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)

The combination of cytokines and cytotoxic drugs offers a new approach to increase the therapeutic index in the treatment of neoplastic diseases. There is no consensus on optimal strategies for combining these agents so far. The molecular mechanisms underlying the interaction, however, should be defined in order to design clinical trials based on preclinical rationales. The broad spectrum of cytotoxic drugs whose activity can be enhanced by cytokines argues for multiple levels of drug interaction in vitro: alteration in the cellular drug uptake, modulation of drug target enzymes, and changes in metabolism or disposition of a drug. In vivo interaction between cytokines and cytotoxic agents involves an additional layer of complexity because of the effects of cytokines on the host immune system and on drug-metabolizing enzymes. A major mechanism involved in the synergistic interaction of interferon (IFN) and 5-fluorouracil (5-FU) seems to be the increase of active 5-FU metabolites by IFN. Moreover, IFN can reverse resistance against 5-FU by inhibiting the overexpression of thymidylate synthase. The absence of cytokinetic effects of IFN and FU argues against the recruitment of Gs cells into the cell cycle. Topoisomerase has emerged as a critical intracellular target of cytotoxic drugs. There is convincing evidence that the synergy between tumor necrosis factor (TNF) and topoisomerase-targeted intercalative (Adriamycin, doxorubicin hydrochloride; m-AMSA, amsacrine; mitoxantrone) and nonintercalative (VM-16, etoposide; VM-26, teniposide) drugs is related to a rapid increase in specific activity of topoisomerase I and II, resulting in enhanced DNA strand breaks and cleavage complex. Furthermore, sensitivity to topoisomerase II targeted drugs can be enhanced by granulocyte colony-stimulating factor (G-CSF) through elevated enzyme activity in tumor cell response to G-CSF. The synergistic interaction between cytokines and cytotoxic agents seems to be sequence dependent. It has recently been demonstrated that newly synthesized metal compounds and IFN are synergistic only after preincubation with cytokines. Cytokines can modulate expression of adhesion receptors on tumor cell lines, thereby influencing their metastatic potential. A considerable number of phase II trials with combination of cytokines and cytotoxic drugs based on these mechanisms have demonstrated promising response rates and tolerable toxicity. Phase III trials are currently in progress to identify enhanced activity combining cytokines and cytotoxic drugs in the treatment of malignancies.
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PMID:Biochemical modulation of cytotoxic drugs by cytokines: molecular mechanisms in experimental oncology. 759 4

In the MCF-7 human breast tumor cell line, the topoisomerase II inhibitor, VM-26, produces a concentration dependent reduction in expression of the oncogene c-myc which parallels growth inhibition. Down-regulation of c-myc expression was examined at transcriptional and post-transcriptional levels. VM-26, at 10 microM, produced a reduction in the transcription rate of both sense and antisense strands of c-myc as determined by nuclear run-off analysis. In contrast, in the presence of the RNA synthesis inhibitor, actinomycin D, VM-26 failed to alter the half-life of the c-myc message. The capacity of VM-26 to reduce c-myc expression was not abrogated in cells pretreated with the protein synthesis inhibitor, cycloheximide (despite superinduction of c-myc expression in both control and VM-26 treated cells); this observation suggests that de novo protein synthesis may not be required to mediate the effects of VM-26 on steady state c-myc transcript levels. An extended analysis of the time course of c-myc expression demonstrated that the decline of steady state c-myc mRNA levels induced by VM-26 was biphasic, 6 h after the initial reduction in c-myc expression to approx. 30% of control levels, c-myc levels rebounded to 70% of control; after 24 h, c-myc expression declined gradually and remained at depressed levels (40% of control) at 48 and 72 h. These observations suggest that the initial transient reduction in c-myc expression associated with inhibition of transcription may represent a component of an early signalling pathway leading to growth arrest in MCF-7 breast tumor cells exposed to VM-26.
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PMID:Transcriptional down-regulation of c-myc expression in the MCF-7 breast tumor cell line by the topoisomerase II inhibitor, VM-26. 759 88

