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)

The expression of the resistance-related proteins P-glycoprotein 170 (P-170), glutathione-S-transferase pi (GST-pi), topoisomerase II (Topo II), thymidylate synthase (TS) and metallothionein (MT) was investigated in leukemic cells of 19 children with newly diagnosed acute nonlymphoblastic leukemia. P-170 was expressed in 84%, GST-pi in 37%, TS in 47%, MT in 68%, and Topo II was downregulated in 37% of the cases investigated. No resistance factors were found in two patients, one positive factor was found in two patients, three factors in three patients, four factors in 7 patients, and all resistance factors investigated were present in one patient. Patients who developed a relapse expressed more than two resistance mechanisms significantly more often than patients who remained in remission (p = 0.005). The probability of continuous first remission was significantly lower where more than two resistance mechanisms were expressed. The results indicate that the higher the number of resistance-related proteins in childhood ANLL the poorer the prognosis of the patients.
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PMID:Multiple resistance mechanisms in acute nonlymphoblastic leukemia (ANLL). 961 93

The expression of seven enzymes involved in the biosynthesis of DNA was measured in HL-60 promyelocytic leukemia cells treated with dimethylsulfoxide (DMSO) or all-trans retinoic acid (RA) to gain information on their role in the termination of proliferation in cells undergoing granulocytic differentiation. The steady-state levels of the mRNAs for topoisomerase I, topoisomerase II. DNA polymerase-alpha, thymidylate synthase, thymidine kinase and hypoxanthine-guanine phosphoribosyltransferase progressively declined from day 3 to day 7 of exposure to the polar solvent or the retinoid suggesting that the expression of these enzymes is coordinately regulated. In contrast, a pronounced difference between the two inducers of differentiation occurred in the expression of the mRNA of the M2 subunit of ribonucleotide reductase, with DMSO causing virtually complete inhibition of the expression of the M2 subunit of the enzyme from day 5 through day 7, with no change in the steady-state levels of the mRNA being produced by retinoic acid. Measurement of the enzymatic activities of two of these catalysts, thymidylate synthase and thymidine kinase, in cells exposed to the two inducers of maturation corroborated the findings at the level of the mRNAs, with corresponding decreases in the activity of these enzymes. The findings collectively demonstrate that the down-regulation of the expression of a relatively wide variety of enzymes involved in DNA replication occurs as late events in the granulocytic differentiation of HL-60 cells, ensuring that cellular replication cannot occur in terminally differentiated cells.
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PMID:Regulation of the expression of enzymes involved in the replication of DNA in chemically-induced granulocytic differentiation of HL-60 leukemia cells. 968 95

The expression of a number of housekeeping enzymes of DNA biosynthesis was measured in HL-60 promyelocytic leukemia cells undergoing monocytic/macrophagic differentiation following treatment with 12-O-tetradecanoylphorbol 13-acetate (TPA) or 1alpha,25-dihydroxyvitamin D3 (vitamin D3). Progressive decreases in the steady-state levels of the mRNAs for thymidylate synthase, topoisomerase II, and hypoxanthine guanine phosphoribosyltransferase occurred following exposure to TPA or vitamin D3. In contrast, the steady-state levels of the mRNAs for thymidine kinase, topoisomerase I, and DNA polymerase-alpha did not decrease until days 3-5 of treatment with vitamin D3 and then progressively declined thereafter. The mRNAs for thymidine kinase and topoisomerase I decreased slightly and the mRNA for DNA polymerase-alpha by 30-40%, and then remained constant between days 1 to 3 of treatment with the phorbol ester. The M2 subunit of ribonucleotide reductase exhibited an even greater difference, with no change in the steady-state concentration of mRNA over 3 days of exposure to TPA or vitamin D3. On days 5-7 of treatment with vitamin D3, essentially complete loss of the expression of the mRNA for the M2 subunit of ribonucleotide reductase occurred. Measurement of the enzymatic activities of thymidylate synthase and thymidine kinase in cells exposed to either of the inducers of maturation corroborated the findings at the level of the mRNAs, with corresponding decreases in the activity of these enzymes. The results indicate that the down-regulation of the expression of housekeeping enzymes of DNA replication occurs as late events in HL-60 cells undergoing monocytic/macrophagic differentiation, implying that the decreases in their gene expression are the result of the termination of proliferation rather than an initiating event in the cessation of DNA biosynthesis.
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PMID:Regulation of the expression of enzymes involved in the replication of DNA in chemically induced monocytic/macrophagic differentiation of HL-60 leukemia cells. 968 96

