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)

Rearrangements involving chromosome band 11q23 are very common in acute leukemia, both lymphoblastic and myeloid (monoblastic), and are less common in lymphoma. Although several different genes have been cloned from 11q23 translocation breakpoints, the great majority involve the MLL (myeloid-lymphoid leukemia) gene. The MLL gene has several different names, ALL1, Htrx, HRX; the central part of the gene codes for multiple zinc fingers which show strong homology to the Drosophila trithorax gene. MLL is involved in four common translocations as well as in 25 uncommon or rare translocations, insertions and deletions. The translocation breakpoints occur within an 8.3kb region which can be detected with a 0.7 kb cDNA probe. Twenty-five percent of patients have a deletion 3' of the breakpoint which includes the zinc finger region. Patients who previously received drugs that inhibit topoisomerase II often develop acute leukemia with translocations involving 11q23. These translocations break MLL in the same 8.3kb region. In the four breakpoints cloned to date, the translocation has led to a fusion gene on the derivative 11 chromosome with a chimeric transcript, consisting of 5' MLL and the 3' segment of the other gene. Although transcripts were also cloned from the other derivative chromosome, all the evidence indicates that the critical fusion gene is on the derivative 11 chromosome. The molecular dissection of these rearrangements will provide insights into the biology of MLL and into the interaction of MLL with topoisomerase II inhibitors. In addition, this research has provided DNA probes that will be important for diagnosis and for monitoring patients during the course of their disease.
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PMID:1993 Robert R. deVilliers Lecture. Chromosome translocations: dangerous liaisons. 815 72

The virulence plasmid pXO1 is responsible for toxin production in Bacillus anthracis. A DNA fragment from pXO1 was isolated and was shown, by sequence analysis, to contain part of a type 1 DNA topoisomerase gene. Attempts to clone the entire wild-type gene, designated topX, in Escherichia coli, were unsuccessful. In order to obtain the complete gene, it was first insertionally inactivated and then cloned in the mutated form. The deduced amino acid sequence of Topo X1 shows similarities to that of the two E. coli type 1 DNA topoisomerases. The N-terminal two-thirds of the putative B. anthracis protein exhibits strongest sequence similarity to topoisomerase III, whereas the C-terminal portion contains cysteine residues that could form three zinc-binding domains, as they do in topoisomerase I. The suggested active-site tyrosine is conserved in all three proteins. The regulation of expression from the topX promoter is modified by addition of a gyrase inhibiting antibiotic. The Topo X1 protein is likely to be involved in the stability of pXO1.
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PMID:Bacillus anthracis pXO1 virulence plasmid encodes a type 1 DNA topoisomerase. 815 71

Etoposide, a DNA topoisomerase II inhibitor, caused a concentration-dependent induction of apoptosis in immature thymocytes. Using a flow cytometric method to separate and quantify normal and apoptotic cells, etoposide-induced apoptosis was inhibited by cycloheximide and actinomycin D but not by zinc. Etoposide induced a marked cleavage of DNA into nucleosomal length fragments or multiples thereof, which was completely inhibited if the thymocytes were also incubated in the presence of zinc. Etoposide, alone, induced the classical ultrastructural features of apoptosis, but in the presence of zinc, the morphological pattern was markedly different and dominated by discrete clumps of condensed chromatin abutting the nuclear membrane. These latter changes resemble those described as the earliest changes in apoptosis. These results support the hypothesis that, in the induction of apoptosis, critical alterations in nuclear chromatin occur prior to endonuclease cleavage of DNA into nucleosomal fragments.
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PMID:Changes in nuclear chromatin precede internucleosomal DNA cleavage in the induction of apoptosis by etoposide. 830 63

As a probe of whether RAG-1 and RAG-2 gene products activate other genes or form part of the recombinase itself, certain mutants of the RAG genes were assayed for their ability to activate variable-diversity-joining region [V(D)J] recombination in a plasmid substrate in fibroblasts. The results indicate that the N-terminal one-third of RAG-1, including a zinc-finger-like domain, and an acidic domain of RAG-2 are dispensable for activating V(D)J recombination in a fibroblast, although they contribute quantitatively. In contrast, deletion of the C-terminal segment of RAG-1, which has homology to a topoisomerase-like protein from yeast, abolished recombination activation. These results do not support the hypothesis that the RAG gene products are transcription factors and suggest the possibility that they are parts of the recombination machinery.
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PMID:Dispensable sequence motifs in the RAG-1 and RAG-2 genes for plasmid V(D)J recombination. 832 89

