Despite the great global public health importance, malaria vaccine development has been slow with only a handful of candidates under development and the most advanced displaying modest efficacy. The current use of bioinformatics for genome/proteome screening, animal models, and more efficient manufacturing facilities for recombinant/synthetic antigens could significantly accelerate the development of novel vaccines. We propose to use these advances to identify and develop vaccine candidates targeting both, Plasmodium falciparum and P. vivax parasites, responsible for >99% of the malaria cases worldwide. Alpha-helical coiled-coil motifs, protein structures widely distributed in the Plasmodium proteomes known to spontaneously fold in aqueous solution, are able to induce antibodies that inhibit parasite growth in vitro. We previously in silico identified 220 proteins in both parasite proteomes, 170 of which were synthesized and tested. High antigenicity with human sera from endemic areas, and immunogenicity in mice led to selection of 145 fragments. Several of them displayed significant P. falciparum/P. vivax cross reactivity, and importantly in vitro parasite inhibition activity, which correlated with malaria clinical immunity. Furthermore, mice immunization with a P. falciparum poly-epitope construct (Pf-181) maintained individual epitopes specificity and parasite inhibitory activity. Our general hypothesis is that ?Plasmodium poly-epitope constructs containing cross-reacting ?-helical coiled-coil motifs induce protection in monkey against homologous and heterologous parasite challenges. The overall goal is to determine the vaccine potential of P. falciparum and P. vivax erythrocytic synthetic poly-epitope constructs containing ?-helical coiled-coil motifs. This goal will be approached through the following specific aims: 1) Design, synthesis and characterization of P. falciparum and P. vivax poly-epitope constructs containing ?-helical coiled-coil motifs; 2) Evaluation of the immunogenicity and protective efficacy of selected P. falciparum and P. vivax poly-epitope constructs against homologous and heterologous parasite challenges in Aotus monkeys; 3) In vitro characterization of ?-helical coiled-coil fragments/constructs and immune responses elicited by animal immunization. Methods proposed are: a) F-moc peptide synthesis of poly-epitope constructs, followed by HPLC purification, mass spectrometry (MS) and circular dichroism (CD) analyses; b) Monkey immunization with selected poly-epitope constructs and evaluation of protective efficacy to homologous and heterologous challenges using P. falciparum (Santa Lucia) and P. vivax (Sal-I) parasite strains; c) ELISA and IFAT test of sera from immunized animals; d) antibody functional in vitro assays (GIA, ADRB,OPA).The innovation of this study is the highly efficient and cost-effective approach, using a comprehensive rational and rigorous epitope down- selection of hundreds of protein fragments from in silico epitope identification in Plasmodium proteomes for poly- epitope design and evaluation of their protective efficacy studies in monkeys. It will accelerate the development of malaria vaccines for human use.
Malaria represents a global public health problem with 2.5 billion people at risk of disease and death. However, during the last two decades, a global decrease of ~60% in malaria incidence has motivated attempts for malaria elimination and malaria vaccines are considered an ideal complementary tool. The ultimate goal of this proposal is to accelerate the development of a malaria vaccine that targets both P. falciparum and P. vivax, the two most abundant malaria parasites. This proposal is based the use of bioinformatics, peptides synthesis, as well as studies in rodents and non-human primates, to identify orthologous cross-reactive protein fragments with ?- helical coiled-coil motifs in both malaria parasite species that could be used to assemble multispecies poly- epitope vaccine constructs targeting both parasites species. This highly efficient and cost-effective approach will greatly contribute to accelerate the development of a universal malaria vaccine that will improve public health in tropical countries and decrease the risk of travelers from non-endemic areas.
