Suppression of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-mediated aryl hydrocarbon receptor transformation and CYP1A1 induction by the phosphatidylinositol 3-kinase inhibitor 2-(4-morpholinyl)-8-phenyl-4H-1- benzopyran-4-one (LY294002).
Numerous flavonoids are ligands of the aryl hydrocarbon receptor (AHR) and function as AHR antagonists and/or agonists. LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one] is a widely used inhibitor of phosphatidylinositol 3-kinase (PI 3-kinase), and is structurally related to members of the flavonoid family. Concentrations of LY294002 >/= 10 microM were cytostatic, but not cytotoxic, to cultures of the immortalized human breast epithelial cell line MCF10A-Neo. Treatment of MCF10A-Neo cultures with the AHR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) stimulated the transcriptional activation of CYP1A1, as monitored by measurements of steady-state CYP1A1 mRNA. Pretreatment of cultures with >/= 10 microM LY294002 suppressed the TCDD activation of CYP1A1 (IC(50) approximately 10 microM). Electrophoretic mobility shift assays employing rat liver cytosol demonstrated that concentrations of LY294002 </= 400 microM did not transform the AHR into a DNA-binding species. However, the addition of LY294002 to cytosol just prior to TCDD addition completely suppressed AHR transformation by TCDD (IC(50) approximately 35 microM). The PI 3-kinase inhibitor Wortmannin was weakly cytostatic, but not cytotoxic to MCF10A-Neo cultures at concentrations </= 500 nM. Exposure of cultures to Wortmannin (10-500 nM) did not suppress TCDD activation of CYP1A1. Analyses of the phosphorylation status of Akt-1, an in vivo substrate of PI 3-kinase, demonstrated that concentrations of LY294002 >/= 50 microM and Wortmannin >/= 10 nM completely suppressed PI 3-kinase activity. Hence, the ability of LY294002 to suppress TCDD-dependent activation of CYP1A1 is unrelated to PI 3-kinase inhibition. Instead, this activity reflects LY294002 functioning as an AHR antagonist. Furthermore, most of the cytostatic activity of LY294002 towards MCF10A-Neo cells is unrelated to the inhibition of PI 3-kinase.
Institute of Chemical Toxicology, Wayne State University, Detroit, MI 48201, USA.
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Reiners
D000730:Androstadienes / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002461:Cell Line, Transformed; D002867:Chromones / Q000494:pharmacology*; D019363:Cytochrome P-450 CYP1A1 / Q000096:biosynthesis* / Q000235:genetics; D004247:DNA / Q000187:drug effects / Q000378:metabolism; D004347:Drug Interactions; D004790:Enzyme Induction; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D009025:Morpholines / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology*; D018336:Receptors, Aryl Hydrocarbon / Q000037:antagonists & inhibitors* / Q000502:physiology; D015533:Transcriptional Activation / Q000187:drug effects; D000077191:Wortmannin
D000730:Androstadienes; D002867:Chromones; D019363:Cytochrome P-450 CYP1A1; D004247:DNA; D004791:Enzyme Inhibitors; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D000077191:Wortmannin; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one