Dmd # 19661 1 Modulation of Human Mrp1 (Abcc1) And
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DMD Fast Forward. Published on December 13, 2007 as DOI: 10.1124/dmd.107.019661 DMD FastThis Forward. article has not Published been copyedited on andDecember formatted. The13, final 2007 version as maydoi:10.1124/dmd.107.019661 differ from this version. DMD # 19661 1 MODULATION OF HUMAN MRP1 (ABCC1) AND MRP2 (ABCC2) TRANSPORT ACTIVITIES BY ENDOGENOUS AND EXOGENOUS GLUTATHIONE- CONJUGATED CATECHOL METABOLITES Andrew J. Slot, Dana D. Wise, Roger G. Deeley, Terrence J. Monks, and Susan P.C. Cole Downloaded from Department of Pathology & Molecular Medicine, and Division of Cancer Biology & Genetics, dmd.aspetjournals.org Queen’s University, Ontario, Canada (A.J.S., R.G.D., S.P.C.C.); Department of Pharmacology & Toxicology, College of Pharmacy, University of Arizona Health Sciences Center, Tucson, Arizona (D.D.W., T.J.M.) at ASPET Journals on September 29, 2021 Copyright 2007 by the American Society for Pharmacology and Experimental Therapeutics. DMD Fast Forward. Published on December 13, 2007 as DOI: 10.1124/dmd.107.019661 This article has not been copyedited and formatted. The final version may differ from this version. DMD # 19661 2 Running title: MRP transport and GSH-conjugated catechol metabolites Corresponding author: Susan P.C. Cole, Division of Cancer Biology & Genetics, Queen’s University Cancer Research Institute, Kingston, ON, Canada, K7L 3N6, Tel.: 613-533-2636; Fax: 613-533-6830, e-mail address: [email protected]. Number of text pages: 33 Downloaded from Number of tables: 3 Number of figures: 6 dmd.aspetjournals.org Number of references: 40 Number of words in the Abstract: 215 Number of words in the Introduction: 732 at ASPET Journals on September 29, 2021 Number of words in the Discussion: 1598 DMD Fast Forward. Published on December 13, 2007 as DOI: 10.1124/dmd.107.019661 This article has not been copyedited and formatted. The final version may differ from this version. DMD # 19661 3 ABBREVIATIONS: 2-GS-CA, 2-(glutathion-S-yl)-caffeic acid; 2-NAC-CA, 5-(N-acetylcystein-S-yl)-caffeic acid; 2-OH-1-GS-E1, 2-hydroxy-1-(glutathion-S-yl)-estrone; 2-OH-1-GS-E2, 2-hydroxy-1-(glutathion-S-yl)-17β-estradiol; 2-OH-4-GS-E2, 2-hydroxy-4-(glutathion-S-yl)-17β-estradiol; Downloaded from 4-OH-2-GS-E2, 4-hydroxy-2-(glutathion-S-yl)-17β-estradiol; 5-GS-N-Me-α-MeDA, 5-(glutathion-S-yl)-N-methyl-α-methyldopamine; 5-NAC-N-Me-α-MeDA, 5-(N-acetylcystein-S-yl)-N-methyl-α-methyldopamine; dmd.aspetjournals.org ABC, ATP-binding cassette; α-MeDA, α-methyldopamine; CA, caffeic acid; at ASPET Journals on September 29, 2021 E217βG, 17β-estradiol 17-(β-D-glucuronide); GSH, glutathione; HEK, human embryonic kidney; LTC4, leukotriene C4; MDMA, methylenedioxymethamphetamine; MRP, multidrug resistance protein; NAC, N-acetylcysteine; N-Me-α-MeDA, N-methyl-α-methyldopamine; DMD Fast Forward. Published on December 13, 2007 as DOI: 10.1124/dmd.107.019661 This article has not been copyedited and formatted. The final version may differ from this version. DMD # 19661 4 Abstract: Members of the multidrug resistance protein (MRP/ABCC) subfamily of ATP-binding cassette proteins transport a wide array of anionic compounds, including sulfate, glucuronide and glutathione (GSH) conjugates. The present study tested the ATP-dependent vesicular transport of leukotriene C4 and 17β-estradiol 17-(β-D-glucuronide) (E217βG) mediated by the MRP1 and MRP2 transporters in the presence of six potential modulators from three different classes of GSH-conjugated catechol metabolites: the ecstasy metabolite 5-(glutathion-S-yl)-N-methyl-α- Downloaded from methyldopamine (5-GS-N-Me-α-MeDA); the caffeic acid metabolite 2-(glutathion-S-yl)-caffeic acid (2-GS-CA); and four GSH conjugates of 2-OH and 4-OH estrogens (GS-estrogens). MRP1- dmd.aspetjournals.org mediated E217βG transport was inhibited in a competitive manner with a relative order of potency of GS-estrogens (IC50 < 1 µM) > 2-GS-CA (IC50 3 µM) > 5-GS-N-Me-α-MeDA (IC50 31 µM). MRP2-mediated transport was inhibited with a similar order of potency, except the 2- at ASPET Journals on September 29, 2021 OH-4-GS-estradiol and 4-OH-2-GS-estradiol conjugates were approximately 50- and 300-fold less potent, respectively. Transport activity was unaffected by N-acetylcysteine conjugates of N- Me-α-MeDA and CA. The position of GSH conjugation appears important since all four GS- estrogen conjugates tested were potent inhibitors of MRP1 transport, but only the 2-OH-1-GS- estradiol and 2-OH-1-GS-estrone conjugates were potent inhibitors of MRP2-mediated transport. In conclusion, we have identified three new classes of MRP1 and MRP2 modulators, and demonstrated that one of these, the estrogen conjugates, shows unanticipated differences in their interactions with the two transporters. DMD Fast Forward. Published on December 13, 2007 as DOI: 10.1124/dmd.107.019661 This article has not been copyedited and formatted. The final version may differ from this version. DMD # 19661 5 Nine multidrug resistance proteins (MRPs) belong to the “C” subfamily of the ATP- binding cassette (ABC) transporter superfamily, and although they share moderate sequence identity (30% – 50%), their substrate specificities only partially overlap (Haimeur et al., 2004). With the exception of the liver, MRP1 is ubiquitously expressed, and in polarized cells it is almost always found on basolateral membranes (Leslie et al., 2005). However, in endothelial cells at the blood-brain barrier, it is expressed on apical membranes (Dallas et al., 2006). Downloaded from Physiologically, MRP1 mediates the release of the pro-inflammatory mediator leukotriene C4 (LTC4) from mast cells (Bartosz et al., 1998; Wijnholds et al., 1997). It also has an established role in mediating the efflux of conjugated phase II metabolites, including many, but not all dmd.aspetjournals.org glucuronide-, sulfate- and GSH-conjugates (Haimeur et al., 2004; Leslie et al., 2005). MRP2 shares 49% sequence identity with MRP1; however, unlike MRP1, it is expressed in a limited number of tissues such as liver, kidney and placenta, and in polarized cells, it localizes to at ASPET Journals on September 29, 2021 apical membranes (Leslie et al., 2005). The most important physiological role of MRP2 is mediating the biliary excretion of organic anion conjugates, primarily bilirubin glucuronides (Konig et al., 1999; Leslie et al., 2005). Despite the differences in their membrane and tissue localization, MRP1 and MRP2 transport many of the same conjugated metabolites, including LTC4 and 17β-estradiol 17-(β-D-glucuronide) (E217βG). However, the relative affinity of MRP2 for these metabolites is significantly lower than the affinity of MRP1 (Cui et al., 1999; Konig et al., 1999). The ring-substituted methamphetamine derivative, methylenedioxymethamphetamine (MDMA, ecstasy) (Fig. 1A) is a popular drug of abuse among Western youth. Although systemic administration of MDMA to rats leads to selective serotonin neurotoxicity, direct injection of MDMA or one of its primary catechol metabolites, α-methyl-dopamine (α-MeDA), DMD Fast Forward. Published on December 13, 2007 as DOI: 10.1124/dmd.107.019661 This article has not been copyedited and formatted. The final version may differ from this version. DMD # 19661 6 fails to produce any long lasting neurotoxic effects (McCann and Ricaurte, 1991). It has now been established that GSH-conjugated metabolites of MDMA contribute to the observed neurotoxicity. Thus, administration of the GSH conjugate 5-GS-α-MeDA to rats reproduces both the behavioral and short term alterations in brain catecholamine levels observed after systemic MDMA administration (Miller et al., 1996). Little is known about the disposition of MDMA metabolites, but previous research has suggested a GSH conjugate uptake mechanism present at the blood-brain barrier is likely involved (Bai et al., 2001). Downloaded from Caffeic acid (3,4-dihydroxycinnaminic acid, CA) (Fig. 1B) is a non-flavonoid catecholic acid found in relatively large quantities in fruits and vegetables. Absorbed CA appears to be mainly dmd.aspetjournals.org glucuronidated, sulfated, or O-methylated to ferulic acid (Mateos et al., 2006). Incubation of CA with liver microsomes and isolated hepatocytes also leads to the formation of several GSH conjugates (Galati et al., 2002; Moridani et al., 2002). Both mono- and di-substituted GSH at ASPET Journals on September 29, 2021 conjugates of caftaric acid and CA are present in relatively high amounts in grape juice and wine (Singleton et al., 1985). Otherwise, little is known about the metabolism and disposition of CA and its metabolites. However, CA can act as an anti-inflammatory agent by inhibiting 5- lipoxygenase, which in turn inhibits the synthesis of pro-inflammatory cytokines, such as the MRP1 substrate LTC4 (Koshihara et al., 1984) . Catechol estrogens (Fig. 1C), although structurally dissimilar from MDMA metabolites or CA, have a similar potential to form conjugates with GSH when oxidized to their corresponding quinones (Butterworth et al., 1996). Prolonged exposure to catechol estrogens is thought to play a role in carcinogenesis, although the underlying mechanisms are unknown. However, 4-OH estrogen quinones are known to form covalent adducts with guanine, leading to depurinating DNA adducts (Abel et al., 2004; Roy and Liehr, 1999; Yue et al., 2003). Furthermore, catechol DMD Fast Forward. Published on December 13, 2007 as DOI: 10.1124/dmd.107.019661 This article has not been copyedited and formatted. The final version may differ from this version. DMD # 19661 7 estrogens can enhance endogenous DNA adduct formation and free-radical generation due to redox cycling between the catechol and quinone forms, which ultimately leads to DNA damage (Roy and Liehr, 1999; Yue et al., 2003). Beyond the shared ability of the three chemically distinct groups of catechol metabolites described above to form conjugates with GSH, relatively little is known about their further disposition.