Escherichia coli DNA topoisomerase I contains three Zn(II) in each enzyme molecule required for relaxation of negatively supercoiled DNA. Apoenzymes were prepared from both the intact topoisomerase (M(r) 97,000) and the truncated active form top85 (M(r) 85,000) that lacks the carboxyl terminal domain but still contains the three Zn(II). Fluorescence and circular dichroism spectroscopy were used to compare the apoenzymes with topoisomerase and top85 reconstituted with Zn2+. The results indicated structural changes affecting the environment of the tryptophan residues and increasing the alpha-helical and beta-sheets content of the protein occurred upon zinc removal. These structural changes probably account for the loss of enzyme activity.
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PMID:Effect of zinc removal on the conformation of Escherichia coli DNA topoisomerase I. 838 Sep 65

Anthracenyl-amino acid conjugates represent a novel chemical class of topoisomerase (topo) inhibitor. NU/ICRF 505 is a lead compound that stabilises topo I cleavable complexes and is actively cytotoxic at low microM concentrations. In this study, endonucleolytic DNA cleavage was used as a marker of apoptosis to investigate mechanisms of cell death produced by this compound. NU/ICRF 505 (5 microM) induced a substantial increase in the level of DNA fragmentation in HL60 cells (up to 30% of total extracted DNA) but only after a 48 and 72 h drug exposure (compared with 6 h after treatment with camptothecin), as determined qualitatively by conventional gel electrophoresis and quantitatively by spectrofluorimetry. This effect was substantially reversed by co-treatment with zinc (1 mM). Subsequent studies with the human lung (NX002), ovarian (A2780) and colon (HT29) cancer cell lines yielded evidence of formation of higher molecular weight DNA fragments in NX002 and A2780 cells in response to NU/ICRF 505 (5 microM). Co-treatment with zinc (1 mM) caused a small decrease in DNA fragmentation. These data suggest that the induction of apoptosis may play an important role in the mechanism of cytotoxicity of NU/ICRF 505 in HL60 cells and that other pathways of cell death may also be operative in NX002 and A2780 in conjunction with apoptosis.
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PMID:Induction of apoptosis in human cancer cell lines by the novel anthracenyl-amino acid topoisomerase I inhibitor NU/ICRF 505. 869 51

A filter binding assay that measures internucleosomal DNA fragmentation associated with apoptosis is described. The assay is based on a novel principle that consists of using simultaneously two kinds of glass fiber filters to harvest [3H]thymidine-prelabeled cells following their incubation with inducers of apoptosis. One filter, which is neutral, traps intact chromatin and high-molecular-weight DNA. The other filter, which is positively charged with DEAE active groups, traps low-molecular-weight DNA fragments. DNA fragmentation is quantified by measuring the radioactivity retained by each of the filters. The assay was evaluated with the histiocytic lymphoma cell line U937 and the topoisomerase inhibitors camptothecin, etoposide, and doxorubicin. These agents caused a dose-dependent decrease of radioactivity in the neutral filter and a parallel increase of radioactivity in the DEAE filter. Irradiation-induced single strand breaks and topoisomerase-mediated primary DNA damage were not detected by this method. Consistent with the detection of internucleosomal DNA fragmentation, the effects measured by this assay were prevented by the endonuclease inhibitor zinc acetate and by the metabolic inhibitor sodium azide. Results obtained using this assay were validated by observation of DNA ladders on agarose gels and by morphologic examination of apoptotic features. Evaluation of the assay in a mock screen demonstrated that the introduction of the DEAE filter increases the assay sensitivity and eliminates false positives. Thus, this assay may be used in high-throughput screening approaches to discover novel modulators of apoptosis.
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PMID:A glass fiber/diethylaminoethyl double filter binding assay that measures apoptotic internucleosomal DNA fragmentation. 893 61

