The calcium channel blockers, 1,4-dihydropyridines, are substrates of the multidrug resistance-linked ABC drug transporter, ABCG2.
The human ATP-binding cassette transporter, ABCG2, confers resistance to multiple chemotherapeutic agents and also affects the bioavailability of different drugs. [(125)I]Iodoarylazidoprazosin (IAAP) and [(3)H]azidopine were used for photoaffinity labeling of ABCG2 in this study. We show here for the first time that both of these photoaffinity analogues are transport substrates for ABCG2 and that [(3)H]azidopine can also be used to photolabel both wild-type R482-ABCG2 and mutant T482-ABCG2. We further used these assays to screen for potential substrates or modulators of ABCG2 and observed that 1,4-dihydropyridines such as nicardipine and nifedipine, which are clinically used as antihypertensive agents, inhibited the photolabeling of ABCG2 with [(125)I]IAAP and [(3)H]azidopine as well as the transport of these photoaffinity analogues by ABCG2. Furthermore, [(3)H]nitrendipine and bodipy-Fl-dihydropyridine accumulation assays showed that these compounds are transported by ABCG2. These dihydropyridines also inhibited the efflux of the known ABCG2 substrates, mitoxantrone and pheophorbide-a, from ABCG2-overexpressing cells, and nicardipine was more potent in inhibiting this transport. Both nicardipine and nifedipine stimulated the ATPase activity of ABCG2, and the nifedipine-stimulated activity was inhibited by fumitremorgin C, suggesting that these agents might interact at the same site on the transporter. In addition, nontoxic concentrations of dihydropyridines increased the sensitivity of ABCG2-expressing cells to mitoxantrone by 3-5-fold. In aggregate, results from the photoaffinity labeling and efflux assays using [(125)I]IAAP and [(3)H]azidopine demonstrate that 1,4-dihydropyridines are substrates of ABCG2 and that these photolabels can be used to screen new substrates and/or inhibitors of this transporter.
Shukla
Suneet
S
Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Robey
Robert W
RW
Bates
Susan E
SE
Ambudkar
Suresh V
SV
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D001386:Azides / Q000378:metabolism* / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000378:metabolism*; D002460:Cell Line; D002734:Chlorophyll / Q000031:analogs & derivatives / Q000378:metabolism; D004095:Dihydropyridines / Q000378:metabolism* / Q000494:pharmacology; D004347:Drug Interactions; D006801:Humans; D008942:Mitoxantrone / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism*; D009529:Nicardipine / Q000378:metabolism / Q000494:pharmacology; D009543:Nifedipine / Q000378:metabolism; D019920:Photoaffinity Labels / Q000378:metabolism; D011224:Prazosin / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology
C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D001386:Azides; D002121:Calcium Channel Blockers; D004095:Dihydropyridines; D009363:Neoplasm Proteins; D019920:Photoaffinity Labels; D002734:Chlorophyll; C017040:8-azidoadenosine 5'-triphosphate; C041130:azidopine; D000255:Adenosine Triphosphate; C041552:azidoprazosin; D008942:Mitoxantrone; D009529:Nicardipine; D009543:Nifedipine; C032623:pheophorbide a; D011224:Prazosin