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)

We have examined the effects of the nitrosoureas, streptozotocin (STZ) and 1,3-bis(chloroethyl)-1-nitrosourea (BCNU), on a human multiple myeloma cell line, RPMI 8226, and its drug-resistant variants. Cell lines selected for doxorubicin (DOX) resistance alone displayed a STZ and BCNU cytotoxicity profile similar to that of the parent cell line. In contrast, two of the drug-resistant variants selected with DOX plus verapamil, an agent which inhibits P-glycoprotein-mediated multidrug resistance, displayed a collateral sensitivity to STZ and BCNU. Verapamil was included in the selection protocol because it has been shown to inhibit the P-glycoprotein-mediated multidrug resistance phenotype and is now in clinical trials as a chemosensitizing agent. The collateral sensitivity to these nitrosoureas seen in the DOX plus verapamil-selected cell lines is due to the functional loss of a DNA repair molecule, O6-Methylguanine DNA methyltransferase (MGMT). The functional loss of MGMT is secondary to the loss of MGMT gene expression. The loss of MGMT gene expression is not due to loss or gross rearrangement of the MGMT-coding region. If this selection pressure applied in vitro reflects the in vivo situation, then new chemotherapeutic strategies may be devised to exploit this phenomenon. These cell lines will serve as useful models for delineating mechanisms which govern MGMT expression.
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PMID:Collateral sensitivity to nitrosoureas in multidrug-resistant cells selected with verapamil. 138 86

The expression of different genes potentially involved in DNA repair and in cell responses to chemotherapy was evaluated in 33 previously untreated ovarian cancer patients. In biopsies of the same patients the expression of repair genes O6-methylguanine DNA methyltransferase (MGMT), 3-methyladenine DNA glycosylase (MAG), ERCC1, MDR-1, DNA topoisomerase I, DNA topoisomerase IIalpha, and glutathione S-transferase-pi (GST-pi) was assessed by Northern blot analysis. No direct statistical correlation was found between the expression of these genes and the response to chemotherapy (mainly platinum-based with or without doxorubicin and cyclophosphamide). Univariate analysis showed a weak negative correlation (P = 0.037) between the expression of ERCC1 and mortality, whereas no statistically significant correlation was found for other parameters. The MDR-1 gene encoding for the P-glycoprotein P-170 was mostly undetectable in these patients (as assessed by Northern blotting), whereas relatively high levels of MAG and MGMT were found in the majority of patients. A statistically significant correlation was found between the expression of DNA topoisomerase I and the expression of either ERCC1 (P = 0.0026) or GST-pi (P = 0.0279).
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PMID:Expression of genes of potential importance in the response to chemotherapy and DNA repair in patients with ovarian cancer. 910 2

Drug resistance genes can protect normal hematopoietic cells from the toxicity of anticancer agents. Because chemotherapeutic agents are often used in combination in current clinical protocols, coexpression of two different drug resistance genes should be useful in protecting normal bone marrow cells from the hematotoxicities caused by combination chemotherapy. In this study, we have combined the human multidrug resistance gene (MDR1) and human O6-methylguanine DNA methyltransferase (MGMT) gene as drug resistance genes. For the coexpression of two drug resistance genes, we have constructed two bicistronic retrovirus vectors. One vector is Ha-MDR-IRES-MGMT, in which translation of the MDR1 cDNA is cap-dependent and MGMT translation is dependent on an internal ribosome entry site (IRES). The other is Ha-MGMT-IRES-MDR, which has cap-dependent MGMT translation and IRES-dependent MDR1 translation. MGMT-negative HeLa derivative (MR) cells transduced with these retroviruses showed resistance to vincristine (from MDR1) and 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosou rea (ACN; from MGMT). Cells transduced with Ha-MDR-IRES-MGMT showed higher resistance to vincristine and lower resistance to ACNU than those transduced with Ha-MGMT-IRES-MDR. In any case, the resistance levels of cells transduced with either vector were high enough to select transduced cells with vincristine or ACNU. The expression levels of P-glycoprotein or MGMT in the transduced cells determined by FACS and Western blot analysis correlated well with the extent of resistance to vincristine and ACNU, respectively. All of the MGMT-transduced cells expressed higher amounts of MGMT than the MGMT-expressing parental cell line HeLa S3. Murine bone marrow cells transduced with Ha-MDR-IRES-MGMT and selected with vincristine also showed simultaneous resistance to vincristine and ACNU. These results suggest that bicistronic retroviral vectors allow the functional coexpression of two different types of drug resistance genes. This strategy could be applicable to any combination of drug resistance genes.
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PMID:Retroviral coexpression of two different types of drug resistance genes to protect normal cells from combination chemotherapy. 981 70

