Project Summary The overall aim of this application is to advance PfCDPK5 and PfGARP as vaccine candidates for falciparum malaria. P.falciparum malaria affects almost one-half of the world's population and causes more than 500,000 deaths annually. Young children in malaria endemic areas of Africa have the highest mortality rate because of their immature immune systems. Global efforts to control the disease have had limited success, and no vaccine has yet been approved for clinical use. Therefore, there is an urgent, unmet need to discover new vaccine candidates. A vaccine against childhood malaria is a priority because children below the age of 5 years are highly vulnerable to the disease. In recent studies, our laboratory discovered Schizont Egress Antigen-1 (PfSEA-1), a 244-kDa-parasite antigen that is crucial for parasite egress from an infected red blood cell (iRBC), which was published as a comprehensive, full-length Research Article in Science. In a parallel approach, we have screened phage display cDNA libraries constructed from parasites isolated at our Tanzanian field site with/without culture adaptation using positive selection with antibodies pooled from resistant two-year-olds and negative selection with antibodies pooled from susceptible children. We identified several independent cDNA clones encoding plasmodium falciparum glutamic acid reach protein (PfGARP) and plant-like calcium-dependent protein kinase (PfCDPK5) that were uniquely recognized by antibodies in resistant, but not susceptible sera. Our preliminary data demonstrate that PfGARP and PfCDPK5 is critical for parasite development in side RBC and egress respectively). PfGARP expresses on the surface of the trophozoite infected RBC and PfCDPK5 is expressed by merozoites as they rupture from erythrocytes. Antibodies against PfGARP and PfCDPK5 block parasite growth up to 99% in vitro, and ortholog vaccine of CDPK5 protect mice from parasitemia, and extend the survival of mice challenged with lethal P. berghei ANKA. Our vaccine discovery program has also identified several known invasion ligands (MSP-4 and MSP-7 collectively referred to as MSPs) In this application, we will evaluate these vaccine candidates with CDPK5 as single fusion protein (PfCDPK5-MSP4 & PfCDPK5-MSP7) in combination with PfGARP in in vitro assays and using multiple adjuvant systems in murine vaccine trials. The lead fusion antigen will be further evaluated for cell mediated immune response using TFRS depletion method in murine model. The deliverables from this study will be an adjuvant optimized tri-valent vaccine ready for Aotus/ P. falciparum challenge and Phase-1 clinical trial in human that targets the entry, intracellular development, and the exit of the parasite cycle in .