Immunogenic properties of a recombinant fusion protein containing the C-terminal 19 kDa of Plasmodium falciparum merozoite surface protein-1 and the innate immunity agonist FliC flagellin of Salmonella typhimurium.

In a recent study, we demonstrated the immunogenic properties of a new malaria vaccine polypeptide based on a 19 kDa C-terminal fragment of the merozoite surface protein-1 (MSP1(19)) from Plasmodium vivax and an innate immunity agonist, the Salmonella enterica serovar Typhimurium flagellin (FliC). Herein, we tested whether the same strategy, based on the MSP1(19) component of the deadly malaria parasite Plasmodium falciparum, could also generate a fusion polypeptide with enhanced immunogenicity. The His(6)FliC-MSP1(19) fusion protein was expressed from a recombinant Escherichia coli and showed preserved in vitro TLR5-binding activity. In contrast to animals injected with His(6)MSP1(19), mice subcutaneously immunised with the recombinant His(6)FliC-MSP1(19) developed strong MSP1(19)-specific systemic antibody responses with a prevailing IgG1 subclass. Incorporation of other adjuvants, such as CpG ODN 1826, complete and incomplete Freund's adjuvants or Quil-A, improved the IgG responses after the second, but not the third, immunising dose. It also resulted in a more balanced IgG subclass response, as evaluated by the IgG1/IgG2c ratio, and higher cell-mediated immune response, as determined by the detection of antigen-specific interferon-gamma secretion by immune spleen cells. MSP1(19)-specific antibodies recognised not only the recombinant protein, but also the native protein expressed on the surface of P. falciparum parasites. Finally, sera from rabbits immunised with the fusion protein alone inhibited the in vitro growth of three different P. falciparum strains. In summary, these results extend our previous observations and further demonstrate that fusion of the innate immunity agonist FliC to Plasmodium antigens is a promising alternative to improve their immunogenicity.

Bargieri DY ，Leite JA ，Lopes SC ，Sbrogio-Almeida ME ，Braga CJ ，Ferreira LC ，Soares IS ，Costa FT ，Rodrigues MM ... -  《-》

New malaria vaccine candidates based on the Plasmodium vivax Merozoite Surface Protein-1 and the TLR-5 agonist Salmonella Typhimurium FliC flagellin.

Bargieri DY ，Rosa DS ，Braga CJ ，Carvalho BO ，Costa FT ，Espíndola NM ，Vaz AJ ，Soares IS ，Ferreira LC ，Rodrigues MM ... -  《VACCINE》

Long-lasting protective immune response to the 19-kilodalton carboxy-terminal fragment of Plasmodium yoelii merozoite surface protein 1 in mice.

Jeamwattanalert P ，Mahakunkijcharoen Y ，Kittigul L ，Mahannop P ，Pichyangkul S ，Hirunpetcharat C ... -  《-》

TLR5-dependent immunogenicity of a recombinant fusion protein containing an immunodominant epitope of malarial circumsporozoite protein and the FliC flagellin of Salmonella Typhimurium.

Camacho AG ，Teixeira LH ，Bargieri DY ，Boscardin SB ，Soares IS ，Nussenzweig RS ，Nussenzweig V ，Rodrigues MM ... -  《-》

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.

Xue X ，Ding F ，Zhang Q ，Pan X ，Qu L ，Pan W ... -  《-》