Understanding Plasmodium falciparum's multidrug resistance protein through the characterization of a novel antimalarial class. - PROJECT SUMMARY Malaria is a deadly disease that affects nearly half the world’s population. In 2021, there was an estimated 619,000 deaths due to malaria. Currently, the front-line treatment for uncomplicated malaria is Artemisinin-based Combination Therapies (ACTs), where an artemisinin-related compound is paired with a slower acting partner drug. Despite successful use of this drug regimen, resistance has been identified to all clinically approved antimalarials. Because of increasing antimalarial resistance, there is an urgent need to discover new targets and chemotherapies potent against malaria, in addition to understanding mechanisms of resistance to antimalarials. A common form of resistance occurs with mutations in the parasite’s multidrug resistance protein (PfMDR1). PfMDR1 is located on the surface of the digestive vacuole, an acidic organelle where the parasite digests hemoglobin. While PfMDR1 is a major driver of antimalarial resistance, we currently do not understand how different mutations of pfmdr1 modulate resistance in the malaria parasite. The current hypothesis is that PfMDR1 can non-specifically import drug into the digestive vacuole, thus making the compound inactive or preventing it from reaching its molecular target. However, this hypothesis is not fully explanatory of the collateral drug sensitivity that we observe in parasites with PfMDR1 mutations. For example, copy number variation and single nucleotide polymorphisms (SNPs) located on this gene can result in resistance to certain antimalarials while simultaneously increasing sensitivity to other compounds. We have discovered a novel compound, PRC1590, potent against malaria that recapitulates this collateral drug sensitivity. Through in vitro selection of resistance to this compound, we have identified that its resistance is due to a SNP on pfmdr1. We hypothesize that the SNP causing resistance to PRC1590 results in decreased import of the drug into the digestive vacuole. In Aim 1, we will characterize resistance of PRC1590 and its localization in the malaria parasite with fluorescence microscopy. We will also perform cross-resistance screening to better understand how compounds like PRC1590 modulate resistance through PfMDR1. For Aim 2, we will use chemoproteomic approaches to determine the mechanism of action (MOA) of PRC1590. A better understanding of PRC1590 resistance, along with its localization and mechanism of action will allow us to better understand how compounds modulate resistance through PfMDR1 mutations, which will be informative for partner drug pairings that exploit collateral drug sensitivity associated with PfMDR1 resistance. This research will take place in Dr. Belen Cassera’s lab at the University of Georgia (UGA), who has expertise in drug discovery, and target identification and validation which will allow us to determine the mechanism of action of PRC1590. Additionally, UGA houses the Center for Tropical and Emerging Global Diseases, which has one of the largest concentrations of parasitologists at a US university. Completion of this project will allow for the trainee to develop into an independent scientist by gaining molecular and chemical biology techniques, and mentorship experience through training of undergraduate researchers.
