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)

Patients with multiple myeloma (MM) commonly become refractory to chemotherapy despite a favorable response to induction treatment. We examined the effectiveness of a previously characterized plasma cell-reactive monoclonal antibody, MM4, in eliminating MM clonogenic colony-forming cells (CCC) with a multidrug-resistant (MDR) phenotype. Experiments were performed using MM cell lines that exhibit 6 (RPMI 8226/DOX6)- and 40 (RPMI 8226/DOX40)-fold resistance to doxorubicin (DOX). Both lines were selected from the chemosensitive MM line RPMI 8226/S and were cross-resistant to mitoxantrone, acronycine, etoposide, and vincristine. Surface marker analysis conducted in this study showed that DOX6 and DOX40 overexpressed the MDR1 gene product p170. Both MDR lines remained reactive to the plasma cell-reactive monoclonal antibodies MM4 and PCA-1 and expressed the relevant cytoplasmic immunoglobulin light chain. Treatment with MM4 and rabbit complement (C') was equally cytotoxic to RPMI 8226/S [80 +/- 5.6% (SD)], DOX6 [74 +/- 8.5], and DOX40 cells [75 +/- 11.3%], based on short-term chromium release studies. Furthermore, MM4 + C' deleted up to 3 logs of CCC colonies from chemosensitive and MDR lines (RPMI 8226/S, 99.87 +/- 0.11%; DOX6, 99.91 +/- 0.08%; DOX40, 99.55 +/- 0.44%). By comparison, the P-glycoprotein-reactive monoclonal antibody MRK-16 and C' inhibited tumor colony formation of MDR cells (8226/DOX6, 95.71 +/- 2.51%; 8226/DOX40, 99.61 +/- 0.43%) but affected that of chemosensitive cells only slightly (8.9 +/- 17.8%). In an attempt to optimize the depletion of myeloma CCC, MM4 was used together with MRK-16. This approach resulted in uniform depletion of myeloma clonogenic colony-forming cells from the chemosensitive (98.32 +/- 1.53%, n = 4) and MDR lines (8226/DOX6, 98.83 +/- 0.08%, n = 4; 8226/DOX40 99.29 +/- 0.62, n = 7) but did not result in enhanced CCC depletion. When DOX40 cells were mixed with normal bone marrow (BM) in the ratio of 90:10 (BM:MM), either MM4 or MRK-16 and C' depleted MM colonies (98.8 +/- 0.71% and 98.10 +/- 1.0%, respectively) without affecting the majority of BM progenitor cells. These observations suggest that either MM4 or MRK-16 is useful for depleting MDR myeloma clonogenic colony-forming cells.
Cancer Res 1989 Sep 01
PMID:Elimination of chemoresistant multiple myeloma clonogenic colony-forming cells by combined treatment with a plasma cell-reactive monoclonal antibody and a P-glycoprotein-reactive monoclonal antibody. 256 59

The calcium channel blocker verapamil has been shown to reverse multidrug resistance (T. Tsuruo et al., Cancer Res. 41: 1967-1972, 1981), but the mechanism of action of this agent has not been fully elucidated. A radioactive photoactive analogue of verapamil, N-[benzoyl-3,5-3H-(+/-)-5-[(3,4-dimethoxyphenetyl)methylamino]-2- (3,4-dimethoxyphenyl)-2-isopropyl-N-p-azidobenzoylpentylamine, was used to label the plasma membranes of a human myelogenous leukemia cell line (K562), a multidrug-resistant subline selected for resistance to Adriamycin (K562/ADM) and its revertant cell (R1-3). Sodium dodecyl sulfate-polyacrylamide gel electrophoretic fluorograms revealed the presence of an intensely radiolabeled Mr 170,000-180,000 protein in the membranes from K562/ADM but not from the drug-sensitive parental K562 and revertant R1-3 cells. The Mr 170,000-180,000 verapamil acceptor was immunoprecipitated by monoclonal antibody MRK16 specific for P-glycoprotein associated with multidrug resistance, indicating that P-glycoprotein in the plasma membrane is a major target of verapamil in K562/ADM cells. The photolabeling of P-glycoprotein with N-[benzoyl-3,5-3H]-(+/-)-5-[(3,4-dimethoxyphenetyl)methylamino]-2- (3,4-dimethoxyphenyl)-2-isopropyl-N-p-azidobenzoylphentylamine was significantly blocked by other calcium channel blockers, nicardipine and diltiazem, that have been shown to overcome multidrug resistance. In addition, the photolabeling was partially blocked by Adriamycin, vincristine, and colchicine, suggesting that the specific binding sites for verapamil on P-glycoprotein are closely related to the binding sites for these calcium channel blockers and antitumor agents. To determine whether verapamil could be a substrate for P-glycoprotein, the cellular accumulation of [3H]verapamil into K562 and K562/ADM was evaluated. The accumulation of [3H]verapamil in the multidrug-resistant cells was 30% of K562 cells and increased when K562/ADM cells were treated with vincristine and nicardipine at 5 microM, indicating that the P-glycoprotein transports verapamil as well as other antitumor agents in the multidrug-resistant cells. These results suggest that verapamil enhances antitumor agent retention through competition for closely related binding sites on P-glycoprotein.
Cancer Res 1989 Sep 15
PMID:Reversal mechanism of multidrug resistance by verapamil: direct binding of verapamil to P-glycoprotein on specific sites and transport of verapamil outward across the plasma membrane of K562/ADM cells. 256 30

