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 purpose of the present study was to evaluate whether intermittent exposure to a constant dose of doxorubicin selects for multidrug resistance (MDR) in RPMI 8226 human myeloma cells and, if so, to determine the molecular mechanism. In an attempt to approximate clinical doxorubicin treatment in vitro, cells were exposed to a fixed dose of doxorubicin for 4 d alternating with growth in drug-free medium for 17 d. An MDR subline emerged, termed 8226/DOXint5, which was 3-4-fold resistant to doxorubicin, etoposide and m-AMSA, and 1.6-fold resistant to vincristine. Sensitivity to docetaxel, melphalan and cisplatin was normal. Verapamil normalized vincristine sensitivity but had little effect on resistance to the other agents. Cellular uptake and retention of daunorubicin and vincristine were reduced by approximately 10%. The 8226/DOXint5 cells showed diminished DNA topoisomerase IIalpha expression and increased expression of the multidrug resistance protein MRP. Expression of MDR1/P-glycoprotein was not detected. Immunostaining showed 70% of the cells to over-express the lung-resistance protein LRP. This new MDR myeloma cell line may prove to be a useful model for the development of strategies to overcome low-level, multifactorial MDR, which might be a common phenomenon in clinical myeloma treated with doxorubicin.
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PMID:Intermittent exposure to doxorubicin in vitro selects for multifactorial non-P-glycoprotein-associated multidrug resistance in RPMI 8226 human myeloma cells. 913 43

We examined the genetic and biochemical bases for drug resistance and the order of appearance of different mechanisms underlying the increasingly more resistant murine erythroleukemia cell lines established in Adriamycin (ADR). In the first-step low-level resistant cell line PC4-A5 (able to grow in 5 ng/mL ADR), there was a 2-fold reduction in topoisomerase IIalpha and topoisomerase IIbeta mRNA levels, as well as topoisomerase IIalpha protein and activity levels as compared with the parental cell line. The topoisomerase IIalpha activity levels remained reduced as the cells became increasingly more resistant. In contrast, the topoisomerase II mRNA and protein levels returned to approximately the parental levels in resistant cells growing in higher drug concentrations (40-160 ng/mL). Parental cells expressed the multidrug resistance protein (MRP), but beginning with PC4-A5 MRP expression decreased and remained reduced in increasingly resistant cell lines. At high levels of ADR resistance, the cells expressed the mdr3 gene concomitant with the appearance of vincristine resistance and energy-dependent daunomycin and vincristine efflux. Glutathione levels, internal pH, and expression of the major vault protein (MVP) remained unchanged in all cell lines. Fluorescence microscopy revealed no alterations in daunomycin distribution or vesicle numbers between the parental and resistant cell lines. Different resistance mechanisms emerge sequentially as cells become more resistant to ADR; the mechanisms are retained during the development of multidrug resistance (MDR). In intermediate-level MDR cell lines (PC4-A10 and PC4-A20), resistance involves an as yet undetermined mechanism(s).
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PMID:Emergence of different mechanisms of resistance in the evolution of multidrug resistance in murine erythroleukemia cell lines. 939 72

Non-small-cell lung cancer (NSCLC) and small-cell lung cancer (SCLC) differ significantly in their clinical response to topoisomerase IIalpha (topo-IIalpha)-directed drugs, such as etoposide and teniposide, as NSCLC is virtually insensitive to single-agent therapy, while SCLC responds in two-thirds of cases. Preclinical studies have indicated that resistance to topo-IIalpha drugs depends on topo-IIalpha content and/or activity, the altered-topo-II multidrug resistance phenotype (at-MDR) and/or one of two different drug efflux pumps, P-glycoprotein (P-gp) and the multidrug resistance protein (MRP). Immunohistochemical analysis on paraffin-embedded tissue from 27 cases of untreated NSCLC and 29 cases of untreated SCLC (of which additional tumour biopsies after treatment with topo-IIalpha-directed drugs were available in ten cases) yielded the following results: NSCLC had significantly less topo-IIalpha than SCLC (P < 0.0001), as only 5 out of 27 NSCLC cases had > 5% positive cells compared with 28 out of 29 SCLC, and 0 out of 27 NSCLC had > 25% positive cells compared with 26 out of 29 SCLC. P-gp was detected in > 5% of cells in only 3 out of 27 NSCLC and in 6 out of 29 SCLC, and MRP in 5 out of 27 of NSCLC and 9 out of 29 SCLC. After treatment of patients with SCLC with either etoposide or teniposide, which are topo-IIalpha-directed drugs, there was an increase in MRP (P < 0.1) and P-gp (P < 0.05) positivity, while topo-IIalpha decreased (P < 0.05). In conclusion, the major difference between untreated NSCLC and SCLC was in topo-IIalpha content. In the small series of ten patients treated for SCLC, all three MDR phenotypes appeared to increase.
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PMID:Immunohistochemical detection of DNA topoisomerase IIalpha, P-glycoprotein and multidrug resistance protein (MRP) in small-cell and non-small-cell lung cancer. 965 63

