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:5.99.1.3 (topoisomerase)
9,911 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Catalytic inhibitors of mammalian DNA topoisomerase II have been found recently in natural and synthetic compounds. These compounds target the enzyme within the cell and inhibit various genetic processes involving the enzyme, such as DNA replication and chromosome dynamics, and thus proved to be good probes for the functional analyses of the enzyme in a variety of eukaryotes from yeast to mammals. Catalytic inhibitors were shown to be antagonists against topoisomerase II poisons. Thus bis(2,6-dioxopiperazines) have a potential to overcome cardiac toxicity caused by potent antitumor anthracycline antibiotics such as doxorubicin and daunorubicin. ICRF-187, a (+)-enantiomer of racemic ICRF-159, has been used in clinics in European countries as cardioprotector. Furthermore, bis(2,6-dioxopiperazines) enhance the efficacy of topoisomerase II poisons by reducing their side effects in preclinical and clinical settings. Bis(2,6-dioxopiperazines) per se among others have antitumor activity, and one of their derivatives, MST-16 or Sobuzoxane, bis(N1-isobutyloxycarbonyloxymethyl-2, 6-dioxopiperazine), has been developed in Japan as an anticancer drug used for malignant lymphomas and adult T-cell leukemia in clinics.
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PMID:Catalytic inhibitors of DNA topoisomerase II. 974 52

A series of topoisomerase-interacting antitumour agents were tested for their ability to differentially inhibit the catalytic activity of either topoisomerase (TOPO) IIalpha or beta, as judged by a DNA decatenation assay. The alpha form, relative to the beta isoform, proved 1 to 3 times more sensitive to nonintercalating complex-stabilizing TOPO II-interacting agents (etoposide and derivatives) and up to 18 times more sensitive to non-complex-stabilizing inhibitors of TOPO II ((+/-)-1,2-bis(3,5-dioxopiperazinyl-1-yl)propane [ICRF 159] and meso-2,3-bis(3,5-dioxopiperazine-1-yl)butane [ICRF 193]). However, the beta form of the enzyme appeared 1 to 3 times more sensitive to intercalating TOPO II-interacting agents (daunorubicin, aclarubicin and mitoxantrone). A possible implication of these data are that tumours preferentially expressing either the alpha or the beta isoform may be differentially responsive to various classes of TOPO II-interacting agents.
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PMID:Differential sensitivities of recombinant human topoisomerase IIalpha and beta to various classes of topoisomerase II-interacting agents. 976 27

Amrubicin is a novel, completely synthetic 9-aminoanthracycline derivative. Amrubicin and its C-13 alcohol metabolite, amrubicinol, inhibited purified human DNA topoisomerase II (topo II). Compared with doxorubicin (DXR), amrubicin and amrubicinol induced extensive DNA-protein complex formation and double-strand DNA breaks in CCRF-CEM cells and KU-2 cells. In this study, we found that ICRF-193, a topo II catalytic inhibitor, antagonized both DNA-protein complex formation and double-strand DNA breaks induced by amrubicin and amrubicinol. Coordinately, cell growth inhibition induced by amrubicin and amrubicinol, but not that induced by DXR, was antagonized by ICRF-193. Taken together, these findings indicate that the cell growth-inhibitory effects of amrubicin and amrubicinol are due to DNA-protein complex formation followed by double-strand DNA breaks, which are mediated by topo II.
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PMID:A new antitumor agent amrubicin induces cell growth inhibition by stabilizing topoisomerase II-DNA complex. 991 93

Doxorubicin induces DNA breakage by stabilizing a cleavable topoisomerase II-DNA complex. In contrast, topoisomerase II catalytic inhibitor ICRF-193 and uncoupling inhibitor aclarubicin interfere with the cleavable complex formation. We analysed combination effects of these drugs using two-dimensional flow cytometry of DNA content and the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labelling assay. Both ICRF-193 and aclarubicin attenuated the cytotoxic effect of doxorubicin on HL-60 cells (85% and 46% maximum reduction, respectively), which suggested that doxorubicin exerts its cytotoxic effect at least partially through the topoisomerase II-dependent DNA cleavage. Doxorubicin and ICRF-193 both induced G2 arrest in HL-60 cells, by which they may have reduced the cytotoxic effect of vincristine. Indeed, although ICRF-193 inhibited doxorubicin-induced apoptosis, ICRF-193 and doxorubicin cooperated in arresting HL-60 cells at G2 phase. These results indicated that G2 arrest was caused not only by DNA damage but also through a DNA damage-free, topoisomerase II inactivation-induced pathway. Western blot analysis showed that both types of G2 arrest were mediated by the inhibition of p34cdc2 dephosphorylation.
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PMID:Inhibition of p34cdc2 dephosphorylation in DNA damage- and topoisomerase II inactivation-induced G2 arrests in HL-60 cells. 1035 37

