The Glycogen Synthase Kinase Inhibitor 3-(2,4-Dichlorophenyl)-4-(1-methyl-1H-indol-3-yl)-1H-pyrrole-2,5-dione (SB216763) Is a Partial Agonist of the Aryl Hydrocarbon Receptor

Abstract

ABSTRACT:Kinase inhibitors are frequently used tools in signal transduction research. 3-(2,4-Dichlorophenyl)-4-(1-methyl-1H-indol-3-yl)-1H-pyrrole-2,5-dione (SB216763), a potent inhibitor of glycogen synthase kinase 3␤ (GSK3␤), is frequently used to activate ␤-catenin signaling by mimicking the action of Wnt molecules. ␤-Catenin is a crucial player in the regulation of hepatic drug metabolism. Thus, it is of particular importance to know whether the tools used to study the effects of ␤-catenin signaling may affect the respective drugmetabolizing target enzymes in an unwanted manner. In this study, we show that SB216763 is able to induce cytochrome P450 1a1 (Cyp1a1) expression in a dose-dependent manner in mouse hepatoma cells. Moreover, SB216763 is able to inhibit Cyp1a1 induction by the prototype aryl hydrocarbon receptor (AhR) ligand 2,3,7,8-tetrachloro-p-dibenzodioxin. Cyp1a1 induction by SB216763 is independent of GSK3␤ and the ␤-catenin pathway. Instead, SB216763 induces Cyp1a1 by activation of AhR-mediated transcription. The present results suggest that SB216763 acts as a partial agonist of the AhR.A variety of cellular signal transduction pathways are regulated by phosphorylation/dephosphorylation events, and a huge number of small molecule kinase inhibitors has been developed allowing for the blockade or activation of distinct signaling cascades. Besides the fact that many kinase inhibitors-often acting in an ATP-competitive manner-do not provide satisfactory specificity toward only one single kinase (Davies et al., 2000;Bain et al., 2007), there are also several reports which demonstrate that some of these molecules can act through additional mechanisms that are different from kinase inhibition. An example of this difference is the interaction with nuclear receptors such as the xenobiotic sensor aryl hydrocarbon receptor (AhR), which is a crucial player in the expression of various drug-metabolizing enzymes. In the absence of a ligand, the AhR is complexed by heat shock proteins in the cytosol. Upon binding of a ligand, mostly a planar hydrophobic molecule, the AhR translocates to the nucleus, releases the chaperones, and dimerizes with the Arnt protein. The AhR/Arnt heterodimer then binds to so-called dioxin response elements (DREs) on the DNA and activates transcription of target genes such as cytochrome P450 1a1 (Cyp1a1). For a recent review on AhR-dependent signal transduction, see Beischlag et al. (2008).The widely used kinase inhibitors U0126, PD98059, and SP600125 are ligands and partial agonists of the AhR (Reiners et al., 1998;Joiakim et al., 2003;Andrieux et al., 2004;Dvorak et al., 2008).Several other kinase inhibitors also affect the process of AhR activation by direct interaction with the AhR or by more complex indirect mechanisms, as reviewed in Braeuning (2009). Because most of the kinase inhibitors available from commercial suppliers have never been tested for a possible interplay with xenobiotic sensors, one might overlook such nonkinase-related properties and attribute the observed eff...

Interplay of β-Catenin with Xenobiotic-Sensing Receptors and its Role in Glutathione S-Transferase Expression

“…This finding was confirmed in murine cells by analyses of AhR mRNA expression in Ctnnb1-mutated mouse liver tumors [37], cultured mouse hepatocytes [15,54,78], and mice with hepatocyte-specific knockout of Ctnnb1 [21,54]; see also Table 5. Moreover, preferential expression of the AhR in perivenous hepatocytes [15,79,80], which exclusively possess activated -catenin in healthy liver [14,81], supports the idea of a stimulatory role of -catenin in the regulation of AhR mRNA expression.…”

“…Again similar to the AhR, PB-and TCPOBOP-induced levels of CAR target genes from phase I and phase II of drug metabolism (including the GST isoforms Gstm2, Gstm3, and Gstm6) were, for the most part, higher in wild type mice, as compared to their Ctnnb1 knockout counterparts (Table 3), indicating higher activity of CAR-dependent transcription in the presence of -catenin [21]. Of note, the induction of hepatocyte AhR mouse liver adenoma, Ctnnb1-mutated mRNA (cDNA microarray) [20,37] AhR mouse hepatocytes, Wnt3a mRNA (real-time RT-PCR) [15,54,78] AhR mouse hepatocytes, SB216763 mRNA (real-time RT-PCR) [15] AhR Ctnnb1 knockout mouse mRNA (real-time RT-PCR) [21,54] AhR mouse hepatocytes, SB216763 mRNA (real-time RT-PCR) [78] CAR Ctnnb1 knockout mouse mRNA (real-time RT-PCR) [21,36,88] CAR mouse liver adenoma, Ctnnb1-mutated mRNA (real-time RT-PCR) [36] CAR Ctnnb1 knockout mouse Protein (Western blotting) [88] PXR Apc knockout mouse mRNA (cDNA microarray) [14] PXR Ctnnb1 knockout mouse * mRNA (real-time RT-PCR) [21] PXR mouse hepatocytes, Wnt3a mRNA (real-time RT-PCR) [54] Arrows indicate up-or down-regulation of respective GST isoforms. *, in female mice only.…”

Cooperation of structurally different aryl hydrocarbon receptor agonists and β-catenin in the regulation of CYP1A expression

“…7C). The latter observation is of special importance, since there are numerous small molecule inhibitors, which act as partial agonists of the AhR, e.g., U0126 (Andrieux et al, 2004) and SB216763 (Braeuning and Buchmann, 2009). Thus, it was necessary to prove that FH535 neither induces basal Cyp1a1 expression nor interferes with AhR agonist-mediated effects.…”

Combination effects of AHR agonists and Wnt/β-catenin modulators in zebrafish embryos: Implications for physiological and toxicological AHR functions

“…11) indicates that XAV could be a partial AHR agonist. This has been suggested for the GSK-3 inhibitor SB216763 (Braeuning and Buchmann, 2009). Also by the CYP1A induction pattern it is suggested that XAV (and possibly also AZP) could be a partial agonist for the AHR.…”