The antineoplastic quinobenoxazines A-62176 and A-74932 were shown to be potent inhibitors of mammalian DNA topoisomerase II in vivo. This was demonstrated by their selective inhibition of the SV40 DNA replication stages that require topoisomerase II. Neither drug stabilized a covalent complex of the enzyme with SV40 DNA, which suggests that they are not poisons of DNA topoisomerase II. A-77601, an analog having little antitumor activity, barely inhibited DNA topoisomerase II in vivo, even at high concentrations. These findings were supported by in vitro studies which showed that A-62176 and A-74932, but not A-77601, strongly inhibited the catalytic activity of mammalian DNA topoisomerase II. A-62176 did not cause topoisomerase II-mediated DNA strand breaks in vitro under conditions in which adriamycin produced extensive DNA breakage. The antineoplastic and topoisomerase inhibitory activities of the quinobenoxazines correlate with their ability to unwind DNA. A-62176 antagonized the poisoning of topoisomerase II by VM-26 in vivo and in vitro, but had no effect on DNA breakage induced by camptothecin, a DNA topoisomerase I poison. A-62176 and A-74932 thus inhibit DNA topoisomerase II reactions at a step prior to the formation of the "cleavable complex" intermediate. These findings indicate that stabilization of the DNA topoisomerase II-DNA cleavable complex is not necessary for the antitumor activity of this class of quinolones and that the catalytic inhibition of DNA topoisomerase II may contribute significantly to the anticancer activity of other DNA topoisomerase II inhibitors.
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PMID:Quinobenoxazines: a class of novel antitumor quinolones and potent mammalian DNA topoisomerase II catalytic inhibitors. 772 84

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

We have examined the DNA cleavage site specificity of human type II DNA topoisomerase in the presence of each of five novel quinolone derivatives. Each quinolone derivative inhibited the human enzyme, inducing double-strand breaks with a four-base stagger. Break sites generated in response to each derivative had a predominance of C in the 3'-terminal position. Consensus sequences derived for cleavage sites induced by each derivative were strikingly similar, not only at the 3'-terminal position, but also at additional positions on either side of the broken phosphodiester bond. Analysis of these consensus sequences yielded information about possible interactions of specific substituents on the quinolone derivatives with DNA and/or topoisomerase. Comparison of the quinolone-based consensus sequences with those derived for cleavage sites generated by the human type II topoisomerase in the presence of either m-AMSA or VM-26, or in the absence of drug, provided compelling evidence that DNA cleavage sites include two domains: one which interacts with drug and a second, larger domain which interacts with topoisomerase.
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PMID:DNA sequence preferences at sites cleaved by human DNA topoisomerase II in response to novel quinolone derivatives. 774 59

A common strongly ordered multi-step-pattern of endogenous DNA degradation was induced in rat liver nuclei and intact thymocytes, prepared in the presence of chelating agents and incubated in the presence of CaCl2 and/or MgCl2. It consisted of sequential generation of 0.3 Mbp, then 0.05 Mbp DNA fragments and finally of oligo- and mononucleosomal DNA. Oligonucleosomal DNA was generated when the genome had already been disintegrated into 0.05 Mbp DNA fragments. ZnCl2 completely inhibited advanced genome cleavage to oligo- and mononucleosomal DNA without affecting the initial generation of large DNA fragments. Therefore, the endonucleolytic activity which produce large DNA fragments is different from Ca2+/Mg2+ endonuclease. The similar pattern of DNA degradation was observed in thymocytes treated with dexamethasone and with the topoisomerase II inhibitor VM-26, the agents known to induce apoptosis. The effect of VM-26 strongly suggests the involvement of topoisomerase II in generation of large DNA fragments. Multi-level organization and regulation of the chromatin structure determine the stepwise process of genome degradation. Detachment of chromatin from the nuclear matrix attachment regions may be one of the possible mechanisms of switching off the genome function and triggering the multi-step process of endogenous chromatin degradation thus leading to cell death in terminal differentiation or stress-induced apoptosis.
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PMID:Comparative study of induction of endogenous DNA degradation in rat liver nuclei and intact thymocytes. 774 35

