Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.6.3.44 (P-glycoprotein)
 13,344 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Topotecan (TPT, 9-dimethylaminomethyl-10-hydroxycamptothecin) is the first topoisomerase I-directed cytotoxic agent to enter clinical trials in the United States in two decades. The effect of P-glycoprotein (Pgp) overexpression on TPT cytotoxicity was examined in CHRC5 (colchicine-resistant) and AuxB1 (parental) Chinese hamster ovary cells. Examination of the IC50 values observed in colony-forming assays revealed that the CHRC5 cells were 15-fold (SD, +/- 3; n = 3) resistant to TPT after a 1-h exposure and 3.2-fold (SD, +/- 1.4; n = 4) resistant in continuous exposure experiments. Band depletion immunoblotting revealed that 4-fold higher concentrations of extracellular TPT were required to induce the formation of topo I-DNA complexes in CHRC5 cells as compared to AuxB1 cells. To assess the role of Pgp in this resistance, drug accumulation and cytotoxicity assays were repeated in the absence and presence of quinidine. Addition of quinidine enhanced TPT accumulation (measured by high-performance liquid chromatography) and diminished the IC50 for TPT to a greater extent in CHRC5 cells than in AuxB1 cells. To examine whether similar effects could be detected in Pgp-expressing human cells, MCF-7/Adriar breast cancer cells and KG1a human acute myelogenous leukemia cells were examined. Quinidine or verapamil enhanced TPT accumulation in both of these cell lines but had no effect in parental MCF-7 cells or a variety of human leukemia cell lines that do not overexpress Pgp. Cytotoxicity measurements performed by counting the number of surviving cells (MCF-7/Adriar) or employing a modified, highly stable tetrazolium dye reduction assay (leukemia cell lines) revealed that quinidine diminished the IC50 for TPT in the Pgp-overexpressing cell lines but not in the control lines. These results suggest that Pgp overexpression diminishes TPT accumulation and TPT cytotoxicity in hamster and human cells. It should be stressed, however, that these effects were substantially smaller than the effects of Pgp overexpression on the accumulation and cytotoxicity of the anthracycline daunorubicin and the epipodophyllotoxin etoposide in the same cell lines.
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PMID:Effect of P-glycoprotein expression on the accumulation and cytotoxicity of topotecan (SK&F 104864), a new camptothecin analogue. 134 48

CPT-11, a semisynthetic derivative of camptothecin, exhibited strong antitumor activity against lymphoma, lung cancer, colorectal cancer, gastric cancer, ovarian cancer, and cervical cancer. CPT-11 is a pro-drug that is converted to an active metabolite, SN-38, in vivo by enzymes such as carboxylesterase. We synthesized a water-soluble and non-pro-drug analog of camptothecin, DX-8951f. It showed both high in vitro potency against a series of 32 malignant cell lines and significant topoisomerase I inhibition. The anti-proliferative activity of DX-8951f, as indicated by the mean GI50 value, was about 6 and 28 times greater than that of SN-38 or SK&F 10486-A (Topotecan), respectively. These three derivatives of camptothecin showed similar patterns of differential response among 32 cell lines, that is, their spectra of in vitro cytotoxicity were almost the same. The antitumor activity of three doses of DX-8951f administered i.v. at 4-day intervals against human gastric adenocarcinoma SC-6 xenografts was greater than that of CPT-11 or SK&F 10486-A. Moreover, it overcame P-glycoprotein-mediated multi-drug resistance. These data suggest that DX-8951f has a high antitumor activity and is a potential therapeutic agent.
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PMID:A new water-soluble camptothecin derivative, DX-8951f, exhibits potent antitumor activity against human tumors in vitro and in vivo. 755 2

We have previously described a mitoxantrone-resistant human breast carcinoma cell line, MCF7/MX, in which resistance was associated with a defect in the energy-dependent accumulation of mitoxantrone in the absence of P-glycoprotein overexpression (M. Nakagawa et al., Cancer Res. 52: 6175-6181, 1992). We now report that this cell line is highly cross-resistant to the camptothecin analogues topotecan (180-fold), 9-aminocamptothecin (120-fold), CPT-11 (56-fold), and SN38 (101-fold), but is only mildly cross-resistant to the parent compound camptothecin (3.2-fold) and 10,11-methylenedioxy-camptothecin (2.9-fold). Topotecan accumulation was decreased in MCF7/MX cells compared to parental MCF7/WT cells, and there was a corresponding reduction in topotecan-mediated stimulation of the enzyme/DNA complex formation in MCF7/MX cells compared to MCF7/WT cells. No overexpression of the multidrug resistance-associated protein was detected compared to parental MCF7/WT cells. Furthermore, both sensitive MCF7/WT and mitoxantrone-resistant MCF7/MX cells contain equal amounts of DNA topoisomerase I protein, and DNA relaxation activities were equal in both cell lines and inhibited to the same extent by topotecan and camptothecin. Thus, these results suggest a novel mechanism of resistance to topoisomerase I inhibitors in cancer cells.
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PMID:Cross-resistance to camptothecin analogues in a mitoxantrone-resistant human breast carcinoma cell line is not due to DNA topoisomerase I alterations. 766 72

