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)

VP-16 (etoposide) has recently been shown to induce topoisomerase II (TOP2)-mediated DNA cleavage within the mixed lineage leukemia (MLL) breakpoint cluster region (bcr), suggesting a role of TOP2 in MLL gene rearrangement. In our current studies, we have compared the induction of DNA cleavage within the MLL bcr in different cell lines after treatment with various anticancer drugs. All anticancer drugs tested including VP-16 (a TOP2-directed drug), camptothecin (a topoisomerase I-directed drug), 5-fluorouracil and methotrexate (antimetabolites), and vinblastine (a microtubule inhibitor) induced the same site-specific cleavage within the MLL bcr. This cleavage was shown to be nuclease-mediated but not TOP2-mediated by the following observations: 1) drug-induced cleavage within the MLL bcr was not protein-linked; 2) unlike TOP2-mediated cleavage, drug-induced DNA cleavage within the MLL bcr was kinetically slow and coincided with the formation of the apoptotic nucleosomal DNA ladder; 3) drug-induced cleavage within the MLL bcr was unaffected in cells with reduced nuclear TOP2; and 4) drug-induced cleavage within the MLL bcr was abolished by the caspase inhibitor, Z-Asp(OCH(3))-Glu(OCH(3))-Val-Asp(OCH(3))-FMK. The possibility that an apoptotic nuclease may be involved in cleavage of the MLL bcr and MLL gene translocation is discussed.
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PMID:Nucleolytic cleavage of the mixed lineage leukemia breakpoint cluster region during apoptosis. 1140 28

The correlation between infant leukemia and in utero exposure to topoisomerase II (topo-II) inhibitor has been clarified. We examined the in vitro effect of topo-II inhibitor (etoposide) on cleavage of the MLL gene in cord and peripheral blood mononuclear cells (MNCs). Southern blot analysis showed cleavage of the MLL gene in peripheral blood MNCs of infants when the MNCs were exposed to etoposide. MNCs were incubated with etoposide at various concentrations (1 to 50 microM), and a ligation-mediated polymerase chain reaction (LM-PCR) was used to detect double strand breaks (DSBs) of DNA in intron 8 of the MLL breakpoint cluster region. PCR products obtained with LM-PCR were subcloned and sequenced to identify the breakpoint in the MLL gene. The PCR products indicated DSBs of the MLL gene were obtained without any difference in the incidence between 3 different samples (cord and peripheral blood from infants and children). Sequencing analysis showed that the DSBs occurred on the telomeric side of intron 8 and near exon 9. There was no evidence that the cord blood was more susceptible to MLL DNA breakage by topo-II inhibitor than were other cells. Instability of the partner gene during the fetal period could be associated with the pathogenesis of infant leukemia.
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PMID:In vitro cleavage of the MLL gene by topoisomerase II inhibitor (etoposide) in normal cord and peripheral blood mononuclear cells. 1213

MLL gene fusions are the hallmark of more than 70% of therapy-related leukemias (t-ML) associated with topoisomerase II inhibitors (e.g., etoposide) and cause leukemia in murine transgenic models. To determine whether Mll genomic fusions can occur after exposure to topoisomerase II inhibitors, we developed a long-distance inverse PCR DNA-based assay for chimeric Mll fusions in mouse embryonic stem cells. We detected Mll fusions at a higher frequency following 100 microM etoposide for 8 h (16x10(-6) cell(-1)) than in no-drug controls (1.0x10(-6) cell(-1), P=0.0002) or after treatment with a comparably cytotoxic exposure to the antimicrotubule drug vincristine (1.0x10(-6) cell(-1), P=0.0047). The fusion points in Mll chimeric products induced by etoposide were localized to a 1.5 kb region between exons 9 and 11, analogous to the MLL breakpoint cluster region in human leukemia. All 49 Mll fusion partners analyzed matched known genomic murine sequences, with 40 (82%) matching annotated genes covering eighteen murine autosomes. One partner was Runx1, the murine homologue of the transcription factor AML-1, a target of human translocations in therapy-related leukemia. These findings indicate that etoposide triggers the formation of Mll gene fusions, a critical step for the development of treatment-induced leukemic transformation.
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PMID:Etoposide induces chimeric Mll gene fusions. 1463 Jun 94

