Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.3.44 (P-glycoprotein)
13,344 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Classic multidrug resistance is characterized by a decrease in the intracellular concentration of drugs in resistant cells as compared to sensitive cells. This is correlated with the presence of P-glycoprotein in the membrane. P-glycoprotein is responsible for an active efflux of drug. In this study we investigated the correlation between P-glycoprotein and influx of daunorubicin. Four Ehrlich ascites tumour cell lines selected in vivo for resistance to daunorubicin were investigated. The sublines EHR2/0.1, EHR2/0.2, passage no. 12 of EHR2/0.8, EHR2/0.4, and passage no. 72 of EHR2/0.8 were 6-, 6-, 5-, 33-, and 35-fold resistant to daunorubicin, respectively. All sublines overexpressed P-glycoprotein as determined with Western blot. Influx was measured over 40 sec. In glucose-enriched medium influx was significantly decreased in all but one of the resistant sublines. A correlation between P-glycoprotein, degrees of resistance, and influx was demonstrated in four sublines. Comparing influx experiments with efflux experiments (Nielsen et al., Biochem Pharmacol 1994, 47, 2125-2135) we found a linear relationship between influx and efflux in the resistant sublines (r = 0.97). Verapamil (5.5 microM, 11.0 microM) increased influx significantly in all resistant sublines, whereas the drug had no effect on sensitive cells. Verapamil (3.3 microM) increased influx in the EHR2/0.8 (passage no. 72) subline to the level of sensitive cells. Comparing this result with efflux experiments, verapamil was found to increase influx preferentially. Depletion of energy (medium without glucose including Na(+)-azide) increased influx in all resistant sublines. In EHR2/0.4 and EHR2/0.8 (passage no. 72) the influx, however, was still significantly decreased after depletion of energy. In these cells further addition of verapamil increased influx to the level of EHR2. These data were consistent with the hypothesis that P-glycoprotein effluxes drug directly from the plasma membrane.
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PMID:Influx of daunorubicin in multidrug resistant Ehrlich ascites tumour cells: correlation to expression of P-glycoprotein and efflux. Influence of verapamil. 764 48

WEHI-3B/NOVO is a cloned murine leukemia cell line selected for resistance to novobiocin that is cross-resistant to the cytotoxic action of etoposide (VP-16) and to a lesser extent to a variety of other topoisomerase II (topo II)-reactive drugs. We have reported previously (Cancer Res. 52: 2782-2790, 1992) that WEHI-3B/NOVO cells exhibit a pronounced decrease in VP-16 induced DNA-topo II cross-link formation compared to the parental WEHI-3B/S cell line in intact cells, in the absence of a significant difference in the P4 unknotting activity of topo II assayed in nuclear extracts. Because the pattern of cross-resistance was suggestive of a topo II-mediated mechanism, we have ascertained whether a change in topo II can account for the multidrug-resistant phenotype of WEHI-3B/NOVO cells. No differences existed between WEHI-3B/S and WEHI-3B/NOVO cells in topo II mRNA and protein levels, as well as in the amount of topo II associated with the nuclear matrix. Neither sensitive nor resistant cells expressed detectable levels of the MDR1 gene; however, VP-16 accumulation in WEHI-3B/NOVO cells was 3-4-fold less than that present in WEHI-3B/S cells, whereas doxorubicin accumulation was the same in both cell lines. Over the first 60 s, no difference existed in the rate of uptake of VP-16 between parental and resistant cells; however, beyond the first 60 s of incubation, [3H]VP-16 accumulated to a greater extent in parental sensitive cells. Thus, an increased rate of efflux of VP-16 was responsible for the lower steady-state concentration of the drug in resistant cells. The efflux Km for VP-16 in WEHI-3B/NOVO cells was 254.7 microM and the Vmax was 10.4 pmol/s/10(7) cells. In the presence of the inhibitors of energy metabolism, sodium azide and deoxyglucose, the efflux of VP-16 was markedly inhibited; readdition of glucose restored the original efflux rate. Northern blot analyses using the human 10.1 probe for the 3'-terminal region of the multidrug-resistance protein (MRP) cDNA revealed a mRNA species of approximately 6 kb in WEHI-3B/NOVO cells but not in WEHI-3B/S cells. Overexpression was associated with amplification of the cognate gene. To ascertain whether the overexpressed gene in WEHI-3B/NOVO cells was the murine MRP or a different member of the same superfamily of ATP-binding ABC cassette transporters, a 341-bp MRP cDNA probe was generated from a murine genomic library.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Increased rate of adenosine triphosphate-dependent etoposide (VP-16) efflux in a murine leukemia cell line overexpressing the multidrug resistance-associated protein (MRP) gene. 767 Dec 47