Over the past decade, DNA topoisomerase I and II appeared to be the targets of some antitumor agents: CPT-11 and Topotecan derived from Camptothecin which interact with topoisomerase I; Actinomycin D, Adriamycin and Daunorubicin, Elliptinium Acetate, Mitoxantrone, Etoposide and Teniposide, Amsacrine which interact with topoisomerase II. The multiple functions of these enzymes are important as they play a role during replication, transcription, recombination, repair and chromatine organisation. Particularly, they relax torsional constraints which appear when intertwined DNA strands are separated while replication fork or RNA polymerases are moving. To some extent, topoisomerase I and II are structurally and functionally different. Moreover, topoisomerase I is not indispensable for a living cell whereas topoisomerase II is. Drug-topoisomerase interaction which probably leads to antitumoral effect of the compounds studied in this review is not a trivial inhibition of the enzyme but rather a poisoning due to stabilization of cleavable complexes between the enzyme and DNA. These stabilized complexes are likely to induce apoptosis-like programmed cell death, which is characterised by DNA fragmentation. However, it appears that it is the collision of the replication fork with the drug-stabilized cleavable complex that is responsible for the cytotoxicity of the drug: poisoning of topoisomerases by antitumor agents leads to a new concept of "dynamic toxicity". Although they interact with a common target, topoisomerase II poisons have differential effects on macromolecules syntheses, cell cycle and chromosome fragmentation; a few compounds may produce free radicals. Because of these differential effects in addition to quantitative and qualitative variations of stabilized cleavable complexes, in particular DNA sequences on which topoisomerase II is stabilized, these antitumor agents do not resemble each other. Cellular resistance to topoisomerases poisons results of two principal types of alteration: target and/or drug transport modification. Decreased ability to form the cleavable complex in resistant cells may be the consequence of both decreased amount of topoisomerase or altered enzyme. On the other hand, overexpression of membrane P-glycoprotein, which pumps drugs out of the cell by an energy dependent process provokes a decreased accumulation of these drugs. Cross resistances to other drugs are mainly under control of these two different mechanisms of resistance. A complete knowledge of their individual effects and mechanisms of resistance would allow a better clinical use of topoisomerases poisons, especially when administered in combination chemotherapy.
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PMID:[Poisons of DNA topoisomerases I and II]. 808 Oct 34

We have previously described a mitoxantrone-resistant MCF7 cell line that is cross-resistant to topotecan, 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxy-camptothecin (CPT-11), and 9-aminocamptothecin, but not to camptothecin. A novel mechanism that resulted in decreased topotecan accumulation in MCF7/MX cells was proposed (Yang et al. Cancer Res 55: 4004-4009, 1995). We now have developed a topotecan-resistant cancer cell line from wild-type MCF7 cells. MCF7/TPT300 cells were 68.9-fold resistant to topotecan, 68.3-fold to 10-hydroxy-7-ethylcamptothecin (SN-38), and 116-fold to mitoxantrone, but only 4.1-fold to camptothecin. Topotecan efflux was increased in MCF7/TPT300 cells compared with MCF7/WT cells, and this increase was reversed upon ATP depletion by sodium azide, suggesting an energy-dependent drug efflux mechanism. However, MCF7/TPT300 cells did not overexpress P-glycoprotein or the multidrug resistance-associated protein (MRP1). In contrast, overexpression of the breast cancer resistance protein (BCRP/MXR/ABCP) was observed in MCF7/TPT300 cells as well as DNA topoisomerase I down-regulation. Our data suggest that enhanced topotecan efflux contributes partly to topotecan resistance in MCF7/TPT300 cells, possibly mediated by BCRP/MXR/ABCP.
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PMID:BCRP/MXR/ABCP expression in topotecan-resistant human breast carcinoma cells. 1093 May 38

