Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:5.99.1.2 (topoisomerase)
 9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The biochemical bases of the multidrug-resistant (MDR) phenotype were investigated in drug-resistant sublines derived from LoVo human colon carcinoma cell lines by doxorubicin (DOX) and teniposide (VM26) selection. In addition to P-glycoprotein-mediated drug extrusion through the plasma-membrane, LoVo MDR cells display a further drug-resistance mechanism. That is, to achieve equitoxic effects, LoVo MDR sublines require much higher intracellular drug concentrations than those required by LoVo drug-sensitive parent cell line. Involvement of mdr1, topoisomerase II and glutathione-S-transferase-pi (GST-pi) drug-resistance systems in intracellular drug resistance was investigated. Pharmacologic and biochemical data indicated that intracellular drug resistance in LoVo MDR sublines is uniquely consequent to the drug-transporting property of intracytoplasmic membrane-bound P-glycoprotein molecules which compartment drugs in vacuole-like structures.
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PMID:P-glycoprotein but not topoisomerase II and glutathione-S-transferase-pi accounts for enhanced intracellular drug-resistance in LoVo MDR human cell lines. 135 86

The modulating effect on drug resistance of amiodarone (AM) and its metabolite desethylamiodarone (DEA) was studied in a P-glycoprotein-positive human colon carcinoma cell line COLO 320, and a human small-cell lung carcinoma cell line GLC4 and its adriamycin (Adr)-resistant subline GLC4-Adr (both P-glycoprotein-negative). AM, DEA and verapamil induced an increase in cytotoxicity of Adr, vincristine and etoposide (VP16) in COLO 320 cells, while in the GLC4 and GLC4-Adr cell line no effect was seen. In the COLO 320 cell line, AM caused more intracellular, and especially intranuclear, fluorescence of Adr and more Adr-induced DNA strand breaks as compared to Adr alone. Moreover, an increase in VP16-induced topoisomerase II-DNA complexes was observed when AM was added. Competition between AM and Adr for the same efflux pump was suggested in efflux studies. The colony-forming unit granulocyte macrophage (CFU-GM) assay showed no increase in cytotoxicity of Adr when AM was added. Fourteen patients with Adr-resistant tumors were treated with Adr and AM. In these patients, peak serum levels of AM plus DEA of 10 microM were reached. Patient serum (20%) obtained after the first i.v. AM infusion induced in vitro significantly more cell kill of Adr in COLO 320 cells. Apart from a transient first-degree AV block in one patient, no cardiac toxicity was observed with the combination of Adr and AM. Bone-marrow toxicity was the same as expected from Adr alone in these patients. One of the 13 evaluable patients obtained a partial remission.
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PMID:In vitro and in vivo modulation of multi-drug resistance with amiodarone. 164 80

Although the stabilization of topoisomerase II cleavable complexes by etoposide (VP-16) has been recognized to be important for cell killing, the lethal events following the formation of cleavable complexes remain to be elucidated. In an attempt to characterize the biochemical requirements for VP-16-induced cytotoxicity, we examined the effects of calcium depletion in Chinese hamster DC3F cells. Four-hour preincubation in calcium-free medium or in complete medium containing 5 mM [ethylenebis(oxyethylenenitrilo)]tetraacetic acid (EGTA) protected against the cytotoxicity of VP-16. Under these same conditions, the VP-16-induced DNA single-strand break frequency in calcium-depleted cells remained similar to that of control cells. Cell-cycle analysis and thymidine pulse incorporation indicated that calcium depletion did not alter DNA synthesis and cell cycle distribution. Drug-induced cytotoxicity was restored progressively within 4-8 hr after calcium-depleted cells were refed with calcium-containing medium. Calcium depletion also protected against the cytotoxicity of camptothecin, hyperthermia and, to a lesser extent, nitrogen mustard and gamma radiation in DC3F cells. Similar results were obtained in human colon carcinoma HT-29 cells. Our results suggest that topoisomerase II-mediated DNA breaks are only potentially lethal and that calcium-dependent cellular processes are required for the cytotoxicity of topoisomerase inhibitors.
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PMID:Cell death induced by topoisomerase inhibitors. Role of calcium in mammalian cells. 164 24

