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)

An overexpression of plasma membrane 170-180 kDa P-glycoproteins is consistently found in multidrug-resistant (MDR) cell lines. In this study MRK-16, a monoclonal antibody (mAb) reacting with P-glycoprotein is used to study the putative functional role of this protein in vincristine (VCR) and daunorubicin (DNR) cellular accumulation in the MDR human ovarian carcinoma cell line 2780AD. We established that this cell line is highly cross-resistant to vincristine and daunomycin, related to a greatly reduced drug accumulation. Verapamil (Vp) (8 microM) caused a 3.6-fold increase in DNR as well as VCR accumulation. Exposition of 2780AD cells to MRK-16 led to an increase of 30% in cellular accumulation of VCR, both in normal growth medium as well as in medium without added glucose and with sodium azide, which largely depleted cellular ATP levels. No increase in DNR accumulation was found under these conditions. However, in the presence of 8 microM Vp, MRK-16 increased not only VCR but also DNR accumulation with about 30%. The relative increase of DNR accumulation was constant in a concentration range of 0.2-4 microM DNR. These data indicate that mAbs against P-glycoprotein might potentiate the action of calcium antagonists like Vp to increase cellular anthracycline accumulation.
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PMID:Increase of daunorubicin and vincristine accumulation in multidrug resistant human ovarian carcinoma cells by a monoclonal antibody reacting with P-glycoprotein. 289 43

A monoclonal antibody (MAb), MRK 16, specific to Adriamycin-resistant human myelogenous leukemia cell line K562, was used to examine whether the antigen molecules (P-glycoprotein) recognized by the MAb are present in the adrenals. The materials examined included 61 human adrenals and several cell lines. Immunohistochemical analysis revealed that almost all of the human adrenal specimens (59 out of 61) were stained positively with MAb MRK 16 and that the antigen was strongly expressed even in cases where anticancer agents had not been given. Immunoprecipitation showed that the Mr 170,000-180,000 glycoprotein was present in all of the adult adrenals but not in fetal and neonatal adrenals. Furthermore, fluorescence image analysis revealed that the P-glycoprotein was more strongly expressed in the cortex than in the medulla, showing a tendency to occur in cell clusters in the latter area. The cell lines derived from animal adrenals (SW-13, Y-1, and PC-12) showed no positive staining with MAb MRK 16. It is suggested that this glycoprotein may be related to maturation of the adrenal, in which it possibly plays a physiological role.
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PMID:Apparent stronger expression in the human adrenal cortex than in the human adrenal medulla of Mr 170,000-180,000 P-glycoprotein. 289 56

One form of multidrug resistance is due to the expression of a 170-kDa energy-dependent drug efflux pump called P-glycoprotein in the plasma membranes of human cancer cells. We have prepared conjugates of Pseudomonas toxin with the anti-P-glycoprotein monoclonal antibody MRK-16. These anti-P-glycoprotein-toxin conjugates specifically kill multidrug-resistant human KB cells. Similar conjugates could be useful in cancer therapy to reduce or eliminate multidrug-resistant tumor populations in tumors intrinsically resistant to chemotherapy or in populations that become resistant during combination chemotherapy.
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PMID:A monoclonal antibody-Pseudomonas toxin conjugate that specifically kills multidrug-resistant cells. 349 6

Expression of the multidrug resistance efflux pump P-glycoprotein (Pgp) was measured in a series of AML patients using two flow cytometry methods. Transport function was assessed by measuring the modulating effect of the Pgp inhibitor cyclosporin A (CsA) on the cellular accumulation of daunorubicin, and Pgp antigen expression by surface immunofluorescence using the MRK-16 antibody. Both methods showed a wide range of values for Pgp expression between individual patients, but in contrast to a series of cell lines expressing Pgp there was no correlation between antigen expression and transport function in the clinical samples. As previously reported for chronic lymphocytic leukemia (CLL), pretreatment with neuraminidase markedly improved MRK-16 staining in some cases, indicating that abnormal glycosylation can cause epitope masking in AML blasts. Because experience with cell lines shows that Pgp expression is a continuous variable which correlates with the level of drug resistance, rather than the 'positive' or 'negative' which are frequently reported by clinical flow cytometry laboratories, we used a calibration procedure to estimate the actual number of Pgp molecules expressed in the AML samples. Despite the additional refinements of neuraminidase treatment and antigen quantification, the correlation between Pgp antigen expression and daunorubicin accumulation remained extremely weak (r = 0.11; P = 0.63). It is suggested that the assay for transport function can detect molecules that affect daunorubicin accumulation but are antigenically distinct from classical P-glycoprotein. Heterogeneity of multidrug resistance efflux pumps might in part explain the relatively weak prognostic significance of immunofluorescence detection of Pgp in AML patients.
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PMID:Discordant P-glycoprotein antigen expression and transport function in acute myeloid leukemia. 747 79

