Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A series of substituted dibenzo[1,4]dioxin-1-carboxamides has been synthesized and evaluated for in vitro and in vivo antitumor activity. The required substituted dibenzo[1,4]dioxin-1-carboxylic acids were prepared by a variety of methods. No regiospecific syntheses were available for many of these, and separation of the mixtures of regioisomers obtained was sometimes difficult. The dibenzo[1,4]dioxin-1-carboxamides are active against wild-type P388 leukemia in vitro and in vivo, with structure-activity relationships resembling those for both the acridine-4-carboxamide and phenazine-1-carboxamide series of DNA-intercalating antitumor agents. In all three series, substituents placed peri to the carboxamide sidechain (the 5-position in the acridines, and the 9-position in the phenazines and dibenzo[1,4]dioxins) enhance activity and potency. The 9-chlorodibenzodioxin-1-carboxamide was also curative against the remotely sited Lewis lung carcinoma. Several of the compounds showed much lower levels of cross-resistance to the P388/AMSA line than classical DNA-intercalating agents, which suggests that their primary mechanism of action may not be via interference with topoisomerase II alpha. This is of interest with regard to the development of drugs to combat resistance mechanisms which arise by the expression of the topo II beta isozyme.
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PMID:Potential antitumor agents. 64. Synthesis and antitumor evaluation of dibenzo[1,4]dioxin-1-carboxamides: a new class of weakly binding DNA-intercalating agents. 131 Jan 19

Topoisomerases catalyse the interconversion of topological isomers of DNA and have key roles in nucleic acid metabolism. Human cells express two distinct type II topoisomerase isozymes, designated topoisomerase II alpha (170 kDa form) and topoisomerase II beta (180 kDa form). We have isolated cDNA clones encoding the beta isozyme from a human B-cell library. The proposed coding region for the topoisomerase II beta protein is 4,863 nucleotides long and would encode a polypeptide with a calculated M(r) of 182,705. The predicted topoisomerase II beta protein sequence shows striking similarity (72% identical residues) to that of the human alpha isozyme, and homology to topoisomerase II proteins from Drosophila, yeast and bacteria. Regions of greatest amino acid sequence divergence lie at the extreme N-terminus and over a C-terminal domain comprising approximately 25% of the total protein. We have quantified the level of topoisomerase II beta mRNA in a panel of human tumour cell lines of different origin using an RNase protection assay, and compared the level to that of topoisomerase II alpha mRNA. Topoisomerase II beta mRNA was expressed in haemopoietic, epithelial and fibroblast cell lines, although to different extents, with U937 cells (promonocytic leukaemia) showing a particularly high level. There was no obvious relationship in terms of level of expression between the topoisomerase II alpha and beta genes. We have localised the gene encoding topoisomerase II beta protein to chromosome 3p24 in the human genome.
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PMID:Isolation of cDNA clones encoding the beta isozyme of human DNA topoisomerase II and localisation of the gene to chromosome 3p24. 133 83

Separate mechanisms underlying the multidrug resistant (MDR) phenotype were identified in 2 independent approaches to select tumour cells resistant to low concentrations of doxorubicin (Dox) from the sensitive T cell leukemia cell line CCRF-CEM. The CEM/A7 cell line was selected at an initial concentration of 0.005 microgram/ml of Dox and maintained at 0.07 microgram/ml. In contrast, the CEM/A5 line was selected using an initial concentration of 0.01 microgram/ml and maintained in Dox at a concentration of 0.05 microgram/ml. P-glycoprotein expression was demonstrated in the CEM/A7 line but not the CEM/A5 line. Amplification of the mdrI gene was not observed in the CEM/A7 cell line. Both cell lines showed cross-resistance to a number of structurally unrelated cytotoxic drugs including anthracyclines and etoposide (VP-16), although only the CEM/A7 line was cross resistant to Vinca alkaloids. Immunoblots of total cell lysates of the CEM/A5 line have revealed almost undetectable levels of topoisomerase II alpha and beta in this line. Cytogenetic analyses of both lines revealed numerous karyotypic abnormalities which were present in the parental cell line as well as both resistant cell lines. The CEM/A7 line also demonstrated a duplication of part of the long arm of chromosome 7 which included the region containing the mdrI gene, a finding not seen in the parental or CEM/A5 line. CEM/A5, however, demonstrated an abnormality of chromosome 7, outside the region of the mdrI gene, and it also contained a deletion of the short arm of chromosome 2. Abnormalities in this latter region of genome have been associated with non-P-glycoprotein-mediated MDR.
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PMID:Cellular and karyotypic characterization of two doxorubicin resistant cell lines isolated from the same parental human leukemia cell line. 751 53

