N/A, Hepatotoxicity, Hepatopathy, including a constellation of observable effects

1746-01-6

HGUFODBRKLSHSI-UHFFFAOYSA-N

2,3,7,8-Tetrachlorodibenzo-p-dioxin

Dioxin Dibenzo[b,e][1,4]dioxin, 2,3,7,8-tetrachloro- 2,3,7,8-TCDD 2,3,7,8-Tetrachlordibenzo[b,e][1,4]dioxin 2,3,7,8-Tetrachloro-1,4-dioxin 2,3,7,8-tetrachlorodibenzo[b,e][1,4]dioxin 2,3,7,8-Tetrachlorodibenzo-1,4-dioxin 2,3,7,8-Tetrachlorodibenzodioxin 2,3,7,8-Tetrachloro-p-dioxin Dibenzo-p-dioxin, 2,3,7,8-tetrachloro- p-Dioxin

DTXSID2021315

60-35-5

DLFVBJFMPXGRIB-UHFFFAOYSA-N

Acetamide

Acetamid acetamida Acetic acid amide Acetimidic acid Ethanamide Ethanimidic acid Methanecarboxamide NSC 25945

DTXSID7020005

D008107

MESH Liver Diseases

PR:000003858

PR aryl hydrocarbon receptor

FMA:63179

FMA Liver cell

CL:0002196

CL hepatic oval stem cell

CL:0002538

CL intrahepatic bile duct epithelial cell

D006528

MESH hepatocellular carcinoma

D001650

MESH Bile Duct Neoplasms

GO:0023052

GO signaling

GO:0019725

GO cellular homeostasis

GO:0006915

GO apoptotic process

GO:0008283

GO cell proliferation

D006965

MESH hyperplasia

WIKI occurrence

WIKI increased

WIKI abnormal

WIKI decreased

Dioxin and dioxin-like compounds The promiscuous nature of the AHR with its ability to bind to a large number of endogenous and exogenous ligands, make the task of constructing an AOP a challenge. For example, many structurally diverse chemicals can bind to and activate the AHR, including dioxin-like chemicals (DLCs), polyaromatic hydrocarbons (PAHs), indole-3-carbinol as found in broccoli and Brussels sprouts, natural flavonoids such as quercetin, galangin and genistein, and endogenous ligands such as indirubicin, equilenin, metabolites of arachidonic acid, heme, tryptophan, and UV photoproducts of tryptophan such as 6-formylindolo[3,2-b]carbazole (FICZ) <a href="#cite_note-Denison_2011-1">[1]</a> <a href="#cite_note-Denison_2003-2">[2]</a> <a href="#cite_note-Gasiewicz_2008-3">[3]</a> It is believed, although not proven, that the endogenous and natural ligands ingested in the diet participate in normal homeostatic control of development and physiology through episodic and short-lived AHR activation. Developmental pathologies in AHR knockout rodents and toxicity associated with sustained AHR activation provide evidence that the AHR has a normal and necessary role. <a href="#cite_note-Denison_2011-1">[1]</a> <a href="#cite_note-Denison_2003-2">[2]</a> Weak agonists also include benzimidazoles such as omeprazole used pharmaceutically as a proton pump inhibitor and antihelminthics such as thiabendazole. Other weak agonists include primaquine, vinclozolin, YH439, phenylthiourea, curcumin and Oltipraz. <a href="#cite_note-Nguyen_2008-4">[4]</a> Therefore, a necessary task for this AOP is to differentiate between AHR ligands that act as rodent liver tumour promoters and those that do not. It may be that ligands resistant to metabolic clearance (e.g., TCDD), or sufficiently high doses of rapidly cleared ligands are able to create the sustained AHR activation required to bring about tumour promotion. Chemical properties for a selection of AHR ligands are provided in Table 3 along with EC50, which itself is a measure of potency and one of the determinants of sustained AHR activation. These representative ligands demonstrate the diversity in chemical properties modelling pharmacokinetics and bioavailability as well as sources, including anthropogenic chemicals, dietary constituents, and endogenous substances formed in vivo. The determinants of sustained AHR activation include: <ul> <li>AHR binding potency, usually measured by an EC50 or ED50 value; </li> <li>AHR binding efficacy or intrinsic activity measured by the maximal response; </li> <li>pharmacokinetic determinants including, <ul> <li>Speed and extent of metabolism/elimination of a particular ligand; </li> <li>Delivery to the target tissue; and, </li> </ul> </li> <li>Magnitude and duration of exposure. </li> </ul> Not all AHR ligands will produce sustained activation. Some ligands may stabilize the AHR, thus keeping it activated for longer times. <a href="#cite_note-Bohonowych_2007-5">[5]</a> DLCs are highly hydrophobic and lipid soluble with half-lives in humans up to seven years or more. <a href="#cite_note-Milbrath_2009-6">[6]</a> Hence, a single high-dose exposure to a highly persistent compound such as TCDD can lead to sustained AHR activation sufficient to trigger some of the early responses. <a href="#cite_note-Saurat_2012-7">[7]</a> In rodents, some of these high-dose responses may lead to the occurrence of KEs. <a href="#cite_note-Stinchcombe_1995-8">[8]</a> Exposure to lower doses over a long time may also produce sustained activation but may not culminate in increased tumour incidence. <a href="#cite_note-NTP_TCDD_2006a-9">[9]</a> Endogenous ligands have very different chemical characteristics than halogenated dibenzo-p-dioxins, dibenzofurans, and dioxin-like PCBs. The endogenous ligand FICZ is a more potent AHR agonist than TCDD but is rapidly metabolized by CYP1A1 in a negative feedback loop. As noted, control of AHR activation by endogenous ligands probably plays a role in development. <a href="#cite_note-Gasiewicz_2008-3">[3]</a> <a href="#cite_note-Wincent_2009-10">[10]</a> <a href="#cite_note-Wincent_2012-11">[11]</a> Ideally, a table similar to Table 3 could be assembled with data on a range of diverse chemicals with results from common assays providing measures of potency, efficacy and metabolism. This information set may be of some utility in predicting the likelihood of a particular chemical to produce sustained AHR activation (the MIE). However, the use of structural alerts or QSAR considerations to predict the occurrence of the MIE remains an area of interest and may become more feasible in the future. <div class="thumb tright"><div class="thumbinner" style="width:182px;"><a href="/wiki/index.php/File:Table2.jpg" class="image"><img alt="Table 3 alt text" src="/wiki/images/thumb/f/fb/Table2.jpg/180px-Table2.jpg" width="180" height="119" class="thumbimage" srcset="/wiki/images/thumb/f/fb/Table2.jpg/270px-Table2.jpg 1.5x, /wiki/images/thumb/f/fb/Table2.jpg/360px-Table2.jpg 2x" /></a> <div class="thumbcaption"><div class="magnify"><a href="/wiki/index.php/File:Table2.jpg" class="internal" title="Enlarge"><img src="/wiki/skins/common/images/magnify-clip.png" width="15" height="11" alt="" /></a></div>Table 3: Application of the dose and temporal concordance Hill considerations for key events in rodents treated with TCDD</div></div></div>

Generally, AHR ligands are relatively large molecules with hydrophobic nature. Hence, these compounds have low volatility and are poorly absorbed through the skin. Therefore, the assumed route of exposure for this AOP is oral. The primary source of dioxin exposure in humans is animal-based food products, and the dioxin levels in foods have been declining over time. <a href="#cite_note-Aylward_2002-12">[12]</a> <a href="#cite_note-Hays_2003-13">[13]</a> <a href="#cite_note-Lorber_2002-14">[14]</a> <a href="#cite_note-Lorber_2009-15">[15]</a> The dose of PCDD/Fs from dietary exposure can be expressed as TCDD toxic equivalents or TEQ. <a href="#cite_note-VanDenBerg_2006-16">[16]</a> Worldwide, this exposure has been estimated at less than 1 pg TEQ/kg/d. <a href="#cite_note-Fromme_2009-17">[17]</a> <a href="#cite_note-Windal_2010-18">[18]</a> Exposures may be somewhat higher in populations consuming relatively more fish and shellfish. <a href="#cite_note-Nakatani_2010-19">[19]</a> Exposure to the remaining universe of exogenous and endogenous AHR ligands is not nearly as well characterised as is exposure to DLCs. A sampling of this universe consists of food products, substances in commercial and consumer products, phytoestrogens, prostaglandins, catechins in green tea, bilirubin and biliverdin, tryptophan, and its metabolites. <a href="#cite_note-Ahn_2008-20">[20]</a> <a href="#cite_note-Bohonowych_2008-21">[21]</a> <a href="#cite_note-Denison_2003-2">[2]</a> <a href="#cite_note-Denison_2011-1">[1]</a> <a href="#cite_note-ElGendy_2012-22">[22]</a> <a href="#cite_note-HeathPag_1998-23">[23]</a> <a href="#cite_note-Hu_2007-24">[24]</a> <a href="#cite_note-Jeuken_2003-25">[25]</a> <a href="#cite_note-Knockaert_2004-26">[26]</a> <a href="#cite_note-Lawrence_2008-27">[27]</a> <a href="#cite_note-Phelan_1998-28">[28]</a> <a href="#cite_note-Seidel_2001-29">[29]</a> <a href="#cite_note-Tiong_2012-30">[30]</a> <a href="#cite_note-Williams_2000-31">[31]</a> <a href="#cite_note-Wincent_2009-10">[10]</a> <a href="#cite_note-Wincent_2012-11">[11]</a> <a href="#cite_note-Zhao_2006-32">[32]</a> <a href="#cite_note-Zhao_2010-33">[33]</a> <a href="#cite_note-Zhao_2013-34">[34]</a> Substantial AHR-activity has been reported in human serum and likely reflects many endogenous and naturally occurring AHR ligands from the diet. <a href="#cite_note-Connor_2008-35">[35]</a> The high level of background TEQ may serve as a ModF for the tumour promotion response to sustained AHR activation initiated by sufficient dosages of chlorinated dibenzo-p-dioxins, dibenzofurans and dioxin-like PCBs.

2016-11-29T18:42:13

2016-11-29T21:19:06

1002698

NCBI Mammalia sp. AVB-2011

435435

NCBI Rattus sp. ABTC 42503

10090

NCBI mouse

10116

NCBI rat

WikiUser_19

ApacheUser rodentia

310402

NCBI Mus sp. 2000082

N/A, Hepatotoxicity, Hepatopathy, including a constellation of observable effects

Tissue

As defined in NTP (2006a), toxic hepatopathy consists of a constellation of different histological observations including, but not limited to, hepatocyte hypertrophy, multinucleated hepatocytes, inflammation, pigmentation, steatosis, portal fibrosis, bile duct cysts, cholangiofibrosis, mitochondrial injury, necrosis, fibrosis, and porphyria, as well as bile duct and oval cell hyperplasia (Boverhof et al., 2005; Chang et al., 2005; Hailey et al., 2005; Jones and Greig, 1975; NTP, 2006a; NTP, 2006b; NTP, 2006c; NTP, 2006d; NTP, 2006e; NTP, 2006f; Walker et al., 2006; Simon et al., 2009). In this AOP, Hepatotoxicity/Hepatopathy is distinguished from Cellular Proliferation/Hyperplasia as the histological observations that comprise toxic hepatopathy but without bile duct hyperplasia and oval cell hyperplasia.

As noted, Hepatotoxicity/Hepatopathy includes the histologically observed effects that comprise toxic hepatopathy save the proliferative effects. Necrosis and inflammation are not observed after 52 weeks of DLC administration but occur during the second year of treatment and are observed at the 2-year termination of the NTP cancer bioassays. Necrosis and inflammation are observed at lower doses than those at which tumors were observed (Hailey et al., 2005). Damage to hepatocytes and cytotoxicity are common features of these histopathological changes. This provides empirical support for the hypothesis that regenerative repair may be a contributor to the proliferative response and the adverse effects observed at the higher TCDD dosages.

The relationship of this constellation of organ-level effects to hepatocellular cancer is well established in mammalian species; for example, steatosis and nonalcoholic steatohepatitis increase the risk of liver cancer in humans and rodents (Ip and Wang, 2014; Nakamura and Terauchi, 2013). Sustained AHR activation induces steatosis and could act in a similar fashion.

