Indomethacin-mediated reversal of multidrug resistance and drug efflux in human and murine cell lines overexpressing MRP, but not P-glycoprotein

Abstract

Summary Decreased accumulation of the fluorescent dye BCECF [2', 7'-bis-(2-carboxyethyl)-5-(6)-carboxyfluorescein] characterized murine and human multidrug-resistant cell lines overexpressing the multidrug resistance protein (MRP). Indomethacin (10 gM), a known cyclooxygenase and glutathione-S-transferase inhibitor as well as a modulator of anion transport, increased accumulation and blocked efflux of BCECF in MRP-expressing murine and human cells. The drug did not affect P-glycoprotein (P-gp)-mediated export of rhodamine 123. The indomethacin effect on BCECF efflux was not reversed by the addition of exogenous prostaglandins, suggesting that the drug acts by a mechanism other than decreasing prostaglandin synthesis. Indomethacin also increased multidrug susceptibility of both murine and human cell lines overexpressing MRP, but not those displaying P-gp-associated resistance. In addition, indomethacin modulated the decreased vincristine accumulation in cells expressing MRP, but not in those expressing P-gp. These data suggest that indomethacin is a specific inhibitor of MRP, possibly functioning by inhibition of glutathione-S-transferase or, alternatively, by direct competition with the drug at the transport site.Keywords: BCECF; chemosensitizer; modulator; transport inhibitors Multidrug resistance in mammalian cells has been associated with altered drug transport. Active efflux or altered intracellular sequestration has been attributed to membrane-associated P-glycoprotein (Pgp) or the multidrug resistance protein (MRP) (Inaba et al, 1979; Zamon et al, 1994). We have previously demonstrated that murine and human cell lines that overexpressed MRP also showed altered accumulation and increased efflux of the free acid form of the fluorescent dye BCECF (Draper et al, 1996). BCECF is loaded into the cells as the membrane-permeable acetoxymethyl (AM) ester, BCECF-AM. This lipophilic ester derivative can cross cell membranes, entering the cytoplasm and various organelles, where the AM ester is hydrolysed by intracellular esterases to release BCECF free acid, which is normally retained by the cell. We used the altered BCECF accumulation phenotype of cell lines overexpressing MRP to look for drugs that could modulate MRP function. One of the drugs examined first was indomethacin as it has previously been reported to modulate efflux of BCECF from cultured epithelial cell monolayers, although the mechanism is unknown (Collington et al., 1992). We report here on the ability of indomethacin, a member of the non-steroidal family of anti-inflammatory drugs (NSAID), to specifically modulate MRP-mediated resistance.

Indomethacin-induced activation of the death receptor-mediated apoptosis pathway circumvents acquired doxorubicin resistance in SCLC cells

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