The recurring chromosome translocation t(11;16)(q23;p13) is detected in leukemia patients, virtually all of whom have received previous chemotherapy with topoisomerase (topo) II inhibitors. In the t(11;16), 3' CBP, on 16p13, is fused to 5' MLL, on 11q23, resulting in an MLL-CBP fusion gene that plays an important role in leukemogenesis. In this study, we cloned genomic breakpoints of the MLL and CBP genes in the t(11;16) in the SN-1 cell line and in five patients with therapy-related leukemia, all of whom had received topo II inhibitors for previous tumors. In all patients except one, both the genomic MLL-CBP and the reciprocal fusions were cloned. Genomic breakpoints in MLL occurred in the 8.3-kb breakpoint cluster region in all patients, whereas the breakpoints in CBP clustered in an 8.2-kb region of intron 3 in four patients. Genomic breakpoints in MLL occurred in intron 11 near the topo II cleavage site in the SN-1 cell line and in one patient, and they were close to LINE repetitive sequences in two other patients. In the remaining two patients, genomic breakpoints were in intron 9 in Alu repeats. Genomic breakpoints in CBP occurred in and around Alu repeats in one and two patients, respectively. In two patients, the breaks were near LINE repetitive sequences, suggesting that repetitive DNA sequences may play a role. No specific recombination motifs were identified at or near the breakpoint junctions. No topo II cleavage sites were detected in introns 2 and 3 of CBP. However, there were deletions and duplications at the breakpoints in both MLL and CBP and microhomologies or nontemplated nucleotides at most of the genomic fusion junctions, suggesting that a nonhomologous end-joining repair mechanism was involved in the t(11;16).
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PMID:Characterization of genomic breakpoints in MLL and CBP in leukemia patients with t(11;16). 1533 49

Exposure to topoisomerase II inhibitors is linked to the generation of leukemia involving translocations of the MLL gene, normally restricted to an 8.3 kbp tract, the breakpoint cluster region (BCR). Using an in vitro assay, apoptotic activators, including radiation and anti-CD95 antibody, trigger site-specific cleavage adjacent to exon 12 within the MLL BCR and promote translocation of the MLL gene in cells that can survive. To explore the mechanism of cleavage and rearrangement in more detail, the entire MLL BCR was placed into the pREP4 episomal vector and transfected into human lymphoblastoid TK6 cells. Episomes containing either the MLL BCR, or deletion constructs of 367 bp or larger, were cleaved at the same position as genomic MLL after exposure to apoptotic stimuli. Further analysis of sequence motifs surrounding the cleaved region of MLL showed the presence of both a predicted nuclear matrix attachment sequence and a potential strong binding site for topoisomerase II, flanking the site of cleavage. Inactivation of topoisomerase II by the catalytic inhibitor merbarone did not inhibit MLL cleavage, suggesting that the initial cleavage step for MLL rearrangement is not mediated by topoisomerase II.
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PMID:Cleavage of the MLL gene by activators of apoptosis is independent of topoisomerase II activity. 1619 84

Phenomena involving the disassembly of chromosomes to approximately 50 kbp double-stranded fragments upon protein denaturing treatments of normal and apoptotic mammalian nuclei as well as yeast protoplasts may be an indication of special, hypersensitive regions positioned regularly at loop-size intervals in the eukaryotic chromatin. Here we show evidence in yeast cell systems that loop-size fragmentation can occur in any phase of the cell cycle and that the plating efficiency of these cells is approximately 100%. The possibility of sequence specificity was investigated within the breakpoint cluster region (bcr) of the human MLL gene, frequently rearranged in certain leukemias. Our data suggest that DNA isolated from yeast cultures or mammalian cell lines carry nicks or secondary structures predisposing DNA for a specific nicking activity, at non-random positions. Furthermore, exposure of MLL bcr-carrying plasmid DNA to S1 nuclease or nuclear extracts or purified topoisomerase II elicited cleavages at the nucleotide positions of nick formation on human genomic DNA. These data support the possibility that certain sequence elements are preferentially involved in the cleavage processes responsible for the en masse disassembly of chromatin to loop-size fragments upon isolation of DNA from live eukaryotic cells.
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PMID:Nick-forming sequences may be involved in the organization of eukaryotic chromatin into approximately 50 kbp loops. 1619 88

Acute leukemias with balanced chromosomal translocations, protean morphologic and immunophenotypic presentations but generally shorter latency and absence of myelodysplasia are recognized as a complication of anti-cancer drugs that behave as topoisomerase II poisons. Translocations affecting the breakpoint cluster region of the MLL gene at chromosome band 11q23 are the most common molecular genetic aberrations in leukemias associated with the topoisomerase II poisons. These agents perturb the cleavage-religation equilibrium of topoisomerase II and increase cleavage complexes. One model suggests that this damages the DNA directly and leads to chromosomal breakage, which may result in untoward DNA recombination in the form of translocations. This review will summarize the evidence for topoisomerase II involvement in the genesis of translocations and extension of the model to acute leukemia in infants characterized by similar MLL translocations.
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PMID:Topoisomerase II and the etiology of chromosomal translocations. 1685 31