High

Boverhof, D.R., Burgoon, L.D., Tashiro, C., Chittim, B., Harkema, J.R., Jump, D.B., Zacharewski, T.R., 2005. Temporal and dose-dependent hepatic gene expression patterns in mice provide new insights into TCDD-Mediated hepatotoxicity. Toxicol. Sci. 85, 1048-1063. Chang, H., Wang, Y.J., Chang, L.W., Lin, P., 2005. A histochemical and pathological study on the interrelationship between TCDD-induced AhR expression, AhR activation, and hepatotoxicity in mice. J. Toxicol. Environ. Heal. A. 68, 1567-1579. Hailey, J.R., Walker, N.J., Sells, D.M., Brix, A.E., Jokinen, M.P., Nyska, A., 2005. Clas- sification of proliferative hepatocellular lesions in harlan sprague-dawley rats chronically exposed to dioxin-like compounds. Toxicol. Pathol. 33, 165-174. Ip, B.C., Wang, X.-D., 2014. Non-alcoholic steatohepatitis and hepatocellular carci- noma: implications for lycopene intervention. Nutrients 6, 124-162. Jones, G., Greig, J.B., 1975. Pathological changes in the liver of mice given 2,3,7,8-tetrachlorodibenzo-p-dioxin. Experientia 31, 1315-1317. Nakamura, A., Terauchi, Y., 2013. Lessons from mouse models of high-fat diet- induced NAFLD. Int. J. Mol. Sci. 14, 21240-21257. NTP, 2006a. NTP technical report on the toxicology and carcinogenesis studies of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (CAS No. 1746-01-6) in female Harlan Sprague-Dawley rats (Gavage Studies). Natl. Toxicol. Program Tech. Rep. Ser. 4-232. NTP, 2006b. NTP toxicology and carcinogenesis studies of 2,3,4,7,8- Pentachlorodibenzofuran (PeCDF) (CAS No. 57117-31-4) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 1-198. NTP, 2006c. NTP toxicology and carcinogenesis studies of 3,3',4,4',5- pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) in female Harlan Sprague-Dawley rats (Gavage Studies). Natl. Toxicol. Program Tech. Rep. Ser. 4-246. NTP, 2006d. NTP technical report on the toxicology and carcinogenesis studies of 2,2',4,4',5,5'-hexachlorobiphenyl (PCB 153) (CAS No. 35065-27-1) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 4-168. NTP, 2006e. NTP toxicology and carcinogenesis studies of a binary mixture of 3,3' ,4,4' ,5-Pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) and 2,20,4,40,5,50-Hexachlorobiphenyl (PCB 153) (CAS No. 35065-27-1) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 1-258. NTP, 2006f. NTP toxicology and carcinogenesis studies of a mixture of 2,3,7,8- tetrachlorodibenzo-p-dioxin (TCDD) (CAS No. 1746-01-6), 2,3,4,7,8- pentachlorodibenzofuran (PeCDF) (CAS No. 57117-31-4), and 3,3',,4,4' ,5- pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 1-180. Simon, T., Aylward, L.L., Kirman, C.R., Rowlands, J.C., Budinsky, R.A., 2009. Estimates of cancer potency of 2,3,7,8-tetrachlorodibenzo(p)dioxin using linear and nonlinear dose-response modeling and toxicokinetics. Toxicol. Sci. 112, 490-506. Walker, N.J., Wyde, M.E., Fischer, L.J., Nyska, A., Bucher, J.R., 2006. Comparison of chronic toxicity and carcinogenicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in 2-year bioassays in female Sprague-Dawley rats. Mol. Nutr. Food Res. 50, 934-944. AOPWiki 2016-11-29T18:41:23

2016-11-29T18:59:10

Activation, Long term AHR receptor driven direct and indirect gene expression changes

Cellular

MIE: Macromolecular Interactions and Sustained Ligand-Activation of Transcription Insight into sustained AHR activation is provided by examining the induction of ethoxyresorufin-O,O-deethylase in liver by three DLCs at three different time points (NTP, 2006a; NTP, 2006b; NTP, 2006c). Plots of the fractional or normalized ethoxyresorufin-O,O- deethylase response from three NTP cancer bioassays are shown in the plots on the left of Fig. 2. Induction of ethoxyresorufin-O,O- deethylase is easily measured and serves as a biomarker of CYP1A1 gene expression. The dose term on the x-axis is the area under the curve (AUC) of liver concentration. The normalized ethoxyresorufin-O,O-deethylase response on the y-axis is similar at 14, 31 and 53 weeks (Left column of Fig. 2). The plots for 31 weeks and 53 weeks are shifted to the right given the dose term on the x-axis is the AUC of hepatic concentration of the three chemicals, TCDD, 4-PeCDF and PCB-126. To obtain a measure of sustained AHR activation, the fractional ethoxyresorufin-O,O- deethylase response is multiplied by the number of weeks. Hence, a fractional response of 50% at 14 weeks would be a sustained AHR activation index of 7. The sustained AHR activation index is plotted versus the AUC of hepatic toxic equivalents (TEQ) for all three chemicals calculated using TEF values of 1.0, 0.3 and 0.1 for TCDD, 4-PeCDF and PCB-126 respectively (Van den Berg et al., 2006) (Fig. 2, right column). The sustained AHR activation index shows a strong relationship to the AUC of hepatic TEQ and thus the sustained AHR activation index reflects the dose, potency and duration of DLCs in the liver. Dose-response modeling can be performed using the sustained AHR activation index as the dose term and the measures of the various KEs or biomarkers as the response. Fig. 4 shows an example in which the well-known Hill dose-response model was used. One of the model parameters is the ED50 or EC50 value e in other words, the effective dose or concentration sufficient to produce a 50% of the maximal response. This parameter is also called the half- maximal dose. When this measure of sustained AHR activation is used, the ESA50 denotes the level of sustained AHR activation required for a half-maximal response. Please also see Becker, R.A., Patlewicz, G., Simon, T.W., Rowlands, J.C., Budinsky, R.A. 2015. The adverse outcome pathway for rodent liver tumor promotion by sustained activation of the aryl hydrocarbon receptor. Regul. Toxicol. Pharmacol. 73, 172-190: PMID: 26145830. The file is open access.

Dose-response modeling can be performed using the sustained AHR activation index as the dose term and the measures of the various KEs or biomarkers as the response. Figure 3 shows an example in which the well-known Hill dose-response model was used. One of the model parameters is the ED50 or EC50 value e in other words, the effective dose or concentration sufficient to produce a 50% of the maximal response. This parameter is also called the half- maximal dose. When this measure of sustained AHR activation is used, the ESA50 denotes the level of sustained AHR activation required for a half-maximal response. Figure 2 <div> <div><a class="image" href="/wiki/index.php/File:CYP1A1_activation_AHR.jpg"><img alt="Figure 2 alt text" class="thumbimage" src="/wiki/images/thumb/9/99/CYP1A1_activation_AHR.jpg/180px-CYP1A1_activation_AHR.jpg" style="height:240px; width:180px" /></a> <div> <div><a class="internal" href="/wiki/index.php/File:CYP1A1_activation_AHR.jpg" title="Enlarge"><img alt="" src="/wiki/skins/common/images/magnify-clip.png" style="height:11px; width:15px" /></a></div> Figure 2: Dose-response of CYP1A1 activation measured by EROD as a measure of AHR activation in response to an AUC measure of dose. The AUC is the hepatic concentration multiplied by the time in weeks. The left hand plots show measured EROD from the NTP bioassays in response to chronic dosing of TCDD (top), 4-PeCDF (middle) or PCB-126 (bottom) (NTP, 2006a, 2006b, 2006c). Dose-dependent EROD levels are sustained over time. The right hand plots show the EROD response on normalized to a zero-to-one scale as a measure of AHR activation level.</div> </div> </div> shows plots of the increase in 7-Ethoxyresorufin-O-deethylase (EROD) versus the area-under-the-curve (AUC) of the hepatic concentration of three dioxin-like chemicals, TCDD, 4-PeCDF and PCB-126 occurring during lifetime dosing. The figure shows corresponding plots of normalized EROD as a measure of AHR activation. In all three cases, the shape of the dose-response remained consistent except that the position along the dose axis increased with increasing time for all three chemicals. This observation indicates that the level of AHR activation remains relatively constant over time with a continuing dose of persistent AHR ligand. A measure of sustained activation (SA) of the AHR can be calculated by multiplying the level of AHR activation observed as fractional CYP1A1 induction by the number of weeks of dosing. SA calculated in this way can be used as a dose surrogate. In order to use the measurements of all three DLCs considered here, their hepatic AUC concentrations were multiplied by their toxic equivalence factors (TEFs) (Van den Berg et al., 2006). SA was plotted against the AUC of hepatic TEQ concentration (Figure 3). <div> <div><a class="image" href="/wiki/index.php/File:AUC_of_AHR.jpg"><img alt="Figure 3 alt text" class="thumbimage" src="/wiki/images/thumb/5/5c/AUC_of_AHR.jpg/180px-AUC_of_AHR.jpg" style="height:135px; width:180px" /></a> <div> <div><a class="internal" href="/wiki/index.php/File:AUC_of_AHR.jpg" title="Enlarge"><img alt="" src="/wiki/skins/common/images/magnify-clip.png" style="height:11px; width:15px" /></a></div> Figure 3: Sustained AHR activation versus the area-under-the-curve of hepatic TEQ. TEQ was calculated for TCDD, PeCDF and PCB126 using TEF values of 1, 0.3 and 0.1 respectively. Data from all three chemicals at 14 weeks are shown with blue markers, those from 31 weeks with green markers and those from 53 weeks with orange markers. Please see narrative for additional details.</div> </div> </div> The plot shows a consistent pattern that can be fit with a Hill function. The curve is shallower than a first-order Hill plot shown by the Hill coefficient less than one. This shallowness may be due to differences in potency and efficacy at different times, possibly stemming from hepatic sequestration of ligand by CYP1A2, especially for PeCDF, thus decreasing the effective hepatic concentration relative to the measured concentration.

At a number of levels of biological organization, differences exist between the human and rodent AHR. Considering toxicodynamics, the human AHR binding affinity is an order of magnitude or more lower than that in rodents that is generally correlated with reduced sensitivity in human hepatocytes relative to rats (Black et al., 2012; Budinsky et al., 2010; Connor and Aylward, 2006). In addition, these species differences include AHR binding affinity, different recruit- ment of co-regulatory proteins, and different patterns of gene regulation (Black et al., 2012; Budinsky et al., 2010; Carlson et al., 2009; Connor and Aylward, 2006; Dere et al., 2011; Flaveny et al., 2010).

