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)

For investigation of relative differences in mRNA expression levels and of correlations in the expression of genes possibly involved in multidrug resistance (MDR) of acute myelogenous leukemias (AML), a complementary DNA polymerase chain reaction (cDNA-PCR) analysis was established for the genes encoding MDR1/P-glycoprotein, the multidrug resistance-associated protein (MRP), topoisomerase II alpha, topoisomerase II beta, topoisomerase I, glutathione S-transferase pi, protein kinase C (PKC) isozymes alpha, beta 1, beta 2, epsilon, eta, theta and cyclin A. In a first descriptive study comprising samples of childhood or adult AML we calculated the mean values from primary (n=14) or relapsed (n=23) states of the diseases, respectively. We found in the latter significant increases of MDR1, MRP, gst pi, and PKC theta gene expression. MDR1 and MRP gene expression levels were generally correlated (rs= +0.4128, P<0.02, n=37), as well as topoisomerase II alpha and cyclin A gene expression levels (rs= +0.8727, P<0.0001, n=35). Within the group of relapsed state AML a significant negative correlation between the gene expression levels of MDR1 and topoisomerase II alpha (rs= -0.5500, P<0.01, n=22) was observed. Remarkably, highly significant positive correlations were found for MDR1/PKC eta (rs= +0.5560, P<0.001, n=32), MRP/PKC theta (rs= +0.6573, P<0.0001, n=34) and MRP/PKC eta (rs= +0.5241, P<0.005, n=32).
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PMID:Expression of PKC isozyme and MDR-associated genes in primary and relapsed state AML. 864 57

P-glycoprotein (P-gp) is an active transporter that can confer multidrug resistance by pumping cytotoxic drugs out of cells and tumors. P-gp is phosphorylated at several sites in the "linker" region, which separates the two halves of the molecule. To examine the role of phosphorylation in drug transport, we mutated P-gp such that it could no longer be phosphorylated by protein kinase C (PKC). When expressed in yeast, the ability of the mutant proteins to confer drug resistance, or to mediate [3H]vinblastine accumulation in secretory vesicles, was indistinguishable from that of wild type P-gp. A matched pair of mammalian cell lines were generated expressing wild type P-gp and a non-phosphorylatable mutant protein. Mutation of the phosphorylation sites did not alter P-gp expression or its subcellular localization. The transport properties of the mutant and wild type proteins were indistinguishable. Thus, phosphorylation of the linker of P-gp by PKC does not affect the rate of drug transport. In light of these data, the use of agents that alter PKC activity to reverse multidrug resistance in the clinic should be considered with caution.
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PMID:Protein kinase C-mediated phosphorylation does not regulate drug transport by the human multidrug resistance P-glycoprotein. 866 68

According to multiple reports, progesterone is not transported by P-glycoprotein (Pgp), which mediates multidrug resistance through active drug efflux. However, progesterone has been shown to block Pgp- mediated efflux of other drugs. To extend these observations and to examine the effect of modulating Pgp phosphorylation, the accumulation of progesterone and 14 other steroids in untreated and calphostin C-treated multidrug-resistant human colon carcinoma SW620 Ad300 cells was compared to the accumulation in parental SW620 cells. However, the accumlation of more hydrophilic steroids was reduced by as much as 50%. Progesterone and progesterone-like compounds, however were potent inhibitors of Pgp-mediated vinblastine efflux; increased antagonism correlated with increased steroid hydrophobicity. Treatment with calphostin C, a PKC inhibitor which decreases Pgp phosphorylation, increased progesterone efflux, modulated Pgp antagonism by steroids, and inhibited photoaffinity labeling of Pgp by progesterone. These results extend previous observations that Pgp can mediate the transport of, and be antagonized by, a variety of steroids and that these properties vary with both steroid's hydrophobicity and the phosphorylated state of Pgp.
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PMID:Steroid treatment, accumulation, and antagonism of P-glycoprotein in multidrug-resistant cells. 866 72