Cisplatin-resistant KCP-4 cells were 12.4- and 31.6-fold more resistant to CPT-11 and SN-38 than parental KB-3-1 cells, respectively. We studied the mechanism of cross-resistance to CPT-11 and SN-38. Our previous study showed that multidrug resistance protein (MRP), canalicular multispecific organic anion transporter (cMOAT) and P-glycoprotein (P-gp) were not expressed in KCP-4 cells (Chen, Z.-S. et al., Exp. Cell Res., 240 (1998) 312-320, and Chuman, Y. et al., Biochem. Biophys. Res. Commun., 226 (1996) 158-165). The accumulation of both CPT-11 and SN-38 in KCP-4 cells was lower than that in KB-3-1 cells. The ATP-dependent efflux of CPT-11 and SN-38 from KCP-4 cells was enhanced compared with that from KB-3-1 cells. DNA topoisomerase (topo) I expression, topo I activity, topo I-mediated cleavable complex, and the sensitivity to SN-38 of DNA topo I in KCP-4 were similar to those in KB-3-1 cells. Furthermore, the conversion of CPT-11 to SN-38 in the two cell lines was also similar. The transport of LTC4 in KCP-4 membrane vesicles was competitively inhibited by bis-(glutathionato)-platinum (II) (GS-Pt), CPT-11 and SN-38. These findings suggested that an unknown transporter distinct from P-gp, MRP or cMOAT is expressed in KCP-4 cells and transports CPT-11 and SN-38.
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PMID:An enhanced active efflux of CPT-11 and SN-38 in cisplatin-resistant human KB carcinoma cells. 1037 68

We established a VP-16-resistant line of human leukemia cells, K562/VP-H2, derived from K562 cells. K562/VP-H2 cells were 44-fold more resistant to VP-16 than were K562 cells. K562/VP-H2 cells were also resistant to doxorubicin, daunorubicin and mitoxantrone, but showed little or no resistance to vincristine, aclarubicin, idarubicin, idarubicinol, cytosine arabinoside, cis-platinum or camptothecin. K562/VP-H2 cells did not over-express P-glycoprotein or multidrug resistance protein, and showed intracellular accumulation of VP-16 similar to that in K562 cells. While the expressions of topoisomerase II-alpha gene and topoisomerase II-beta gene, or catalytic activity in nuclear extract of K562/VP-H2 cells were similar to that of K562 cells, the VP-16 induced DNA cleavage was reduced in K562/VP-H2 cells compared to K562 cells, suggesting that the reduction of topoisomerase II-mediated DNA cleavage through qualitative alteration of topoisomerase II may be the main mechanism of acquired multidrug resistance for K562/VP-H2 cells. The K562/VP-H2 cell line is an interesting model for studying resistance to antileukemia drugs targeting topoisomerase II.
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PMID:Multidrug resistance due to impaired DNA cleavage in a VP-16-resistant human leukemia cell line. 1069 18

