Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:5.99.1.2 (topoisomerase)
 9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Drug-induced DNA hypermethylation was observed to constitute one component of the response of human tumor cells to toxic concentrations of commonly used cancer chemotherapy agents. In both human lung adenocarcinoma cells (HTB-54) and human rhabdomyosarcoma cells (CCl-136), pulse exposures to the topoisomerase II inhibitors etoposide and nalidixic acid; to the antibiotic doxorubicin; to the microtubule inhibitors vincristine, vinblastine, and colchicine; to the DNA cross-linking agent cisplatinum; to hydroxyurea; and to the antimetabolites 1-beta-D-arabinofuranosylcytosine, 5-fluorouracil, 5-fluorodeoxyuridine, and methotrexate were associated with profound drug-induced DNA hypermethylation. Exposure of human T-lymphocytes (MOLT-4) to toxic pulse doses of 3'-azidodideoxythymidine was associated with similar drug-induced DNA hypermethylation. In every case, drug-induced DNA hypermethylation was observed only when the degree of DNA synthesis inhibition caused by the drug exceeded 90% and when drug levels or duration of exposure was sufficient to kill 90-100% of exposed cells. Drug-induced DNA hypermethylation was shown not to represent a tissue culture phenomenon, since it occurred in vivo during high-dose 1-beta-D-arabinofuranosylcytosine and hydroxyurea treatments in two leukemic patients. Drug-induced alterations in DNA methylation were frequently biphasic, with hypomethylation occurring at drug concentrations which produced mild DNA synthesis inhibition and which killed less than 50% of exposed cells. Exposure to the alkylating agents 1,3-bis(2-chloroethyl)-1-nitrosourea and cyclophosphamide and to the antimetabolites 5-azadeoxycytidine and 6-thioguanine was associated with DNA hypomethylation at all studied concentrations in HTB-54 cells. Drug-induced DNA hypermethylation could be blocked by preexposure to hypomethylating agents administered at nontoxic to mildly toxic concentrations. Drug-induced DNA hypermethylation may be capable of creating drug-resistant phenotypes by inactivating genes the products of which are required for drug cytotoxicity. Perhaps paradoxically, drug-induced DNA hypermethylation may also produce a second class of drug-resistant tumor cells, characterized by overexpression of particular gene products, by potentiating the process of gene amplification.
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PMID:Drug-induced DNA hypermethylation and drug resistance in human tumors. 279 Jul 94

The schedule-dependent cytotoxic effects of topotecan were evaluated in tissue culture experiments with Chinese hamster V79 cells. One hour exposure to topotecan resulted in a typical phase-specific cell killing curve in which increasing concentrations kill progressively more cells and then reach a plateau when all susceptible cells are killed. In contrast, exposure for 24 h results in a steep concentration-response curve with no plateau. Other S-phase agents such as hydroxyurea or aphidicolin antagonized cytotoxicity when administered by simultaneous exposure with topotecan. Combinations of melphalan, BCNU (1,3 bis(2-chloroethyl)-1-nitrosourea), or cisplatinum with topotecan were most effective when cells were exposed to the alkylating agent or platinating agent during the first hour of a 24-h topotecan exposure. Combinations of topotecan with etoposide or adriamycin produce more cytotoxicity when topotecan is administered by prolonged exposure; however, there is no significant difference depending on whether the topoisomerase II inhibitor is added at the beginning or end of the topotecan exposure. These studies show the importance of appropriate dose scheduling to obtain optimal interaction of chemotherapeutic agents given in combination with topotecan.
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PMID:Schedule-dependent cytotoxicity of topotecan alone and in combination chemotherapy regimens. 786 2

