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:3.5.4.4 (adenosine deaminase)
5,136 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

High levels of deoxyadenosine and deoxyguanosine in patients with inherited deficiency of either adenosine deaminase or purine-nucleoside phosphorylase, respectively, are considered to be responsible for the associated immunological disorder. The mechanism involves phosphorylation to the corresponding deoxyribonucleoside triphosphates which subsequently inhibit the CDP-reducing activity of ribonucleotide reductase. Addition of deoxycytidine protects cells from the cytotoxic effects of deoxyadenosine and deoxyguanosine by competition for phosphorylation and by replenishing dCTP, the apparent limiting DNA precursor. Addition of cytidine, but not uridine, led to a reversal of deoxyguanosine and thymidine growth inhibition, comparable to that obtained with deoxycytidine. Analysis of the intracellular nucleotide pools showed that increased levels of cytidine ribonucleotides were sufficient to overcome the inhibitory effects of dGTP and dTTP on CDP reduction, thereby circumventing a depletion of the dCTP pool. A partial reversal of deoxyadenosine toxicity was also obtained with addition of cytidine. In this case little change in the dCTP level was observed, but a decreased dGTP pool appeared to be correlated with growth inhibition. High cytidine ribonucleotide levels partially prevented this effect. The present results may encourage the use of cytidine in combination with deoxycytidine as a pharmacological regime in treatment of immunodeficiency disease associated with increased deoxyribonucleotide levels.
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PMID:On the mechanism of deoxyribonucleoside toxicity in human T-lymphoblastoid cells. Reversal of growth inhibition by addition of cytidine. 387 78

Three general questions regarding nucleosides and lymphocytes are discussed: (a) Why are so many measurements being made of adenosine deaminase activity, what do the results mean, and why is there still disagreement about some of the conclusions; (b) what do we understand about nucleosides and lymphocyte death; and (c) to what extent do we really understand nucleoside and nucleotide metabolism in lymphocytes? Experimental studies show that treatment of mice with deoxycoformycin, to produce accumulation of deoxyadenosine, leads to rapid thymus involution, elevated dATP concentrations in thymus and liver, and inhibition of adenosylhomocysteine hydrolase in these tissues. Deoxyguanosine inhibits the growth of mouse lymphoma L5178Y cells, and this toxicity is prevented by deoxycytidine plus adenine. In cells treated with deoxyguanosine, concentrations of both GTP and dGTP are elevated, and this is not affected by deoxycytidine. Adenine, however, reduces GTP concentrations to normal, and prevents most of the elevation in dGTP concentrations. Contrary to previous belief, it has been demonstrated that lymphocytes and nucleated bone marrow cells will synthesize purine nucleotides de novo if incubated in an appropriate medium; carbon dioxide is particularly important for this process.
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PMID:Regulation of purine metabolism in lymphocytes. 387 99

We have studied the effects of various immunosuppressive drugs on the growth of human-derived T (MOLT-4) and B (MGL-8) lymphoblasts. In addition, we have examined whether the lymphotoxic effect of any of these drugs could be attributed to inhibition of either adenosine deaminase (ADA) or purine nucleoside phosphorylase (PNP). Results indicated that 1-beta-D-arabinofuranosylcytosine (Ara-C), methotrexate and chlorambucil were four to seven times more toxic for T than for B cells, while azathioprine, 6-thioguanine, 6-mercaptopurine, and 5-fluorouracil were highly toxic for both T and B cells. Cyclophosphamide and oxisuran were lymphotoxic only at concentrations exceeding 300 microM. Deoxyadenosine (50 microM), deoxyguanosine (10 microM) and deoxycoformycin (10 microM) failed to enhance T cell toxicity when individually combined with each drug. None of the drugs tested inhibited T or B lymphoblast ADA or PNP activity. With the exception of Ara-C, neither dATP nor dGTP accumulated in T lymphoblasts incubated in the presence of any of the drugs. We conclude that the cell culture system used in this investigation is useful for identifying lymphotoxic and T cell-specific immunosuppressive agents. However, none of the drugs studied appeared to function as an inhibitor of, or a competitive substrate for, either ADA or PNP.
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PMID:Effect of immunosuppressive agents on human T and B lymphoblasts. 640 81

