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)

Antifolates have been shown to increase the DNA strand breaks produced by the topoisomerase inhibitor etoposide. PT523 is a potent new antifolate that cannot be polyglutamated. Human SCC-25 squamous carcinoma cells were exposed to methotrexate, trimetrexate or PT523 at a concentration of 5 microM for 24 h along with various concentrations of etoposide or novobiocin during the final 2 h. Isobologram analysis of the treatment combinations indicated that exposure of the cells to PT523/etoposide, methotrexate/etoposide, PT523/novobiocin, methotrexate/novobiocin and trimetrexate/novobiocin resulted in greater than additive cytotoxicity. DNA alkaline elution studies with the same drug combinations indicated that there were three- to four-fold increases in the radiation equivalent (rad equivalent) strand breaks in the cellular DNA with etoposide or novobiocin along with the antifolate compared with the topoisomerase II inhibitors alone. Tumor growth delay studies were carried out in the murine SCC VII squamous carcinoma. PT523 (0.5 mg/kg) and methotrexate (2 mg/kg) were administered by 7-day continuous infusion while trimetrexate (3.75 mg/kg) was administered intraperitoneally daily on days 7-9. Etoposide (10 mg/kg) and novobiocin (100 mg/kg) were administered intraperitoneally on alternate days (7, 9, 11). The combinations of PT523 with etoposide or novobiocin were significantly more effective than methotrexate and etoposide or novobiocin, producing tumor growth delays of 8.4 days and 6.9 days, respectively. Overall, the antifolate/topoisomerase II inhibitor treatment combinations produced tumor growth delays that were apparently additive to greater than additive.
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PMID:Antifolates can potentiate topoisomerase II inhibitors in vitro and in vivo. 776 54

Etoposide, an inhibitor of the breakage-reunion reaction associated with cellular type II DNA topoisomerases, was shown to inhibit plaque formation of vaccinia virus. This drug had a major effect on the segregation of newly replicated DNA concatemers. Gene expression and the initiation and elongation phases of viral DNA replication were essentially unaffected. Pulsed-field gel electrophoresis of viral DNA replicated in the presence of etoposide revealed two major classes of DNA: the mature monomeric linear genome and DNA that failed to enter the gel (the relative proportions depending on the concentrations of drug). Restriction enzyme analysis showed a severe defect in telomere resolution. In addition, slowly migrating restriction fragments were suggestive of a general recombination defect. We have isolated several etoposide-resistant mutants and used marker rescue and DNA sequencing to localize the resistance-causing mutation to the amino terminus of the viral DNA ligase gene. Inactivation of the DNA ligase also resulted in an etoposide-resistant phenotype, but to a lesser extent. The telomere resolution and segregation defects were corrected both in the drug-resistant mutants and in the DNA ligase knockout mutants. Reinsertion of wild-type or mutant DNA ligase in the viral thymidine kinase locus confirmed the role of the viral DNA ligase in conferring sensitivity not only to etoposide but also to another topoisomerase II inhibitor, 4'-(9-acridinylamino) methanesulphon-m-anisidide (mAMSA). The data suggest that the nonessential DNA ligase is involved in telomere resolution, possibly as part of a general recombinase.
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PMID:An etoposide-induced block in vaccinia virus telomere resolution is dependent on the virus-encoded DNA ligase. 788 54

Etoposide (VP-16) is used as an antineoplastic drug in humans. It inhibits topoisomerase II(topoII) activity by forming a ternary complex (DNA-etoposide-topoII). This complex prevents the DNA-strand rejoining activity of topo II, which results in DNA-strand breaks and the formation of structural chromosome aberrations. Topo II activity is also required for removing regions of DNA catenation prior to chromosome segregation. The possibility exists that patients undergoing etoposide chemotherapy may incur genetic damage and, consequently, may be at a greater risk for developing secondary tumors and having genetically abnormal offspring. We studied the ability of etoposide for inducing both structural chromosome aberrations and aneuploidy in mouse oocytes. Different dosages of etoposide were given to female mice at various times before and after human chronic gonadotrophin injection, and ovulated oocytes were collected 17 h later. The proportions of chromatid acentric fragments and of hyperploid metaphase II oocytes were significantly higher (P < 0.01) in the etoposide groups than in concurrent controls. These results indicate that both structural and numerical aberrations can be induced without direct interaction with DNA or with the various organelles associated with chromosome segregation. Additionally, unlike other compounds (vinblastine, colchicine, benomyl, and griseofulvin) that induce both meiotic delay (ovulated metaphase I oocytes and polyploidy) and aneuploidy, etoposide did not cause meiotic delay in oocyte maturation.
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PMID:Preferential pericentric lesions and aneuploidy induced in mouse oocytes by the topoisomerase II inhibitor etoposide. 791 Apr 18