Combined modalities are currently used for cancer therapy, although their mechanisms of activity remain incompletely deciphered. The design of new drug combinations suffers from our inability to anticipate accurately their efficacy or toxicity. They can be evaluated in vivo, using human tumors grafted into immunodeficient mice, as we did here with combined protocols used in the clinical setting. Xenografts of small cell lung carcinoma (SCLC) from eight patients were used to test the tumor sensitivity to etoposide (VP16; 12-16 mg/kg/days, days 1, 2, and 3), cisplatin (CDDP; 6-9 mg/kg/day, day 1) and ifosfamide (IFO; 90-210 mg/kg/day, days 1, 2, and 3) as single agents and to evaluate the efficacy of the two-drug or three-drug combinations. Five xenografts came from untreated patients (SCLC-61, SCLC-6, SCLC-10, SCLC-41, and SCLC-96) and three after treatment (SCLC-74, SCLC-101, and SCLC-108). p53 was inactivated in all of them. Tumor growth inhibition, growth delay, and the survival rate of tumor-bearing mice reflected individual SCLC chemosensitivity. As single agents, IFO inhibited tumor growth in a dose-dependent manner, whereas CDDP and VP16 had little or no effect. Both CDDP and IFO potentiated VP16, inducing complete regressions in the most sensitive SCLCs; VP16-IFO was more effective than VP16-CDDP, with complete regressions in six versus three of the eight tumors tested, respectively. CDDP-IFO was less effective than VP16-IFO, with three of eight SCLCs giving complete regressions. The three-drug combination led to modest improvement over the best two-drug combination but only for sensitive SCLCs. Because drug-responses distinguished two classes of SCLCs, as sensitive or refractory, MDR1, glutathione S-transferase pi, lung-related multidrug resistance protein, multidrug resistance protein, and topoisomerase IIalpha mRNA expression was studied by semiquantitative reverse transcription. There was no correlation with SCLC sensitivity; topoisomerase IIalpha and multidrug resistance protein was expressed in all cases, lung-related multidrug resistance protein and glutathione S-transferase pie in seven of eight, and MDR1 gene in four of eight. In conclusion, these SCLC xenografts displayed a pattern of chemotherapy response close to that observed in patients. This model confirmed that in two-drug combinations, each component potentiated the effects of the other, with VP16-IFO tending to be the best two-drug combination, both of which were more effective than VP16-CDDP and better tolerated than CDDP-IFO. The addition of a third agent gave a modest, if any, therapeutic benefit in the responders but none in refractory SCLCs. There was no correlation between the extent of response and resistance markers.
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PMID:Distinctive potentiating effects of cisplatin and/or ifosfamide combined with etoposide in human small cell lung carcinoma xenografts. 1081 35

BN 80915 is the lead compound from a novel class of E-ring modified camptothecin analogues, the homocamptothecins, which show potent antitumor activities in animal models. Here, we report that BN 80915 induces up to 2-fold more cleavable complexes between plasmid DNA and purified human topoisomerase I than SN-38 and camptothecin. BN 80915 also induces DNA-topoisomerase I complexes in living HT-29 colon carcinoma cells, as shown by the in vivo link assay. BN 80915 is an extremely potent inducer of DNA-protein complexes in these cells starting at a concentration of 5 nM in the media. BN 80915 is clearly more potent than SN-38, because at least 20 times more SN-38 is needed to induce comparable levels of cleavable complexes. Kinetic experiments show that BN 80915 induces cleavable complexes within minutes that remain stable for at least 6 h in the presence of drug. Whereas the majority of the complexes are reversed within 15 min after drug removal, a substantial fraction (30%) persists for at least 4 h, in contrast with SN-38-treated cells, where all complexes have disappeared by this time. BN 80915 shows strong antiproliferative effects toward HT-29 cells with an IC50 of 0.3 nM compared with 20 nM for SN-38 and 40 nM for topotecan. BN 80915 is also potent against other colon carcinoma cells as well as toward cells growing in three dimensions as multicellular spheroids. HL-60 cells expressing functional P-glycoprotein or multidrug resistance protein show no cross-resistance toward BN 80915. Taken together, our results show that BN 80915 is unusually potent toward human colon carcinoma cells because of the formation of high levels of stable, covalent DNA-topoisomerase complexes.
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PMID:Unusual potency of BN 80915, a novel fluorinated E-ring modified camptothecin, toward human colon carcinoma cells. 1130 74

