Abstract
The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor and a member of the basic helix-loop-helix PER/ARNT/SIM family of chemosensors and developmental regulators. The AhR is widely known as a mediator of dioxin toxicity; however, it also suppresses cancer cell proliferation and recent findings have implicated its role as a tumor suppressor. We conducted a chemical library screen to identify nontoxic AhR ligands with anti-cancer effects and discovered flutamide (Eulexin) as a putative AhR ligand. Flutamide is an androgen receptor (AR) antagonist approved by the United States Food and Drug Administration for the treatment of prostate cancer. We found that flutamide inhibited the growth of several cancer cell lines independent of AR status, and that suppression of AhR expression reversed the anti-proliferative effects of flutamide. We investigated the AhR-dependent mechanism of action of flutamide in human hepatocellular carcinoma cells and identified that transforming growth factor-β1 (TGF-β1) is induced by flutamide in an AhR-dependent manner. In contrast, the potent AhR agonist 2,3,7,8-Tetrachlorodibenzo-p-dioxin had no effect on TGF-β1 expression, indicating the ligand specificity of AhR activation. We also determined that TGF-β1 induction is required for the AhR-dependent growth inhibitory effects of flutamide. Therefore, flutamide may be effective in AhR-positive cancers that are sensitive to TGF-β1 signaling, such as hepatocellular carcinoma.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 50 print issues and online access
$259.00 per year
only $5.18 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Burbach KM, Poland A, Bradfield CA . Cloning of the Ah-receptor cDNA reveals a distinctive ligand-activated transcription factor. Proc Natl Acad Sci USA 1992; 89: 8185–8189.
Denison MS, Fisher JM, Whitlock JP . Inducible, receptor-dependent protein-DNA interactions at a dioxin-responsive transcriptional enhancer. Proc Natl Acad Sci USA 1988; 85: 2528–2532.
Perdew GH, Bradfield CA . Mapping the 90 kDa heat shock protein binding region of the Ah receptor. Biochem Mol Biol Int 1996; 39: 589–593.
Felts SJ, Toft DO . P23, a simple protein with complex activities. Cell Stress Chaperones 2003; 8: 108–113.
Perdew GH . Association of the Ah receptor with the 90-kda heat-shock protein. J Biol Chem 1988; 263: 13802–13805.
Carver LA, Bradfield CA . Ligand-dependent interaction of the aryl hydrocarbon receptor with a novel immunophilin homolog in vivo. J Biol Chem 1997; 272: 11452–11456.
Meyer BK, Pray-Grant MG, Vanden Heuvel JP, Perdew GH . Hepatitis B virus X-associated protein 2 is a subunit of the unliganded aryl hydrocarbon receptor core complex and exhibits transcriptional enhancer activity. Mol Cell Biol 1998; 18: 978–988.
Beischlag TV, Morales JL, Hollingshead BD, Perdew GH . The aryl hydrocarbon receptor complex and the control of gene expression. Crit Rev Eukaryot Gene Expr. 2008; 18: 207–250.
Hu W, Sorrentino C, Denison MS, Kolaja K, Fielden MR . Induction of cyp1a1 is a nonspecific biomarker of aryl hydrocarbon receptor activation: results of large scale screening of pharmaceuticals and toxicants in vivo and in vitro. Mol Pharmacol 2007; 71: 1475–1486.
Puga A, Ma C, Marlowe JL . The aryl hydrocarbon receptor cross-talks with multiple signal transduction pathways. Biochem Pharmacol 2009; 77: 713–722.
Safe S . Molecular biology of the Ah receptor and its role in carcinogenesis. Toxicol Lett 2001; 120: 1–7.
Bertazzi PA, Zocchetti C, Guercilena S, Consonni D, Tironi A, Landi MT et al. Dioxin exposure and cancer risk: a 15-year mortality study after the ''Seveso accident''. Epidemiology 1997; 8: 646–652.
Warner M, Eskenazi B, Mocarelli P, Gerthoux P, Samuels S, Needham L et al. Serum dioxin concentrations and breast cancer risk in the Seveso Women’s Health Study. Am J Epidemiol 2002; 155: s102–s102.
Hsu EL, Yoon D, Choi HH, Wang F, Taylor RT, Chen N et al. A proposed mechanism for the protective effect of dioxin against breast cancer. Toxicol Sci 2007; 98: 436–444.
