title: Development of a rapid and reliable assay for in vitro determination of compound cidality against the asexual stages of Plasmodium falciparum.
abstract: The pace of anti-malarial drug discovery is often impeded due to the lack of tools to determine the cidality of compounds in vitro. An anti-malarial compound must have a cidal mode of action, i.e. kill parasites, in order to quickly reduce parasite load. A static compound that merely inhibits growth must be identified early on in the discovery cascade. In this paper, we describe a high-throughput fluorescent assay for determination of the cidality of an anti-malarial compound. The assay works on the principle that cultures treated with a static compound will exhibit re-growth while treatment with a cidal compound leads to a marked reduction in parasite number. Parasite cultures are treated with the drug for 48 or 72 h following which the drug is washed off. Cultures are allowed to recover in drug-free media for 72 h and DNA content estimated using the fluorescent dye SyBR Green I. Following estimation of IC50 and IC99 values, we find that the IC99/IC50 ratio is a reliable indicator of the cidality of a compound. Cidal compounds like artemisinin and chloroquine display an IC99/IC50 ratio <5 while the ratio for a static compound like atovaquone is <5. This correlation holds true for various anti-malarial drugs with known modes of action. Importantly, the IC99/IC50 ratio drops to <5 when a compound becomes cidal in action with longer duration of treatment. The assay is robust, reliable and provides a fast and effective means for prioritizing cidal compounds for progression along the drug discovery cascade.
authors: Pavithra
authors: P
authors: Viswanath
authors: Sapna
authors: S
authors: Morayya
authors: Nikhil
authors: N
authors: Rautela
authors: Achyut
authors: A
authors: Sinha
mesh_terms: D000962:Antimalarials / Q000494:pharmacology*; D052160:Benzothiazoles; D003959:Diamines; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical; D020128:Inhibitory Concentration 50; D061848:Optical Imaging; D009930:Organic Chemicals; D010963:Plasmodium falciparum / Q000502:physiology*; D011804:Quinolines
chemical_list: D000962:Antimalarials; D052160:Benzothiazoles; D003959:Diamines; D009930:Organic Chemicals; D011804:Quinolines; C098022:SYBR Green I