A Novel Fluorescence Resonance Energy Transfer-Based Screen in High-Throughput Format To Identify Inhibitors of Malarial and Human Glucose Transporters.
Antimicrob Agents Chemother
; 60(12): 7407-7414, 2016 12.
Article
in En
| MEDLINE
| ID: mdl-27736766
ABSTRACT
The glucose transporter PfHT is essential to the survival of the malaria parasite Plasmodium falciparum and has been shown to be a druggable target with high potential for pharmacological intervention. Identification of compounds against novel drug targets is crucial to combating resistance against current therapeutics. Here, we describe the development of a cell-based assay system readily adaptable to high-throughput screening that directly measures compound effects on PfHT-mediated glucose transport. Intracellular glucose concentrations are detected using a genetically encoded fluorescence resonance energy transfer (FRET)-based glucose sensor. This allows assessment of the ability of small molecules to inhibit glucose uptake with high accuracy (Z' factor of >0.8), thereby eliminating the need for radiolabeled substrates. Furthermore, we have adapted this assay to counterscreen PfHT hits against the human orthologues GLUT1, -2, -3, and -4. We report the identification of several hits after screening the Medicines for Malaria Venture (MMV) Malaria Box, a library of 400 compounds known to inhibit erythrocytic development of P. falciparum Hit compounds were characterized by determining the half-maximal inhibitory concentration (IC50) for the uptake of radiolabeled glucose into isolated P. falciparum parasites. One of our hits, compound MMV009085, shows high potency and orthologue selectivity, thereby successfully validating our assay for antimalarial screening.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Plasmodium falciparum
/
Monosaccharide Transport Proteins
/
Protozoan Proteins
/
Fluorescence Resonance Energy Transfer
/
Small Molecule Libraries
/
High-Throughput Screening Assays
/
Glucose
/
Antimalarials
Type of study:
Prognostic_studies
Limits:
Humans
Language:
En
Journal:
Antimicrob Agents Chemother
Year:
2016
Document type:
Article