| dc.description.abstract | Malaria is an entirely preventable and treatable disease, yet it is endemic in nearly half of the countries around the world. One of the most important drug targets for this disease is the hemozoin formation pathway, a heme detoxification process found in the malaria parasite. In the search for novel hemozoin inhibitors, we developed a target based assay that closely mimics the biological conditions and screened three compound libraries. Hits were then tested in a phenotypic screen for antiplasmodial activity. These experiments, however, do not provide the information required to fully comprehend the in vivo drug mechanism of action. To validate hemozoin inhibition, we analyzed three types of parasitic heme: hemoglobin, intracellular free heme, and hemozoin. Compounds were confirmed hits upon observation of a rise in free heme and a decrease in hemozoin percentage, corresponding to parasite death. Our screening efforts resulted in high hit rates compared to other β-hematin inhibition assays. To explain this discrepancy, we examined the physiochemical properties required for β-hematin formation using detergents as models for lipids within the digestive vacuole. A longer hydrophilic tail on the detergents allowed for more efficient heme sequestration and solubilization in lipophilic regions, which formed hemozoin-like crystals. Furthermore, even though drug resistance is climbing as the parasite adapts to antimalarial treatment, the hemozoin formation pathway is still a valid target since the mechanism of inhibition is separate from the mechanism of resistance. Hemozoin inhibitors bind to the µ-oxo heme dimer, preventing crystallization throughout the ring stage of the parasite life cycle, when hemozoin begins to form as a result of hemoglobin degradation. However, the location and exact mechanism of heme-drug interaction may vary among hemozoin inhibitors. Additionally, we investigated how these compounds specifically interact with their biological target using fluorescent probes. In drug discovery, it is not only important to find molecules that possess in vitro antimalarial activity, but also to understand how drugs interact with their biological target in order to aid in compound optimization. | |