Selection of human cells for resistance to vincristine or doxorubicin often induces overexpression of the multidrug resistance 1 gene (MDR1), which encodes the cell surface P-glycoprotein, as a result of gene amplification or transcriptional activation. However, the precise mechanism underlying such transcriptional activation of MDR1 remains unclear. The relation between methylation status of CpG sites in the MDR1 promoter region and transcriptional activation of MDR1 has now been investigated. The P-glycoprotein-overexpressing, multidrug-resistant KB/VJ300 and KB-C1 cells, which were established from human cancer KB3-1 cells, were examined; MDR1 is transcriptionally activated but not amplified in KB/VJ300 cells, whereas it is amplified in KB-C1 cells. Determination of the methylation status revealed that the MDR1 promoter region was hypomethylated in KB/VJ300 and KB-C1 cells, but hypermethylated in KB3-1 cells. Prior treatment of KB3-1 cells with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine resulted in a 90-fold increase in the frequency of vincristine-resistance. Of three lines, KB/CdR-1, KB/CdR-2, and KB/CdR-3, established from KB3-1 cells after exposure to 5-aza-2'-deoxycytidine, MspI/HpaII sites in the MDR1 promoter region were hypomethylated in KB/CdR-1 and KB/CdR-2 cells, but not in KB/CdR-3 cells. MDR1 mRNA expression was detected in KB/CdR-1 and KB/CdR-2 cells, but not in KB/CdR-3 cells. The binding of YB-1 and Sp1, transcription factors implicated in MDR1 expression, in the MDR1 promoter was not affected by the methylation status of a neighboring CpG sites. The MDR1 promoter region in KB/VJ300 cells showed an increased sensitivity to DNase I compared with that in KB3-1 cells, suggesting an altered chromatin structure. The methylation status of the promoter region may plays an important role in MDR1 overexpression and in acquisition of the P-glycoprotein-mediated multidrug resistance phenotype.
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PMID:Association of 5' CpG demethylation and altered chromatin structure in the promoter region with transcriptional activation of the multidrug resistance 1 gene in human cancer cells. 1041 57

Cancer chemotherapy is the principal approach for urogenital cancers. However, the acquisition of resistance to anticancer agents is a critical factor that limits the successful treatment of malignancies. The multidrug resistant (MDR) phenotype has been widely recognized in cancer chemotherapy in urogenital tumors and the mechanisms underlying MDR have also been extensively studied. One of the principle mechanisms in MDR is caused by the overexpression of P-glycoprotein (P-gp), encoded by the multidrug resistance gene (MDR1). It functions as an ATP-dependent active efflux pump of chemotherapeutic agents in human cancer cells. Recently, other drug resistance proteins, including multidrug resistance-associated protein (MRP1) and cMOAT (or MRP2), were also identified from multidrug resistant cells. A functional analysis of MRP1 has shown that MRP1 may have the potential to act as a transporter of glutathione conjugates, which has been known as a central detoxification pathway in anticancer agents. Furthermore, several other resistance-related proteins (e.g. glutathione S-transferase, metallothionein, thioredoxin, topoisomerase I, II, O6-alkylguanine-DNA methyltransferase, etc.) have been found to be up- or down-regulated in resistant cells and these molecules are believed to contribute to the resistant phenotype as well. Based on the molecular characteristics identified in MDR, several experimental and clinical approaches have been studied to overcome MDR. One of these strategies is to reverse MDR by using such P-gp inhibitors as verapamil and cyclosporine A. In this review, we summarize the recent advances in MDR-related molecules and clinical trials to circumvent MDR in urogenital carcinomas.
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PMID:Mechanisms of drug resistance in chemotherapy for urogenital carcinoma. 1051 Aug 88

