The phosphatidylinositol 3-kinase inhibitor LY294002 binds the estrogen receptor and inhibits 17β-estradiol-induced transcriptional activity of an estrogen sensitive reporter gene

Estrogen receptors (ERs) are members of the superfamily of ligand-activated transcription factors. In addition to the classical, hormone-mediated activation, ERs may alternatively be activated in a ligand-independent manner by a variety of agents including growth factors, neurotransmitters and cAMP. It has been demonstrated that the phosphatidylinositol 3 (PI3)-dependent kinase/Akt pathway may activate the ER a by increasing the activity of both estrogen independent activation function-1 and estrogen-dependent activation function-2 domains. The Akt phosphorylation site in the ER is Serl67. Phosphorylation of this residue is inhibited by LY294002, which blocks the P13-kinase/Akt pathway. In the course of studies examining the effects of LY294002 on ligand-independent activation of ERs in L cells, we found that LY294002 exhibits antiestrogenic effects in a dose-dependent manner. By competition binding assays, we found that LY294002 specifically displaced radiolabelled estradiol from ERs with an IC50 of 11 +/- 0.06 nM, being an estradiol competitor as effective as the antiestrogens ICI182,780 (IC50, 21 +/- 0.13) and 4-OH-tamoxifen (IC50, 15 +/- 0.09). Further, LY294002 irreversibly blocked estrogen-induced transactivation of an estradiol-sensitive reporter gene. These findings are of particular importance in the interpretation of studies demonstrating ERs inactivation by the PI3-kinase inhibitor. Our studies show that an apparent block of ER activation cannot be dissociated from inhibition of ligand-mediated events. Thus, this effect can be the result of the ability of LY294002 to bind the ERs and inhibit transactivation of estrogen-regulated genes. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.