Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.6.3.44 (P-glycoprotein)
 13,344 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The topoisomerase II inhibitor, VP-16 (etoposide), is an important component in many chemotherapeutic regimens. To characterize resistance to this drug, the human melanoma cell line, FEM-X, was selected in multiple steps with VP-16. To prevent the development of typical multidrug resistance, an inhibitor of P-glycoprotein, the tiapamil analog, RO-11-2933, was added to the selections. The resultant clone FVP3 is 56-fold resistant to VP-16 and cross-resistant to doxorubicin (Adriamycin) (9-fold) and VM-26 (27-fold). These cells are also two- to four-fold resistant to m-AMSA, daunorubicin, and mitoxantrone. FVP3 is not resistant to the P-glycoprotein substrates vinblastine, does not express the MDR1 gene at detectable levels, and does not show reduced 3H-VP-16 accumulation. Unlike other cell lines that exhibit resistance to inhibitors of topoisomerase II, FVP3 has the same level of topoisomerase II expression and activity as FEM-X. Using live cells treated with VP-16, band depletion assays and KCI/SDS precipitation assays show that topoisomerase II from FVP3 is much less susceptible to drug-induced cleavable complex formation than is that from FEM-X. This difference in sensitivity to VP-16 is also detected using lysates from disrupted cells, but not with isolated nuclei devoid of cytoplasmic and membrane components. In addition, the topoisomerase II present in nuclear extracts from FVP3 is not resistant to the effects of VP-16 as measured by: (1) inhibition of strand passing activity during decatenation of kinetoplast DNA, (2) drug-induced linearization of plasmid DNA, and (3) immunodepletion by VP-16. These results suggest that some component of the cytoplasm or cellular membranes, or a factor depleted from nuclei during their isolation, is responsible for the resistance to VP-16 in FVP3.
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PMID:Characterization of an unusual mutant of human melanoma cells resistant to anticancer drugs that inhibit topoisomerase II. 809 46

K562 leukaemia cells were selected for resistance using 0.5 microM etoposide (VP-16). Cloned K/VP.5 cells were 30-fold resistant to growth inhibition by VP-16 and 5- to 13-fold resistant to m-AMSA, adriamycin and mitoxantrone. K/VP.5 cells did not overexpress P-glycoprotein; VP-16 accumulation was similar to that in K562 cells. VP-16-induced DNA damage was reduced in cells and nuclei from K/VP.5 cells compared with K562 cells. Topoisomerase II protein was reduced 3- to 7-fold and topoisomerase II alpha and topoisomerase II beta mRNAs were each reduced 3-fold in resistant cells. After drug removal, VP-16-induced DNA damage disappeared 1.7 times more rapidly and VP-16-induced DNA-topoisomerase II adducts dissociated 1.5 times more rapidly in K/VP.5 cells than in K562 cells. ATP (1 mM) was more effective in enhancing VP-16-induced DNA damage in nuclei isolated from sensitive cells than in nuclei from resistant cells. In addition, ATP (0.3-5 mM) stimulated VP-16-induced DNA-topoisomerase II adducts to a greater extent in K562 nuclei than in K/VP.5 nuclei. Taken together, these results indicate that resistance to VP-16 in a K562 subline is associated with a quantitative reduction in topoisomerase II protein and, in addition, a distinct qualitative alteration in topoisomerase II affecting the stability of drug-induced DNA-topoisomerase II complexes.
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PMID:Altered stability of etoposide-induced topoisomerase II-DNA complexes in resistant human leukaemia K562 cells. 814 56

Human colon (HCT116/VP48) and lung (A549B/VP29) adenocarcinoma cell lines selected for resistance to etoposide exhibited modified patterns of multi-drug resistance (MDR) that included a differential sensitivity to other DNA topoisomerase II inhibitors and to the plant alkaloids homoharringtonine, vinblastine, and vincristine. The resistance and cross-resistance drug phenotype of the A549B/VP29 cell line was different from that of the HCT116/VP48 cell line. The HCT116/VP48 cell line was 50-fold resistant to etoposide and 30-fold resistant to teniposide. The degree of resistance to other DNA topoisomerase II inhibitors was of a lower magnitude: Adriamycin, 9-fold; daunomycin, 3-fold; 4'-[(9-acridinyl)-amino]-methanesulfone-m-anisidide (m-AMSA), 3-fold; and actinomycin D, 6-fold. The HCT 116/VP48 cell line exhibited a 7-fold resistance to vincristine and a 2-fold resistance to vinblastine but was sensitive to homo-harringtonine. The A549B/VP29 cell line was 5-fold resistant to etoposide and 2-fold resistant to teniposide. The A549B/VP29 cell line exhibited a 2-fold resistance to Adriamycin but was sensitive to daunomycin and showed a 3-fold resistance to m-AMSA. This cell line was sensitive to actinomycin D. The A549B/VP29 cell line was 2-fold resistant to vinblastine and sensitive to homoharringtonine. Both cell lines (HCT116/VP48 and A549/VP29) exhibited no amplification of the human mdr1 DNA sequence, the 4.3-kb P-glycoprotein transcript, or the membrane P-glycoprotein. The sensitivity of cells exhibiting an MDR phenotype not mediated by P-glycoprotein suggests a potential use for homoharringtonine in treating tumors with this type of drug resistance.
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PMID:Etoposide-resistant human colon and lung adenocarcinoma cell lines exhibit sensitivity to homoharringtonine. 826 74

