Multiple antigen constructs (MACs): induction of sterile immunity against sporozoite stage of rodent malaria parasites, Plasmodium berghei and Plasmodium yoelii.

We prepared multiple antigen constructs (MACs) using circumsporozoite (CS) protein-based B-epitopes from Plasmodium berghei, (PPPPNPND)2 and Plasmodium yoelii, (QGPGAP)3QG, along with a P. berghei T-helper epitope KQIRDSITEEWS. Mice were immunized with individual MACs in oil-in-water or water-in-oil vehicles containing block copolymer (P1005) and detoxified RaLPS (RaLPS) as well as other adjuvants. Sporozoite challenge results demonstrated that MACs in adjuvant could induce antibodies capable of active and passive protection. Water-in-oil vaccines induced the highest level of protection in mice immunized with either P. berghei or P. yoelii MACs. In a study aimed at co-eliciting immunity against P. berghei and P. yoelii, three immunizations with MACs induced protective antibodies against P. berghei but not P. yoelii parasite challenge. Therefore, it can be concluded that individually MACs are capable of inducing strong and protective immune responses to either species of rodent malaria, and that protection can be passively transferred. When MAC formulations were used together as a combined vaccine, P. berghei MACs induced a strong protective antibody response while P. yoelii MACs induced a weaker nonprotective response.

Rapid onset of malaria-induced mortality by immunizations with lipo-peptides: an experimental model to study deleterious immune responses and immunopathology in malaria.

We have recently shown that circumsporozoite (CS) protein-based cytotoxic T-cell epitope of Plasmodium berghei coupled to monoplamitic and tripalmitic acid was able to induce cytotoxic T-cell responses. In the present study, we investigated whether lipopeptide derivatized CS protein B and T helper epitopes in different combinations will be able to induce protective immune responses against sporozoite challenge. Several P. berghei CS peptides with monopalmitic fatty acid tails were prepared, suspended in an oil-in-water emulsion, and used to immunize and boost female A/J mice. The mice were challenged iv. with viable sporozoites of P. berghei (ANKA) two weeks after the last immunization. While immunization with some of these vaccine formulations induced protective immune responses, others shifted the typical bimodal pattern of P. berghei sporozoite induced death toward a rapid onset of death in a peptide specific manner. Therefore, demonstration that immunization with formulations of malarial peptides can cause enhanced malaria-related death provides an experimental model to delineate characteristics of deleterious immune responses.

Re-investigation of the circumsporozoite protein-based induction of sterile immunity against Plasmodium berghei infection.

Although the circumsporozoite protein (CSP) of the malaria parasite is the most immunologically characterized protein, the goal of using this protein in an effective vaccine has not yet been realized. Monoclonal antibody against the repetitive immunodominant B-epitope of the CSP can protect mice from malaria, but vaccines that induce antibody against this epitope do not consistently induce protection. Toward developing a rationale for a CSP-based effective vaccine, we have re-investigated the ability of anti-CSP repeat antibodies, as induced by different CSP vaccine formulations with several adjuvants, to confer sterile immunity against sporozoite challenge. Using Plasmodium berghei rodent malaria model and several CSP subunit vaccine constructs, we found that a formulation consisting of the P. berghei CSP repetitive epitope, (DPPPPNPN)2 (CS), conjugated to BSA by carbodiimide, formulated in a block copolymer and detoxified lipopolysaccharide (RaLPS) adjuvant, was particularly promising. Mice were immunized and boosted with vaccines that contain varying malarial peptide-carrier ratios of 6:1 (CS6-BSA), 55:1 (CS55-BSA) and 170:1 (CS170-BSA). Following immunization, the animals were challenged with live sporozoites. Two types of effects were observed in vaccinated mice. First, sterile immunity was induced in 100%, 50% and 29% of mice that were immunized with the CS170-BSA, CS55-BSA, and CS6-BSA vaccine conjugates, respectively. The second effect of immunization was observed with the CS170-BSA conjugate vaccine primed mice; a boost in IFA titers followed sporozoite challenge. In addition, we observed that IgG1 isotype titer against the surface of the sporozoite, as measured by IFA, and antibody avidity parallel sterile immunity. These findings reiterate the potential of the CSP as a malaria vaccine candidate antigen, and suggest that the induction of sterile immune responses depends on inducing antibody of the appropriate isotype, avidity and specificity.

Inhibition of malaria parasite development in mosquitoes by anti-mosquito-midgut antibodies.

The mosquito midgut plays a central role in the development and subsequent transmission of malaria parasites. Using a rodent malaria parasite, Plasmodium berghei, and the mosquito vector Anopheles stephensi, we investigated the effect of anti-mosquito-midgut antibodies on the development of malaria parasites in the mosquito. In agreement with previous studies, we found that mosquitoes that ingested antimidgut antibodies along with infectious parasites had significantly fewer oocysts than mosquitoes in the control group. We also found that the antimidgut antibodies inhibit the development and/or translocation of the sporozoites. Together, these observations open an avenue for research toward the development of a vector-based malaria parasite transmission-blocking vaccine.