Quantitative high throughput screen for delayed death inhibitors of the malarial parasite plastid, 96 hour incubation
The goal of this screen is to discover new antimalarial compounds that act by inhibiting the development of the apicoplast in the malarial parasite Plasmodium falciparum. The biochemical processes that make this organelle essential for erythrocytic stage parasites are not well understood. However, antibiotics such as azithromycin and tetracycline, which target the apicoplast translational more ..
BioActive Compounds: 165
Depositor Specified Assays
NIH Molecular Libraries Probe Production Network [MLPCN]
NIH Chemical Genomics Center [NCGC]
MLPCN Grant: R21 NS059500
Assay Provider: David Fidock and Eric Ekland, Columbia University
The goal of this screen is to discover new antimalarial compounds that act by inhibiting the development of the apicoplast in the malarial parasite Plasmodium falciparum. The biochemical processes that make this organelle essential for erythrocytic stage parasites are not well understood. However, antibiotics such as azithromycin and tetracycline, which target the apicoplast translational machinery, have a potent antimalarial effect. The killing caused by these drugs is unusual in that it does not appear to affect the first generation of parasites that are exposed to the drug, but rather manifests itself in their progeny.
We have developed a cell-based assay that measures parasite growth based on the expression of an integrated copy of a firefly luciferase reporter. To detect small molecules that cause this 'delayed death' phenotype, erythrocytes infected with the luciferase-expressing parasites were incubated with compounds for either one or two generations, corresponding to 48 and 96 hours, respectively. Compounds that inhibit parasite growth in the second generation, but not the first, should be enriched in antimalarials that target the apicoplast. Growth inhibition is detected by a decrease in luciferase activity
Four microliters of culture medium (RPMI 1640 with 0.5% w/v Albumax (GIBCO), 24 mM sodium bicarbonate and 10 ug/mL gentamycin) were dispensed by a Multi-drop Combi into white solid 1536-well plates (Grenier) and 23 nL compound was added by a pin tool. Four microliters of infected erythrocytes (2% hematocrit, 0.1% parasitemia final concentration) in culture medium were dispensed and the plates incubated for 96 hours at 37 C in 5% CO2. Two microliters of luciferase detection reagent was added and luminescence was detected by a ViewLux (PerkinElmer) reader
1. Compounds are first classified as having full titration curves, partial modulation, partial curve (weaker actives), single point activity (at highest concentration only), or inactive. See data field "Curve Description". For this assay, apparent inhibitors are ranked higher than compounds that showed apparent activation.
2. For all inactive compounds, PUBCHEM_ACTIVITY_SCORE is 0. For all active compounds, a score range was given for each curve class type given above. Active compounds have PUBCHEM_ACTIVITY_SCORE between 40 and 100. Inconclusive compounds have PUBCHEM_ACTIVITY_SCORE between 1 and 39. Fit_LogAC50 was used for determining relative score and was scaled to each curve class' score range.
* Activity Concentration. ** Test Concentration.
Data Table (Concise)