A series of substituted angular benzophenazines were prepared using a new synthetic route via a novel regiocontrolled condensation of 1,2-naphthoquinones and 2,3-diaminobenzoic acids. The synthesis and biological activity of this new series of substituted 8,9-benzo[a]phenazine carboxamide systems are described. The analogues were evaluated against the H69 parental human small cell lung carcinoma cell line and H69/LX4 resistant cell line which overexpresses P-glycoprotein. Selected analogues were evaluated against the COR-L23 parental human non small cell lung carcinoma cell line and the COR-L23/R resistant cell line which overexpresses multidrug resistance protein. This series of novel angular benzophenazines were potent cytotoxic agents in these cell lines and may be able to circumvent multidrug resistance mechanisms which result in the lack of efficacy of many drugs in cancer chemotherapy. These compounds show dual inhibition of topoisomerase I and topoisomerase II and thus target two key enzymes responsible for the topology of DNA that are active at different points in the cell cycle. The introduction of chirality into the carboxamide side chain of these novel benzophenazine carboxamides has resulted in the discovery of a potent enantiospecific series of cytotoxic agents, exemplified by 4-methoxy-benzo[a]phenazine-11-carboxylic acid (2-(dimethylamino)-1-(R)-methyl-ethyl)-amide, XR11576 ((R)-4j' '). In vivo activity has been demonstrated for 4-methoxy-benzo[a]phenazine-11-carboxylic acid (2-(dimethylamino)-1-(R)-methyl-ethyl)-amide, XR11576, after intravenous administration to female mice, and this compound has been selected as a development candidate for further evaluation.
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PMID:Novel angular benzophenazines: dual topoisomerase I and topoisomerase II inhibitors as potential anticancer agents. 1180 24

Studies on multidrug resistance (MDR) require a sensitive and quantitative assay of mRNA expression in clinical tumor samples. Based on the small size, heterogenity, and the possibility of partial degradation of clinical specimens, unambiguous data are often difficult to obtain. The aim of the present study was to develop a multiplex polymerase chain reaction (PCR) in combination with nested PCR for quantitative analyses of mRNA expression of MDR1, MRP (multidrug resistance protein), and DNA topoisomerase IIalpha in small amounts of tumor tissue. RNA samples extracted from the human cell line RPMI 8226 and its MDR sublines 8226/Dox6 and DOXint40c, that overexpress MDR1 and MRP, respectively, were used as model substrates. In the first step, cDNAs of the three genes as well as of the housekeeping gene beta-actin were simultaneously amplified in single tubes using 20 cycles of PCR after random-primed reverse transcription. When necessary, a second amplification step of the preamplified PCR products was employed using nested primer pairs. Primer competition was evaluated by analyses of serially diluted amounts of cDNA and at different numbers of PCR cycles. Based on the results obtained, this multiplex/nested PCR approach may provide a base for quantitative analyses of MDR1, MRP, and topoisomerase IIalpha mRNA expression in clinical tumor biopsies.
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PMID:A diagnostic tool for monitoring multidrug resistance expression in human tumor tissues. 1223 60

Ras-homologous GTPases are involved in the regulation of genotoxic stress-induced gene expression and cell death. Since they need C-terminal isoprenylation for correct intracellular localization and function, we investigated whether depletion of cells from isopren precursor moieties using the HMG-CoA reductase inhibitor lovastatin affects cellular sensitivity to DNA damaging drugs. Here we show that lovastatin renders cells highly resistant to the tumor-therapeutic compound doxorubicin. Desensitization by lovastatin was reverted by co-treatment with GGPP indicating that inhibition of protein geranylgeranylation is involved in acquired doxorubicin resistance. Lovastatin does not influence cellular sensitivity to DNA damaging compounds such as cisplatin, methyl methanesulfonate and ionizing radiation. The frequency of apoptotic cell death induced by doxorubicin was not affected by lovastatin as shown by both annexin V and DNA fragmentation assay. However, lovastatin releases cells from doxorubicin induced G2 blockage. Furthermore, lovastatin protects cells from doxorubicin-induced DNA strand breakage without affecting drug uptake or the expression of multidrug resistance protein (mdr-1). Since lovastatin confers cross-resistance to the topoisomerase II specific inhibitor etoposide, we suggest desensitization by the statin to be related to topoisomerase II function. The finding that lovastatin renders cells resistant to doxorubicin and etoposide by reducing their genotoxic and cytotoxic effects might have clinical implications for cancer therapy.
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PMID:The HMG-CoA reductase inhibitor lovastatin protects cells from the antineoplastic drugs doxorubicin and etoposide. 1223 96


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