Identification of two new protective pre-erythrocytic malaria vaccine antigen candidates

<p>Abstract</p> <p>Background</p> <p>Despite years of effort, a licensed malaria vaccine is not yet available. One of the obstacles facing the development of a malaria vaccine is the extensive heterogeneity of many of the current malaria vaccine antigens. To counteract this antigenic diversity, an effective malaria vaccine may need to elicit an immune response against multiple malaria antigens, thereby limiting the negative impact of variability in any one antigen. Since most of the malaria vaccine antigens that have been evaluated in people have not elicited a protective immune response, there is a need to identify additional protective antigens. In this study, the efficacy of three pre-erythrocytic stage malaria antigens was evaluated in a <it>Plasmodium yoelii</it>/mouse protection model.</p> <p>Methods</p> <p>Mice were immunized with plasmid DNA and vaccinia virus vectors that expressed one, two or all three <it>P. yoelii </it>vaccine antigens. The immunized mice were challenged with 300 <it>P. yoelii </it>sporozoites and evaluated for subsequent infection.</p> <p>Results</p> <p>Vaccines that expressed any one of the three antigens did not protect a high percentage of mice against a <it>P. yoelii </it>challenge. However, vaccines that expressed all three antigens protected a higher percentage of mice than a vaccine that expressed PyCSP, the most efficacious malaria vaccine antigen. Dissection of the multi-antigen vaccine indicated that protection was primarily associated with two of the three <it>P. yoelii </it>antigens. The protection elicited by a vaccine expressing these two antigens exceeded the sum of the protection elicited by the single antigen vaccines, suggesting a potential synergistic interaction.</p> <p>Conclusions</p> <p>This work identifies two promising malaria vaccine antigen candidates and suggests that a multi-antigen vaccine may be more efficacious than a single antigen vaccine.</p

Sterile Protection against Plasmodium knowlesi in Rhesus Monkeys from a Malaria Vaccine: Comparison of Heterologous Prime Boost Strategies

Using newer vaccine platforms which have been effective against malaria in rodent models, we tested five immunization regimens against Plasmodium knowlesi in rhesus monkeys. All vaccines included the same four P. knowlesi antigens: the pre-erythrocytic antigens CSP, SSP2, and erythrocytic antigens AMA1, MSP1. We used four vaccine platforms for prime or boost vaccinations: plasmids (DNA), alphavirus replicons (VRP), attenuated adenovirus serotype 5 (Ad), or attenuated poxvirus (Pox). These four platforms combined to produce five different prime/boost vaccine regimens: Pox alone, VRP/Pox, VRP/Ad, Ad/Pox, and DNA/Pox. Five rhesus monkeys were immunized with each regimen, and five Control monkeys received a mock vaccination. The time to complete vaccinations was 420 days. All monkeys were challenged twice with 100 P. knowlesi sporozoites given IV. The first challenge was given 12 days after the last vaccination, and the monkeys receiving the DNA/Pox vaccine were the best protected, with 3/5 monkeys sterilely protected and 1/5 monkeys that self-cured its parasitemia. There was no protection in monkeys that received Pox malaria vaccine alone without previous priming. The second sporozoite challenge was given 4 months after the first. All 4 monkeys that were protected in the first challenge developed malaria in the second challenge. DNA, VRP and Ad5 vaccines all primed monkeys for strong immune responses after the Pox boost. We discuss the high level but short duration of protection in this experiment and the possible benefits of the long interval between prime and boost

Adenovirus-5-Vectored P. falciparum Vaccine Expressing CSP and AMA1. Part B: Safety, Immunogenicity and Protective Efficacy of the CSP Component

