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)

Fostriecin is a new antitumor antibiotic which is being developed further as an anticancer agent based on its marked activity in murine leukemias. Its mechanism of action, however, has thus far remained unknown. The present study demonstrates that fostriecin inhibits the catalytic activity of partially purified type II topoisomerase from Ehrlich ascites carcinoma. Under the experimental conditions employed, fostriecin completely inhibited the enzyme at 100 microM. A general kinetic analysis showed that fostriecin inhibited topoisomerase in an uncompetitive manner with a Ki,app of 110 microM and produced kinetics that were distinctly different from those of VM-26 which exhibited noncompetitive inhibition. Fostriecin did not cause DNA strand breaks in L1210 cells, suggesting that it did not stabilize a cleavable complex as do other known inhibitors of this enzyme. Fostriecin, however, did partially inhibit DNA strand breaks produced by amsacrine. An analysis by flow cytometry showed that L1210 cells exposed to 5 microM fostriecin for 12 hr caused a block in the G2 phase of the cell cycle. These studies thus suggest that the mechanism by which fostriecin produces its antitumor effects may be through inhibition of topoisomerase II and that the type of inhibition is markedly different from existing antitumor agents which inhibit this enzyme.
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PMID:Inhibition of type II topoisomerase by fostriecin. 284 52

It has recently been suggested that topoisomerases could be important targets for several DNA intercalating drugs used in cancer therapy. This prompted us to purify and characterize a type II topoisomerase in a highly tumorigenic transplantable rabbit tumor isolated from a skin carcinoma associated with cottontail rabbit papillomavirus. We have found that the decatenating activity present in tumor cells was 40-100 times higher than that in the rabbit liver, while no activity could be found in skin extracts. The type II topoisomerases purified from tumor and liver cells consist of two subunits with molecular masses of about 160 kDa. The conditions of the reactions of relaxation, unknotting and decatenation catalyzed by these topoisomerases II were found to be similar to those observed with enzymes of other eukaryotic cells. In the course of the purification of the VX2 enzyme, we isolated and characterized a protein of about 30 kDa in whose presence the topoisomerase II was able to catenate very efficiently supercoiled DNA molecules. This protein has the same electrophoretic mobility as an H1-2 histone, and cross-reacts with an anti-H1 antiserum. The VX2 topoisomerase II as well as the VX2 tumor should constitute useful models for assays of antitumoral drugs.
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PMID:A type-II DNA topoisomerase and a catenating protein from the transplantable VX2 carcinoma. 298 98

Evidence suggests that the anticancer agents etoposide (VP16-213) and teniposide (VM26) produce DNA breaks and cytotoxicity by interaction with type II topoisomerase. Therefore, levels of type II topoisomerase may influence sensitivity to VP16-213 and VM26. We have characterized four lung carcinoma-derived cell lines for natural sensitivity or resistance to VP16-213 and VM26. Included in this study were two small cell lung carcinoma lines (SW900 and SW1271), an adenocarcinoma line (A549), and a large cell carcinoma (H157). SW1271 was the most sensitive line with a median inhibitory concentration for cell proliferation of 0.5 microM for VM26 and 2.7 microM for VP16-213, and SW900 was the most resistant with median inhibitory concentration values of 2.0 and 16 microM, respectively. A549 and H157 cells were intermediate in sensitivity to these drugs. Alkaline elution techniques were used to study in vivo formation and repair of single and double strand DNA breaks. Single strand DNA breaks were observed in SW1271 cells exposed to as little as 10 nM VM26 or 100 nM VP16-213 for 1 h, whereas SW900 cells required exposure to 10-fold higher concentrations of VM26 or VP16-213 to produce similar results. Single strand DNA breaks predominated only in SW1271 and A549 cells and then, only at low drug concentrations, whereas the ratios between single and double strand DNA breaks decreased at higher drug concentrations. Plots of cytotoxicity versus single and double strand DNA breakage revealed that cytotoxicity produced by both drugs was more closely related to double strand DNA break formation in all four cell lines. DNA breaks appeared rapidly upon addition of drug, reaching plateaus in DNA breaks within 30 min, and repair of both single and double strand DNA breaks occurred rapidly with time to repair one-half of the DNA breaks of 20 to 60 min in all four cell lines upon removal of drug, arguing against repair as a mechanism for drug resistance. DNA breakage was also observed in nuclei isolated from SW900 and SW1271 cells in similar magnitude to that observed in the respective cells. Results indicate that DNA breakage plateaus may reflect a steady-state equilibrium established between the drug and its nuclear target, possibly type II topoisomerase, and suggest that natural resistance to VP16-213 and VM26 may be due to different enzyme levels in sensitive and naturally resistant cells.
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PMID:DNA breakage in human lung carcinoma cells and nuclei that are naturally sensitive or resistant to etoposide and teniposide. 301 77