We studied transepithelial transport of 3H-labeled hydrophobic cationic drugs in epithelia formed by wild-type and by drug-resistant Madin-Darby canine kidney (MDCk) cells that had been infected with a retrovirus carrying the multidrug-resistance (MDR1) cDNA which encodes the P-glycoprotein. P-glycoprotein is an ATP consuming plasma membrane multidrug transporter responsible for the efflux of cytotoxic chemotherapeutic drugs from resistant cancer cells. Wild-type MDCK cells have small amounts of P-glycoprotein detected by immunoprecipitation. Net transepithelial transport across wild-type MDCK epithelia was demonstrated. Basal to apical flux of 100 nM vinblastine was about six times higher than apical to basal flux. Addition of unlabeled vinblastine reduced basal to apical flux of tracer and increased apical to basal flux of tracer, a pattern expected if there is a saturable pump that extrudes vinblastine at the apical plasma membrane. Daunomycin, vincristine, and actinomycin D were also actively transported and at 20 microM these agents inhibited transport of vinblastine, suggesting that wild-type MDCK cells have a common transporter for all these drugs. Vinblastine transport was also inhibited by 20 microM verapamil, which inhibits the multidrug transporter and reverses multidrug-resistance in non-polarized cells. Net transepithelial transport of all these cytotoxic drugs and of verapamil was much higher in epithelia formed by MDCK cells infected with a human MDR1 virus (MDR-MDCK) which is expressed on the apical surface of MDR-MDCK monolayers. Because the transport of these cytotoxic drugs and verapamil is increased in MDR-MDCK epithelia compared to wild-type MDCK epithelia, transport in both these cell populations can be attributed to P-glycoprotein. These results are consistent with a role for P-glycoprotein in multidrug secretory transport across the epithelium of the proximal tubule since P-glycoprotein is normally expressed on the apical membrane of proximal tubule cells.
J Biol Chem 1989 Sep 05
PMID:Transepithelial transport of drugs by the multidrug transporter in cultured Madin-Darby canine kidney cell epithelia. 257 70

The biosynthesis, processing, and half-life of the drug efflux pump, P-glycoprotein, were studied in human multidrug-resistant KB (KB-C2) cells selected for resistance to colchicine. An antibody directed against a synthetic oligopeptide corresponding to the amino-acid sequence (Glu-393-Lys-408) of P-glycoprotein from human mdr1 cDNA was prepared in rabbits. With immunoblotting and immunoprecipitation, we detected a 140-170 kDa protein in KB-C2 cells but not in parental sensitive KB cells. KB-C2 cells made a 125 kDa precursor that was slowly processed (t1/2 = 45 min) to the mature form of 140-150 kDa. The processing rate of P-glycoprotein was slower than that of low-density lipoprotein receptor. We detected another 160-180 kDa smear band, which might be a completely denatured form of P-glycoprotein. With immunoblotting, a minor band of high molecular mass (greater than 500 kDa) was also detected and this form increased after the cells were treated with chemical cross-linker, 1,5-difluoro-2,4-dinitrobenzene. The half-life of P-glycoprotein was long; no significant loss of P-glycoprotein was observed within 24 h after synthesis. Cells treated with tunicamycin produced a 120 kDa form of P-glycoprotein which was no longer processed but showed stability similar to that of the mature 140-150 kDa form. Agents that reverse multidrug resistance, phorbol ester and transport substrate did not affect the stability of P-glycoprotein.
Biochim Biophys Acta 1989 Sep 15
PMID:Biosynthesis, processing and half-life of P-glycoprotein in a human multidrug-resistant KB cell. 257 Jun 11

