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)

In the present report, we show evidence that the membrane from the protozoan parasite Leishmania tropica (LRC-L39), in vitro resistant to 1 mM of methotrexate (MTX), has a significative increased ATPase activity with respect to wild-type line. This ATPase activity is vanadate sensitive, a characteristic of the P-type ATPases included in the ATP-binding casette (ABC) superfamily of transporters, such as P-glycoprotein involved in the multidrug resistance in mammalian cells. Also, this ATPase activity is not modified by MTX, ammonium molybdate and other detergents such as Triton X-100, Brij 58 and lysophosphatidylcholine. However, unlike the P-glycoprotein, we have observed that the ATPase activity is not stimulated by the drugs verapamil and puromycin. This significative ATPase activity could be related to the overexpressed putative P-glycoprotein, with unknown function in these MTX-resistant parasites.
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PMID:Increased P-type ATPase activity in Leishmania tropica resistant to methotrexate. 790 68

Daunomycin, an anti-neoplastic agent, is known to be sequestered by acidic organelles in normal and multidrug-resistant cells [Willingham, M.C., Cornwell, M.M., Cardarelli, C.O., Gottesman, M.M., & Pastan, I. (1986) Cancer Res. 46, 5941-5946]. We studied the mechanism of accumulation of daunomycin into acidic organelles using chromaffin granule vesicles and proteoliposomes reconstituted with purified F-type H(+)-ATPase as model systems. Radiolabeled daunomycin was taken up by chromaffin vesicles upon addition of ATP. Its ATP-dependent uptake was stimulated about 1.4- to 1.8-fold by valinomycin plus K+, but was inhibited by ammonium chloride (10 mM) and nigericin plus K+. Quinidine (5 microM), verapamil (5 microM), or vanadate (0.5 mM), inhibitors of P-glycoprotein, had no effect on its uptake. Daunomycin was also taken up by liposomes reconstituted with F-type H(+)-ATPase. Furthermore, doxorubicin and vinblastine were taken up by these vesicles, whereas colchicine and rhodamine 123 were not. The accumulations of daunomycin and doxorubicin in acidic organelles of cultured cells were decreased by inhibiting vacuolar ATPase by addition of bafilomycin A1 or concanamycin A, or by increasing the internal pH by addition of nigericin. Melittin and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide dissipated the delta pH and inhibited accumulation of daunomycin in the membrane vesicles and acidic organelles in cultured cells. These results indicate that the delta pH established by vacuolar-type ATPase drives the uptake of daunomycin, doxorubicin or vinblastine into acidic organelles, and that no specific transporters are involved in their uptakes.
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PMID:ATP-dependent uptake of anti-neoplastic agents by acidic organelles. 820 70

The mechanism by which chlorpromazine (2-chloro-10-(3-dimethylaminopropyl)-phenothiazine) reverses P-glycoprotein (P-gp2) mediated multidrug resistance was investigated using membrane vesicles prepared from human CCRF-CEM leukaemia cells. Chlorpromazine was transported in an ATP-dependent manner into membrane vesicles prepared from vinblastine resistant (VBL1000) cells but not from drug-sensitive cells. The chlorpromazine uptake was sensitive to osmotic pressure, indicating true transport into the vesicle lumen. The ATP-dependent chlorpromazine uptake was inhibited about 30% by the addition of ammonium chloride, indicating that a pH or electrical gradient could not account for the majority of ATP-dependent chlorpromazine uptake. The results of this study show that chlorpromazine is actively transported my P-glycoprotein and that chemosensitization by phenothiazines may occur by competition of these agents for active transport of anticancer agents by P-glycoprotein.
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PMID:Chlorpromazine transport in membrane vesicles from multidrug resistant CCRF-CEM cells. 884 36

The basic distinguishing feature of all cells expressing functional P-glycoprotein-multidrug resistance (P-gp-MDR) is a decrease in steady-state accumulation drug levels as compared to drug-sensitive controls. In an attempt to identify mechanism(s) by which MDR can be circumvented, we examined the cellular accumulation, in resistant cells, of 4'-O-tetrahydropyranyl-doxorubicin (pirarubicin) alone and in conjunction with various molecules belonging to three different classes: the crown ethers, the tetraalkylammonium salts, and the polyoxethylene amphiphiles. The present study was performed using a spectrofluorometric method which enabled us to follow the uptake and release of fluorescent molecules by living cells while the cells were being incubated with the drug. Erythroleukemia K562 cell lines were used. Our data show that the compounds of these three completely different classes were able to increase the incorporation of pirarubicin provided they had a minimum degree of lipophilicity. Study of the growth inhibitory activity of these compounds revealed that cross-resistance to the tetraalkyl ammonium salt increased with the lipophilicity and was equal to 58 for tetraoctylammonium salt, the most lipophilic compound of this series. This demonstrates that neither the presence of a positive charge nor an aromatic moiety is required for MDR recognition.
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PMID:The effect of crown ethers, tetraalkylammonium salts, and polyoxyethylene amphiphiles on pirarubicin incorporation in K562 resistant cells. 884 34