The metabolism of 8-14C-labelled 2'-deoxyadenosine (dAR) and 2'-deoxyguanosine (dGR) has been investigated using lymphocytes in long-term culture transformed by Epstein-Barr (EB) virus (B-cells) from eight patients with different inherited purine enzyme defects. The use of such lines enabled accurate mapping of the route of metabolism by acting as a 'trap' for the radiolabel at specific points. With either substrate (25 microM) most of the label was recovered in the medium. Using dAR, less than 30% of the radiolabel was incorporated into cellular nucleotides. For dGR, values were less than 18%. Studies with dAR alone confirmed the principal route of metabolism was to hypoxanthine, with further metabolism (by lines with intact salvage pathways) to ATP and GTP in the ratio of approximately 4:1. Lack of accumulation of deoxyinosine in the purine nucleoside phosphorylase (PNP) deficient line, or hypoxanthine in the hypoxanthine guanine phosphoribosyltransferase (HGPRT) deficient line, using dAR together with the adenosine deaminase (ADA) inhibitor 2'-deoxycoformycin (dCF) at 10 microM, confirmed the effectiveness of ADA inhibition. Nevertheless, some ATP was still formed by all lines in the presence of dCF by a route as yet unknown. Only the ADA deficient lines formed dATP with dAR alone. However, some dATP was formed by all lines in the presence of dCF. A partially HGPRT deficient line formed extremely high dATP levels, well in excess of those formed by the T-cell line CEM. Studies with dGR revealed some interesting differences, a large proportion of the substrate being metabolized predominantly to xanthine by most enzyme deficient lines. In the PNP deficient line most of the substrate remained unmetabolized, but some dGTP was formed. No other enzyme deficient line formed any dGTP--with or without the PNP inhibitor 8-aminoguanosine (8-NH2GR)--with one exception. Again this was the partially HGPRT deficient line, which with the inhibitor again formed more dGTP than the T-cell line. Within the cells most of the substrate was metabolized to GTP, except in the PNP, and totally HGPRT deficient lines. Levels of GTP formed were not altered by the inhibitor, reflecting the lack of effective PNP inhibition by 8-NH2GR. Some counts were also found in ATP and IMP, confirming the existence of this route in mammalian cells of lymphoid origin. The results also support previous studies by us using cell lines with intact purine pathways, which demonstrated that, contrary to current beliefs, some B-cell lines are capable of accumulating high levels of deoxynucleotides.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Metabolism of deoxynucleosides by lymphocytes in long-term culture deficient in different purine enzymes. 642 79

Enzyme inhibitors used to simulate the inherited immunodeficiency diseases, adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP) deficiency, have been assessed in cultured human lymphocytes. Only 2'-deoxycoformycin (dCF) completely inhibited ADA in T and B cells at concentrations in excess of 5 microM. Erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) and 8-amino guanosine (8-NH2GR) did not inhibit ADA or PNP completely at any concentration. Detailed metabolic experiments comparing viability and deoxynucleotide accumulation showed that B cell lines of malignant origin also accumulated high levels of dATP from 2'-deoxyadenosine (dAR), and dGTP from 2'-deoxyguanosine (dGR) as effectively as T cells--even without inhibitors, however, dAR reduced cell viability only when ADA was inhibited by dCF, whilst dGR was equally toxic with or without inhibitor, even to a line which accumulated no dGTP. These experiments indicate that cultured lymphocytes, using either EHNA or 8-NH2GR as enzyme inhibitor, are not valid models of the toxicity to the immune system in inherited ADA or PNP deficiency. They demonstrate that the ability to accumulate high levels of dATP or dGTP is not exclusive to T cells and that the in vitro toxicity of dAR or dGR could relate to the use of excess substrate and/or accumulation in different nucleotide, not deoxynucleotide pools.
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PMID:B cells as well as T cells form deoxynucleotides from either deoxyadenosine or deoxyguanosine. 642 86