Leishmania tarentolae cells selected for resistance to the oxyanions pentavalent or trivalent antimonials or to trivalent arsenicals exhibited cross-resistance to the other oxyanions. The basis for resistance in these mutants was studied by transport experiments using radioactive arsenite. All mutants exhibiting high level resistance to arsenite showed a marked decrease in the steady-state accumulation of arsenite. Decreased accumulation was also observed in antimonials-resistant mutants cross-resistant to various concentrations of arsenite. Cells depleted of endogenous energy reserves with metabolic inhibitors were loaded with radioactive arsenite; following addition of glucose, rapid efflux of arsenite was observed from arsenite mutant cells. Mutants resistant to high levels of arsenicals exhibited amplification of the P-glycoprotein related gene ltpgpA or of a linear amplicon of unknown function. However, the efflux-mediated arsenite resistance did not correlate with the amplification of the ltpgpA gene or with the presence of the linear amplicon. The calcium channel blocker verapamil and arsenite act in synergy in cells exhibiting the efflux system. Overall the oxyanion efflux system in Leishmania shares several properties with other resistance efflux systems mediated by transporters.
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PMID:High level arsenite resistance in Leishmania tarentolae is mediated by an active extrusion system. 783 83

Primary monolayer cultures of winter flounder renal proximal tubule epithelium mounted in Ussing chambers were used to characterize transepithelial transport of daunomycin (Dau). Control tissues performed active net secretion of Dau (0.064 +/- 0.027 nmol.cm-2.h-1). Mild heat shock (5 degrees C elevation for 6-8 h followed by return to normal temperature) almost doubled Dau secretion (0.114 +/- 0.026 nmol.cm-2.h-1). This response was inhibited approximately 40% by addition of the protein synthesis inhibitor, cycloheximide. Dau secretion was inhibited by verapamil, vinblastine, cyclosporin A, and to a lesser degree by the organic cation, tetraethylammonium. In addition, tetraethylammonium secretion was inhibited by vinblastine. Dau secretion was not inhibited by the organic anion, p-aminohippurate, and p-aminohippurate secretion was not inhibited by vinblastine. The transepithelial reabsorptive flux of Dau and the electrical characteristics of the tissues, including rheogenic glucose transport, were unaffected by any of the above treatments. Reaction of tissues with a monoclonal antibody to P-glycoprotein (C219) revealed the presence of this transporter on only apical microvilli. The data indicate that flounder possess an active mechanism for the renal excretion of Dau that is stimulated by mild heat shock. This mechanism is distinct from organic anion, but not organic cation, transport and has characteristics consistent with transport by an apical P-glycoprotein.
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PMID:Heat-shock-stimulated transepithelial daunomycin secretion by flounder renal proximal tubule primary cultures. 784 Feb 40

In order to address the association of enhanced drug efflux with the multidrug-resistant (MDR) phenotype, we have studied the cellular pharmacokinetics of anthracyclines in the P-glycoprotein (Pgp)-positive MDR cell lines H69/LX4 (human small cell lung cancer) and EMT6/AR1.0 (mouse mammary tumour). Both doxorubicin (DOX) and daunorubicin (DNR) were accumulated to a lesser extent and effluxed at a higher rate by MDR cells than by their drug-sensitive counterparts. In contrast, the 9-alkyl substituted compound, aclacinomycin A (ACL), was accumulated and effluxed from parent and MDR cells at an identical rate. In experiments designed to examine energy-dependent efflux, DOX and DNR were shown to be efficiently effluxed against the concentration gradient in the presence of glucose. However, in the same experiments the analogues ACL and Ro 31-3294 (9-alkyl and morpholinyl substituted), which have previously been shown to retain activity against MDR cell lines, were accumulated and effluxed at identical rates in parent and MDR EMT6 cells. Hence, 9-alkyl and morpholinyl substituted compounds appear to behave less favourably as substrates for energy-driven drug efflux by Pgp-positive MDR cells than do DOX or DNR. Resistance modifiers verapamil and cyclosporin A appeared to abolish energy-dependent efflux for DOX and DNR in both the EMT6 and H69 MDR lines whereas they had no effect on the cellular efflux of ACL. The altered cellular pharmacology in MDR cell lines may provide a rational basis for the use of modified anthracycline analogues (e.g. 9-alkyl and morpholinyl (substituted) and resistance of modifying agent in the treatment of tumours expressing a Pgp-mediated phenotype.
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PMID:The efflux of anthracyclines in multidrug-resistant cell lines. 790 89