Multidrug resistance (MDR) is a major problem in patients with hematological malignancies. Although drug-resistance is known to be induced by the expression of P-glycoprotein (P-gp) encoded by the MDR-1 gene, little is known about the mechanisms regulating this gene. Herein, we studied the DNA methylation patterns at the enhancer and repressor binding sites of the MDR-1 gene using the human erythroleukemia cell line K562 and its multidrug resistant derivative K562/ADM (adriamycin). Direct DNA sequence analysis demonstrated methylation to be present at the repressor site (minus 110 GC-box) of the MDR-1 gene in K562/ADM cells, but not in parental K562 cells. Methylation-specific PCR (MSP) analysis yielded similar results. Treatment of K562/ADM cells with 5-Aza-2'-deoxycytidine (decitabine; DAC), an inhibitor of DNA methyltransferase, caused demethylation of the repressor binding site of MDR-1 gene, as assessed by MSP, and also decreased P-gp expression, as assessed by flow cytometric and Northern blot analysis. Although it is generally accepted that DAC upregulates gene expression by demethylating the activator binding sites, our present results suggest that DAC induces down-regulation of P-gp expression as a result of demethylation at the repressor binding site in K562/ADM cells. In this regard, methylation-dependent regulation of the MDR-1 gene in K562/ADM cells is unique.
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PMID:Decitabine (5-Aza-2'-deoxycytidine) decreased DNA methylation and expression of MDR-1 gene in K562/ADM cells. 1106 27

Overexpression of the human multidrug resistance gene 1 (MDR1) is a negative prognostic factor in leukemia. Despite intense efforts to characterize the gene at the molecular level, little is known about the genetic events that switch on gene expression in P-glycoprotein-negative cells. Recent studies have shown that the transcriptional competence of MDR1 is often closely associated with DNA methylation. Chromatin remodeling and modification targeted by the recognition of methylated DNA provide a dominant mechanism for transcriptional repression. Consistent with this epigenetic model, interference with DNA methyltransferase and histone deacetylase activity alone or in combination can reactivate silent genes. In the present study, we used chromatin immunoprecipitation to monitor the molecular events involved in the activation and repression of MDR1. Inhibitors of DNA methyltransferase (5-azacytidine [5aC]) and histone deacetylase (trichostatin A [TSA]) were used to examine gene transcription, promoter methylation status, and the chromatin determinants associated with the MDR1 promoter. We have established that methyl-CpG binding protein 2 (MeCP2) is involved in methylation-dependent silencing of human MDR1 in cells that lack the known transcriptional repressors MBD2 and MBD3. In the repressed state the MDR1 promoter is methylated and assembled into chromatin enriched with MeCP2 and deacetylated histone. TSA induced significant acetylation of histones H3 and H4 but did not activate transcription. 5aC induced DNA demethylation, leading to the release of MeCP2, promoter acetylation, and partial relief of repression. MDR1 expression was significantly increased following combined 5aC and TSA treatments. Inhibition of histone deacetylase is not an overriding mechanism in the reactivation of methylated MDR1. Our results provide us with a clearer understanding of the molecular mechanism necessary for repression of MDR1.
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PMID:Precipitous release of methyl-CpG binding protein 2 and histone deacetylase 1 from the methylated human multidrug resistance gene (MDR1) on activation. 1186 62