High High

Abel, J., Haarmann-Stemmann, T., 2010. An introduction to the molecular basics of aryl hydrocarbon receptor biology. Biol. Chem. 391, 1235e1248. http:// dx.doi.org/10.1515/BC.2010.128. Andersen, M.E., Preston, R.J., Maier, A., Willis, A.M., Patterson, J., 2014. Dose- response approaches for nuclear receptor-mediated modes of action for liver carcinogenicity: results of a workshop. Crit. Rev. Toxicol. 44, 50e63. http:// dx.doi.org/10.3109/10408444.2013.835785. Angrish, M.M., Jones, A.D., Harkema, J.R., Zacharewski, T.R., 2011. Aryl hydrocarbon receptor-mediated induction of stearoyl-CoA desaturase 1 alters hepatic fatty acid composition in TCDD-elicited steatosis. Toxicol. Sci. 124, 299e310. http:// dx.doi.org/10.1093/toxsci/kfr226. Beischlag, T.V., Luis Morales, J., Hollingshead, B.D., Perdew, G.H., 2008. The aryl hydrocarbon receptor complex and the control of gene expression. Crit. Rev. Eukaryot. Gene. Expr. 18, 207e250, 6f28b0540a5e6e63,5ec7b3e06964879d [pii]. Bendall, S.C., Nolan, G.P., 2012. From single cells to deep phenotypes in cancer. Nat. Biotechnol. 30, 639e647. <a class="external free" href="http://dx.doi.org/10.1038/nbt.2283" rel="nofollow" target="_blank">http://dx.doi.org/10.1038/nbt.2283</a>. Black, M.B., Budinsky, R.A., Dombkowski, A., Lecluyse, E.L., Ferguson, S.S., Thomas, R.S., Rowlands, J.C., 2012. Cross-species comparisons of transcriptomic alterations in human and rat primary hepatocytes exposed to 2,3,7,8- tetrachlorodibenzo-p-dioxin. Toxicol. Sci. 127, 199e215. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.1093/toxsci/kfs069. Boutros, P.C., Bielefeld, K.A., Pohjanvirta, R., Harper, P.A., 2009. Dioxin-dependent and dioxin-independent gene batteries: comparison of liver and kidney in AHR- null mice. Toxicol. Sci. 112, 245e256 kfp191 [pii] 10.1093/toxsci/kfp191. Boutros, P.C., Moffat, I.D., Franc, M.A., Tijet, N., Tuomisto, J., Pohjanvirta, R., Okey, A.B., 2004. Dioxin-responsive AHRE-II gene battery: identification by phylogenetic footprinting. Biochem. Biophys. Res. Commun. 321, 707e715. Boutros, P.C., Yao, C.Q., Watson, J.D., Wu, A.H., Moffat, I.D., Prokopec, S.D., Smith, A.B., Okey, A.B., Pohjanvirta, R., 2011. Hepatic transcriptomic responses to TCDD in dioxin-sensitive and dioxin-resistant rats during the onset of toxicity. Toxicol. Appl. Pharmacol. 251, 119e129. <a class="external free" href="http://dx.doi.org/10.1016/" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/</a> j.taap.2010.12.010. S0041-008X(10)00467-9 [pii]. Boverhof, D.R., Burgoon, L.D., Tashiro, C., Chittim, B., Harkema, J.R., Jump, D.B., Zacharewski, T.R., 2005. Temporal and dose-dependent hepatic gene expression patterns in mice provide new insights into TCDD-Mediated hepatotoxicity. Toxicol. Sci. 85, 1048e1063. Boverhof, D.R., Burgoon, L.D., Tashiro, C., Sharratt, B., Chittim, B., Harkema, J.R., Mendrick, D.L., Zacharewski, T.R., 2006. Comparative toxicogenomic analysis of the hepatotoxic effects of TCDD in Sprague Dawley rats and C57BL/6 mice. Toxicol. Sci. 94, 398e416. Brauze, D., Rawłuszko, A.A., 2012. The effect of aryl hydrocarbon receptor ligands on the expression of polymerase (DNA directed) kappa (Polk), polymerase RNA II (DNA directed) polypeptide A (PolR2a), CYP1B1 and CYP1A1 genes in rat liver. Environ. Toxicol. Pharmacol. 34, 819e825. <a class="external free" href="http://dx.doi.org/10.1016/" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/</a> j.etap.2012.09.004. Budinsky, R.A., LeCluyse, E.L., Ferguson, S.S., Rowlands, J.C., Simon, T., 2010. Human and rat primary hepatocyte CYP1A1 and 1A2 induction with 2,3,7,8- tetrachlorodibenzo-p-dioxin, 2,3,7,8-tetrachlorodibenzofuran, and 2,3,4,7,8- pentachlorodibenzofuran. Toxicol. Sci. 118, 224e235 kfq238 [pii] 10.1093/ toxsci/kfq238. Budinsky, R.A., Schrenk, D., Simon, T., Van den Berg, M., Reichard, J.F., Silkworth, J.B., Aylward, L.L., Brix, A., Gasiewicz, T., Kaminski, N., Perdew, G., Starr, T.B., Walker, N.J., Rowlands, J.C., 2014. Mode of action and dose-response framework analysis for receptor-mediated toxicity: the aryl hydrocarbon receptor as a case study. Crit. Rev. Toxicol. 44, 83e119. <a class="external free" href="http://dx.doi.org/10.3109/" rel="nofollow" target="_blank">http://dx.doi.org/10.3109/</a> 10408444.2013.835787. Connor, K.T., Aylward, L.L., 2006. Human response to dioxin: aryl hydrocarbon re- ceptor (AhR) molecular structure, function, and dose-response data for enzyme induction indicate an impaired human AhR. J. Toxicol. Environ. Health B Crit. Rev. 9, 147e171. Carlson, E.A., McCulloch, C., Koganti, A., Goodwin, S.B., Sutter, T.R., Silkworth, J.B., 2009. Divergent transcriptomic responses to aryl hydrocarbon receptor agonists between rat and human primary hepatocytes. Toxicol. Sci. 112, 257e272. http:// dx.doi.org/10.1093/toxsci/kfp200 kfp200 [pii]. Chia, N.-Y., Ng, H.-H., 2012. Stem cell genome-to-systems biology. Wiley Interdiscip. Rev. Syst. Biol. Med. 4, 39e49. <a class="external free" href="http://dx.doi.org/10.1002/wsbm.151" rel="nofollow" target="_blank">http://dx.doi.org/10.1002/wsbm.151</a>. Dere, E., Lee, A.W., Burgoon, L.D., Zacharewski, T.R., 2011. Differences in TCDD- elicited gene expression profiles in human HepG2, mouse Hepa1c1c7 and rat H4IIE hepatoma cells. BMC Genomics 12, 193. <a class="external free" href="http://dx.doi.org/10.1186/1471-" rel="nofollow" target="_blank">http://dx.doi.org/10.1186/1471-</a> 2164-12-193. Denison, M.S., Soshilov, A.A., He, G., DeGroot, D.E., Zhao, B., 2011. Exactly the same but different: promiscuity and diversity in the molecular mechanisms of action of the aryl hydrocarbon (dioxin) receptor. Toxicol. Sci. 124, 1e22. http:// dx.doi.org/10.1093/toxsci/kfr218. Dietrich, C., Kaina, B., 2010. The aryl hydrocarbon receptor (AhR) in the regulation of cell-cell contact and tumor growth. Carcinogenesis 31, 1319e1328. http:// dx.doi.org/10.1093/carcin/bgq028 bgq028 [pii]. Ellinger-Ziegelbauer, H., Gmuender, H., Bandenburg, A., Ahr, H.J., 2008. Prediction of a carcinogenic potential of rat hepatocarcinogens using toxicogenomics analysis of short-term in vivo studies. Mutat. Res. 637, 23e39. <a class="external free" href="http://dx.doi.org/10.1016/" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/</a> j.mrfmmm.2007.06.010. Ellinger-Ziegelbauer, H., Stuart, B., Wahle, B., Bomann, W., Ahr, H.J., 2005. Com- parison of the expression profiles induced by genotoxic and nongenotoxic carcinogens in rat liver. Mutat. Res. 575, 61e84. <a class="external free" href="http://dx.doi.org/10.1016/" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/</a> j.mrfmmm.2005.02.004. Fielden, M.R., Adai, A., Dunn, R.T., Olaharski, A., Searfoss, G., Sina, J., Aubrecht, J., Boitier, E., Nioi, P., Auerbach, S., Jacobson-Kram, D., Raghavan, N., Yang, Y., Kincaid, A., Sherlock, J., Chen, S.-J., Car, B., Predictive Safety Testing Consortium, Carcinogenicity Working Group, 2011. Development and evaluation of a genomic signature for the prediction and mechanistic assessment of non- genotoxic hepatocarcinogens in the rat. Toxicol. Sci. 124, 54e74. http:// dx.doi.org/10.1093/toxsci/kfr202. Fielden, M.R., Brennan, R., Gollub, J., 2007. A gene expression biomarker provides early prediction and mechanistic assessment of hepatic tumor induction by nongenotoxic chemicals. Toxicol. Sci. 99, 90e100. <a class="external free" href="http://dx.doi.org/10.1093/" rel="nofollow" target="_blank">http://dx.doi.org/10.1093/</a> toxsci/kfm156. Flaveny, C.A., Murray, I.A., Perdew, G.H., 2010. Differential gene regulation by the human and mouse aryl hydrocarbon receptor. Toxicol. Sci. 114, 217e225. http:// dx.doi.org/10.1093/toxsci/kfp308 kfp308 [pii]. Fletcher, N., Wahlstrom, D., Lundberg, R., Nilsson, C.B., Nilsson, K.C., Stockling, K., Hellmold, H., Hakansson, H., 2005. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) alters the mRNA expression of critical genes associated with cholesterol metabolism, bile acid biosynthesis, and bile transport in rat liver: a microarray study. Toxicol. Appl. Pharmacol. 207, 1e24. Forgacs, A.L., Burgoon, L.D., Lynn, S.G., LaPres, J.J., Zacharewski, T., 2010. Effects of TCDD on the expression of nuclear encoded mitochondrial genes. Toxicol. Appl. Pharmacol. 246, 58e65. <a class="external free" href="http://dx.doi.org/10.1016/j.taap.2010.04.006" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/j.taap.2010.04.006</a>. S0041- 008X(10)00133-X [pii]. Franc, M.A., Moffat, I.D., Boutros, P.C., Tuomisto, J.T., Tuomisto, J., Pohjanvirta, R., Okey, A.B., 2008. Patterns of dioxin-altered mRNA expression in livers of dioxin- sensitive versus dioxin-resistant rats. Arch. Toxicol. 82, 809e830. http:// dx.doi.org/10.1007/s00204-008-0303-0. Furness, S.G., Whelan, F., 2009. The pleiotropy of dioxin toxicityexenobiotic misappropriation of the aryl hydrocarbon receptor's alternative physiological roles. Pharmacol. Ther. 124, 336e353. <a class="external free" href="http://dx.doi.org/10.1016/j.pharm-" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/j.pharm-</a> thera.2009.09.004. S0163-7258(09)00187-9 [pii]. Gasiewicz, T.A., Henry, E.C., Collins, L.L., 2008. Expression and activity of aryl hy- drocarbon receptors in development and cancer. Crit. Rev. Eukaryot. Gene Expr. 18, 279e321, 4de8c9fe5b6af78b,714c550a3fb330d1 [pii]. George, C.L., Lightman, S.L., Biddie, S.C., 2011. Transcription factor interactions in genomic nuclear receptor function. Epigenomics 3, 471e485. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.2217/epi.11.66. Hailey, J.R., Walker, N.J., Sells, D.M., Brix, A.E., Jokinen, M.P., Nyska, A., 2005. Clas- sification of proliferative hepatocellular lesions in harlan sprague-dawley rats chronically exposed to dioxin-like compounds. Toxicol. Pathol. 33, 165e174. <a class="external free" href="http://dx.doi.org/10.1080/01926230590888324" rel="nofollow" target="_blank">http://dx.doi.org/10.1080/01926230590888324</a>. TBE21PK0FLEJ66J7 [pii]. Harrill, J.A., Hukkanen, R.R., Lawson, M., Martin, G., Gilger, B., Soldatow, V., Lecluyse, E.L., Budinsky, R.A., Rowlands, J.C., Thomas, R.S., 2013. Knockout of the aryl hydrocarbon receptor results in distinct hepatic and renal phenotypes in rats and mice. Toxicol. Appl. Pharmacol. 272, 503e518. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.1016/j.taap.2013.06.024. Heise, T., Schug, M., Storm, D., Ellinger-Ziegelbauer, H., Ahr, H.J., Hellwig, B., Rahnenfuhrer, J., Ghallab, A., Guenther, G., Sisnaiske, J., Reif, R., Godoy, P., Mielke, H., Gundert-Remy, U., Lampen, A., Oberemm, A., Hengstler, J.G., 2012. In vitro e in vivo correlation of gene expression alterations induced by liver carcinogens. Curr. Med. Chem. 19, 1721e1730. Ikeda, H., Nishi, S., Sakai, M., 2004. Transcription factor Nrf2/MafK regulates rat placental glutathione S-transferase gene during hepatocarcinogenesis. Bio- chem.. J. 380, 515e521. <a class="external free" href="http://dx.doi.org/10.1042/BJ20031948" rel="nofollow" target="_blank">http://dx.doi.org/10.1042/BJ20031948</a>. Jaramillo, M.C., Zhang, D.D., 2013. The emerging role of the Nrf2-Keap1 signaling pathway in cancer. Genes. Dev. 27, 2179e2191. <a class="external free" href="http://dx.doi.org/10.1101/gad.225680.113" rel="nofollow" target="_blank">http://dx.doi.org/10.1101/gad.225680.113</a>. Julien, E., Boobis, A.R., Olin, S.S., Ilsi Research Foundation Threshold Working Group, 2009. The key events dose-response framework: a cross-disciplinary mode-of- action based approach to examining dose-response and thresholds. Crit. Rev. Food. Sci. Nutr. 49, 682e689. <a class="external free" href="http://dx.doi.org/10.1080/10408390903110692" rel="nofollow" target="_blank">http://dx.doi.org/10.1080/10408390903110692</a>. Kumar, M.B., Ramadoss, P., Reen, R.K., Vanden Heuvel, J.P., Perdew, G.H., 2001. The Q-rich subdomain of the human Ah receptor transactivation domain is required for dioxin-mediated transcriptional activity. J. Biol. Chem. 276, 42302e42310. <a class="external free" href="http://dx.doi.org/10.1074/jbc.M104798200" rel="nofollow" target="_blank">http://dx.doi.org/10.1074/jbc.M104798200</a>. M104798200 [pii]. Kumar, R., Wang, R.-A., Barnes, C.J., 2004. Coregulators and chromatin remodeling in transcriptional control. Mol. Carcinog. 41, 221e230. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.1002/mc.20056. Kwak, M.K., Itoh, K., Yamamoto, M., Sutter, T.R., Kensler, T.W., 2001. Role of tran- scription factor Nrf2 in the induction of hepatic phase 2 and antioxidative enzymes in vivo by the cancer chemoprotective agent, 3H-1, 2-dimethiole-3- thione. Mol. Med. 7, 135e145. Le Vee, M., Jouan, E., Fardel, O., 2010. Involvement of aryl hydrocarbon receptor in basal and 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced expression of target genes in primary human hepatocytes. Toxicol. In Vitro 24, 1775e1781. http:// dx.doi.org/10.1016/j.tiv.2010.07.001. Lee, K.C., Lee Kraus, W., 2001. Nuclear receptors, coactivators and chromatin: new approaches, new insights. Trends Endocrinol. Metab. 