Development, expression, and murine testing of a multistage Plasmodium falciparum malaria vaccine candidate

A synthetic gene encoding twelve B cell epitopes, six T-cell proliferative epitopes, and three cytotoxic T lymphocyte (CTL) epitopes from nine stage-s pecific antigens, representing the sporozoite, liver stage, asexual blood-s tage, and sexual-stage antigens of Plasmodium falciparum, was constructed b y assembling overlapping oligonucleotides followed by PCR extension and ann ealing. A three-step PCR protocol using twelve long oligonucleotides was em ployed to generate a 1053 base-pair synthetic gene, the identity of which w as confirmed by sequencing. This synthetic gene, named CDC/NII MAL VAC-1, w as cloned, and the recombinant protein was expressed in the Baculovirus Exp ression Vector System (BEVS). The selection of malarial epitopes for inclus ion in this vaccine construct was based on immunoepidemiological studies in malaria endemic area, in vitro, and in vivo protection studies in model sy stems. The 41 kDa BEVS-expressed recombinant protein reacted with mouse ant ibodies specific for individual B cell epitopes in the vaccine construct an d with sera from clinically immune Kenyan adults. An immunization study in three strains of mice that differ at the H-2 locus demonstrated that the BE VS-expressed recombinant protein is immunogenic; the candidate vaccine anti gen induced high titer antibodies, and lymphocyte proliferative and IFN-gam ma responses. These results demonstrate that individual B and T cell epitop es can be assembled to create synthetic genes that encode proteins capable of eliciting specific antibody and T cell responses. Published by Elsevier Science Ltd.