Dexniguldipine-HCl is a potent allosteric inhibitor of [3H]vinblastine binding to P-glycoprotein of CCRF ADR 5000 cells

Malkhandi, Joy; Ferry, David; Boer, Rainer; Gekeler, Volker; Ise, W; Keer, David J.

doi:10.1016/0922-4106(94)90015-9

Cited by 30 publications

(20 citation statements)

References 21 publications

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“…Indeed, alterations of drug resistance profiles, particularly segregation of vinblastine resistance from daunomycin and colchicine resistance, have been observed in mutant P-glycoproteins (41)(42)(43). This, together with the observations of Tamai and Safa (44) that azidopine noncompetitively interacts with vinblastine and cyclosporin A binding to P-glycoprotein and observations of Ferry and co-workers (45,46), that the vinca alkaloid binding site is allosterically coupled to binding sites for dihydropyridines and taxanes, might reflect the presence of different drugbinding sites on P-glycoproteins for different classes of transported substrates. In this perspective it is interesting to note that colchicine noncompetitively inhibits Pdr5p-mediated rhodamine 6G fluorescence quenching in yeast plasma membranes whereas the vinca alkaloids vinblastine and vincristine were competitive inhibitors.…”

Section: Resultsmentioning

confidence: 78%

Anticancer Drugs, Ionophoric Peptides, and Steroids as Substrates of the Yeast Multidrug Transporter Pdr5p

Kołaczkowski

Rest

Cybularz-Kolaczkowska

et al. 1996

Journal of Biological Chemistry

299

288

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Pdr5p is the yeast Saccharomyces cerevisiae ATPbinding cassette transporter conferring resistance to several unrelated drugs. Its high overproduction in Pdr1p transcription factor mutants allows us to study the molecular mechanism of multidrug transport and substrate specificity. We have developed new in vivo and in vitro assays of Pdr5p-mediated drug transport. We show that in spite of little sequence homology, and inverted topology in respect to that of mammalian P-glycoproteins, Pdr5p shares with them common substrates. Pdr5p extrudes rhodamines 6G and 123, from intact yeast cells in an energy-dependent manner. Plasma membrane preparations from a Pdr5p-overproducing strain exhibit ATP hydrolysis-dependent, osmotically sensitive rhodamine 6G fluorescence quenching. The quenching is competitively inhibited by micromolar concentrations of many anticancer drugs, such as vinblastine, vincristine, taxol, and verapamil, and of ionophoric peptides as well as steroids. In contrast, other anticancer drugs, like colchicine and some multidrug resistance modifiers, such as quinidine, exert noncompetitive inhibition. Our experimental system opens new possibilities for the analysis of structure-function relationship of multidrug transporter substrates and inhibitors.

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Section: Resultsmentioning

confidence: 78%

Anticancer Drugs, Ionophoric Peptides, and Steroids as Substrates of the Yeast Multidrug Transporter Pdr5p

Kołaczkowski

Rest

Cybularz-Kolaczkowska

et al. 1996

Journal of Biological Chemistry

299

288

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“…Several lines of evidence measuring transport, binding and ATPase activity of P-gp have pointed to the presence of multiple drug interaction sites (Ferry et al, 1992(Ferry et al, , 1995Spoelstra et al, 1992;Malkhandi et al, 1994;Ayesh et al, 1996;Orlowski et al, 1996;Martin et al, 1997). CP100-356 is a diaminoquinazoline and amongst the most potent MDR-modulating agents reported (Kajiji et al, 1994), hence providing specificity and selectivity of binding displacement.…”

Section: Discussionmentioning

confidence: 99%

“…The diaminoquinazoline, CP100-356, is clearly a high affinity ligand for P-gp, with K i values of 58 ± 2 nM and 94 ± 21 nM for the P-gp1a and P-gp1b isoforms respectively. Nicardipine, a 1,4-dihydropyridine, which binds to a distinct site allosterically-linked to the vinblastine site (Malkhandi et al, 1994;Martin et al, 1997) (K i = 128 ± 20 nM) with approximately twice the affinity observed for P-gp1b-containing membranes (K i = 239 ± 48 nM; P < 0.05). In contrast to CP100-356, progesterone was found to have a low and almost identical affinity for both isoforms (K i = 1.9 ± 0.3 µM and 1.8 ± 0.2 µM for P-gp1a and P-gp1b respectively).…”

Section: Displacement Of [ 3 H]vinblastine Binding By Pharmacologicalmentioning

confidence: 99%

The equilibrium and kinetic drug binding properties of the mouse P-gp1a and P-gp1b P-glycoproteins are similar

