Antimalarial cytoskeletal targeting with broad apicomplexan activity.

Abstract

Malaria is a devastating disease that resulted in an estimated 610,000 deaths in 2024, the majority being children under the age of five. Here, we use KNX-115 to illustrate multistage antiparasitic activity upon targeting the cytoskeletal enzyme <i>Plasmodium falciparum</i> myosin A (PfMyoA). KNX-115 inhibits purified actin-activated ATPase with a potency in the low nanomolar range and >50-fold selectivity against cardiac, skeletal, and smooth muscle myosins. KNX-115 traps PfMyoA in a state that binds weakly to actin. A 2.35 Å resolution structure of KNX-115 bound to PfMyoA reveals critical interactions contributing to its mechanism of action. Importantly, in vitro evolution data reveal that KNX-115 engages PfMyoA as a sole cellular target. Inhibiting PfMyoA blocks the development of the blood and liver stages of laboratory strains of <i>P. falciparum</i>, with no liver cell toxicity, sporozoite cell traversal and motility, and sporozoite development in the mosquito. Inhibiting PfMyoA completely kills parasites after 96 h of treatment. Furthermore, KNX-115 is equally effective at inhibiting a panel of <i>Plasmodium</i> strains resistant to experimental and marketed antimalarials and shows inhibitory activity against <i>P. falciparum</i> circulating isolates from the Brazilian Amazon. Inhibiting PfMyoA with KNX-115 also blocks the blood stage of a laboratory strain of <i>Plasmodium vivax</i>. In line with the evolutionary identity of MyoA among various apicomplexan parasites, KNX-115 also inhibits <i>Cryptosporidium</i> and <i>Eimeria</i> MyoA in vitro and is an effective inhibitor of <i>Cryptosporidium</i>, <i>Toxoplasma,</i> and <i>Eimeria</i> cellular growth, with EC₅₀s similar to those found for blood and liver stage <i>Plasmodium</i>.