Expression of Insulin-Like Growth Factor Binding Protein-2 by MCF-7 Breast Cancer Cells Is Regulated through the Phosphatidylinositol 3-Kinase/AKT/Mammalian Target of Rapamycin Pathway

IGF binding protein-2 (IGFBP-2) has been implicated in the development and spread of a number of tumor types, and its abrogation in experimental models of cancer is associated with decreased tumor growth. This suggests that targeted inhibition of IGFBP-2 expression in some cancers may have therapeutic benefit. In this study, we investigated signaling pathways involved in extracellular IGFBP-2 expression in an IGF- and estrogen-responsive breast cancer cell line, MCF-7. IGFBP-2 was present at approximately 150 ng per 10(6) cells in serum-free MCF-7-conditioned medium and constituted the predominant IGFBP. Inhibition of the phosphatidylinositol 3-kinase signaling pathway using LY294002, or the downstream signaling intermediate mammalian target of rapamycin using rapamycin, markedly reduced IGFBP-2 in conditioned medium to approximately 25% of untreated levels (P < 0.001); there was no effect of inhibition of p38 MAPK, and an inhibitor of p44/42 MAPK activation, PD98059, caused only a slight reduction in extracellular IGFBP-2. IGFBP-2 levels were increased 25-30% by estradiol, whereas IGF-I (100 ng/ml) increased IGFBP-2 levels 2-fold (P < 0.001) by a type 1 IGF receptor (IGFR1)-dependent mechanism. Estradiol enhanced the effect of IGF-I on IGFBP-2 levels, and this was associated with increased phosphorylation of IGFR1. Basal, IGF-, or estradiol-stimulated IGFBP-2 was abrogated by LY294002 and rapamycin and an inhibitor of IGFR1 tyrosine kinase activity, AG1024. Modulation of intracellular hypoxia-inducible factor-1alpha had no effect on IGFBP-2 expression. These findings indicate that IGFBP-2 is regulated predominantly through the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin pathway, the target of a number of anticancer agents currently in clinical trial and use.