In previous studies, we found that VP-16 (etoposide) induced cytotoxicity and protein-concealed strand break formation was prevented in a small cell lung cancer (SCLC) cell line, when the cells were incubated with aclarubicin prior to treatment with VP-16. In the present work, we studied the effect of adding aclarubicin to the cell suspension after VP-16. In a clonogenic assay, we found that the cytotoxicity induced by VP-16 in SCLC cells was inhibited when cells were postincubated with aclarubicin. The addition of aclarubicin at any time in relation to VP-16 was able to stop further cytotoxicity induced by the topoisomerase II (topo-II) targeting drug. Aclarubicin was also found to antagonize the cytotoxicity induced by VM-26 (teniposide), and m-AMSA. With the alkaline elution technique we found that postincubating the cells with aclarubicin inhibited VP-16-induced DNA strand break formation. In an in vitro system with purified topo-II and naked DNA we likewise found, that postincubation with aclarubicin prevented VP-16 induced cleavage. In the same in vitro system, also baseline cleavage induced by topo-II was inhibited when aclarubicin was present. Importantly, aclarubicin exerted the antagonism to topo-II targeting drugs both when administered prior to and after the topo-II targeting agents. Thus, our data suggest that sequential rather than simultaneous administration of aclarubicin and topo-II targeting agents may be superior with respect to net-cytotoxicity.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Postincubation with aclarubicin reverses topoisomerase II mediated DNA cleavage, strand breaks, and cytotoxicity induced by VP-16. 777 29

The relative content of topoisomerase II (topo II) and the induction of topo-II-mediated DNA damage and cellular abnormalities have been characterized in developing spermatogenic cells of Xenopus laevis to gain an insight into the role of topo II during spermatogenesis. Decatenation assays identified topo II activity in nuclear extracts from spermatocytes and pre-elongate spermatids, but not in extracts from elongate spermatids or sperm. Extracts from early-mid spermatids contained 14% (per cell) of the decatenation activity found in spermatocyte extracts. Immunoblots of SDS extracts from whole cells and nuclei from both spermatocytes and pre-elongate spermatids, but not elongate spermatids or sperm, resolved a 180 kDa polypeptide that reacts with polyclonal antisera to Xenopus oocyte topo II, an antipeptide antibody (FHD29) to human topo II alpha and beta, and an antipeptide antibody to human topo II alpha, suggesting homology between Xenopus spermatogenic cell topo II and mammalian topo II alpha. Immunofluorescence microscopy of topo II in testis cryosections revealed the presence of topo II in nuclei of all spermatogenic stages, but not in sperm. The relative levels of topo II estimated from fluorescence intensity were highest in spermatogonia and spermatocytes, then early-mid spermatids, followed by elongate spermatids and somatic cells. Incubation of isolated spermatogenic cells with teniposide (VM-26), a topo II-targetted drug, resulted in a dose-dependent induction of DNA breaks in all spermatocytes and spermatid stages to nuclear elongation stages, as analyzed by alkaline single cell gel electrophoresis. Addition of 0.5-50 microM VM-26 to spermatogenic cell cultures for 27 hours resulted in stage-dependent abnormalities. Mid-late spermatid stages were relatively resistant to VM-26-induced damage. In contrast, meiotic division stages were arrested and spermatogonia B were killed by VM-26, and VM-26 induced abnormal chromosome condensation in pachytene spermatocytes. The results of these studies show that cellular levels of topo II are stage-dependent during spermatogenesis, that most spermatogenic stages are sensitive to topo II-mediated DNA damage, and that spermatogonia B, meiotic divisions and pachytene spermatocytes are particularly sensitive to induction of morphological abnormalities and cell death during acute exposure to topo II-targetted drugs.
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PMID:Topoisomerase II expression and VM-26 induction of DNA breaks during spermatogenesis in Xenopus laevis. 787 55

