H.-J. Lee, H. Azargoshasb, P.G. Vekilov
University of Houston,
United States
Keywords: malaria, hematin crystallization, nonclassical inhibition mechanisms
Summary:
Malaria remains a significant public health challenge in equatorial regions of the world largely owing to the parasite's emerging resistance to the recently introduced drugs of the artemisinin (ART) family. In the human body most ART-derivative drugs are metabolized to dihydroartemisinin (DHA), which, in the parasite, after activation by heme, can form a hematin–dihydroartemisinin adduct (H-DHA). In search of strategies to circumvent malaria parasites drug resistance, we explore the mechanism employed by H-DHA to inhibit the synthesis of hemozoin crystals, the main constituent of the heme detoxification pathway of malaria parasites residing in the red blood cells. We supplement these insights with an examination of how tafenoquine (TQ), an antimalarial approved by the Food and Drug Administration in 2018 as a constituent of combinational antimalarial treatments, interacts with hemozoin formation. We show that both H-DHA and TQ block a fraction of the crystal growth sites, the kinks, and mildly inhibit the classical growth mode of beta-hematin, the synthetic analogue of hemozoin, by generation and spreading of layers. H-DHA and TQ exert opposing effects on the nucleation of hematin crystals: H-DHA strongly enhances crystal nucleation, whereas TQ potently suppresses it. The two drugs accomplish their nucleation control missions by enhancing or impeding the formation of mesoscopic hematin-rich clusters, the precursors of hematin crystal nucleation. The diverging effects on crystal nucleation and the nucleation precursors trigger in two distinct nonclassical pathways to suppress hematin crystal growth that transform H-DHA and TQ into potent hematin growth inhibitors. With H-DHA we find that hematin crystallites, whose nucleation is promoted by H-DHA, incorporate into large hematin crystals and suppress their growth, likely by straining the crystal lattice. A second consequence of H-DHA is the generation of macrosteps on hematin crystal surfaces that hinder growth. Importantly, the induced growth suppression is irreversible and persists even in the absence of H-DHA. TQ suppresses a nonclassical growth mode, which activates at elevated hematin concentrations, by impeding the formation of the hematin-rich clusters, the shared precursors of both crystal nucleation and nonclassical growth. Our findings suggest that the resistance to artemisinin-class drugs may rely on the reduced concentrations of hematin and H-DHA, which deactivate the dual non-classical mode of action of the adduct in the delayed-clearance parasite strains. Our findings on TQ suggest that this drug exerts a stage-specific inhibitory effect: during the ring stage of intraerythrocytic stage, it suppresses hematin nucleation, whereas in the trophozoite stage, it inhibits classical and nonclassical crystal growth. This dual-action mechanism highlights TQ’s ability to disrupt parasite survival at multiple points during the blood stage of infection.