Multidrug Resistance Protein (MRP) 1 and MRP3 Attenuate Cytotoxic and Transactivating Effects of the Cyclopentenone Prostaglandin, 15-Deoxy-Δ^12,14Prostaglandin J₂ in MCF7 Breast Cancer Cells

Abstract

One of the most potent cyclopentenone prostaglandins, 15-deoxy-Delta(12,14)prostaglandin J(2) (15-d-PGJ(2)), has been shown to be cytotoxic in some tumor cells and, as a ligand of peroxisome proliferator activated receptor gamma (PPARgamma), to influence the transcriptional regulation of several genes. We examined whether a glutathione conjugate of 15-d-PGJ(2), 15-d-PGJ(2)-SG, is formed and if the glutathione conjugate efflux pumps, MRP1 and MRP3, could transport this conjugate, thereby attenuating the cytotoxicity and transactivating activity of 15-d-PGJ(2) in MCF7 breast cancer cells. Formation of 15-d-PGJ(2)-SG was demonstrated both in vitro and in cells, and its structure was determined by ESI/MS and NMR. Expression of MRP1 and MRP3 was achieved by stable transduction of parental MCF7 cells. Membrane vesicles derived from these cells supported efficient, ATP-dependent transport of 15-d-PGJ(2)-SG (K(M) 1.4 and 2.9 microM for MRP1 and MRP3, respectively). When compared with parental, MRP-minus MCF7 cells, expression of MRP1 and MRP3 conferred approximately 2-fold protection from 15-d-PGJ(2) cytotoxicity. 15-d-PGJ(2)-mediated transcriptional activation was evaluated in cells transiently transfected with a reporter gene under the transcriptional control of a PPAR responsive element. Treatment of parental MCF7 cells with 15-d-PGJ(2) resulted in a time-dependent induction of reporter gene activity-induction that was measurable with concentrations of added 15-d-PGJ(2) as low as 100 nM. In contrast, expression of MRP1 or MRP3 abolished 15-d-PGJ(2)-dependent reporter gene induction. Depletion of intracellular glutathione reversed MRP1- and MRP3-mediated attenuation of 15-d-PGJ(2) cytotoxicity and transactivation. These data indicate that MRP1 and MRP3 can modulate the biological effects of 15-d-PGJ(2), and likely other cyclopentenone prostaglandins, in a glutathione-dependent manner. The results are consistent with a mechanism for the attenuation of the biological activities of 15-d-PGJ(2) that involves the formation and active efflux of its glutathione conjugate, 15-d-PGJ(2)-SG.

Casein Kinase 2α Regulates Multidrug Resistance-Associated Protein 1 Function via Phosphorylation of Thr249

“…Media containing viral particles was collected from above the cells and used to transduce MCF7/WT cells. Transduced cells were grown under G418 selection, and clones were selected and analyzed for MRP1 expression via Western blotting as described previously (Paumi et al, 2003). At least two clones for each mutation were analyzed in cytotoxicity, transport, and doxorubicin accumulation assays that yielded similar results.…”

“…Cell lysates were then placed in a new tube containing 50 l of 6ϫ SDS-PAGE loading buffer and frozen at Ϫ20°C. Frozen samples were later thawed and incubated at 37°C for 1 h; 70 l of each sample was run on a 7% SDS-PAGE gel followed by Western blotting procedure as described previously (Paumi et al, 2003). MRP1 protein was probed with rat MRP1r1 antibody (1:1000).…”

Intestinal transporters for endogenic and pharmaceutical organic anions: the challenges of deriving in-vitro kinetic parameters for the prediction of clinically relevant drug–drug interactions

“…p-Aminohippuric acid [92] 372 Bilirubin biglucuronide [93] 0.8 a Daunorubicin [94,95] 1.4 Atazanavir [59] Bilirubin bisglucuronide [93] Bilirubin monoglucuronide [93] 0.4 a Grepafloxacin [96] Benzbromarone [97,98] Bilirubin monoglucuronide [93] E217bG [99] 22 Irinotecan [100] Benzylpenicillin [98] DHEAS [101] 5 Estradiol sulfate [102] 0.4 Methotrexate [98,103,104] 930/2150 Daunorubicin [99]~8a DNP-GS [105] 3.6 E23SO417bG [99] 1.7 Doxorubicin [99]~5 0 a E217bG [99,[105][106][107] 1.5-4.8 E316bG [99] 45 Mitoxantrone [108] Flurbiprofen [109] Estrone-3-sulfate [102] 0.7/4.2 E317bG [99] 1.4 Saquinavir [110] Furosemide [98] Folic acid [103] Estrone-3-sulfate [102] 0.5 Vinblastine [111] Glibenclamide [77] Glutathione [97,112] Glycholate [99] Vincristine [95,113] Indinavir [114] Leucovorin [103] Glycolithocholate-3-sulfate [99] 1.4 Indometacin …”

ABC drug transporter expression and functional activity in trophoblast-like cell lines and differentiating primary trophoblast

“…The ATP-binding cassette (ABC) superfamily of transporters appears to be particularly important in this context, being specifically dedicated to removal of xenobiotics and toxic endogenous metabolites from the fetal organism (11,22). Although the ABC pumps were initially described as drug efflux pumps, conferring resistance to cancer cells from druginduced apoptosis (13), they have more recently been shown to be implicated in cellular protection from various proapoptotic injuries, such as exposure to endogenous toxins or hypoxic damage (10,19). Human placenta expresses several members of the ABC superfamily; of these, the most effective for drug transfer are P-glycoprotein (Pgp), encoded by the multidrug resistance 1 (MDR1) gene, multidrug resistance proteins (MRPs) 1 and 2, and the breast cancer resistance protein (BCRP; see Refs.…”