2-ClATP exerts anti-tumoural actions not mediated by P2 receptors in neuronal and glial cell lines

Nadia D'Ambrosi, Stefano Costanzi, Daniela F. Angelini, Rosaria Volpini, Giuseppe Sancesario, Gloria Cristalli, Cinzia Volonté

Research output: Contribution to journalArticlepeer-review

Abstract

We investigated the effects of the ATP analogue and P2 receptor agonist 2-ClATP on growth and survival of different neuronal (PC12, PC12nnr5 and SH-SY5Y) and glial (U87 and U373) cell lines, by the use of direct count of intact nuclei, fluorescence microscopy, fluorescence-activated cell sorter analysis (FACS) and high pressure liquid chromatography (HPLC). 2-ClATP lowered the number of cultured PC12nnr5, SH-SY5Y, U87 and U373 cells to almost 5%, and of PC12 cells to about 35% after 3-4 days of treatment. EC50 was in the 5-25μM range, with 2-ClATP behaving as a cytotoxic or cytostatic agent. Analysis of the biological mechanisms demonstrated that pyridoxalphosphate-6- azophenyl-2′,4′-disulphonic acid (P2 receptor antagonist and nucleotidases inhibitor), but not Caffeine or CGS-15493 (P1 receptor antagonists) effectively prevented 2-ClATP-induced toxicity. 2-ClATP metabolic products (2-ClADP, 2-ClAMP, 2-Cladenosine) and new synthesis derivatives (2-CldAMP, 2-Cldadenosine-3′,5′-bisphosphate and 2-CldATP) exerted similar cytotoxic actions. Inhibition of both serum nucleotidases and purine nucleoside transporters strongly reduced 2-ClATP-induced cell death, which was conversely increased by the nucleotide hydrolyzing enzyme apyrase. The adenosine kinase inhibitor 5-iodotubericidin totally prevented 2-ClATP or 2-Cladenosine-induced toxicity. In summary, our findings indicate that 2-ClATP exerts either cell cycle arrest or cell death, acting neither on P2 nor on P1 receptors, but being extracellularly metabolized into 2-Cladenosine, intracellularly transported and re-phosphorylated.

Original languageEnglish
Pages (from-to)621-630
Number of pages10
JournalBiochemical Pharmacology
Volume67
Issue number4
DOIs
Publication statusPublished - Feb 15 2004

Keywords

  • Adenosine kinase
  • Cell death
  • PC12 cells
  • SH-SY5Y cells
  • U87, U373 cells

ASJC Scopus subject areas

  • Pharmacology

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