Optimization and Characterization of the Antimalarial of Activity Aryl N-Acetamides that are Susceptible to Mutations in ROM8 and CSC1

New antimalarials are needed due to the threat of emerging resistance against existing antimalarial therapies. A screen for compounds that block the development of P. falciparum asexual ring stage parasites uncovered the N-aryl acetamide structural class. We investigated the structure-activity relationship and defined key modifications comprising a N-substituted biaryl linker, a N-methyl amino acetamide, and either a 3-endocyclic nitrogen or a 2-fluoro on the central ring which produced WEHI-326 with potent antimalarial activity. WEHI-326 was found to have a moderate barrier to resistance and a moderate rate of sexual kill and potently inhibited both gametocytes and gametes, and in-turn blocked the transmission of parasites to the mosquito. The optimization of the metabolic stability and aqueous solubility of the N-aryl acetamide class will be a future challenge to achieve improved efficacy in a malaria mouse model. Forward genetics and cross-resistance profiling determined that parasites resistant to the N-aryl acetamide analogs had mutations in rhomboid protease 8 (ROM8) and a putative cation channel, CSC1. Structural modelling showed that the mutations mapped to the periphery of the catalytic cleft of ROM8 and a parasitophorous vacuole-facing region of CSC1, but the significance of the mutation locations and role of these proteins in parasite survival remains unclear. WEHI-326 will be an important tool to unravel the role of ROM8 and CSC1 in P. falciparum asexual and sexual stage development.