Activation of the Aryl Hydrocarbon Receptor by the Calcium/Calmodulin-Dependent Protein Kinase Kinase Inhibitor 7-Oxo-7H-benzimidazo[2,1-a]benz[de]isoquinoline-3-carboxylic Acid (STO-609) (original) (raw)
Research ArticleArticle
Drug Metabolism and Disposition December 2008, 36 (12) 2556-2563; DOI: https://doi.org/10.1124/dmd.108.023333
Abstract
This study was designed to analyze the effects of the Ca2+/calmodulin-dependent protein kinase kinase (CaMKK) inhibitor STO-609 (7-oxo-7_H_-benzimidazo[2,1-_a_]benz[de]isoquinoline-3-carboxylic acid) toward the aryl hydrocarbon receptor (AhR) pathway because Ca2+/calmodulin-dependent protein kinase (CaMK) Iα, known as a downstream CaMKK effector, has been recently shown to contribute to the AhR cascade. STO-609 failed to alter up-regulation of the AhR target CYP1A1 in response to the potent AhR ligand 2,3,7,8-tetrachlorodibenzo-_p_-dioxin (TCDD) in MCF-7 cells. STO-609, used at a 25 μM concentration known to fully inhibit CaMKK activity, was surprisingly found to markedly induce CYP1A1 expression and activity by itself in MCF-7 cells; it similarly up-regulated various other AhR target genes in human macrophages. STO-609-related CYP1A1 induction was prevented by chemical inhibition or small interfering RNA-mediated knockdown expression of AhR. Moreover, STO-609 was demonstrated to physically interact with the ligand-binding domain of AhR, as assessed by TCDD binding competition assay, and to induce AhR translocation to the nucleus. As already reported for AhR agonists, STO-609 triggered the increase of [Ca2+]i and activation of CaMKIα, whose inhibition through the use of the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,_N_′,_N_′-tetraacetic acid-acetoxymethyl ester or the CaMK inhibitor KN-93 (2-[_N_-(2-hydroxyethyl)]-_N_-(4-methoxybenzenesulfonyl)]amino-_N_-(4-chlorocinnamyl)-_N_-methylbenzylamine), respectively, prevented STO-609-mediated CYP1A1 activity induction. Taken together, these results demonstrate that the CaMKK inhibitor STO-609 can act as an AhR ligand and, in this way, fully activates the Ca2+/CaMKIα/AhR cascade. Such data, therefore, make unlikely any contribution of CaMKK activity to the AhR pathway and, moreover, suggest that caution may be required when using STO-609 as a specific inhibitor of CaMKKs.
Footnotes
This work was supported by grants from the Ligue Nationale contre le Cancer and from the Agence Francaise de Sécurité Sanitaire de l'Environnement et du Travail. P.M. received a fellowship from the Ligue Nationale contre le Cancer.
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
doi:10.1124/dmd.108.023333.
ABBREVIATIONS: AhR, arylhydrocarbon receptor; TCDD, 2,3,7,8-tetrachlorodibenzo-_p_-dioxin; PAH, polycyclic aromatic hydrocarbon; XRE, xenobiotic-responsive element; MAPK, mitogen-activated protein kinase; CaMK, Ca2+/calmodulin-dependent protein kinase; CaMKK, Ca2+/calmodulin-dependent protein kinase kinase; STO-609, 7-oxo-7_H_-benzimidazo[2,1-_a_]benz[de]isoquinoline-3-carboxylic acid; U0126, 1,4-diamino-2,3-dicyano-1,4-bis(2-aminophynyltio)butadiene; PD98059, 2′-amino-3′-methoxyflavone; 2-APB, 2-aminoethoxydiphenylborate; αNF, α-naphthoflavone; BAPTA, 1,2-bis(2-aminophenoxy)ethane-N,N,_N_′,_N_′-tetraacetic acid; AM, acetoxymethyl ester; KN-93, 2-[_N_-(2-hydroxyethyl)]-_N_-(4-methoxybenzenesulfonyl)]amino-_N_-(4-chlorocinnamyl)-_N_-methylbenzylamine; W7, _N_-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide; Ab, antibody; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; AMPK, AMP-activated protein kinase; FITC, fluorescein isothiocyanate; DMSO, dimethyl sulfoxide; EROD, ethoxyresorufin _O_-deethylase; RT, reverse transcription; qPCR, real-time quantitative polymerase chain reaction; IL, interleukin; ITG, integrin; siRNA, small interfering RNA; iAhR, small interfering RNA targeting AhR; iNT1, nontargeting siRNA; a.u., arbitrary unit(s); SP600125, 1,9-pyrazoloanthrone.
- Received July 11, 2008.
- Accepted August 26, 2008.
The American Society for Pharmacology and Experimental Therapeutics
Log in using your username and password
Purchase access
You may purchase access to this article. This will require you to create an account if you don't already have one.