1999

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SummaryThe gene encoding the multidrug resistance P-glycoprotein (P-gp) is duplicated in rodent species and the functional basis for this remains unresolved. Despite a high sequence similarity, the mouse P-gp1a and P-gp1b isoforms show distinct patterns of tissue distribution which suggest a specific role of the P-gp1b isoform in steroid transport. In the present study possible biochemical differences between the isoforms were directly investigated at the level of drug interaction. There was no detectable difference in the affinity or binding capacity of the two isoforms towards [ 3 H]vinblastine at equilibrium. Similarly, the rate at which [ 3 H]vinblastine associates with P-gp was indistinguishable between the two isoforms. Some modest differences were observed in the relative abilities of the multidrug-resistant (MDR) reversing agents CP100-356, nicardipine and verapamil to displace equilibrium [ 3 H]vinblastine binding to P-gp1a and P-gp1b. The steroid hormone progesterone displayed a low affinity (K i = 1.2 ± 0.2 µM for P-gp1a and 3.5 ± 0.5 µM for P-gp1b), suggesting an unlikely role as a physiological substrate. Thus the mouse isoforms do not appear to exhibit functional differences at the level of initial substrate interaction with protein.

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“…From our data it seems obvious, however, that chemosensitisers binding to P-gp with high affinity (such as DNIG) may be particularly valuable for clinical use as MDR modulators. Nonetheless, the bisindolylmaleide represents a new tool structure possibly binding to a site on P-gp different from the binding site of DNIG, which supposedly acts as an allosteric inhibitor of P-gp according to Ferry et al (1992) and Malkhandi et al (1994 …”

Section: Western Immunoblottingmentioning

confidence: 99%

Effects of the selective bisindolylmaleimide protein kinase C inhibitor GF 109203X on P-glycoprotein-mediated multidrug resistance

Gekeler

Boer²,

Überall³

et al. 1996

Br J Cancer

113

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Summary Inhibition of protein kinase C (PKC) is discussed as a new approach for overcoming multidrug resistance (MDR) in cancer chemotherapy. For evaluation of this concept we applied the bisindolylmaleimide GF 109203X, which shows a highly selective inhibition of PKC isozymes a, /31, ,B2, y, 6 and e in vitro. The efficacy of this compound in modulation of MDR was examined using several P-glycoprotein (P-gp)-overexpressing cell lines including a MDRI-transfected HeLa clone, and was compared with the activities of dexniguldipine-HCl (DNIG) and dexverapamil-HCl (DVER), both of which essentially act via binding to P-gp. As PKCa has been suggested to play a major role in P-gp-mediated MDR, cell lines exhibiting different expression levels of this PKC isozyme were chosen. On crude PKC preparations or in a cellular assay using a cfos(-71 I)CAT-transfected NIH 3T3 clone, the inhibitory qualities of the bisindolylmaleimide at submicromolar concentrations were demonstrated. At up to 1 gm final concentrations of the PKC inhibitor GF 109203X, a concentration at which many PKC isozymes should be blocked substantially, no cytotoxic or MDR-reversing effects whatsoever were seen, as monitored by 72 h tetrazolium-based colorimetric MTT assays or a 90 min rhodamine 123 accumulation assay. Moreover, depletion of PKCa by phorbol ester in HeLa-MDR1 transfectants had no influence on rhodamine 123 accumulation after 24 or 48 h. MDR reversal activity of GF 109203X was seen at higher final drug concentrations, however. Remarkably, [3H]vinblastine-sulphate binding competition experiments using P-gp-containing crude membrane preparations demonstrated similar dose dependencies as found for MDR reversion by the three modulators, i.e. decreasing efficacy in the series dexniguldipine-HCl>dexverapamil-HCl>GF 109203X. Similar interaction with the P-gp in the micromolar concentration range was revealed by competition of GF 109203X with photoincorporation of [3H]azidopine into P-gp-containing crude membrane preparations. No significant effect of the PKC inhibitor on MDR1 expression was seen, which was examined by cDNA-PCR. Thus, the bisindolylmaleimide GF 109203X probably influences MDR mostly via direct binding to P-gp. Our work identifies the bisindolylmaleimide GF 109203X as a new type of drug interacting with P-gp directly, but does not support the concept of a major contribution of PKC to a P-gp-associated MDR, at least using the particular cellular model systems and the selective, albeit general, PKC inhibitor GF 109203X.

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Dexniguldipine-HCl is a potent allosteric inhibitor of [3H]vinblastine binding to P-glycoprotein of CCRF ADR 5000 cells

Cited by 30 publications

References 21 publications

Anticancer Drugs, Ionophoric Peptides, and Steroids as Substrates of the Yeast Multidrug Transporter Pdr5p

Anticancer Drugs, Ionophoric Peptides, and Steroids as Substrates of the Yeast Multidrug Transporter Pdr5p

The equilibrium and kinetic drug binding properties of the mouse P-gp1a and P-gp1b P-glycoproteins are similar

Effects of the selective bisindolylmaleimide protein kinase C inhibitor GF 109203X on P-glycoprotein-mediated multidrug resistance

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