F) and TeNT using the DNA-based SFV replicon vectors, and evaluated their immunogenicity and protective capability against challenge with the BoNTs and TeNT mixture in mice. Results Individual replicon vaccine for TeNT The immunogenicity and protective ability of replicon DNA or VRP vaccine against TeNT were investigated and compared with the pVAX1STHc encoding the THc antigen. As shown in Table 1, these vaccines induced strong ELISA and neutralizing antibody responses in BALB/c mice following 3 immunizations. Mice immunized 3 times with pSCARSTHc, VRP-THc, or pVAX1STHc were completely protected Prostaglandin E1 (PGE1) against 1000 or 10? 000 mouse LD50 of biologically active TeNT, and 2 injections with low antibody responses provided partial protection against 1000 LD50 of TeNT, while the negative control mice succumbed to intoxication and died within 12 h as the native mice. Table 1. Sera antibody titers, neutralization titers, and survival of mice following vaccination with pSCARSTHC, pVAX1STHC, or RVP-THC 0.05) indicate significant anti-THc antibody titer difference between pSCARSTHC- and pVAX1STHC-vaccination groups with 3 injections; bPooled sera from each group of mice were diluted initially 1:32 (500 L) and then 2-fold for serum neutralization titers. Due to the limited amount of serum available, serum from each group of mice was pooled, and so only the average neutralization titer (IU/mL) of Prostaglandin E1 (PGE1) the group against TeNT could be assayed; cBALB/c mice alive (8 mice/group) after s.c. challenge with a dose 1000 LD50 of TeNT 4 wk after the last injection; dBALB/c mice alive (8 mice/group) after s.c. challenge with a dose 10?000 LD50 of TeNT 4 wk after the last injection; eNumber of vaccinations. To further define the protective potencies of these replicon vaccines, we rechallenged the mice of 3 vaccination 1 Prostaglandin E1 (PGE1) wk later with a higher TeNT doses (100?000 LD50) and observed no mice deaths in the pSCARSTHc or VRP-THc-immunized group. However, the pVAX1STHc vaccine provided no protection. Notably, the mean ELISA and neutralizing antibody titers in the 3 doses of pSCARSTHc or VRP-THc-immunized group Prostaglandin E1 (PGE1) were higher than that of pVAX1STHc-immunized group ( 0.05). Protective effects correlated directly with the ELISA and neutralizing antibody titers to TeNT. Therefore, these results indicated that the replicon DNA or VRP encoding the THc antigen is an effective vaccine against TeNT. Pentavalent plasmid replicon DNA vaccines for BoNTs and TeNT Then, the individual replicon DNA TeNT vaccines were combined with tetravalent vaccines against BoNTs24 to prepare pentavalent replicon DNA vaccines against BoNTs and TeNT. We evaluated 2 types of pentavalent replicon DNA vaccine candidates: pentavalent DNA-I (pSCARSAHc + pSCARSBHc + pSCARSEHc + pSCARSFHc + pSCARSTHc) and pentavalent DNA-II (pSCARSA/BHc + pSCARSE/FHc + pSCARSTHc). As shown in Table 2, the 2 2 types of pentavalent replicon DNA vaccines induced high ELISA IgG antibody titers to each Hc and efficacious neutralizing antibody Prostaglandin E1 (PGE1) levels against TeNT in mice. And these vaccinations provided nearly complete protective effects against challenge with 100 or 1000 LD50 of BoNTs (serotypes A, B, E, and F) and TeNT mixtures (Table 2). Table?2. Sera antibody titers, neutralization titers, and survival of mice following 3 vaccinations i.m. with pentavalent replicon DNA vaccines against BoNTs and TeNT and/or virus are being assessed in our laboratory. Thus, we have described an efficient strategy to design and develop multivalent vaccines against multi-agent pathogens using DNA-based SFV replicon vectors. In.