Effect of cyclosporin and verapamil on the cellular kinetics of daunorubicin

Hu, Xiangdong; Luise, M. De; Martin, Tanya; Zalcberg, John

doi:10.1016/0277-5379(90)90159-q

Cited by 16 publications

(9 citation statements)

References 7 publications

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“…Mean fluorescence intensity was recorded for each tested population (after correcting for non-specific binding) to provide an estimate of relative MRK-16 binding. (Hu et al, 1990b). Cells were washed three times with PBS and resuspended in fresh medium for 1 h at 37°C in a humidifier before performing drug accumulation studies.…”

Section: Pgp Expressionmentioning

confidence: 99%

Induction of MDR1 gene expression by anthracycline analogues in a human drug resistant leukaemia cell line

Slater

Rischin

et al. 1999

Br J Cancer

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SummaryThe effects of 4-demethoxydaunorubicin (idarubicin, IDA) and MX2, a new morpholino-anthracycline, on up-regulation of the MDR1 gene in the low-level multidrug resistant (MDR) cell line CEM/A7R were compared at similar concentrations (IC 10 , IC 50 and IC 90 ) over a short time exposure (4 and 24 h). The chemosensitivity of each drug was determined by a 3-day cell growth inhibition assay. Compared with epirubicin (EPI), IDA and MX2 were 17-and eightfold more effective in the CEM/A7R line respectively. No cross-resistance to 5-FU was seen in the CEM/A7R line. Verapamil (5 µM) and PSC 833 (1 µM), which dramatically reversed resistance to EPI in the CEM/A7R line, had no sensitizing effect on the resistance of this line to MX2, but slightly decreased resistance to IDA. The sensitivity to 5-FU was unchanged by these modulators. The induction of MDR1 mRNA expression by IDA, MX2 and 5-FU was analysed by Northern blotting and semiquantitatively assessed by scanning Northern blots on a phosphorimager. The relative level of MDR1 expression was expressed as a ratio of MDR1 mRNA to the internal RNA control glyceraldehyde-3-phosphate dehydrogenase (GAPDH). IDA, MX2 and 5-FU differentially up-regulated MDR1 mRNA in the CEM/A7R line in a dose-dependent manner. Both IDA and MX2 induced MDR1 expression within 4 h. 5-FU up-regulated MDR1 expression only when drug exposure was prolonged to 24 h. Based on MRK 16 binding, flow cytometric analysis of P-glycoprotein (Pgp) expression paralleled the increase in MDR1 mRNA levels. For the three anthracyclines, the increase in MDR1 expression was stable in cells grown in the absence of drug for more than 3 weeks after drug treatment. The induction of MDR1 expression by 5-FU was transient, associated with a rapid decrease in the increased Pgp levels which returned to baseline 72 h after the removal of 5-FU. This study demonstrates that MDR1 expression can be induced by analogues of anthracyclies not pumped by Pgp, and that this induction appears to be stable despite a 3-week drug-free period.

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Section: Pgp Expressionmentioning

confidence: 99%

Induction of MDR1 gene expression by anthracycline analogues in a human drug resistant leukaemia cell line

Slater

Rischin

et al. 1999

Br J Cancer

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“…The discrepancy remains, however, even when measurements are made using the same procedure. Thus, Hu et al (1990) ABBREVIATIONS: P-gp, P-glycoprotein; EATC, Ehrlich ascites tumor cell; AM, acetoxymethyl ester; K app , apparent half-saturation concentration (e.g., IC 50 ); I, inhibitor concentration; k i , the pump rate for pumping from the inside; k i,0 , the uninhibited pump rate for pumping from the inside; K i , intrinsic affinity for P-gp at the inner leaflet of the cell membrane; k o , the pump rate for (preemptive) pumping from the outer face of the membrane; k o,0 , the uninhibited pump rate for pumping from the outer face of the membrane; K o , intrinsic affinity for P-gp at the outer face of the membrane (preemptive pumping); p, the leak rate of a compound that crosses the membrane by passive diffusion; S i , intracellular substrate concentration; S o , extracellular substrate concentration; GF120918, elacridar; XR9576, tariquidar. …”

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confidence: 99%

“…4 M (CEM/ vinblastine1000 cells) (Hu et al, 1990), with a range of intermediate values (Cardarelli et al, 1995;Lan et al, 1996;Litman et al, 1997). For verapamil, the IC 50 for reversal of daunorubicin extrusion by P-gp ranges from 0.4 M (SW2780AD cells) (Javaheri et al, 1983) to 10 M (CEM/ vinblastine1000 cells) (Hu et al, 1990), and the in vivo situation is further complicated because oxidative metabolites may act as substrates and inhibitors of P-gp (Pauli-Magnus et al, 2000).Part of the variability can be ascribed to different measures of the reverser's effect on drug pumping, either by assessing the effect of the blocker on reversing resistance to cytotoxicity (Lee et al, 1994), or on reversing the drug accumulation deficit due to P-gp (Lan et al, 1996). The discrepancy remains, however, even when measurements are made using the same procedure.…”