Etoposide (VP-16) is one of the most important anticancer agents available and is used in many chemotherapeutic regimens. To characterize resistance to this drug, we established a VP-16-resistant human ovarian cancer cell line, SKOV3/VP, by continuous stepwise exposure of SKOV3 cells to VP-16. The degree of resistance to VP-16 of SKOV3/VP was about 25 times that of the parent cell line (SKOV3), and SKOV3/VP showed cross-resistance to teniposide, adriamycin, CPT-11, and vincristine. The accumulation of [3H]-VP-16 observed in SKOV3/VP cells was about half that seen in SKOV3 cells, and the accumulation of Adriamycin by this resistant cell line was also lower than that of its parent. Overexpression of neither the multidrug resistance gene mdr-1, the multidrug-resistance-associated protein (mrp) gene, nor P-glycoprotein was detected using reverse transcriptase-polymerase chain reaction analysis and flow cytometry with MRK-16, a monoclonal antibody against P-glycoprotein. The topoisomerase II activity of nuclear extracts from SKOV3/VP cells was lower than that from the parental cells, as was the amount of DNA topoisomerase II, demonstrated by immunoblotting. These results suggest that the mechanism responsible for the multidrug resistance of this cell line may be attributable to changes on its DNA topoisomerase II and to its reduced accumulation of the drugs as compared with the parental line SKOV3.
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PMID:Characterization of an etoposide-resistant human ovarian cancer cell line. 791 42

We have studied the ability of 8-methoxycaffeine (8-MOC)--one of the most effective caffeine derivatives in inducing chromosomal aberrations--to induce DNA double strand breaks (DSB) in purified human T lymphocytes during the cell cycle. Etoposide- or ellipticine-mediated DNA break frequency was used as a parameter of topoisomerase II activity. DNA-DSB induced by either 8-MOC or VP16 or ellipticine rose co-ordinately with the level of DNA topoisomerase II and with the onset of DNA replication. At concentrations between 10 and 50 microM 8-MOC was approximately 75% as active in terms of DSB as VP16 and ellipticine. By contrast with VP16 and ellipticine, 8-MOC was not cytotoxic. In conclusion, our data suggest that 8-MOC is an agent that efficiently induces DNA-DSB at non-toxic concentrations, and without direct inhibition of topoisomerase II.
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PMID:Induction of DNA double-strand breaks by 8-methoxycaffeine: cell cycle dependence and comparison with topoisomerase II inhibitors. 795 97

The genotoxic and cytotoxic effects of etoposide (VP-16), a topoisomerase II inhibitor, on male rat spermatogenic cells were studied by analysing induction of micronuclei during meiosis. Micronuclei (MN) were scored in early spermatids after different time intervals corresponding to exposure of different stages of meiotic prophase. Etoposide had a strong effect on diplotene-diakinesis I cells harvested 1 day after exposure, and a significant effect also on late pachytene cells harvested 3 days after exposure. The effect at 18 days corresponding to exposure of preleptotene stage of meiosis (S-phase) was weaker but also statistically significant. Adriamycin was used as a positive control in this study. The results indicate a different mechanism of action of etoposide compared with adriamycin and other chemicals studied previously with the spermatid micronucleus test. DNA flow cytometry was carried out to assess cytotoxic damage at the same time intervals (1, 3, and 18 days after treatment) at stages I and VII of the seminiferous epithelial cycle allowing a study of cytotoxicity to different spermatogenic cell stages. Damage of differentiating spermatogonia was observed by a decrease in the cell numbers of the 2C peak 1 and 3 days after treatment and by a reduction of the number of 4C cells (primary spermatocytes) 18 d after etoposide treatment. Adriamycin also killed differentiating spermatogonia. Since the cell population which showed a high induction of MN by etoposide was not reduced in number, the genotoxic effect is remarkable. We conclude that etoposide is a potent inducer of genotoxicity and patients treated with this agent during cancer chemotherapy are at a risk of genetic damage.
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PMID:Etoposide (VP-16) is a potent inducer of micronuclei in male rat meiosis: spermatid micronucleus test and DNA flow cytometry after etoposide treatment. 795 23

