Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The in vitro transcription of viral specific DNA sequences in nuclei and chromatin isolated from mouse cells chronically infected with Moloney murine leukemia virus (Mo-MuLV) has been studied. The in vitro RNA synthesized by Escherichia coli RNA polymerase has been isolated by sulfhydryl affinity column following reaction in the presence of 5-mercuriuridine triphosphate. By comparison of the Crt curves of the in vitro RNA with that of 70S viral RNA, the content of viral sequences is found to be 1.3% in nuclei product and 0.24% in chromatin product which is lower than the 2.5% found in chromatin associated RNA. This latter value, however, is very close to the in vivo viral RNA content in pulse-labeled [3H]RNA of the infected cells. Unexpectedly, it is observed that over 20% of the chromatin associated RNA prelabeled in vivo with [5-3H]uridine is elongated and tagged with Hg atoms during RNA synthesis catalyzed by the exogenous E. coli RNA polymerase in the presence of Hg-UTP. The elongation reaction is dependent on the presence of all four nucleotide triphosphates and appears to be due to E. coli RNA polymerase per se. It is suggested that most of the viral specific sequences observed in the in vitro RNA products are very likely initiated and derived from the chromatin associated species. The implication of the present findings for in vitro RNA synthesis in nuclei and chromatin as related to regulation of gene expression is discussed.
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PMID:In vitro transcription of Moloney leukemia virus genes in infected cell nuclei and chromatin: elongation of chromatin associated ribonucleic acid by Escherichia coli ribonucleic acid polymerase. 32 7

A density cut method was used to prepare two subpopulations of bone marrow cells which were enriched (BE) and depleted (BD) respectively of blast cells. Marrow aspirates were obtained from 32 patients, 26 of whom had acute leukaemia. The uptake of a variety of chemotherapeutic agents by these two subpopulations and the acid-soluble ribonucleotide profiles of the populations were compared and significant differences were found in drug uptake by BE and BD subpopulations. For some drugs such as cytosine arabinoside, uptake was greatest by the BE cells, while for 5-azacytidine the BD subpopulation took up the greatest amount of the drug. The preparation of the BE subpopulation also permitted the recognition of several patients with acute leukaemia whose blast cells possess nondetectable levels of ATP, UTP, and GTP components in their acid-soluble fractions. The studies presented demonstrate the necessity of using purified cell populations when characterizing the drug uptake patterns and the soluble ribonucleotide profiles of the leukaemic cells. A simple method for enriching bone marrow aspirates for leukaemic cells is also presented.
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PMID:Drug uptake and ribonucleotide profiles of blast-enriched and blast-depleted human bone marrow cell populations. 60 55

Metabolic effects and mode of cytotoxicity of 5-deazaacyclotetrahydrofolate (5-DACTHF, BW543U76), a glycineamide ribonucleotide transformylase inhibitor, were studied in MOLT-4 cells, a human T-cell leukemia line. 5-DACTHF inhibits purine synthesis with 50% inhibitory concentration values of 0.5 microM and 0.08 microM following 6- or 24-h exposure to drug, respectively. At 6 h, adenine nucleotide synthesis is preferentially inhibited over guanine nucleotide synthesis. A similar effect was observed with another glycineamide ribonucleotide transformylase inhibitor, 5,10-dideazatetrahydrofolate. GTP was depleted to 40% of control and ATP to 10% of control by 5 microM 5-DACTHF. After a transitory increase, UTP and CTP were depleted to 30% of control. Deoxynucleotides were also depleted by the drug; dCTP was depleted to the greatest extent, followed by dATP, dTTP, and dGTP, respectively. MOLT-4 cell growth was inhibited by 5-DACTHF with a 50% inhibitory concentration of 0.066 microM. Complete reversal was effected by hypoxanthine, and there was no reversal by thymidine. The drug was cytotoxic to MOLT-4 cells in the range 0.25 to 5.0 microM, but a minimum of 48 h was required for trypan blue-staining dead cells to appear. The rate and extent of kill with the thymidylate synthase inhibitor 2-methyl-10-propargyl-5,8-dideazafolate was greater than with 5-DACTHF, which indicates that kill by inhibition of thymidylate synthase is more effective than that by inhibition of purine synthesis. Electron microscopy of MOLT-4 cells exposed to 5-DACTHF showed electron-dense mitochondria and nuclear changes reminiscent of apoptosis. These morphological changes were accompanied by the appearance of DNA strand breaks at approximately 180-base pair intervals (internucleosomal breaks). Concomitant proteolysis of nuclear proteins poly(ADP-ribose) polymerase and lamin B was observed.
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PMID:Metabolic effects and kill of human T-cell leukemia by 5-deazaacyclotetrahydrofolate, a specific inhibitor of glycineamide ribonucleotide transformylase. 151 46