Previous investigations have revealed that the human TE-671 MR human rhabdomyosarcoma xenograft selected in vivo for melphalan resistance (M. C. Rosenberg, et al., Cancer Res., 49: 6917-6922, 1989) is cross-resistant to a wide variety of alkylating agents and to bleomycin, but is collaterally sensitive to etoposide. Although glutathione levels were noted to be elevated in TE-671 MR compared to the melphalan-sensitive parental TE-671 xenograft, treatment with buthionine sulfoximine to deplete glutathione levels did not fully restore melphalan sensitivity in the TE-671 MR xenograft. The present studies were undertaken to search for additional mechanisms of resistance in the TE-671 MR xenograft. Drug sensitivity testing performed at the dose of agents that was lethal to 10% of the animals revealed that the TE-671 MR xenograft maintained resistance to the bifunctional cross-linking agent 1,3-bis(2-chloroethyl)-1-nitrosourea and was cross-resistant to the topoisomerase I poison topotecan. Treatment with buthionine sulfoximine did not sensitize the TE-671 MR xenograft to 1,3-bis(2-chloroethyl)-1-nitrosourea. Further, even though O6-alkylguanine-DNA alkyltransferase levels were high in both the TE-671 and TE-671 MR xenografts, depletion of O6-alkylguanine-DNA alkyltransferase activity by treatment with O6-benzylguanine substantially sensitized the TE-671 xenografts but not the TE-671 MR xenografts, suggesting an additional mechanism of resistance. Measurement of additional enzyme activities that might be involved in DNA repair revealed significant elevations in DNA polymerase alpha (46 +/- 8 (SD) units/mg protein in TE-671, 69 +/- 6 units/mg protein in TE-671 MR, P < 0.05) and DNA polymerase beta (0.43 +/- 0.01 units/mg protein in TE-671, 0.78 +/- 0.12 units/mg protein in TE-671 MR, P < 0.05) but not DNA polymerase delta or total DNA ligase. Examination of topoisomerases by activity assays and Western blotting revealed a 2-fold increase in topoisomerase II and a 2-fold decrease in topoisomerase I in the TE-671 MR xenograft compared to the parental xenograft, apparently explaining the collateral sensitivity to etoposide and cross-resistance to topotecan. These results suggest that TE-671 MR xenografts contain multiple changes in activities of DNA repair-related proteins and other nuclear proteins that could contribute to alkylating agent resistance.
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PMID:Elevated DNA polymerase alpha, DNA polymerase beta, and DNA topoisomerase II in a melphalan-resistant rhabdomyosarcoma xenograft that is cross-resistant to nitrosoureas and topotecan. 801 71

By altering the accessibility of DNA sequences for alkylation or platination, and/or for subsequent repair, topoisomerase II can potentially affect the level of DNA interstrand cross-links induced in cells by bifunctional agents. In this study, we investigated the extent to which inhibition of topoisomerase II activity in a human glioblastoma multiforme cell line alters the kinetics of both the formation and the repair of total genomic DNA interstrand cross-links, as well as the sensitivity of the tumor cells to cis-diamminedichloroplatinum II (cis-DDP) and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). Cells were incubated with and without 200 microM novobiocin, a known topoisomerase II inhibitor, for 24 h, followed by exposure to 50 microM BCNU and 25 microM cis-DDP. DNA interstrand cross-linking was determined at various time points over 72 h, using a modified ethidium bromide-DNA binding assay. Sensitivity of the cells to cis-DDP and BCNU was also determined with and without novobiocin pretreatment with 200 microM novobiocin. This concentration of novobiocin showed no significant direct cytotoxicity, although it inhibited topoisomerase II activity in tumor cell nuclear extracts by 73%. A significant decrease in the rate of repair of both cis-DDP and BCNU induced DNA interstrand cross-links, with a corresponding decrease in the clonogenic survival of the cells, was observed following novobiocin exposure. Although the peak cross-link indices of novobiocin-treated cells relative to controls were not significantly increased, residual DNA cross-linking in the cells after 72 h was increased by 1.4-fold for BCNU and 3-fold for cells treated with cis-DDP, thus, indicating a greater effect of topoisomerase II on cross-link repair than on cross-link formation. These data suggest that inhibition of topoisomerase II may provide a potentially effective clinical strategy for sensitizing human brain tumors, and possibly other tumors as well, to DNA cross-linking anticancer agents.
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PMID:Topoisomerase II inhibition and altered kinetics of formation and repair of nitrosourea and cisplatin-induced DNA interstrand cross-links and cytotoxicity in human glioblastoma cells. 824 21

