Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:5.99.1.3 (topoisomerase)
 9,911 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In a P-glycoprotein-negative cell line, GLC4-Adr90, a 75-fold acquired Adriamycin (Adr) resistance coincided with a reduced cellular Adr level, an increased detoxifying capacity (glutathione (GSH) and glutathione S-transferase (GST) elevated), and a reduced topoisomerase-II (topo-II) activity compared with the parent cell line GLC4. The effect on Adr resistance of buthionine sulfoximine (BSO, GSH synthesis inhibitor), was studied alone or in combination with verapamil (drug-efflux inhibitor), docosahexaenoic acid (membrane lipid domain affector), ethacrynic acid (GST inhibitor), aphidicolin (DNA-polymerase-alpha inhibitor) or novobiocin (NOV, topo-II inhibitor). Cytotoxicity was tested using a microculture tetrazolium assay. In GLC4-Adr90, BSO and NOV increased Adr-induced cytotoxicity 12.9-fold and 1.8-fold respectively. The combination of BSO plus NOV showed an additive effect, decreasing the Adr resistance factor from 75 to 2.7. Combination of modulators of Adr resistance directed at different resistance mechanisms appears promising in vitro.
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PMID:Combined in vitro modulation of adriamycin resistance. 168 Aug 15

The cyanomorpholino derivative of doxorubicin (MRA-CN) is a DNA intercalator and alkylator that is a highly potent cytotoxin, non-cross-resistant in multidrug-resistant cells, and noncardiotoxic in comparison with doxorubicin. To further examine mechanisms of action and resistance to MRA-CN, a cell line resistant to MRA-CN, ES-2R, was established by growing a human ovarian carcinoma cell line, ES-2, in increasing concentrations of the drug. The resistant subline was 4-fold resistant to MRA-CN and cross-resistant to other DNA cross-linking agents, cisplatin (7-fold) and carmustine (3-fold), as well as to the DNA strand-breaking agents etoposide (6-fold), doxorubicin (2-fold), bleomycin (5-fold), and ionizing radiation (2-fold). In contrast, ES-2R cells were not cross-resistant to vinblastine. Several months of additional growth of ES-2R cells in MRA-CN did not yield higher, stable levels of drug resistance. A low level of P-glycoprotein was detectable in the ES-2R cells. However, the extent of intracellular accumulation of [3H]MRA-CN by this resistant cell line was identical to that of the sensitive line. The number of DNA cross-links formed by cisplatin in ES-2R was only 50% of that of the ES-2 cells and was associated with a 50% increase in the rate of repair of these cross-links in the resistant cells. Ionizing radiation induced similar amounts of single- and double-strand breaks in the ES-2 line as well as in the ES-2R cells. There was no apparent difference between the two cell lines in the rate and extent of repair of these DNA breaks. Thus, enhanced DNA repair cannot explain the phenomenon of cross-resistance to radiation. Comparisons of glutathione (GSH) content and the enzymes involved in GSH homeostasis showed significant differences. Resistant cells contained 1.5-fold more GSH, a 2.2-fold increase in gamma-glutamyltranspeptidase activity, and a 2.4-fold increase in GSH reductase compared with ES-2 cells (all P less than 0.05). Total glutathione-S-transferase (GST) activity was 2.6-fold higher (P less than 0.01) in the ES-2R line. The pi-class GST subunit by Western blotting and GST activity toward ethacrynic acid were increased 2-fold in the resistant cells. Depletion of GSH levels in ES-2R cells by buthionine sulfoximine restored the sensitivity of ES-2R to MRA-CN. These findings implicate a role for GSH metabolism in the resistance phenotype of ES-2R cells. We have previously reported that these cells have an increased generation time and decreased topoisomerase II content. Thus, the ES-2R cell line exhibits a complex phenotype of broad cross-resistance, which is likely to involve multiple mechanisms, and includes enhanced DNA repair and increased GSH content and GST activity.
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PMID:Multifactorial mechanisms associated with broad cross-resistance of ovarian carcinoma cells selected by cyanomorpholino doxorubicin. 171 40

