ABSTRACT To discover leads for next-generation chemoprotective antimalarial drugs, we tested >500,000 compounds for their ability to inhibit liver-stage development of luciferase-expressing Plasmodium parasites (681 with an IC50 < 1 µM). Cluster analysis identified potent and previously unreported scaffold families as well as other series previously associated with chemoprophylaxis. Further testing through multiple phenotypic assays that predict stage-specific and multispecies antimalarial activity revealed compound classes that are likely to provide symptomatic relief from blood-stage and others that only prevent malaria. Target identification using functional assays, in vitro evolution or metabolic profiling of the most potent blood stage-active scaffolds revealed multiple mitochondrial inhibitors but also compounds with likely new mechanisms of action. The total dataset provided hundreds of new chemotypes and scaffold families (compounds sharing a core structure) that may be used in further drug development for the treatment and prevention of malaria. Here we propose to perform hit to lead optimization on 1-2 scaffold families per year, synthesizing approximately ~750 compounds. Select compounds (10%) will be tested for microsomal stability, protein binding, hERG, and CYP inhibition. Pharmacokinetic and prophylactic animal model testing will be performed on ~10 compounds per year. In addition, we will explore whether promising leads can be converted to a prodrug form that can be given as intramuscular injection which provides long- lasting chemoprevention. Hit to lead optimization will be accompanied by investigation into the mechanism of action of six scaffold families using a suite of different methods including in vitro evolution and whole genome analysis. We aim to develop treatments that may prevent malaria from developing, that are convenient to use and which are superior to current medicines. .