Biphasic cytotoxic mechanism of extracellular ATP on U-937 human histiocytic leukemia cells: involvement of adenosine generation

Since extracellular ATP can exhibit cytotoxic activity in vivo and in vitro , its application has been proposed as an alternative anticancer therapy. I n this study we investigated the mechanisms of ATP-induced cytotoxicity in a human leukemic cell line (U-937). ATP added as a single dose exceeding 50 CIM was cytostatic or even cytotoxic for U-937 cells. Interestingly. growt h inhibition by ATP (50-3500 muM) showed a biphasic dose response. Up to 80 0 muM. ATP was cytotoxic in a dose-dependent manner (EC50 90 muM). In a ran ge between 800 and 2000 muM, cell count was markedly higher despite the hig her ATP concentrations. The cytotoxic effect of ATP could be antagonized by addition of uridine as a pyrimidine source and, alternatively, by addition of the nucleoside transmembrane inhibitor dipyridamole, The apoptosis-indu cing adenosine A(3) receptor was not involved in measurable quantities, sin es (1) adenosine did not lead to an elevation of intracellular calcium leve ls, and (2) tin unselective A(1-3) antagonist (ULS-II-80) could net abrogat e the cytotoxic effect. Experiments monitoring extracellular nucleotide met abolism confirmed the assumption that the long-term production and continuo us uptake of adenosine, which is extracellularly generated by degradation o f ATP, led to an intracellular nucleotide imbalance with pyrimidine starvat ion. The biphasic dose response to higher ATP concentrations could be expla ined by the rapid degradation of lower ATP concentrations (300 muM) to aden osine by serum-derived enzymes, whereas higher concentrations (900 muM) onl y produced small amounts of adenosine due to forward inhibition of AMP hydr olysis by prolonged high ADP levels. FAGS analysis revealed that at lower a denosine concentrations (300 muM) a reversible G(1) phase attest of the cel l cycle in as induced, whereas higher concentrations (1000 muM) triggered a poptosis. Considering ATP as a potential cytostatic drug, our data have imp ortant implications concerning metabolic interactions of administered nucle otides. (C) 2001 Elsevier Science B.V. Ah rights reserved.