New drugs are urgently required for the treatment of malaria, due to the emergence of multi-drug resistant forms of the causative parasite, Plasmodium falciparum. Recent drug discovery programs utilising high-throughput phenotypic screens have identified thousands of small molecules that have antiparasitic activity against P. falciparum in vitro, and a representative subset of these, known as the ‘Malaria Box’, have been released openly to the research community . However, little is known about the mechanism of action of these compounds, restricting further development of these hit compounds. 

Systems biology techniques offer an ideal platform to investigate mechanisms of drug action in a hypothesis-free manner. This approach is ideal for the de novo identification of drug mechanisms where no prior knowledge about the mode of action is available. Specifically, metabolomics provides a snapshot of the metabolic state of the parasite after drug treatment, and can reveal the direct impact of a test compound on metabolism. 

LCMS-based untargeted metabolomics methods for P. falciparum cultures were optimised for 96-well plate format, allowing analysis of the metabolic phenotype induced by 100 known and novel antimalarials. Many of the novel compounds induced metabolic phenotypes that clustered with known antimalarials. In particular, inhibition of pyrimidine biosynthesis was a common mechanism of antimalarial action. Furthermore, depletion of unique parasite peptides and lipids indicated inhibition of parasite-specific pathways that may provide attractive new targets for drug discovery. This work demonstrates the suitability of metabolomics approaches for medium-throughput screening of the impact of drug candidates on cellular biochemistry, and revealed mechanisms of action associated with many of these novel antimalarial compounds.