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:5.99.1.2 (topoisomerase)
9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ellipticine derivatives have been shown to induce DNA strand breaks by trapping DNA-topoisomerase II (Topo II) in an intermediary covalent complex between Topo II and DNA which could be related to their cytotoxic effects. We report here that Celiptium and Detalliptinium, two ellipticine derivatives clinically used for their antitumoral properties against breast cancer, exhibit the highest in vitro activity on Topo II DNA cleavage reaction and decatenation among a series of 14 ellipticine derivatives. The in vitro cleavage site specificity in pBR 322 plasmid DNA and in a human c-myc gene inserted in a lambda phage DNA is identical for both ellipticines, but different from m-AMSA, another Topo II related antitumoral agent. Recently, it has been shown that the ellipticine derivative Celiptium presents a strong cytotoxic activity in vitro on different human tumors including small cell lung carcinoma (SCLC). However, the studies that involved Topo II as a target for ellipticine derivatives have been performed only by using animal tumor cell lines. Therefore we have studied the in vivo DNA cleavage activity of Celiptium and Detalliptinium on a human SCLC cell line, NCI N417, comparatively to that obtained with m-AMSA. The respective IC50 on cell growth are 9, 8 and 1 microM for Celiptium, Detalliptinium and m-AMSA, respectively. Using the alkaline elution technique, we have observed that Celiptium and Detalliptinium exhibit a weak cleavage activity on genomic DNA from whole cells. The ellipticines are about 50 times less potent than m-AMSA in inducing DNA strand breaks. Analysis of in vivo c-myc gene cleavage by Southern blot hybridization also demonstrates a lack of activity of the ellipticine derivatives as no gene cleavage could be detected up to 50 microM of the drug. With m-AMSA, c-myc gene cleavage is detected at a concentration of 0.2 microM, which indicates that this methodology is less sensitive in detecting DNA strand breaks than is the alkaline elution. Further studies of the drug effect on isolated nuclei by alkaline elution also show that the DNA cleavage activity of Celiptium and Detalliptinium is increased when compared to whole cells. Our data indicate that these two drugs have a weaker cytotoxic effect than m-AMSA on NCI N417 cell line, due to a limited access to the cell nucleus rather than to a lack of activity on Topo II as assessed by in vitro and isolated nuclei experiments.
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PMID:Topoisomerase II-mediated DNA cleavage activity induced by ellipticines on the human tumor cell line N417. 254 83

Aza-ellipticines are DNA intercalative ellipticine analogues with antitumor activity that induce protein-linked DNA breaks in NIH 3T3 cells in culture. The effects of two aza-ellipticine congeners (BD-40 and BR-76) on the activity of purified Calf Thymus type II topoisomerase were studied using pUC13 DNA as substrate. DNA cleavage was stimulated by both molecules at those doses required for inducing lethal effects in cells (DE5O). This effect was reversed by high salt treatment, indicating that it was actually mediated by Topo II. Mapping of cleavage sites on linearized and 3' end-labelled pUC13 DNA showed that ellipticine and aza-ellipticines stimulated the same sites, which differed from those stimulated by m-AMSA. Decatenating activity of Topo II on Trypanosoma cruzi kDNA was both inhibited by ellipticine and BD-40 at concentrations much higher than DE50 concentrations. Activity of aza-ellipticines was also investigated on isolated nuclei. Unlike ellipticine which promoted DNA-breaking activity, BD-40 and BR-76 were repeatedly inactive. Prior treatment of DNA by Proteinase K did not reveal hidden breaks which are formed in intact cells treated with BD-40 (Vilarem et al., 1984, Nucleic Ac. Res. 12, 8653). Concordant with these data, BD-40 did not impair DNA-synthetic activity in isolated nuclei, while Ellipticine largely decreased it. These results indicate that lesions induced in DNA by Aza-ellipticines are mediated by Topo II. The absence of effect of these drugs on isolated nuclei compared to that of Ellipticine may be due to some specific features of the association between Topo II and Aza-ellipticines or reflect a bioactivation step as a prerequisite for in vivo activity.
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PMID:The in vitro involvement of topoisomerase II in the activity of aza-ellipticine analogues is not correlated with drug activity on isolated nuclei. 301 46

I have found that antineoplastic drugs which are known to be inhibitors of mammalian DNA topoisomerases have pronounced and selective effects on simian virus 40 DNA replication. Ellipticine, 4'-(9-acridinylamino)methanesulfon-m-aniside, and Adriamycin blocked decatenation of newly replicated simian virus 40 daughter chromosomes in vivo. The arrested decatenation intermediates produced by these drugs contained single-strand DNA breaks. Ellipticine in particular produced these catenated dimers rapidly and efficiently. Removal of the drug resulted in rapid reversal of the block and completion of decatenation. The demonstration that these drugs interfere with decatenation suggests that they may exert their cytotoxic and antineoplastic effects by preventing the separation of newly replicated cellular chromosomes. Camptothecin rapidly breaks replication forks in growing Cairns structures. It is likely that the target of camptothecin is the "swivel" topoisomerase required for DNA replication and that it is located at or very near the replication fork in vivo. Evidence is presented that many of the broken Cairns structures are in fact half-completed sister chromatid exchanges. One pathway for the resolution of these structures is completion of the sister chromatid exchange to produce a circular head-to-tail dimer.
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PMID:Topoisomerase inhibitors can selectively interfere with different stages of simian virus 40 DNA replication. 302 45