In multidrug-resistant murine J774.2 cells, the mdr1a and mdr1b genes encode the 120- and 125-kDa P-glycoprotein precursors, respectively (Hsu, S. I., Lothstein, L., and Horwitz, S.B. (1989) J. Biol. Chem. 264, 12053-12062). It is shown here that a J774.2 cell line selected for vinblastine resistance (J7.V3) switched from the 125- to 120-kDa precursor when cells that were maintained in 20 nM vinblastine were grown in 40 nM vinblastine for 20 months. The rate of switching was accelerated by growing cells in higher levels of vinblastine. These findings suggest that cells which express mdr1a have a selective growth advantage compared to cells which express mdr1b. Consistent with this hypothesis, the switching event that occurs in cells maintained at 40 nM vinblastine was correlated with 3.5-5-fold higher levels of resistance to vinblastine, taxol, and doxorubicin in the absence of any detectable increase in the amount of immunoreactive P-glycoprotein. These findings suggest that P-glycoproteins derived from mdr1a and mdr1b are functionally distinct.
J Biol Chem 1989 Sep 25
PMID:Alternate overexpression of two P-glycoprotein [corrected] genes is associated with changes in multidrug resistance in a J774.2 cell line. 257 Jul 78

The human MDR (P-glycoprotein) gene family is known to include two members, MDR1 and MDR2. The product of the MDR1 gene, which is responsible for resistance to different cytotoxic drugs (multidrug resistance), appears to serve as an energy-dependent efflux pump for various lipophilic compounds. The function of the MDR2 gene remains unknown. We have examined the structure of the human MDR gene family by Southern hybridization of DNA from different multidrug-resistant cell lines with subfragments of MDR1 cDNA and by cloning and sequencing of genomic fragments. We have found no evidence for any other cross-hybridizing MDR genes. The sequence of two exons of the MDR2 gene was determined from genomic clones. Hybridization with single-exon probes showed that the human MDR1 gene is closely related to two genes in mouse and hamster DNA, whereas MDR2 corresponds to one rodent gene. The human MDR locus was mapped by field-inversion gel electrophoresis, and both MDR genes were found to be linked within 330 kilobases. The expression patterns of the human MDR genes were examined by enzymatic amplification of cDNA. In multidrug-resistant cell lines, increased expression of MDR1 mRNA was paralleled by a smaller increase in the levels of MDR2 mRNA. In normal human tissues, MDR2 was coexpressed with MDR1 in the liver, kidney, adrenal gland, and spleen. MDR1 expression was also detected in colon, lung, stomach, esophagus, muscle, breast, and bladder.
Mol Cell Biol 1989 Sep
PMID:Structure and expression of the human MDR (P-glycoprotein) gene family. 257 Oct 78

Tumor cell resistance to anthracyclines, epipodophyllotoxins and vinca alkaloids, called multi-drug resistance (MDR) is intimately linked to changes in the plasma membrane which facilitate an increased energy dependent drug extrusion in the resistant cell compared to the wild type cell. Isolated plasma membrane vesicles from wild type Ehrlich ascites tumor cells (EHR2) and the daunorubicin (DNR) resistant subline EHR2/DNR+ were utilised to study binding and possible transport of DNR and vincristine (VCR). A significant ATP enhanced increase in VCR binding to vesicles from EHR2/DNR+ compared to EHR2 was demonstrated. Furthermore, an increase in ATP enhanced VCR binding in proportion to content of the MDR associated P-glycoprotein was seen in plasma membrane vesicles prepared from various benign human endocrine tumors. VCR binding to EHR2/DNR+ vesicles was inhibited by other vinca alkaloids greater than actinomycin D greater than colchicine greater than anthracyclines, with 35-75 microM concentrations of anthracyclines needed for 50% inhibition. VCR binding to EHR2/DNR+ vesicles was pH and temperature dependent with an activation energy of -30 kJ/mol and was decreased by replacement of Na+ with K+ and by addition of Ca2+. Preincubation of vesicles with monoclonal antibody against the C terminal of P-glycoprotein had no effect on VCR binding and osmolality tests failed to show genuine transmembranal transport of VCR. DNR binding was similar in plasma membrane vesicles from both cell lines, and showed none of the characteristics mentioned for VCR. Furthermore, a radiolabeled N-hydroxysuccinimide ester derivative of doxorubicin, which inhibited VCR binding to EHR2/DNR+ membranes to an even greater extent than doxorubicin, labeled plasma membrane proteins from EHR2 and EHR2/DNR+ identically and did not demonstrate any binding to P-glycoprotein. Therefore, even though the study confirms the close link between vinca alkaloid binding and P-glycoprotein, it could not detect a similar association between anthracyclines and P-glycoprotein thus attesting to the complexity of the MDR phenotype.
Biochem Pharmacol 1989 Sep 15
PMID:Daunorubicin and vincristine binding to plasma membrane vesicles from daunorubicin-resistant and wild type Ehrlich ascites tumor cells. 257 33