DNA topoisomerase II is a major protein of the nuclear matrix. The enzyme appears to have a central role in both DNA organization and replication. The importance of nuclear matrix topoisomerase II alpha as a target for certain anticancer agents was evaluated in CEM human leukemia cells. Studies were done to determine the extent to which the alpha (170 kDa) and beta (180 kDa) isozymes of topoisomerase II form covalent enzyme-DNA complexes in whole cells and in the nuclear matrix and nonmatrix fractions of CEM cells that are either sensitive or resistant to topoisomerase II-active anticancer agents. Topoisomerase II alpha was detected in both the high salt-soluble (nonmatrix) and matrix fractions of nuclei from parental CEM cells. Most of the matrix topoisomerase II alpha was tightly bound to DNA in cells incubated with VM-26. In contrast, topoisomerase II beta was detected only in the high salt-soluble (nonmatrix) fraction of the nucleus. The subnuclear distribution of the alpha and beta topoisomerase II isozymes in CEM/VM-1 cells resistant to topoisomerase-active drugs was similar to that in drug-sensitive CEM cells. However, the amount and activity of topoisomerase II alpha in nuclear matrices of CEM/VM-1 cells were decreased 3- to 6-fold relative to that of CEM cells. The differences observed in the subnuclear distribution and DNA binding pattern of the topoisomerase II isozymes support the hypotheses that each isozyme has a distinct cellular function. Furthermore, these results provide evidence that topoisomerase II alpha is the nuclear matrix target for VM-26, and that depletion of the nuclear matrix isozyme contributes to cellular resistance to this anticancer agent.
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PMID:DNA topoisomerase II isozymes involved in anticancer drug action and resistance. 757 48

Topoisomerase II alpha is an essential nuclear enzyme involved in DNA replication and a target for many of the clinically useful antineoplastic agents. In a mitoxantrone-selected human leukemia cell line, HL-60/MX2, cellular topoisomerase II (topo II) catalytic activity is decreased, in association with the finding of reduced nuclear topo II alpha and beta protein levels. In addition, HL-60/MX2 cells contain a novel M(r) 160,000 topo II alpha-related protein that localizes predominantly to the cell cytoplasm (W. G. Harker et al., Biochemistry, 30: 9953-9961, 1991). In these studies, we have investigated the molecular mechanisms underlying the altered expression of the topo II alpha protein(s) in these cells. Three topo II alpha mRNAs, 7.2, 6.3, and 4.8 kb, were identified in the HL-60/MX2 cells, with the 6.3 and 4.8 kb transcripts being present in roughly equivalent amounts, while the 7.2-kb mRNA represents < 7% of the total topo II alpha-specific mRNA. Portions of the 3'-coding and 3'-untranslated regions were found to be missing from the 7.2- and 4.8-kb topo II alpha mRNAs by Northern blot analysis. Sequences encoding the 3' regions of the normal and truncated forms of the topo II alpha enzyme were obtained from the HL-60/MX2 cells through the use of a 3'-rapid amplification of cDNA ends strategy. Approximately 1321 nucleotides are missing from the 3'-coding and 3'-untranslated regions of the 4.8-kb mRNA and are replaced by 122 nucleotides that contain an in-frame stop codon and consensus polyadenylation signal. The translation product of the truncated 4388-bp topo II alpha transcript would have a predicted M(r) of 157,850, with 108 COOH-terminal amino acids being replaced by 13 novel residues. Immunoblot analysis confirmed that amino acids in the COOH-terminal region of topo II alpha were missing from the M(r) 160,000 HL-60/MX2 protein, and antisera generated to a synthetic peptide representing the 13 unique amino acids identified a M(r) 160,000 protein in nuclear extracts from these cells. PCR evaluation of the organization of the 3' region of the topo II alpha gene revealed that the 4.8-kb mRNA found in HL-60/MX2 cells diverges from that of the 6.3-kb mRNA at a consensus exon-intron splice donor site. The 122-bp novel nucleotides identified in the truncated transcript appear to originate from an adjacent intron as a result of altered RNA processing. These studies suggest that as a result of the disruption of the carboxy terminus of the topo II alpha protein and the putative nuclear targeting sequences identified therein, cellular localization of the protein is altered, which may confer a growth advantage for the HL-60/MX2 cells in the presence of mitoxantrone.
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PMID:Selective use of an alternative stop codon and polyadenylation signal within intron sequences leads to a truncated topoisomerase II alpha messenger RNA and protein in human HL-60 leukemia cells selected for resistance to mitoxantrone. 758 37

