Activation of the Aryl Hydrocarbon Receptor by the Calcium/Calmodulin-Dependent Protein Kinase Kinase Inhibitor 7-Oxo-7<i>H</i>-benzimidazo[2,1-<i>a</i>]benz[de]isoquinoline-3-carboxylic Acid (STO-609)

Monteiro, Patricia; Gilot, David; Langouët, Sophie; Fardel, Olivier

doi:10.1124/dmd.108.023333

Cited by 29 publications

(27 citation statements)

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“…In addition, to compare the effects of ITPKA and PNCK knock-down on EROD activity to our previous published data with CaMKIα [13], [16], [19], we also used 2 other well-characterized siRNA sequence targeting this kinase [16], [19] (siCaMKIa-1 and a-2, respectively, Fig. 4A).…”

Section: Resultsmentioning

confidence: 99%

“…These studies are recurrently based on chemical inhibition of these signaling mediators [10], although chemical inhibitors are rarely specific of only one family of kinases [12], [13]. Moreover, we and others have demonstrated that caution may be required when using chemical inhibitors to study AhR signaling since many of them are agonist/antagonist of AhR and/or inhibitor of CYP1A1 activity [14], [15], [16]. Ideally, a combination of chemical and genetic (knock-down or knock-out) inhibitions should be used to demonstrate the role of a candidate.…”

Section: Introductionmentioning

confidence: 99%

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RNAi-Based Screening Identifies Kinases Interfering with Dioxin-Mediated Up-Regulation of CYP1A1 Activity

et al. 2011

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BackgroundThe aryl hydrocarbon receptor (AhR) is a transcription factor activated by several environmental pollutants, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and involved in carcinogenesis and various physiological processes, including immune response and endocrine functions. Characterization of kinases-related AhR transduction pathway remains an important purpose.ResultsWe performed a kinome-wide siRNA screen in human mammary MCF-7 cells to identify non redundant protein kinases implicated in the up-regulation of cytochrome P-450 (CYP) 1A1 activity, an AhR referent target, in response to TCDD exposure. To this aim, we monitored CYP1A1-related ethoxyresorufin-O-deethylase (EROD) activity and quantified cell density. This normalization was crucial since it allowed us to focus only on siRNA affecting EROD activity and discard siRNA affecting cell density. Analyses of the cell density data allowed us to identify several hits already well-characterized as effectors of the cell cycle and original hits. Collectively, these data fully validated the protocol and the siRNA library. Next, 22 novel candidates were identified as kinases potentially implicated in the up-regulation of CYP1A1 in response to TCDD, without alteration of cell survival or cell proliferation. The siRNA library screen gave a limited number of hits (approximately 3%). Interestingly, four of them are able to bind calmodulin among which the IP3 kinase A (ITPKA) and pregnancy up-regulated non-ubiquitously expressed CaM kinase (PNCK, also named CaMKIβ). Remarkably, for both proteins, their kinase activity depends on the calmodulin binding. Involvement of ITPKA and PNCK in TCDD-mediated CYP1A1 up-regulation was further validated by screening-independent expression knock-down. PNCK was finally shown to regulate activation of CaMKIα, a CaMKI isoform previously reported to interplay with the AhR pathway.ConclusionsThese data fully support a role for both IP3-related kinase and CaMK isoforms in the AhR signaling cascade. More generally, this study also highlights the interest of large scale loss-of-function screens for characterizing the molecular mechanism of action of environmental contaminants.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

RNAi-Based Screening Identifies Kinases Interfering with Dioxin-Mediated Up-Regulation of CYP1A1 Activity

et al. 2011

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“…We found that STO-609 was a weak AhR agonist with an IC50 of 45 µM and thus we did not pursue its ability to induce Tregs. However, in previous studies designed to determine the role of Ca 2+ in AhR signaling, STO-609 was discovered to be a weak, albeit full AhR agonist in MCF-7 cells and in human macrophages [29]. Interestingly, AhR activation occurred at the same concentration (25 µM) that was known to fully inhibit CaMKK [26], opening up the possibility that some of the effects attributed to CaMKK inhibition could be due to changes in expression of AhR-regulated genes.…”

