Abstract
ABC multidrug transporters are key players in cancer multidrug resistance and in general xenobiotic elimination, thus their functional assays provide important tools for research and diagnostic applications. In this study we have examined the potential interactions of three key human ABC multidrug transporters with PhenGreen diacetate (PGD), a cell permeable fluorescent metal ion indicator. The non-fluorescent, hydrophobic PGD rapidly enters the cells and, after cleavage by cellular esterases, in the absence of quenching metal ions, PhenGreen (PG) becomes highly fluorescent. We found that in cells expressing functional ABCG2, ABCB1, or ABCC1 transporters, cellular PG fluorescence is strongly reduced. This fluorescence signal in the presence of specific transporter inhibitors is increased to the fluorescence levels in the control cells. Thus the PG accumulation assay is a new, unique tool for the parallel determination of the function of the ABCG2, ABCB1, and ABCC1 multidrug transporters. Since PG has very low cellular toxicity, the PG accumulation assay also allows the selection, separation and culturing of selected cell populations expressing either of these transporters.
Highlights
Several members of the ATP-binding cassette (ABC) superfamily of membrane transporters are working as efflux pumps for a large variety of xenobiotics and drugs
The three key ABC efflux transporters involved in human cancer drug resistance and drug metabolism are the ABCB1 (P-glycoprotein, Pgp), the ABCC1 and the ABCG2 proteins, their evaluation has a major importance in drug development and clinical diagnostics [1,2,3,4,5,6]
A new fluorescent assay for parallel measurements of the ABCG2, ABCB1 and ABCC1 transporter functions
Summary
Several members of the ATP-binding cassette (ABC) superfamily of membrane transporters are working as efflux pumps for a large variety of xenobiotics and drugs. These transporters are important players in multidrug resistance against anti-cancer therapeutic compounds, and significantly modify the absorption, distribution, metabolism, excretion and toxicity (ADME-Tox) parameters for numerous therapeutic agents. The three key ABC efflux transporters involved in human cancer drug resistance and drug metabolism are the ABCB1 (P-glycoprotein, Pgp), the ABCC1 (multidrug resistance protein 1, MRP1) and the ABCG2 (breast cancer resistance protein, BCRP) proteins, their evaluation has a major importance in drug development and clinical diagnostics [1,2,3,4,5,6]. Due to the promiscuity of these proteins in drug binding and transport, the molecular mechanisms of drug interactions
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