Funatake CJ, Marshall NB, Kerkvliet NI . 2,3,7,8-tetrachlorodibenzo-p-dioxin afters the differentiation of alloreactive CD8(+) T cells toward a regulatory T cell phenotype by a mechanism that is dependent on aryl hydrocarbon receptor in CD4(+) T cells. J Immunotoxicol 2008; 5: 81–91.
Fernandez-Salguero P, Pineau T, Hilbert DM, McPhail T, Lee SS, Kimura S et al. Immune system impairment and hepatic fibrosis in mice lacking the dioxin-binding Ah receptor. Science 1995; 268: 722–726.
Bock KW, Kohle C . Ah receptor: dioxin-mediated toxic responses as hints to deregulated physiologic functions. Biochem Pharmacol 2006; 72: 393–404.
Lai IK, Dhakal K, Gadupudi GS, Li M, Ludewig G, Robertson LW et al. N-acetylcysteine (NAC) diminishes the severity of PCB 126-induced fatty liver in male rodents. Toxicology 2012; 302: 25–33.
Egeland GM, Sweeney MH, Fingerhut MA, Wille KK, Schnorr TM, Halperin WE . Total serum testosterone and gonadotropins in workers exposed to dioxin. Am J Epidemiol 1994; 139: 272–281.
Rier SE, Martin DC, Bowman RE, Dmowski WP, Becker JL . Endometriosis in rhesus-monkeys (Macaca-Mulatta) following chronic exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Fund Appl Toxicol 1993; 21: 433–441.
Abbott BD, Perdew GH, Birnbaum LS . Ah receptor in embryonic mouse palate and effects of TCDD on receptor expression. Toxicol Appl Pharm 1994; 126: 16–25.
Schmidt JV, Su GHT, Reddy JK, Simon MC, Bradfield CA . Characterization of a murine Ahr null allele: involvement of the Ah receptor in hepatic growth and development. Proc Natl Acad Sci USA 1996; 93: 6731–6736.
Fernandez-Salguero PM, Hilbert DM, Rudikoff S, Ward JM, Gonzalez FJ . Aryl-hydrocarbon receptor-deficient mice are resistant to 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced toxicity. Toxicol Appl Pharm 1996; 140: 173–179.
Gasiewicz TA, Henry EC, Collins LL . Expression and activity of aryl hydrocarbon receptors in development and cancer. Crit Rev Eukaryot Gene Expr 2008; 18: 279–321.
Mitsuhashi T, Yonemoto J, Sone H, Kosuge Y, Kosaki K, Takahashi T . In utero exposure to dioxin causes neocortical dysgenesis through the actions of p27(Kip1). Proc Natl Acad Sci USA 2010; 107: 16331–16335.
Weiss C, Kolluri SK, Kiefer F, Gottlicher M . Complementation of Ah receptor deficiency in hepatoma cells: negative feedback regulation and cell cycle control by the Ah receptor. Exp Cell Res 1996; 226: 154–163.
Puga A, Xia Y, Elferink C . Role of the aryl hydrocarbon receptor in cell cycle regulation. Chem Biol Interact 2002; 141: 117–130.
Fritz WA, Lin TM, Cardiff RD, Peterson RE . The aryl hydrocarbon receptor inhibits prostate carcinogenesis in TRAMP mice. Carcinogenesis 2007; 28: 497–505.
Fan YX, Boivin GP, Knudsen ES, Nebert DW, Xia Y, Puga A . The aryl hydrocarbon receptor functions as a tumor suppressor of liver carcinogenesis. Cancer Res 2010; 70: 212–220.
Barretina J, Caponigro G, Stransky N, Venkatesan K, Margolin AA, Kim S et al. The Cancer Cell Line Encyclopedia enables predictive modelling of anticancer drug sensitivity. Nature 2012; 483: 603–607.
Hall JM, Barhoover MA, Kazmin D, McDonnell DP, Greenlee WF, Thomas RS . Activation of the aryl-hydrocarbon receptor inhibits invasive and metastatic features of human breast cancer cells and promotes breast cancer cell differentiation. Mol Endocrinol 2010; 24: 359–369.
Kolluri SK, Weiss C, Koff A, Gottlicher M . p27(Kip1) induction and inhibition of proliferation by the intracellular Ah receptor in developing thymus and hepatoma cells. Gene Dev 1999; 13: 1742–1753.