After twenty years, understanding the mechanisms of tumor cells kill by anthracyclines still remains an active area of research. Of many mechanisms described for this class of drugs, efforts in the last year have focused on defining the role of free radical formation, topoisomerase II-induced DNA breakage, and P-170-dependent cellular accumulation of anthracyclines in tumor cell kill and resistance. First, in a number of tumor cell lines, the formation of free radical species from anthracyclines has been implicated in the cell killing. Modulation of detoxification pathways in a drug-resistant cell line e.g depletion of GSH, a substrate for peroxidase and transferase, enhanced both the formation of oxy-radicals and adriamycin cytotoxicity. It should be noted, however, that these findings are not true for every cell line examined, and free radical-mediated tumor kill may be cell- or tissue-specific. Second, anthracyclines-mediated topo II-dependent DNA cleavage was observed in most cell lines and reduced breaks were found in resistant cells. The decrease in single-strand breaks, however, neither correlated with the degree of resistance nor with differences in the relative topo II activity, which was in most cases only two-fold less in resistant cells than in sensitive cells. Finally, the reduced accumulation of the drug does not appear to be the only contributing factor in multidrug resistant cells and P-170 is not the only protein overexpressed in certain cells, e.g., an 85,000 Da protein may also be linked to adriamycin resistance. Although GST protein is overexpressed in most adriamycin resistant cells along with mdr1 gene, current evidence suggests that this protein may not be directly involved in adriamycin resistance. Taken together, both the mechanism of action and resistance to this class of drug likely vary among cell lines. Clinical studies in the past year have brought about interesting refinements in anthracycline-containing chemotherapy; ICRF-187 (by itself also cytotoxic) seems to offer protection against cardiac toxicity, while implicating iron in the mediation of cardiac damage. Out of a large number of newer anthracycline derivatives, clinical evidence indicates only a modest increase in therapeutic index with a few analogs, perhaps idarubicin and epirubicin. It is not yet clear that being able to receive more milligrams (or more cycles) of anthracycline eventually translates into a significantly better response rate or in a survival advantage. Much less clear is whether patients refractory to adriamycin may derive any benefit from newer anthracyclines.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Anthracyclines. 222 2

Chronic lymphocytic leukemia is a neoplastic disease in which drug resistance invariably occurs. We have studied the uptake and interaction with molecular targets of two drugs, chlorambucil and adriamycin, in CLL lymphocytes and CHO cell lines. Resistance does not appear related to uptake for either drug. Exposure to CLB causes DNA cross-links in the sensitive but not in the resistant cell line. The GSH content of B-CLL lymphocytes is depleted after a 20-hr incubation. An inability to maintain its GSH content may contribute to this cell's vulnerability to CLB. The resistance of CLL lymphocytes to ADR may be related to the undetectable levels of its target enzyme DNA topoisomerase II. Future approaches may involve study of novel anthracyclines, DNA topoisomerase I inhibitors and the development of in vitro predictive tests.
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PMID:Studies on drug resistance in chronic lymphocytic leukemia. 256 Dec 48

Exposure to benzene, a human and animal carcinogen, results in the formation of structural chromosomal aberrations in the bone marrow and blood cells of animals and humans. The mechanisms underlying these clastogenic effects are unknown. Inhibition of enzymes involved in DNA replication and repair, such as topoisomerase enzymes, by the metabolites of benzene represents a potential mechanism for the formation of chromosomal aberrations. To test this hypothesis, the inhibitory effects of various phenolic and quinone metabolites of benzene on the activity of human topoisomerases I and II were studied in vitro. No inhibition of topoisomerase I was seen with any of the tested metabolites. Inhibitory effects on topoisomerase II were not observed for hydroquinone, phenol, 2,2'-biphenol, 4,4'-biphenol and catechol at concentrations as high as 500 microM. 1,4-Benzoquinone and 1,2,4-benzenetriol inhibited topoisomerase II at relatively high 500 and 250 microM concentrations, respectively. However following bioactivation using a peroxidase/H2O2 system, inhibitory effects were seen at concentrations as low as 50 microM for both phenol and 2,2'-biphenol and 10 microM for 4,4'-biphenol. The addition of reduced glutathione (GSH) to the 4,4'-biphenol and horseradish peroxidase reaction system protected topoisomerase II from inhibition suggesting that diphenoquinone or another oxidation product formed from 4,4'-biphenol might be the reactive species. These in vitro results indicate that inhibition of topoisomerase II may contribute to the clastogenic and carcinogenic effects of benzene. In addition, metabolites formed from these phenolic compounds appear to represent several new types of topoisomerase II-inhibiting compounds.
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PMID:Topoisomerase inhibition by phenolic metabolites: a potential mechanism for benzene's clastogenic effects. 758 26