The toxicity of the deoxyribonucleosides, 2'-deoxyadenosine, 2'-deoxyguanosine, and thymidine, for human T lymphoblasts is mediated by the accumulation of the corresponding deoxyribonucleoside triphosphate (dATP, dGTP, or dTTP, respectively). We have examined whether leukemic cells of non-T-cell origin are capable of accumulating deoxyribonucleotides in culture and whether this capability correlates with the activities of purine metabolizing enzymes in these cells. We have found that non-T, non-B acute lymphoblastic leukemia cells with low ecto-5'-nucleotidase and high adenosine deaminase activities increase their dATP pools by greater than tenfold when exposed to deoxyadenosine and an inhibitor of adenosine deaminase in culture. Cells from 2 of 9 patients with chronic lymphocytic leukemia and 4 of 11 patients with acute nonlymphoblastic leukemia achieved similar elevations in dATP, but there was no relationship between dATP accumulation and adenosine deaminase, purine nucleoside phosphorylase, or ecto-5'-nucleotidase activities. Treatment of four individuals with acute lymphoblastic leukemia with the adenosine deaminase inhibitor, 2'-deoxycoformycin, resulted in elevations in plasma deoxyadenosine concentrations and in increments in lymphoblast dATP levels that were similar to those measured in lymphoblasts cultured with deoxyadenosine and deoxycoformycin prior to treatment. In vitro incubations of leukemic cells with deoxyribonucleosides may provide a rational basis for the use of these compounds as chemotherapeutic agents.
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PMID:Deoxyribonucleoside triphosphate accumulation by leukemic cells. 660 41

The biochemical mechanism of lymphocyte dysfunction with adenosine deaminase deficiency has been investigated using cultured phytohemagglutinin stimulated normal peripheral blood lymphocytes and the adenosine deaminase (ADA) inhibitor 2'-deoxycoformycin. The addition of deoxyadenosine to ADA-inhibited (but not to uninhibited) cells generated increased dATP pools (up to 50-fold greater than controls) and depressed the mitogen response. dATP Accumulation was accompanied by depletion of the other three deoxynucleoside triphosphate (dNTP) pools (dTTP, dCTP, and dGTP). Suppression of the mitogen response could be prevented ("reversed") to 90% of control levels by the addition of deoxynucleoside precursors for the depleted dNTPs at the initiation of mitogen stimulation. "Reversal" restored the dTTP and possibly the dGTP pools. Thus the mechanism of toxicity in this model appears to be inhibition of ribonucleotide reductase by massive accumulation of dATP, resulting in starvation for the other three deoxyribonucleoside triphosphates. "Reversibility" of this toxicity by providing sources for the missing three deoxynucleoside triphosphates argues for ribonucleotide reductase inhibition rather than other mechanisms of deoxyadenosine toxicity in this model.
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PMID:The mechanism of inhibition and "reversal" of mitogen-induced lymphocyte activation in a model of adenosine deaminase deficiency. 661 Apr 85

An inherited deficiency of adenosine deaminase (Ado deaminase; adenosine aminohydrolase, EC 3.5.4.4) causes severe combined immunodeficiency disease in humans. A similar deficiency in purine nucleoside phosphorylase (Puo phosphorylase; purine-nucleoside:orthophosphate ribosyltransferase, EC 2.4.2.1) engenders a selective cellular immune deficit. To elucidate the possible metabolic basis for the contrasting immunologic phenotypes, we compared the toxicity toward mature resting human lymphocytes of the Ado deaminase substrates deoxyadenosine and adenosine and the Puo phosphorylase substrate deoxyguanosine. When Ado deaminase was inhibited, micromolar concentrations of deoxyadenosine progressively killed nondividing helper and suppressor-cytotoxic T cells, but not B cells. The toxicity required phosphorylation, with subsequent dATP formation. The deoxyadenosine analogs 2-chlorodeoxyadenosine, 2-fluorodeoxyadenosine, and adenine arabinonucleoside also killed resting T cells. Cell death was unrelated to inhibition of adenosylhomocysteinase (EC 3.3.1.1) but was preceded by a gradual decline in ATP levels. As much as 1 mM deoxyguanosine did not impair resting lymphocyte viability, despite the synthesis of dGTP. The combination of 200 microM adenosine plus 500 microM homocysteine thiolactone killed dividing lymphocytes but had no discernible toxic effect toward resting T cells, which accumulated adenosylhomocysteine over a 4-hr period but thereafter excreted the nucleoside into the culture medium. The different clinical syndromes associated with genetic deficiencies of Ado deaminase and Puo phosphorylase may be explained by the ability of dATP to kill mature resting T lymphocytes by depleting ATP levels.
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PMID:Possible metabolic basis for the different immunodeficient states associated with genetic deficiencies of adenosine deaminase and purine nucleoside phosphorylase. 680 16