Ellipticine (EPC), a natural alkaloid extracted from Aspidosperma williansii (Apocynaceae), is known to have antitumor and cytotoxic activities on various types of tumors. This drug showed a strong clastogenic effect on bone marrow cells of Wistar rats treated in vivo (7.75-31.00 mg/kg body weight). EPC was also tested in vitro using the human peripheral blood lymphocyte system, at concentrations 100 times lower than those used in the in vivo test on rats, since the cytotoxic effect on lymphocytes was very strong. At the 2 highest concentrations used (7.75 X 10(-1) and 1.55 X 10(-1) micrograms/ml culture medium), EPC induced a statistically significant increase in the frequency of chromosome aberrations and sister-chromatid exchanges in lymphocytes. Based on data reported in the literature, we have tried to establish relationships between the clastogenic effect observed and the process of EPC intercalation into DNA and the formation of protein-associated DNA-strand breaks probably promoted by topoisomerase enzymes.
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PMID:Clastogenic effect of the plant alkaloid ellipticine on bone marrow cells of Wistar rats and on human peripheral blood lymphocytes. 336 53

Previous work has shown that exposing mouse L1210 cells to intercalating agents such as adriamycin, ellipticine and actinomycin D results in DNA single-stranded breaks and DNA-protein crosslinks. To characterize further the interaction between these drugs and intracellular DNA we have employed a modification of the alkaline elution technique which allows the detection of DNA double-stranded breaks. Ellipticine (1.25-5.0 microgram/ml) adriamycin (0.5-3.0 microgram/ml) and actinomycin D (1.5-3.0 microgram/ml) all caused double-stranded breaks in DNA from L1210 cells following a 1 h treatment. The number of double-stranded breaks found per single strand break was highest for ellipticine, despite the fact that this is least cytotoxic of the three drugs. By comparing the single and double strand break frequency caused by radiation to that caused by ellipticine, it appears that most if not all of the drug-induced single strand breaks observed actually represent double-strand breaks. We suggest that these double-strand breaks may result from the action of an intracellular enzyme, perhaps topoisomerase, which breaks both strans in concert to relieve the topological strain caused by drug intercalation.
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PMID:DNA double-stranded breaks in mammalian cells after exposure to intercalating agents. 727 6

Ellipticine is a potent antitumor agent whose mechanism of action is considered to be based mainly on DNA intercalation and/or inhibition of topoisomerase II. Using [3H]-labeled ellipticine, we observed substantial microsome (cytochrome P450)-dependent binding of ellipticine to DNA. In rat, rabbit, minipig, and human microsomes, in reconstituted systems with isolated cytochromes P450 and in Supersomes containing recombinantly expressed human cytochromes P450, we could show that ellipticine forms a covalent DNA adduct detected by [32P]-postlabeling. The most potent human enzyme is CYP3A4, followed by CYP1A1, CYP1A2, CYP1B1, and CYP2C9. Another minor adduct is formed independent of enzymatic activation. The [32P]-postlabeling analysis of DNA modified by activated ellipticine confirms the covalent binding to DNA as an important type of DNA modification. The DNA adduct formation we describe is a novel mechanism for the ellipticine action and might in part explain its tumor specificity.
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PMID:The anticancer agent ellipticine on activation by cytochrome P450 forms covalent DNA adducts. 1175 21

Ellipticine is a potent antineoplastic agent whose mechanism of action is considered to be based mainly on DNA intercalation and/or inhibition of topoisomerase II. Recently, we found that ellipticine also forms covalent DNA adducts and that the formation of the major adduct is dependent on the activation of ellipticine by cytochrome P450 (CYP). We examined a panel of genetically engineered V79 cell lines including the parental line V79MZ and recombinant cells expressing the human CYP enzymes CYP1A1, CYP1A2 or CYP3A4 for their ability to activate ellipticine. The extent of activation was determined by analysing DNA adducts by 32P-postlabelling. Ellipticine was found to be toxic to all V79 cell lines with IC(50) values ranging from 0.25 to 0.40 microM. The nuclease P1 version of the 32P-postlabelling assay yielded a similar pattern of ellipticine-DNA adducts with two major adducts in all cells, the formation of only one of which was dependent on CYP activity. This pattern is identical to that detected in DNA reacted with ellipticine and the reconstituted CYP enzyme system in vitro as confirmed by HPLC of the isolated adducts. Total adduct levels ranged from 2 to 337 adducts per 10(8) nucleotides, in the parental line and in V79 expressing CYP3A4, respectively. As in vitro, human CYP1A2 and CYP1A1 were less active. The results presented here are the first report showing the formation of CYP-mediated covalent DNA adducts by ellipticine in cells in culture, and confirm the formation of covalent DNA adducts as a new mechanism of ellipticine action.
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PMID:Covalent binding of the anticancer drug ellipticine to DNA in V79 cells transfected with human cytochrome P450 enzymes. 1212 50

