Characterization of the commercially-available fluorescent chloroquine-BODIPY conjugate, LynxTag-CQGREEN, as a marker for chloroquine resistance and uptake in a 96-well plate assay

Loh, Cheryl C. Y. and Suwanarusk, Rossarin and Lee, Yan Quan and Chan, Kitti W. K. and Choy, Kit-Ying and Rénia, Laurent and Russell, Bruce and Lear, Martin J. and Nosten, François H. and Tan, Kevin S. W. and Chow, Larry M. C. and Snounou, Georges (2014) Characterization of the commercially-available fluorescent chloroquine-BODIPY conjugate, LynxTag-CQGREEN, as a marker for chloroquine resistance and uptake in a 96-well plate assay. PLoS ONE, 9 (10). e110800. ISSN 1932-6203

Full content URL: http://dx.doi.org/10.1371/journal.pone.0110800

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Characterization of the commercially-available fluorescent chloroquine-BODIPY conjugate, LynxTag-CQGREEN, as a marker for chloroquine resistance and uptake in a 96-well plate assay
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Abstract

Chloroquine was a cheap, extremely effective drug against Plasmodium falciparum until resistance arose. One approach to reversing resistance is the inhibition of chloroquine efflux from its site of action, the parasite digestive vacuole. Chloroquine accumulation studies have traditionally relied on radiolabelled chloroquine, which poses several challenges. There is a need for development of a safe and biologically relevant substitute. We report here a commercially-available green fluorescent chloroquine-BODIPY conjugate, LynxTag-CQGREEN, as a proxy for chloroquine accumulation. This compound localized to the digestive vacuole of the parasite as observed under confocal microscopy, and inhibited growth of chloroquine-sensitive strain 3D7 more extensively than in the resistant strains 7G8 and K1. Microplate reader measurements indicated suppression of LynxTag-CQGREEN efflux after pretreatment of parasites with known reversal agents. Microsomes carrying either sensitive or resistant-type PfCRT were assayed for uptake; resistant-type PfCRT exhibited increased accumulation of LynxTag-CQGREEN, which was suppressed by pretreatment with known chemosensitizers. Eight laboratory strains and twelve clinical isolates were sequenced for PfCRT and Pgh1 haplotypes previously reported to contribute to drug resistance, and pfmdr1 copy number and chloroquine IC50s were determined. These data were compared with LynxTag-CQGREEN uptake/fluorescence by multiple linear regression to identify genetic correlates of uptake. Uptake of the compound correlated with the logIC50 of chloroquine and, more weakly, a mutation in Pgh1, F1226Y.

Additional Information:Editor: Georges Snounou, Université Pierre et Marie Curie, France
Keywords:chloroquine, drug labelling, bioassays, drug resistance, commercialisation, JCOpen
Subjects:F Physical Sciences > F165 Biomolecular Chemistry
C Biological Sciences > C720 Biological Chemistry
C Biological Sciences > C500 Microbiology
Divisions:College of Science > School of Chemistry
ID Code:17096
Deposited On:17 Apr 2015 08:10

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