12, 191e197. Lees, M.J., Whitelaw, M.L., 1999. Multiple roles of ligand in transforming the dioxin receptor to an active basic helix-loop-helix/PAS transcription factor complex with the nuclear protein Arnt. Mol. Cell. Biol. 19, 5811e5822. Liby, K.T., Sporn, M.B., 2012. Synthetic oleanane triterpenoids: multifunctional drugs with a broad range of applications for prevention and treatment of chronic disease. Pharmacol. Rev. 64, 972e1003. <a class="external free" href="http://dx.doi.org/10.1124/" rel="nofollow" target="_blank">http://dx.doi.org/10.1124/</a> pr.111.004846. Lu, H., Cui, W., Klaassen, C.D., 2011. Nrf2 protects against 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD)-induced oxidative injury and steatohepatitis. Toxicol. Appl. Pharmacol. 256, 122e135. <a class="external free" href="http://dx.doi.org/10.1016/j.taap.2011.07.019" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/j.taap.2011.07.019</a>. Ma, Q., Baldwin, K.T., Renzelli, A.J., McDaniel, A., Dong, L., 2001. TCDD-inducible poly(ADP-ribose) polymerase: a novel response to 2,3,7,8-tetrachlorodibenzo- p-dioxin. Biochem. Biophys. Res. Commun. 289, 499e506. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.10 06/bbrc.20 01.5987. Ma, Q., 2011. Influence of light on aryl hydrocarbon receptor signaling and conse- quences in drug metabolism, physiology and disease. Expert. Opin. Drug. Metab. Toxicol. 7, 1267e1293. <a class="external free" href="http://dx.doi.org/10.1517/17425255.2011.614947" rel="nofollow" target="_blank">http://dx.doi.org/10.1517/17425255.2011.614947</a>. Ma, Q., He, X., 2012. Molecular basis of electrophilic and oxidative defense: promises and perils of Nrf2. Pharmacol. Rev. 64, 1055e1081. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.1124/pr.110.004333. Matthews, J., Gustafsson, J.A., 2006. Estrogen receptor and aryl hydrocarbon re- ceptor signaling pathways. Nucl. Recept. Signal 4, e016. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.1621/nrs.04016. Matthews, J., Wihlen, B., Thomsen, J., Gustafsson, J.A., 2005. Aryl hydrocarbon receptor-mediated transcription: ligand-dependent recruitment of estrogen receptor alpha to 2,3,7,8-tetrachlorodibenzo-p-dioxin-responsive promoters. Mol. Cell Biol. 25, 5317e5328. <a class="external free" href="http://dx.doi.org/10.1128/MCB.25.13.5317-" rel="nofollow" target="_blank">http://dx.doi.org/10.1128/MCB.25.13.5317-</a> 5328.2005, 25/13/5317 [pii]. Moffat, I.D., Boutros, P.C., Chen, H., Okey, A.B., Pohjanvirta, R., 2010. Aryl hydro- carbon receptor (AHR)-regulated transcriptomic changes in rats sensitive or resistant to major dioxin toxicities. BMC Genomics 11, 263. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.1186/1471-2164-11-263. Nie, A.Y., McMillian, M., Parker, J.B., Leone, A., Bryant, S., Yieh, L., Bittner, A., Nelson, J., Carmen, A., Wan, J., Lord, P.G., 2006. Predictive toxicogenomics ap- proaches reveal underlying molecular mechanisms of nongenotoxic carcino- genicity. Mol. Carcinog. 45, 914e933. <a class="external free" href="http://dx.doi.org/10.1002/mc.20205.NTP" rel="nofollow" target="_blank">http://dx.doi.org/10.1002/mc.20205.NTP</a>, 2006a. NTP technical report on the toxicology and carcinogenesis studies of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (CAS No. 1746-01-6) in female Harlan Sprague-Dawley rats (Gavage Studies). Natl. Toxicol. Program Tech. Rep. Ser. 4e232. NTP, 2006b. NTP toxicology and carcinogenesis studies of 2,3,4,7,8- Pentachlorodibenzofuran (PeCDF) (CAS No. 57117-31-4) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 1e198. NTP, 2006c. NTP toxicology and carcinogenesis studies of 3,30,4,40,5- pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) in female Harlan Sprague-Dawley rats (Gavage Studies). Natl. Toxicol. Program Tech. Rep. Ser. 4e246. NTP, 2006d. NTP technical report on the toxicology and carcinogenesis studies of 2,20,4,40,5,50-hexachlorobiphenyl (PCB 153) (CAS No. 35065-27-1) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 4E168. NTP, 2006e. NTP toxicology and carcinogenesis studies of a binary mixture of 3,30 ,4,40 ,5-Pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) and 2,20,4,40,5,50-Hexachlorobiphenyl (PCB 153) (CAS No. 35065-27-1) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 1E258. NTP, 2006f. NTP toxicology and carcinogenesis studies of a mixture of 2,3,7,8- tetrachlorodibenzo-p-dioxin (TCDD) (CAS No. 1746-01-6), 2,3,4,7,8- pentachlorodibenzofuran (PeCDF) (CAS No. 57117-31-4), and 3,30 ,4,40 ,5- pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) in female Harlan Sprague-Dawley rats (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 1E180. NTP, 2010. Toxicology and carcinogenesis studies of 2,30 ,4,40 ,5-pentachlorobiphenyl (PCB 118) (CAS No. 31508-00-6) in female harlan Sprague-Dawley rats (gavage studies). Natl. Toxicol. Program. Tech. Rep. Ser. 1E174. Osburn, W.O., Yates, M.S., Dolan, P.D., Chen, S., Liby, K.T., Sporn, M.B., Taguchi, K., Yamamoto, M., Kensler, T.W., 2008. Genetic or pharmacologic amplification of nrf2 signaling inhibits acute inflammatory liver injury in mice. Toxicol. Sci. 104, 218e227. <a class="external free" href="http://dx.doi.org/10.1093/toxsci/kfn079" rel="nofollow" target="_blank">http://dx.doi.org/10.1093/toxsci/kfn079</a>. Ovando, B.J., Ellison, C.A., Vezina, C.M., Olson, J.R., 2010. Toxicogenomic analysis of exposure to TCDD, PCB126 and PCB153: identification of genomic biomarkers of exposure to AhR ligands, BMC. Genomics 11, 583. <a class="external free" href="http://dx.doi.org/10.1186/" rel="nofollow" target="_blank">http://dx.doi.org/10.1186/</a> 1471-2164-11-583, 1471-2164-11-583 [pii]. Ovando, B.J., Vezina, C.M., McGarrigle, B.P., Olson, J.R., 2006. Hepatic gene down- regulation following acute and subchronic exposure to 2,3,7,8- tetrachlorodibenzo-p-dioxin. Toxicol. Sci. 94, 428e438. Partch, C.L., Gardner, K.H., 2010. Coactivator recruitment: a new role for PAS do- mains in transcriptional regulation by the bHLH-PAS family. J. Cell. Physiol. 223, 553e557. <a class="external free" href="http://dx.doi.org/10.1002/jcp.22067" rel="nofollow" target="_blank">http://dx.doi.org/10.1002/jcp.22067</a>. Perissi, V., Rosenfeld, M.G., 2005. Controlling nuclear receptors: the circular logic of cofactor cycles. Nat. Rev. Mol. Cell. Biol. 6, 542e554. <a class="external free" href="http://dx.doi.org/10.1038/" rel="nofollow" target="_blank">http://dx.doi.org/10.1038/</a> nrm1682. Reen, R.K., Cadwallader, A., Perdew, G.H., 2002. The subdomains of the trans- activation domain of the aryl hydrocarbon receptor (AhR) inhibit AhR and es- trogen receptor transcriptional activity. Arch. Biochem. Biophys. 408, 93e102. Rowlands, J.C., McEwan, I.J., Gustafsson, J.A., 1996. Trans-activation by the human aryl hydrocarbon receptor and aryl hydrocarbon receptor nuclear translocator proteins: direct interactions with basal transcription factors. Mol. Pharmacol. 50, 538e548. Rowlands, J.C., Budinsky, R., Gollapudi, B., Novak, R., Abdelmegeed, M., Cukovic, D., Dombkowski, A., 2011. Transcriptional profiles induced by the Aryl hydrocarbon receptor agonists 2,3,7,8-tetrachlorodibenzo-p-dioxin, 2,3,7,8- tetrachlorodibenzofuran and 2,3,4,7,8-pentachlorodibenzofuran in primary rat hepatocytes. Chemosphere. <a class="external free" href="http://dx.doi.org/10.1016/j.chemo-" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/j.chemo-</a> sphere.2011.06.026. S0045e6535(11)00669-2 [pii]. Sato, S., Shirakawa, H., Tomita, S., Tohkin, M., Gonzalez, F.J., Komai, M., 2013. The aryl hydrocarbon receptor and glucocorticoid receptor interact to activate human metallothionein 2A. Toxicol. Appl. Pharmacol. 273, 90e99. <a class="external free" href="http://dx.doi.org/" rel="nofollow" target="_blank">http://dx.doi.org/</a> 10.1016/j.taap.2013.08.017. Shelton, P., Jaiswal, A.K., 2013. The transcription factor NF-E2-related factor 2 (Nrf2): a protooncogene? FASEB. J. 27, 414e423. <a class="external free" href="http://dx.doi.org/10.1096/fj.12-217257" rel="nofollow" target="_blank">http://dx.doi.org/10.1096/fj.12-217257</a>. Simon, T.W., Simons, S.S., Preston, R.J., Boobis, A.R., Cohen, S.M., Doerrer, N.G., Fenner-Crisp, P.A., McMullin, T.S., McQueen, C.A., Rowlands, J.C., RISK21 Dose- Response Subteam, 2014. The use of mode of action information in risk assessment: quantitative key events/dose-response framework for modeling the dose-response for key events. Crit. Rev. Toxicol. 44 (Suppl. 3), 17e43. http:// dx.doi.org/10.3109/10408444.2014.931925. Soshilov, A., Denison, M.S., 2011. Ligand displaces heat shock protein 90 from overlapping binding sites within the aryl hydrocarbon receptor ligand-binding domain. J. Biol. Chem. 286, 35275e35282. <a class="external free" href="http://dx.doi.org/10.1074/" rel="nofollow" target="_blank">http://dx.doi.org/10.1074/</a> jbc.M111.246439. Tanaka, M., Itoh, T., Tanimizu, N., Miyajima, A., 2011. Liver stem/progenitor cells: their characteristics and regulatory mechanisms. J. Biochem. 149, 231e239. <a class="external free" href="http://dx.doi.org/10.1093/jb/mvr001" rel="nofollow" target="_blank">http://dx.doi.org/10.1093/jb/mvr001</a>. Tappenden, D.M., Hwang, H.J., Yang, L., Thomas, R.S., Lapres, J.J., 2013. The aryl- hydrocarbon receptor protein interaction network (AHR-PIN) as identified by tandem affinity purification (TAP) and mass spectrometry. J. Toxicol. 2013, 279829. <a class="external free" href="http://dx.doi.org/10.1155/2013/279829" rel="nofollow" target="_blank">http://dx.doi.org/10.1155/2013/279829</a>. Tsai, C.-C., Fondell, J.D., 2004. Nuclear receptor recruitment of histone-modifying enzymes to target gene promoters. Vitam. Horm. 68, 93e122. http:// dx.doi.org/10.1016/S0083-6729(04)68003-4. van Delft, J.H.M., van Agen, E., van Breda, S.G.J., Herwijnen, M.H., Staal, Y.C.M., Kleinjans, J.C.S., 2005. Comparison of supervised clustering methods to discriminate genotoxic from non-genotoxic carcinogens by gene expression profiling. Mutat. Res. 575, 17e33. <a class="external free" href="http://dx.doi.org/10.1016/" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/</a> j.mrfmmm.2005.02.006. Van den Berg, M., Birnbaum, L.S., Denison, M., De, V.M., Farland, W., Feeley, M., Fiedler, H., Hakansson, H., Hanberg, A., Haws, L., Rose, M., Safe, S., Schrenk, D., Tohyama, C., Tritscher, A., Tuomisto, J., Tysklind, M., Walker, N., Peterson, R.E., 2006. The 2005 world health organization reevaluation of human and mammalian toxic equivalency factors for dioxins and dioxin-like compounds. Toxicol. Sci. 93, 223e241. Vezina, C.M., Walker, N.J., Olson, J.R., 2004. Subchronic exposure to TCDD, PeCDF, PCB126, and PCB153: effect on hepatic gene expression. Environ. Heal. Perspect. 112, 1636e1644. Wang, L., He, X., Szklarz, G.D., Bi, Y., Rojanasakul, Y., Ma, Q., 2013. The aryl hydro- carbon receptor interacts with nuclear factor erythroid 2-related factor 2 to mediate induction of NAD(P)H:quinoneoxidoreductase 1 by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Arch. Biochem. Biophys. 537, 31e38. http:// dx.doi.org/10.1016/j.abb.2013.06.001. Watanabe, T., Suzuki, T., Natsume, M., Nakajima, M., Narumi, K., Hamada, S., Sakuma, T., Koeda, A., Oshida, K., Miyamoto, Y., Maeda, A., Hirayama, M., Sanada, H., Honda, H., Ohyama, W., Okada, E., Fujiishi, Y., Sutou, S., Tadakuma, A., Ishikawa, Y., Kido, M., Minamiguchi, R., Hanahara, I., Furihata, C., 2012. Discrimination of genotoxic and non-genotoxic hepatocarcinogens by statistical analysis based on gene expression profiling in the mouse liver as determined by quantitative real-time PCR. Mutat. Res. 747, 164e175. http:// dx.doi.org/10.1016/j.mrgentox.2012.04.011. Yao, C.Q., Prokopec, S.D., Watson, J.D., Pang, R., P'ng, C., Chong, L.C., Harding, N.J., Pohjanvirta, R., Okey, A.B., Boutros, P.C., 2012. Inter-strain heterogeneity in rat hepatic transcriptomic responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Toxicol. Appl. Pharmacol. 260, 135e145. <a class="external free" href="http://dx.doi.org/10.1016/" rel="nofollow" target="_blank">http://dx.doi.org/10.1016/</a> j.taap.2012.02.001. Yates, M.S., Kensler, T.W., 2007. Keap1 eye on the target: chemoprevention of liver cancer. Acta Pharmacol. Sin. 28, 1331e1342. <a class="external free" href="http://dx.doi.org/10.1111/j.1745-" rel="nofollow" target="_blank">http://dx.doi.org/10.1111/j.1745-</a> 7254.2007.00688.x. Yates, M.S., Tauchi, M., Katsuoka, F., Flanders, K.C., Liby, K.T., Honda, T., Gribble, G.W., Johnson, D.A., Johnson, J.A., Burton, N.C., Guilarte, T.R., Yamamoto, M., Sporn, M.B., Kensler, T.W., 2007. Pharmacodynamic characterization of che- mopreventive triterpenoids as exceptionally potent inducers of Nrf2-regulated genes. Mol. Cancer Ther. 6, 154e162. <a class="external free" href="http://dx.doi.org/10.1158/1535-7163.MCT-" rel="nofollow" target="_blank">http://dx.doi.org/10.1158/1535-7163.MCT-</a> 06-0516. Zhang, Q., Andersen, M.E., 2007. Dose response relationship in anti-stress gene regulatory networks. PLoS Comput. Biol. 3, e24. <a class="external free" href="http://dx.doi.org/10.1371/" rel="nofollow" target="_blank">http://dx.doi.org/10.1371/</a> journal.pcbi.0030024. AOPWiki 2016-11-29T18:41:23