Multiple myeloma is basically an incurable cancer. Most patients respond initially to chemotherapy with reduction in bone marrow (BM) plasma cells and monoclonal Ig levels, but the disease nearly always recurs and becomes refractory to therapy. The objective of this study was to characterize the expression of the multidrug transport pump, P-glycoprotein 170 (P-gp), in myeloma. The great majority of B cells from peripheral blood mononuclear cells (PBMCs) in myeloma express P-gp, detected by the monoclonal antibody MRK-16. P-gp+ blood B cells exhibit extensive DNA hyperdiploidy, suggesting replicative abnormality characteristic of malignant growth. We speculate these represent a stem cell population in myeloma. The proportion of B cells expressing P-gp was comparable among untreated myeloma patients and those treated with chemotherapy, biologic response modifiers, or off treatment. Among BM cells, P-gp was absent or low in untreated myeloma patients but was expressed at high levels on BM cells from patients previously treated with chemotherapy. For untreated patients the majority of B/plasma cells expressing P-gp are located in PBMCs, not the BM cells. Flow cytometric analysis of rhodamine 123 dye efflux indicated a functional P-gp that was efficiently blocked by verapamil or cyclosporin A (CsA). Both the CD11bhi CD19+ B cells and the T cells in myeloma PBMCs had active CsA-inhibited dye efflux, but monocytes lacked the ability to efflux dye. Nearly all CD38hi plasma cells from myeloma BM cells retained dye, indicating their lack of a functional transport pump. Thus, PBMC B cells may be the predominant set of drug-resistant tumor cells. Myeloma PBMC B cells were cultured with Adriamycin with or without CsA and drug toxicity evaluated by the induction of apoptosis, using flow cytometry to quantitate DNA disruption. No apoptosis was detectable at 0.01 microgram/mL adriamycin, the in vivo steady-state level, with or without CsA. With 0.1 microgram adriamycin, no apoptosis was detectable in the absence of CsA, but with CsA, 66% of B cells initiated DNA disruption, whereas most T cells were spared. This work suggests that currently used drug dosages are too low to effect P-gp+ B-cell death, even in the presence of CsA. We suggest that blood B cells comprise a highly drug-resistant subset of the myeloma B lineage that escapes conventional chemotherapy and may underlie the almost uniform fatal relapse in myeloma patients.
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PMID:Circulating monoclonal B cells expressing P glycoprotein may be a reservoir of multidrug-resistant disease in multiple myeloma. 750 31

Multidrug resistance (MDR) in tumors is associated with P-glycoprotein (Pgp) expression. In vivo quantitation of Pgp may allow MDR to be evaluated noninvasively prior to treatment planning. The purpose of this study was to radiolabel MRK-16, a monoclonal antibody that targets an external epitope of P-glycoprotein, and perform in vivo quantitation of P-glycoprotein in a MDR xenograft nude mouse model. MRK-16 was labeled with 125I by the iodogen method, with subsequent purification by size exclusion chromatography. Groups of 10 Balb c mice were each xenografted with colchicine-resistant or sensitive neuroblastoma cell lines, respectively. Whole body clearance and tumor uptake over time was quantitated by gamma camera imaging, and biodistribution studies were performed with [125I]MRK-16 and an isotype matched control antibody, A33. Quantitative autoradiography and immunohistochemistry analysis of tumors was also evaluated to confirm specific targetting of [125I]MRK-16. Peak tumor uptake was at 2-3 days post-injection, and was significantly greater in resistance compared to sensitive tumors (mean % injected dose/g +/- SD) (18.76 +/- 2.94 vs 10.93 +/- 0.96; p < 0.05). Quantitative autoradiography verified these findings (19.13 +/- 0.622 vs 12.08 +/- 0.38, p < 0.05). Specific binding of [125I]MRK-16 was confirmed by comparison to [131I]A33 in biodistribution studies, and localized to cellular components of tissue stroma by comparison of histologic and autoradiographic sections of sensitive and resistant tumors. Immunoblot analysis demonstrated a 4.5-fold difference in P-glycoprotein expression between sensitive and resistant cell lines without colchicine selective pressure. We conclude that in vivo quantitation of P-glycoprotein in MDR tumors can be performed with [125I]MRK-16.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:In vivo imaging and specific targeting of P-glycoprotein expression in multidrug resistant nude mice xenografts with [125I]MRK-16 monoclonal antibody. 755 27

To study the molecular function of the multidrug-resistance gene product P-glycoprotein, we purified and reconstituted it into liposomes. Twelve detergents were examined in an attempt to solubilize and reconstitute the transport activity of K562/ADM membrane proteins containing P-glycoprotein. We found that transport activity was effective reconstituted after solubilization with cholate, glycocholate and taurocholate. Other detergents, such as CHAPS, Triton X-100 and deoxycholate, diminished the transport activity. The K562/ADM membrane was solubilized by 1% glycocholate, and P-glycoprotein was purified by MRK-16 immunoaffinity column chromatography to a homogeneous single band on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The purified P-glycoprotein was reconstituted by detergent dialysis into liposomes composed of phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine. The reconstituted P-glycoprotein specifically bound [3H]azidopine and had an ATPase activity that was slightly stimulated when vincristine was added. Furthermore, though its activity was reduced, the reconstituted P-glycoprotein was shown to be an ATP-dependent transporter of vincristine.
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PMID:Reconstitution of purified P-glycoprotein into liposomes. 755 41

