Malaria parasites need to traffic proteins to various cellular regions such as organelles, membranes and even onto the cell exterior. Blood-stage Plasmodium parasites have established an additional destination for their proteins which is out into the cytoplasmic compartment of their host red blood cell (RBC). P. falciparum parasites invest almost 10% of their proteome into modifying their RBC to acquire plasma nutrients, avoid host immunity and likely to perform a myriad of yet-undefined functions. The Plasmodium Translocon of Exported Proteins (PTEX) is an essential protein complex which resides in the parasitophorous vacuole membrane enveloping the intracellular parasites. PTEX acts as a conduit through which exported proteins are transported into the RBC cytoplasm. Due to the essential nature of PTEX and protein trafficking more generally, this pathway could be developed as an excellent drug target.
The Medicines for Malaria Venture (MMV) Malaria Box is a library of 400 compounds with anti-malarial activity. We screened the MMV Malaria Box for inhibitors of P. falciparum protein secretion and export. For this screen parasites were transfected with a bioluminescent Nanoluciferase fusion protein containing an export sequence, allowing trafficking of the protein into the host RBC. After drug treatment, differential fractionation of the cells allowed measurement of the fraction of bioluminescent protein trafficked into the parasite, parasitophorous vacuole and RBC compartments. Using this screen, several compounds were shortlisted for their ability to inhibit protein export to the host RBC. In this work we used a variety of assays to further characterise the activity of these shortlisted export inhibitors and to examine their effect on parasite activity. Further, we aim to discover the targets and mechanism of action of these compounds in hopes of developing them into future antimalarials.