The Adriamycin-resistant small cell lung carcinoma cell line, GLC4/ADR, showed large differences in cross-resistance to drugs such as Adriamycin, etoposide (VP-16), teniposide (VM-26), 4'-(9-acridinylamino)-methanesulfon-m-anisidide (m-AMSA), and mitoxantrone, which stimulate the formation of topoisomerase (Topo) II-DNA complexes. GLC4/ADR cells demonstrated a reduced Topo II activity and no detectable levels of the P-glycoprotein compared to the parental GLC4 cells (S. De Jong et al., Cancer Res., 50: 304-309, 1990). In the present study, the resistance to VM-26 (59.5-fold) and to m-AMSA (4-fold) of GLC4/ADR after a 1-h incubation was further analyzed. Using the K(+)-sodium dodecyl sulfate precipitation assay, a reduction in VM-26- and m-AMSA-induced cleavable complex formation was found in GLC4/ADR cells compared to GLC4 cells that was related to the degree of resistance to each drug. Cellular accumulation of the VM-26 analogues VP-16 was 3- to 8-fold less and the accumulation of m-AMSA 1- to 2-fold less in GLC4/ADR cells than in the parental cells. Following the removal of VM-26, the cleavable complexes in GLC4/ADR cells disappeared at least 2-fold faster than in GLC4 cells, while the efflux of VP-16 was also enhanced in the resistant cells. On the contrary, no differences in cleavable complex disappearance or drug efflux between these cell lines were observed with m-AMSA. Efflux of both drugs, however, occurred at a much higher rate than cleavable complex disappearance. Using isolated nuclei, a reduction in cleavable complexes in GLC4/ADR was still observed with VM-26 as well as m-AMSA compared to GLC4. The resistant nuclei and nuclear extracts showed a 3-fold decrease in M(r) 170,000 Topo II by immunoblotting. No differences in cleavable complex formation were found between nuclear extracts of both cell lines, when the Topo II activities were equalized. These findings suggest that the cross-resistance to m-AMSA is due to a decreased amount of Topo II and decreased drug accumulation, while in addition to these mechanisms an increased rate of cleavable complex disappearance is involved in the cross-resistance to VM-26 of the GLC4/ADR cell line.
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PMID:Topoisomerase II as a target of VM-26 and 4'-(9-acridinylamino)methanesulfon-m-aniside in atypical multidrug resistant human small cell lung carcinoma cells. 838 51

We examined the expression of the genes encoding topoisomerases I and II and those associated with V(D)J [variable(diversity)joining] recombination in two human T-cell acute lymphoblastic leukemia (T.ALL) cell lines, CEM and CEM/DOX. In CEM/DOX cells, which are resistant to doxorubicin, the topoisomerase I gene was found to be 4-fold overexpressed and nuclear topoisomerase I relaxation activity was 2-fold greater in CEM/DOX than in CEM cells. Furthermore, the cleavable complex reaction induced by camptothecin, a specific topoisomerase I inhibitor, was found to be 2.5-increased in the presence of topoisomerase I extracted from CEM/DOX, in comparison to that in CEM cells. Conversely, the topoisomerase II mRNA levels, nuclear decatenation activities and (mAMSA) 4'(9-acridinylamino)methanesulfon-m-anisidide-induced cleavable complex formation in CEM/DOX were similar to those of the doxorubicin-sensitive cells. The results indicate that topoisomerase I activity is elevated in CEM/DOX cells. Nevertheless, CEM/DOX cells were 11-fold more resistant to camptothecin than were CEM cells, and cross-resistance to camptothecin was not reversed by verapamil. Furthermore, using an intact cell assay for DNA-protein complexes, we found that camptothecin-stimulated cleavable complexes formed in CEM/DOX cells were increased in correlation with the elevated topoisomerase I activity. These results suggest that camptothecin resistance in CEM/DOX cells is due to different mechanism(s) than topoisomerase- or P-glycoprotein-associated multidrug resistance. The recombination activating gene, RAG1, which is one of the components of the site-specific V(D)J recombination complex, was 20-fold overexpressed in CEM/DOX cells. In contrast, RAG2 and T160 gene transcripts, other components of the V(D)J complex, were at best poorly detected in both sensitive and resistant cells. No specific V(D)J recombinase activity was found in CEM or CEM/DOX cells when the pJH201 transfection assay was used. The results indicate that CEM/DOX cells failed to generate V(D)J recombination although RAG1 gene is overexpressed. The mechanism of the RAG1 gene activation was not gene amplification, and no rearrangement was detected in the RAG1 gene locus. RAG1 presents homology with the yeast gene HPR1, itself homologous to yeast topoisomerase I and responsible for the control of recombination in somatic cells. Since DNA topoisomerases are themselves involved in the control of DNA topology, recombination and DNA repair, the possible coactivation of RAG1 and topoisomerase I genes in CEM/DOX cells is discussed.
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PMID:Altered topoisomerase I activity and recombination activating gene expression in a human leukemia cell line resistant to doxorubicin. 839 37

