Stability and potency of the Plasmodium falciparum MSP1-19/AMA-1(III) chimeric vaccine candidate with Montanide ISA720 adjuvant.

The Plasmodium falciparum AMA-1(III) and MSP1-19 proteins have been expressed as a chimera (PfCP-2.9), adjuvanted with Montanide ISA720 and developed as a vaccine candidate tested in human. The PfCP-2.9 protein contains 18 cysteine residues that form nine intramolecular disulfide bonds. The protective immune responses induced by the chimeric protein were dependent on its disulfide bond-based conformation. In this study, we developed a sandwich ELISA to assess the nature of the protein in the emulsion over time (6, 12 and 18 months). Our results showed that the OD(450) values corresponding to vaccine storages were within the 95% confidence interval, indicating that the conformation of the protein in the emulsion stored for up to 18 months at 4 degrees C was unchanged. Furthermore, no protein degradation was detected by Coomassie blue, silver staining, and Western blot analysis for samples stored at 4 degrees C for up to 2 years. Although some protein aggregation was observed in the emulsion preparations, these aggregates were only a small percentage of the total protein in the sample (7.6%). Moreover, the protein multimers maintained their conformational epitope. The potency assay of the formulation showed no significant differences in ED(50) values (50% effective dose for achieving seroconversion) between fresh vaccine formulations (ED(50)=0.057+/-0.024 microg) and formulations stored for up to 6 (ED(50)=0.046 microg) or 12 months (ED(50)=0.040 microg). Importantly, the immune sera of rabbits immunized with formulations stored for 0, 3, 6, 9 and 12 months effectively inhibited parasite growth in vitro at similar levels. These data indicated that the vaccine emulsion was stable over long periods of storage and maintained both its physical and biological properties.