Cross-resistance between different cytostatic agents which are structurally and functionally dissimilar is a common phenomenon called multidrug resistance (MDR). The best characterized mechanism of MDR involves P-glycoprotein. However, this does not completely explain MDR. Within the last few years, two new genes that can confer MDR have been identified (MRP and LRP). Furthermore, topoisomerase II has been associated with a special form of MDR. During the past several years, considerable interest has been shown in strategies to reverse MDR by using pharmacological compounds, monoclonal antibodies, immunotoxins, bispecific antibodies, antisense oligodeoxynucleotides, ribozymes, and albumin-conjugated drugs in in vitro and in vivo assays. All these experimental assays demonstrated that MDR can be circumvented. Two agents that have received the most attention in the clinic are verapamil and cyclosporin A. Despite some promising results (especially in hematological malignancies), the results obtained in the treatment of solid tumors with modulators have so far been quite disappointing. This may be explained by the fact that the MDR phenotype alone does not completely account for the resistance of human cancer. Several other resistance-related proteins (e.g., glutathione S-transferase, metallothionein, O6-alkylguanine-DNA-alkyltransferase, thymidylate synthase, dihydrofolate reductase, heat shock proteins) can be also expressed in resistant tumors. Additionally, cell proliferation, vascularization and apoptosis are involved in resistance.
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PMID:Multidrug resistance and its reversal. 971 85

New drug development requires simple in vitro models that resemble the in vivo situation more in order to select active drugs against solid tumours and to decrease the use of experimental animals. In this paper, we review the characteristics and scope of a relatively simple cell-culture system with a three-dimensional organisation pattern - the multilayered postconfluent cell culture model. Solid tumour cell lines from diverse origins when grown in V-bottomed microtiter plates reach confluence in 3-5 days and then start to form multilayers. The initial exponential growth of the culture is followed by a plateau phase when cells reach confluence. This produces changes in the morphology of the cells. For some cell lines, it is possible to observe cell differentiation. A substantial advantage of the system is the use of the sulforodamine B (SRB) assay to determine relative cell growth or viability, which allows semiautomation of the experiments. Several experiments were performed to assess the differences and similarities between cells cultured as monolayers and multilayers, and eventually, compared with the results for solid tumours and some other models such as spheroids. Cell-cycle analysis for multilayers showed a lower S-phase arrest, which is accompanied by a decrease in the expression of cell-cycle-related proteins and a decrease in cellular nucleotide pools. Gene and protein expression of topoisomerase I, topoisomerase II and thymidylate synthase expression were lower for multilayers, but no substantial changes were observed for the expression of DT-diaphorase. P53 expression increased. Multilayer cultures present distinctive properties for drug transport across the membrane, drug accumulation and retention. In fact, the transport of antifolates across the membrane, accumulation of topotecan and gemcitabine-triphosphate are reduced in multilayers when compared with monolayers, which may be related to a decrease in drug penetration to the inner regions of the multilayers. Alteration of these pharmacodynamic parameters is directly related to a decrease in drug activity. The most powerful application of multilayers is in the assessment of cytotoxicity. Solid tumour cell lines from different origins have been treated with several conventional and investigational anticancer drugs. The data show that multilayers are more resistant to the drugs than the corresponding monolayers, but there are substantial differences between the drugs depending on culture conditions, e.g. the difference was rather small for a drug such as cisplatin, miltefosine and EO9, a drug, which is activated under hypoxic conditions. Gemcitabine was active against ovarian cancer but not against colon cancer, resembling the in vivo situation. This observation was not evident with monolayer experiments. Another interesting application is the possibility to perform drug combination studies. The combination of gemcitabine and cisplatin proved to produce selective cell kill in H322 cells (non-small cell lung cancer cell line). Neither of the drugs was independently able to produce similar effects. In summary, multilayer cultures are relatively simple three-dimensional systems to study the effect of microenvironmental conditions on anticancer drug activity. The model might serve as a base for a more rigorous secondary in vitro screening.
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PMID:The multilayered postconfluent cell culture as a model for drug screening. 1103 3

