The broad long-term objective of this project is to evaluate the potential of synthetic linear peptide chimeras (LPCs) containing Plasmodium universal T cell epitopes and immunodominant B cell epitopes as effective malaria vaccine components. We propose to study a series of promiscuous T cell epitopes that we have identified in two Plasmodium vivax merozoite proteins. These putative universal T cell epitopes were identified by using in vitro peptide-binding assays to soluble DRB1* molecules. Sets of synthetic peptides, representing the entire amino acid sequences of both proteins, were individually tested. Helper activity induced by immunization with promiscuous T cell epitopes was confirmed by designing LPCs containing individual T cell epitopes from one of the two characterized proteins and the NANP repeat sequence from the P. falciparum circumsporozoite (CS) protein, as a standard B cell epitope for these evaluations. Very high antipeptide and antiparasite antibodies and cellular anti-peptide Th1 and Th2 cytokine profiles were elicited after immunization with the entire group of constructs tested. Further, genetic restriction to the NANP sequence was overcome in the typically non-responder BALB/c mouse. Importantly, the antibodies elicited by the LPC immunizations were also able to inhibit the invasion of viable sporozoites to human hepatocytes. We now aim to investigate further the in vivo relevance of this approach for the design of subunit malaria vaccines. New peptide constructs will be designed containing other promiscuous T cell epitopes, selected from those we have identified, and tested for immunogenicity in mice with diverse genetic backgrounds. We hypothesize that a number of these promiscuous T cell epitopes are in fact universal T cell epitopes for their ability to bind several MHC class II molecules. The simplicity of the LPC method for designing multimeric and multispecies constructs facilitates the screening of several variables including: the physical association of more that one T cell epitope, the intrinsic activity of the particular T cell epitopes on the polarization of the immune response towards a Th1 or Th2 phenotype, the inclusion of different B cell epitopes, and the definition of minimal T cell epitopes required for T helper cell activity. These studies will form the foundation for designing and testing the effectiveness of selected Plasmodium-based LPCs in rhesus monkeys, an animal model close to humans. Potential LPC immunogens will be evaluated in peptide-binding assays for their ability to bind Mamu-DR alleles, and then for immunogenicity and recall responses in monkeys experimentally infected with P. cynomolgi and P. coatneyi: simian malaria models for P. vivax and P. falciparum, respectively. These approaches will provide strong data to evaluate the LPC approach as a ways forward for clinical trials, whether used singly or in combination with other malaria vaccine platforms.
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