In order to simulate drug resistance observed in the clinic, two cisplatin-resistant cell lines were produced from a murine ovarian reticulosarcoma, M5076 (M5), by pulse (M5/CDDP) and continuous (M5/CDDPc) treatment with cis-diamminedichloroplatinum(II)(CDDP). These cell lines showed a similar stable low level of resistance (approximately 3-fold) to CDDP and cross-resistance to carboplatin, iproplatin and the new alkylating agent tallimustine, but not to L-PAM (L-phenylalanine mustard) and BCNU (1,3-bis(2-chloroethyl)-1-nitrosourea). Collateral sensitivity to two inhibitors of topoisomerase II, VP16 (etoposide) and doxorubicin (Dox), but cross-resistance to the topoisomerase I inhibitor, camptothecin, were observed. The two cell lines were also sensitive to 5-fluorouracil. No increase in the level of glutathione or activity of glutathione S-transferase could be observed in resistant cells compared with the parental M5 cells. Total DNA platination immediately after treatment was similar in the parental and resistant cell lines. Repair of total DNA platination, measured after 24 h of recovery, was undetectable in M5 and M5/CDDP cells, but was 33% in M5/ CDDPc cells. Initial DNA-interstrand cross-links (DNA-ISC) were six times higher in M5 than in M5/CDDP cells, but 24 h after treatment, both lines had completely repaired this damage. M5/ CDDPc cells did not show formation of DNA-ISC at any time after treatment. The two resistant cell lines were tumorigenic when implanted in mice and resistant to CDDP treatment in vivo. The CDDP resistant tumours were not cross-resistant in vivo to L-PAM, BCNU and Dox, which had been active in vitro, nor to tallimustine, which had been cross-resistant in vitro. Mechanisms of resistance in M5/CDDP and M5-CDDPc seem to be based on a lower formation of DNA-ISC combined, for the latter cell line, with a higher repair capacity for total DNA platination.
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PMID:In vitro and in vivo characterisation of low-resistant mouse reticulosarcoma (M5076) sublines obtained after pulse and continuous exposure to cisplatin. 894 89

We have shown previously that NAD/poly(ADP-ribose) polymerase-deficient cells that overexpress Mr 78,000 glucose-regulated stress protein (GRP78) are resistant to topoisomerase II inhibitors, such as etoposide, m-amsacrine, and doxorubicin. However, these cells have been found to be hypersensitive to DNA cross-linking agents, including melphalan, cisplatin, and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). These observations prompted us to examine whether overexpression of GRP78 is associated with modulation of cytotoxicity of clinically useful DNA-cross-linking agents such as melphalan, BCNU, and cisplatin. We up-regulated GRP78 in V79 Chinese hamster cells by 2-5-fold using two independent approaches that include exposure to 6-aminonicotinamide, or 2-deoxyglucose. Subsequently, these GRP78-overexpressing cells were trypsinized, plated in regular medium without GRP78-inducing agents, and allowed a 5-h attachment time before being treated with melphalan, BCNU, or cisplatin for 1 h to determine clonogenic survivals. In addition, repair of DNA cross-links induced by those agents were determined by alkaline elution assay. Our results show that the GRP78-overexpressing V79 cells are hypersensitive to DNA cross-linking agents compared to the control V79 cells. Furthermore, repair of drug-induced DNA cross-links appears to be considerably slower in these cells relative to that found in control V79 cells. Thus, our results suggest that (a) up-regulation of GRP78 is associated with an impairment of DNA cross-link repair, (b) up-regulation of GRP78 is associated with potentiation of cytotoxicity induced by alkylating and platinating agents, and (c) up-regulation of GRP78 can be considered as a potentially useful tool to modulate the cytotoxicity of clinically useful alkylating and platinating agents.
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PMID:Hypersensitivity to DNA cross-linking agents associated with up-regulation of glucose-regulated stress protein GRP78. 937 11

Two major obstacles in the treatment of patients with central nervous system malignancies are drug resistance and host toxicity. The goal of combination chemotherapy is to achieve therapeutic effects that are more favorable than using a single drug alone, but without an increase in normal organ toxicity. The study reported here examined the combination of a topoisomerase I inhibitor, irinotecan (CPT-11), with three different alkylating agents: 1,3-bis(2-chloroethyl)-1-nitrosourea, busulfan, and cyclophosphamide. We evaluated the antitumor effects of these three combinations against a panel of human tumor xenografts derived from central nervous system malignancies, including adult high-grade gliomas (D-54 MG, D-245 MG) and a childhood ependymoma (D-612 EP). In replicate experiments, the alkylating agents were given on day 1 in doses varying from 10% to 75% of the dose lethal to 10% of the animals, and CPT-11 was given on days 1-5 and 8-12 in doses varying from 10% to 100% of the dose lethal to 10% of the animals. The antitumor effects of the various combinations ranged from less than additive (7.61 days below additive with 0.5 CPT-11 + 0.75 cyclophosphamide in D-54 MG) to statistically significant (P < 0.001) supraadditive effects (18.80 days above additive with 0.5 CPT-11 + 0.5 1,3-bis(2-chloroethyl)-1-nitrosourea in D-54 MG). These studies show that the combination of the topoisomerase inhibitor CPT-11 and alkylating agents may increase the antitumor effect in some cases well above additive with no increase in host toxicity (0/10 deaths in both experiments cited above) and should be considered for combination chemotherapy of central nervous system malignancies.
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PMID:Enhancement of irinotecan (CPT-11) activity against central nervous system tumor xenografts by alkylating agents. 955 93