The possible regulation of the multidrug-resistant (MDR) phenotype and P-glycoprotein by protein kinase C (PKC) was investigated in the doxorubicin (Dox)-resistant MCF-7 cell line (MCF-7/Dox). In a clonogenic assay, cells exposed to 100 nM phorbol 12-myristate 13-acetate (PMA) for 1 hr were about 3-fold more resistant to Dox than were cells exposed to Dox alone. The PKC inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7, 30 microM) completely blocked the PMA-induced effect, but did not reverse the MDR phenotype. Complete down-regulation of PKC from MCF-7/Dox cells by 24-hr preincubation with PMA did not alter the degree of Dox resistance. Intracellular accumulation of [14C]Dox decreased from a baseline of 28 pmol/10(6) cells to 15 pmol/10(6) cells in the presence of 100 nM PMA. The reduced Dox accumulation in the presence of PMA was not blocked by pretreatment of cells with H7. Following a 24-hr pretreatment with PMA, the cells accumulated almost equal amounts of [14C]Dox in the absence or presence of PMA. Cells from PMA-treated colonies showed significantly higher levels of expression of P-glycoprotein when compared with those from control colonies. H7 did not affect the basal level of P-glycoprotein in cells from control colonies or PMA-induced overexpression of P-glycoprotein in cells from PMA-treated colonies. Upon stimulation with PMA (100 nM), PKC alpha and beta translocated to the cell membrane and nucleus and PKC delta and epsilon to the perinuclear membrane and the nucleus, respectively. H7 (30 microM) completely inhibited PMA-induced translocations of PKC delta and epsilon, whereas it only partially blocked the translocations of PKC alpha and beta. These results suggest that PMA appears to alter Dox resistance and intracellular Dox accumulation in a PKC-dependent manner and to induce increased expression of P-glycoprotein in MCF-7/Dox cells. Differential effects of H7 on the PMA-induced changes suggest that different isoforms of PKC may be involved in cell growth and drug accumulation processes as well as P-glycoprotein expression.
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PMID:Selective inhibition of the effects of phorbol ester on doxorubicin resistance and P-glycoprotein by the protein kinase C inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7) in multidrug-resistant MCF-7/Dox human breast carcinoma cells. 868 92

Dexniguldipine hydrochloride (DNIG) is a potent antineoplastic agent with well-documented anti-(protein kinase C) activity and an ability to reverse multidrug resistance. Given the importance of protein kinase C (PKC) activity in proliferation and differentiation, we examined the effect of DNIG on several parameters of Friend erythroleukemia cell (FELC) activity. Particular attention was paid to proliferation, hexamethylene-bisacetamide-(HMBA)-induced differentiation, nuclear localization of protein kinase C, and nuclear protein phosphorylation. P-glycoprotein expression was also followed as an indicator of changes in multidrug resistance. At 2.5 microM, DNIG caused a significant decrease in the rate of FELC proliferation, while maintaining a cellular viability of greater than 80%, whether exposure to the drug was continuous over 96 h or took the form of a 6-h pulse/chase. DNA synthesis was decreased in cells exposed to DNIG for 20 h. Flow cytometry showed a marked increase in the percentage of cells in S phase of the cell cycle. Phosphorylation studies revealed decreased phosphorylation of two nuclear proteins (80 kDa and 47 kDa) following a 4-h exposure to the drug. HMBA-induced differentiation was significantly inhibited with continuous exposure to DNIG, and this effect appears to be a pre-commitment one, as 6-h pulse/chase exposures also resulted in inhibition of differentiation. Cells induced to differentiate with HMBA also demonstrated a decrease in the quantity of the 80-kDa phosphoprotein. Western blotting revealed that, even in the face of decreased phosphorylation, exposure to this PKC inhibitor resulted in an increase in the amount of nuclear PKC alpha. Finally, levels of P-glycoprotein were decreased in the presence of this drug. Our work identifies several effects of the PKC inhibitor DNIG on FELC and suggests several roles for PKC in regulating FELC proliferation and differentiation. Additionally, these results suggest that this PKC inhibitor may increase the effect of other chemotherapeutic drugs, particularly S-phase-specific ones, by increasing the length of S phase and decreasing multidrug resistance. The possibility of combination therapy with DNIG and other antineoplastic agents should be investigated further in light of these findings.
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PMID:Dexniguldipine hydrochloride, a protein-kinase-C-specific inhibitor, affects the cell cycle, differentiation, P-glycoprotein levels, and nuclear protein phosphorylation in Friend erythroleukemia cells. 869 46

