Immunization of mice withPasnodiumyoeli sporozoite surface protein 2 (PySSP2) and cfrcumorozoite protein protects completely against P. yoelii. The amino acid sequence of PySSP2 suggested that the thrombospondinrelated anonymous protein (TRAP) [Robson, K.

Cytotoxic T lymphocytes (CTL) against the circumsporozoite (CS) protein of malaria sporozoites protect against malaria in rodents. Although there is interest in developing human vaccines that induce CTL against the Plasmodiumfalk4iarm CS protein, humans have never been shown to produce CTL against any Plasmodium species protein or other parasite protein. We report that when peripheral blood mononuclear cells (PBMC) from three of four volunteers immunized with irradiated P. falkiparum sporozoites were stimulated in vitro with a recombinant vaccinia virus expressing the P.fakiparum CS protein or a peptide including only amino acids 368-390 of the P. falciparum CS protein ], the PBMC lysed autologous Epstein-Barr virus-transformed B cells transfected with the P. falciparum CS protein gene or incubated with CS-(368-390) tricosapeptide. Activity was antigen specific, genetically restricted, and dependent on CD8 T cells. In one volunteer, seven peptides reflecting amino acids 311-400 were tested, and, as in B1O.BR mice, CTL activity was only associated with the CS-(368-390) peptide. Development of an assay for studying human CTL against the CS and other malaria proteins and a method for constructing target cells by direct gene transfection provide a foundation for studying the role of CTL in protection against malaria.After inoculation by mosquitoes, mature sporozoites pass through the blood to the liver where each uninucleate sporozoite can develop within a hepatocyte during -1 week to a schizont containing 1-5 x 104 uninucleate merozoites. When this schizont ruptures, each merozoite can invade an erythrocyte initiating the stage responsible for human disease.

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Immunization of mice with irradiated malaria sporozoites induces protection that is dependent on CD8+ T cells, and adoptive transfer of CD8+ cytotoxic T lymphocyte (CTL) clones against rodent malaria circumsporozoite (CS) protein and sporozoite surface protein 2 completely protects against sporozoite challenge.Thus, there are now efforts to develop vaccines that induce CTL against the CS protein and sporozoite surface protein 2. Until recently, it was thought that induction of CTL required production of target proteins within cells, breakdown of the proteins to peptides in the cytoplasm, and transport of the peptides to the cell surface in combination with class I major histocompatibility complex molecules. It has now been shown that immunization with peptides in Freund's complete adjuvant and with soluble protein in liposomes can induce CTL. To determine whether we could induce CTL against the Plasmodium falciparum CS protein by immunization with soluble protein, B10.BR mice were immunized intravenously, intraperitoneally, or intramuscularly with a recombinant P. falciparum CS protein called RLF mixed with the adjuvant DETOX (monophosphoryl lipid A, cell wall skeleton ofMycobacteria phlei, and squalane). Two weeks after the last dose, spleen cells from mice immunized intravenously, but not intraperitoneally or intramusculariy, had peptidespecific, major histocompatibility complex-restricted, CD8+ T-cell-dependent cytolytic activity against peptide 368-390 from the 7G8 P.falciparum CS protein. To determine whether the adjuvant was required for induction of the cytolytic activity, mice were immunized with RLF without adjuvant, and similar cytolytic activity was demonstrated. The finding that we could induce CTL by administration of soluble protein without adjuvant markedly broadens the possibilities for vaccinologists working to develop methods of inducing CTL in humans.