Exposure of mouse L1210 leukemia cells to 25 microM brequinar for 4 h results in large accumulations of N-carbamyl-L-aspartate and L-dihydroorotate to cellular concentrations of 8.5 mM and 0.8 mM, respectively, while UTP and CTP decrease to 4% of their initial levels; incorporation of [14C]bicarbonate into nucleic acids (DNA and RNA) was decreased to 47%. These data provide direct evidence for inhibition of DHO dehydrogenase by brequinar in growing cells. Exposure of leukemia cells to 200 microM ciprofloxacin for 4 h did not affect de novo pyrimidine nucleotide biosynthesis or the incorporation of [14C]bicarbonate into nucleic acids but resulted in a general decrease in nucleoside triphosphates, with concomitant accumulation of nucleoside mono- and diphosphates (the adenylate energy charge decreased from 0.89 to 0.69), consistent with inhibition of the electron transport chain or uncoupling of oxidative phosphorylation.
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PMID:Effects of brequinar and ciprofloxacin on de novo nucleotide biosynthesis in mouse L1210 leukemia. 196 81

6-L-Thiodihydroorotate (TDHO) and 2-oxo-1,2,3,6-tetrahydropyrimidine-4,6-dicarboxylate (HDDP) are potent inhibitors of mammalian dihydroorotase in vitro (R. I. Christopherson, K. J. Schmalzl, E. Szabados, R. J. Goodridge, M. C. Harsanyi, M. E. Sant, E. M. Algar, J. E. Anderson, A. Armstrong, S. C. Sharma, W. A. Bubb, and S. D. Lyons, Biochemistry, 28: 463-470, 1989). Using human CCRF-CEM leukemia cells growing in culture, TDHO and HDDP as the free acids have 50% inhibitory concentration (IC50) values of 32 microM and greater than 1000 microM, respectively, whereas for TDHO methyl ester, the IC50 value is 25 microM, and for HDDP dimethyl ester, the IC50 value is 21 microM. These IC50 values were not affected by addition of dihydroorotate, uridine, or deoxycytidine to the culture medium. TDHO methyl ester (25 microM) had only slight inhibitory effects upon the dihydroorotase reaction of de novo pyrimidine biosynthesis in growing leukemia cells, cells arrested in G2 + M phases of the cell cycle. At 250 microM TDHO methyl ester, analysis of cell extracts by high-performance liquid chromatography showed that after 4 h carbamyl aspartate had accumulated from undetectable levels to 760 microM, whereas UTP decreased from 580 to 110 microM and CTP from 350 to 86 microM, indicating inhibition of dihydroorotase in growing leukemia cells. IMP accumulated from 63 to 350 microM, total guanylates increased while adenylates decreased, and the adenylate energy charge decreased from 0.91 to 0.69 after 4 h. The cellular concentration of 5-phosphoribosyl 1-pyrophosphate increased from 180 to 290 microM due to sparing from pyrimidine nucleotide biosynthesis resulting in complementary stimulation of the de novo purine pathway. HDDP dimethyl ester at concentrations of up to 250 microM had no discernable effect upon pyrimidine or purine nucleotide biosynthesis. At 25 microM HDDP-dimethyl ester, cells arrested in G2 + M phases initially, with accumulation of cells in G1/G0 at later times. These data suggest that the primary mechanisms of growth inhibition for TDHO and HDDP involve inhibition of cell cycle progression from late G2 or M phase to G1 phase and that blockade of the pyrimidine pathway by TDHO is a secondary effect found at higher concentrations.
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PMID:Cytotoxic effects of dihydroorotase inhibitors upon human CCRF-CEM leukemia. 197 49