Indirect evidence has suggested that P-glycoprotein (P-gp), the multidrug transporter, is phosphorylated by protein kinase C (PKC) and that phosphorylation modulates its transport function. To address the first premise more directly, ie., that P-gp is phosphorylated by PKC, we investigated the interaction between P-gp and PKC in sensitive and multidrug resistant MCF-7 and KB human carcinoma cell lines. We found that P-gp and PKC were coimmunoprecipitated from the multidrug-resistant cell lines MCF-7/AdrR and KB-V-1, using antibodies to either protein. The association between the two proteins was enhanced by phorbol 12-myristate 13-acetate, an analogue of diacylglycerol that induces translocation of PKC to the plasma membrane. The anti-P-gp immunoprecipitates contained PKC activity as measured by direct phosphorylation reactions. The interaction of PKC with P-gp displayed isozyme specificity: PKC-alpha, -beta, gamma, -epsilon, and -phi, but not -delta, -mu, -zeta, -lambda, were found to coimmunoprecipitate with P-gp. These studies indicate that P-gp closely interacts with PKC and serves as a substrate, and that specific isozymes of this kinase may be involved in the phosphorylation of the multidrug transporter.
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PMID:Interaction of P-glycoprotein with protein kinase C in human multidrug resistant carcinoma cells. 875 35

To investigate the role of protein kinase C (PKC) in the regulation of multidrug resistance and P-glycoprotein (P-gp) phosphorylation, the natural isomer of sphingosine (SPH), D-erythro sphingosine (De SPH), and its three unnatural stereoisomers were synthesized. The SPH isomers showed similar potencies as inhibitors of in vitro PKC activity and phorbol binding, with IC50 values of approximately 50 microM in both assays. Treatment of multidrug-resistant MCF-7ADR cells with SPH stereoisomers increased vinblastine (VLB) accumulation up to 6-fold at 50 microM but did not alter VLB accumulation in drug-sensitive MCF-7 wild-type (WT) cells or accumulation of 5-fluorouracil in either cell line. Phorbol dibutyrate treatment of MCF-7ADR cells increased phosphorylation of P-gp, and this increase was inhibited by prior treatment with SPH stereoisomers. Treatment of MCF-7ADR cells with SPH stereoisomers decreased basal phosphorylation of the P-gp, suggesting inhibition of PKC-mediated phosphorylation of P-gp. Most drugs that are known to reverse multidrug resistance, including several PKC inhibitors, have been shown to directly interact with P-gp and inhibit drug binding. SPH stereoisomers did not inhibit specific binding of [3H] VLB to MCF-7ADR cell membranes or [3H]azidopine photoaffinity labeling of P-gp or alter P-gp ATPase activity. These results suggest that SPH isomers are not substrates of P-gp and suggest that modulation of VLB accumulation by SPH stereoisomers is associated with inhibition of PKC-mediated phosphorylation of P-gp.
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PMID:Effects of sphingosine stereoisomers on P-glycoprotein phosphorylation and vinblastine accumulation in multidrug-resistant MCF-7 cells. 875 33

The multidrug resistant (MDR) phenotype is a well-studied subject that has been recognized as a determinant underlying specific types of drug resistance in human cancer. Although it is clear that the P-glycoprotein plays a major role in MDR, it is not clear whether post-translational modifications such as phosphorylation have any major impact on its modulation. The laboratory of Dr. Bruce Chabner was one of the first to describe increased expression and activity of protein kinase C (PKC) associated with the MDR phenotype. Since that time, a similar correlation has been observed in many other MDR cell lines. Most of these studies have been performed with doxorubicin-selected cells that have acquired MDR and have shown increased PKC activity, mainly for PKC-alpha isoenzyme. Intrinsic MDR in human renal cell carcinoma lines has been shown to correlate directly with PKC activity, but further studies with intrinsic MDR cell lines are needed before any conclusions can be drawn. More recent evidence suggests that there is a complex biochemical process by which PKC isoenzymes differentially phosphorylate specific serine residues in the linker region of P-glycoprotein which may lead to alterations in P-glycoprotein ATPase and drug-binding functions. To further complicate matters, PKC plays an important role in anti-apoptotic pathways, which can confound the dissection and elucidation of drug-resistance mechanisms. However, these areas are still under active investigation and not fully answered. Further studies are needed to specifically answer the question of whether PKC directly modulates basal and/or drug-stimulated P-glycoprotein function. This manuscript reviews the majority of the literature on PKC and MDR, as well as offers caveats for interpretation of these studies to answer the above questions.
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PMID:P-glycoprotein, multidrug resistance and protein kinase C. 882 Sep 51

