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)

HL-60-R, a multi-drug-resistant (MDR) subclone of the human leukemia cell line HL-60, was selected in continuous culture in doxorubicin (DOX) in the absence of mutagenic agents. When compared to the parent line HL-60, HL-60-R showed greater relative resistance to vinblastine than to etoposide, or to the selecting agent DOX. Co-exposure to verapamil, a known modulator of MDR, partially increased its sensitivity to DOX and vinblastine. The HL-60-R cell line stained positively with the P-glycoprotein-specific monoclonal antibody (MAb), C219, whereas the HL-60 parent was negative. Southern analysis showed 32-fold amplification of the mdrI gene in HL-60-R, whereas slot-blot analysis demonstrated 70-fold over-expression of the specific mdrI message in HL-60-R compared to HL-60. Northern blot analysis revealed the presence of 2 species of messenger RNA of sizes 5.1 kb and 4.5 kb. No transcripts were detectable in the parent. Flow cytometric analysis showed significantly reduced cellular retention of DOX as well as rapid efflux from the drug-resistant cell line. HL-60-R proved to be nearly 4 times more resistant to hydrogen peroxide than its parent, and 1,000 times more resistant to inhibition of cellular glutathione synthesis by D,L-buthionine sulfoximine (BSO). Verapamil modulated DOX resistance in HL-60-R incompletely but, in the presence of glutathione depletion, nearly completely reversed DOX resistance. Elevated levels of glutathione and glutathione-peroxidase activity were demonstrated, thereby implicating enhanced activity of the glutathione/glutathione-peroxidase cycle as an additional basis for its resistance to DOX. These findings suggest that an enhanced capacity for detoxifying oxyradicals may contribute to anthracycline resistance in acute leukemia.
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PMID:P-glycoprotein and alterations in the glutathione/glutathione-peroxidase cycle underlie doxorubicin resistance in HL-60-R, a subclone of the HL-60 human leukemia cell line. 809 91

We have studied the pharmacological parameters of doxorubicin resistance in three lines of murine cells selected by long-term culture in the presence of this drug or vincristine. A line originating from rat hepatoma spontaneously presented an intrinsic doxorubicin resistance as compared to the other lines, originating from a rat glioblastoma and from simian-virus-40-transformed mouse hepatocytes. This intrinsic resistance, as well as the doxorubicin resistance exhibited by the vincristine-selected glioblastoma variant, could be entirely attribute to decreased drug accumulation due to drug efflux. In contrast, the doxorubicin-selected variants of the three lines exhibited an intracellular tolerance to this drug. Despite a reduction in drug accumulation when exposed to the same amount of doxorubicin, they accumulated 6-12 times more doxorubicin than wild lines when submitted to equitoxic exposures. Verapamil could restore in these lines the doxorubicin accumulation observed in sensitive lines but could not restore doxorubicin cytotoxicity. Quantitative evaluation of P-glycoprotein expression by Western blotting with the C219 antibody indicated that the wild hepatoma line overexpressed P-glycoprotein by a factor of 5 in comparison with the other wild lines, and that the vincristine-selected glioblastoma variant overexpressed this protein almost as much as the doxorubicin-selected variants. These observations favor the existence of P-glycoprotein-independent mechanisms of doxorubicin resistance, which are added to the classical multidrug-resistant phenotype in doxorubicin-selected highly resistant variant cell lines.
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PMID:Pharmacological and molecular characterization of intrinsic and acquired doxorubicin resistance in murine tumor cell lines. 810 Aug 23

The ability of P-glycoprotein (P-gp) to transport naturally occurring and synthetic opiate analgesics was investigated. Multidrug-resistant Chinese hamster ovary cells (B30) were found to accumulate significantly lower amounts of morphine than their drug-sensitive counterparts (B1). This decreased accumulation was reversed upon depletion of cellular stores of ATP. In addition, morphine bound to plasma membranes from B30 cells in a specific, saturable fashion. Verapamil and vinblastine, compounds transported by P-gp, were able to increase accumulation and displace the binding of morphine in B30 cells. In turn, the synthetic opiates meperidine, pentazocine, and methadone were able to increase the accumulation of vinblastine in resistant cells. These compounds were also able to displace the specific binding of vinblastine and the photoaffinity labeling of P-gp in plasma membranes by the radioiodinated anthracycline, iodomycin. The implications of these findings in relation to the distribution, tolerance, and gastrointestinal side effects of opiates are discussed.
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PMID:Synthetic and natural opiates interact with P-glycoprotein in multidrug-resistant cells. 810 46