A series of doxorubicin-resistant variants of the human LS174T colon carcinoma cell line was generated by stepwise selection. These variants also exhibited increased resistance to vinblastine, etoposide, cis-platinum, and melphalan, suggesting that resistance was multifactorial. The parental LS174T cell line and 3 resistant variants were examined for over-expression of P-glycoprotein, changes in total cellular glutathione content, and the level of topoisomerase-II expression. Changes in all of these parameters were observed in the doxorubicin-selectants, along with a marked shift in the intracellular distribution of doxorubicin. P-glycoprotein RNA and protein levels were increased 2- to 3-fold in the resistant variants, while total glutathione levels increased 1.4- to 2.1-fold. Treatment with DL-buthionine-[S,R]-sulfoximine, an inhibitor of glutathione biosynthesis, was able to reverse resistance to cis-platinum and melphalan in these variants, but had little effect on doxorubicin resistance. Immunoblot analysis of cell extracts indicated that the level of DNA topoisomerase II (EC 5.99.1.3) in the doxorubicin-resistant LS174T cells was decreased by approximately 50% compared with the parental cell line. Doxorubicin was mainly localized to the cytoplasm in resistant cells, while in the parent line it was mostly found in the nucleus. This constellation of changes suggests that selection with doxorubicin activated several mechanisms of resistance involving drug transport, metabolism, and ability to reach nuclear target sites.
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PMID:Multifactorial resistance in LS174T human colon carcinoma cells selected with doxorubicin. 168 Aug 16

Stable acquired resistance to etoposide (VP-16) or teniposide (VM-26) in HCT116 human colon carcinoma cells and A549 human lung adenocarcinoma cells, was previously obtained by weekly 1-h exposures to either drug (B. H. Long, Natl. Cancer Inst. Monogr., 4: 123-127, 1987). The purpose of this study was to identify possible mechanisms of resistance present in these cells by using human mdr1 and topoisomerase II DNA probes, antibodies to these gene products, and P4 phage unknotting assay for topoisomerase II activities. HCT116(VP)35 cells were 9-, 7-, and 6-fold resistant to VP-16, VM-26, and Adriamycin, respectively, and showed no cross-resistance to colchicine and actinomycin D. These cells had no differences in mdr1 gene, mdr1 mRNA, or P-glycoprotein levels but displayed decreased levels of topoisomerase II mRNA and enzyme activity without any alteration of drug sensitivity displayed by the enzyme. HCT116(VM)34 cells were 5-, 7-, and 21-fold resistant to VP-16, VM-26, and Adriamycin; were cross-resistant to colchicine (7-fold) and actinomycin D (18-fold); and possessed a 9-fold increase in mdr1 mRNA and increased P-glycoprotein without evidence of mdr1 gene amplification. No alterations in topoisomerase II gene or mRNA levels, enzyme activity, or drug sensitivity were observed. A549(VP)28 and A549(VM)28 cells were 8-fold resistant to VP-16 and VM-26 and 3-fold resistant to Adriamycin. Both lines were not cross-resistant to colchicine or actinomycin D but were hypersensitive to cis-platinum. No alterations in mdr1 gene, mdr1 mRNA, or P-glycoprotein levels, but lower topoisomerase II mRNA levels and decreased enzyme activities, were observed. Of the four acquired resistant cell lines, resistance is likely related to elevated mdr1 expression in one line and to decreased topoisomerase II expression in the other three lines.
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PMID:Mechanisms of resistance to etoposide and teniposide in acquired resistant human colon and lung carcinoma cell lines. 171 44

