Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023473 (chronic myeloid leukemia)
 18,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cytidine deaminase, an enzyme that catalyses the deamination of both cytidine and its nucleoside analogues including the antineoplastic agents cytosine arabinoside (ara-C) and 5-azacytidine (5-azaC), has been partially purified from normal and leukemic human granulocytes. The purification procedure included heat precipitation at 70 degrees C, ammonium sulfate precipitation, calcium phosphate gel ion exchange, and Sephadex G-150 gel filtration. The enzyme has mol wt 51,000, isoelectric pH of 4.8, and maximum activity over a broad pH range of 5-9.5. The enzyme is stabilized by the presence of the sulfhydryl reagent, dithiothreitol. Cytidine deaminase from normal human granulocytes has a greater affinity for its physiologic substrate cytidine (K(m) = 1.1 x 10(-5) M) than for ara-C (8.8 x 10(-5) M) or 5-azaC (4.3 x 10(-4) M). Halogenated analogues such as 5-fluorocytidine and 5-bromo-2'-deoxycytidine also exhibited substrate activity, with maximum velocities greater than that of the physiologic substrates cytidine and deoxycytidine. No activity was observed with nucleotides or deoxynucleotides. The relative maximum velocity of the enzyme for cytidine and its nucleoside analogues remained constant during purification, indicating that a single enzyme was responsible for deamination of these substrates. Tetrahydrouridine (THU) was found to be a strong competitive inhibitor of partially purified deaminase with a K(i) of 5.4 x 10(-8) M. The biochemical properties of partially purified preparations of cytidine deaminase from normal and leukemic cells were compared with respect to isoelectric pH, molecular weight, and substrate and inhibitor kinetic parameters, and no differences were observed. However, normal circulating granulocytes contained a significantly greater concentration of cytidine deaminase (3.52+/-1.86 x 10(3)/mg protein) than chronic myelocytic leukemia (CML) cells (1.40+/-0.70 x 10(3) U/mg protein) or acute myelocytic leukemia (AML) cells (0.19+/-0.17 x 10(3) U/mg protein). To explain these differences in enzyme levels in leukemic versus normal cells, the changes in cytidine deaminase levels associated with maturation of normal granulocytes were studied in normal human bone marrow. Myeloid precursors obtained from bone marrow aspirates were separated into mature and immature fractions by Ficoll density centrifugation. Deaminase activity in lysates of mature granulocytes was 3.55-14.2 times greater than the activity found in the lysates of immature cells. Decreased enzyme activity was also found in immature myeloid cells from a patient with CML as compared to mature granulocytes from the same patient. These observations support the conclusion that the greater specific activity of cytidine deaminase in normal mature granulocytes as compared to leukemic cells is related to the process of granulocyte maturation rather than a specific enzymatic defect in leukemic cells.
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PMID:Purification and properties of cytidine deaminase from normal and leukemic granulocytes. 452 17