2016-12-02T11:22:58

Changes/Inhibition, Cellular Homeostasis and Apoptosis

Cellular

Tumor promotion requires a perturbation in the balance between cell gain via mitosis and cell loss via apoptosis (Roberts et al., 1997). Indirectly, the inhibition of apoptosis in either damaged or initiated cells favors their survival, and inhibition of apoptosis af- fords initiated cells an increased opportunity for clonal expansion and autonomous growth with the chance to acquire additional mutations during the process of tumor progression. AHR activation inhibits apoptosis in altered hepatic foci (i.e., initiated hepatic cells), and this inhibition affords cells within altered hepatic foci a sur- vival advantage and increases the likelihood that these cells will acquire additional mutations.

For this KE, initiation-promotion studies provide indirect evidence of inhibition of intrafocal apoptosis due to sustained AHR activation and direct evidence of a threshold for the clonal expansion of altered hepatic foci (Dragan and Schrenk, 2000; Teeguarden et al., 1999). Although increases of cell proliferation could contribute to the increase in size and volume fraction of altered hepatic foci, the greater magnitude of inhibition of apoptosis suggests it is the primary factor contributing to clonal expansion of altered hepatic foci, at least early on (Luebeck et al., 2000; Moolgavkar et al., 1996; Stinchcombe et al., 1995). AHR activators inhibit apoptosis produced by UV light exposure of human cell lines and rat primary hepatocytes (Ambolet-Camoit et al., 2010; Chopra et al., 2009, 2010; Schwarz et al., 2000). Inhibition of apoptosis in primary rat hepatocytes was mediated through phosphorylation and inactivation of p53 and modulation of Mdm2, Tfgb1/4, and AGR2; in addition, inhi- bition of apoptosis required protein synthesis (Ambolet-Camoit et al., 2010; Chopra and Schrenk, 2011; Chopra et al., 2009, 2010; Davis et al., 2001; Franc et al., 2008; Paajarvi et al., 2005; Worner and Schrenk, 1996, 1998). Cytotoxicity appears to occur after intrafocal apoptosis inhibition is measured and inflammation-driven cell proliferation is a somewhat later event. What remains unknown is whether a proliferative response in stem and stellate cells occurs earlier or at the same time as intrafocal apoptosis inhibition. Quantitative stereology is used to quantify the growth of AHF and such studies provide a measure of this KE (Hendrich et al., 1987; Dragan et al., 1997; Teeguarden et al., 1999; Viluksela et al., 2000).

Rodents are highly susceptible to the hepatotoxic, proliferative, and carcinogenic effects of sustained AHR activation induced by TCDD and other dioxin-like chemicals (Hailey et al., 2005; Goodman and Sauer, 1992; Kociba et al., 1978). The sustained AHR activation rodent liver tumor promotion AOP appears to be a pathway that very likely requires exceedance of a threshold for sustained AHR activation for liver cancers to occur in rodents.

CL:0000182

CL hepatocyte

Not Specified Not Specified

Ambolet-Camoit, A., Bui, L.C., Pierre, S., Chevallier, A., Marchand, A., Coumoul, X., Garlatti, M., Andreau, K., Barouki, R., Aggerbeck, M., 2010. 2,3,7,8- tetrachlorodibenzo-p-dioxin counteracts the p53 response to a genotoxicant by upregulating expression of the metastasis marker agr2 in the hep- atocarcinoma cell line HepG2. Toxicol. Sci. 115, 501-512. Chopra, M., Dharmarajan, A.M., Meiss, G., Schrenk, D., 2009. Inhibition of UV-C light-induced apoptosis in liver cells by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicol. Sci. 111, 49-63. Chopra, M., Gahrs, M., Haben, M., Michels, C., Schrenk, D., 2010. Inhibition of apoptosis by 2,3,7,8-tetrachlorodibenzo-p-dioxin depends on protein biosyn- thesis. Cell Biol. Toxicol. 26, 391-401. Chopra, M., Schrenk, D., 2011. Dioxin toxicity, aryl hydrocarbon receptor signaling, and apoptosis-Persistent pollutants affect programmed cell death. Crit. Rev. Toxicol. 41:292-320. Davis W., J.W., Lauer, F.T., Burdick, A.D., Hudson, L.G., Burchiel, S.W., 2001. Prevention of apoptosis by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the MCF-10A cell line: correlation with increased transforming growth factor alpha production. Cancer. Res. 61, 3314-3320. Dragan, Y. P., Campbell, H. A., Xu, X. H., Pitot, H. C., 1997. Quantitative stereological studies of a 'selection' protocol of hepatocarcinogenesis following initiation in neonatal male and female rats. Carcinogenesis. 18, 149-58. Dragan, Y.P., Schrenk, D., 2000. Animal studies addressing the carcinogenicity of TCDD (or related compounds) with an emphasis on tumour promotion. Food. Addit. Contam. 17, 289-302. Franc, M.A., Moffat, I.D., Boutros, P.C., Tuomisto, J.T., Tuomisto, J., Pohjanvirta, R., Okey, A.B., 2008. Patterns of dioxin-altered mRNA expression in livers of dioxin- sensitive versus dioxin-resistant rats. Arch. Toxicol. 82, 809-830. Goodman, D.G., Sauer, R.M., 1992. Hepatotoxicity and carcinogenicity in female Sprague-Dawley rats treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD): a pathology working group reevaluation. Regul. Toxicol. Pharmacol. 15, 245-252. Hailey, J.R., Walker, N.J., Sells, D.M., Brix, A.E., Jokinen, M.P., Nyska, A., 2005. Clas- sification of proliferative hepatocellular lesions in harlan sprague-dawley rats chronically exposed to dioxin-like compounds. Toxicol. Pathol. 33, 165-174. Hendrich, S., Campbell, H. A., Pitot, H. C., 1987. Quantitative stereological evaluation of four histochemical markers of altered foci in multistage hepatocarcinogenesis in the rat. Carcinogenesis. 8, 1245-50. Kociba, R.J., Keyes, D.G., Beyer, J.E., Carreon, R.M., Wade, C.E., Dittenber, D.A., Kalnins, R.P., Frauson, L.E., Park, C.N., Barnard, S.D., Hummel, R.A., Humiston, C.G., 1978. Results of a two-year chronic toxicity and oncogenicity study of 2,3,7,8-tetrachlorodibenzo-p-dioxin in rats. Toxicol. Appl. Pharmacol. 46, 279-303. Luebeck, E.G., Buchmann, A., Stinchcombe, S., Moolgavkar, S.H., Schwarz, M., 2000. Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on initiation and promotion of GST-P-positive foci in rat liver: a quantitative analysis of experimental data using a stochastic model. Toxicol. Appl. Pharmacol. 167, 63-73. Moolgavkar, S.H., Luebeck, E.G., Buchmann, A., Bock, K.W., 1996. Quantitative analysis of enzyme-altered liver foci in rats initiated with diethylnitrosamine and promoted with 2,3,7,8-tetrachlorodibenzo-p-dioxin or 1,2,3,4,6,7,8- heptachlorodibenzo-p-dioxin. Toxicol. Appl. Pharmacol. 138, 31-42. Paajarvi, G., Viluksela, M., Pohjanvirta, R., Stenius, U., Hogberg, J., 2005. TCDD ac- tivates Mdm2 and attenuates the p53 response to DNA damaging agents. Carcinogenesis 26, 201-208. Roberts, R.A., Nebert, D.W., Hickman, J.A., Richburg, J.H., Goldsworthy, T.L., 1997. Perturbation of the mitosis/apoptosis balance: a fundamental mechanism in toxicology. Fundam. Appl. Toxicol. 38, 107-115. Schwarz, M., Buchmann, A., Stinchcombe, S., Kalkuhl, A., Bock, K., 2000. Ah receptor ligands and tumor promotion: survival of neoplastic cells. Toxicol. Lett. 112e113, 69-77. Stinchcombe, S., Buchmann, A., Bock, K.W., Schwarz, M., 1995. Inhibition of apoptosis during 2,3,7,8-tetrachlorodibenzo-p-dioxin-mediated tumour pro- motion in rat liver. Carcinogenesis 16, 1271-1275. Teeguarden, J.G., Dragan, Y.P., Singh, J., Vaughan, J., Xu, Y.H., Goldsworthy, T., Pitot, H.C., 1999. Quantitative analysis of dose- and time-dependent promotion of four phenotypes of altered hepatic foci by 2,3,7,8-tetrachlorodibenzo-p- dioxin in female Sprague-Dawley rats. Toxicol. Sci. 51, 211-223. Viluksela, M., Bager, Y., Tuomisto, J. T., Scheu, G., Unkila, M., Pohjanvirta, R., Flodstrom, S., Kosma, V. M., Maki-Paakkanen, J., Vartiainen, T., Klimm, C., Schramm, K. W., Warngard, L., Tuomisto, J., 2000. Liver tumor-promoting activity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in TCDD-sensitive and TCDD-resistant rat strains 27. Cancer Res. 60, 6911-6920. Worner, W., Schrenk, D., 1996. Influence of liver tumor promoters on apoptosis in rat hepatocytes induced by 2-acetylaminofluorene, ultraviolet light, or transforming growth factor beta 1. Cancer. Res. 56, 1272-1278. AOPWiki 2016-11-29T18:41:27