We had previously shown that 10,11-methylenedioxy-20-(RS)-camptothecin (MDO-CPT) is a more potent inhibitor of purified DNA topoisomerase I than 20-(S)-camptothecin (CPT). The current studies compared the cytotoxicity and DNA damage induced by MDO-CPT and CPT in the human colon carcinoma cell line, HT-29. MDO-CPT was 7- to 10-fold more potent than CPT both for cytotoxicity (ID50 = 25 vs. 180 nM) and production of DNA single-strand breaks (SSB). Kinetics of SSB formation and reversal were similar for MDO-CPT and CPT. DNA-protein crosslinks (DPC) were also produced by both drugs with a SSB/DPC ratio of 1/1. Moreover, no SSB were detected under non-deproteinizing conditions, indicating that both CPT and MDO-CPT produced protein-linked DNA single-strand breaks. A good correlation between cytotoxic potency and protein-linked DNA single-strand break production was observed for CPT and MDO-CPT, implying a causal relationship between drug-induced cytotoxicity and topoisomerase I inhibition. The sensitivity of human colon HT-29 cancer cells to camptothecins may be a selective phenomenon since these cells normally express natural resistance to current chemotherapeutic drugs, including topoisomerase II inhibitors.
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PMID:10,11-Methylenedioxycamptothecin, a topoisomerase I inhibitor of increased potency: DNA damage and correlation to cytotoxicity in human colon carcinoma (HT-29) cells. 217 90

A series of analogs based on a novel template, 11-aza-(20S)-camptothecin, were obtained from total synthesis and tested as potential anticancer drugs in the topoisomerase I enzyme cleavable complex assay. The parent compound 11-aza-(20S)-camptothecin (8) was derived from a Friedlander condensation between the known aminopyridine derivative 3-(3-amino-4-picolylidene)-p-toluidine and optically active tricyclic ketone 7. Compound 8 had activity approximately twice that of (20S)-camptothecin in the calf thymus topoisomerase I cleavable complex assay. Compounds were prepared wherein the 11-aza nitrogen atom was quaternized as either the corresponding N-oxide or methyl iodide. Compounds with quaternized N-11 showed improved water solubility and were equipotent to the clinically investigated camptothecin analog topotecan in the cleavable complex assay. These compounds were evaluated in vivo in nude mice bearing HT-29 human colon carcinoma xenografts. The analog 11-aza-(20S)-camptothecin 11-N-oxide was found to significantly retard tumor growth when compared to untreated controls. Finally, 7,10-disubstituted 11-azacamptothecin analogs were synthesized using Pd(0) coupling reactions of 10-bromo-7-alkyl-11-aza-(20S)-camptothecins 19 and 20, which in turn were available from a Friedlander condensation of the novel bromopyridine derivatives 17a and 17b with 7. Among the 10-substituted series, a number of analogs displayed extremely high in vitro potency against topoisomerase I and improved aqueous solubility. A significant number of the compounds were found to be active in whole cell cytotoxicity assays and several were evaluated in nude mice bearing the HT-29 tumor xenografts. The most effective of these proved to be (S)-11-aza-7-ethyl-10-(aminohydroximinomethyl)camptothecin trifluoracetic acid salt (27), a potent topoisomerase I inhibitor which demonstrated excellent efficacy in both short term and in extended in vivo assays. A comparison between in vitro enzyme data and in vivo data from nude mouse studies in other compounds in this series revealed a poor overall correlation between topoisomerase inhibition in vitro and antitumor efficacy in vivo.
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PMID:Synthesis, topoisomerase I inhibitory activity, and in vivo evaluation of 11-azacamptothecin analogs. 770 14

Cisplatin (CDDP) resistance mechanisms were studied in a model of three germ cell tumour and three colon carcinoma cell lines representing intrinsically CDDP-sensitive and -resistant tumours respectively. The CDDP sensitivity of the cell lines mimicked the clinical situation. The glutathione levels of the cell lines correlated with CDDP concentrations inhibiting cell survival by 50% (IC50); total cellular sulphydryl content (TSH) was unexpectedly inversely correlated with IC50. IC50 correlated neither with glutathione S-transferase (GST) nor with GST pi expression, topoisomerase I or II activity. Immediately after 4 h incubation with CDDP, platinum (Pt) accumulation and Pt bound to DNA were not correlated, but after another 24 h drug-free culture, Pt binding to DNA in germ cell tumour but not in colon carcinoma cell lines correlated with IC50. With the exception of in vitro sensitivity and TSH, none of the parameters studied discriminated between the two groups of cell lines. Correction of CDDP sensitivity parameters for phenotypical differences did not influence statistical correlations. Analysis of variance revealed a correlation between IC50 and the combination of glutathione, GST activity and Pt bound to DNA. But at other CDDP cytotoxicity levels sensitivity was also correlated with Pt accumulation, topoisomerase II activity and TSH in various combinations. This model of intrinsic CDDP resistance showed that multiple parameters ought to be studied to explain CDDP resistance, but did not elucidate the cause of the unique sensitivity of germ cell carcinoma, although the unexpected values of TSH deserve further attention.
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PMID:Cellular basis for differential sensitivity to cisplatin in human germ cell tumour and colon carcinoma cell lines. 771 Sep 29