Catalysis of ATP hydrolysis by two NH(2)-terminal fragments of yeast DNA topoisomerase II was studied in the absence and presence of DNA, and in the absence and presence of inhibitor ICRF-193. The results indicate that purified Top2-(1-409), a fragment containing the NH(2)-terminal 409 amino acids of the yeast enzyme, is predominantly monomeric, with a low level of ATPase owing to weak association of two monomers to form a catalytically active dimer. The ATPase activity of Top2-(1-409) is independent of DNA in a buffer containing 100 mM NaCl, in which intact yeast DNA topoisomerase II exhibits robust DNA-dependent ATPase and DNA transport activities. Purified Top2-(1-660), a fragment containing the NH(2)-terminal 660 amino acid of the yeast enzyme, appears to be dimeric in the absence or presence of DNA, and the ATPase activity of the protein is significantly stimulated by DNA. These results are consistent with a model in which binding of an intact DNA topoisomerase II to DNA places the various subfragments of the enzyme in a way that makes the intramolecular dimerization of the ATPase domains more favorable. We believe that this alignment of subfragments is mainly achieved through the binding of the enzyme to the DNA segment within which the enzyme makes transient breaks. The ATPase activity of Top2-(1-409) is inhibited by ICRF-193, suggesting that the bisdioxopiperazine class of DNA topoisomerase II inhibitors directly interacts with the paired ATPase domains of the enzyme.
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PMID:Catalysis of ATP hydrolysis by two NH(2)-terminal fragments of yeast DNA topoisomerase II. 1041 79

A screening procedure which permits identification of compounds based on their activities against specific biological targets directly in a living organism, Saccharomyces cerevisiae, has been established as part of our new drug discovery programme. Use of this assay has provided the first direct evidence that TOP1 and RAD52 proteins are involved in the mode of action of bisdioxopiperazine ICRF compounds, which thus express a mode of action quite distinctive from the other known TOP2 inhibitors evaluated. The functional assay is based on a comparison of pairs of yeast differing in their phenotypes by specific traits: the expression or lack of expression of ectopic human DNA topoisomerase I, with or without that of the RAD52 gene. Amongst a series of anticancer agents, inhibitors of topoisomerase I (camptothecin) were identified as such in yeast expressing human topoisomerase I, whilst the presence or absence of RAD52 protein permitted the discrimination of compounds generating double-stranded DNA breaks, either directly (bleomycin) or involving DNA adduct formation (cisplatin), or indirectly with DNA damage mediated via inhibition of the topoisomerase II enzyme (etoposide). Notably, however, both the RAD52 protein and the lack of TOP1 enzyme appeared implicated in the cytotoxic activities of the series of bisdioxopiperazine ICRF compounds tested. This functional assay in a living organism therefore appears to provide a valuable tool for probing distinctive and specific mode(s) of action of diverse anticancer agents.
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PMID:Differential expression of topoisomerase I and RAD52 protein in yeast reveals new facets of the mechanism of action of bisdioxopiperazine compounds. 1055 49

Bisdioxopiperazines are a unique class of topoisomerase II inhibitors that lock topoisomerase II at a point in the enzyme reaction cycle where the enzyme forms a closed clamp around DNA. We examined cell killing by ICRF-187 and ICRF-193 in yeast cells expressing human topoisomerase II alpha (htop-IIalpha). Expression of htop-IIalpha in yeast cells sensitizes them to both ICRF-187 and ICRF-193, compared with cells expressing yeast topoisomerase II. ICRF-193 is still able to exert growth inhibition in the presence of genes encoding both ICRF-193-resistant and ICRF-193-sensitive htop-IIalpha enzymes, indicating that sensitivity to bisdioxopiperazines is dominant. Killing by ICRF-193 occurs more rapidly, than the killing in yeast cells due to a temperature-sensitive yeast topoisomerase II incubated at the non-permissive temperature. These results are reminiscent of a top-II poison such as etoposide. However, the killing caused by ICRF-193 and ICRF-187 is not enhanced by mutations in the RAD52 pathway. The levels of drug-induced DNA cleavage observed with htop-IIalpha in vitro is insufficient to explain the sensitivity induced by this enzyme in yeast cells. Finally, arrest of cells in G(1) does not protect cells from ICRF-193 lethality, a result inconsistent with killing mechanisms due to catalytic inhibition of top-II or stabilization of a cleavable complex. We suggest that the observed pattern of cell killing is most consistent with a poisoning of htop-II by ICRF-193 by a novel mechanism. The accumulation of closed clamp conformations of htop-II induced by ICRF-193 that are trapped on DNA might interfere with transcription, or other DNA metabolic processes, resulting in cell death.
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PMID:A novel mechanism of cell killing by anti-topoisomerase II bisdioxopiperazines. 1063 19