Ge NL, Elferink CJ . A direct interaction between the aryl hydrocarbon receptor and retinoblastoma protein—Linking dioxin signaling to the cell cycle. J Biol Chem 1998; 273: 22708–22713.
Kharat I, Saatcioglu F . Antiestrogenic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin are mediated by direct transcriptional interference with the liganded estrogen receptor. Cross-talk between aryl hydrocarbon- and estrogen-mediated signaling. J Biol Chem 1996; 271: 10533–10537.
O’Donnell EF, Saili KS, Koch DC, Kopparapu PR, Farrer D, Bisson WH et al. The anti-inflammatory drug leflunomide is an agonist of the aryl hydrocarbon receptor. PLos One 2010; 5: e13128.
Raghow S, Kuliyev E, Steakley M, Greenberg N, Steiner MS . Efficacious chemoprevention of primary prostate cancer by flutamide in an autochthonous transgenic model. Cancer Res 2000; 60: 4093–4097.
Bisson WH, Koch DC, O’Donnell EF, Khalil SM, Kerkvliet NI, Tanguay RL et al. Modeling of the aryl hydrocarbon receptor (AhR) ligand binding domain and its utility in virtual ligand screening to predict new AhR ligands. J Med Chem 2009; 52: 5635–5641.
O’Donnell EF, Kopparapu PR, Koch DC, Jang HS, Phillips JL, Tanguay RL et al. The aryl hydrocarbon receptor mediates leflunomide-induced growth inhibition of melanoma cells. PLos One 2012; 7: e40926.
Robson CN, Gnanapragasam V, Byrne RL, Collins AT, Neal DE . Transforming growth factor-beta 1 up-regulates p15, p21 and p27 and blocks cell cycling in G1 in human prostate epithelium. J Endocrinol 1999; 160: 257–266.
Mullen AC, Orlando DA, Newman JJ, LovÈn J, Kumar RM, Bilodeau S et al. Master transcription factors determine cell-type-specific responses to TGF-β signaling. Cell 2011; 147: 565–576.
Kolluri SK, Balduf C, Hofmann M, Gottlicher M . Novel target genes of the Ah (dioxin) receptor: transcriptional induction of N-myristoyltransferase 2. Cancer Res 2001; 61: 8534–8539.
Punj S, Kopparapu P, Jang HS, Phillips JL, Pennington J, Rohlman D et al. Benzimidazoisoquinolines: a new class of rapidly metabolized aryl hydrocarbon receptor (AhR) ligands that induce AhR-dependent Tregs and prevent murine graft-versus-host disease. PLoS One 2014; 9: e88726.
O’Donnell EF, Koch DC, Bisson WH, Jang HS, Kolluri SK . The aryl hydrocarbon receptor mediates raloxifene-induced apoptosis in estrogen receptor-negative hepatoma and breast cancer cells. Cell Death Dis 2014; 5: e1038.
Hannon GJ, Beach D . pl5INK4B is a potentia| effector of TGF-[beta]-induced cell cycle arrest. Nature 1994; 371: 257–261.
Massague J, Blain SW, Lo RS . TGF beta signaling in growth control, cancer, and heritable disorders. Cell 2000; 103: 295–309.
Reynisdottir I, Polyak K, Iavarone A, Massague J . Kip/Cip and Ink4 Cdk inhibitors cooperate to induce cell-cycle arrest in response to TGF-beta. Gene Dev 1995; 9: 1831–1845.
Fanayan S, Firth SM, Baxter RC . Signaling through the Smad pathway by insulin-like growth factor-binding protein-3 in breast cancer cells—relationship to transforming growth factor-beta 1 signaling. J Biol Chem 2002; 277: 7255–7261.
Fan Y, Boivin GP, Knudsen ES, Nebert DW, Xia Y, Puga A . The aryl hydrocarbon receptor functions as a tumor suppressor of liver carcinogenesis. Cancer Res 2010; 70: 212–220.
Gomez-Duran A, Carvajal-Gonzalez JM, Mulero-Navarro S, Santiago-Josefat B, Puga A, Fernandez-Salguero PM . Fitting a xenobiotic receptor into cell homeostasis: how the dioxin receptor interacts with TGF beta signaling. Biochem Pharmacol 2009; 77: 700–712.