Resistance may limit the clinical usefulness of a variety of chemotherapeutic drugs including mitomycin C (MMC). The MMC-sensitive HT-29 colon cancer cell line and its MMC-resistant subline, HT-29R13, were studied in vitro under aerobic conditions to help characterize the mechanisms associated with MMC resistance. HT-29R13 cells exhibit approximately 2-fold resistance to MMC compared with HT-29 cells and lack the typical multidrug-resistance pattern; resistance is stable in the absence of drug exposure. Levels of glutathione (GSH) and total glutathione-S-transferase (GST) activity were not different between the two cell lines; however, levels of GSH reductase and GSH peroxidase were increased significantly in HT-29R13. Although total GST activity was unchanged, GST-pi and GST-alpha isoenzyme expression as measured using western blot were increased significantly in HT-29R13 compared with HT-29. DT-diaphorase levels and topoisomerase II activity were decreased significantly in HT-29R13. Both cell lines had equal P-glycoprotein expression. Multiple drug resistance mechanisms are present in HT-29R13 including decreased drug activation (decreased DT-diaphorase), increased drug detoxification (increased GST-pi and GST-alpha, GSH reductase, GSH peroxidase), and decreased accessibility of DNA targets (decreased topoisomerase II). Further work will be necessary to determine the degree to which each of these mechanisms contribute to MMC resistance in this model.
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PMID:Biochemical characterization of a mitomycin C resistant colon cancer cell line variant. 790 34

We have reported the establishment of a mitomycin-C (MMC)-resistant non-small-cell lung-cancer cell line, PC-9/MC4. As determined by an MTT assay, this resistant cell line was found to be 4 times more sensitive to adriamycin (ADM) than was the parental PC-9. There were no significant differences in sensitivity to etoposide, mitoxantrone, daunomycin, epirubicin, pirarubicin, 9-aminoanthracycline or 3'-deamino-3'-morpholino-13-deoxo-10-hydroxy carminomycin. These data suggest that neither qualitative or quantitative changes in DNA topoisomerase II nor the enhanced repair of DNA can explain the differing sensitivity to ADM observed. No significant differences were found in the accumulation of ADM and glutathione (GSH) in these cell lines. Although total glutathione-S-transferase (GST) activity in PC-9/MC4 cells was lower than that observed in PC-9 cells and treatment with ethacrynic acid (EA) reduced sensitivity to ADM in both cell lines, relative resistance was unaffected. NADH-cytochrome b5 reductase (B5R) activity in PC-9/MC4 cells showed a 3-fold greater decrease than that in PC-9 cells, and DT-diaphorase (DTD) activity in PC-9/MC4 cells showed an approximately 200-fold greater decrease than that in PC-9 cells. Addition of dicumarol, an inhibitor of DTD, decreased the sensitivity of ADM of PC-9 but not of PC-9/MC4. DTD activity in the PC-9 cell line was inhibited by treatment with dicumarol while in PC-9/MC4 it remained unchanged. These data suggest that DT-diaphorase is a determinant of sensitivity to ADM in the 2 cell lines.
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PMID:DT-diaphorase as a determinant of sensitivity to adriamycin in non-small-cell lung-cancer cell lines. 792 20

This study describes characteristics of a human bladder cancer cell line J82/MMC that is 6-fold more resistant to mitomycin C (MMC) than the parental cells. The J82/MMC subline was isolated by repeated continuous exposures of the J82/WT cells to increasing concentrations of MMC. The J82/MMC cell line showed (1) collateral sensitivity to taxol, 5-FU and topoisomerase II inhibitors; and (2) cross-resistance to cisplatin, melphalan and MMC analogues BMY 25282 and BMY 25067. Levels of two key MMC activation enzymes, NADPH cytochrome P450 reductase and DT-diaphorase, were significantly lower in J82/MMC cells compared with J82/WT, suggesting that lower sensitivity of J82/MMC cells to MMC may result from deficient drug activation. Further support is indicated by: 1) reduction in the differential in toxicity between the 2 cell lines by BMY 25282; and 2) a higher effect of DT-diaphorase inhibitor dicumarol on the wild-type cells compared with J82/MMC. Although glutathione (GSH) levels did not differ in these cells, a small but significant increase in GSH transferase (GST) activity was noticed in J82/MMC cells. GST inhibitor ethacrynic acid significantly enhanced MMC cytotoxicity in the J82/MMC cell line. A small but significant increase in the level of anti-oxidative enzyme catalase, but not GSH peroxidase, was also observed in J82/MMC cell line compared with J82/WT. Thus, the possibility that relatively lower sensitivity of J82/MMC cells to MMC may result from reduced oxygen radical generation cannot be ruled out. MMC-induced DNA interstrand cross-linking was markedly lower in the J82/MMC cell line compared with J82/WT. Our results suggest that the MMC resistance in the J82/MMC cell line may be multifactorial.
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PMID:Characterization of a human bladder cancer cell line selected for resistance to mitomycin C. 807 54

