Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The expression of beta-actin, gamma-actin, alpha-tubulin, and beta-tubulin mRNA during the lectin activation of human peripheral blood lymphocytes was examined with specific cDNA clones. The resting lymphocyte has a low level of both alpha- and beta-tubulin mRNAs, and these increase 10-fold after 72 h of lectin stimulation in which maximum cell transformation is achieved. Although there is a slight increase in tubulin mRNA during the first 6 h, most of the increase occurs between 6 and 24 h as the cells start to increase their RNA content and progress from G0 into G1. Both beta- and gamma-actin mRNAs are more abundant than the tubulin mRNAs in resting cells, with beta-actin mRNA being the major species. Upon activation, beta-actin mRNA increases threefold, whereas gamma-actin mRNA increases almost sixfold. Both beta- and gamma-actin mRNA are elevated 2.5-fold as early as 6 h, the gamma-actin mRNA level then increasing more than beta-actin between 6 and 24 h, resulting in the reduced beta-actin/gamma-actin mRNA ratio. The lectin-stimulated lymphocyte has a similar beta-actin/gamma-actin mRNA ratio as that of the human leukemic T-lymphoblast cell line CCRF-CEM. These increases are over and above the general increase in polyadenylated RNA content upon lectin activation. On returning to a noncycling state, the levels of these cytoskeletal mRNAs decrease. There were two beta-tubulin mRNAs present in lymphocyte cytoplasm, one of 1.8 kilobases and one of 2.8 kilobases in length. The nongrowing lymphocytes had relatively lower levels of the larger sized mRNA. Upon stimulation, the relative level of the larger mRNA was increased, and at 72 h the cells had approximately equal levels of both mRNAs as did the leukemic lymphoblasts.
Mol Cell Biol 1984 Sep
PMID:Changes in levels of actin and tubulin mRNAs upon the lectin activation of lymphocytes. 654 47

Resistance to dexamethasone (1 microM) was measured in glucocorticoid-sensitive diploid and tetraploid clones of the human leukaemic cell line CCRF-CEM (clone C7) during continuous culture and after X-ray or chemical mutagenesis. In continuous culture resistant diploid cells accumulated at a rate of about one cell per 10(5) divisions, while the rate for tetraploid cells was less than one per 10(7) divisions. Chemical and X-ray mutagenesis caused a marked increase in the number of resistant diploid cells but had very little effect on tetraploid cells. These results are consistent with a mutational basis for the acquisition of the glucocorticoid-resistant phenotype in human lymphoid cells.
Mol Cell Endocrinol 1983 Jan
PMID:Decreased acquisition of glucocorticoid resistance in tetraploid human lymphoid cells. 657 61

We studied the accumulation of [3H]vinblastine (VLB) by lines of CCRF-CEM cultured human leukemic lymphoblasts that were either sensitive or resistant to the drug. Neither cell line metabolized VLB, nor selectively retained any radioactive impurities. There was an apparent "instantaneous" accumulation of VLB by cells of both lines, resulting in cell to medium ratios greater than 1.0 within 1 sec after drug addition. Experiments between 0 and 60 sec revealed that the presumed undirectional initial rate of VLB accumulation by resistant cells, termed CEM/VLB100, was about one-half that of sensitive CEM cells. In experiments carried out over 60 min, the VLB-resistant cells accumulated considerably less [3H]VLB than did the sensitive cells. Drug accumulation by both cell lines was temperature-sensitive, since incubation of cells at 4 degrees resulted in only minimal uptake beyond that observed at zero time. CEM/VLB100 cells retained less drug than did CEM cells, apparently because of a larger fraction of readily releasable VLB compared with CEM cells. The accumulation of VLB by either cell line was related in part to cellular levels of ATP. Although depletion of ATP was associated with decreased accumulation of VLB by CEM cells, it was related to enhanced drug accumulation by CEM/VLB100 cells. Restoration of ATP levels to near control values by addition of glucose also had opposite effects on the two cell lines, causing further accumulation of VLB by the sensitive line but leading to apparent drug efflux from the resistant line. Potentially competing substrates (VM-26, colchicine, daunorubicin, and doxorubicin) failed to block this glucose-mediated release of VLB from the CEM/VLB100 cells. In experiments with energy-depleted CEM/VLB100 cells preloaded with VLB and then incubated in drug-free medium, initial drug loss was shown to be independent of cellular metabolism, being roughly the same for both metabolically intact and metabolically depleted cells. Glucose (energy) was required only for subsequent release of what appeared to be a more tightly bound cell-associated fraction of VLB. Results of zero-time binding studies tended to confirm that VLB binding by resistant cells has two components, one requiring and the other not requiring metabolic energy. Differences in the proportions of these two components between the sensitive and resistant cells suggest a mechanism for resistance to VLB and similar compounds.
Mol Pharmacol 1983 Nov
PMID:Energy-dependent reduced drug binding as a mechanism of Vinca alkaloid resistance in human leukemic lymphoblasts. 657 44

