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:3.1.27.5 (RNase)
17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The expression of multidrug resistance (mdr) genes was investigated in the livers of transgenic mice that express the human hepatitis B virus large envelope polypeptide under the transcriptional control of a liver-specific promoter. These mice develop a storage disease due to the accumulation of a nonsecretable form of hepatitis B surface antigen in the hepatocyte. Liver cell injury is followed by a hepatocellular proliferative response, dysplasia, microscopic nodular hyperplasia, and finally hepatocellular carcinoma. The expression of mdr1, mdr2, and mdr3 genes was analyzed in livers at different stages of the disease by RNase protection assay, Western blot, and immunohistochemistry. RNase protection assay revealed that mdr3 mRNA expression was moderately increased in tissue with microscopic nodular hyperplasia and significantly overexpressed in hepatocellular carcinoma but undetectable in earlier stages of the disease. Western blot using isoform-specific anti-mdr3 antibody demonstrated that the expression of mdr3 protein reflected the steady-state level of mdr3 mRNA. Immunohistochemical analyses using anti-mdr3 isoform-specific antibody and monoclonal antibody C219, which recognizes all the three mdr isoforms, demonstrated selective overexpression in preneoplastic foci during the stage of microscopic nodular hyperplasia as well as in neoplastic hepatocytes in hepatocellular carcinoma. No consistent activation of mdr1 and mdr2 (but occasional coactivation with mdr1) genes during hepatocarcinogenesis was observed. Our results suggest that the hepatocellular mdr3-specific activation mechanism is associated with the late events of hepatocarcinogenesis in this model. The predictable kinetics of mdr gene expression in this transgenic tumor model suggest that it is suitable for future studies of the mechanism of mdr gene activation and the possible pharmacological consequences for mdr3 gene expression of hepatocellular carcinoma.
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PMID:Activation of multidrug resistance (P-glycoprotein) mdr3/mdr1a gene during the development of hepatocellular carcinoma in hepatitis B virus transgenic mice. 135 18

A mitoxantrone-resistant human MCF-7 breast cancer subline (MCF/MX) which is approximately 4000-fold resistant to mitoxantrone was isolated by serial passage of the parental wild-type MCF-7 cells (MCF/WT) in stepwise increasing concentrations of drug. MCF/MX cells were also approximately 10-fold cross-resistant to doxorubicin and etoposide but were not cross-resistant to vinblastine. Intracellular accumulation of radiolabeled mitoxantrone was markedly reduced in MCF/MX cells relative to that in the drug-sensitive MCF/WT cells. This decrease in intracellular drug accumulation into MCF/MX cells was associated with enhanced drug efflux, which was reversed when cells were incubated in the presence of sodium azide and 2, 4-dinitrophenol, suggesting an energy-dependent process. Incubation of MCF/MX cells with verapamil did not affect either the accumulation of mitoxantrone or the level of resistance in these cells. Furthermore, RNase protection and Western blot analyses failed to detect the expression of the mdr1 RNA or P-glycoprotein, a drug efflux pump known to be associated with the development of multidrug resistance in vitro. However, a polyclonal antibody directed against a synthetic peptide corresponding to the putative ATP binding domain of P-glycoprotein reacted with two (M(r) 42,000 and 85,000) membrane proteins from MCF/MX cells which were not found in MCF/WT. Functional assays and Western blot analysis for topoisomerase II revealed no differences in topoisomerase II activity or protein levels in MCF/MX cells. Thus, resistance in this cell line is apparently associated with enhanced drug efflux involving a pathway distinct from the mdr1-encoded multidrug transporter P-glycoprotein.
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PMID:Reduced intracellular drug accumulation in the absence of P-glycoprotein (mdr1) overexpression in mitoxantrone-resistant human MCF-7 breast cancer cells. 135 31