Ellipticine is a potent antineoplastic agent, whose mode of action is considered to be based mainly on DNA intercalation and/or inhibition of topoisomerase II. Recently, we found that ellipticine also forms covalent DNA adducts and that the formation of the major adduct is dependent on the activation of ellipticine by cytochrome P450 (P450). We examined rat, rabbit, and human hepatic microsomal samples for their ability to activate ellipticine. The extent of activation was determined by binding of 3H-labeled ellipticine to DNA and by analyzing DNA adducts by 32P-postlabeling. We demonstrate that cytochrome P450 of human hepatic microsomes activating ellipticine to species binding to DNA is analogous to that of rats, but not of rabbits. Most of the ellipticine activation in rat and human hepatic microsomes is attributed to P450 enzymes of the same subfamily, P450 3A1/2 and P450 3A4, respectively, while the orthologous enzyme in rabbit hepatic microsomes, P450 3A6, is much less efficient. With purified enzymes, the major role of P450 3A1 and 3A4 in ellipticine-DNA adduct formation was confirmed. We identified deoxyguanosine as the target for P450-mediated ellipticine binding to DNA using polydeoxyribonucleotides and deoxyguanosine 3'-monophosphate. The results strongly suggest that rats are more suitable models than rabbits mimicking the metabolic activation of ellipticine in humans.
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PMID:Rat microsomes activating the anticancer drug ellipticine to species covalently binding to deoxyguanosine in DNA are a suitable model mimicking ellipticine bioactivation in humans. 1269 29

Ellipticine is a potent antineoplastic agent whose mode of action is considered to be based mainly on DNA intercalation and/or inhibition of topoisomerase II. Recently, we found that ellipticine also forms covalent DNA adducts in vitro and that the formation of the major adduct is dependent on the activation of ellipticine by cytochrome P450 (CYP). Here, we investigated the capacity of ellipticine to form DNA adducts in vivo. Male Wistar rats were treated with ellipticine, and DNA from various organs was analyzed by (32)P postlabeling. Ellipticine-specific DNA adduct patterns, similar to those found in vitro, were detected in most test organs. Only DNA of testes was free of the ellipticine-DNA adducts. The highest level of DNA adducts was found in liver (19.7 adducts per 10(7) nucleotides), followed by spleen, lung, kidney, heart and brain. One major and one minor ellipticine-DNA adducts were found in DNA of all these organs of rats exposed to ellipticine. Besides these, 2 or 3 additional adducts were detected in DNA of liver, kidney, lung and heart. The predominant adduct formed in rat tissues in vivo was identical to the deoxyguanosine adduct generated in DNA by ellipticine in vitro as shown by cochromatography in 2 independent systems. Correlation studies showed that the formation of this major DNA adduct in vivo is mediated by CYP3A1- and CYP1A-dependent reactions. The results presented here are the first report showing the formation of CYP-mediated covalent DNA adducts by ellipticine in vivo and confirm the formation of covalent DNA adducts as a new mode of ellipticine action.
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PMID:DNA adduct formation by the anticancer drug ellipticine in rats determined by 32P postlabeling. 1460 Oct 46

Ellipticine is an antineoplastic agent, the mode of action of which is considered to be based on DNA intercalation and inhibition of topoisomerase II. We found that ellipticine also forms the cytochrome P450 (CYP)-mediated covalent DNA adducts. We now identified the ellipticine metabolites formed by human CYPs and elucidated the metabolites responsible for DNA binding. The 7-hydroxyellipticine, 9-hydroxyellipticine, 12-hydroxyellipticine, 13-hydroxyellipticine, and ellipticine N(2)-oxide are generated by hepatic microsomes from eight human donors. The role of specific CYPs in the oxidation of ellipticine and the role of the ellipticine metabolites in the formation of DNA adducts were investigated by correlating the levels of metabolites formed in each microsomal sample with CYP activities and with the levels of the ellipticine-derived deoxyguanosine adducts in DNA. On the basis of this analysis, formation of 9-hydroxyellipticine and 7-hydroxyellipticine was attributable to CYP1A1/2, whereas production of 13-hydroxyellipticine and ellipticine N(2)-oxide, the metabolites responsible for formation of two major DNA adducts, was attributable to CYP3A4. Using recombinant human enzymes, oxidation of ellipticine to 9-hydroxyellipticine and 7-hydroxyellipticine by CYP1A1/2 and to 13-hydroxyellipticine and N(2)-oxide by CYP3A4 was corroborated. Homologue modeling and docking of ellipticine to the CYP3A4 active center was used to explain the predominance of ellipticine oxidation by CYP3A4 to 13-hydroxyellipticine and N(2)-oxide.
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PMID:The anticancer drug ellipticine forms covalent DNA adducts, mediated by human cytochromes P450, through metabolism to 13-hydroxyellipticine and ellipticine N2-oxide. 1554 7


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