We have compared the pharmacological and molecular characteristics of 2 cell lines derived from the C6 rat glioblastoma, and selected for resistance either to doxorubicin (C6 0.5 line) or to vincristine (C6 IV line). Each line displays a preferential 400-fold resistance towards the drug used for selection, the C6 IV line being especially weakly resistant to doxorubicin (13-fold). Verapamil completely restored doxorubicin sensitivity in the C6 IV line as well as vincristine resistance in the C6 0.5 line, but could not completely reverse doxorubicin resistance in the C6 0.5 line or vincristine resistance in the C6 IV line. This suggests that specific mechanisms of resistance against each drug were added to a common P-glycoprotein-mediated multidrug-resistance mechanism. Doxorubicin efflux was total within 2 hr in the C6 IV line, whereas it remained 8 to 10% of drug in the C6 0.5 line 4 hr after drug removal, despite a more rapid efflux of the drug in the first 30 min. This 2-compartment behavior could be related to a special sub-cellular distribution of doxorubicin in C6 0.5 cells. Northern and Western blot analysis of the mdrI gene and of the P-glycoprotein expressed by the 2 resistant cell lines made it possible to quantify their degree of over-expression; when compared with the C6 wild strain, the C6 0.5 line over-expressed both the mdrI gene and the P-glycoprotein to a slightly higher level than the C6 IV line. Northern and Western blot analysis also suggested that C6 0.5 cell preferentially over-expressed the mdrIa gene, whereas the C6 IV cells preferentially over-expressed the mdrIb gene. This differential over-expression was confirmed after polymerase-chain-reaction amplification of the cDNA sequences transcribed from total RNA extracted from the 2 lines. It can be concluded therefore that the mdrIa gene product is more efficient than the mdrIb gene product in extruding anti-cancer drugs from the cells; and that the mdrIb gene product might preferentially extrude vincristine rather than doxorubicin.
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PMID:Differential over-expression of mdr1 genes in multidrug-resistant rat glioblastoma cell lines selected with doxorubicin or vincristine. 810 27

This work was designed to discern the frequency of expression of classical multidrug resistance (MDR) in acute myeloid leukemia (AML) at the time of diagnosis, using Western blotting for P-glycoprotein (Pgp) and functional assays for an MDR phenotype (enhancement of daunorubicin [DNR] accumulation/retention and cytotoxicity by the known MDR modulators verapamil, cyclosporin A, and progesterone). Blast cells were studied from 49 newly diagnosed AML patients who were subsequently treated with the "3 and 7" combination of cytosine arabinoside (ara-C) and DNR. DNR accumulation (1 microgram/mL, 3 hours) and retention (16 hours) were determined by flow cytometry. Cyclosporin A (CsA, 5 mumol/L) or verapamil (6.6 mumol/L) each caused significant enhancement of DNR accumulation and retention in these blast cell samples (P < .001, Wilcoxon's test). Verapamil or CsA caused greater than 20% enhancement of DNR accumulation or retention in over 25% or 50% of these patients, respectively; however, there was no correlation with the presence or degree of enhancement and response to treatment. Progesterone (10 mumol/L) caused no significant enhancement of DNR accumulation or retention. The effects of the MDR modulators on the cytotoxicity of DNR was also determined in blast cells from 40 of the patients, using a flow cytometric assay. CsA alone was cytotoxic (caused an approximate 20% decrease in cell survival compared with control, P < .001); CsA or verapamil caused enhancement of 1 mumol/L DNR cytotoxicity (P < .001). Greater than 40% enhancement of cell kill by CsA or verapamil was observed in over 75% of patients studied. There was no difference in the degree of enhancement of cytotoxicity between patients clinically sensitive or resistant to treatment. Progesterone caused no enhancement in DNR cytotoxicity. In contrast to the functional assays, highly sensitive immunoblots using the C219 antibody to Pgp showed evidence of low level expression of Pgp in blast cells from only 3 of these patients: 1 was chemotherapy resistant, 2 were sensitive. Thus, although the functional assays suggest a high frequency of expression of a classic MDR phenotype in AML patients at the time of diagnosis, with enhancement by CsA obtained at a clinically relevant concentration (5 mumol/L), the frequency of Pgp expression detectable by C219 Western blots was low in these patients. This could be interpreted either that the method used was not sufficiently sensitive to detect Pgp in all of the blast cell specimens that actually overexpressed mdr1, or that the accumulation/efflux-based MDR phenotype observed is not always mediated by Pgp in these previously untreated patients.
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PMID:Enhancement of daunorubicin accumulation, retention, and cytotoxicity by verapamil or cyclosporin A in blast cells from patients with previously untreated acute myeloid leukemia. 810 61

