The fourth group of rabbits was immunized with a cocktail of the three PfAMA1 alleles, while the fifth group was immunized with a mixture of the three Diversity covering proteins (DiCo mix) at all immunization time points

The fourth group of rabbits was immunized with a cocktail of the three PfAMA1 alleles, while the fifth group was immunized with a mixture of the three Diversity covering proteins (DiCo mix) at all immunization time points. focused on shared epitopes by exclusively boosting these common determinants through immunization of rabbits with different PfAMA1 alleles in sequence. The in vitro parasite growth inhibition assay was used to further evaluate the functional effects of the broadened antibody response that is characteristic of multi-allele vaccine strategies. Results A mixed antigen immunization protocol elicited humoral responses that were functionally similar to those elicited by a sequential immunization protocol (p > 0.05). Sequential exposure to the different PfAMA1 allelic variants induced immunological recall of responses to previous alleles and yielded functional cross-strain antibodies that would be capable of optimal growth inhibition of variant parasites at high enough concentrations. Conclusions These findings may have implications for the current understanding of the natural acquisition of clinical immunity to malaria as well as for rational vaccine design. Background Malaria caused by parasites of the Plasmodium spp. continues to be a major public health problem with half of the world’s population at risk of infection [1]. The greatest risk of disease and fatality in Plasmodium falciparum-endemic areas occurs in children under 5 years and in first-time pregnant women. Natural immunity to clinical malaria is believed to develop in an age- and exposure-dependent manner, after repeated contamination by a number of (different) parasite strains [2-4]. Even in adults who have had several parasite encounters, NITD008 acquired clinical immunity is partial and is believed to be dependent on constant or periodic exposure to low-level parasitaemia [3,5]. The natural ability to acquire immunity to malaria, although partial, is a strong indication of the feasibility of developing at least an anti-disease vaccine directed against the blood stages of Plasmodium. Antigenic variation in immunogenic parasite targets however provides an immune escape route for parasites. Polymorphism in such well-known vaccine targets as the Merozoite Surface Proteins (MSPs) and Apical Membrane Antigen 1 (AMA1) have been associated with host immune pressure on parasites [6-10]. This presents malaria vaccine researchers with a formidable challenge since immunization with one variant of these polymorphic antigens induces antibodies that show NITD008 limited cross-inhibition/recognition of parasites expressing other allelic variants of the same antigen. This has been exhibited extensively in animal models [11,12] and to some extent in human field studies [13-16]. There is growing interest in multi-allele/multi-antigen malaria vaccines and the potential of such vaccines for the induction of broad inhibitory antibody responses has been exhibited [17-19]. The broadened response most likely results from diluting out strain-specific epitopes in the antigen mixture, with the bulk of remaining epitopes being those that are common to the vaccine component alleles [20]. The hypothesis that immunization of rabbits with different PfAMA1 alleles in sequence would NITD008 result in boosting of only antibodies to epitopes that are common to all antigens was tested in this study. Antibodies to highly specific epitopes would not be boosted, and this is usually expected to further increase the proportion of induced cross-strain antibodies in comparison with antibodies induced by a multi-allele vaccine that incorporates the same allelic antigens. Such a mechanism of cross-strain antibody production would be based on the concept of original antigenic sin (clonal imprinting). Original antigenic sin results when prior exposure to one strain/antigen diverts the antibody response to shared epitopes following exposure to a second closely related strain/antigen such that the newly elicited antibodies still react strongly with the priming antigen [21-23]. A sequential immunization protocol may mimic the development of natural clinical immunity and provide some insight into its acquisition in the field where over time an individual is usually uncovered (sequentially) to a number of variant parasite strains. The generated data shows that a sequential immunization protocol may not be materially different from a mixed Rabbit Polyclonal to OR10H2 antigen protocol with respect to the proportions of elicited strain-specific and cross-strain antibodies. As expected, antibody production in the sequential immunization groups was through associative immune recall of previous antigen encounter. This data is relevant to the current understanding of the acquisition of clinical immunity against malaria in endemic areas, as well as for rational vaccine design. Methods Antigen production, rabbit immunization and antibody purification The full ectodomain of the AMA1 allelic forms from P. falciparum strains FVO, HB3, 3D7 and CAMP, as well as the in silico-designed Diversity Covering antigens (DiCo 1, DiCo 2 and DiCo 3) [24], were expressed as.