Etoposide (VP-16) is one of several DNA-damaging agents that induce subcellular structural changes associated with apoptosis. VP-16 exerts its DNA-damaging and cytotoxic effects subsequent to interference with DNA topoisomerase II activity. VP-16 also stimulates c-jun and c-fos mRNA expression in some cell lines, including human leukemia K562 and HL-60 cells. To compare the temporal relationship between drug-induced c-jun expression and apoptosis, we examined cell morphology, cell viability, DNA integrity, and c-jun induction during VP-16 treatment of K562 and HL-60 cells. VP-16 (10 microM)-induced internucleosomal DNA damage and nuclear fragmentation were readily apparent within 6 hr in HL-60 cells but were absent in K562 cells treated for up to 24 hr. Some internucleosomal DNA damage was observed in K562 cells but only after treatment with 100 microM VP-16 for 24 hr. In contrast, VP-16-induced DNA single-strand breaks, VP-16-induced topoisomerase II/DNA covalent complex formation, and VP-16-mediated growth inhibition were similar in K562 and HL-60 cells. Also, the time course of VP-16-induced c-jun mRNA expression was comparable for both K562 and HL-60 cell lines. Western blot analysis of whole-cell lysates showed that Bcl-2 protein levels were 13-fold greater in HL-60 cells than in K562 cells. Thus, the resistance of VP-16-treated K562 cells to apoptosis was not attributable to protection by Bcl-2. Furthermore, the relatively high levels of Bcl-2 in HL-60 cells were not sufficient to protect these cells against apoptosis. Together, our results indicate that the temporal coupling of VP-16-induced DNA damage, c-jun expression, and apoptosis is cell type specific and suggest that different signaling pathways for apoptosis are operating in these two human leukemia cell lines.
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PMID:Differential induction of etoposide-mediated apoptosis in human leukemia HL-60 and K562 cells. 796 39

The mechanism by which etoposide, a topoisomerase II inhibitor, killed replicating mouse L929 fibroblasts was investigated. Etoposide at 10 microM killed 70% of the cells within 4 days, a result that was accompanied by DNA fragmentation. A characteristic "ladder" pattern of DNA fragmentation was confirmed by agarose gel electrophoresis. Simultaneous exposure of the cells to 10 microM etoposide plus 1 microM cycloheximide reduced both the extent of cell killing and the fragmentation of DNA. Delayed addition of cycloheximide protected cells only if cycloheximide was added 1-6 hr after exposure to etoposide. When added 6-24 hr after treatment with etoposide, cycloheximide lost the ability to protect cells. Cell growth was completely inhibited by either etoposide or cycloheximide. Furthermore, DNA synthesis was inhibited by either etoposide or cycloheximide within 6 hr. Protein synthesis, however, was not inhibited by etoposide. Thus, the ability of cycloheximide to protect cells correlated with inhibition of protein synthesis, rather than inhibition of DNA synthesis. A 1-hr exposure to 2.5 mM N-methyl-N-nitrosourea similarly inhibited DNA synthesis within 6 hr. without affecting protein synthesis. However, no loss of viability accompanied N-methyl-N-nitrosourea treatment. Thus, an imbalance between protein synthesis and DNA synthesis cannot explain the cell killing by etoposide. H-7, a protein kinase C inhibitor, prevented the cell killing and DNA fragmentation, whereas aurintricarboxylic acid, an endonuclease inhibitor, reduced the extent of DNA fragmentation but did not have an effect on cell killing. The data document that the killing of replicating mouse fibroblasts by etoposide represents an example of programmed cell death (apoptosis) that depends on protein synthesis. Although protein synthesis is required during the first 24 hr of exposure to etoposide, cell death is delayed until several days later.
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PMID:Programmed cell death (apoptosis) of mouse fibroblasts is induced by the topoisomerase II inhibitor etoposide. 796 76