Verapamil, usually given as a racemic mixture, decreases in vivo and in vitro digoxin renal tubular secretion, which is suggested to be mediated by P-glycoprotein, an ATP-dependent multidrug efflux pump. Importantly, the two enantiomers of verapamil have been reported to similarly inhibit P-glycoprotein-mediated transport of chemotherapeutic agents. In this study, we examined effects of enantiomers of verapamil on digoxin transport across an LLC-PK1 cell monolayer, a model of proximal renal tubular cells. The results indicate that verapamil inhibition of digoxin transport is non-stereospecific. Furthermore, the verapamil-digoxin interaction is not competitive. The two drugs may not share a common initial step in the P-glycoprotein-mediated transport.
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PMID:The mechanism of the verapamil-digoxin interaction in renal tubular cells (LLC-PK1). 824 76

The doxorubicin-selected multidrug resistant (MDR) human large cell lung cancer line COR-L23/R, lacks P-glycoprotein but shows a drug accumulation deficit. It does however overexpress a 190k membrane protein which shares an epitope with, but is otherwise distinct from, P-glycoprotein. The resistant cells show only a small sensitisation to vincristine and daunorubicin on treatment with cyclosporin A and its more potent analogue, PSC-833 despite an increase in drug accumulation. Verapamil, another effective resistance modifier in P-glycoprotein MDR cells, is slightly more effective. Fluorescent daunorubicin distributes in the cytoplasm and nucleus of sensitive parent COR-L23 cells but is confined to cytoplasmic perinuclear vesicles in resistant cells. Addition of cyclosporin A or PSC-833 slightly increases cytoplasmic fluorescence whereas verapamil also increases nuclear fluorescence. Resistance in this non-P-glycoprotein MDR line, COR-L23/R where these resistance modifiers have little effect may be associated with expression of the 190k protein.
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PMID:Chemosensitisation and drug accumulation effects of cyclosporin A, PSC-833 and verapamil in human MDR large cell lung cancer cells expressing a 190k membrane protein distinct from P-glycoprotein. 839 42

Verapamil reverses multidrug resistance acquired by cancer cells during treatment with chemotherapeutic agents such as doxorubicin by inhibiting the function of P-glycoprotein. Verapamil has also been suggested to potentiate the cardiotoxicity of doxorubicin. We have recently demonstrated that selective inhibition of cardiac muscle gene expression is among the earliest events in doxorubicin cardiotoxicity. To explore the influence of verapamil on doxorubicin cardiotoxicity, we evaluated [14C]-doxorubicin accumulation, cardiac muscle gene expression by Northern blot analysis, and ultrastructural changes in cultured cardiomyocytes in the presence and absence of verapamil. Treatment with a combination of doxorubicin and verapamil for 24 h did not augment doxorubicin accumulation in cardiomyocytes, although substantial augmentation of doxorubicin accumulation by verapamil in cardiac fibroblasts was observed. Further, treatment with verapamil for 24 h did not augment the decrease in expression of muscle genes induced by doxorubicin (myosin light chain 2 slow, troponin I, M isoform creatine kinase). However, we found that verapamil reduced alpha-actin gene expression in a direct, doxorubicin-independent manner. Furthermore, the effect of doxorubicin plus verapamil on alpha-actin gene expression was additive over a wide range of doxorubicin and verapamil concentrations, resulting in a selective augmentation of doxorubicin-induced inhibition of gene expression for this single muscle protein gene. This was reflected in a substantial increase in cardiac myocyte damage when treatment with verapamil and doxorubicin was compared to treatment with doxorubicin alone by thin section electron microscopy. This suggests a possible mechanism by which verapamil may potentiate doxorubicin cardiotoxicity.
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PMID:Effect of verapamil on doxorubicin cardiotoxicity: altered muscle gene expression in cultured neonatal rat cardiomyocytes. 840 43