(1) Synthesis of deoxythymidine by either direct transfer of deoxyribosyl to thymine (pyrimidine deoxyribosyltransferase) or by a coupled deoxynucleoside phosphorylase mechanism is approximately twofold greater with normal leukocyte extracts (55 to 88% granulocytes) than with extracts prepared from leukocytes obtained from patients with chronic myelogenous leukemia. Activities in lymphocytes (normal or leukemic) are one-fifth the activity of normal granulocytes.(2) The lower activity in chronic myelogenous leukemia remains at 50% of normal even when patients are in hematologic remission with a normal per cent mature granulocytes in the peripheral blood.(3) The leukemic enzyme could not be distinguished from the normal by pH optima, thermal stability, or kinetic properties. The Km's for the deoxyribosyl acceptor and deoxyribosyl donors were identical for both enzymes. Both are subject to substrate inhibition by thymine and to inhibition by purine bases with similar Ki's. In addition, the transferase component of both the leukemic and the normal cell enzyme is activated by phosphate and arsenate. It appears, therefore, that there is no qualitative difference between the enzyme obtained from leukocytes of patients with chronic myelogenous leukemia and the enzyme obtained from normal leukocytes, suggesting that the difference in total cell activity is due to an actual decrease in amount of enzyme in chronic myelogenous leukemia or to a mixed cell population, one with a normal quantity of enzyme and the other with little or no active enzyme.(4) In both the normal cell and the leukemic cell extracts, transferase and phosphorylase activities could not be separated. The ratio of the two activities remained constant over a 140- and a 230-fold purification in normal and leukemic cell extracts, respectively. These and other observations indicate that transferase and phosphorylase activities are associated with the same protein.(5) The metabolism of pyrimidine and purine deoxynucleosides is similar for normal and leukemic cells. Catabolism of all deoxynucleosides tested was by direct phosphorolysis, except for deoxyadenosine which required initial deamination to deoxyinosine before phosphorolysis. In contrast to the greater rates of pyrimidine deoxynucleoside synthesis and cleavage with normal leukocyte extracts, the rates of purine deoxynucleoside synthesis and cleavage were approximately twofold greater with extracts prepared from cells of patients with chronic myelogenous leukemia. There was no significant difference in the rate of phosphorolytic cleavage of pyrimidine nucleosides (uridine) between the CML and normal leukocyte extracts.
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PMID:The enzymatic mechanisms for deoxythymidine synthesis in human leukocytes. IV. Comparisons between normal and leukemic leukocytes. 525 32

Lysosomal arylsulfatases A and B of peripheral leukocytes from patients with chronic myelogenous leukemia and from healthy subjects were studied. Two enzyme activities of leukemia cells were significantly higher than those of cells from healthy subjects, irrespective of total and differential counts of leukemic cells. Upon anion-exchange chromatography, the arylsulfatases of chronic myelogenous leukemia cells and normal leukocytes were separated into the basic B enzyme and its anionic variant (B1) and A enzyme. However, the amount of B1 enzyme relative to B enzyme or the activity ratio of B1 enzyme to total arylsulfatase B (B + B1) was higher in chronic myelogenous leukemia cells than in normal cells. The anionic property of the enzyme was found to be due to phosphate groups bound to the carbohydrate moiety of the arylsulfatase, based on the following results. When B1 enzyme was treated with alkaline phosphatase followed by isoelectric focusing, it was changed to a less anionic enzyme with heterogeneous components which are ascribed to phosphodiester groups linked to the heterogeneous carbohydrate moiety of the enzyme; no effect was observed by sialidase treatment. Upon treatment of B1 enzyme with endo-beta-N-acetylglucosaminidase H, which cleaves sugar chains of a high mannose type in glycoproteins, the anionic heterogeneous components were converted to the basic component similar to B enzyme. From our present and previous observations, it can be concluded that the increase of phosphorylated forms of the lysosomal hydrolase represents one characteristic of rapidly proliferating neoplastic cells.
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PMID:Lysosomal arylsulfatases of human leukocytes: increment of phosphorylated B variants in chronic myelogenous leukemia. 613 78

We have used enzyme specific inhibitors and heat inactivation to distinguish Leukocyte alkaline phosphate (LAP) from other organ-specific alkaline phosphatases as well as to compare LAP from normal granulocytes and leukemic cells with elevated LAP. The heat inactivation and inhibition curves of LAP are quite different from those of other organ-specific alkaline phosphatases. The inhibition curves and heat inactivation characteristics of LAP from normal granulocytes and that obtained from chronic granulocytic leukemia (CGL) blast phase cells with elevated LAP are identical. These data suggest that LAP is distinct from other organ-specific alkaline phosphatases, particularly placental alkaline phosphatase. We also conclude that the LAP present in cells with elevated levels is very similar or identical to that of normal granulocytes.
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PMID:Leukocyte alkaline phosphatase: another organ-specific alkaline phosphatase. 657 85