The effect of lowering intracellular glutathione (GSH) concentrations on the toxicity of alkylating agents, and RNA synthesis inhibitor and topoisomerase 1 and 2 inhibitors to a number of human leukaemic cell lines were evaluated. By using the GSH synthesis inhibitor DL-buthionine-(S,R)-sulfoximine (BSO), GSH levels were artificially reduced. Cells with low GSH concentrations were exposed to a number of cytotoxic agents and the resultant mode of cell death was analysed using morphological and biochemical criteria. It was found that untreated cells exposed to the above drugs underwent apoptosis to varying extents. However, the toxicity of alkylating agents was dramatically increased to all cell lines on lowering GSH levels, with the mode of cell death switching from apoptosis to necrosis. The reduction of GSH levels had no effect on the toxicity of actinomycin-D, camptothecin or etoposide, nor did it affect the mode of cell death induced by these agents. These observations suggest that modulation of GSH levels effect the toxicity of alkylating agents and that GSH influences the mode of cell death induced by alkylating agents.
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PMID:Apoptosis or necrosis: intracellular levels of glutathione influence mode of cell death. 808 Apr 40

We have previously obtained, by exposure to near continuous increasing concentrations of cisplatin, a panel of human ovarian cancer cell lines that exhibit a wide range of primary resistance to the drug (9- to > 400-fold). These cells had strikingly increased (4- to 50-fold) levels of glutathione (GSH) as compared with the drug-sensitive cells of origin (A. K. Godwin et al., Proc. Natl. Acad. Sci. USA, 89: 3070-3074, 1992). Utilizing this panel of resistant cell lines, we evaluated cross-resistance to classical alkylating agents, natural product drugs, and irradiation. We observed that cross-resistance to carboplatin paralleled that of cisplatin, culminating in approximately 250-fold resistance. Similarly, melphalan cross-resistance continued to increase to > 400-fold and again paralleled the primary cisplatin resistance. Cell lines with low to very high levels of resistance to cisplatin are 8- to 850-fold resistant to the epipodophyllotoxin derivative etoposide. Cross-resistance is also observed for other natural product drugs, including Adriamycin (approximately 80-fold), mitoxantrone (approximately 440-fold), and taxol (approximately 40-fold). Cross-resistance to irradiation is, however, modest (< 2-fold). The cells with the greatest primary resistance to cisplatin most commonly had the highest cross-resistance to the other drugs examined. The cross-resistance to the natural product category drugs was found not to be mediated by the products of either the multidrug resistance 1 (MDR1) or multidrug resistance-associated protein (MRP) genes based on lack of coordinate increased expression or amplification of these genes as assessed by Northern and Southern blot analyses. Furthermore, verapamil failed to markedly increase drug sensitivity. Although there was no indication that these natural product drug efflux pumps were operative, we observed decreased doxorubicin accumulation in these cell lines cross-resistant to natural products. In addition, alternations in DNA topoisomerase II mRNA levels, which have been observed in a variety of human tumor cell lines selected in vitro for resistance to etoposide or teniposide, were not detected. Only intracellular levels of GSH correlated with cross-resistance to these diverse anticancer agents and partial loss of resistance was associated with a marked decrease in glutathione levels. In the absence of alternative mechanisms, we speculate that the very broad clinically relevant cross-resistance seen in this model system may, at least in part, be the direct result of GSH-mediated drug inactivation or may be due to a combination of GSH conjugation to drug and conjugate efflux mediated by the putative ATP-dependent glutathione S-conjugate export pump.
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PMID:Cross-resistance to diverse drugs is associated with primary cisplatin resistance in ovarian cancer cell lines. 810 43


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