Small cell lung carcinomas (SCLC) are characterised by chemosensitivity to diverse antitumoral compounds. However, responses are transitory and relapses are commonly observed. We examined the ability of verapamil, a reverser of P-glycoprotein (Pgp)-related resistance, to improve the efficacy of CyCAV combined chemotherapy (Cy, cyclophosphamide (CPA); C, cisplatin (CDDP); A, doxorubicin (ADM);V, etoposide (VP16)), as currently administered to SCLC patients at Institut Gustave-Roussy, France, and adapted to the treatment of nude mice implanted with these tumours. Although Pgp encoded by the MDR1 (multidrug resistance) gene is not the only mechanism for multidrug resistance (MDR), and not all drugs included in this regimen are recognised by Pgp, we anticipated a therapeutic benefit. Four different SCLC lines, expressing the MDR1 gene and recently grafted into nude mice, were used. SCLC-75, SCLC-6 and SCLC-41 originated from untreated patients, and SCLC-74T was derived from a patient treated with a combination of ADM, CPA and VP16. SCLC-41% and SCLC-6T tumours were used after having undergone, respectively, five and nine cycles of in vivo passage and CyCAV treatment of the tumour-bearing nude mice, to reinforce their chemoresistance. The efficacy of the CyCAV regimen, associated with or without verapamil (given 24 h before CyCAV on days 1-5), was tested on the growth of these SCLC. Verapamil (25 mg/kg) improved the antitumour effect of CyCAV in mice bearing SCLC-6T, SCLC-41T and SCLC-75 tumours, although toxicity was observed. Verapamil modestly delayed the plasma clearance of ADM. Two daily injections of 10 mg/kg of verapamil, administered at a 3 h interval, proved to be effective, whereas the same total dose administered as a bolus was not. These results indicate that the association of some reversers of MDR, including drugs possibly interacting with Pgp, might potentiate SCLC combined chemotherapy.
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PMID:Adding a reverser (verapamil) to combined chemotherapy overrides resistance in small cell lung cancer xenografts. 854 Nov 14

Vesicular neurotransmitter transporters function in synaptic vesicles and other subcellular organelles and they were thought to be involved only in neurotransmitter storage. Several findings have led us to test novel aspects of their function. Cells expressing a c-DNA coding for one of the rat monoamine transporters (VMAT1) become resistant to the neurotoxin N-methyl-4-phenylpyridinium (MPP+) [Liu et al. (1992) Cell, 70, 539-551]. The basis of the resistance is the VMAT1-mediated transport and sequestration of the toxin into subcellular compartments. In addition, the deduced sequence of VMAT1 predicts a protein that shows a distinct homology to a class of bacterial drug resistance transporters (TEXANs) that share some substrates with mammalian multidrug resistance transporters (MDR) such as the P-glycoprotein. These findings induced us to test whether compounds that are typically transported by MDR interact also with vesicular transporters. The use of [3H]reserpine binding to determine drug interactions with VMAT allowed assessment of the ability of various drugs to bind to the substrate site of the transporter. Cytotoxic compounds such as ethidium, isometamidium, tetraphenylphosphonium, rhodamine, tacrine and doxorubicin, interact specifically with vesicular monoamine transporters. Verapamil, a calcium channel blocker, is also a competitive inhibitor of transport. In the case of rhodamine, fluorescence measurements in digitonin-permeabilized cells demonstrated ATP-dependent VMAT-mediated transport. The results imply that even though the bacterial and vesicular transporters are structurally different from the P-glycoprotein, they share a similar substrate range. These findings suggest a novel possible way of protection from the effects of toxic compounds by removal to subcellular compartments.
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PMID:The pharmacological profile of the vesicular monoamine transporter resembles that of multidrug transporters. 854 51


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