2017-09-16T10:15:01

Alterations, Cellular proliferation / hyperplasia

Tissue

Over the time period of high levels of sustained AHR activation, DLCs produce a complex pattern of cell proliferative responses. Teeguarden et al. (1999) observed that rats initiated with diethylnitrosamine (DEN) and then dosed with either 0.1 or 1 ng/ kg-d TCDD for one month exhibited a reduced labeling index relative to controls, and reduced BrdU-labeling was also observed following the 0.1 ng/kg-d TCDD dose after three months. Maronpot et al. (1993) observed a reduction in BrdU labeling index in hepatocytes at a low dose of 3.5 ng/kg-d TCDD in DEN-initiated rats after 30 weeks of TCDD administration but, at a dose of 125 ng/kg/d the labeling index was increased. Both parenchymal calls and liver stem cells are likely involved in the organ-level response to sustained AHR activation. Early acti- vation of the AHR in zone 3 of the liver acinus causes decreased hepatocyte replication and may act as an indirect proliferative stimulus for stem cells and hepatoblasts (Andersen et al., 1997; Conolly and Andersen, 1997; Tritscher et al., 1992). Oval cells in the periportal region likely function as a source of replacement of hepatocytes after inhibition of normal hepatocyte replication- replacement (Paku et al., 2001; Sahin et al., 2008; Tanaka et al., 2011; Wang et al., 2003). While hepatocyte replication is considered the normal means for replacement of liver parenchyma, inhibition of hepatocyte replication in centrilobular regions induced by TCDD may induce normally quiescent liver stem cells to proliferate. Following longer period of sustained AHR activation, organ- level increases in cell proliferation ensue, demonstrated by an in- crease in BrdU labeling and likely reflecting the regenerative response to organ-wide toxicity (Hailey et al., 2005). Non-parenchymal cells, including stem cells, hepatoblasts, biliary cells, stellate cells, endothelial cells, and Kuppfer cells, play a role in this AOP. In rodents, TCDD elicits a fibrogenic and bile duct proliferative response that requires pathological alteration of stellate cell function and increased differentiation and growth of hepatoblasts and bile duct cells before 33 weeks of exposure. Retinoid depletion induces stellate cell proliferation, production of extracellular matrix components, and the transition to fibroblast; stellate cells maintain vitamin A homeostasis and respond to liver injury with formation of proliferative cytokines such as TGF-a and EGF (Friedman, 2008; Pintilie et al., 2010; Senoo et al., 2010). TCDD induces loss of retinoid content (presumably from stellate cells) and may disrupt the extensive communication between various liver cell types (Fletcher et al., 2001; Hoegberg et al., 2005; Pierre et al., 2014; Schmidt et al., 2003). Thus, TCDD-induced retinol loss from hepatic stellate cells may contribute to cell proliferation, biliary fibrosis, and cholangiolarcarcinoma (Fattore et al., 2000; Friedman, 2008; Hakansson and Hanberg, 1989; Schmidt et al., 2003). AHR activation also induces changes in stem/oval cells. All of the rats receiving 100 ng/kg/day TCDD, the highest dose group animals in the 2-year cancer bioassay, developed oval cell hyperplasia with clear statistical increases in this endpoint at 22 ng/kg/day or greater (Hailey et al., 2005). Evidence points to the involvement of TNF-alpha regulation in the proliferative response of hepatic stem cells; this is likely mediated through modulation of the levels of TNF-alpha, altered beta-catenin signaling, and inhibition of cell-to-cell contact (Knight et al., 2000; Umannova et al., 2007; Vondracek et al., 2009; Dietrich et al., 2002; Prochazkova et al.,2011; Weiss et al., 2008). TNF-alpha is an inflammatory cytokine with an important role in liver tumor promotion. More research on how sustained AHR activation dysregulates normal TNF-alpha activity could be very impactful on evolving the AOP.

Bile duct hyperplasia and oval cell hyperplasia are measured histopathological observations using frequency of occurrence and a severity index.

The proliferative response of the liver appears to occur in rodents but not humans.

UBERON:0002107

UBERON liver

Not Specified

Andersen, M.E., Birnbaum, L.S., Barton, H.A., Eklund, C.R., 1997. Regional hepatic CYP1A1 and CYP1A2 induction with 2,3,7,8-tetrachlorodibenzo-p-dioxin eval- uated with a multicompartment geometric model of hepatic zonation. Toxicol. Appl. Pharmacol. 144, 145-155. Conolly, R.B., Andersen, M.E., 1997. Hepatic foci in rats after diethylnitrosamine initiation and 2,3,7,8-tetrachlorodibenzo-p-dioxin promotion: evaluation of a quantitative two-cell model and of CYP 1A1/1A2 as a dosimeter. Toxicol. Appl. Pharmacol. 146, 281-293. Dietrich, C., Faust, D., Budt, S., Moskwa, M., Kunz, A., Bock, K.W., Oesch, F., 2002. 2,3,7,8-tetrachlorodibenzo-p-dioxin-dependent release from contact inhibition in WB-F344 cells: involvement of cyclin A. Toxicol. Appl. Pharmacol. 183, 117-126. Fletcher, N., Hanberg, A., Håkansson, H., 2001. Hepatic vitamin a depletion is a sensitive marker of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure in four rodent species. Toxicol. Sci. 62, 166-175. Hailey, J.R., Walker, N.J., Sells, D.M., Brix, A.E., Jokinen, M.P., Nyska, A., 2005. Clas- sification of proliferative hepatocellular lesions in harlan sprague-dawley rats chronically exposed to dioxin-like compounds. Toxicol. Pathol. 33, 165-174. Hoegberg, P., Schmidt, C.K., Fletcher, N., Nilsson, C.B., Trossvik, C., Gerlienke Schuur, A., Brouwer, A., Nau, H., Ghyselinck, N.B., Chambon, P., Håkansson, H., 2005. Retinoid status and responsiveness to 2,3,7,8-tetrachlorodibenzo-p- dioxin (TCDD) in mice lacking retinoid binding protein or retinoid receptor forms. Chem. Biol. Interact. 156, 25-39. Knight, B., Yeoh, G.C., Husk, K.L., Ly, T., Abraham, L.J., Yu, C., Rhim, J.A., Fausto, N., 2000. Impaired preneoplastic changes and liver tumor formation in tumor necrosis factor receptor type 1 knockout mice. J. Exp. Med. 192, 1809-1818. Maronpot, R.R., Foley, J.F., Takahashi, K., Goldsworthy, T., Clark, G., Tritscher, A.,Portier, C., Lucier, G., 1993. Dose response for TCDD promotion of hep- atocarcinogenesis in rats initiated with DEN: histologic, biochemical, and cell proliferation endpoints 8. Environ. Heal. Perspect. 101, 634-642. Paku, S., Schnur, J., Nagy, P., Thorgeirsson, S.S., 2001. Origin and structural evolution of the early proliferating oval cells in rat liver. Am. J. Pathol. 158, 1313-1323. Pierre, S., Chevallier, A., Teixeira-Clerc, F., Ambolet-Camoit, A., Bui, L.-C., Bats, A.-S., Fournet, J.-C., Fernandez-Salguero, P., Aggerbeck, M., Lotersztajn, S., Barouki, R., Coumoul, X., 2014. Aryl hydrocarbon receptor-dependent induction of liver fibrosis by dioxin. Toxicol. Sci. 137, 114-124. Pintilie, D.G., Shupe, T.D., Oh, S.-H., Salganik, S.V., Darwiche, H., Petersen, B.E., 2010. Hepatic stellate cells' involvement in progenitor-mediated liver regeneration. Lab. Invest. 90, 1199-1208. Prochazkova, J., Kabatkova, M., Bryja, V., Umannova, L., Bernatík, O., Kozubík, A., Machala, M., Vondracek, J., 2011. The interplay of the aryl hydrocarbon receptor and b-catenin alters both AhR-dependent transcription and Wnt/b-catenin signaling in liver progenitors. Toxicol. Sci. 122, 349-360. Sahin, M.B., Schwartz, R.E., Buckley, S.M., Heremans, Y., Chase, L., Hu, W.-S., Verfaillie, C.M., 2008. Isolation and characterization of a novel population of progenitor cells from unmanipulated rat liver. Liver Transpl. 14, 333-345. Schmidt, C.K., Hoegberg, P., Fletcher, N., Nilsson, C.B., Trossvik, C., Hakansson, H., Nau, H., 2003. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) alters the endoge- nous metabolism of all-trans-retinoic acid in the rat. Arch. Toxicol. 77, 371-383. Senoo, H., Yoshikawa, K., Morii, M., Miura, M., Imai, K., Mezaki, Y., 2010. Hepatic stellate cell (vitamin A-storing cell) and its relativeepast, present and future. Cell. Biol. Int. 34, 1247-1272. Tanaka, M., Itoh, T., Tanimizu, N., Miyajima, A., 2011. Liver stem/progenitor cells: their characteristics and regulatory mechanisms. J. Biochem. 149, 231-239. Teeguarden, J.G., Dragan, Y.P., Singh, J., Vaughan, J., Xu, Y.H., Goldsworthy, T., Pitot, H.C., 1999. Quantitative analysis of dose- and time-dependent promotion of four phenotypes of altered hepatic foci by 2,3,7,8-tetrachlorodibenzo-p- dioxin in female Sprague-Dawley rats. Toxicol. Sci. 51, 211-223. Tritscher, A.M., Goldstein, J.A., Portier, C.J., McCoy, Z., Clark, G.C., Lucier, G.W., 1992. Dose-response relationships for chronic exposure to 2,3,7,8- tetrachlorodibenzo-p-dioxin in a rat tumor promotion model: quantification and immunolocalization of CYP1A1 and CYP1A2 in the liver. Cancer. Res. 52, 3436-3442. Umannova, L., Zatloukalova, J., Machala, M., Krcmar, P., Majkova, Z., Hennig, B., Kozubík, A., Vondracek, J., 2007. Tumor necrosis factor-alpha modulates effects of aryl hydrocarbon receptor ligands on cell proliferation and expression of cytochrome P450 enzymes in rat liver “stem-like” cells. Toxicol. Sci. 99, 79-89. Vondracek, J., Krcmar, P., Prochazkova, J., Trilecova, L., Gavelova, M., Skalova, L., Szotakova, B., Buncek, M., Radilova, H., Kozubik, A., Machala, M., 2009. The role of aryl hydrocarbon receptor in regulation of enzymes involved in metabolic activation of polycyclic aromatic hydrocarbons in a model of rat liver progenitor cells. Chem. Biol. Interact. 180, 226-237. Wang, X., Foster, M., Al-Dhalimy, M., Lagasse, E., Finegold, M., Grompe, M., 2003. The origin and liver repopulating capacity of murine oval cells. Proc. Natl. Acad. Sci. U. S. A. 100 (Suppl. 1), 11881-11888. Weiss, C., Faust, D., Schreck, I., Ruff, A., Farwerck, T., Melenberg, A., Schneider, S., Oesch-Bartlomowicz, B., Zatloukalova, J., Vondracek, J., Oesch, F., Dietrich, C., 2008. TCDD deregulates contact inhibition in rat liver oval cells via Ah receptor, JunD and cyclin A. Oncogene 27, 2198-2207. AOPWiki 2016-11-29T18:41:27

2017-09-16T10:15:02

Formation, Hepatocellular and Bile duct tumors

Organ

If AHR activation is sustained for a period of more than 30 weeks or 30% of a rat's lifespan, hepatocellular adenomas/carcinomas and cholangiocarcinomas develop.. These two tumors are also part of the organ-level response and are the adverse outcome. Adenomas may arise from altered hepatic foci that are derived from hepato- cytes or hepatoblasts whereas hepatocellular carcinomas and cholangiocarcinomas likely arise from initiated stem cells. How- ever, the actual cellular origin of the various liver tumor types is not known with certainty and involvement of both liver stem cells and hepatocyte-like cells have been observed in hepatocellular adenomas (Libbrecht et al., 2001; Libbrecht, 2006).

Histopathological examination is a necessary part of lifetime cancer bioassays. This type of examination is used to detect tumors.