Topoisomerase (topo) inhibitors induce enzyme-linked DNA breaks. Resulting DNA damage can lead to cell cycle arrest and/or cell death by apoptosis. The sensitivity of five human leukemic cell lines to topo I (camptothecin or CPT) and topo II (etoposide or VP-16) inhibitors varied widely (100-fold for CPT and 30-fold for VP-16). Three cell lines were more sensitive (BV173, HL60, U937) and two cell lines were resistant (K562, KCL22) to both drugs. None of these cell lines were selected for drug resistance and overexpressed mdr1 gene. Their sensitivity was not related to their doubling time nor to cell cycle repartition. The initial DNA damage (cleavable complexes) induced by topo I and II inhibitors was measured as DNA-protein crosslinks (DPC) using alkaline elution. Neither DPC level induced by 30-min treatment with CPT or VP-16 nor the levels of topo 1, topo II alpha and topo II beta mRNA were related to sensitivity. Electron microscopy and DNA fragmentation measured by filter elution and agarose gel electrophoresis demonstrated that apoptosis was induced by both drugs in the five cell lines. The kinetics of DNA fragmentation was related to cell sensitivity. At drug concentrations higher than IC50, DNA fragmentation increased very rapidly in the three sensitive, compared with the two resistant, cell lines. Continuous exposure to both drugs induced cell cycle arrest in either G2 or S phase that was related both to cell sensitivity and drug concentration. Comparison between cell lines indicated that the ability of cells to arrest cell cycle in G2 or S phase was related to their drug sensitivity and increased with cell resistance. In a given cell line, cell cycle progression was observed to be progressively inhibited by increasing drug concentrations. Treatment of synchronized cells demonstrated that highly cytotoxic drug concentration induced a complete inhibition of cell cycle progression. Altogether, these data suggest that the ability of leukemic cell lines to regulate cell cycle progression and to trigger apoptosis is more indicative of their sensitivity to topoisomerase poisons than cleavable complexes induced by these drugs.
Leukemia 1995 Jun
PMID:The role of cell cycle regulation and apoptosis triggering in determining the sensitivity of leukemic cells to topoisomerase I and II inhibitors. 759 66

A human HL-60 leukemia cell line selected for resistance to mitoxantrone, HL-60/MX2, displays cross-resistance only to agents whose cytotoxicities result from interaction with the nuclear enzyme DNA topoisomerase II (topo II). The topo II catalytic activity is reduced 2-fold in the drug-resistant cell line in association with the absence of the M(r) 180,000 isoform of topo II and the finding of novel M(r) 160,000 topo II alpha-related immunoreactive protein in these cells by immunoblot. The topo II alpha (M(r) 170,000) protein levels in nuclear extracts from the HL-60/MX2 cells were noted on average to be approximately 40% lower than in comparable HL-60 nuclei. Studies of the subcellular localization of topo II by immunohistochemical and fractional extraction techniques demonstrated that the M(r) 160,000 topo II alpha-related protein is primarily localized in the cytoplasm. Levels of the 6.3-kilobase topo II alpha mRNA were noted to be reduced 2-fold in the HL-60/MX2 cells in association with the finding of a novel 4.8-kilobase topo II alpha-related mRNA transcript that was present in HL-60/MX2 but not HL-60 cells. The absence of topo II beta protein in nuclear and whole cell extracts from the HL-60/MX2 cells was associated with the virtual absence of detectable topo II beta mRNA in those cells by Northern blot analysis. Using a reverse transcription-PCR assay we were able to demonstrate the presence of very low levels of topo II beta mRNA in HL-60/MX2 cells, representing < 1% of that found in the HL-60 cells. In contrast, the nuclear catalytic activity and cellular mRNA levels of the related nuclear enzyme DNA topoisomerase I were nearly identical in the two cell types. Southern blot analysis of DNA extracted from the drug-sensitive and drug-resistant cells revealed a structural alteration in one topo II alpha allele in the HL-60/MX2 cells, but there was no evidence of rearrangement or hypermethylation of the topo II beta locus. These results indicate that the reduced levels of topo II alpha and beta isoenzymes observed in mitoxantrone-resistant HL-60/MX2 cells are related to changes in the levels of their respective mRNA transcripts. The identification of structural changes in one topo II alpha allele in the HL-60/MX2 cell line suggests that the altered allele may serve as the source of the unique 4.8-kilobase topo II alpha-related mRNA transcript and the M(r) 160,000 protein discovered in those cells.
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PMID:Alterations in the topoisomerase II alpha gene, messenger RNA, and subcellular protein distribution as well as reduced expression of the DNA topoisomerase II beta enzyme in a mitoxantrone-resistant HL-60 human leukemia cell line. 771 79