P-glycoprotein 170 (P-gp), the multidrug transport pump, excludes drugs from the interior of cells and is inhibited by agents such as cyclosporin A (CsA), verapamil, and FK-506, which are also substrates for the P-gp pump. This work documents the age- and differentiation-related changes in P-gp on T and B lymphocytes from human blood or spleen, and its absence on most thymus and bone marrow cells. Analysis of rhodamine 123 (Rh123) dye efflux, and its inhibition by cyclosporin A, was used as a quantitative measure of functional P-gp, and reactivity with MRK-16 was used as a measure of P-gp surface expression. The dye efflux and phenotypic expression of P-gp+ PBMC appeared equivalent to that of a moderately drug-resistant cell line, although efflux is prolonged. The sensitivity to inhibition by CsA, cyclosporin G (CsG), and PSC833 of P-gp on PBMC, thymocytes, or T-cell lines varied with apparent cell-type specificity. Normal blood and splenic T- or B-cells included 50-80% of cells with surface P-gp (MRK-16+), which mediated CsA-sensitive dye export. The proportion of P-gp+ T- and B-cells was lowest among children under age 10 years, increased in adulthood, and decreased after age 60. Thymus included 30% of P-gp+ cells mediating CsA-sensitive dye export, including CD3-4-8- progenitors and mature CD3hi CD4+8- or CD4-8+ thymocytes. Mature T-cells in cord or adult blood, spleen, and bone marrow included a large proportion (50-60%) with efficient CsA-sensitive dye export, preferentially among the CD45RA+ subset. Monocytes from all tissue sources, and most bone marrow cells, expressed surface P-gp but retained Rh123, suggesting the absence of a functional dye export mechanism. In vitro mitogen-stimulated PBMC T and B lymphocytes lost P-gp function within 4-24 hr, consistent with the observation that P-gp was reduced on antigen-experienced CD45R0+ T-cells in vivo. Drug export by P-gp may protect lymphocytes from toxic effects of CsA, and may contribute to the immunosuppressive effects of such drugs. The developmentally regulated expression of P-gp function on lymphocytes, and its modulation on activated T- or B-cells, suggest an important role in normal immune development.
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PMID:Multidrug transporter P-glycoprotein 170 as a differentiation antigen on normal human lymphocytes and thymocytes: modulation with differentiation stage and during aging. 763 78

Multidrug resistant (MDR) phenotype is characterized by a defect in drug accumulation caused by overexpression of a transmembrane glycoprotein, the P-glycoprotein (P-gp). MDR phenotype can be characterized either with monoclonal antibodies raised against P-gp or with functional tests, most often based on the incorporation of fluorescent compounds. In the present study, data obtained with the monoclonal antibodies C219, JSB1 and MRK16 are compared to those of functional tests performed by flow cytometry including uptake of daunorubicin (DNR), Rhodamine 123 (Rh 123) or Hoechst 33342. Sensitive and resistant cell lines K562S, K562R, KBA1 and KB31, derived either from a human chronic myeloid leukemia or from a human epithelial carcinoma, were used. In resistant cells, P-gp expression was revealed with either the monoclonal antibodies C219, JSB1 or MRK-16. The most specific results were obtained with MRK-16. With functional tests, no matter which dyes were used, the fluorescence was always stronger in sensitive than in resistant cells. However, with DNR and Hoechst 33342, an incorporation of these dyes was exhibited in resistant cells. This phenomenon was not observed with Rh 123, which makes it possible to distinguish clearly between sensitive and resistant cells and to detect as few as 1% of resistant cells. Because of its high sensitivity, the functional test involving incorporation of Rh 123 was successfully used in acute myeloid leukemia to detect multichemoresistant cells.
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PMID:Evaluation of multidrug resistant phenotype by flow cytometry with monoclonal antibodies and functional tests. 765 50

Anti-P-glycoprotein antibody (MRK-16)-dependent cell-mediated cytotoxicity (ADCC) by blood mononuclear cells (MNC) was examined in patients with small cell lung cancer (SCLC) before and after systemic chemotherapy. The effect of in vitro treatment of MNC with interleukin (IL)-2 and macrophage-colony-stimulating factor (M-CSF) was also examined. The ADCC reaction was assessed by a 6 h 51Cr-release assay using a multidrug-resistant (MDR) SCLC cell line (H69/VP cells). The MRK-16 monoclonal antibody was able to augment spontaneous cytotoxicity by MNC, even in SCLC patients. Pretreatment of MNC with IL-2 significantly augmented their ADCC ability in SCLC patients, while M-CSF had no effect on ADCC activity. After the first cycle of systemic chemotherapy, the ADCC activity tended to decline, but ADCC of MNC pretreated with IL-2 was not affected. The results suggest that anti-P-glycoprotein antibody, in combination with a cytokine such as IL-2, may be therapeutically useful against human SCLC resistant to chemotherapeutic drugs.
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PMID:Influence of systemic chemotherapy on anti-P-glycoprotein antibody-dependent cell-mediated cytotoxicity in patients with small cell lung cancer. 766 88


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