Tumor cell resistance due to enhanced efflux of drugs with diverse structures and/or mechanisms of action is termed multidrug resistance (MDR), and modulation of the MDR phenotype by calcium blockers or calmodulin inhibitors is suggested to involve P-glycoprotein. In drug-sensitive (S) and 5-fold doxorubicin (DOX)-resistant (R0) L1210 mouse leukemia cells, no obvious differences in mdr mRNA or P-glycoprotein expression or alterations in cellular uptake, retention, or cytotoxicity of vincristine (VCR) were observed. However, in the 10-fold (R1) and 40-fold (R2) DOX-resistant sublines, expression of P-glycoprotein was correlated with the level of resistance (R2 greater than R1). An RNase protection assay revealed that elevated levels of mdr1 and mdr2 mRNA were detected in R1 and R2 cells, with an additional increase in mdr3 mRNA in the R2 subline. Further, in the R1 and R2 sublines, no VCR dose-dependent cytotoxicity was apparent, and cell kill of greater than 40% was not achievable following a 3-hr drug exposure. Cellular uptake and retention of VCR were 2- to 4-fold lower in the R1 and R2 sublines, compared with similarly treated S or R0 cells. Potentiation of VCR cytotoxicity by a noncytotoxic concentration of 5 microM trifluoperazine (TFP) was greater than 2-fold in S and R0 cells and less than 1.3-fold in the R1 and R2 sublines. Modulation of VCR uptake by 5 microM TFP in the S and R0 cells was 2-fold and it was 4- to 7-fold in the R1 and R2 sublines. The presence of 5 microM TFP, by competing for efflux, enhanced VCR retention 1.5-fold in S and R0 cells and 2- to 4-fold in the R1 and R2 sublines. In contrast to these results with VCR, dose-dependent cytotoxicity of DOX was apparent in all the resistant sublines, and modulation of DOX cytotoxicity by 5 microM TFP was dependent on the level of resistance. Cellular accumulation of DOX was 20 and 50% lower in the R1 and R2 sublines, respectively, compared with similarly treated S or R0 cells. Marked increases (greater than 1.5-fold) in cellular accumulation of DOX by TFP were apparent only in the R2 subline. Results suggest that a relationship between overexpression of P-glycoprotein isoforms and their role in affecting cellular drug levels and consequent cytotoxicity in MDR L1210 cells determines resistance to VCR but not DOX.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Relationship between expression of P-glycoprotein and efficacy of trifluoperazine in multidrug-resistant cells. 167 Sep 62

In an effort to better understand the preferential resistance to actinomycin D displayed by the multidrug-resistant Chinese hamster lung cell line DC-3F/ADX, we have cloned from those cells a number of cDNAs representing p-glycoprotein gene transcripts. Of the 12 clones isolated, all represent pgp1 transcripts and one, pADX165, contains a 4304-base pair insert with an open reading frame encoding a 1276-amino acid protein that is the homolog of the mouse mdr3/mdr1a gene product. A domain by domain comparison of this protein with p-glycoproteins capable of supporting multidrug resistance, i.e. human mdr1, mouse mdr1/mdr1b, and mouse mdr3/mdr1a, shows that, in addition to the ATP binding sites, the second, fourth, and eleventh transmembrane domains and the four small intracellular loops, IC-1, IC-2, IC-4, and IC-5, are highly conserved and are therefore likely to be important for the maintenance of p-glycoprotein function. Of the remaining 11 cDNA clones, 9 were found to be truncated versions of pADX165. Two others, however, pADX185 and pADX124, contained internal deletions resulting in open reading frames capable of encoding lnovel forms of p-glycoprotein. S1 nuclease and RNase protection analysis demonstrated that these cDNAs represent transcripts present in a number of different multidrug-resistant Chinese hamster lung cell lines. Hence, both are considered to be splicing variants of the hamster pgp1 gene primary transcript.
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PMID:Full length and alternatively spliced pgp1 transcripts in multidrug-resistant Chinese hamster lung cells. 167 63

