adenosine-kinase and Leukemia--Lymphoid

Cell extracts from human leukemic T lymphoblasts and myeloblasts were chromatographed on DEAE-cellulose columns to separate purine deoxyribonucleoside, deoxyadenosine (dAdo) and deoxyguanosine (dGuo), phosphorylating activities. Three distinct purine deoxyribonucleoside kinases, a deoxycytidine (dCyd) kinase, an adenosine (Ado) kinase, and a deoxyguanosine (dGuo) kinase (the latter appears to be localized in mitochondria), were resolved. dCyd kinase contained the major phosphorylating activity for dAdo, dGuo, and 9-beta-D-arabinofuranosyladenine (ara-A). Ado kinase represented a second kinase for dAdo and ara-A while a third kinase for dAdo was found in mitochondria. dCyd kinase was purified about 2000-fold with ion-exchange, affinity, and hydrophobic chromatographies. On gel electrophoresis, both dCyd and dAdo phosphorylating activities comigrated, indicating that the activities are associated with the same protein. The enzyme showed a broad pH optimum ranging from pH 6.5 to pH 9.5. Divalent cations Mg2+, Mn2+, and Ca2+ stimulated dCyd kinase activity; Mg2+ produced the maximal activity. dCyd kinase from either lymphoid or myeloid cells showed broad substrate specificity. The enzyme used several nucleoside triphosphates, but ATP, GTP, and dTTP were the best phosphate donors. dCyd was the best nucleoside substrate, since dCyd kinase had an apparent Km of 0.3, 85, 90, and 1400 microM for dCyd, dAdo, dGuo, and ara-A, respectively. The enzyme exhibited substrate activation with both pyrimidine and purine deoxyribonucleosides, suggesting that there is more than one substrate binding site on the kinase. These studies show that, in lymphoblasts and myeloblasts, purine deoxyribonucleosides and their analogues are phosphorylated by dCyd kinase, Ado kinase, and dGuo kinase.

Topics: Adenosine Kinase; Cell Line; Child; Enzyme Activation; Humans; Kinetics; Leukemia, Lymphoid; Leukemia, Myeloid; Mitochondria; Phosphorylation; Phosphotransferases; Phosphotransferases (Alcohol Group Acceptor); Purine Nucleosides; Pyrimidine Nucleosides; Substrate Specificity

Adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP), 5'nucleotidase (5'NT), ecto-5'NT, hypoxanthine-guanine phosphoribosyltransferase(HGPRT), adenine phosphoribosyltransferase(APRT), adenosine kinase(AK), AMP deaminase (AMPD) and adenylate kinase(AdKin) activities were assayed in leukemic cells from bone marrow and/or peripheral blood of 43 newly diagnosed children with acute lymphoblastic leukemia(ALL). These enzyme activities have been investigated in relation to some immunological markers. ADA activity was higher in E-rosette positive leukemia(E+ ALL), while HGPRT, APRT, PNP, 5'NT, ecto-5'NT and AdKin activities were found to be lower in E+ ALL as compared to E- ALL. In common ALL (cALL) antigen positive leukemia, mean ADA activity was significantly lower as compared to cALL- leukemia, whereas PNP, 5'NT, ecto-5'NT and AdKin activities were significantly higher. cALL cells with cytoplasmic immunoglobulin M(IgM) heavy chains were found to have mean 5'NT activities twice as high as cALL cells lacking cytoplasmic IgM heavy chains. In two patients who had surface immunoglobulins on their cell membranes, low 5'NT activities were found. When measuring enzyme activities after 2-4 days of prednisone monotherapy, only mean ADA and HGPRT activities decreased in non-B, non-T ALL. These decreases were not significant in T-ALL patients. Mean enzyme activities in the leukemic cells of five patients with relapse were comparable to those in newly diagnosed patients, except for 5'NT, which was found to be within the activity range of control peripheral blood lymphocytes. It is concluded that ADA and AdKin activities are suitable as markers for E+ ALL and cALL+ leukemias respectively. 5'NT might help to distinguish between cALL cells having and lacking pre-B characteristics. Since 5'NT activity may also be decreased in B-ALL, it is not suitable as a T-ALL marker. Enzymes of purine metabolism in leukemic relapse need further investigation.

Topics: 5'-Nucleotidase; Adenine Phosphoribosyltransferase; Adenosine Deaminase; Adenosine Kinase; Adenylate Kinase; Adolescent; AMP Deaminase; Bone Marrow; Child; Child, Preschool; Humans; Hypoxanthine Phosphoribosyltransferase; Infant; Leukemia, Lymphoid; Nucleotidases; Purine-Nucleoside Phosphorylase; Purines

Topics: Adenosine Kinase; Cell Line; Deoxyadenine Nucleotides; Deoxyadenosines; Deoxycytidine; Deoxycytidine Kinase; Humans; Kinetics; Leukemia, Lymphoid; Mutation; Phosphotransferases; T-Lymphocytes

Deoxycoformycin (DCF) is an inhibitor of adenosine deaminase (ADA). Twenty-one courses of DCF were administered to 13 patients ranging in age from 15 to 78 yr. Eight patients had T-cell disorders, and five patients had non-T-cell malignancies. The i.v. bolus dose was escalated from 5 to 30 mg/sq m/day, and the duration of the courses ranged from 1 to 5 days. The DCF plasma half-life ranged from 4.9 to 6.2 hr and was independent of dose. The dose-limiting toxicities involved the central nervous system (CNS) and the kidneys. Other toxicities included bronchitis, decreases in hematocrit, arthralgias, and myalgias. Mortality was encountered in three patients. These toxic effects may have been secondary to the accumulation of the metabolites adenosine and deoxyadenosine. Deoxyadenosine and adenosine were both detectable in plasma (10(-6) M) and in urine (10(-3) M). Two partial remissions were observed: one in a patient with T-cell ALL and another in a patient with mycosis fungoides. Minimal responses characterized by either declines in peripheral blast counts or partial resolution of adenopathy were observed in five other patients. No responses were observed in six patients. These observations suggest that DCF is effective in the treatment of T-cell lymphoid malignancies.