Multidrug resistance (MDR) is one of the major obstacles to long term successful cancer chemotherapy. The use of MDR reversal (MDRR) agents is a promising approach to overcome the undesired MDR phenotype. To design more effective MDRR agents that are urgently needed for clinical use, a data set of 609 diverse compounds tested for MDRR activity against P388/ADR-resistant cell lines was submitted to the MULTICASE computer program for structure-activity analysis. Some substructural features related to MDRR activity were identified. For example, the CH2-CH2-N-CH2-CH2 group was found in most of the active compounds, and the activity was further enhanced by the presence of (di)methoxylphenyl groups, whereas the presence of a stable quaternary ammonium salt, a carboxylic, a phenol, or an aniline group was found to be detrimental to activity. Possible explanations for these observations are proposed. Some physicochemical properties, e.g., the partition coefficient (log P) and the graph index (which in some sense measures the "complexity" of a molecule) were also found to be relevant to activity. Their role in MDRR was also rationalized. Based on our quantitative structure-activity relationship study of MDRR agents, some compounds with desired substructural features and activity were identified from the MACCS-II and National Cancer Institute DIS databases and tested experimentally. Our study may also help the rational design of anti-cancer drugs. Based on this study and on observations by other researchers, we postulate that P-glycoprotein-mediated resistance to paclitaxel could probably be eliminated by proper substitution of its benzamido and phenyl groups. Several novel compounds with the paclitaxel skeleton are proposed, which may lead to a new generation of paclitaxel anti-cancer drugs with less MDR potential.
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PMID:Quantitative structure-activity relationship of multidrug resistance reversal agents. 927 56

1. We have used mice with homozygously disrupted mdr1a and mdr1b genes (mdr1a/1b (-/-) mice) to study the role of the mdr1-type P-glycoprotein (P-gp) in the elimination of cationic amphiphilic compounds from the body. These mice lack drug-transporting P-gps, but show no physiological abnormalities under laboratory conditions and have normal bile flow. 2. 3H-labelled cationic drugs were administered intravenously (i.v.) to mice as a single bolus dose and the disposition of the studied cationic drugs was investigated by focusing on drug secretion into bile, intestinal lumen and urine. 3. Hepatobiliary secretion of the investigated cationic drugs was profoundly reduced in mice devoid of the mdr1-type P-gps. In fact, the cumulative biliary output, measured during 1 h, of the small type 1 compounds tri-butylmethyl ammonium (TBuMA) and azidoprocainamide methoiodide (APM), as well as of the more bulky type 2 cationic drug vecuronium, was reduced by at least 70% in the mdrla/lb (-/-) mice compared to wild-type. 4. The intestinal secretion of TBuMA, APM and vecuronium was also profoundly reduced in mdrla/lb (-/-) mice compared to wild-type mice. The absence of the mdrl-type P-gp resulted in virtual elimination of intestinal secretion of TBuMA and APM (>90% reduced as compared to wild-type (P=0.0001 and 0.0022, respectively)). The intestinal secretion of the type 2 cation drug vecuronium was reduced by 58% (P=0.0004) compared to the wild-type mice. 5. Increased renal clearances of both the type 1 compounds TBuMA and APM and also of the type 2 cationic compound vecuronium in the mdrla/lb (-/-) mice were observed. Furthermore, the balance between hepatic, intestinal and renal clearances of small type 1 organic cations clearly shifted towards a predominant role for renal clearance. Increased renal clearance may be explained by (over)expression of additional mechanisms for renal organic cation secretion, alternatively they may also point to an as yet undefined role of P-glycoprotein in kidney physiology and renal secretory function. 6. We conclude that the elimination from the body of a broad spectrum of cationic amphiphilic drugs via liver and intestine, is largely dictated by the activity of mdrl-type P-glycoproteins.
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PMID:Hepatobiliary and intestinal clearance of amphiphilic cationic drugs in mice in which both mdr1a and mdr1b genes have been disrupted. 964 61

We have developed and validated a sensitive and selective method for the determination of the P-glycoprotein modulator GF120918 in murine and human plasma. Chlorpromazine is used as internal standard. Sample pretreatment involves liquid-liquid extraction with tert-butyl methyl ether. Chromatographic separation is achieved by reversed-phase high-performance liquid chromatography using a Symmetry C18 column and detection was accomplished with a fluorescence detector set at excitation and emission wavelengths of 260 and 460 nm, respectively. The mobile phase consists of acetonitrile-50 mM ammonium acetate buffer, pH 4.2 (35:65, v/v). To achieve good separation from endogenous compounds and to improve the peak shape the counter-ion 1-octane sulfonic acid (final concentration 0.005 M) was added to the mobile phase. The lower limit of quantitation was 5.7 ng/ml using 200 microl of human plasma and 23 ng/ml using 50 microl of murine plasma. Within the dynamic range of the calibration curve (5.7-571 ng/ml) the accuracy was close to 100% and within-day and between-day precision were within the generally accepted 15% range. The stability of GF120918 was tested in plasma and blood from mice and humans incubated at 4 degrees C, room temperature, and 37 degrees C for up to 4 h. No losses were observed under these conditions. This method was applied to study the pharmacokinetics of orally administered GF120918 in humans and mice. The sensitivity of the assay was sufficient to determine the concentration in plasma samples obtained up to 24 h after drug administration.
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PMID:Bioanalysis and preliminary pharmacokinetics of the acridonecarboxamide derivative GF120918 in plasma of mice and humans by ion-pairing reversed-phase high-performance liquid chromatography with fluorescence detection. 1149 17