Overall, empirical evidence supporting the applicability of this AOP to humans is absent. Only a single KE has been observed in humans with respect to the binding and activation of the AHR by DLCs with accompanying hepatic CYP1A induction (Abraham et al., 2002; Budinsky et al., 2010; Coenraads et al., 1999; Lambert et al., 2006; Tang et al., 2008). For perspective purposes regarding an unequivocal dioxin effect occurring in highly exposed human subjects, high levels of AHR activation in humans alter the growth and differentiation of keratinocytes and produce chloracne (Forrester et al., 2014; Geusau et al., 2001; Ju et al., 2011; Moses and Prioleau, 1985; Saurat and Sorg, 2010; Saurat et al., 2012; Sorg, 2014; Sutter et al., 2009, 2010, 2011). In contrast, the epidemiological evidence for TCDD- associated liver cancer in humans is negative or equivocal (Akintobi et al., 2007; Bertazzi et al., 1989; Du et al., 2006; Geusau et al., 2005; Hankinson, 2009; Loertscher et al., 2001). Trichlorophenol workers exposed to TCDD show no increase in liver or biliary cancer (Collins et al., 2009; McBride et al., 2009). The occurrence of chloracne indicates high levels of exposure to DLCs and significant AHR activation; even in such cases, no evidence of liver injury or cancer has been reported (Ghezzi et al., 1982; Mocarelli et al., 1986, 1991; Pocchiari et al., 1979; Reggiani, 1980). In other trichlorophenol workers, transient changes in liver enzyme levels were reported in alcohol consumers only (Calvert et al., 1992). The unique Yusho and Yucheng rice oil poisonings are confounded by exposures to a mixture of complex PCBs, polychlorinated dibenzofurans, and mixtures of quarterphenyl, and terphenyl compounds. Clearly, these compounds possess dioxin- like properties and the mixture was sufficiently potent to induce a chloracne-like condition in some individuals (Lambert et al., 2006). An increase in mortality from cirrhosis and chronic liver disease has been observed among the victims of the Yusho poisoning incident, whereas liver cancer was not elevated (Onozuka et al., 2009). The rate of mortality from chronic liver disease was increased in men only among the victims of the similar Yucheng poisoning incident without excess liver cancer in either sex (Tsai et al., 2007). In contrast to humans, rodents are highly susceptible to the hepatotoxic, proliferative, and carcinogenic effects of TCDD (Hailey et al., 2005; Goodman and Sauer, 1992; Kociba et al., 1978). To summarize, the sustained AHR activation rodent liver tumor promotion AOP appears to be a pathway that very likely requires exceedance of a threshold for sustained AHR activation for liver cancers to occur in rodents (e.g. Fig. 4). In humans, increases in liver cancer have not been observed even in highly exposed populations, and no population level data in humans are available showing an increased liver cancer response, even in individuals with chloracne and obvious high exposure to DLCs. However, as is often the case for evaluations of chemicals which lead to tumor formation in laboratory animals, for regulatory purposes, assumptions are made that potential risks to human health can be estimated from animal studies. In many cases where there is sparse data, a default linear no-threshold extrapolation method is used. However, in applying this AOP for such an assessment, the extensive body of scientific evidence clearly indicates that liver tumor promotion by DLCs only occurs after a threshold level of sustained AHR activation is exceeded. These thresholds also become apparent in Figs. 3B and 4. Therefore, a quantitative application to derive an exposure guidance value for humans to address the potential for tumor promotion by DLCs should be based on a threshold mode of action (e.g., Simon et al., 2009). To the extent humans have been inadvertently, accidentally, or intentionally exposed to TCDD, no evidence of increased liver cancer or even liver injury have been observed, consistent with rats being more sensitive than humans. Given that tumorigenic responses in rodents only occur when AHR activation is sustained for a period approximating 30% of the life- span, and the steep slopes corresponding to responses elicited when this apparent threshold of AHR activation is exceeded, risk assessments for humans using this AOP should employ a threshold model. As noted above, binding to the AHR is insufficient to infer activity leading to the adverse outcome of liver tumors. Moreover, there is considerable scientific debate as to whether the rat liver tumori- genic responses induced by TCDD are relevant endpoints for human health. WHO indicates that cancer may not be the most sensitive response in either humans or animals and EPA's latest assessment is based on non-cancer effects in humans (sperm deficits among young males exposed between the ages of 1e9 and increased TSH levels in 72-hour neonates born of Seveso mothers with elevated serum TCDD concentrations). Nonetheless, the utility of the AOP is the identification and ordering of effects, demonstration of dose-response concordance and illustrating that rodent liver tumor promotion by sustained AHR is a threshold phenomenon.

UBERON:0002423

UBERON hepatobiliary system

Not Specified

2017-09-16T10:15:02

<upstream-id>2a414f09-3780-48e5-8337-aecfc3a128bc</upstream-id> <downstream-id>75aac401-7b65-462e-b716-964ecc9d1195</downstream-id> Sustained AHR activation inhibits apoptosis in altered hepatic foci (i.e., initiated hepatic cells), and this inhibition affords cells within altered hepatic foci a survival advantage and increases the likelihood that these cells will acquire additional mutations.

The weight of evidence descriptor for this KER is strong.

All the elements in this AOP are strongly associated with the biological steps and elements of carcinogenesis (Hanahan and Weinberg, 2011). First, there is extensive body of mechanistic evidence in support the biological plausibility of this MOA (see recent review by Budinsky et al., 2014). Further, the relationships between sustained AHR activation and changes in cellular growth homeostasis / apoptosis, has been used for many years in initiation-promotion studies to understand early events in tumor formation (Dragan et al. 1992; Dragan and Schrenk, 2000; Luebeck et al. 2000; Maronpot et al. 1993; Teeguarden et al. 1999).

AHR activation appears to cause inhibition of apoptosis in altered hepatic foci (Luebeck et al., 2000; Paajarvi et al., 2005; Schrenk et al., 1994, 2004; Stinchcombe et al., 1995). For this KE, initiation-promotion studies provide indirect evidence of inhibition of intrafocal apoptosis due to sustained AHR activation and direct evidence of a threshold for the clonal expansion of altered hepatic foci (Dragan and Schrenk, 2000; Teeguarden et al., 1999). Also, changes in cellular growth homeostasis / apoptosis are measured by changes in AHF reflecting changes in the apoptosis/proliferation balance occurs earliest in dose and time, reflected by the measure of SAA shown in the Dose-Time concordance table on the main page of this AOP.

There are few, if any, uncertainties or inconsistencies regarding this KER.

Using the measure of SAA that incorporates both dose and time allows understanding of the quantitative relationship between the sustained AHR activation and changes in cellular growth homeostasis / apoptosis. Figure 4 shows a plot of the increase in volume fraction of ATPase-negative AHF versus sustained AHR activation. The Hill model fit to these data had an ESA50 value of 17.6. <div class="thumb tright"><div class="thumbinner" style="width:182px;"><a href="/wiki/index.php/File:Figure_2-AHR-AOP-ppr.jpg" class="image"><img alt="Figure 4 alt text" src="/wiki/images/thumb/e/e1/Figure_2-AHR-AOP-ppr.jpg/180px-Figure_2-AHR-AOP-ppr.jpg" width="180" height="76" class="thumbimage" srcset="/wiki/images/thumb/e/e1/Figure_2-AHR-AOP-ppr.jpg/270px-Figure_2-AHR-AOP-ppr.jpg 1.5x, /wiki/images/thumb/e/e1/Figure_2-AHR-AOP-ppr.jpg/360px-Figure_2-AHR-AOP-ppr.jpg 2x" /></a> <div class="thumbcaption"><div class="magnify"><a href="/wiki/index.php/File:Figure_2-AHR-AOP-ppr.jpg" class="internal" title="Enlarge"><img src="/wiki/skins/common/images/magnify-clip.png" width="15" height="11" alt="" /></a></div>Figure 4: Rat liver tumor promotion by persistent AHR ligands. A) Simplified diagram showing changes in both cell proliferation and apoptosis appear to promote tumors. B) Plot of the increase in volume fraction of ATPase-deficient hepatic foci versus sustained AHR activation (SAA) index. Data from Teeguarden et al. (1999).</div></div></div>

#<Reference::ActiveRecord_Associations_CollectionProxy:0x00005eb8a0d76128> AOPWiki 2016-11-29T18:41:35

2016-11-29T20:41:21

<upstream-id>2a414f09-3780-48e5-8337-aecfc3a128bc</upstream-id> <downstream-id>aa51ef0d-fcd7-4f27-9e87-6230fb555362</downstream-id> It is not exactly known just how sustained AHR activation leads to hepatotoxicity. Nonetheless, the constellation of different histopathological alterations included in toxic hepatopathy are highly associated with tumor formation (Simon et al. 2009).

The quantitative relationship discussed in the sustained AHR activation (MIE) page and also presented below in common to dioxin-like chemicals (NTP, 2006a, 2006b, 2006c, 2006d, 2006e, 2006f). In addition, rats fed indole-3-carbinol for eight weeks in an initiation-promotion medium term assay showed the development of oxidative stress, likely due to induction of CYP1A and other phase I enzymes. The development of AHF, here noted as KE#1 was also enhanced (Shimamoto et al. 2011).

Many two-year bioassays of dioxin-like chemicals showed both sustained AHR activation measured by CYP induction as well as toxic hepatopathy (NTP, 2006a, 2006b, 2006c, 2006d, 2006e, 2006f).

While the exact mechanism of how sustained AHR activation leads to toxic hepatopathy, a large number of observations lend certainty to the relationship.

<div class="thumb tleft"><div class="thumbinner" style="width:182px;"><a href="/wiki/index.php/File:Figure_4-AHR-AOP-ppr.jpg" class="image"><img alt="Figure 4 alt text" src="/wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/180px-Figure_4-AHR-AOP-ppr.jpg" width="180" height="208" class="thumbimage" srcset="/wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/270px-Figure_4-AHR-AOP-ppr.jpg 1.5x, /wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/360px-Figure_4-AHR-AOP-ppr.jpg 2x" /></a> <div class="thumbcaption"><div class="magnify"><a href="/wiki/index.php/File:Figure_4-AHR-AOP-ppr.jpg" class="internal" title="Enlarge"><img src="/wiki/skins/common/images/magnify-clip.png" width="15" height="11" alt="" /></a></div>Figure 4: Indirect key event relationships. Sustained AHR activation was identified as the MIE and determined as the product of fractional AHR activation measured by CYP1A1 induction and time in weeks. The frequency of histopathology and tumors observed in rats in the NTP bioassays was plotted against sustained AHR activation. A) Relationships between the MIE and aspects of KE2 (hepatopathy and hyperplasia; all response shown as filled circles). B) Relationships between the MIE and the two tumor types representing the adverse outcome (AO) (responses shown as an X). The Hill coefficients, BMD21 and TDV21 values are shown in the table below. The values shown as open circles in the two plots in B show tumor responses in animals in the stop-exposure group that were exposed to the maximum DLC concentration for 31 weeks (see text for discussion). In each plot in B, three values, one each for TCDD, 4-PeCDF and PCB-126, are shown for the stop-exposure experiments and only two are not obscured by the other responses. The numerical ESA50 values are the half-maximal level of sustained AHR activation similar to an EC50 and described in the text.</div></div></div> The quantitative KER linking the sustained AHR activation to Hepatotoxicity/Hepatopathy is shown in Fig. 4 at the left. Indirect key event relationships. Sustained AHR activation was identified as the MIE and determined as the product of fractional AHR activation measured by CYP1A1 induction and time in weeks. The frequency of histopathology and tumors observed in rats in the NTP bioassays was plotted against sustained AHR activation. A) Relationships between the MIE and aspects of Hepatotoxicity/Hepatopathy and Cellular Proliferation / Hyperplasia; all response shown as filled circles). B) Relationships between the MIE and the two tumor types representing the adverse outcome (AO) (responses shown as an X). The Hill coefficients, BMD21 and TDV21 values are shown in the table below. The values shown as open circles in the two plots in B show tumor responses in animals in the stop-exposure group that were exposed to the maximum DLC concentration for 31 weeks. In each plot in B, three values, one each for TCDD, 4-PeCDF and PCB-126, are shown for the stop-exposure experiments and only two are not obscured by the other responses. The numerical ESA50 values are the half-maximal level of sustained AHR activation similar to an EC50 and described in the text.Using methods described in Simon et al. (2014), the transitional dose values of the sustained AHR activation index based on a 21% response level are shown in the table below Here, the transitional dose value is the projection from the 21% response level to the background response level using the slope of the dose-response at this same 21% response level (Sand et al., 2006; Simon et al., 2014). While not definitive thresholds, transitional dose values based on the 21% response level can be used as an approximation of a threshold. As noted, toxic hepatopathy includes a constellation of effects and the transitional dose value has the lowest value of the three effects representing the Hepatotoxicity/Hepatopathy and the Cellular Proliferation / Hyperplasia. Hill Model parameters and Transitional Dose Values (TDV) for Hepatotoxicity/Hepatopathy, Cellular Proliferation / Hyperplasia and the occurrence of Hepatocellular and Bile Duct Tumors based on the Quantitative Measure of the MIE or Sustained AHR Activation in ppb-weeks <table border="1"> <tr> <td> Endpoint </td> <td> Hill Coeff. </td> <td> Kd in SAA units </td> <td> BMD21 in SAA units </td> <td> TDV21 in SAA units </td></tr> <tr> <td colspan="5" align="center"> KE#2 </td></tr> <tr> <td align="center"> Toxic Hepatopathy </td> <td align="center"> 2.139 </td> <td align="center"> 61.92 </td> <td align="center"> 33.2 </td> <td align="center"> 29.2 </td></tr> </table> ESA50 (Fig. 4) is similar to an EC50 - it is the effective level of sustained AHR activation necessary to achieve a 50% response. As described, the level of sustained AHR activation can be calculated by multiplying the fractional level of AHR activation measured by CYP1A1 induction by the number of weeks of dosing. Bile duct hyperplasia occurs at a higher level of sustained AHR activation and oval cell hyperplasia at an even higher level (Fig. 5A, middle and bottom plots). The dose-response for oval cell hyperplasia is much steeper than the other two histopathological effects that comprise KE#3. Although speculative, this higher level of sustained AHR activation needed for bile duct hyperplasia may be the reason why Kociba et al. (1978) failed to observe bile duct tumors whereas they were observed in NTP (2006a). Possibly, the distinction between the two studies may be that dosage regimen (diet vs. gavage) or changes in the Sprague-Dawley strain over time. In the rats used in Kociba et al. (1978), the degree of sustained AHR activation needed for promotion of bile duct tumors may not have been achieved.