The cytotoxic efficacy of antitumor drugs targeted at DNA topoisomerase II (topo II) in many cases varies in direct proportion to cellular topo II content. To investigate the transcriptional control of the predominant alpha form of topo II, the 5' flanking region of the human topo II alpha gene (positions -562 to +90) was subcloned into a firefly luciferase reporter plasmid and transiently transfected into HL-60 human leukemia cells, a line capable of monocytic differentiation after treatment with various agents. Early in phorbol-12-myristate-13-acetate (30 nM)-induced differentiation (18-24 hr after treatment), an unexpected 3-5-fold activation of topo II alpha gene promoter activity was observed. Activation was observed in HL-60 cells and U-937 cells, but not in HeLa human cervical carcinoma cells. Sodium butyrate (NaB) (0.4 mM) also led to activation (4-17-fold) of the topo II alpha promoter in HL-60 and U-937 cells. Promoter sequences between position -90 and position +90 mediated the inducing effects of NaB. This NaB-dependent promoter-reporter induction was partly mirrored by a transient approximately 2-fold increase in endogenous topo II alpha enzyme. The stimulus for promoter activation could be partly attributed to a 2-fold increase in DNA synthesis at 16 hr for NaB, but not phorbol-12-myristate-13-acetate. Regardless of the primary stimulus for topo II alpha promoter trans-activation, it could be bypassed by treatment of HL-60 cells with NaB for 48 hr before transfection, revealing the expected 60-70% suppression of topo II alpha promoter activity. Further study of topo II alpha promoter down-regulation later in monocytic differentiation may serve as a model for elucidating the transcriptional mechanisms that may also be exploited by tumor cells expressing intrinsic or acquired resistance to topo II-directed drugs.
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PMID:Topoisomerase II alpha promoter trans-activation early in monocytic differentiation of HL-60 human leukemia cells. 772 30

In a series of 60 ALL samples drawn during different stages of the disease we used a cDNA-PCR approach to analyse the relative mRNA levels of the MDR-associated genes encoding mdr1/P-glycoprotein, mrp, and the topoisomerase II isozymes alpha and beta. Expression analysis of the cyclin A gene was included to examine cellular proliferation activity. The expression of gapdh served as an internal standard. Calculating the mean values we found: (i) a distinctly lower mdr1 gene expression in primary ALL and first relapses compared to bone marrow from healthy donors, (ii) no change in mdr1 and mrp, but a decreased topoisomerase II alpha gene expression in first relapses of ALL compared to the primary leukaemia, and (iii) increased mdr1 and mrp levels combined to decreased topoisomerase II alpha levels in recurrent relapses of ALL showing significant correlations (mdr1/mrp: rs = +0.6833, P < 0.05; mdr1/topoII alpha: rs = -0.6727, P < 0.05). The expression of the topoisomerase II alpha gene was correlated to that of cyclin A, indicating a link of its expression to cellular proliferation. Our findings suggest that a multifactorial MDR including mrp appears particularly in recurrent relapses of ALL, which often do not respond to chemotherapy. Nonetheless, some individual samples showed gene expression levels very different from the mean values calculated for a particular state of the leukaemia, indicating the need of an individual expression analysis of MDR-associated genes.
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PMID:Expression of mdr1, mrp, topoisomerase II alpha/beta, and cyclin A in primary or relapsed states of acute lymphoblastic leukaemias. 787 86

The induced expression of multiple drug resistance (MDR)-associated genes as a direct response of tumor cells to antineoplastic drugs could be an important factor influencing the success of cancer chemotherapy. We investigated the effects of such compounds on mdr1/P-glycoprotein (P-gp) gene expression and drug sensitivities in the T-lymphoblastoid human cell line CCRF-CEM and MDR sublines. Thereby, we observed that actinomycin D or adriamycin administered at sublethal concentrations induced increases of mdr1 mRNA levels and resistance within 72 h. Furthermore, on leukemia cell samples collected before and after chemotherapy we checked by a complementary DNA polymerase chain reaction (cDNA-PCR) approach for similar alterations in the relative expression levels of the MDR-associated genes (a) mdr1/P-gp (b) mrp (MDR related protein), and (c) the topoisomerase II isoforms alpha and beta. We found a concomitant increase in mdr1 and mrp gene expression combined with a decreased expression of topoisomerase II alpha in the course of the second relapse of an acute lymphoblastic leukemia (ALL). This points to the emergence of at least three different MDR mechanisms in this type of leukemia unresponsive to chemotherapy. A chronic myeloid leukemia (CML) in blast crisis, however, showed combined increases in mdr1 (about 20-fold) and mrp (about four fold) gene expression after intense but unsuccessful chemotherapy over a 6-month period. Our results indicate the occurrence of induced resistance in vitro and in vivo and suggest a contribution of the newly identified ATP-binding cassette (ABC) transporter MRP in MDR.
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PMID:Drug-induced changes in the expression of MDR-associated genes: investigations on cultured cell lines and chemotherapeutically treated leukemias. 791 48


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