A study was made of the influence of inhibitors of poly(ADP-ribose)polymerase, topoisomerase I and topoisomerase II on the frequency of gene targeting of hprt gene as well as on the frequency of random integration of targeting vector pRV9.1 into genome of mouse F9 teratocarcinoma cells. We found that the treatment of cells with the inhibitor of poly(ADP-ribose)polymerase 3-aminobenzamide after electroporation resulted in 3-4-times increase of homologous integration of exogenic vector into chromosomal DNA, and did not affect the frequency of random insertion of transfected DNA. The treatment of cells after electroporation with inhibitors of topoisomerases VP-16, ICRF-193 enhanced random integration of transfected DNA but exerted no effect on the frequency of gene targeting in this experimental system.
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PMID:[Effect of inhibitors of topoisomerases and poly(ADP-ribosylation) on homologous and non-homologous integration of exogenous DNA in genes of mammalian somatic cells]. 1064 51

DNA topoisomerase II uses a complex, sequential mechanism of ATP hydrolysis to catalyze the transport of one DNA duplex through a transient break in another. ICRF-193 is a catalytic inhibitor of topoisomerase II that is known to trap a closed-clamp intermediate form of the enzyme. Using steady-state and rapid kinetic ATPase and DNA transport assays, we have analyzed how trapping this intermediate by the drug perturbs the topoisomerase II mechanism. The drug has no effect on the rate of the first turnover of decatenation but potently inhibits subsequent turnovers with an IC(50) of 6.5 +/- 1 microM for the Saccharomyces cerevisiae enzyme. This drug inhibits the ATPase activity of topoisomerase II by an unusual, mixed-type mechanism; the drug is not a competitive inhibitor of ATP, and even at saturating concentrations of drug, the enzyme continues to hydrolyze ATP, albeit at a reduced rate. Topoisomerase II that was specifically isolated in the drug-bound, closed-clamp form continues to hydrolyze ATP, indicating that the enzyme clamp does not need to re-open to bind and hydrolyze ATP. When rapid-quench ATPase assays were initiated by the addition of ATP, the drug had no effect on the sequential hydrolysis of either the first or second ATP. By contrast, when the drug was prebound, the enzyme hydrolyzed one labeled ATP at the uninhibited rate but did not hydrolyze a second ATP. These results are interpreted in terms of the catalytic mechanism for topoisomerase II and suggest that ICRF-193 interacts with the enzyme bound to one ADP.
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PMID:Steady-state and rapid kinetic analysis of topoisomerase II trapped as the closed-clamp intermediate by ICRF-193. 1064 21

ICRF-187 is a bisdioxopiperazine anticancer drug that inhibits the catalytic activity of DNA topoisomerase (topo) II without stabilizing DNA-topoII cleavable complexes. To better understand the mechanisms of action of and resistance to topoII catalytic inhibitors, human leukemic CEM cells were selected for resistance to ICRF-187. The clones CEM/ICRF-8 and CEM/ICRF-18 are approximately 40- and 69-fold resistant to ICRF-187, and 12- and 67-fold cross-resistant to ICRF-193, respectively, but are sensitive to other topoII catalytic inhibitors (merbarone and aclarubicin), as well as collaterally sensitive to the DNA-topoII complex-stabilizing drug etoposide (VP-16). Both the number of VP-16- induced DNA-topoII complexes formed and the amount of in vitro topoII catalytic activity are enhanced in the drug-resistant cells. The ICRF-187-resistant clones contain approximately 5-fold increase in topoIIalpha protein levels and approximately 2.2-fold increase in topoIIalpha mRNA levels. Furthermore, CEM/ICRF-8 expresses approximately 3.5-fold increase in topoIIalpha promoter activity, suggesting that up-regulation of topoIIalpha in this clone occurs at the transcriptional level. Treatment of the drug-resistant or -sensitive cells with equitoxic doses of merbarone or teniposide results in a G(2)/M arrest. In marked contrast, when treated with equitoxic ICRF-187 doses, the drug-resistant clones exhibit either a transient arrest or completely lack the G(2)/M checkpoint compared with the drug-sensitive cells. This aberrant cell cycle profile is associated with a 48-h delay in drug-induced apoptotic cell death, as revealed by fluorescent-end labeling of DNA and poly (ADP-ribose) polymerase cleavage. In summary, resistance to ICRF-187 in CEM cells is associated with increased levels of catalytically active topoIIalpha and altered G(2)/M checkpoint and apoptotic responses.
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PMID:Selection of human leukemic CEM cells for resistance to the DNA topoisomerase II catalytic inhibitor ICRF-187 results in increased levels of topoisomerase IIalpha and altered G(2)/M checkpoint and apoptotic responses. 1064 39


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