An improved method for the detection of deoxythymidine kinase (TK) in human sera is reported. The method which utilizes 125I-iododeoxyuridine (IdUrd) as a substrate was used to measure TK in sera from patients with different diseases. Sera collected during the acute stage of infectious mononucleosis were found to contain elevated levels of TK, in most cases 10-40 times the normal value. The serum TK activity disappeared gradually and reached a normal level within 4 weeks. Sera from patients with other viral infections contained in most cases normal serum TK levels except in connection with measles, rubella, varicella, herpes simplex virus and cytomegalovirus infections. Additional studies revealed that sera from patients with different types of advanced lymphomas, acute leukemias, chronic granulocytic leukemia and lung cancer of the small-cell type with metastases, contained high TK levels which fluctuated in parallel with alterations in activity of the disease. The TK activity in sera from patients with both mononucleosis and tumor disease was characterized by electrophoresis and by its ability to utilize cytidine triphosphate as the phosphate donor. The results showed that the serum TK has the same properties as the human cytosolar TKI, except in connection with varicella.
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PMID:Application of an in vitro assay for serum thymidine kinase: results on viral disease and malignancies in humans. 669 95

The adenine nucleoside analogue fludarabine phosphate in combination with cytosine-arabinoside (Ara-C) and granulocyte-colony stimulating factor (G-CSF) has recently proved effective in the treatment of poor-prognosis acute non-lymphoid leukaemia. We used this triple combination in a case of Ph1+ chronic myeloid leukaemia (CML) unresponsive to alpha interferon that had progressed to acute phase after 5 months of treatment with 6-mercaptopurine plus hydroxyurea. The patient was treated with four courses of fludarabine 30 mg/m2 + Ara-C 2 g/m2 (days 1-5) and G-CSF (from day 0 to polymorphonuclear (PMN) recovery). Bone marrow blasts decreased from 80% to less than 5%, and karyotyping showed a progressive clearance of Ph1+ metaphases (from 100% to 9% after the fourth course). The patient is presently receiving autologous bone marrow transplantation (ABMT). This therapeutic success in a patient for whom conventional treatment would usually be ineffective makes this combination worthy of further studies, in view of its wider use as a preparative regimen to ABMT in CML.
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PMID:FLAG (fludarabine+cytosine arabinoside+G-CSF) induces complete remission in acute-phase chronic myeloid leukaemia: a case report. 751 68

Glycoxidation products (GOPs), such as N epsilon-(carboxymethyl)lysine (CML) and pentosidine, are formed during reaction of glucose with protein under oxidative conditions in vitro. It is uncertain whether these GOPs are derived from oxidation of Amadori adducts on protein or from oxidation of glucose or intermediates formed prior to the Amadori rearrangement. To address this question, we reacted collagen with 250 mM glucose in 200 mM phosphate buffer, pH 7.4, under antioxidative conditions, yielding a protein rich in Amadori adducts, but with only traces of GOPs. This "preglycated" collagen was then exposed to [13C6]glucose under oxidative conditions, producing both natural and [13C2]-CML. At 200 mM phosphate buffer, [13C2]-CML was the major product, even at low (5 mM) [13C6]glucose concentration, indicating a limited role for Amadori compounds in formation of CML in high phosphate. The relative yields of natural and [13C2]-CML varied with phosphate concentration, becoming similar at more physiological (10 mM) phosphate. We conclude that during glycation of proteins at high phosphate concentrations in vitro, GOPs are formed primarily by oxidation of free glucose or rapidly-formed intermediates preceding the Amadori rearrangement, such as carbinolamine or Schiff base adducts. In contrast, at lower phosphate and glucose concentrations in vivo, the Amadori adduct may be the more significant precursor of GOPs. The fact that glycoxidation reactions proceed by multiple routes must be considered in the development of therapeutic approaches for inhibiting the Maillard reaction in diabetes.
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PMID:Pathways of formation of glycoxidation products during glycation of collagen. 757 27

