UC San Diego

The antimalarial resistance arms race is alive and well. In 2022, the Plasmodium genus of parasites were responsible for 249 million cases of malaria, 608,000 of which were fatal (WHO, 2023). This increase in cases is attributed to lack of an effective vaccine and restricted access to preventative and treatment measures coming after the COVID-19 pandemic, but especially to the ongoing development of antimalarial resistant parasites. Thus, there is a dire need to create medicines that possess novel modes of action. And if malaria were ever to be eradicated, we need those that inhibit parasite growth at every stage of their life cycle. So, how do we know which compounds exhibit antimalarial activity in a novel way?

This dissertation highlights and applies strategies of antimalarial target identification. Chapter 1 describes omics approaches: in-vitro evolution and whole genome analysis, proteomic affinity chromatography, cellular thermal shift assay, metabolomic profiling. Chapter 2 uses these methods on imidazolopiperazines, a new class of antimalarials that possess a novel mode of action and are active against multiple stages of the parasite life cycle. Previous work evolving parasites against the GNF179 analog only presented multidrug resistance mechanisms; metabolomic profiling with the KAF156 analog did not present clear perturbations. Using proteomic affinity chromatography, thermal shift assay, and surface plasmon resonance, we identify GNF179 interaction with a putative dynamin-like GTPase. Having only been predicted to be essential in parasites, we also confirm that it is an essential P. falciparum gene via conditional knockdown. Molecular docking and GTPase activity assay suggest imidazolopiperazine binding to the N-terminal GTPase domain, yet a nonsense mutation that removes residues from the C-terminal end confers resistance. In Chapter 3, we investigate the significance of the transmembrane domains and C-terminal tail of this GTPase protein to imidazolopiperazine interaction. Through this research, we can better guide imidazolopiperazine development and anticipate resistant alleles.