The relative rate of kill of the MMV Malaria Box compounds provides links to the mode of antimalarial action and highlights scaffolds of medicinal chemistry interest

Objectives: Rapid rate-of-kill (RoK) is a key parameter in the target candidate profile 1 (TCP1) for the next-generation antimalarial drugs for uncomplicated malaria, termed Single Encounter Radical Cure and Prophylaxis (SERCaP). TCP1 aims to rapidly eliminate the initial parasite burden, ideally as fast as artesunate, but minimally as fast as chloroquine. Here we explore whether the relative RoK of the Medicine for Malaria Venture (MMV) Malaria Box compounds are linked to their mode of action (MoA) and identify scaffolds of medicinal chemistry interest. Methods: We used a Bioluminescence Relative RoK (BRRoK) assay over 6 and 48h, with exposure to equipotent-IC50 concentrations, to compare the cytocidal effects of Malaria Box compounds to benchmark antimalarials. Results: BRRoK assay data demonstrate the following relative RoK from fast to slow: inhibitors of 31 PfATP4 > parasite hemoglobin catabolism > DHFR-TS > DHODH > bc1 complex. Core scaffold clustering analyses reveal intrinsic rapid cytocidal action for diamino-glycerols and 2- (aminomethyl)phenol, but slow action for 2-phenylbenzimidazoles, 8-hydroxyquinolines, and triazolopyrimidines. Conclusions: This study provides proof of principle that a compound’s RoK is related to its MoA and that the target’s intrinsic RoK is also modified by factors affecting a drug’s access to it. Our findings highlight that as we use medicinal chemistry to improve potency, we can also improve the RoK for some scaffolds. Our BRRoK assay provides the necessary throughput for drug discovery and a critical decision-making tool to support development campaigns. Finally, two scaffolds, diamino-glycerols and 2-phenylbenzimidazoles, exhibit fast cytocidal action, inviting medicinal chemistry improvements towards TCP1 candidates.