Various analogs of etoposide have been studied and compared in different tests in order to identify which tests best correlate with antitumor activity. These tests included DNA breakage assays using standard alkaline elution procedures as a means of studying topoisomerase II inhibition in intact cells, cytotoxicity studies in naturally sensitive and resistant human carcinoma cell lines, in vitro assays of the effect of the different congeners on topoisomerase II activity, and a preliminary evaluation of the ability of etoposide and teniposide to induce resistance. As in previous studies, a direct correlation was seen between double strand DNA breakage and cytotoxicity but not between single strand DNA breakage and cytotoxicity. Analogs with blocked 4'-hydroxyl groups were poor antitumor agents but were still capable of inhibiting topoisomerase II as evidenced by the production of DNA breaks. However, this DNA breakage was qualitatively different from that produced by VP16. None of the analogs were able to overcome either naive or acquired drug resistance. The dihydroxy analog of VP16, a possible bioactivated analog, was much less potent and possibly less stable than VP16. A model is proposed for the inhibition of topoisomerase II by demethylepipodophyllotoxins that may explain the relationship between double strand DNA breakage and cytotoxicity.
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PMID:Structure-activity relationships of podophyllin congeners that inhibit topoisomerase II. 304 Dec 39

DNA-topoisomerase catalyzing the conversion of a superhelical circular covalently closed DNA molecule into a super-helix free circular molecule, was isolated from mouse Ehrlich ascites carcinoma cells and purified 209-fold. The optimal conditions for the action and stability of the enzyme were elaborated. Using polyacrylamide gel electrophoresis under non-denaturating conditions as well as in the presence of Na-DS the heterogeneity of purified DNA-topoisomerase was established. This heterogeneity implies the presence of three active forms of the enzyme with Mr of 97 000, 81 000 and 69 000, respectively. Using one-dimensional fingerprint method and limited proteolysis with Staphylococcus aureus protease, it was demonstrated that the low molecular weight enzyme forms are products of limited proteolysis of the highest molecular weight form of DNA-topoisomerase.
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PMID:[Isolation, some properties and heterogeneous nature of DNA-topoisomerase (relaxing enzyme) from Ehrlich ascites carcinoma cells]. 629 17

We have previously shown that a DNA topoisomerase I from mouse mammary carcinoma cells is inhibited by heparin. Taking advantage of this enzyme-heparin interaction, we developed a rapid and efficient method of purification of this enzyme to near homogeneity by extraction of chromatin with 0.15 M phosphate buffer followed by two-step column chromatography on heparin-Sepharose and phenyl-Sepharose. Electrophoresis on sodium dodecyl sulfate-polyacrylamide gels revealed that the final preparation is composed of two polypeptides with apparent Mr approximately 98,000 (p98) and 102,000 (p102), p98 comprising 70% and p102 30%. Extraction and renaturation of the polypeptides from the gel shows that both p98 and p102 seem to possess topoisomerase activity. Partial proteolytic digestion of p98 and p102 with Staphylococcus aureus V8 and chymotrypsin yielded a series of identical peptides, indicating that the two polypeptides are structurally related. The enzyme sedimented through sucrose density gradient with s20,w of 4.0 S, and thus is monomeric in solution.
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PMID:Rapid purification and characterization of DNA topoisomerase I from cultured mouse mammary carcinoma FM3A cells. 631 74