Deoxycytidylate deaminase has been highly purified (1232-fold) from human leukemia CCRF-CEM cells. The native molecular weight of the enzyme is 108 000 and subunit molecular weight 50 500, suggesting that the native enzyme exists as a dimer. The enzyme exhibits a sigmoidal initial velocity vs substrate concentration curve and is regulated by allosteric effectors, dCTP and TTP. The curve relating substrate concentration to initial velocity was changed from a sigmoidal shape to a hyperbolic one by the activator dCTP, while the inhibitor TTP increased the sigmoidicity of the curve. The molecular weight of deoxycytidylate deaminase was unchanged in the presence of allosteric effectors, indicating that aggregation-disaggregation is not the basis of regulation. Deoxycytidylate deaminase exhibited the greatest affinity for the substrate dCMP, with lesser affinity for ara-CMP, and least affinity for CMP. Ara-CMP was an effective substrate in the presence of dCTP concentrations exceeding 4 microM. These data indicate that human neoplastic cell deoxycytidylate deaminase is a highly regulated allosteric enzyme, which is likely to have a significant influence on cellular dUMP, dCTP and TTP pools. These findings further suggest, that the enzyme through its influence on dUMP levels is likely to modulate the biochemical effects of pyrimidine antimetabolites active against the thymidylate synthetase reaction and in the presence of elevated dCTP pools will promote deamination of ara-CMP to the inactive ara-UMP.
Mol Cell Biochem 1983
PMID:Kinetic behaviour and allosteric regulation of human deoxycytidylate deaminase derived from leukemic cells. 658 81

Expression of class I histocompatibility antigens (HLA-A and B) on hybrids of human T and B lymphoblastoid cell lines (LCL) was examined. The T-LCL CEM expressed low levels of HLA-A and B antigens. CEMR and CEMR .3, two 8-azaguanine- and ouabain-resistant sublines of CEM used for fusion, expressed no detectable HLA-B antigens and expressed HLA-A antigens at a level below that of CEM. The three B-LCL studied expressed class I histocompatibility antigens at levels 50- to 80-fold in excess of that found on CEM as assessed by indirect immunofluorescence and flow cytometry. Total levels of class I histocompatibility antigens on hybrids of CEMR and CEMR .3 with B-LCL were similar to those found on the B-LCL. CEM-encoded HLA-A and B antigens were expressed on the hybrids at levels much greater than those seen on CEM itself; expression by the hybrids of CEM-encoded and B-LCL-encoded class I antigens was comparable. By RNA-DNA filter hybridization, CEMR .3 was found to have extremely low levels of class I heavy-chain mRNA compared with two B-LCL and with HSB, a T-LCL that expresses high levels of class I histocompatibility antigens. Thus, the paucity of HLA-A and B expression by CEMR .3 (and by inference, CEMR and CEM), as well as the enhancement of CEM-encoded HLA-A and B antigen expression on B-LCL X T-LCL hybrids, must be due, at least in part, to modulation of the level of transcripts encoding HLA class I heavy chains.
Somat Cell Mol Genet 1984 May
PMID:Expression of class I histocompatibility antigens on human T-B lymphoblast hybrids. 660 40

Fifty-four independent dexamethasone-resistant clones were isolated from the clonal, glucocorticoid-sensitive human leukemic T-cell line CEM-C7. Resistance to 1 microM dexamethasone was acquired spontaneously at a rate of 2.6 X 10(-5) per cell per generation as determined by fluctuation analysis. After mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), the phenotypic expression time for dexamethasone resistance was determined to be 3 days. Spontaneous acquisition of resistance to 0.1 mM 6-thioguanine appeared to occur at a much slower rate, 1.6 X 10(-6) per cell per generation. However, the expression time after MNNG mutagenesis for this resistant phenotype was greater than 11 days, suggesting that the different rates of acquisition for the two phenotypes measured by fluctuation analysis were the results of the disparate expression times. The mutagens ICR 191 and MNNG were effective in increasing the dexamethasone-resistant fraction of cells in mutagenized cultures; ICR 191 produced a 35.6-fold increase, and MNNG produced an 8.5-fold increase. All the spontaneous dexamethasone-resistant clones contained glucocorticoid receptors, usually less than half of the amount found in the parental clone. They are therefore strikingly different from dexamethasone-resistant clones derived from the mouse cell lines S49 and W7. Dexamethasone-resistant clones isolated after mutagenesis of CEM-C7 contained, on the average, lower concentrations of receptor than did those isolated spontaneously, and one clone contained no detectable receptor. These results are consistent with a mutational origin for dexamethasone resistance in these human cells at a haploid or functionally hemizygous locus. They also suggest that this is a useful system for mutation assay.
Mol Cell Biol 1981 Jun
PMID:Isolation and characterization of dexamethasone-resistant mutants from human lymphoid cell line CEM-C7. 696 6