We determined the expression levels of the mdr1 and mdr3 multidrug-resistance genes (also known as PGY1 and PGY3, respectively) in peripheral blood cells from 69 adult patients with acute and chronic leukemias, using an RNase protection assay. Expression of mdr1 was found in samples from patients with acute nonlymphocytic leukemia (13 of 17), chronic myelocytic leukemia (CML, chronic phase, 10 of 10; blast crisis, three of four), acute lymphocytic leukemia (ALL, eight of 11), B-cell chronic lymphocytic leukemia (B-CLL, 17 of 17), hairy cell leukemia (HCL, one of two), and T-cell prolymphocytic leukemia (one of one), but not in B-cell prolymphocytic leukemia (B-PLL, 0 of seven). Expression of mdr3 was only detected in samples from B-cell lymphocytic leukemias: CML, lymphoid blast crisis (one of one), B-cell ALL (two of two), B-CLL (17 of 17), B-PLL (seven of seven), and HCL (two of two). In vitro drug uptake studies by on-line flow cytometry showed that in leukemia cells expressing either mdr1 or mdr3, the steady-state accumulation of daunorubicin could be significantly increased by addition of cyclosporine and, to a lesser extent, by verapamil. Because cyclosporine and verapamil are known as inhibitors of the mdr1-encoded P-glycoprotein drug-efflux pump, and because the mdr1 and mdr3 genes are highly homologous, our data suggest that the mdr3 gene encodes a functional drug pump in B-cell lymphocytic leukemias. The results of this study may have implications for clinical therapy for acute or chronic leukemias expressing the mdr1 or mdr3 gene, in particular, treatment with combinations of cytotoxic drugs plus agents that reverse multidrug resistance. Since mdr1 and mdr3 are frequently expressed in untreated as well as treated leukemia, such combination therapy should be considered for untreated patients as well as treated patients.
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PMID:Expression of mdr1 and mdr3 multidrug-resistance genes in human acute and chronic leukemias and association with stimulation of drug accumulation by cyclosporine. 197 61

Overexpression of a family of plasma membrane glycoproteins, known as P-glycoproteins, is commonly associated with multidrug resistance in animal cells. In rodents, three multidrug resistance (mdr or pgp) genes have been identified, but only two can confer the multidrug resistance phenotype upon transfection into animal cells. Using the RNase protection method, we demonstrated that the levels of three mdr gene transcripts differ among mouse tissues, confirming a previous report that the expression of these genes is tissue specific (J.M. Croop, M. Raymond, D. Huber, A. DeVault, R. J. Arceci, P. Gros, and D. E. Housman, Mol. Cell. Biol. 9:1346-1350, 1989). The levels of mdr transcripts were determined for mouse liver tumors spontaneously arising in both C3H/HeN and transgenic animals containing the hepatitis B virus envelope gene and for tumors induced by two different carcinogenic regimens in C57BL/6N and B6C3-F1 mice. The mdr3 gene was overexpressed in all 22 tumors tested. Our results demonstrate that overexpression of the mdr3 gene in mouse liver tumors does not require exposure of the animals to carcinogenic agents and suggest that its overexpression is associated with a general pathway of hepatic tumor development. The overexpression of the mdr3 gene, which is the homolog of human mdr1 gene, in hepatocellular carcinomas may be responsible for the poor response of these tumors to cancer chemotherapeutic agents.
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PMID:Overexpression of the multidrug resistance gene mdr3 in spontaneous and chemically induced mouse hepatocellular carcinomas. 212 32

P-glycoprotein (PGP), a transporter conferring multidrug resistance to cancer cells, is expressed in the kidney. C219 monoclonal antibody binding revealed PGP in proximal tubules and mesangium of mouse kidneys. A cell line (TKPTS) expressing PGP was developed from proximal tubules of the 8Tg(SV40E)Bri7 mouse. Northern blot analysis demonstrated a 5.0-kb message identified as mdr1 by ribonuclease protection assay. Cyclosporin A (CSA) at 0.15 and 10 microM increased cellular accumulation of verapamil (VRP) by 32 and 121%, respectively (P < 0.001). VRP at 5 microM increased steady-state cellular accumulation of CSA by 46% (P = 0.02). Basal-to-apical transport of the PGP substrate vinblastine was inhibited by VRP. Rhodamine-123 (R-123) influx was rapid and independent of PGP. R-123 efflux was inhibited by VRP and CSA. Inhibition of PGP transport by VRP, CSA, and PSC-833 decreased the 50% effective dose of adriamycin. The concomitant administration of VRP and CSA was not deleterious and coincided with preferential accumulation of VRP over CSA. Inhibition of PGP-mediated transport is demonstrated as a mechanism of renal cell toxicity.
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PMID:Expression and function of P-glycoprotein in a mouse kidney cell line. 765 14