A sensitive high-performance liquid chromatographic (HPLC) method was developed for the determination of paclitaxel in micro-samples of rat plasma in order to study the mechanism of enhanced systemic exposure of paclitaxel co-administered with P-glycoprotein inhibitors. The assay involved solid-phase extraction procedures using 2'-methylpaclitaxel as the internal standard. Chromatographic separations were achieved using a ZORBAX ODS C18 column and mobile phase consisting of acetonitrile, methanol and ammonium acetate buffer (10 mM, pH 5.0) (48.5:16.5:35) pumped at 0.8 ml/min. The effluents were measured for UV absorption at 227 nm, with retention times of 8.5 and 11.0 min for paclitaxel and 2'-methylpaclitaxel, respectively. The chromatographic separation was excellent, with no endogenous interference. The standard curves showed a good linearity (r=0.9994) over the concentration ranges of 10-1,000 ng/ml. At 1,000 ng/ml, the absolute recoveries of paclitaxel and 2'-methylpaclitaxel are 89 and 90%, respectively. The intra- and inter-day variabilities of paclitaxel were both less than 15%. This validated method for the assay of paclitaxel in micro-sample rat plasma made it feasible to study the pharmacokinetics of the drug in a single rat.
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PMID:Quantitation of paclitaxel in micro-sample rat plasma by a sensitive reversed-phase HPLC assay. 1260 67

The objective of the current investigation was to examine the transport characteristics of choline, an endogenous quaternary ammonium compound, into human intestinal Caco-2 cells; the transport of choline has not been characterized in human intestine. The cellular accumulation of choline was independent of an inwardly directed Na(+) gradient and demonstrated temperature dependence and saturability. Using the initial uptake rates, choline accumulation was best characterized by a Michaelis-Menten equation and a diffusion component with a K(m) and V(max) of 110 +/- 3 micro mol/L and 2800 +/- 250 pmol/(mg protein. 10 min), respectively. Choline uptake was significantly inhibited by an excess of choline itself and by hemicholinium-3, a structural analog of choline. However other hydrophilic organic cations, such as tetraethylammonium (TEA) and N-methylnicotinamide (NMN), did not affect choline uptake in Caco-2 cells. Additionally, two typical p-glycoprotein substrates, daunomycin and verapamil, both inhibited choline accumulation. However the opposite was not true: choline did not inhibit DNM accumulation in Caco-2 cells. These results indicate the presence of a carrier-mediated transport system for choline in Caco-2 cells. The substrate specificity of this carrier is unlike that seen in the rat intestinal epithelium, and the human transport protein is distinct from those for TEA and NMN. P-glycoprotein substrates may inhibit choline uptake through specific or nonspecific interactions with the choline transporter.
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PMID:Choline uptake in human intestinal Caco-2 cells is carrier-mediated. 1288 45

We have developed and validated a sensitive and selective method for the quantitative determination of the P-glycoprotein inhibitor zosuquidar (LY335979) in human and murine plasma using only 50 microl sample volumes. Sample pretreatment involved liquid-liquid extraction with tert-butyl methyl ether. Zosuquidar and the internal standard chlorpromazine were separated using a narrow bore column (2.1 mm x 150 mm) packed with 3.5 microm symmetry C(18) material. The mobile phase consisted of 38% (v/v) acetonitrile in 50mM ammonium acetate buffer pH 3.8 containing 0.005 M 1-octyl sulfonic acid and was delivered at 0.2 ml/min. Detection was performed with a fluorescence detector set at an excitation wavelength of 260 nm and an emission wavelength of 460 nm. The calibration curve was prepared in blank human plasma and was linear over the dynamic range (10-1000 ng/ml). The lower limit of quantitation was 20 ng/ml. The validation results showed that the assay was selective and reproducible. Within the range of the calibration curve the accuracy was close to 100% and within-day and between-day precision were within the generally accepted 15% range. This method was applied to study the pharmacokinetics of i.v. administered zosuquidar in mice. The sensitivity of the assay was sufficient to determine the drug concentration in plasma samples obtained up to 24 h after administration.
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PMID:Bioanalysis of zosuquidar trihydrochloride (LY335979) in small volumes of human and murine plasma by ion-pairing reversed-phase high-performance liquid chromatography. 1463 Mar 60


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