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Updated: May 24, 2026
Characterization of Membrane Transporters by Heterologous Expression in E. coli and Production of Membrane Vesicles
Published on: December 31, 2019
Impact of probe compound in MRP2 vesicular transport assays
Heidi Kidron1, Gloria Wissel, Nenad Manevski
1Centre for Drug Research, Faculty of Pharmacy, University of Helsinki, P.O. Box 56, 00014 Helsinki, Finland. heidi.kidron@helsinki.fi
Abstract:
MRP2 is an efflux transporter that is expressed mainly in the canalicular membrane of hepatocytes, where it expels polar and ionic compounds into the bile. MRP2 is also present in the apical membrane of enterocytes and epithelial cells of proximal tubules of the kidney. Inhibition of MRP2 transport can lead to the accumulation of metabolites and other MRP2 substrates up to toxic levels in these cells. The transport properties of MRP2 are frequently studied with the vesicular transport assay. The assay identifies compounds that interact with MRP2 by measuring the effect of a compound on the transport of a radioactively labeled or fluorescent probe. We have compared the effect of eight selected test compounds (quercetin, disopyramide, paracetamol, indomethacin, diclofenac, estrone-3-sulfate, budesonide, and thioridazine) on the MRP2-mediated transport of three commonly used probes: 5(6)-carboxy-2,7-dichlorofluorescein, leukotriene C4 and estradiol-17-β-d-glucuronide (E217βG). Five of the test compounds had different probe-dependent effects on the MRP2-mediated transport, suggesting differences in the transport mechanism of the probes. Our results underline the complexity of substrate recognition by these efflux transporters and the difficulties in directly comparing results obtained with different assays, especially when different probes are used.
Insights
Multidrug resistance-associated protein 2 (MRP2) transport varies based on the probe used. This study highlights probe-dependent effects on MRP2 activity, complicating direct assay comparisons.
Area of Science:
- Pharmacology
- Biochemistry
- Cell Biology
Background:
- Multidrug resistance-associated protein 2 (MRP2) is a key efflux transporter in hepatocytes, enterocytes, and kidney proximal tubules.
- MRP2 expels polar and ionic compounds into bile, and its inhibition can cause toxic metabolite accumulation.
- Vesicular transport assays are commonly used to study MRP2 function by measuring probe transport inhibition.
Purpose of the Study:
- To investigate the probe-dependent effects of selected compounds on MRP2-mediated transport.
- To assess the complexity of substrate recognition by MRP2.
- To evaluate challenges in comparing MRP2 assay results using different probes.
Main Methods:
- Comparison of eight test compounds (quercetin, disopyramide, paracetamol, indomethacin, diclofenac, estrone-3-sulfate, budesonide, thioridazine).
- Assessment of effects on MRP2-mediated transport of three probes: 5(6)-carboxy-2,7-dichlorofluorescein, leukotriene C4, and estradiol-17-β-d-glucuronide (E217βG).
- Utilized vesicular transport assay methodology.
Main Results:
- Five test compounds exhibited probe-dependent effects on MRP2 transport.
- Observed variations in compound interactions with MRP2 depending on the probe used.
- Indicated distinct transport mechanisms or interactions for different probes.
Conclusions:
- MRP2 substrate recognition is complex and influenced by probe selection.
- Direct comparison of MRP2 transport assay results is challenging when using different probes.
- Results emphasize the need for careful consideration of probe choice in MRP2 studies.