Chang XQ, Fan YX, Karyala S, Schwemberger S, Tomlinson CR, Sartor MA et al. Ligand-independent regulation of transforming growth factor beta 1 expression and cell cycle progression by the aryl hydrocarbon receptor. Mol Cell Biol 2007; 27: 6127–6139.
Elizondo G, Fernandez-Salguero P, Sheikh MS, Kim GY, Fornace AJ, Lee KS et al. Altered cell cycle control at the G(2)/M phases in aryl hydrocarbon receptor-null embryo fibroblast. Mol Pharmacol 2000; 57: 1056–1063.
Carvajal-Gonzalez JM, Roman AC, Cerezo-Guisado MI, Rico-Leo EM, Martin-Partido G, Fernandez-Salguero PM . Loss of dioxin-receptor expression accelerates wound healing in vivo by a mechanism involving TGF beta. J Cell Sci 2009; 122: 1823–1833.
Dohr O, Abel J . Transforming growth factor-beta(1) coregulates mRNA expression of aryl hydrocarbon receptor and cell-cycle-regulating genes in human cancer cell lines. Biochem Biophys Res Commun 1997; 241: 86–91.
Perkins A, Phillips J, Kerkvliet N, Tanguay R, Perdew G, Kolluri S et al. A structural switch between agonist and antagonist bound conformations for a ligand-optimized model of the human aryl hydrocarbon receptor ligand binding domain. Biology 2014; 3: 645–669.
Murray IA, Patterson AD, Perdew GH . Aryl hydrocarbon receptor ligands in cancer: friend and foe. Nat Rev Cancer 2014; 14: 801–814.
Safe S, Lee SO, Jin UH . Role of the aryl hydrocarbon receptor in carcinogenesis and potential as a drug target. Toxicol Sci 2013; 135: 1–16.
Zhang S, Kim K, Jin UH, Pfent C, Cao HJ, Amendt B et al. Aryl hydrocarbon receptor agonists induce microRNA-335 expression and inhibit lung metastasis of estrogen receptor negative breast cancer cells. Mol Cancer Ther 2012; 11: 108–118.
Jin UH, Lee SO, Safe S . Aryl hydrocarbon receptor (AhR)-active pharmaceuticals are selective AhR modulators in MDA-MB-468 and BT474 breast cancer cells. J Pharmacol Exp Ther 2012; 343: 333–341.
Acknowledgements
This work was supported by American Cancer Society (RSG-13-132-01-CDD), the National Institute of Environmental Health Sciences (NIEHS) grant numbers ES016651 and ES019000, The US Army Medical Research and Material Command and The Cell Imaging and Analysis Facility Cores of the Environmental Health Sciences Center, NIEHS grant number P30 ES00210. DCK was supported by a National Research Service Award (1F31CA144571-01) pre-doctoral fellowship from the National Cancer Institute. DCK is currently supported by the Burroughs Wellcome Fund post-doctoral enrichment program. EFO was supported by a pre-doctoral fellowship from the Department of Defense Breast Cancer Research Program (W81XWH-10-1-0160). DCK and EFO were previously supported by NIEHS training grant (T32ES07060). The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript. We thank Samuel Bradford for excellent flow cytometry assistance, Sammy Khalil, Cathy Duong and Viktor Dikov for laboratory assistance.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no conflict of interest.
Additional information
Supplementary Information accompanies this paper on the Oncogene website
Supplementary information
Rights and permissions
About this article
Cite this article
Koch, D., Jang, H., O'Donnell, E. et al. Anti-androgen flutamide suppresses hepatocellular carcinoma cell proliferation via the aryl hydrocarbon receptor mediated induction of transforming growth factor-β1. Oncogene 34, 6092–6104 (2015). https://doi.org/10.1038/onc.2015.55
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/onc.2015.55
This article is cited by
-
Role of miR-653 and miR-29c in downregulation of CYP1A2 expression in hepatocellular carcinoma
Pharmacological Reports (2022)
-
Discovery and Mechanistic Characterization of a Select Modulator of AhR-regulated Transcription (SMAhRT) with Anti-cancer Effects
Apoptosis (2021)
-
Design and synthesis of anticancer 1-hydroxynaphthalene-2-carboxanilides with a p53 independent mechanism of action
Scientific Reports (2019)
-
Role of the aryl hydrocarbon receptor in carcinogenesis and potential as an anti-cancer drug target
Archives of Toxicology (2017)