2,3-(Methylenedioxy)-5-methyl-7-hydroxy-8-methoxybenzo[c]phenanthr idinium hydrogensulfate dihydrate, called NK109, is a benzo[c]phenanthridine derivative, which inhibits DNA topoisomerase II activity by stabilizing the DNA-enzyme-drug complex, and shows strong growth-inhibitory effects on several human cancer cells. In the present study, NK109 treatment induced DNA fragmentation and a rise in the level of cytoplasmic nucleosomes, which are markers of apoptosis, in human small-cell lung carcinoma SBC-3 cells. These effects were inhibited by zinc ions and enhanced by cycloheximide or actinomycin D. Dose-dependent single- and double-strand DNA breaks were observed, using alkaline and neutral elution assays, in SBC-3 cells treated with more than 0.2 microM NK109 for 4 h. Treatment with NK109 caused more DNA single- and double-strand breaks than treatment with an equimolar amount of VP-16. These results suggest that NK109 induces DNA strand breaks and apoptosis. In addition, it appears that this process does not require protein or RNA synthesis, but involves a specific endonuclease which is inhibited by zinc ions.
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PMID:A topoisomerase II inhibitor, NK109, induces DNA single- and double-strand breaks and apoptosis. 895 68

GL331 is a semisynthetic topoisomerase II inhibitor derived from a plant toxin podophyllotoxin. In 72-h exposure assays, LD50 values of GL331 range from 0.5 to 2 microM, which are three- to ten-fold lower than those of its homologous compound etoposide (VP-16), depending on different cancer cell lines including nasopharyngeal, hepatocellular, gastric, cervical and colon cancer types. Apoptotic DNA ladders could be detected when cancer cells were treated with GL331 for 24 h even if the Bcl-2 and Bax protein levels were not altered during the period. Besides acting as topoisomerase II inhibitors, both GL331 and VP-16 decrease the cellular protein tyrosine kinase (PTK) activities in cancer cells. The activities of protein tyrosine phosphatase (PTP) are significantly increased after GL331 treatment but are not affected by VP-16. GL331-induced internucleosomal cleavage can be efficiently prevented by two inhibitors of PTP, sodium orthovanadate and zinc chloride, but not by okadaic acid, which inhibits serine/threonine phosphatase activity. These results indicate that GL331 may induce apoptotic cell death, and that activation of protein tyrosine phosphatases may be involved in this process.
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PMID:Protein tyrosine phosphatase activities are involved in apoptotic cancer cell death induced by GL331, a new homolog of etoposide. 901 84

HER2 (erbB-2) proto-oncogene amplification and/or overexpression correlate with poor prognosis in many malignancies. The precise biological role of this oncogenic signaling pathway (which also involves the HER4 gene) in breast cancer is unclear. One property conferred by this oncogene relates to response to drug therapy. Clinical studies support an association between HER2 overexpression and resistance to alkylating agents (cisplatinum and cyclophosphamide). Data from the Cancer and Leukemia Group B 8869/8541 study indicate enhanced dose responsiveness to doxorubicin (Adriamycin) in patients who overexpress the HER2 receptor. Heregulin beta-2, a naturally occurring ligand that activates the HER2 receptor by inducing its heterodimerization with the HER4 receptor, has recently been cloned. The ability of this ligand to phosphorylate the HER2 receptor exogenously allows us to study the effect of HER2 activation on cancer cell behavior. To study the relationship between chemotherapy response and activation of HER2, MCF-7 cells expressing biologically active heregulin were assessed for response to doxorubicin and etoposide, both of which are topoisomerase IIalpha (topo IIalpha) inhibitors. Several clones show markedly increased sensitivity to these drugs. In addition, the same wild-type MCF-7 cells transfected with heregulin beta-2 under the control of an inducible promoter also show this dose-response relationship to doxorubicin after the expression of heregulin beta-2 is activated by zinc. The modulation of topo IIalpha was studied in the cell lines transfected with heregulin. topo IIalpha mRNA and protein (total protein and enzymatic decatenating activity) were found to be up-regulated in heregulin beta-2-transfected cells. Moreover, topo IIalpha promoter activity was also modestly increased in heregulin beta-2-transfected cells. Because up-regulation of topo IIalpha in vitro and in clinical specimens is associated with increased response to doxorubicin (presumptively by an increase in drug substrate), this may be the mechanism of the increased sensitivity to doxorubicin seen in heregulin beta-2-transfected cells. This implies that activation of HER2 or one of the other members of the receptor family may increase sensitivity to doxorubicin by up-regulation of topo IIalpha. This finding suggests the use of receptor/ligand expression to direct patient-specific therapeutic choices (e.g., doxorubicin versus alkylator-based regimens) and the use of biological agents (such as heregulin) in combination with certain chemotherapeutic agents to enhance response to treatment in breast cancer patients.
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PMID:Induction of sensitivity to doxorubicin and etoposide by transfection of MCF-7 breast cancer cells with heregulin beta-2. 956 96


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