Resistance to the cytotoxic actions of antineoplastic drugs, whether intrinsic or acquired, remains a barrier to the establishment of curative chemotherapy regimens for advanced breast cancer. Over-expression of P-glycoprotein (P-gp), encoded by the MDR1 gene and known to mediate resistance to many antineoplastic drugs, may contribute to poor breast cancer treatment outcome. Nonetheless, the precise molecular mechanisms responsible for high or low level P-gp expression in breast cancer cells have not been established. We assessed the role of DNA hypermethylation near the MDR1 transcriptional regulatory region in MDR1 expression in MCF-7 breast cancer cells, which fail to express MDR1 mRNA, and MCF-7/ADR cells, known to express high MDR1 mRNA levels. When compared to MCF-7/ADR cells, MCF-7 cells manifested markedly diminished MDR1 transcription rates by nuclear run-off assay, but equivalent MDR1 promoter trans-activation activity in transient transfection experiments, indicating that cis factors were most likely responsible for the differences in MDR1 transcription between MCF-7/ADR cells and MCF-7 cells. Bisulfite genomic sequencing analyses revealed substantially less extensive MDR1 promoter methylation in MCF-7/ADR cells than in MCF-7 cells, suggesting that CpG dinucleotide methylation might contribute to the observed MDR1 transcription differences. Chromatin immunoprecipitation analyses indicated an inactive MDR1 chromatin conformation in MCF-7 cells, with a paucity of acetylated histones and the presence of 5-mC-binding proteins MeCP2 and MBD2, and an active MDR1 chromatin conformation in MCF-7/ADR cells, with an abundance of acetylated histones and the presence of the transcriptional trans-activator YB-1. Stable MCF-7 sublines which had been treated with the DNA methyltransferase inhibitor 5-azacytidine, exhibited a reduction in MDR1 promoter methylation and a complex MDR1 chromatin configuration, characterized by the simultaneous presence of transcriptional activators and repressors. In this state, MDR1 expression was markedly sensitive to treatment with the histone deacetylase inhibitor trichostatin A.
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PMID:MDR1 promoter hypermethylation in MCF-7 human breast cancer cells: changes in chromatin structure induced by treatment with 5-Aza-cytidine. 1525 26

The resistance of cancer cells to chemotherapeutic agents is a major clinical problem and an important cause of treatment failure in cancer. Mechanisms that have developed to guard cancer cells against anti-cancer drugs are major barriers to successful anti-cancer therapy. Therefore, the identification of novel mechanisms of cellular resistance holds the promise of leading to better treatments for cancer patients. In the present study, we used human MCF-7 breast adenocarcinoma cell line and its doxorubicin-resistant variant MCF-7/R to determine the role of alterations of DNA methylation of chemoresitance-related genes, such as multidrug resistance 1 (MDR1), glutathione-S-transferase (GSTpi), O(6)-methylguanine DNA methyltransferase (MGMT), and urokinase (Upa), in the development of drug resistance. The promoter regions of MDR1, GSTpi, MGMT, and Upa genes were highly methylated in MCF-7 cell line but not in its MCF-7/R drug resistant variant. The hypomethylated status of MDR1 gene was associated with overexpression of P-glycoprotein. We hypothesize that acquirement of doxorubicin resistance of MCF-7 cells is associated with DNA hypomethylation of the promoter regions of the MDR1, GSTpi, MGMT, and Upa genes.
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PMID:Role of DNA hypomethylation in the development of the resistance to doxorubicin in human MCF-7 breast adenocarcinoma cells. 1635 34

Information of genome structure with its size variation may provide important clues for evolutionary processes at lower taxon level in eukaryotes. Here, we analyzed the compact genome structure of the monogonont rotifer, Brachionus koreanus in the light of transphyletic genome comparison and economic genome usage. To confirm the genome compactness of B. koreanus, we compared the genomic structure of several selected genes with those of human and pufferfish. For example, one of the large genes, DNA-dependent protein kinase (DNA-PK) with dimeric protein Ku70 and Ku80, showed high similarity, even though genomic DNA lengths were quite different. The replication protein As (RPAs) as a heterotrimeric protein also showed a compact genomic structure including all the essential domains and motifs in B. koreanus. Regarding transmembrane protein-containing genes, the B. koreanus P-glycoprotein (P-gp) showed exactly the same topology of the TM domain compared to those of human and pufferfish, even though it had a compact genome structure. In addition, the gene structure of an inducible repair enzyme O(6)-methylguanine DNA methyltransferase (O(6)-MGMT) of B. koreanus showed the highest compactness among the genes tested. The objective of this report is to evaluate the potential for whole genome sequencing and functional genomic research using the monogonont rotifer B. koreanus as a non-model organism that plays important roles in aquatic food-webs. Subsequently, we discussed possible reasons for compact genome structures as well as small and fewer introns from several perspectives. We conclude that the small size genome of B. koreanus would make this species potentially useful for comparative genome structure analysis of non-model species through whole genome sequencing and genetic mapping.
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PMID:Genomic organization of selected genes in the small monogonont rotifer, Brachionus koreanus. 2267 67


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