The intracellular concentration of 1-beta-D-arabinofuranosylcytosine (ara-C) for half-maximal phosphorylation by leukemic blasts obtained directly from patients was 2.1 +/- 2.5 microM (median, 1.3 microM, N = 25), and the rate of ara-C accumulation actually declined at concentrations above 20 microM in 35% of these cell populations. These apparent Km values for cellular phosphorylation were an order of magnitude lower than the Km of deoxycytidine (dCyd) kinase for ara-C with ATP as phosphate donor. dCyd kinase was purified from human leukemia cells and assayed for [3H]ara-C kinase activity with a mixture of 7 nucleotides at their approximate cellular concentrations or with a single nucleotide deleted. At low or high ara-C concentrations, ATP, GTP, CTP, or dTTP could be eliminated without significantly altering the rate. The only potential phosphate donor that was clearly important was UTP, since its deletion reduced the rate to only 25% of that with the complete mix. As anticipated, eliminating dCTP, the end product of this salvage pathway, moderately increased the rate by 50% at 0.4 microM ara-C or by 26% at 40 microM ara-C. At 40 microM ara-C, deleting UDP from the mix increased the rate more than deleting dCTP. dCTP was less inhibitory against 1 mM UTP (50% inhibitory concentration, 26 microM) than against 4 mM ATP (50% inhibitory concentration, 2.2 microM). In kinetic assays with 4 mM ATP and variable ara-C, UDP was a potent uncompetitive inhibitor with a Ki of 4 microM; the Ki for ADP was 1000-fold higher. Direct fit of kinetic data to the Michaelis equation yielded a Km for ara-C of 49 microM with 4 mM ATP as the phosphate donor; however, there was evidence of negative cooperativity with a Hill coefficient of 0.7. High ara-C Km values were also obtained with GTP and CTP, but with no evidence of cooperativity. With 1 mM UTP, the Km was 1.5 microM with moderate substrate inhibition; thus the kinetic data with UTP were similar to those for ara-C phosphorylation by intact cells. UDP was less potent versus UTP than versus ATP. It lowered the Vmax and enhanced the ara-C substrate inhibition without altering the Km. When 1 mM UTP and 4 mM ATP were mixed, the kinetic pattern was similar to that for UTP alone. The Km for UTP with [3H]dCyd as the phosphate acceptor of 0.8 microM was 25-fold lower than the Km for ATP of 20 microM.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:A critical role for uridine nucleotides in the regulation of deoxycytidine kinase and the concentration dependence of 1-beta-D-arabinofuranosylcytosine phosphorylation in human leukemia cells. 202 37

Myeloid differentiated human leukaemia (HL-60) cells contain a soluble phospholipase C that hydrolysed phosphatidylinositol 4.5-bisphosphate and was markedly stimulated by the metabolically stable GTP analogue guanosine 5'-[gamma-thio]triphosphate (GTP[S]). Half-maximal and maximal (up to 5-fold) stimulation of inositol phosphate formation by GTP[S] occurred at 1.5 microM and 30 microM respectively. Other nucleotides (GTP, GDP, GMP, guanosine 5'-[beta-thio]diphosphate. ATP, adenosine 5'-[gamma-thio]triphosphate, UTP) did not affect phospholipase C activity, GTP[S] stimulation of inositol phosphate accumulation was inhibited by excess GDP, but not by ADP. The effect of GTP[S] on inositol phosphate formation was absolutely dependent on and markedly stimulated by free Ca2+ (median effective concn. approximately 100 nM). Analysis of inositol phosphates by anion-exchange chromatography revealed InsP3 as the major product of GTP[S]-stimulated phospholipase C activity. In the absence of GTP[S], specific phospholipase C activity was markedly decreased when tested at high protein concentrations, whereas GTP[S] stimulation of the enzyme was markedly enhanced under these conditions. As both basal and GTP[S]-stimulated inositol phosphate formation were linear with time whether studied at low or high protein concentration, these results suggest that (a) phospholipase C is under an inhibitory constraint and (b) GTP[S] relieves this inhibition, most likely by activating a soluble GTP-binding protein.
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PMID:Guanosine 5'-[gamma-thio]triphosphate-stimulated hydrolysis of phosphatidylinositol 4,5-bisphosphate in HL-60 granulocytes. Evidence that the guanine nucleotide acts by relieving phospholipase C from an inhibitory constraint. 217 6