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confidence: 99%

mentioning

confidence: 99%

“…The discrepancy remains, however, even when measurements are made using the same procedure. Thus, Hu et al (1990) measured the effect of cyclosporin A and verapamil on daunorubicin accumulation in two resistant variants of CEM/CCRF cells and found, that for cyclosporin A, IC 50 increased from 1.7 M in the moderately resistant subline CEM/vinblastine100 to 4 M for the highly resistant CEM/vinblastine1000 line. For another P-gp blocker, GF120918, Chen et al (2000) found that its IC 50 for reversal of Tc-tetrofosmin transport increased from 40 to 385 nM for a low P-gp-expressing and a high P-gp-expressing drug-resistant carcinoma cell line, respectively.…”

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confidence: 99%

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Pumping of Drugs by P-Glycoprotein: A Two-Step Process?

Litman

Skovsgaard²,

Stein³

2003

J Pharmacol Exp Ther

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The apparent inhibition constant, K app , for the blockade of P-glycoprotein (P-gp) by four drugs, verapamil, cyclosporin A, XR9576 (tariquidar), and vinblastine, was measured by studying their ability to inhibit daunorubicin and calcein-AM efflux from four strains of Ehrlich cells with different levels of drug resistance and P-gp content. For daunorubicin as a transport substrate, K app was independent of [P-gp] for verapamil but increased strictly linearly with [P-gp] for vinblastine, cyclosporin A, and XR9576. A theoretical analysis of the kinetics of drug pumping and its reversal shows that K app for inhibition should increase linearly with the amount of pumps present in the membrane for a reverser that inhibits pumping from the cytoplasmic face. In contrast, if the reverser acts by blocking transport from the outer face, i.e., preemptively, K app should be independent of the number of pumps present. The experimental data suggest that verapamil blocks pumping at the extracellular face of the membrane, whereas the other three blockers act on pumping from the cytoplasmic phase. The maximum degree of inhibition was the same for all four blockers; thus, they do not act in parallel but rather, in serial, i.e., a drug that is pumped from the cytoplasmic phase has to pass the preemptive route upon leaving the cell. Our results are consistent with the Sauna-Ambudkar two-step model for pumping by P-gp. We suggest that the vinblastine/cyclosporin A/XR9576-binding site accepts daunorubicin at the cytoplasmic face and transfers it to the verapamil-binding site, from where daunorubicin is emptied at the extracellular surface.Multidrug resistance (MDR) constitutes a major barrier to effective chemotherapy of cancer (Bates et al., 1996). Although alternate resistance mechanisms exist (Borst et al., 2000;Litman et al., 2001;Sparreboom et al., 2003), the MDR phenotype is most often characterized by a reduced intracellular drug level and overexpression of the ATP-binding cassette transporter P-glycoprotein (P-gp) that is capable of transporting a wide range of xenobiotic compounds out of tumor cells (Chabner and Fojo, 1998;Seelig and Landwojtowicz, 2000). The transport activity of P-gp can be blocked and multidrug resistance reversed by the addition of substrate analogs, or so-called chemosensitizers or modulators (Skovsgaard et al., 1984;Krishna and Mayer, 2001).In the literature, variable values for IC 50 (the drug concentration that gives half-maximal inhibition) for P-glycoprotein reversal are reported for the same drug in different models.For example, for the blocker cyclosporin A, the IC 50 for inhibition of daunorubicin pumping by P-gp ranges from 0.04 M (P388/ADR cells) to ca. 4 M (CEM/ vinblastine1000 cells) (Hu et al., 1990), with a range of intermediate values (Cardarelli et al., 1995;Lan et al., 1996;Litman et al., 1997). For verapamil, the IC 50 for reversal of daunorubicin extrusion by P-gp ranges from 0.4 M (SW2780AD cells) (Javaheri et al., 1983) to 10 M (CEM/ vinblastine1000 cells) (Hu et al., 1990),...

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Irreversibility of drug resistance in VAD‐refractory myeloma

Dimopoulos

Alexanian

1992

American J Hematol

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Effect of cyclosporin and verapamil on the cellular kinetics of daunorubicin

Cited by 16 publications

References 7 publications

Induction of MDR1 gene expression by anthracycline analogues in a human drug resistant leukaemia cell line

Induction of MDR1 gene expression by anthracycline analogues in a human drug resistant leukaemia cell line

Pumping of Drugs by P-Glycoprotein: A Two-Step Process?

Irreversibility of drug resistance in VAD‐refractory myeloma

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