We assessed the effect of the protein kinase C inhibitor 2,6-diamino-N-([1-(1-oxotridecyl)-2-piperidinyl]methyl)hexanami de (NPC 15437) on the action of anthracyclines, epipodophyllotoxins and vinca alkaloids in P-glycoprotein (Pgp)-expressing CH(R)C5 hamster ovary and MCF-7/Adria(R) human breast cancer cells. Flow microfluorimetry revealed that treatment of CH(R)C5 cells with 75 microM NPC 15437 for 1 h resulted in a 6- to 10-fold increase in the nuclear accumulation of daunorubicin. Colony forming assays revealed that treatment with 75 microM NPC 15437 was associated with a 4-fold decrease in the LD90 for etoposide and a 2.5-fold decrease in the LD50 for vincristine. At higher concentrations of NPC 15437, greater modulation of anthracycline accumulation was observed; but NPC 15437 itself inhibited subsequent colony formation. Similar effects on drug accumulation and cytotoxicity were observed in MCF-7/Adria(R) cells. Experiments designed to investigate the mechanism by which NPC 15437 exerts these effects revealed that treatment with the protein kinase C activator phorbol-12-myristate 12-acetate partially reversed the effect of NPC 15437, suggesting that NPC 15437 was exerting an effect through protein kinase C. Photoaffinity labeling experiments revealed that NPC 15437 also inhibited the binding of [3H]-azidopine to Pgp in isolated membrane vesicles. These results identify NPC 15437 [correction of NPC15437] as the prototype of a new class of potential Pgp modulators but indicate that the effects of this agent as a modulator are potentially limited by its cytotoxicity.
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PMID:Evaluation of 2,6-diamino-N-([1-(1-oxotridecyl)-2-piperidinyl]methyl)- hexanamide (NPC 15437), a protein kinase C inhibitor, as a modulator of P-glycoprotein-mediated resistance in vitro. 882 46

Inhibition of protein kinase C (PKC) is discussed as a new approach for overcoming multidrug resistance (MDR) in cancer chemotherapy. For evaluation of this concept we applied the bisindolylmaleimide GF 109203X, which shows a highly selective inhibition of PKC isozymes alpha, beta 1, beta 2, gamma, delta and epsilon in vitro. The efficacy of this compound in modulation of MDR was examined using several P-glycoprotein (P-gp)-overexpressing cell lines including a MDR1-transfected HeLa clone, and was compared with the activities of dexniguldipine-HCI (DNIG) and dexverapamil-HC1 (DVER), both of which essentially act via binding to P-gp. As PKC alpha has been suggested to play a major role in P-gp-mediated MDR, cell lines exhibiting different expression levels of this PKC isozyme were chosen. On crude PKC preparations or in a cellular assay using a cfos(-711)CAT-transfected NIH 3T3 clone, the inhibitory qualities of the bisindolylmaleimide at submicromolar concentrations were demonstrated. At up 1 microM final concentrations of the PKC inhibitor GF 109203X, a concentration at which many PKC isozymes should be blocked substantially, no cytotoxic or MDR-reversing effects whatsoever were seen, as monitored by 72 h tetrazolium-based colorimetric MTT assays or a 90 min rhodamine 123 accumulation assay. Moreover, depletion of PKC alpha by phorbol ester in HeLa-MDR1 transfectants had no influence on rhodamine 123 accumulation after 24 or 48 h. MDR reversal activity of GF 109203X was seen at higher final drug concentrations, however. Remarkably, [3H]vinblastine-sulphate binding competition experiments using P-gp-containing crude membrane preparations demonstrated similar dose dependencies as found for MDR reversion by the three modulators, i.e. decreasing efficacy in the series dexniguldipine-HCl > dexverapamil-HCl > GF 109203X. Similar interaction with the P-gp in the micromolar concentration range was revealed by competition of GF 109203X with photoincorporation of [3H]azidopine into P-gp-containing crude membrane preparations. No significant effect of the PKC inhibitor on MDR1 expression was seen, which was examined by cDNA-PCR. Thus, the bisindolylmaleimide GF 109203X probably influences MDR mostly via direct binding to P-gp. Our work identifies the bisindolylmaleimide GF 109203X as a new type of drug interacting with P-gp directly, but does not support the concept of a major contribution of PKC to a P-gp-associated MDR, at least using the particular cellular model systems and the selective, albeit general, PKC inhibitor GF 109203X.
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PMID:Effects of the selective bisindolylmaleimide protein kinase C inhibitor GF 109203X on P-glycoprotein-mediated multidrug resistance. 882 55


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