The highly proliferating phenotype of mammary carcinoma is known to be associated with a particularly aggressive clinical course. We have been interested in the underlying molecular causes that give rise to the increased proliferative activity Proliferative activity was determined immunohistochemically by the detection of topoisomerase II-alpha (Ki-S1). The subgroup of highly proliferating tumors with a Ki-S1 index exceeding 30% was characterized by a high frequency of c-myc amplification and aberrant p53 expression, whereas tumors, with a low mitotic activity rarely exhibited gene amplification or an altered p53 expression. We conclude, that the highly proliferating phenotype is not capable of regular replication and tends to develop gene amplifications. One of the causes might be a defective cell cycle control by p53.
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PMID:[Proliferative activity and defective replication in breast cancer]. 753 9

Azatoxin (NSC 640737), a synthetic molecule, was rationally designed as a topoisomerase-II inhibitor and was shown to be a potent cytotoxic agent that inhibits both tubulin and topoisomerase II. A structure-activity relationship study allowed to select 3 derivatives that inhibit either tubulin (methylazatoxin) only or topoisomerase II (fluoroanilinoazatoxin and nitroanilino-azatoxin) in MTT assays performed on K562 and K562/ADM cells; the latter, expressing P-glycoprotein, indicated cross-resistance of K562/ADM cells to all 4 compounds. DNA double-strand breaks induced by the 3 azatoxins that inhibit topoisomerase II in vitro were decreased in K562/ADM as compared with K562 cells. Nitroanilino-azatoxin was the only compound for which resistance and reduced DNA damage observed in K562/ADM cells was partially reversed by verapamil, suggesting that nitroanilinoazatoxin was a substrate for P-glycoprotein. These results were confirmed by testing the cytotoxic activity of azatoxins on P-glycoprotein-expressing rat colon-carcinoma DHDK12/TRb cells in the absence and the presence of verapamil. Cell-cycle and mitotic-index studies indicated that azatoxin- and methyl-azatoxin-induced M-phase arrest was less in K562/ADM than in K562 cells. The G2 block induced by fluoro- and nitroanilinoazatoxins was delayed in K562/ADM cells. Verapamil increased cell-cycle inhibition induced by nitroanilinoazatoxin in K562/ADM cells without modifying cell-cycle effects of fluoroanilinoazatoxin. These results (i) are consistent with the specific inhibition of topoisomerase II or tubulin by azatoxin derivatives in cells; (ii) indicate that the nitro group of nitroanilinoazatoxin allows recognition and efflux by the P-glycoprotein; and (iii) suggest that cross-resistance of K562/ADM cells to other azatoxin derivatives is not mediated by P-glycoprotein.
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PMID:Cellular pharmacology of azatoxins (topoisomerase-II and tubulin inhibitors) in P-glycoprotein-positive and -negative cell lines. 759 Dec 16

The combination of cis-diamminedichloroplatinum(II) (CDDP) and 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11), a topoisomerase-I inhibitor, has been shown to be synergistic in vitro and clinically active against several human cancers refractory to chemotherapy. To elucidate the mechanism of the synergistic cytotoxicity of CDDP and 7-ethyl-10-hydroxycamptothecin (SN-38), an active metabolite of CPT-11, we studied the interaction of these agents using an HST-1 human squamous-carcinoma cell line. Cells were exposed to the IC50 concentration of SN-38 (5.0 ng/ml) for 1 hr and various concentrations of CDDP for 1 hr in several different treatment schedules. SN-38 augmented the anti-tumor activity of CDDP in all schedules, with maximal synergy observed with simultaneous administration. Evaluation of the kinetics of the removal of DNA interstrand cross-links, measured by alkaline elution, showed significant reduction of this removal in the cells exposed to SN-38 and CDDP, as compared with the cells exposed to CDDP alone. No differences, however, were found in the initially attained level of DNA interstrand cross-links induced by CDDP between these cells. Moreover, the intracellular accumulation of platinum measured by atomic-absorption spectrophotometry, was virtually identical between these cells. These results indicate that SN-38 can modulate the removal of platinum-DNA adducts, thereby potentiating the cytotoxicity of CDDP, suggesting a critical role for topoisomerase I in the repair of DNA interstrand cross-links.
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PMID:Inhibition of cis-diamminedichloroplatinum (II)-induced DNA interstrand cross-link removal by 7-ethyl-10-hydroxy-camptothecin in HST-1 human squamous-carcinoma cells. 760 70

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


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