We report a highly sensitive time-resolved immunofluorometric method for quantification of polymerase chain reaction (PCR)-amplified mRNA sequences. The PCR primers are labeled at their 5' ends, one with biotin and the other with a hapten. The modified primers are incorporated, during PCR, in the amplified product. The PCR product is captured, through its biotin moiety, to a streptavidin-coated solid phase and subsequently is detected with an alkaline phosphatase-labeled antibody. The phosphate ester of fluorosalicylic acid is used as a substrate. The fluorosalicylate produced forms a highly fluorescent ternary complex with Tb(3+)-EDTA, which is measured by time-resolved fluorometry. We chose the determination of PCR-amplified chronic myelogenous leukemia-specific mRNA as a model system. mRNA molecules corresponding to 0.1 leukemic cell in the presence of 0.5 million normal cells may be detected (signal-to-background ratio of 1.5). The method provides a sensitive and rapid nonisotopic alternative to Southern blot and hybridization with radioactive probes.
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PMID:Time-resolved immunofluorometric determination of specific mRNA sequences amplified by the polymerase chain reaction. 784 30

A new sensitive method for the measurement of 1-beta-D-arabinofuranosyl-CTP (ara-CTP), an intracellular active metabolite of 1-beta-D-arabinofuranosylcytosine (ara-C), in human materials in vivo has been established. An acid-soluble fraction containing ara-CTP was extracted from blastic cells by ara-C treatment with trichloroacetic acid (final concentration, 0.3 M) neutralized with an equal volume of cold freon containing 0.5 M tri-n-octylamine. The ara-CTP fraction was separated from the acid -soluble fraction by high-performance liquid chromatography (TSK gel diethylaminoethyl-2 SW column) eluted with 0.05 M phosphate buffer (pH 6.9) and 20% acetonitrile. ara-CTP was lyophilized, dephosphorylated to ara-C by incubation with 10 units alkaline phosphatase for 12 h at 55 degrees C, and measured by RIA using anti-ara-C serum. Recovery through the whole procedure was 92%. In the human chronic myelogenous leukemia cell line K562, the intracellular ara-CTP levels produced when the cells were incubated with ara-c were assayed as above, and they showed a linear increase depending on Ara-C concentrations from 0.01 to 10 microns, demonstrating a very close correlation with the labeled ara CTP levels yielded by cells on incubation with radiolabeled ara-C (r2 = 0.99). The detection limit was 0.1 pmol/5 x 10(6) cells, and a sample amount of only 5 x 10(6) cells was enough for each assay. In the clinical applications, our method proved capable of detecting a wide concentration range of ara-CTP produced when patients were treated with ara-C or its derivatives from very low to intermediate doses. No radiolabeled drug was necessary. The method was very useful for in vivo pharmacodynamic studies of ara-C therapy.
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PMID:A new sensitive method for determination of intracellular 1-beta-D-arabinofuranosylcytosine 5'-triphosphate content in human materials in vivo. 862 Apr 96

Nepsilon-(Carboxymethyl)lysine (CML) is an advanced glycation end product formed on protein by combined nonenzymatic glycation and oxidation (glycoxidation) reactions. We now report that CML is also formed during metal-catalyzed oxidation of polyunsaturated fatty acids in the presence of protein. During copper-catalyzed oxidation in vitro, the CML content of low density lipoprotein increased in concert with conjugated dienes but was independent of the presence of the Amadori compound, fructoselysine, on the protein. CML was also formed in a time-dependent manner in RNase incubated under aerobic conditions in phosphate buffer containing arachidonate or linoleate; only trace amounts of CML were formed from oleate. After 6 days of incubation the yield of CML in RNase from arachidonate was approximately 0.7 mmol/mol lysine compared with only 0.03 mmol/mol lysine for protein incubated under the same conditions with glucose. Glyoxal, a known precursor of CML, was also formed during incubation of RNase with arachidonate. These results suggest that lipid peroxidation, as well as glycoxidation, may be an important source of CML in tissue proteins in vivo and that CML may be a general marker of oxidative stress and long term damage to protein in aging, atherosclerosis, and diabetes.
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PMID:The advanced glycation end product, Nepsilon-(carboxymethyl)lysine, is a product of both lipid peroxidation and glycoxidation reactions. 862 37


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