Aryl N-acetamide compounds exert antimalarial activity by acting as agonists of rhomboid protease Pf ROM8 and cation channel Pf CSC1

With resistance to current frontline antimalarials spreading globally, new drug candidates need to be discovered to populate the antimalarial drug development pipeline. We previously screened the Medicines for Malaria Venture Pathogen Box for compounds that prevent Plasmodium falciparum parasites from exiting and invading human erythrocytes, steps essential for the proliferation of parasites in the blood, which causes disease. Compound MMV020512 (M-512) was identified in this screen and live cell imaging here established that it does not specifically inhibit invasion but likely inhibits intraerythrocytic parasite growth. M-512 resistance selection in parasites led to the identification of mutations in the membrane protease Pf ROM8 and the cation ion channel Pf CSC1. Pf ROM8 was validated as a target of M-512 when a L562R putative resistance mutation was engineered into wildtype parasites reproducing the resistance phenotype. Knockdown of wildtype Pf ROM8, the L562R mutant and CSC1 reduced parasite growth, indicating the proteins are functionally important. Counterintuitively, the Pf ROM8 and Pf CSC1 knockdown parasites became more resistant to M-512 suggesting that the compound is an agonist of both proteins which may form a functional complex and that dysregulation of this complex is deleterious to parasite growth.