Rapid kinetic techniques were used to study the transport and salvage of uridine and other nucleosides in mouse spleen cells. Spleen cells express two nucleoside transport systems: (1) the non-concentrative, symmetrical, Na+-independent transporter with broad substrate specificity, which has been found in all mammalian cells and is sensitive to inhibition by dipyridamole and nitrobenzylthioinosine; and (2) a Na+-dependent nucleoside transport, which is specific for uridine and purine nucleosides and resistant to inhibition by dipyridamole and nitrobenzylthioinosine. The kinetic properties of the two transporters were determined by measuring uridine influx in ATP-depleted cells and dipyridamole-treated cells, respectively. The Michaelis-Menten constants for Na+-independent and -dependent transport were about 40 and 200 microM, respectively, but the first-order rate constants were about the same for both transport systems. Nitrobenzylthioinosine-sensitivity of the facilitated nucleoside transporter correlated with the presence of about 10,000 high-affinity (Kd = 0.6 nM) nitrobenzylthioinosine-binding sites per cell. The turnover number of the nitrobenzylthioinosine-sensitive nucleoside transporter was comparable to that of mouse P388 leukemia cells. The activation energy of this transporter was 20 kcal/mol. Entry of uridine via either of the transport routes was rapidly followed by its phosphorylation and conversion to UTP. The Michaelis-Menten constant for the in situ phosphorylation of uridine was about 50 microM and the first-order rate constants for phosphorylation and transport were about the same. The spleen cells also efficiently salvaged adenosine, adenine, and hypoxanthine, but not thymidine.
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PMID:Na+-dependent and -independent transport of uridine and its phosphorylation in mouse spleen cells. 273 Sep 9

DUP 785 (NSC 368390; Brequinar sodium) is a new inhibitor of pyrimidine de novo biosynthesis with antitumor activity against several experimental tumors. DUP 785 inhibits the mitochondrial enzyme dihydroorotate dehydrogenase, blocking the conversion of dihydroorotate to orotate. We examined the influence of exposure time to DUP 785 on its growth-inhibitory effects in L1210 murine leukemia and WiDR human adenocarcinoma cells and the effects of pyrimidine (deoxy) nucleosides on reversal of growth-inhibition. The results were correlated with changes in intracellular pyrimidine nucleotide pools and cell cycle distribution. In L1210 cells, a continuous exposure to 25 microM DUP 785 up to 96 hr caused complete growth inhibition. A 2 hr exposure of cells to the drug did not affect growth. In WiDR cells, exposure to the drug for 1-24 hr, followed by cultivation in drug-free medium resulted in recovery of growth. However, cells exposed to the drug for 48 hr or longer were not able to resume growth when recultured in drug-free medium. Reversal studies were performed to know whether selective depletion of one of the pyrimidine (deoxy) nucleotides might be related to the growth-inhibitory effects of DUP 785. Neither thymidine, deoxycytidine alone, deoxycytidine plus tetrahydrouridine; nor cytidine plus tetrahydrouridine added after 24 hr were able to reverse cell growth inhibition induced by 25 microM DUP 785. However, uridine and cytidine alone reversed growth inhibition. UTP and CTP pools in L1210 cells decreased to about 30-40% of control levels after 4 hr of drug exposure, while dTTP and dCTP pools decreased to about 30% of control levels. There were no significant changes in purine nucleotide pools. In WiDR cells, UTP and CTP pools decreased rapidly after drug exposure and were substantially depleted after 24 hr. Reculture of cells in drug-free medium resulted in a significant recovery of UTP and CTP levels only for cells exposed to DUP 785 for 1-24 hr. For cells exposed to the drug for 48 and 72 hr recovery of nucleotide pools was minimal. In L1210 cells, a 12-hr exposure to the drug caused an accumulation of cells in the early S-phase. In WiDR cells, there was a clear accumulation of cells in the S-phase of the cell cycle after 24 hr drug exposure.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:DUP 785 (NSC 368390): schedule-dependency of growth-inhibitory and antipyrimidine effects. 284 Sep 10

In vivo effects of DL-alpha-difluoromethylornithine (DFMO) on the metabolism of polyamines and nucleotide phosphates were monitored in P388/S leukemia cells grown intraperitoneally in BDF1 inbred male mice. Inhibiting the ornithine decarboxylase (ODC) activity DFMO depleted putrescine and spermidine to 30-50 and 50-60%, respectively, and increased spermine to 25-60% compared with the controls, when given as 2% solution in drinking water of the tumor-bearing animals. DFMO treatment caused a parallel 56% elevation of total nucleotide content in tumor cells with distinct and significant increase of some nucleotide phosphates. The most pronounced alterations were shown in the intracellular UTP (202%), CTP (103%), ADP (92%) and ATP (71%) concentrations. Changes in polyamine and nucleotide phosphate metabolisms were dependent on tumor progression. A possible explanation of the metabolic events induced by DFMO is discussed.
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PMID:In vivo effects of DL-alpha-difluoromethylornithine on the polyamine and nucleotide phosphate metabolism in P388/S leukemia cells. 308 89


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