The effects of methotrexate (MTX) in the presence or absence of exogenous thymidine (dThd, 10(-5) M) or hypoxanthine (Hx, 10(-4) M) on cell cycle kinetics and deoxyribonucleoside triphosphate pools (dNTP) were studied in cultured human leukemic T-cells (CCRF-CEM). MTX cytotoxicity was found to increase linearly with drug dose for MTX concentrations between 10(-9) M and 10(-7) M. No further increase in cytoxicity was observed with much higher MTX concentrations (10(-7) M-10(-4) M). A similar dose-response relationship was found for both MTX-induced inhibition of DNA synthesis and changes in dTTP and dGTP pools but not for either MTX-induced inhibition of purine synthesis or changes in dATP and dCTP pools. Exogenous dThd reduced MTX cytotoxicity, at all MTX concentrations examined, but Hx reduced cytotoxicity only at MTX concentrations less than 6 X 10(-8) M and potentiated toxicity with higher MTX concentrations. This potentiation of cytotoxicity was accompanied by substantial elevation of dATP pools. In all instances where dThd or Hx reduced MTX cytotoxicity, a concomitant increase in both dTTP and dGTP levels and in the rate of DNA synthesis was observed. These results suggest a close correlation between MTX-induced alterations of dNTP and inhibition of DNA synthesis and subsequent MTX cytotoxicity. The possible modulation of MTX cytotoxicity by purines is discussed.
Mol Pharmacol 1982 Jan
PMID:Biochemical and cell cycle perturbations in methotrexate-treated cells. 698 94

Methylthioadenosine phosphorylase (MTAP) is an enzyme that functions in a salvage pathway for adenine synthesis. The locus that encodes MTAP activity has been mapped to human chromosome 9 (9q12-9pter) by analysis of mouse x human somatic cell hybrids. Cells that have MTAP activity will stop proliferating, and eventually die in the presence of azaserine, an inhibitor of de novo purine synthesis, but can be rescued by the addition of methylthioadenosine (MTA) to the culture medium. Some mouse and human tumor cells lack MTAP activity and can not grow in the presence of azaserine and MTA. We fused MTAP competent human fibroblast cells to MTAP deficient mouse L-cells and selected for somatic cell hybrids, containing MTAP activity, in medium containing azaserine and MTA. In a separate experiment, a CHO cell x human fibroblast somatic cell hybrid, containing a normal copy of human chromosome 9, was used to prepare microcells, which were fused to an MTAP-deficient human leukemic cell line, CCRF-CEM. Somatic cell and microcell hybrids were shown to retain human chromosome 9 by fluorescence in situ hybridization using probes that hybridize to the interferon-alpha and -beta 1 genes on human chromosome 9 (9p21), and the centromere of human chromosome 9. This is the first report of complementation for MTAP activity being used to select for somatic cell hybrids and microcell hybrids that retain a human chromosome 9.
Somat Cell Mol Genet 1993 Sep
PMID:The use of methylthioadenosine phosphorylase activity to select for human chromosome 9 in interspecies and intraspecies hybrid cells. 750 66