A human breast cancer cell line (MCF7/WT) was selected for resistance to etoposide (VP-16) by stepwise exposure to 2-fold increasing concentrations of this agent. The resulting cell line (MCF7/VP) was 28-, 21-, and 9-fold resistant to VP-16, VM-26, and doxorubicin, respectively. MCF7/VP cells also exhibited low-level cross-resistance to 4'-(9-acridinylamino)-methanesulfon-m-anisidide, mitoxantrone, and vincristine and no cross-resistance to genistein and camptothecin. Furthermore, these cells were collaterally sensitive to the alkylating agents melphalan and chlorambucil. DNA topoisomerase II levels were similar in both wild-type MCF7/WT and drug-resistant MCF7/VP cells. In contrast, topoisomerase II from MCF7/VP cells appeared to be 7-fold less sensitive to drug-induced cleavable complex formation in whole cells and 3-fold less sensitive in nuclear extracts than topoisomerase II from MCF7/WT cells. Although this suggested that the resistant cells may contain a qualitatively altered topoisomerase II, no mutations were detected in either the ATP-binding nor the putative breakage/resealing regions of either DNA topoisomerase II alpha or II beta. In addition, the steady-state intracellular VP-16 concentration was reduced by 2-fold in the resistant cells, in the absence of detectable mdr1/P-gp expression and without any change in drug efflux. In contrast, expression of the gene encoding the MRP was increased at least 10-fold in resistant MCF7/VP cells as compared to sensitive MCF7/WT cells. These results suggest that resistance to epipodophyllotoxins in MCF7/VP cells is multifactorial, involving a reduction in intracellular drug concentration, possibly as a consequence of MRP overexpression, and an altered DNA topoisomerase II drug sensitivity.
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PMID:Multidrug resistance-associated protein gene overexpression and reduced drug sensitivity of topoisomerase II in a human breast carcinoma MCF7 cell line selected for etoposide resistance. 790 2

A number of clinically important drugs such as the epipodophyllotoxins etoposide (VP-16) and teniposide (VM-26), the anthracycline daunorubicin and doxorubicin (Adriamycin), and the aminoacridine amsacrine exert their cytotoxic action by stabilizing the cleavable complex formed between DNA and the nuclear enzyme topoisomerase II. We have previously demonstrated in several in vitro assays that the anthracycline aclarubicin (aclacinomycin A) inhibits cleavable-complex formation and thus antagonizes the action of drugs such as VP-16 and daunorubicin. The present study was performed to validate these in vitro data in an in vivo model. At nontoxic doses of 6 and 9 mg/kg, aclarubicin yielded a marked increase in the survival of non-tumor-bearing mice given high doses of VP-16 (80-90 mg/kg) in six separate experiments. In therapy experiments on mice inoculated with Ehrlich ascites tumor cells, aclarubicin given at 6 mg/kg roughly halved the increase in median life span induced by VP-16 at doses ranging from 22 to 33 mg/kg. An attempt to determine a more favorable combination of VP-16 and aclarubicin by increasing VP-16 doses failed, as the two drugs were always less effective than VP-16 alone. The way in which VP-16-induced DNA strand breaks lead to cell death remains unknown. However, VP-16 has been reported to cause apoptosis (programmed cell death) in several cell lines. To ascertain whether the protection given by aclarubicin could have a disruptive effect on the apoptotic process, we used the small intestine as an in vivo model. Whereas VP-16-induced apoptosis in crypt stem cells was detectable at a dose as low as 1.25 mg/kg, aclarubicin given at up to 20 mg/kg did not cause apoptosis. Indeed, aclarubicin caused a statistically significant reduction in the number of cells rendered apoptotic by VP-16. The present study thus confirms the previous in vitro experiments and indicates the value of including an in vivo model in a preclinical evaluation of drug combinations.
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PMID:In vivo inhibition of etoposide-mediated apoptosis, toxicity, and antitumor effect by the topoisomerase II-uncoupling anthracycline aclarubicin. 792 61


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