Cellular and <i>in Vitro</i> Transport of Glutathione Conjugates by MRP

Shen, Hongxie; Paul, Saptarshi; Breuninger, Lisa M.; Ciaccio, Paul J.; Laing, Naomi; Helt, Marija; Tew, Kenneth D.; Kruh, Gary D.

doi:10.1021/bi960098n

Cited by 48 publications

(34 citation statements)

References 24 publications

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“…Using membrane vesicle transport assays, we demonstrate in this report that MRP also pumps a spectrum of natural product cytotoxic drugs. The specificity of MRP for both classes of compounds is consistent with our observations that daunorubicin transport is inhibited by drug glutathione conjugates and that APA-SG transport is inhibited by natural product drugs (29). Analysis of the kinetics of inhibition further suggests that these diverse substrates may share a common binding site on the transporter.…”

Section: Resultssupporting

confidence: 87%

“…This possibility was examined by determining the effect of drug glutathione conjugates on daunorubicin transport. We have observed in vitro transport of the glutathione analog APA-SG by MRP (29) and transport of the drug glutathione conjugate DNP-SG has also been reported (15,16). Fig.…”

Section: Resultssupporting

confidence: 58%

“…Competitive inhibition of daunorubicin uptake by these compounds was consistent with the broad specificity of the transporter and suggests that they share a common binding site with lipophilic drugs. (14)(15)(16)29). Using membrane vesicle transport assays, we demonstrate in this report that MRP also pumps a spectrum of natural product cytotoxic drugs.…”

Section: Resultsmentioning

confidence: 62%

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RETRACTED: ATP-dependent uptake of natural product cytotoxic drugs by membrane vesicles establishes MRP as a broad specificity transporter.

Paul

Breuninger

Tew

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

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Section: Resultssupporting

confidence: 87%

Section: Resultssupporting

confidence: 58%

Section: Resultsmentioning

confidence: 62%

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RETRACTED: ATP-dependent uptake of natural product cytotoxic drugs by membrane vesicles establishes MRP as a broad specificity transporter.

Paul

Breuninger

Tew

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

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“…Mutations of lysine residues in the Walker A motif of the first and second nucleotide binding domains of MRP decreased drug resistance levels compared with wild-type MRP (5), implying that MRP functions as an ATP-dependent drug pump. Its function has been studied widely using membrane vesicles prepared from MRP-expressing cells (11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23). It was found that anticancer drugs, such as vincristine, cannot be transported by MRP, unless glutathione is also present (13).…”

mentioning

confidence: 99%

Stimulation of ATPase Activity of Purified Multidrug Resistance-associated Protein by Nucleoside Diphosphates

Chang

Hou

Riordan

1998

Journal of Biological Chemistry

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Membrane vesicles prepared from cells expressing the multidrug resistance-associated protein (MRP) transport glutathione S-conjugates of hydrophobic substrates in an ATP dependent manner. Purified MRP possesses ATPase activity which can be further stimulated by anticancer drugs or leukotriene C4. However, the detailed relationship between ATP hydrolysis and drug transport has not been established. How the ATPase activity of MRP is regulated in the cell is also not known. In this report, we have examined the effects of different nucleotides on the ATPase activity of purified MRP. We have found that pyrimidine nucleoside triphosphates have little effect on enzymatic activity. In contrast, purine nucleotides dATP, dGTP, and adenosine 5-(␤,␥-imido)triphosphate function as competitive inhibitors. Somewhat unexpectedly, low concentrations of all the nucleoside diphosphates (NDPs) tested, except UDP, stimulate the ATPase activity severalfold. ADP or GDP at higher concentrations was inhibitory, reflecting NDP binding to the substrate site. On the other hand, the enhancement of hydrolysis at low NDP concentrations must reflect interactions with a separate site. Therefore, we postulate the presence of at least two types of nucleotide binding sites on the MRP, a catalytic site(s) to which ATP preferentially binds and is hydrolyzed and a regulatory site to which NDPs preferentially bind and stimulate hydrolysis. Interestingly, the stimulatory effects of drugs transported by MRP and NDPs are not additive, i.e. drugs are not able to further stimulate the NDP-activated enzyme. Hence, the two activation pathways intersect at some point. Since both nucleotide binding domains of MRP are likely to be required for drug stimulation of ATPase activity, the two sites that we postulate may also involve both domains.The multidrug resistance-associated protein (MRP) 1 gene was cloned from a multidrug-resistant cell line, H69AR, which was obtained by stepwise selection in increasing concentration of doxorubicin (1). A single open reading frame of 1531 amino acids was defined, and the protein was found to be a member of the ABC superfamily of transport systems (1) and to contain three membrane-associated domains and two nucleotide binding domains (2). As predicted, cells transfected with MRP cDNA become multidrug-resistant (3-10). Mutations of lysine residues in the Walker A motif of the first and second nucleotide binding domains of MRP decreased drug resistance levels compared with wild-type MRP (5), implying that MRP functions as an ATP-dependent drug pump. Its function has been studied widely using membrane vesicles prepared from MRP-expressing cells (11-23). It was found that anticancer drugs, such as vincristine, cannot be transported by MRP, unless glutathione is also present (13). Several natural substrates, such as the glutathione S-conjugate, LTC4 (11-13, 16, 20) and other conjugates including glutathione S-conjugated prostaglandin A1 and A2 (23), 17-␤-estradiol 17-␤-D-glucuronide (14), bile salt, and glucuronosylhyodeoxycholate...

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“…However, there is no direct evidence that MRP can bind unmodified forms of the drugs to which it confers resistance, and there are conflicting reports of its ability to transport these compounds (17)(18)(19). MRP can function as a primary active ATP-dependent transporter of cysteinyl leukotrienes, other organic glutathione conjugates including oxidized glutathione, as well as etoposide glucuronide, certain steroid glucuronides, and bile salt derivatives (18 -24).…”

mentioning

confidence: 99%

Reconstitution of ATP-dependent Leukotriene C4 Transport by Co-expression of Both Half-molecules of Human Multidrug Resistance Protein in Insect Cells

Gao

Loe²,

Grant³

et al. 1996

Journal of Biological Chemistry

128

111

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Multidrug resistance protein (MRP) confers a multidrug resistance phenotype similar to that associated with overexpression of P-glycoprotein. Unlike P-glycoprotein, MRP has also been shown to be a primary active ATP-dependent transporter of conjugated organic anions. The mechanism(s) by which MRP transports these compounds and increases resistance to natural product drugs is unknown. To facilitate studies on the structure and function of MRP, we have determined whether a baculovirus expression system can be used to produce active protein.

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Cellular and in Vitro Transport of Glutathione Conjugates by MRP

Cited by 48 publications

References 24 publications

RETRACTED: ATP-dependent uptake of natural product cytotoxic drugs by membrane vesicles establishes MRP as a broad specificity transporter.

RETRACTED: ATP-dependent uptake of natural product cytotoxic drugs by membrane vesicles establishes MRP as a broad specificity transporter.

Stimulation of ATPase Activity of Purified Multidrug Resistance-associated Protein by Nucleoside Diphosphates

Reconstitution of ATP-dependent Leukotriene C4 Transport by Co-expression of Both Half-molecules of Human Multidrug Resistance Protein in Insect Cells

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