Because the aberrantly activated phosphoinositide 3-kinase (PI3K)/Akt pathway renders tumor cells resistant to cytotoxic insults, including those related to anticancer drugs, inhibition of the pathway may possibly restore or augment the effectiveness of chemotherapy. Using the human malignant glioma cell lines U87, A172, LN18, and LN229, we examined effects of the PI3K inhibitor LY294002 on both apoptosis and cytotoxicity induced by chemotherapeutic agents, including antimicrotubule agents vincristine and paclitaxel, an alkylating agent 1,3-bis(2-chloroethyl)-1-nitrosourea, a topoisomerase II inhibitor etoposide, and a DNA cross-linking agent cisplatin (cis-diamminedichloroplatinum), and we compared the LY294002-induced enhancement of effects of those agents. Ten to 20 micro M LY294002 augmented both apoptosis and caspase 3-like activity caused by antimicrotubule agents to a larger extent than induced by 1,3-bis(2-chloroethyl)-1-nitrosourea, etoposide, and cisplatin in all four malignant glioma cell lines examined. The same doses of LY294002 enhanced cytotoxicity more efficiently with antimicrotubule agents than with other chemotherapeutic agents. Quantitative analyses using a modified isobologram and median effect plot method revealed that enhancement by LY294002 of vincristine- or paclitaxel-induced cytotoxicity was synergistic, whereas enhancement by the PI3K inhibitor of the other chemotherapeutic agent-induced cytotoxicity was additive. Our study indicates that the synergistic augmentation of the cytotoxicity by LY294002 occurs specifically with antimicrotubule agents, at least partially through an increase in caspase 3-dependent apoptosis, and we suggest that inhibitors of the PI3K/Akt pathway in combination with antimicrotubule agents may induce cell death effectively and be a potent modality to treat patients with malignant gliomas.
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PMID:Synergistic augmentation of antimicrotubule agent-induced cytotoxicity by a phosphoinositide 3-kinase inhibitor in human malignant glioma cells. 1287 4

Apurinic/apyrimidinic (AP) endonuclease 1 (APE1) is the primary enzyme in mammals for the repair of abasic sites in DNA, as well as a variety of 3' damages that arise upon oxidation or as products of enzymatic processing. If left unrepaired, APE1 substrates can promote mutagenic and cytotoxic outcomes. We describe herein a dominant-negative form of APE1 that lacks detectable nuclease activity and binds substrate DNA with a 13-fold higher affinity than the wild-type protein. This mutant form of APE1, termed ED, possesses two amino acid substitutions at active site residues Glu(96) (changed to Gln) and Asp(210) (changed to Asn). In vitro biochemical assays reveal that ED impedes wild-type APE1 AP site incision function, presumably by binding AP-DNA and blocking normal lesion processing. Moreover, tetracycline-regulated (tet-on) expression of ED in Chinese hamster ovary cells enhances the cytotoxic effects of the laboratory DNA-damaging agents, methyl methanesulfonate (MMS; 5.4-fold) and hydrogen peroxide (1.5-fold). This MMS-induced, ED-dependent cell killing coincides with a hyperaccumulation of AP sites, implying that excessive DNA damage is the cause of cell death. Because an objective of the study was to identify a protein reagent that could be used in targeted gene therapy protocols, the effects of ED on cellular sensitivity to a number of chemotherapeutic compounds was tested. We show herein that ED expression sensitizes Chinese hamster ovary cells to the killing effects of the alkylating agent 1,3-bis(2-chloroethyl)-1-nitrosourea (also known as carmustine) and the chain terminating nucleoside analogue dideoxycytidine (also known as zalcitabine), but not to the radiomimetic bleomycin, the nucleoside analogue beta-D-arabinofuranosylcytosine (also known as cytarabine), the topoisomerase inhibitors camptothecin and etoposide, or the cross-linking agents mitomycin C and cisplatin. Transient expression of ED in the human cancer cell line NCI-H1299 enhanced cellular sensitivity to MMS, 1,3-bis(2-chloroethyl)-1-nitrosourea, and dideoxycytidine, demonstrating the potential usefulness of this strategy in the treatment of human tumors.
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PMID:A dominant-negative form of the major human abasic endonuclease enhances cellular sensitivity to laboratory and clinical DNA-damaging agents. 1725 46