Transmembrane ion flux modifiers verapamil and ouabain modulate cytotoxic effects of extracellular ATP on human tumor cells in vitro

Extracellular ATP can often induce tumor cell cytotoxicity; however, the molecular mechanisms of these effects are mostly unknown. We investigated whether modifications in transmembrane ion fluxes are involved in the determination of ATP-cytotoxicity. We have found that cultured human tumor cells derived from colon (LoVo) and lung (A549) carcinomas are resistant to ATP, while LoVo-Dx cells (a doxorubicin-resistant derivative of LoVo cells) and melanoma GLL-19 cells are highly sensitive to this nucleotide. 48 h exposure to 100 nM verapamil increases sensitivity of LoVo and A549 cells to ATP. This effect is completely reverted by the addition of the calcium ionophore A23187. Conversely, 4 h exposure to 100 nM ouabain, which blocks the Na⁺/K⁺ exchange pump, neutralyzes ATP cytotoxicity against LoVo-Dx and GLL-19 cells. Furthermore, ATP-mediated cytotoxic effects on these cells are completely antagonized by ADP-β-S, a non hydrolyzable analogue of ATP. These findings suggest that transmembrane ion flux modifications play a critical role in ATP cytotoxicity and that ATP binding on surface receptors and probably nucleotide hydrolysis are needed for inducing cytotoxicity on human tumor cells.