The decrease of the intracellular concentration of drug in resistant cells as compared to sensitive cells is, in most of cases, correlated with the presence, in the membrane of resistant cells, of a 170-kDa P-glycoprotein (P-gp) responsible for an active efflux of the drug. The fluorescence emission spectra from anthracycline-treated cells suspended in buffer have been used to follow the P-gp-associated efflux of these drugs in the absence or presence of verapamil. In the present study, 4'-o-tetrahydro-pyranyladriamycin (THP-adriamycin) was used. Two different methods were used to determine the kinetics of active efflux of THP-adriamycin: (1) at the steady-state, (2) directly, after the addition of glucose to cells first incubated with THP-adriamycin in the presence of N3- and in the absence of glucose. Kinetic analysis indicates: (1) a saturation of the active efflux when the cytosolic free drug concentration increased (the Michaelis constant Km = 0.5 +/- 0.3 microM) and (2) that the inhibitory effect of verapamil on P-gp-associated efflux of THP-adriamycin in living cells is non-competitive.
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PMID:Non-competitive inhibition of P-glycoprotein-associated efflux of THP-adriamycin by verapamil in living K562 leukemia cells. 790 85

The MDR1 gene product, P-glycoprotein, has been localized to the apical surface of the renal proximal tubule, but its functional role in the kidney is unknown. We studied renal luminal and antiluminal uptake of three known substrates of P-glycoprotein: vinblastine, vincristine and colchicine, by using the single pass multiple indicator dilution method under control conditions and in the presence of increasing concentrations of cyclosporin A, a potent inhibitor of P-glycoprotein. A bolus of [125I]albumin (plasma reference), L-[14C]glucose (extracellular and glomerular reference) and tracer 3H-substrate was injected into the left renal artery of anesthetized dogs and timed serial samples were collected from the left renal vein and left and right ureters. In a single pass, approximately 38, 13 and 8% of [3H]vinblastine, [3H] vincristine and [3H]colchicine, respectively, was extracted from the postglomerular circulation. Drug binding to plasma proteins was determined to be 81% for [3H]vinblastine, 71% for [3H] vincristine and 23% for [3H]colchicine. Despite the high degree of drug protein binding, the urine recoveries of [3H]vinblastine, [3H]vincristine and [3H]colchicine relative to L-[14C]glucose were 0.75 +/- 0.06, 0.69 +/- 0.06 and 0.94 +/- 0.02, confirming net secretion of each of these substrates. Infusion of cyclosporin A (0.1-5 microM) significantly decreased the urine recovery of [3H] vinblastine and [3H]vincristine relative to L-[14C]glucose in a dose-dependent manner. The renal excretion of [3H]colchicine was not affected by cyclosporin A at the concentrations tested (1-2 microM). The evidence suggests that net secretion of [3H]vinblastine and [3H]vincristine occurs across the luminal membrane of the renal cell.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Renal secretion of vinblastine, vincristine and colchicine in vivo. 790 30

The effects of cyclosporin A (CSA) and cellular energy depletion on daunorubicin (DAU) transport kinetics were investigated in a human erythroid leukaemia cell line K562 c1.6 selected for resistance to daunorubicin. K562 c1.6/DAU resistant cells displayed high levels of P-glycoprotein and a high level of multidrug resistance against several antitumour drugs. The resistance factors of K562 c1.6/DAU cells to DAU, doxorubicin, vinblastine and etoposide were 106, 114, 85 and 13 respectively. A 1.6-fold decrease (P < 0.01, n = 8) in DAU accumulation and a 4-fold increase (P < 0.001, n = 8) in DAU efflux were shown in the resistant cells when compared to K562 c1.6 drug-sensitive parental cells. K562 c1.6/DAU cells were also shown to reach a DAU saturation level (SL) 8-fold faster (P < 0.001, n = 8) than the parental cells. Addition of CSA to the resistant cells led to a dose-dependent increase in cellular DAU retention, while no such effect was observed in the sensitive cells by the introduction of CSA. Resistance to the antitumour drugs could be reduced to various extents by CSA. The patterns of changes and modulations of DAU transport kinetics, as well as chemosensitivity in K562 c1.6/DAU cells were found to be similar to a vinblastine-resistant leukaemia cell line CEM/VLB100. However, K562 c1.6/DAU cells were more resistant to DAU, doxorubicin and etoposide than the CEM/VLB100 cells. An increase in DAU accumulation, intracellular SL and the time to reach 90% saturation level (SL90), and a decrease in DAU efflux in the resistant but not the sensitive cells were found in response to ATP depletion by sodium azide. These effects could be completely reversed by addition of glucose. Our results suggest that the presence of an energy-dependent effluxing mechanism responsible for the decreased drug accumulation and enhanced drug efflux may make a major contribution to the mechanism of resistance in K562 c1.6/DAU resistant cells.
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PMID:Characterization and modulation of drug transport kinetics in K562 c1.6 daunorubicin-resistant cell line. 804 36