Section: Discussionmentioning

confidence: 99%

Benzimidazoisoquinolines: A New Class of Rapidly Metabolized Aryl Hydrocarbon Receptor (AhR) Ligands that Induce AhR-Dependent Tregs and Prevent Murine Graft-Versus-Host Disease

et al. 2014

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The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that plays multiple roles in regulation of immune and inflammatory responses. The ability of certain AhR ligands to induce regulatory T cells (Tregs) has generated interest in developing AhR ligands for therapeutic treatment of immune-mediated diseases. To this end, we designed a screen for novel Treg-inducing compounds based on our understanding of the mechanisms of Treg induction by the well-characterized immunosuppressive AhR ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). We screened a ChemBridge small molecule library and identified 10-chloro-7H-benzimidazo[2,1-a]benzo[de]Iso-quinolin-7-one (10-Cl-BBQ) as a potent AhR ligand that was rapidly metabolized and not cytotoxic to proliferating T cells. Like TCDD,10-Cl-BBQ altered donor CD4+ T cell differentiation during the early stages of a graft versus host (GVH) response resulting in expression of high levels of CD25, CTLA-4 and ICOS, as well as several genes associated with Treg function. The Treg phenotype required AhR expression in the donor CD4+ T cells. Foxp3 was not expressed in the AhR-induced Tregs implicating AhR as an independent transcription factor for Treg induction. Structure-activity studies showed that unsubstituted BBQ as well as 4, 11-dichloro-BBQ were capable of inducing AhR-Tregs. Other substitutions reduced activation of AhR. Daily treatment with 10-Cl-BBQ during the GVH response prevented development of GVH disease in an AhR-dependent manner with no overt toxicity. Together, our data provide strong support for development of select BBQs that activate the AhR to induce Tregs for treatment of immune-mediated diseases.

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“…Specific binding of BaP to the AHR has been measured in vitro directly using radiolabeled substrate (Monteiro et al, 2008), indirectly utilizing luciferase reporters containing the AHR response element (Malik et al, 2013) and by monitoring the expression of a known AHR target gene, Cyp1a1 (Ovesen et al, 2011; Table 5). Activation of the AHR by BaP leads to its own metabolism (Section 2.3.3) creating reactive metabolites that produce its subsequent genotoxic MOA.…”

Section: Genomics Approaches (Ra2 and Ra3)mentioning

confidence: 99%

Comparison of toxicogenomics and traditional approaches to inform mode of action and points of departure in human health risk assessment of benzo[a]pyrene in drinking water

Moffat

Chepelev

Labib

et al. 2015

Critical Reviews in Toxicology

138

100

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Toxicogenomics is proposed to be a useful tool in human health risk assessment. However, a systematic comparison of traditional risk assessment approaches with those applying toxicogenomics has never been done. We conducted a case study to evaluate the utility of toxicogenomics in the risk assessment of benzo[a]pyrene (BaP), a well-studied carcinogen, for drinking water exposures. Our study was intended to compare methodologies, not to evaluate drinking water safety. We compared traditional (RA1), genomics-informed (RA2) and genomics-only (RA3) approaches. RA2 and RA3 applied toxicogenomics data from human cell cultures and mice exposed to BaP to determine if these data could provide insight into BaP's mode of action (MOA) and derive tissue-specific points of departure (POD). Our global gene expression analysis supported that BaP is genotoxic in mice and allowed the development of a detailed MOA. Toxicogenomics analysis in human lymphoblastoid TK6 cells demonstrated a high degree of consistency in perturbed pathways with animal tissues. Quantitatively, the PODs for traditional and transcriptional approaches were similar (liver 1.2 vs. 1.0 mg/kg-bw/day; lung 0.8 vs. 3.7 mg/kg-bw/day; forestomach 0.5 vs. 7.4 mg/kg-bw/day). RA3, which applied toxicogenomics in the absence of apical toxicology data, demonstrates that this approach provides useful information in data-poor situations. Overall, our study supports the use of toxicogenomics as a relatively fast and cost-effective tool for hazard identification, preliminary evaluation of potential carcinogens, and carcinogenic potency, in addition to identifying current limitations and practical questions for future work.

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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)

Abstract: ABSTRACT:This study was designed to analyze the effects of the Ca

Cited by 29 publications

References 38 publications

RNAi-Based Screening Identifies Kinases Interfering with Dioxin-Mediated Up-Regulation of CYP1A1 Activity

RNAi-Based Screening Identifies Kinases Interfering with Dioxin-Mediated Up-Regulation of CYP1A1 Activity

Benzimidazoisoquinolines: A New Class of Rapidly Metabolized Aryl Hydrocarbon Receptor (AhR) Ligands that Induce AhR-Dependent Tregs and Prevent Murine Graft-Versus-Host Disease

Comparison of toxicogenomics and traditional approaches to inform mode of action and points of departure in human health risk assessment of benzo[a]pyrene in drinking water

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