LY294002 and LY303511 sensitize tumor cells to drug-induced apoptosis via intracellular hydrogen peroxide production independent of the phosphoinositide 3-kinase-Akt pathway.

The phosphoinositide 3-kinase (PI3K)-Akt pathway is constitutively active in many tumors, and inhibitors of this prosurvival network, such as LY294002, have been shown to sensitize tumor cells to death stimuli. Here, we report a novel, PI3K-independent mechanism of LY-mediated sensitization of LNCaP prostate carcinoma cells to drug-induced apoptosis. Preincubation of tumor cells to LY294002 or its inactive analogue LY303511 resulted in a significant increase in intracellular hydrogen peroxide (H2O2) production and enhanced sensitivity to non-apoptotic concentrations of the chemotherapeutic agent vincristine. The critical role of intracellular H2O2 in LY-induced death sensitization is corroborated by transient transfection of cells with a vector containing human catalase gene. Indeed, overexpression of catalase significantly blocked the amplifying effect of LY pretreatment on caspase-2 and caspase-3 activation and cell death triggered by vincristine. Furthermore, the inability of wortmannin, another inhibitor of PI3K, to induce an increase in H2O2 production at doses that effectively blocked Akt phosphorylation provides strong evidence to unlink inhibition of PI3K from intracellular H2O2 production. These data strongly support death-sensitizing effect of LY compounds independent of the PI3K pathway and underscore the critical role of H2O2 in creating a permissive intracellular milieu for efficient drug-induced execution of tumor cells.