Gefitinib inhibits the ATP-binding site of the tyrosine kinase associated with the epidermal growth factor receptor. It is conceivable that gefitinib may inhibit functions of ATP-binding cassette (ABC) transporters by binding at their ATP-binding sites. The aim of this study is to systematically explore the combined effect of gefitinib and chemotherapeutic agents in gefitinib-insensitive multidrug resistant (MDR) cells that overexpress ABC transporters. MCF7 breast carcinoma cells and CL1 lung adenocarcinoma cells were both insensitive to gefitinib. MDR cancer cells were developed by stepwise escalating concentrations of each chemotherapeutic agent in culture media. Cells that overexpress P-glycoprotein (MCF7/Adr and CL1/Pac), breast cancer-resistant protein (MCF7/TPT and CL1/Tpt), and MDR-associated protein 1 (MCF7/Vp) were used in this study. All resistant mutants were insensitive to gefitinib. Gefitinib (0.3-3 micromol/L) added to culture media had no effect on IC50 values of paclitaxel, topotecan, doxorubicin, or etoposide in wild-type MCF7 or CL1 cells. In contrast, these concentrations of gefitinib caused a dose-dependent reversal of resistance to paclitaxel in CL1/Pac cells, to doxorubicin in MCF7/ADR cells, and to topotecan in CL1/Tpt and MCF7/TPT cells. Gefitinib had no influence on sensitivity to etoposide in MDR-associated protein1 overexpressing MCF7/VP cells. Topotecan efflux was inhibited and accumulation was partially restored in CL1/Tpt and MCF7/TPT cells when cells were incubated simultaneously with gefitinib. Our results suggest that the interaction of gefitinib and chemotherapeutic agents does occur in cells expressing one of these two proteins.
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PMID:Gefitinib reverses chemotherapy resistance in gefitinib-insensitive multidrug resistant cancer cells expressing ATP-binding cassette family protein. 1606 79

Topotecan (TPT) is a semisynthetic water-soluble derivative of camptothecin (CPT) used as second-line therapy in patients with metastatic ovarian carcinoma, small cell lung cancer, and other malignancies. However, both dose-limiting toxicity and tumor resistance hinder the clinical use of TPT. The mechanisms for resistance to TPT are not fully defined, but increased efflux of the drug by multiple drug transporters including P-glycoprotein (PgP), multidrug resistance associated protein 1 (MRP1) and breast cancer resistance protein (BCRP) from tumor cells has been highly implicated. This study aimed to investigate whether overexpression of human MRP4 rendered resistance to TPT by examining the cytotoxicity profiles using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazonium bromide (MTT) assay and cellular accumulation of TPT in HepG2 cells stably overexpressing MRP4. Two kinds of cell lines, HepG2 with insertion of an empty vector plasmid (V/HepG2), HepG2 cells stably expressing MRP4 (MRP4/HepG2), were exposed to TPT for 4 or 48 hr in the absence or presence of various MRP4 inhibitors including DL-buthionine-(S,R)-sulphoximine (BSO), diclofenac, celecoxib, or MK-571. The intracellular accumulation of TPT and paclitaxel (a PgP substrate) by V/HepG2 and MRP4/HepG2 cells was determined by incubation of TPT with the cells and the amounts of the drug in cells were determined by validated HPLC methods. The study demonstrated that MRP4 conferred a 12.03- and 6.86-fold resistance to TPT in the 4- and 48-hr drug-exposure MTT assay, respectively. BSO, MK-571, celecoxib, or diclofenac sensitised MRP4/HepG2 cells to TPT cytotoxicity and partially reversed MRP4-mediated resistance to TPT. In addition, the accumulation of TPT was significantly reduced in MRP4/HepG2 cells compared to V/HepG2 cells, and one-binding site model was found the best fit for the MRP4-mediated efflux of TPT, with an estimated K(m) of 1.66 microM and V(max) of 0.341 ng/min/106 cells. Preincubation of MRP4/HepG2 cells with BSO (200 microM) for 24 hr, celecoxib (50 microM), or MK-571 (100 microM) for 2 hr significantly increased the accumulation of TPT over 10 min in MRP4/HepG2 cells by 28.0%, 37.3% and 32.5% (P < 0.05), respectively. By contrast, there was no significant difference in intracellular accumulation of paclitaxel in V/HepG2 and MRP4/HepG2 cells over 120 min. MRP4 also rendered resistance to adefovir dipivoxil (bis-POM-PMEA) and methotrexate, two reported MRP4 substrates. MRP4 did not exhibit any significant resistance to other model drugs including vinblastine, vincristine, etoposide, carboplatin, cyclosporine and paclitaxel in both long (48 hr) and short (4 hr) drug-exposure MTT assays. These findings indicate that MRP4 confers resistance to TPT and TPT is the substrate for MRP4. Further studies are needed to explore the role of MRP4 in resistance to, toxicity and pharmacokinetics of TPT in cancer patients.
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PMID:Topotecan is a substrate for multidrug resistance associated protein 4. 1645 95