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

New synthetic routes to a series of tetra- and pentacyclic acridines related in structure to marine natural products are reported. The novel water-soluble agent dihydroindolizino[7,6,5-kl]acridinium chloride 14 has inhibitory activity in a panel of non-small-cell lung and breast tumor cell lines exceeding that of m-AMSA. The salt inhibited the release of minicircle products of kDNA confirming that disorganization of topoisomerase II partly underlies the activity of the compound. COMPARE analysis of the NCI mean graph profile of compound 14 at the GI(50) level corroborates this conclusion with Pearson correlation coefficients (>0.6) to clinical agents of the topoisomerase II class: however, this correlation was not seen at the LC(50) level. The inhibitory action of 14 on Saccharomyces cerevisiae transfected with human topoisomerase II isoforms showed a 3-fold selectivity against the IIalpha isoform over the IIbeta isoform. Unlike m-AMSA, 14 is not susceptible to P-glycoprotein-mediated drug efflux and retains activity in lung cells with derived resistance to the topoisomerase II inhibitor etoposide.
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PMID:Antitumor polycyclic acridines. 7. Synthesis and biological properties of DNA affinic tetra- and pentacyclic acridines. 1078 Sep 13

DNA topoisomerase II has been shown to be an important therapeutic target in cancer chemotherapy. Here, we describe studies on the antitumor activity of a novel topoisomerase II inhibitor, ER-37328 [12,13-dihydro-5-[2-(dimethylamino)ethyl]-4H-benzo[c]pyrimido[5,6,1- jk]carbazole-4,6,10(5H,11H)-trione hydrochloride]. ER-37328 inhibited topoisomerase II activity at 10 times lower concentration than etoposide in relaxation assay and induced double-strand DNA cleavage within 1 h in murine leukemia P388 cells, in a bell-shaped manner with respect to drug concentration. The maximum amount of DNA cleavage was obtained at 2 microM. Like etoposide, ER-37328 (2 microM) induced topoisomerase II-DNA cross-linking in P388 cells. A spectroscopic study of ER-37328 mixed with DNA demonstrated that ER-37328 has apparent binding activity to DNA. ER-37328 showed potent growth-inhibitory activity against a panel of 21 human cancer cell lines [mean (50% growth-inhibitory concentration) GI50 = 59 nM]. COMPARE analysis according to the National Cancer Institute screening protocol showed that the pattern of the growth-inhibitory effect of ER-37328 was similar to that of etoposide, but different from that of doxorubicin. Studies on etoposide-, amsacrine [4'-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA)]-, and camptothecin-resistant P388 cell lines showed that: (a) etoposide- and m-AMSA-resistant P388 cell lines were partially resistant to ER-37328 compared with the parental cell line; and (b) a camptothecin-resistant cell line showed no cross-resistance to ER-37328. In addition, ER-37328 overcame P-glycoprotein-mediated resistance. In vivo, ER-37328 produced potent tumor regression of Colon 38 carcinoma inoculated s.c., and its activity was superior to that of etoposide or doxorubicin. These results indicate that ER-37328 inhibits topoisomerase II activity through the formation of topoisomerase II-DNA cleavable complex and has potent antitumor activity both in vitro and in vivo.
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PMID:Antitumor activity of ER-37328, a novel carbazole topoisomerase II inhibitor. 1246 11

Multidrug resistance (MDR) mediated by P-glycoprotein is one of the best characterized transporter-mediated barriers to successful cancer chemotherapy. In an attempt to find MDR-reversing agents, a series of novel acridine derivatives were synthesized and evaluated for their in vitro antiproliferative activities against K562 and K562/ADM cells. Some of these compounds showed superior MDR-reversing activities than Amsacrine, the reference compound. Structure-activity relationships (SAR) of these compounds indicated that the N, N-diethylamine moiety had an affect on the in vitro antiproliferative activity. Interestingly, the compounds bearing N, N-diethylamine moiety showed higher growth-inhibitory activity against K562/ADM cells than K562 cells. The high duplex DNA binding affinity and inhibition of topoisomerase of these acridine compounds are maintained which were confirmed by fluorescent quenching and DNA topoisomerase II cleavage assay, respectively. Moreover, several compounds were examined for their ability to increase the accumulation of rhodamine 123 in K562 and K562/ADM cells, and the result suggested that they may be inhibitors for P-glycoprotein. Our study suggested that acridine framework is a potentially interesting scaffold for developing novel MDR-reversing agents.
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PMID:Synthesis, structure-activity relationship and biological activity of acridine derivatives as potent MDR-reversing agents. 2389 91


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