Effect Of Ado Research Articles

MECHANISM OF ADENOSINE TOXICITY TO ADENOSINE KINASE DEFICIENT MAMMALIAN CELLS: 30

Somatic cell genetics has been used to probe the mechanism of adenosine (Ado) toxicity to mammalian cells deficient in Ado deaminase and Ado kinase. Ado resistant clones of an Ado kinase deficient murine lymphoblastoid cell line (R1.1) were isolated and characterized. In deoxycoformycin supplemented medium, the mutant clones were 10-30 fold more resistant than parental cells to the anti-proliferative actions of Ado, 3-deaza-Ado, carbocyclic-Ado, adenine, 5'-methylthioadenosine, and several other Ado analogs. The mutants were normally sensitive to the toxic effects of deoxyadenosine and adenine arabinoside. Levels of S-adenosylmethionine (SAM) were 50 pmols/106 cells in the parental line, compared to 250-350 pmois/106 cells in the mutant. Methionine adenosyitransferase activity was 1.5-3.2 fold higher in the Ado-resistant cells than in parental lymphoblasts, and varied inversely with the medium methionine concentration. Ado induced the accumulation of equivalent amounts of S-adenosylhomocystelne (SAH) in both cell types. However, the SAH to SAM ratio in the Ado-resistant mutants never exceeded 0.1. These results show that (i) methionine adenosyltransferase is an inducible enzyme in mammalian cells, (ii) the toxic effects of Ado, adenine, and many Ado analogs, can be averted by increasing the velocity of SAM synthesis.

Effect of adenosine on gluconeogenesis in the perfused liver of chicken and guinea-pig

1. 1. Glucose formation from lactate by the perfused liver of 48 hr starved chickens was strongly inhibited by adenosine (Ado); the half-maximal inhibition was attained at 40 μM. This effect was paralleled by a four- to five-fold increase of ATP content as determined in freeze-clamped liver. 2. 2. In chicken liver homogenate gluconeogenesis from precursors such as alanine, glutamate, glutamine and aspartate, which are not converted into glucose by the perfused chicken liver, proceeded at rates equal to or higher than that with lactate, being markedly inhibited by Ado. 3. 3. In the perfused guinea-pig liver glucose synthesis with lactate, propionate, glycerol and fructose was also inhibited by Ado; however, when precursors such as pyruvate, glutamine and a mixture of lactate + pyruvate were supplied to the liver Ado did not inhibit gluconeogenesis. 4. 4. Assay of adenine nucleotides in the perfused guinea-pig liver, stopped by freeze-clamping technique in a number of experimental variants, revealed no correlation between the rate of gluconeogenesis and the changes induced by Ado in the adenine nucleotide pool. 5. 5. In the perfused liver of both chicken and guinea-pig Ado produced an increase of the lactate to pyruvate ratio and, in general, a diminution of the content of malate-aspartate shuttle intermediates. 6. 6. The results are interpreted as suggesting that the inhibitory effect of Ado on hepatic gluconeogenesis is not necessarily mediated by the changes in the adenine nucleotide pool.

Inhibition of immune cell function by adenosine: biochemical studies.

Adenosine (Ado) has been shown to inhibit the cytolysis of tumor cells by specifically-sensitized mouse lymphocytes1. The mechanism of this immunosuppressive effect of Ado is of interest in view of the apparent causal relationship between Ado deaminase deficiency and severe combined immunodeficiency disease2. Thus far, Ado has been shown to cause two distinct biochemical effects in the cytolytic lymphocytes: (i) an elevation of adenosine 3′:5′-monophosphate (cAMP)1; and (ii) an elevation of S-adenosylhomocysteine (AdoHcy)3. Ado has little or no effect on the pool sizes of CTP, UTP, ATP or GTP in these cells1,4. Studies with other agents have shown that a selective elevation of either cAMP, as caused by prostaglandin E 1 5 , or AdoHcy, as caused by 3-deazaadenosine3, is sufficient to inhibit the cytolytic activity of these lymphocytes. The present studies were therefore undertaken in an attempt to discern the relative importance of cAMP and AdoHcy to this immunosuppressive action of Ado.

Biochemical and pharmacologic studies with 1-β- d-arabinofuranosylcytosine 5′-adamantoate (NSC-117614), a depot form of cytarabine

In the treatment of L1210 leukemic mice, the antitumor activity observed with 1-β- d-arabinofuranosylcytosine 5′-adamantoate (AdO- ara-C) suggested that the agent represents a molecular depot form or sustained action form of 1-β- d-arabinofurano-sylcytosine ( ara-C, cytarabine). The results of the present study provide strong support for this hypothesis. Many biochemical similarities between ara-C and AdO- ara-C were observed, suggesting similar or identical modes of action. Cytotoxicity of both agents in cell culture could be prevented with deoxycytidine. Like ara-C, AdO- ara-C markedly inhibited DNA synthesis with little effect on RNA or protein synthesis. Cross-resistance (L1210 cells in culture) was also observed. The differences observed in vitro (e.g. lower intrinsic Cytotoxicity, lower extent of uptake by L1210 cells, slower kinetics of inhibition of DNA synthesis with the derivative) were also consistent with the hypothesis that hydrolysis of AdO- ara-C to ara-C is required for cytotoxic activity. Direct evidence for hydrolysis was obtained in studies of the metabolism in vitro of AdO- ara-C in mammalian plasma and in plasma level studies in mice. Inhibition of enzymatic hydrolysis with an esterase inhibitor (eserine sulfate) markedly reduced the Cytotoxicity of AdO- ara-C towards L1210 cells in culture. Plasma level and excretion studies indicated that i.p. administration of AdO- ara-C to mice yielded cytotoxic ara-C levels which persisted for much longer than is possible with a single dose of the parent compound itself. These data, when considered with those concerning the effects of low levels of ara-C in contact with cells in culture for long periods help to explain the unusual therapeutic effects of AdO- ara-C.