Background: A protective malaria vaccine will likely need to elicit both cell-mediated and antibody responses. As adenovirus vaccine vectors induce both these responses in humans, a Phase 1/2a clinical trial was conducted to evaluate the efficacy of an adenovirus serotype 5-vectored malaria vaccine against sporozoite challenge.\ud \ud Methodology/Principal Findings: NMRC-MV-Ad-PfC is an adenovirus vector encoding the Plasmodium falciparum 3D7 circumsporozoite protein (CSP). It is one component of a two-component vaccine NMRC-M3V-Ad-PfCA consisting of one adenovector encoding CSP and one encoding apical membrane antigen-1 (AMA1) that was evaluated for safety and immunogenicity in an earlier study (see companion paper, Sedegah et al). Fourteen Ad5 seropositive or negative adults received two doses of NMRC-MV-Ad-PfC sixteen weeks apart, at 1x1010 particle units per dose. The vaccine was safe and well tolerated. All volunteers developed positive ELISpot responses by 28 days after the first immunization (geometric mean 272 spot forming cells/million[sfc/m]) that declined during the following 16 weeks and increased after the second dose to levels that in most cases were less than the initial peak (geometric mean 119 sfc/m). CD8+ predominated over CD4+ responses, as in the first clinical trial. Antibody responses were poor and like ELISpot responses increased after the second immunization but did not exceed the initial peak. Pre-existing neutralizing antibodies (NAb) to Ad5 did not affect the immunogenicity of the first dose, but the fold increase in NAb induced by the first dose was significantly associated with poorer antibody responses after the second dose, while ELISpot responses remained unaffected. When challenged by the bite of P. falciparum-infected mosquitoes, two of 11 volunteers showed a delay in the time to patency compared to infectivity controls, but no volunteers were sterilely protected.\ud \ud Significance: The NMRC-MV-Ad-PfC vaccine expressing CSP was safe and well tolerated given as two doses, but did not provide sterile protection

Construction and characterization of adenovirus vectors expressing optimized blood stage antigens of Plasmodium falciparum

Malaria is the most devastating parasitic disease affecting humans. Each year there are 300-500 million new infections and 1-3 million deaths, primarily of children in sub-Saharan Africa. The feasibility of a malaria vaccine is supported by the demonstration of protective immunity following exposure to the intact Plasmodium parasite and the decrease in incidence, prevalence, and density of infection with age and exposure. In the latter, the protective immune mechanism is thought to be mediated primarily by antibodies directed against antigens expressed during the blood-stage. Immunization with subunit vaccines incorporating blood stage antigens has the potential to induce protective antibody responses. Adenovectors offer great potential for the next generation of molecular vaccines. They induce strong and protective immune responses in multiple disease systems and multiple animal models including mice and non-human primates. Moreover, adenovectors are now undergoing clinical testing for application as an HIV vaccine. Our strategy is to develop a bivalent adenovector that expresses optimized forms of two P. falciparum blood stage antigens, PfAMA1 and PfMSP142. To maximize the potential of these antigens to induce strong antibody responses, we have designed adenovectors to express these antigens either intracellularly or at the cell surface. In addition, as the malaria parasite does not glycosylate its proteins efficiently, we generated mutants of both PfAMA1 and PfMSP142 with conservative substitutions in all of the potential glycosylation sites. These variant antigens were then built into adenovectors and evaluated for cell surface expression, glycosylation and their capacity to induce antibody and T-cell responses in mice. The higher apparent molecular weight of the secreted/glycosylated (SG) versions of both antigens observed on immunoblots suggested that these antigens were post-translationally modified. The glycosylation status of both antigens was confirmed by treatment with endoglycosidases Endo H and PNGase F. Immunofluorescence assays (IFA) indicated that the PfAMA1 (SG) antigen was located at the cell surface and the secreted/non-glycosylated (SNG) and non-secreted (NS) versions were expressed preferentially inside the cell. Protease digestion of intact infected cells confirmed these findings, suggesting that the PfAMA1 (SG) antigen is present at the cell surface in a conformation that is recognized by the 4G2 antibody and sensitive to trypsin digestion. All variants of the PfMSP142 protein from infected A549 cells were preferentially associated with the cells and not secreted into the media when examined by immunoblotting. One variant of PfMSP142, PfMSP142 (DSA), which contained the decay-accelerating factor (DAF) signal sequence and GPI anchor domain was shown to preferentially associate with the cell surface by FACS analysis using the 5.2 mAb. These vectors are currently being evaluated for their capacity to induce antibody and T cell responses to the PfAMA1 and PfMSP142 antigens. Results of this analysis will be presented

Correction: Vaccine Strain-Specificity of Protective HLA-Restricted Class 1 P. falciparum Epitopes.

[This corrects the article DOI: 10.1371/journal.pone.0163026.]