Amplification and expression of the mdr1 gene encoding P-glycoprotein have been studied in H69/LX4 a multidrug resistant variant (MDR) of small cell lung cancer (SCLC) cell line NCI-H69. Recently a second independently derived MDR variant of this cell line designated H69/AR was found by others not to show amplification, rearrangement or over-expression of the mdr1 gene. The present study reports that in marked contrast to H69/AR, H69/LX4 shows amplification and expression of the P-glycoprotein gene and raises the possibility that P-glycoprotein hyperexpression may be a clinically relevant component of MDR in some SCLC tumours.
Br J Cancer 1989 Sep
PMID:Amplification and expression of mdr1 gene in a multidrug resistant variant of small cell lung cancer cell line NCI-H69. 257 55

The development of multidrug resistance in MCF7 human breast cancer cells is associated with overexpression of P-glycoprotein, changes in activities of several detoxication enzymes, and loss of hormone sensitivity and estrogen receptors (ERs). We have cloned the cDNA for one of the drug-detoxifying enzymes overexpressed in multidrug-resistant MCF7 cells (AdrR MCF7), the anionic isozyme of glutathione S-transferase (GST pi). Hybridization with this GST pi cDNA, GST pi-1, demonstrated that increased GST pi activity in AdrR MCF7 cells is associated with overexpression but not with amplification of the gene. We mapped the GST pi gene to human chromosome 11q13 by in situ hybridization. Since multidrug resistance and GST pi overexpression are associated with the loss of ERs in AdrR MCF7 cells, we examined several other breast cancer cell lines that were not selected for drug resistance. In each of these cell lines we found an inverse association between GST pi expression and ER content. We also examined RNA from 21 primary breast cancers and found a similar association between GST pi expression and ER content in vivo. GST pi mRNA content in 11 ER-positive tumors (less than or equal to 10 fmol/mg of protein) was significantly different from the GST pi content of 10 ER-negative tumors (P = 0.002; Mann-Whitney Wilcoxon test for two independent samples). The finding of similar patterns of expression of a drug-detoxifying enzyme and of ERs in vitro as well as in vivo suggests that ER-negative breast cancer cells may have greater protection against antineoplastic agents conferred by GST pi than ER-positive tumors.
Proc Natl Acad Sci U S A 1988 Sep
PMID:Isolation of the human anionic glutathione S-transferase cDNA and the relation of its gene expression to estrogen-receptor content in primary breast cancer. 284 75

Overexpression of an immunologically conserved, cell-surface glycoprotein (P-glycoprotein) is consistently associated with multidrug resistance in cell lines in vitro. A preliminary survey of specimens from 12 solid tumor types in our laboratories indicates significant overexpression of P-glycoprotein in some sarcomas. When tested by immunoblotting with monoclonal antibodies directed against P-glycoprotein; tumors from six of 25 sarcoma patients displayed elevated levels of P-glycoprotein. Three of the sarcoma patients exhibiting P-glycoprotein had not previously been exposed to chemotherapy, implying that overexpression of this marker and possible concomitant multidrug resistance may not depend only on selection during prior drug treatments. The P-glycoprotein overexpression in the sarcoma specimens is evidence for the presence of multidrug resistant cells in these tumors; thus, our data suggest that this mode of resistance may have clinical significance in sarcoma patients.
J Clin Oncol 1987 Sep
PMID:P-glycoprotein in human sarcoma: evidence for multidrug resistance. 288 42


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