To study the mechanisms of the acute induction of drug resistance in cancer cells, we have established a model system in which adriamycin (ADM) induces immediate drug resistance. In this system, human colon carcinoma HT-29 cells were pretreated for 1 h with a subtoxic dose of ADM (0.3 micrograms/ml) and incubated for 24 h in drug-free medium. Then the cells were treated for 1 h with ADM, and the cell survival was determined in terms of colony-forming ability. The survival of the pretreated cells was increased up to 100-fold, as compared with that of untreated cells. Such increased survival, however, was observed only after high doses of ADM (2 to 8 micrograms/ml); more than 99% of the cells were killed. These results indicate that only a small fraction of ADM-pretreated cells acquire the ADM-resistant phenotype. Similar induced resistance was observed in five of seven subclones isolated from HT-29 cells by limiting dilution, suggesting that the majority of cells in the parental HT-29 population could acquire the ADM-resistant phenotype. In the subclone HT-29T9, the ADM pretreatment induced concomitant resistance to daunomycin, VP-16, and VM-26 but not to agents other than topoisomerase II inhibitors. The ADM-induced drug resistance did not accompany MDR1 gene expression and could not be overcome by verapamil, a P-glycoprotein inhibitor. The present system could be useful to study the acute induction mechanism(s) of ADM-resistance, which could be relevant to clinical resistance in patients.
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PMID:Acute induction of adriamycin-resistance in human colon carcinoma HT-29 cells exposed to a sublethal dose of adriamycin. 773 Jan 48

The in vitro cytotoxic effects of docetaxel (Taxotere; RP56976, NSC688503) proved both time and concentration dependent. Amongst thirteen human cell lines from various tumor types, exposure to increasing concentrations of docetaxel over 24 hrs resulted in a plateau-shaped dose response curve, suggesting that increased cell kill becomes more dependent on increased exposure duration than on concentration. IC50 concentrations (reducing survival by 50%) ranged from 0.13-3.3 ng/ml, with three neuroblastoma lines proving most sensitive and three breast and two colon carcinoma lines showing least sensitivity. There was significant cross-resistance to docetaxel in the classic multidrug resistant (MDR) Chinese hamster ovarian (CHO) CHRC5 line and the human lymphoblastoid CCRF-CEMVLB1000 line, as well as in two vincristine (VCR)-selected MDR MCF-7 sublines. All four of these MDR sublines overexpress P-glycoprotein (Pgp), as did a 6-fold docetaxel-selected resistant CHO subline. As an apparent corollary, in two human teratoma lines selected for etoposide resistance and showing some cross-resistance to VCR and in two CHO sublines expressing low levels of VCR resistance, yet all proving Pgp positive, no docetaxel cross-resistance was identified. Verapamil modulated docetaxel resistance only in sublines expressing resistance to the drug and overexpressing Pgp. Four other human tumor sublines selected for resistance to 5-fluorouracil, cisplatin or teniposide, showed a lack of cross-resistance to docetaxel. Furthermore, cross-resistance to docetaxel was not apparant in four epipodophyllotoxin-selected resistant sublines with alterations in topoisomerase II, indicating its effectiveness against tumor cells expressing the topoisomerase II-related MDR phenotype. Our observation that docetaxel cross-resistance was not automatically expressed by classic MDR tumour cells appears of interest and of potential clinical relevance.
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PMID:Differential cytotoxic effects of docetaxel in a range of mammalian tumor cell lines and certain drug resistant sublines in vitro. 789 35


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