The main objective of this study to analyze which of 31 cellular factors (resistance proteins, proliferative factors, apoptotic factors, angiogenic factors, proto-oncogenes) most accurately predict the resistance of non-small cell lung carcinomas. To this purpose, we used a short-term in vitro test that measures changes in the rate at which radioactive nucleic acid precursors are incorporated into tumor cells after the addition of doxorubicin to determine the response to doxorubicin in 94 non-small cell lung carcinomas. The results obtained by the short-term test were related to the various cellular factors which were in turn determined by immunohistochemistry and flow cytometry. A significant correlation was found between the data obtained by the short-term test and the expression of P-glycoprotein 170 (P = 0.00004), glutathione-S-transferase-pi (P = 0.0002), metallothionein (P = 0.0008), thymidylate synthase (P = 0.002), O6-methylguanine-DNA-methyltransferase (P = 0.008) and lung resistance-related protein (LRP, P = 0.03). There was only a weak correlation between heat shock proteins (HSP70) and no correlation between the expression of topoisomerase II or catalase and the short-term test results. To measure the proliferative activity, the following were determined: PCNA, cyclin A, cyclin D and cdk2. Only a weak relationship was found between the expression of cdk2 (P = 0.04) and PCNA (P = 0.05) and the doxorubicin response in vitro. Of the investigated pro-apoptotic factors (Fas/CD95, Fas ligand, caspase-3), only Fas/CD95 is significantly associated with the drug response (P = 0.007). The apoptotic index also reveals a significant correlation (P = 0.03). Angiogenesis, as measured by the microvessel density and the angiogenic factors, is inversely correlated to the resistance of non-small cell lung cancer. Platelet-derived endothelial cell growth factor (PD-ECGF) and vascular endothelial growth factor (VEGF) exhibit a significant relationship to the drug resistance (P = 0.0006 and P = 0.004, respectively). Of the investigated proto-oncogenes (Fos, Jun, ErbB-1, ErbB-2, Myc, Ras), only ErbB-2 is weakly associated with the in vitro short term test. In order to determine whether combining factors can result in improved predictive information, combinations of the factors (pairs, triplets) were analyzed. The systematic investigation of these combinations yields an improvement in the predictive information. With one factor up to 76.6% of the tumors, with two factors up to 85.4% and with three factors up to 89.5% of the tumors could be correctly diagnosed.
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PMID:Cellular predictive factors for the drug response of lung cancer. 1113 47

mRNA levels of several Crithidia fasciculata genes involved in DNA metabolism have previously been found to cycle as cells progress through the cell cycle. Octamer consensus sequences in the 5' untranslated regions (5' UTRs) of these transcripts were shown to be required for cycling of these mRNAs. The KAP3 gene encodes a kinetoplast histone H1-like DNA binding protein, and its mRNA levels cycle in parallel with those of the kinetoplast DNA topoisomerase (TOP2), dihydrofolate reductase-thymidylate synthase (DHFR-TS), and the large subunit of the nuclear single-stranded DNA binding protein (RPA1). KAP3 mRNA contains two octamer consensus sequences in its 3' UTR but none in its 5' UTR. Mutation of these octamer sequences was not sufficient to prevent cycling of a sequence-tagged KAP3 mRNA expressed from a plasmid. Mutation of an octamer sequence contained on the precursor transcript but not on the mRNA, in addition to mutation of the two octamer sequences in the 3' UTR, was necessary to abolish cycling of the mRNA. The requirement for a sequence not present on the mature mRNA indicates that regulation of the mRNA levels by the octamer sequences occurs at or prior to splicing of the transcript. Incompletely processed RNAs containing octamer sequences were also found to accumulate during the cell cycle when the mRNA levels were lowest. These RNA species hybridize to both the KAP3 coding sequence and that of the downstream drug resistance gene, indicating a lack of processing within the intergenic region separating these genes. We propose a cell cycle-dependent interference in transcript processing mediated by octamer consensus sequences as a mechanism contributing to the cycling of such transcripts.
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PMID:Sequence elements in both the intergenic space and the 3' untranslated region of the Crithidia fasciculata KAP3 gene are required for cell cycle regulation of KAP3 mRNA. 1291 86