Topics: Adenosine; Adenosine Deaminase Inhibitors; Adenosine Kinase; Adolescent; Adult; Aged; Bronchitis; Coformycin; Coma; Conjunctivitis; Deoxyadenosines; Female; Humans; Kidney Diseases; Leukemia, Lymphoid; Lymphoma; Male; Middle Aged; Mycosis Fungoides; Nucleoside Deaminases; Pentostatin; Phosphotransferases; Phosphotransferases (Alcohol Group Acceptor); Ribonucleosides

Cultured human T-cell leukemia lymphocytes have enhanced sensitivity to growth inhibition by deoxyadenosine. We have used flow cytometry to investigate the mechanism of deoxyadenosine toxicity in cultured T-leukemic cells. Comparative studies on deoxyadenosine-resistant Epstein-Barr virus-transformed B-lymphocyte cell lines were also performed. After exposure of T-cells to low concentrations of deoxyadenosine (3 microM), in the presence of an adenosine deaminase inhibitor (erythro-9-[3-(2-hydroxynonyl)]adenosine), accumulation of cells of cells with a G1 DNA content was demonstrated. In contrast, B-cell lines showed a similar degree of growth inhibition after exposure to 200 to 400 microM deoxyadenosine but were blocked in S phase. The T-cell G1 block was associated with a rise in the deoxyadenosine triphosphate pool, and both these phenomena were prevented by the addition of deoxycytidine. The biochemical mechanism of this G1 block induced by deoxyadenosine in T-cells is not understood.

Topics: Adenosine Kinase; B-Lymphocytes; Cell Cycle; Cell Transformation, Viral; Cells, Cultured; Deoxyadenosines; Deoxycytidine; Deoxyguanosine; Deoxyribonucleosides; Flow Cytometry; Herpesvirus 4, Human; Humans; Hydroxyurea; Leukemia, Lymphoid; T-Lymphocytes; Thymidine

Activities of adenosine deaminase (ADA), adenosine kinase (AK), adenine phosphoribosyltransferase (APRT), hypoxanthine guanine phosphoribosyltransferase (HGPRT), and purine nucleoside phosphorylase (PNP), all enzymes of the purine interconversion system, were determined in lymphocytes of 25 patients with chronic lymphatic leukemia (CLL) and in 23 controls. A statistically significant decrease of PNP activities and a reduction of ADA activities at borderline levels were found in the patients, whereas for the other enzymes assayed no deviation from normal values was observed.

Topics: Adenine Phosphoribosyltransferase; Adenosine Deaminase; Adenosine Kinase; Aged; Female; Humans; Hypoxanthine Phosphoribosyltransferase; Leukemia, Lymphoid; Male; Middle Aged; Nucleoside Deaminases; Pentosyltransferases; Purine-Nucleoside Phosphorylase

Purine metabolism and reutilization pathways were studied as they applied to normal and leukemic leukocytes. The enzyme activities were expressed in terms of the quantity of protein extracted and per 10(10) cells. Whereas the protein extracted and the enzyme activities from normal lymphocytes were relatively constant, considerable variation was noted in cases of chronic lymphocytic leukemia (CLL). This variability in the properties of the leukemic cells suggests that the difference may be useful in the subclassification of the leukemias. The studies of the complete enzyme system were done with 300 million cells. The extraction of 350,000 normal lymphocytes/mul gave a soluble protein concentration of 1.46+/-0.16 mg protein per ml, and the yield from the same number of CLL lymphocytes varied between 0.72 and 8.32 mg protein per ml. The 5'-nucleotidase activity gave an inverse correlation with the amount of extractable protein. In individual cases of CLL, the protein concentrations and the 5'-nucleotidase activities were found on either side of the normal values. In most cases, the adenosine deaminase of CLL lymphocytic cell extracts was lower than normal, and the adenosine kinase was higher; in the CLL cells, these two enzymes gave a positive correlation with one another. Little or no difference was observed in the activities of the purine nucleoside phosphorylases in extracts of normal or leukemic lymphocytes and granulocytes. The hypoxanthine-guanine and adenine phosphoribosyltransferase activities increased in the leukemic granulocytes but almost always showed a decrease in the CLL lymphocytes when compared with the normal cells. Most of the leukemic cells had greater than normal activities of the enzymes synthesizing phosphoribosyl pyrophosphate when tested with the purines. The total nucleotide produced from adenine and guanine with adenine- and hypoxanthine-guanine phosphoribosyltransferase was about equal in normal and leukemic lymphocytes, but the proportion of the adenosine 5'-triphosphate in the product was much greater with the leukemic cells. This suggested that the ribosyltransferase activities were the same in both types of cells, but the nucleoside kinases and the nucleoside diphosphate kinases were more active in the leukemic cells. Inosine monophosphate dehydrogenase was less active than normal in the CLL cell extracts and was not directly related to the amount of inosine monophosphate generated from hypoxanthine.

Topics: Adenine Phosphoribosyltransferase; Adenosine Deaminase; Adenosine Kinase; Glucosephosphates; Granulocytes; Humans; Hypoxanthine Phosphoribosyltransferase; IMP Dehydrogenase; Inosine Monophosphate; Leukemia, Lymphoid; Leukocytes; Lymphocytes; Nucleotidases; Phosphotransferases; Purine Nucleotides; Purine-Nucleoside Phosphorylase; Purines; Ribose-Phosphate Pyrophosphokinase; Ribosemonophosphates