Two Friend leukemic multidrug-resistance (MDR) cell lines were established by exposure to stepwise increased concentrations of rhodamine-123 (RHO) (cell line RR-30) or Adriamycin (ADR) (cell line ARN-15). RR-30 displays preferential resistance to RHO, whereas ARN-15 is more resistant to ADR. The levels of resistance to other MDR drugs and reversibility by verapamil between these two MDR cell lines were somewhat different. Southern blot, RNase protection, and Western blot analysis using gene-specific probes demonstrated that RR-30 and ARN-15 cells preferentially amplified the mdr1 and mdr3 genes, respectively, leading to overexpression of the corresponding P-glycoproteins (p-gp). Our results suggest that members of the mdr gene family can be amplified independently by using different selecting agents, which could be responsible for the differences in the sensitivities to these selecting agents as well as to these MDR drugs.
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PMID:Two multidrug-resistant Friend leukemic cell lines selected with different drugs exhibit overproduction of different P-glycoproteins. 790 27

Considerable evidence has accumulated indicating that overexpression of P-glycoproteins encoded by the multidrug-resistance (mdr) genes is responsible for the development of collateral resistance to a number of structurally and functionally dissimilar cytotoxic compounds in animal cells. There are three mdr genes (mdr1, mdr2, and mdr3) in the mouse genome and two (MDR1 and MDR2) in the human genome; however, only two mouse genes (mdr1 and mdr3) and one human gene (MDR1) can confer multidrug resistance upon transfection into otherwise drug-sensitive cells. Using RNase protection assay we report here that the steady-state levels of mdr1 and mdr3 messenger RNA were elevated in mouse hepatoma cells treated with dexamethasone (Dex); whereas no induction of mdr2 gene was found. Western blot analyses using anti-mdr1 and anti-mdr3 antibodies revealed that the encoded proteins appeared to be increased, but at much reduced levels. The induction was time and Dex concentration dependent. Nuclear run-on experiments demonstrated that the induction was at least in part by transcriptional control. The induction apparently required new protein synthesis since no increases in mdr1 and mdr3 transcripts was found when cultured cells were simultaneously treated with Dex and cycloheximide. Neither mdr1 nor mdr3 gene was induced in the Dex-treated nonhepatoma cell lines, LMtk- and NIH3T3. Similarly, MDR1 messenger RNA levels were elevated in the Dex-treated human hepatoma line, HepG2, but not in the nonhepatoma, HeLa. This study demonstrated that the hormonal regulation of mdr gene expression is gene and cell type specific.
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PMID:Modulation of multidrug resistance gene expression by dexamethasone in cultured hepatoma cells. 810 93

Multidrug resistance (MDR) and more specifically the expression of P-glycoprotein (Pgp) have been studied extensively in vitro. Unfortunately, it appears that the predictive value of MDR recognized in vitro is mostly an incorrect measure to determine the responsiveness of a particular tumour in the clinic. This misunderstood or overvalued role of MDR might explain the failure of strategies to reverse Pgp function by the use of modulators in solid tumours. To obtain more insight in in vivo drug resistance we investigated a panel of 15 human ovarian cancer xenografts consisting of the most common histological subtypes known in ovarian cancer patients. The response rate to cisplatin, cyclophosphamide and doxorubicin in the xenografts resembled the results of phase II trials with these agents in ovarian cancer patients. This resemblance justifies drug resistance studies in this experimental in vivo human tumour system. We determined the expression levels of MDR 1, MRP 1, LRP and topoisomerase IIalpha mRNA by the RNase protection assay and the presence of MRP1 and LRP proteins by immunohistochemistry. The S-phase fraction was investigated as a separate parameter by flow cytometry. In none of the 15 ovarian cancer xenografts was MDR 1 expression detectable. The expression levels of MRP 1 and LRP were low to moderate and resembled the presence of the MRP1 and LRP proteins. There was a weak, inverse relationship between the expression levels of LRP and sensitivity to cisplatin and cyclophosphamide (r = -0.44 and -0.45), but not to doxorubicin. The levels of topoisomerase IIalpha varied among the xenografts (0.73-2.66) and failed to correlate with doxorubicin resistance (r = 0.14). The S-phase fraction, however, showed a relation with the sensitivity to cisplatin (r = 0.66). Among the determinants studied in ovarian cancer in vivo, LRP mRNA and the S-phase fraction were the best predictive factors for drug response and most specifically for the activity of cisplatin.
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PMID:Drug resistance features and S-phase fraction as possible determinants for drug response in a panel of human ovarian cancer xenografts. 1097 Jun 95


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