A simple, sensitive, specific and high-resolution reversed-phase liquid chromatographic method utilizing ultraviolet detection has been developed and validated for simultaneous determination of topotecan and four intestinal permeability markers (atenolol, antipyrine, propranolol and furosemide) as suggested by US-FDA. Chromatography was carried out on C-18 column with mobile phase comprising water (pH 3.0) and acetonitrile gradient pumped at a flow rate of 1 ml min(-1). The validation parameters included specificity, accuracy, precision, sensitivity and stability studies. Topotecan, an anti-cancer drug widely used in metastatic carcinoma, is a P-glycoprotein substrate having oral bioavailability of 30% with large inter-patient variability. The present method was successfully applied for demonstrating P-gp mediated transport of topotecan and its inhibition using verapamil in Caco-2 cell monolayer. The method can be used in identification of novel P-gp inhibitors for topotecan and estimating the contribution of P-gp in affecting oral bioavailability of topotecan. The other applications of method include its use in validation of Caco-2 monolayer assay for getting biowaiver based on Biopharmaceutic Classification System and its extrapolation to in situ and/or in vivo studies.
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PMID:Concurrent determination of topotecan and model permeability markers (atenolol, antipyrine, propranolol and furosemide) by reversed phase liquid chromatography: utility in Caco-2 intestinal absorption studies. 1793 93

Topotecan is a substrate of the ATP-binding cassette transporters P-glycoprotein (P-gp/MDR1) and breast cancer resistance protein (BCRP). To define the role of these transporters in topotecan penetration into the ventricular cerebrospinal fluid (vCSF) and brain parenchymal extracellular fluid (ECF) compartments, we performed intracerebral microdialysis on transporter-deficient mice after an intravenous dose of topotecan (4 mg/kg). vCSF penetration of unbound topotecan lactone was measured as the ratio of vCSF-to-plasma area under the concentration-time curves. The mean +/- SD ratios for wild-type, Mdr1a/b(-/-), Bcrp1(-/-), and Mdr1a/b(-/-)Bcrp1(-/-) mice were 3.07 +/- 0.09, 2.57 +/- 0.17, 1.63 +/- 0.12, and 0.86 +/- 0.05, respectively. In contrast, the ECF-to-plasma ratios for wild-type, Bcrp1(-/-), and Mdr1a/b(-/-)Bcrp1(-/-) mice were 0.36 +/- 0.06, 0.42 +/- 0.06, and 0.88 +/- 0.07. Topotecan lactone was below detectable limits in the ECF of Mdr1a/b(-/-) mice. When gefitinib (200 mg/kg) was preadministered to inhibit Bcrp1 and P-gp, the vCSF-to-plasma ratio decreased to 1.29 +/- 0.09 in wild-type mice and increased to 1.13 +/- 0.13 in Mdr1a/b(-/-)Bcrp1(-/-) mice, whereas the ECF-to-plasma ratio increased to 0.74 +/- 0.14 in wild-type and 1.07 +/- 0.03 in Mdr1a/b(-/-)Bcrp1(-/-) mice. Preferential active transport of topotecan lactone over topotecan carboxylate was shown in vivo by vCSF lactone-to-carboxylate area under the curve ratios for wild-type, Mdr1a/b(-/-), Bcrp1(-/-), and Mdr1a/b(-/-)Bcrp1(-/-) mice of 5.69 +/- 0.83, 3.85 +/- 0.64, 3.61 +/- 0.46, and 0.78 +/- 0.19, respectively. Our results suggest that Bcrp1 and P-gp transport topotecan into vCSF and out of brain parenchyma through the blood-brain barrier. These findings may help to improve pharmacologic strategies to treat brain tumors.
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PMID:Compartment-specific roles of ATP-binding cassette transporters define differential topotecan distribution in brain parenchyma and cerebrospinal fluid. 1956 73