A validated bioluminescence-based assay for the rapid determination of the initial rate of kill for discovery antimalarials

Objectives:

A future treatment for uncomplicated malaria will contain at least one component that exerts a rapid rate of kill (RoK). We describe here the validation and application of a simple, robust and rapid bioluminescence-based assay for the determination of the initial RoK in intraerythrocytic asexual stages of Plasmodium falciparum.

Methods:

A modification to the concentration-response bioluminescence (here termed bioluminescence relative rate of kill, BRRoK) assay, utilizing exposure to fold-IC50 concentrations (0.33x to 9x), is used to monitor the immediate cytocidal effect of 372 open source compounds for antimalarial drug discovery available through the Medicine for Malaria Venture’s Malaria Box.

Results:

Antimalarial drugs that exert a rapid cytocidal effect produce a concentration dependent loss of bioluminescence signal that correlates with available in vitro and in vivo estimates of parasite clearance time and parasite reduction ratio. Following the measurement of IC50 for the Malaria Box compounds in Dd2luc30 , the BRRoK assay was used to identify and rank 372 compounds for their initial cytocidal activity. Fifty three compounds in the Malaria Box show an initial relative RoK greater than that of chloroquine, with 17 of these with an initial relative RoK greater than that of dihydroartemisinin.

Conclusion:

The BRRoK assay provides a rapid assay format for the estimation of a key pharmacodynamic property of antimalarial drug action. The simplicity and robustness of the assay suggests it would be readily scalable for high throughput screening and a critical decision-making tool for antimalarial drug development.