The suspect human carcinogen, etoposide, is known to be genotoxic, producing both gene and chromosomal mutations, probably by virtue of its ability to inhibit topoisomerase II activity. The present paper describes assays conducted using the Salmonella assay, the mouse lymphoma tk+/- assay (gene and chromosomal mutation analysis and molecular analysis of tk-/- mutants) and the mouse bone marrow micronucleus assay. Nonreproducible, weak, dose-related increases in mutation frequency in strain TA98 (but not TA1538 or TA1537) of Salmonella typhimurium were observed. Etoposide was highly mutagenic at the heterozygous thymidine kinase (tk+/-) locus of L5178Y mouse lymphoma cells at concentrations below 0.1 micrograms/ml. Mostly small colony mutants were induced, consistent with the potent clastogenicity also observed. Molecular analysis of mutants indicated that 83% and 92% of large and small colony mutants, respectively, had lost the entire target gene sequence. Chromosomally aberrant L5178Y cells were approximately 2 to 600-fold more prevalent than small tk-/- mutant colonies. This suggests that the viable target for etoposide-mediated clastogenesis in the selective assay is approximately one-fifth of chromosome 11b, itself being approximately one-fortieth of the mouse genome. An unusually potent response was observed for etoposide in the mouse bone marrow micronucleus assay (63.1 +/- 18 MPE/1,000 PE 24 hours after an oral dose of 1 mg/kg). The minimum detectable dose level in the assay was between 0.01 and 0.1 mg/kg. At dose levels between 1 and 15 mg/kg, an inverse dose response was observed. This reduction in assay response was not due to the small concommitant decrease in the incidence of polychromatic erythrocytes, a conclusion based on studies with N-methyl-N-nitrosourea. Animals sampled 48 hours after dosing with etoposide (10 mg/kg) had no polychromatic erythrocytes in the bone marrow. These observations for the micronucleus assay await explanation. The chemical structure of etoposide is displayed and discussed within the context of such strong mutagenic activity being associated with a nonelectrophilic agent.
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PMID:Potent clastogenicity of the human carcinogen etoposide to the mouse bone marrow and mouse lymphoma L5178Y cells: comparison to Salmonella responses. 773 7

Etoposide has demonstrated highly significant clinical activity against a wide variety of neoplasms, including germ-cell malignancies, small-cell lung cancer, non-Hodgkin's lymphomas, leukemias, Kaposi's sarcoma, neuroblastoma, and soft-tissue sarcomas. It is also one of the important agents in the preparatory regimens given prior to bone marrow and peripheral stem-cell rescue. Despite its high degree of efficacy in a number of malignancies, the optimal dose, schedule, and dosing form remain to be defined. It is possible that continuous or prolonged inhibition of the substrate, i. e., topoisomerase II, may be the key factor for the cytotoxic effects of etoposide. Clinical studies have shown the activity of etoposide to be schedule-dependent, with prolonged dosing, best accomplished by the oral dosing form, offering a therapeutic advantage. This benefit awaits validation by prospective randomized studies, some of which are in progress. Recent clinical investigations have focused on the use of etoposide in combination with (a) cytokines to ameliorate myelosuppression, the dose-limiting toxicity of etoposide; (b) agents such as cyclosporin A and verapamil to alter the p-glycoprotein (mdr1) function; and (c) topoisomerase I inhibitors to modulate the substrate upon which it acts. There is continued interest in the development of etoposide to its maximal clinical dimensions and in the examination of alternative biochemical and mechanistic approaches to further our understanding of this highly active agent.
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PMID:Etoposide: current status and future perspectives in the management of malignant neoplasms. 807 20


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