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)

DNA cleavage stimulated by different topoisomerase II inhibitors shows in vitro a characteristic sequence specificity. Since chromatin structure and genome organization are expected to influence drug-enzyme interactions and repair of drug-induced DNA lesions, we investigated topoisomerase II DNA cleavage sites stimulated by teniposide (VM-26), 4-demethoxy-3'-deamino-3'-hydroxy-4'-epi-doxorubicin (dh-EPI, a doxorubicin derivative), 4'-(9-acridinylamino)-methanesulfon-m-anisidide, and amonafide in the histone gene locus and satellite III DNA of Drosophila cells with Southern blottings and genomic sequencing by primer extension. VM-26 stimulated cleavage in the satellite III DNA, whereas the other studied drugs did not. All four drugs stimulated cleavage in the histone gene cluster, but they yielded drug-specific cleavage intensity patterns. Cleavage sites by dh-EPI and VM-26 were sequenced in the histone H2A gene promoter and were shown to be distinct. DNA cleavage analysis in cloned DNA fragments with Drosophila topoisomerase II showed that drugs stimulated the same sites in vivo and in vitro. Strand cuts were in vivo staggered by 4 bases, and base sequences at major dh-EPI and VM-26 sites completely agreed with known in vitro drug sequence specificities. Moreover, DNA cleavage reverted faster in the satellite III than in the histone repeats. While stimulating similar levels of DNA breakage in bulk genomic DNA, dh-EPI and VM-26 markedly differed for cleavage extent and reversibility in specific chromatin loci. The results demonstrate a high heterogeneity in the localization, extent, and reversibility of drug-stimulated DNA cleavage in the chromatin of living cells.
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PMID:Drug-specific sites of topoisomerase II DNA cleavage in Drosophila chromatin: heterogeneous localization and reversibility. 862 May 4

To define the sites of topoisomerase II activity in two genomic regions of Drosophila melanogaster Kc cells, we have investigated in vivo DNA cleavage sites stimulated by three poisons with diverse sequence specificity, clerocidin, VM-26 and dh-EPI (an anthracycline analog). DNA cleavage was studied by PFGE (pulse-field gel electrophoresis), standard gel electrophoresis, and by genomic primer extension. Poisons stimulated specific intensity patterns of cleavage in the two genomic regions studied. At the centromeric satellite III DNA, fragments of about 270-310 and 385-430 kb could be detected specifically after treatment with clerocidin, suggesting a complex DNA loop organisation, which may correspond with a centromere-specific higher-order chromatin structure. Clerocidin-dependent DNA fragmentation was detectable by PFGE but not by standard agarose gel electrophoresis; while VM-26-dependent cleavage was detected with either method, dh-EPI was ineffective at this locus. Thus, clerocidin DNA cleavage sites were rarer than those of VM-26 at the satellite locus. In the histone H2A-H2B intergenic region, clerocidin and dh-EPI stimulated cleavage whereas VM-26 was only weakly effective. Some clerocidin cleavage sites did not undergo spontaneous reversion, indicating that this agent can stimulate irreversible cleavage in vivo. Direct genomic sequencing showed that many clerocidin and dh-EPI sites, although distinct, mapped to the transcription start and to the proximal promoter of the H2A gene, suggesting that the region is highly accessible to topoisomerase II. Thus, the enzyme may play a role in maintaining a highly accessible chromatin structure under normal cell growth conditions, possibly mediated by specialised protein complexes. This study demonstrates that the use of distinct poisons greatly improves the definition of genomic sites of topoisomerase II activity.
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PMID:Genomic sites of topoisomerase II activity determined by comparing DNA breakage enhanced by three distinct poisons. 987 28

Anthracyclines exert antitumor activity by stimulating site-selective DNA cleavage by topoisomerase II (top2). DNA cleavage sites stimulated by two anthracycline analogues, dh-EPI and da-IDA, were investigated at the histone gene cluster of cultured Drosophila Kc cells. The two agents stimulated analogue-specific patterns of double-stranded DNA cleavage in Kc cell chromatin. Analyses of 47 base sequences of dh-EPI sites showed that the analogue largely followed the in vitro selectivity rule, the requirement of (5')TA at 3' ends of cleaved strands. da-IDA was more selective than dh-EPI, and thus fewer sites could be collected. Nevertheless, base sequences were consistent with its in vitro base preferences. DNA cleavage was then studied in vitro with Drosophila and human top2 isoforms. The tested drugs stimulated distinct in vitro patterns that corresponded to the in vivo patterns. Human top2alpha promoted cleavage patterns that were much more similar to those of Drosophila top2 (both in vitro and in vivo) than human top2beta. Moreover, da-IDA showed a marked site-dependent preference for human top2beta. Thus, DNA site selection in vivo is different for the test anthracyclines, and together with a degree of beta-form specificity, may affect drug activity in human cells.
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PMID:In vivo site specificity and human isoenzyme selectivity of two topoisomerase II-poisoning anthracyclines. 1091 49

Trastuzumab is used for breast cancer patients with high expression levels of HER2 (human epidermal growth factor receptor 2)/neu; however, it has no effect on cancers with low levels of HER2/neu. SM (solamargine), a major steroidal alkaloid glycoside purified from Solanum incanum, triggered apoptosis of breast cancer cells (MCF-7 and SK-BR-3 cells) and non-cancerous breast epithelial cells (HBL-100 cells) within 3 h. To extend the application of trastuzumab in breast cancer patients, the regulation of HER2/neu expression by SM was investigated. SM significantly up-regulates HER2/neu expression in breast cancer cells with low and high expression levels of HER2/neu, and synergistically enhanced the effect of trastuzumab in inhibiting cell proliferation. Additionally, HER2/neu and TOP2A [TopoII (topoisomerase II) alpha] genes share the same amplicon on an identical chromosome. Notably, SM co-regulates HER2/neu and TopoIIalpha expression markedly, and enhances TopoII inhibitor-EPI (epirubicin)-induced cytotoxicity to breast cancer cells.
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PMID:Solamargine induces apoptosis and enhances susceptibility to trastuzumab and epirubicin in breast cancer cells with low or high expression levels of HER2/neu. 1869 74