Recurring chromosome abnormalities are strongly associated with certain subtypes of leukemia, lymphoma and sarcomas. More recently, their potential involvement in carcinomas, i.e. prostate cancer, has been recognized. They are among the most important factors in determining disease prognosis, and in many cases, identification of these chromosome abnormalities is crucial in selecting appropriate treatment protocols. Chromosome translocations are frequently observed in both de novo and therapy-related acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS). The mechanisms that result in such chromosome translocations in leukemia and other cancers are largely unknown. Genomic breakpoints in all the common chromosome translocations in leukemia, including t(4;11), t(9;11), t(8;21), inv(16), t(15;17), t(12;21), t(1;19) and t(9;22), have been cloned. Genomic breakpoints tend to cluster in certain intronic regions of the relevant genes including MLL, AF4, AF9, AML1, ETO, CBFB, MYHI1, PML, RARA, TEL, E2A, PBX1, BCR and ABL. However, whereas the genomic breakpoints in MLL tend to cluster in the 5' portion of the 8.3 kb breakpoint cluster region (BCR) in de novo and adult patients and in the 3' portion in infant leukemia patients and t-AML patients, those in both the AML1 and ETO genes occur in the same clustered regions in both de novo and t-AML patients. These differences may reflect differences in the mechanisms involved in the formation of the translocations. Specific chromatin structural elements, such as in vivo topoisomerase II (topo II) cleavage sites, DNase I hypersensitive sites and scaffold attachment regions (SARs) have been mapped in the breakpoint regions of the relevant genes. Strong in vivo topo II cleavage sites and DNase I hypersensitive sites often co-localize with each other and also with many of the BCRs in most of these genes, whereas SARs are associated with BCRs in MLL, AF4, AF9, AML1, ETO and ABL, but not in the BCR gene. In addition, the BCRs in MLL, AML1 and ETO have the lowest free energy level for unwinding double strand DNA. Virtually all chromosome translocations in leukemia that have been analyzed to date show no consistent homologous sequences at the breakpoints, whereas a strong non-homologous end joining (NHEJ) repair signature exists at all of these chromosome translocation breakpoint junctions; this includes small deletions and duplications in each breakpoint, and micro-homologies and non-template insertions at genomic junctions of each chromosome translocation. Surprisingly, the size of these deletions and duplications in the same translocation is much larger in de novo leukemia than in therapy-related leukemia. We propose a non-homologous chromosome recombination model as one of the mechanisms that results in chromosome translocations in leukemia. The topo II cleavage sites at open chromatin regions (DNase I hypersensitive sites), SARs or the regions with low energy level are vulnerable to certain genotoxic or other agents and become the initial breakage sites, which are followed by an excision end joining repair process.
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PMID:Chromatin structural elements and chromosomal translocations in leukemia. 1689 85

Etoposide-induced treatment-related acute myelogenous leukemia (t-AML) is characterized by rearrangements of the mixed lineage leukemia (MLL) gene with one of its >50 partner genes, most probably as a consequence of etoposide-induced DNA double-strand breaks (DSBs). Recent studies have shown that etoposide-induced DSBs occur predominantly within the breakpoint cluster region (bcr) of the MLL gene. However, bcr-specific DSBs induced by etoposide are not topoisomerase II-linked but the result of apoptotic nuclease-mediated DNA cleavage. Here, we test the involvement of caspase-activated DNase (CAD) and other apoptotic components in etoposide-induced gene rearrangements using two methods. First, we measured the effect of etoposide on the integration frequency of a transfected plasmid. Etoposide strongly stimulated plasmid integration in CAD cDNA-complemented mouse embryonic fibroblasts (MEFs) but not in CAD knockout (KO) MEFs. Consistently, down-regulation of ICAD (inhibitor of CAD, also required for proper folding of CAD) in an HT29-derived cell line, which leads to decreased CAD activity, significantly reduced etoposide-induced plasmid integration. Second, we used long-template inverse PCR to focus on gene rearrangements at the MLL locus. Etoposide stimulated MLL fusion product formation in CAD cDNA-complemented MEFs but not in CAD KO MEFs. Together, these results suggest that CAD and other apoptotic components may play an important role in etoposide-induced t-AML.
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PMID:Role of apoptotic nuclease caspase-activated DNase in etoposide-induced treatment-related acute myelogenous leukemia. 1698 37

Using semi-quantitative PCR-based approach, we have shown that the breakpoint cluster region of the AML1 gene was associated with the nuclear matrix. We have demonstrated that inhibition of topoisomerase II by etoposide stimulates the appearance of histone gammaH2AX foci, an indicator for the presence of DNA double-strand breaks. Furthermore, the major part of these foci was associated with the nuclear matrix. We also visualized nuclear matrix--associated multiprotein complexes involved in topoisomerase II--induced DNA double-strand break repair. Colocalization studies have demonstrated that these complexes included the principal components of the non-homologous end joining repair system (Ku80, DNA-PKcs and DNA ligase IV). Thus, it is reasonable to suggest that the non-homologous DNA end joining is a possible mechanism of topoisomerase II--induced chromosomal rearrangements.
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PMID:[Illegitimate recombination as a possible mechanism of DNA-topoisomerase II induced chromosomal rearrangements]. 1708 89


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