Abstract
The human multidrug resistance proteins MRP4 and MRP5 are organic anion transporters that have the unusual ability to transport cyclic nucleotides and some nucleoside monophosphate analogs. Base and nucleoside analogs used in the chemotherapy of cancer and viral infections are potential substrates. To assess the possible contribution of MRP4 and MRP5 to resistance against these drugs, we have investigated the transport mediated by MRP4 and MRP5. In cytotoxicity assays, MRP4 conferred resistance to the antiviral agent 9-(2-phosphonomethoxyethyl)adenine (PMEA) and high-performance liquid chromatography analysis showed that, like MRP5, MRP4 transported PMEA in an unmodified form. MRP4 also mediated substantial resistance against other acyclic nucleoside phosphonates, whereas MRP5 did not. Apart from low-level MRP4-mediated cladribine resistance, the cytotoxicity of clinically used anticancer nucleosides was not influenced by overexpression of MRP4 or MRP5. In contrast, MRP5 mediated efflux of the pyrimidine-based antiviral 2′,3′-dideoxynucleoside 2′,3′-didehydro-2′,3′-dideoxythymidine 5′-monophosphate (d4TMP) and its phosphoramidate derivative alaninyl-d4TMP from cells loaded with the 2′,3′-didehydro-2′,3′-dideoxythymidine prodrugs cyclosaligenyl-d4TMP and aryloxyphosphoramidate d4TMP (So324), respectively. Moreover, only inside-out membrane vesicles derived from MRP5-overexpressing cells accumulated alaninyl-d4TMP. Cellular efflux and vesicular uptake studies were carried out to further compare transport mediated by MRP4 and MRP5 and showed that dipyridamole, dilazep, nitrobenzyl mercaptopurine riboside, sildenafil, trequinsin and MK571 inhibited MRP4 more than MRP5, whereas cyclic nucleotides and monophosphorylated nucleoside analogs were equally poor inhibitors of both pumps. These results strongly suggest that the affinity of MRP4 and MRP5 for nucleotide-based substrates is low.
Footnotes
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↵1 Current address: Netherlands Ophthalmic Research Institute, KNAW, Meibergdreef 47, 1105 BA Amsterdam, the Netherlands.
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This work was supported by Dutch Cancer Society grants NKI 2001–2473 to P.B and J.W., 1998–1764 to P.B. and a grant of the European Commission (HPAW-CT-2002–90001 and QLRT-2000–30291) to J.B.
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G.R. and P.W. contributed equally to this work.
- Abbreviations:
- MRP
- multidrug resistance protein
- bis-POM-PMEA
- bis(pivaloyloxymethyl)-9-(2-phosphonomethoxyethyl)adenine
- cGMP
- guanosine 3′,5′-cyclic monophosphate
- cPr-PMEDAP
- cyclopropyl-PMEDAP
- cycloSAL-d4TMP
- cyclosaligenyl-d4T-5′-monophosphate
- ddC
- 2′,3′-dideoxycytidine
- d4T
- 2′,3′-didehydro-2′,3′-dideoxythymidine
- d4TMP
- 2′,3′-didehydro-2′,3′-dideoxythymidine 5′-monophosphate
- DMEM
- Dulbecco's modified Eagle's medium
- E217βG
- estradiol glucuronide
- HEK
- human embryonic kidney
- HPLC
- high-performance liquid chromatography
- MK571
- 3-[[3-[2-(7-chloroquinolin-2-yl)vinyl]phenyl]-(2-dimethylcarbamoylethylsulfanyl)methylsulfanyl] propionic acid
- PMEA
- 9-(2-phosphonomethoxyethyl)adenine
- PMEDAP
- 9-(2-phosphonomethoxyethyl)-2,6-diaminopurine
- PMEG
- 9-(2-phosphonomethoxyethyl)guanine
- PDE
- phosphodiesterase
- So324
- aryloxyalaninylphosphoramidate of d4T-5′-monophosphate (d4TMP)
- Received November 18, 2002.
- Accepted January 31, 2003.
- The American Society for Pharmacology and Experimental Therapeutics
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