The retinoblastoma tumor suppressor protein (RB) is targeted for inactivation in the majority of human tumors, underscoring its critical role in attenuating cellular proliferation. RB inhibits proliferation by repressing the transcription of genes that are essential for cell cycle progression. To repress transcription, RB assembles multiprotein complexes containing chromatin-modifying enzymes, including histone deacetylases (HDACs). However, the extent to which HDACs participate in transcriptional repression and are required for RB-mediated repression has not been established. Here, we investigated the role of HDACs in RB-dependent cell cycle inhibition and transcriptional repression. We find that active RB mediates histone deacetylation on cyclin A, Cdc2, topoisomerase IIalpha, and thymidylate synthase promoters. We also demonstrate that this deacetylation is HDAC dependent, since the HDAC inhibitor trichostatin A (TSA) prevented histone deacetylation at each promoter. However, TSA treatment blocked RB repression of only a specific subset of genes, thereby demonstrating that the requirement of HDACs for RB-mediated transcriptional repression is promoter specific. The HDAC-independent repression was not associated with DNA methylation or gene silencing but was readily reversible. We show that this form of repression resulted in altered chromatin structure and was dependent on SWI/SNF chromatin remodeling activity. Importantly, we find that cell cycle inhibitory action of RB is not intrinsically dependent on the ability to recruit HDAC activity. Thus, while HDACs do play a major role in RB-mediated repression, they are dispensable for the repression of critical targets leading to cell cycle arrest.
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PMID:Histone deacetylation of RB-responsive promoters: requisite for specific gene repression but dispensable for cell cycle inhibition. 1456 17

The novel antitumor compound NC-190 strongly inhibited the growth of FM3A cells with an IC50 of 0.019 microg/ml (0.042 microM) when cultured with NC-190 for 48 h. NC-190 potently suppressed DNA synthesis, with 90% inhibition observed at 0.1 microg/ml of NC-190. RNA and protein syntheses were also suppressed under the same conditions, but to a lesser extent. We then measured the cellular enzymatic activities of DNA polymerase alpha, RNA polymerase, thymidine kinase, thymidylate synthase and Leu-tRNA synthetase of FM3A cells cultured with or without NC-190. Of these 5 enzymes, the activity of thymidine kinase was most strongly suppressed by NC-190, by 77%. Although NC-190 did not directly inhibit the activitiy of thymidine kinase in a cell-free system, expression of mRNA of thymidine kinase was suppressed by 75% in NC-190-treated cells. These results indicate that NC-190 can suppress the expression of the gene for thymidine kinase and the inhibition of thymidine kinase contributes to the inhibition of cell growth by NC-190 together with the inhibition of topoisomerase II.
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PMID:The topoisomerase II-inhibitor NC-190 reduces the level of thymidine kinase mRNA in murine tumor cells. 1463 16

While several studies have reported that thymidylate synthase (TS) tumour expression can be a reliable predictive marker of clinical response to 5-Fluorouracil (5-FU) for advanced colorectal cancer patients, only a few studies that searched for predictive factors of irinotecan (CPT-11) clinical response are available. The aim of the present study has been to verify the predictive value of immunohistochemical topoisomerase-I (Topo-I) and TS primary tumour expression in a consecutive series of 62 advanced colorectal cancer patients that received a first line 5-FU/CPT-11 chemotherapy. TS and Topo-I immunostaining was observed in 76% and 43% of tumours, respectively, resulting in a significant relationship within each tumour (r=0.365, p<0.004). Patients with different TS tumour expression showed a similar percentage of Objective Clinical Response, OR (40% vs. 28% of OR in low and high TS-expressing tumours, respectively, p=ns); also, patients with different Topo-I tumour expression did not show a different probability of OR (39% vs. 29% of OR in high and low Topo-I expressing tumours, respectively; p=ns). The tumour expression of these 2 biomarkers also did not impact on time to progression and overall survival of patients. Furthermore, the combined analysis of TS and Topo-I tumour status did not permit to individualize subgroups of patients with different probability of OR. With multivariate analysis, only patient Performance Status significantly impacted on OS (Hazard ratio 4.87; p=0.02) of these patients. We can conclude that high TS tumour expression seems not to preclude a clinical activity for 5-FU/CPT-11 polichemotherapy in advanced colorectal cancer patients; furthermore, clinical response and prognosis of colorectal cancer patients treated with 5-FU/CPT-11 regimen do not differ in tumours with different TS or Topo-I expression.
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PMID:Topoisomerase-I, thymidylate synthase primary tumour expression and clinical efficacy of 5-FU/CPT-11 chemotherapy in advanced colorectal cancer patients. 1519 79


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