Malaria causes at least 250 million cases and nearly 1 million deaths per year. GSK's malaria vaccine, RTS, S/AS01 is being tested in a Phase 3 clinical trial, and is likely to be licensed for use in children in the developing world, if safe ad effective. This vaccine is based entirely on the repeat region and carboxy terminus of the Plasmodium falciparum (Pf) circumsporozoite protein (CSP). It is administered with an adjuvant AS01, which includes liposomes, monophosphoryl lipid A, and a purified plant extract, QS21, but was initially developed with an oil in water-based adjuvant. Downselection of adjuvants for clinical trials was done through a series of iterative studies in mice and non-human primates (NHPs). In its final formulation this vacccine protects 50% of volunteers against experimental challenge with Pf for 2 weeks after last dose, and 22% of volunteers for 6 months. Protection is thought to be primarily mediated by antibodies against the repeat region of PfCSP and possibly CD4+ T cell responses against the C' terminus of the PfCSP. The vaccine does not induce meaningful CD8+ T cell responses. However, many malariologists believe that long- term protection will be dependent on induction of Pf-specific CD8+ T cell immunity, as has been obseved in mice and NHPs immunized with irradiated sporozoites. RTS, S/AS01 is not being considered for non-immune travelers and military personnel, because its protective efficacy is too low. A vaccine for this population needs to provide >80% protective immunity for at least 6 months to have a substantial market. We hypothesize that by adding highly functional, protective CD8+ T cell responses to antibody responses against the PfCSP, such protective immunity can be achieved. Recombinant adenovirus (Ad) expressing proteins like the PfCSP is currently a popular method for inducing CD8+ T cell responses in humans. However, despite the induction of antigen-specific CD8+ T cell responses of very high magnitude such Ad-based vaccines have not been highly protective in humans, especially against malaria. Recently, it was shown in mice that recombinant attenuated Listeria monocytogenes (Lm) induced much higher quality (functional) CD8+ T cell responses than did recombinant Ad5. We will use a heterologous prime-boost regimen combining an adjuvanted recombinant PfCSP protein (rPfCSP) and Lm expressing PfCSP (Lm-PfCSP). The goal of this strategy is to induce PfCSP- specific protective antibodies and protective CD8+ and CD4+ T cell responses that provide >80% protection that is sustained for at least 6 months. In Phase I we will identify combinations of rPfCSP, adjuvant and Lm- PfCSP that induce high level antibodies, and CD8+ and CD4+ T cell responses in mice. In Phase II we will take the approach used by GSK, and use immunogenicity in NHPs to downselect combinations for clinical trials of a vaccine that is intended to have efficacy adequate to prevent >80% of vaccinees from developing Pf parasitemia; a vaccine suitable for the potential multi-billion dollar non-immune traveler, business, and military markets, and for eliminating Pf in geographically focused campaigns in the developing world.