Studies on the mode of action of PT523 [N alpha-(4-amino-4-deoxypteroyl)-N delta-hemiphthaloyl-L-ornithine], a potent nonpolyglutamatable antifolate, were carried out in sensitive and resistant H35 rat hepatoma cell lines in culture, to compare it with other antifolates, including three dihydrofolate reductase (DHFR) inhibitors, i.e., methotrexate (MTX), gamma-fluoro-MTX, and trimetrexate (TMQ), two thymidylate synthase inhibitors, i.e., N10-propargyl-5,8- dideazafolate (PDDF) and 2-desamino-2-methyl-N10-propargyl-5,8-dideazafolate (dmPDDF), and the glycinamide ribonucleotide formyltransferase inhibitor 5,10-dideaza-5,6,7,8-tetrahydrofolate. PT523 was the most active compound in this group against the parental H35 cells, with an IC50 ranging from 2.5 nM for 72 hr of treatment to 0.21 microM for 2 hr of treatment. Sublines resistant to MTX by virtue of a transport defect or a combination of defective transport and increased DHFR activity were resistant to PT523 and MTX but not to PDDF, whereas sublines resistant to fluoropyrimidines by virtue of increased thymidylate synthase activity were resistant to PDDF but not to PT523, TMQ, or MTX. Inhibition of H35 cell growth by PT523 was associated with a concentration- and time-related decrease in de novo dTMP and purine biosynthesis. Growth inhibition by PT523, MTX, and TMQ was prevented by leucovorin or a combination of thymidine (dThd) and hypoxanthine but not by dThd or hypoxanthine alone; in contrast, growth inhibition by dmPDDF was prevented by dThd alone. Intracellular reduced folate polyglutamate pools were markedly altered by PT523 treatment, with the most pronounced effect being an increase in 7,8-dihydrofolate mono- and polyglutamates and a decrease in 5,10-methylene-5,6,7,8-tetrahydrofolate mono- and polyglutamates, 5,6,7,8-tetrahydrofolate mono- and polyglutamates, and 10-formyl-5,6,7,8-tetrahydrofolate mono- and polyglutamates. This pattern was qualitatively similar to that observed with MTX and TMQ but different from that observed with dmPDDF or 5,10-dideaza-5,6,7,8-tetrahydrofolate, which resulted in little or no change in the folate species. Uptake of [3H]MTX and [3H]folinic acid, but not [3H]folic acid, by H35 cells was inhibited in a dose-related manner by PT523, suggesting that penetration of the cell probably involves, at least in part, active transport by the MTX/reduced folate carrier. To determine whether the potent cellular effects of PT523 might be due to chemical or enzymic clevage to N'-(4-amino-4-deoxypteroyl)-L-ornithine, a potent inhibitor of folylpolyglutamate synthetase, the formation of [3H]MTX polyglutamates in CCRF-CEM lymphoblasts pulsed with [3H]MTX after preincubation with PT523 was examined.(ABSTRACT TRUNCATED AT 400 WORDS)
Mol Pharmacol 1994 Apr
PMID:Biochemical studies on PT523, a potent nonpolyglutamatable antifolate, in cultured cells. 751 64

Oltipraz, an inhibitor of human immunodeficiency virus type 1 replication in vitro (ED50 approximately 10 microM), undergoes extensive metabolism in vivo. Most of the orally administered drug undergoes opening of the dithiolethione ring, reduction, recyclization, and methylation to form 7-methyl-6,8-bis(methylthio)pyrrolo[1,2-a]pyrazine ("metabolite III"). We report here that metabolite III inhibits viral replication in vitro (ED50 approximately 25 microM) in acutely infected H9 and CEM T cell lymphoma cell lines. Although both metabolite III and oltipraz were able to inhibit phorbol-12-myristate-13-acetate-stimulated viral replication in the chronically infected U1 promonocytic leukemia cell line, only metabolite III was able to inhibit phorbol-12-myristate-13-acetate-stimulated viral replication in chronically infected ACH-2 T cell lymphoma cells. The results with ACH-2 cells suggest that oltipraz inhibits an early stage of the viral life cycle, whereas metabolite III affects human immunodeficiency virus type 1 replication at a step distal to viral integration. This is consistent with the finding that oltipraz inhibits reverse transcriptase, whereas metabolite III does not. Although the mean ED50 for metabolite III in acutely infected peripheral blood mononuclear cells was 18 microM, the ED50 was below 5 microM in three of eight independent experiments. Studies of metabolite III in combination with oltipraz in acutely infected peripheral blood mononuclear cells demonstrated significant antiviral synergy. These results raise the possibility that the in vitro potency of oltipraz may underestimate its antiretroviral activity in vivo. Based on these results, the pharmacokinetics of oltipraz and metabolite III will be compared with the pharmacodynamic effects of orally administered oltipraz in a forthcoming phase I/II trial of oltipraz in patients with p24 antigenemia.
Mol Pharmacol 1995 Jul
PMID:Inhibition of human immunodeficiency virus type 1 replication by 7-methyl-6,8-bis(methylthio)pyrrolo[1,2-a]pyrazine, an in vivo metabolite of oltipraz. 754 38


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