We studied the transport mechanism of pirarubicin (THP) in HL60 and its THP-resistant (HL60/THP) cells, which showed no expression of mdr1 mRNA on Northern blot analysis. Under physiological conditions, the uptake of THP by both types of cell was time- and temperature-dependent. The amount of drug transport in the resistant cells was significantly less than that in the parent cells within 3 min of incubation. THP uptake was significantly higher in the presence than in the absence of 4 mM 2,4-dinitrophenol (DNP) in glucose-free Hanks' balanced salt solution in both HL60 and HL60/THP cells and the increases were approximately equal. In the presence of DNP, the uptake of THP by both types of cell was concentration-dependent, and there were no significant differences in the apparent kinetic constants (Michaelis constant (Km), maximum velocity (Vmax) and Vmax/Km) for THP uptake between HL60 and HL60/THP cells. Additionally, THP transport was competitively inhibited by its analogue doxorubicin. The efflux of THP from HL60/THP cells was significantly greater than that from HL60 cells, and the release from both types of cell was completely inhibited by decreasing the incubation temperature to 0 degrees C and by treatment with DNP in glucose-free medium. In contrast, the P-glycoprotein inhibitors verapamil and cyclosporin A did not inhibit THP efflux. However, genistein, which is a specific inhibitor of multidrug resistance-associated protein (MRP), increased the THP remaining in the resistant cells, and the value was approximately equal to that of the control group in the sensitive cells. These results suggest that THP is taken up into HL60 and HL60/THP cells via a common carrier by facilitated diffusion, and then pumped out in an energy-dependent manner. Furthermore, the accelerated efflux of THP by a specific mechanism, probably involving MRP, other than the expression of P-glycoprotein, resulted in decreased drug accumulation in the resistant cells, and was responsible, at least in part, for the development of resistance in HL60/THP cells.
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PMID:Transport mechanism of anthracycline derivatives in human leukemia cell lines: uptake and efflux of pirarubicin in HL60 and pirarubicin-resistant HL60 cells. 854 74

The mechanisms of intestinal permeation of several beta-lactam antibiotics and anionic compounds were studied in vitro using excised rat intestinal segments. Permeation of cefazolin through jejunum, ileum and colon was highly secretory-oriented; serosal-to-mucosal permeation rates were two- to three-fold greater than mucosal-to-serosal permeation rates. Serosal-to-mucosal permeation decreased in the absence of D-glucose, and mucosal-to-serosal permeation increased, indicating that the preferential secretory transport of cefazolin is energy dependent. Ampicillin permeation across rat jejunum also favored secretion, whereas the permeation of cefaclor and cephradine favored absorption. Because cefazolin is anionic, several structurally unrelated anionic compounds were also tested. Of these only phenol red exhibited preferential serosal-to-mucosal permeation. The intestinal permeation of phenol red was concentration dependent and glucose dependent. Verapamil and a monoclonal antibody to P-glycoprotein only modestly and inconsistently affected the permeation of cefazolin, ampicillin and phenol red. Probenecid and guanidine were much more effective inhibitors of cefazolin and phenol red secretion. Mutual interactions between cefazolin and phenol red were also observed. These results show that the rat intestine has the capability for net secretory transport of some hydrophilic, anionic compounds. Transport of these compounds has some of the characteristics of organic anion and organic cation transport systems.
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PMID:The secretory intestinal transport of some beta-lactam antibiotics and anionic compounds: a mechanism contributing to poor oral absorption. 876 53


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