Biochemical and immunological properties of lymphocytes were measured repetitively over a period of 40 mo during enzyme replacement by transfusion in a child with adenosine deaminase (ADA) deficiency and severe combined immunodeficiency disease. Catalytically defective ADA protein is present in the child's cells. ADA activity in his lymphocytes is 7 nmol/min per 10(8) cells with 51 ng of ADA protein/10(8) cells by radioimmunoassay. ADA activities in normal cord and adult lymphocytes average 193 and 92 nmol/min per 10(8) cells, respectively, with 429 and 223 ng of ADA protein/10(8) cells. Deoxy(d)ATP accumulates in the patient's erythrocytes and lymphocytes. Transfusion of irradiated packed erythrocytes partially corrects the metabolic defects. Frank metabolic relapse occurs if transfusions are discontinued for several months. The amounts of dATP in erythrocytes and lymphocytes averaged 13 and 2 times normal, respectively, during periods when transfusions were administered every 2-4 wk. Deoxyguanosine triphosphate and deoxycytidine triphosphate in lymphocytes were normal on 11 occasions, but deoxyribosylthymine triphosphate was ninefold increased. On 11 occasions dATP was measured in lymphocytes and erythrocytes isolated simultaneously. There was a positive, but statistically insignificant, correlation between amounts of dATP in the two types of cells (r = 0.25,P > 0.1). The absolute peripheral lymphocyte count was correlated with the activity of ADA in circulating erythrocytes and with the response of lymphocytes to phytohemagglutinin (r = 0.64, P < 0.01; r = 0.49, P < 0.05). Response of lymphocytes to stimulation by phytohemagglutinin in vitro and absolute peripheral lymphocyte counts were not significantly correlated with levels of dATP in the erythrocyte or lymphocyte during periods of intensive therapy. Although there was objective improvement during enzyme replacement, the child remained immunodeficient and biochemically abnormal.
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PMID:Biochemical and functional abnormalities in lymphocytes from an adenosine deaminase-deficient patient during enzyme replacement therapy. 726 61

Template-independent nucleotide additions (N regions) generated at sites of V(D)J recombination by terminal deoxynucleotidyl transferase (TdT) increase the diversity of antigen receptors. Two inborn errors of purine metabolism, deficiencies of adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP), result in defective lymphoid development and aberrant pools of 2'-deoxynucleotides that are substrates for TdT in lymphoid precursors. We have asked whether selective increases in dATP or dGTP pools result in altered N regions in an extrachromosomal substrate transfected into T-cell or pre-B-cell lines. Exposure of the transfected cells to 2'-deoxyadenosine and an ADA inhibitor increased the dATP pool and resulted in a marked increase in A-T insertions at recombination junctions, with an overall decreased frequency of V(D)J recombination. Sequence analysis of VH-DH-JH junctions from the IgM locus in B-cell lines from ADA-deficient patients demonstrated an increase in A-T insertions equivalent to that found in the transfected cells. In contrast, elevation of dGTP pools, as would occur in PNP deficiency, did not alter the already rich G-C content of N regions. We conclude that the frequency of V(D)J recombination and the composition of N-insertions are influenced by increases in dATP levels, potentially leading to alterations in antigen receptors and aberrant lymphoid development. Alterations in N-region insertions may contribute to the B-cell dysfunction associated with ADA deficiency.
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PMID:Nucleotide pool imbalance and adenosine deaminase deficiency induce alterations of N-region insertions during V(D)J recombination. 1007 4


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