The relationship between Hepatoxicity/Hepatopathy and downstream KEs of Cellular Proliferation / Hyperplasia and Hepatocellular and Bile Duct Tumors does not appear to occur in humans; however, a comprehensive assessment of this KER in humans has not been conducted in a fashion appropriate for this AOP.

#<Reference::ActiveRecord_Associations_CollectionProxy:0x00005eb8a0770af8> AOPWiki 2016-11-29T18:41:33

2016-11-29T19:58:17

<upstream-id>2a414f09-3780-48e5-8337-aecfc3a128bc</upstream-id> <downstream-id>96acf976-c1e9-4ca9-af06-697bed1db181</downstream-id> Both the development of altered hepatic foci, which are preneoplastic lesions, and the highly proliferative environment induced in the liver by Hepatoxicity/Hepatopathy induce cell proliferation. Replacement of liver cells normally occurs by hepatocyte replication (Paku et al, 2001). However, Hepatoxicity/Hepatopathy may produce sufficient liver damage that stem cell replication also contributes to hepatocyte replacement (Alison, 2005). In general, increased stem cell replication appears to be associated with tumor formation (Tomasetti and Vogelstein, 2015).

The quantitative relationship discussed in the Sustained AHR Activation (MIE) page and also presented on the KER page for Sustained AHR Activation --> Hepatotoxicity/Hepatopathy is common to dioxin-like chemicals (NTP, 2006a, 2006b, 2006c, 2006d, 2006e, 2006f). In addition, rats fed indole-3-carbinol for eight weeks in an initiation-promotion medium term assay showed the development of oxidative stress, likely due to induction of CYP1A and other phase I enzymes. The development of AHF, here noted as Changes in Cellular Growth Homeostasis / Apoptosis was also enhanced (Shimamoto et al. 2011).

While the exact mechanism of how sustained AHR activation leads to a strong hepato-proliferative response, a large number of observations lend certainty to the relationship.

<div class="thumb tleft"><div class="thumbinner" style="width:182px;"><a href="/wiki/index.php/File:Figure_4-AHR-AOP-ppr.jpg" class="image"><img alt="Figure 4 alt text" src="/wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/180px-Figure_4-AHR-AOP-ppr.jpg" width="180" height="208" class="thumbimage" srcset="/wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/270px-Figure_4-AHR-AOP-ppr.jpg 1.5x, /wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/360px-Figure_4-AHR-AOP-ppr.jpg 2x" /></a> <div class="thumbcaption"><div class="magnify"><a href="/wiki/index.php/File:Figure_4-AHR-AOP-ppr.jpg" class="internal" title="Enlarge"><img src="/wiki/skins/common/images/magnify-clip.png" width="15" height="11" alt="" /></a></div>Figure 4: Indirect key event relationships. Sustained AHR activation was identified as the MIE and determined as the product of fractional AHR activation measured by CYP1A1 induction and time in weeks. The frequency of histopathology and tumors observed in rats in the NTP bioassays was plotted against sustained AHR activation. A) Relationships between the MIE and aspects of KE2 (hepatopathy and hyperplasia; all response shown as filled circles). B) Relationships between the MIE and the two tumor types representing the adverse outcome (AO) (responses shown as an X). The Hill coefficients, BMD21 and TDV21 values are shown in the table below. The values shown as open circles in the two plots in B show tumor responses in animals in the stop-exposure group that were exposed to the maximum DLC concentration for 31 weeks (see text for discussion). In each plot in B, three values, one each for TCDD, 4-PeCDF and PCB-126, are shown for the stop-exposure experiments and only two are not obscured by the other responses. The numerical ESA50 values are the half-maximal level of sustained AHR activation similar to an EC50 and described in the text.</div></div></div> The quantitative KER linking the sustained AHR activation to Hepatotoxicity/Hepatopathy is shown in Fig. 4 at the left. Indirect key event relationships. Sustained AHR activation was identified as the MIE and determined as the product of fractional AHR activation measured by CYP1A1 induction and time in weeks. The frequency of histopathology and tumors observed in rats in the NTP bioassays was plotted against sustained AHR activation. A) Relationships between the MIE and aspects of Hepatotoxicity/Hepatopathy and Cellular Proliferation / Hyperplasia; all response shown as filled circles). B) Relationships between the MIE and the two tumor types representing the adverse outcome (AO) (responses shown as an X). The Hill coefficients, BMD21 and TDV21 values are shown in the table below. The values shown as open circles in the two plots in B show tumor responses in animals in the stop-exposure group that were exposed to the maximum DLC concentration for 31 weeks. In each plot in B, three values, one each for TCDD, 4-PeCDF and PCB-126, are shown for the stop-exposure experiments and only two are not obscured by the other responses. The numerical ESA50 values are the half-maximal level of sustained AHR activation similar to an EC50 and described in the text.Using methods described in Simon et al. (2014), the transitional dose values of the sustained AHR activation index based on a 21% response level are shown in the table below Here, the transitional dose value is the projection from the 21% response level to the background response level using the slope of the dose-response at this same 21% response level (Sand et al., 2006; Simon et al., 2014). While not definitive thresholds, transitional dose values based on the 21% response level can be used as an approximation of a threshold. As noted, toxic hepatopathy includes a constellation of effects and the transitional dose value has the lowest value of the three effects representing the Hepatotoxicity/Hepatopathy and the Cellular Proliferation / Hyperplasia. Hill Model parameters and Transitional Dose Values (TDV) for Hepatotoxicity/Hepatopathy, Cellular Proliferation / Hyperplasia and the occurrence of Hepatocellular and Bile Duct Tumors based on the Quantitative Measure of the MIE or Sustained AHR Activation in ppb-weeks <table border="1"> <tr> <td> Endpoint </td> <td> Hill Coeff. </td> <td> Kd in SAA units </td> <td> BMD21 in SAA units </td> <td> TDV21 in SAA units </td></tr> <tr> <td colspan="5" align="center"> KE#3 </td></tr> <tr> <td align="center"> Bile duct hyperplasia </td> <td align="center"> 2.28 </td> <td align="center"> 87.05 </td> <td align="center"> 48.7 </td> <td align="center"> 43.0 </td></tr> <tr> <td align="center"> Oval cell hyperplasia </td> <td align="center"> 7.44 </td> <td align="center"> 91.2 </td> <td align="center"> 76.3 </td> <td align="center"> 73.6 </td></tr> </table> ESA50 (Fig. 4) is similar to an EC50 - it is the effective level of sustained AHR activation necessary to achieve a 50% response. As described, the level of sustained AHR activation can be calculated by multiplying the fractional level of AHR activation measured by CYP1A1 induction by the number of weeks of dosing. Bile duct hyperplasia occurs at a higher level of sustained AHR activation and oval cell hyperplasia at an even higher level (Fig. 5A, middle and bottom plots). The dose-response for oval cell hyperplasia is much steeper than the other two histopathological effects that comprise KE#3. Although speculative, this higher level of sustained AHR activation needed for bile duct hyperplasia may be the reason why Kociba et al. (1978) failed to observe bile duct tumors whereas they were observed in NTP (2006a). Possibly, the distinction between the two studies may be that dosage regimen (diet vs. gavage) or changes in the Sprague-Dawley strain over time. In the rats used in Kociba et al. (1978), the degree of sustained AHR activation needed for promotion of bile duct tumors may not have been achieved.

This relationship occurs in rodents and possibly other animals but not in humans.

#<Reference::ActiveRecord_Associations_CollectionProxy:0x00005eb8a0c8de78> AOPWiki 2016-11-29T18:41:35

2016-11-29T20:41:26

<upstream-id>2a414f09-3780-48e5-8337-aecfc3a128bc</upstream-id> <downstream-id>387be3d0-8933-48dc-b673-faa0ecee7669</downstream-id> Replacement of liver cells normally occurs by hepatocyte replication (Paku et al, 2001). However, KE#2 may produce sufficient liver damage that stem cell replication also contributes to hepatocyte replacement (Alison, 2005). KE#3, hyperplasia/proliferation likely includes both hepatocytes and liver stem cells. In humans, liver damage from a range of stressors (e.g., alcohol consumption) is relation to tumor development and, in general, increased stem cell replication appears to be associated with tumor formation (Tomasetti and Vogelstein, 2015).

The quantitative relationship discussed in the MIE page and also presented on the KER page for MIE --> KE#2 is common to dioxin-like chemicals (NTP, 2006a, 2006b, 2006c, 2006d, 2006e, 2006f). Note the two plots on the left of the figure on the KER page for MIE --> KE#2 show the relationship between the MIE and the AO. Liver tumors in rodents occur with sustained activation of the AHR (NTP, 2006a, 2006b, 2006c, 2006d, 2006e, 2006f; Shimamoto et al. 2011).

Many two-year bioassays of dioxin-like chemicals showed both sustained AHR activation measured by CYP induction as well as an extensive proliferative/regenerative response in the liver (NTP, 2006a, 2006b, 2006c, 2006d, 2006e, 2006f). The AO of liver tumors occurs toward the end of the lifespan and requires an MIE level of around 100 pbb-weeks.

A large body of evidence cited throughout this AOP supports the relationship between the MIE and AO.

<div class="thumb tleft"><div class="thumbinner" style="width:182px;"><a href="/wiki/index.php/File:Figure_4-AHR-AOP-ppr.jpg" class="image"><img alt="Figure 4 alt text" src="/wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/180px-Figure_4-AHR-AOP-ppr.jpg" width="180" height="208" class="thumbimage" srcset="/wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/270px-Figure_4-AHR-AOP-ppr.jpg 1.5x, /wiki/images/thumb/e/eb/Figure_4-AHR-AOP-ppr.jpg/360px-Figure_4-AHR-AOP-ppr.jpg 2x" /></a> <div class="thumbcaption"><div class="magnify"><a href="/wiki/index.php/File:Figure_4-AHR-AOP-ppr.jpg" class="internal" title="Enlarge"><img src="/wiki/skins/common/images/magnify-clip.png" width="15" height="11" alt="" /></a></div>Figure 4: Indirect key event relationships. Sustained AHR activation was identified as the MIE and determined as the product of fractional AHR activation measured by CYP1A1 induction and time in weeks. The frequency of histopathology and tumors observed in rats in the NTP bioassays was plotted against sustained AHR activation. A) Relationships between the MIE and aspects of KE2 (hepatopathy and hyperplasia; all response shown as filled circles). B) Relationships between the MIE and the two tumor types representing the adverse outcome (AO) (responses shown as an X). The Hill coefficients, BMD21 and TDV21 values are shown in the table below. The values shown as open circles in the two plots in B show tumor responses in animals in the stop-exposure group that were exposed to the maximum DLC concentration for 31 weeks (see text for discussion). In each plot in B, three values, one each for TCDD, 4-PeCDF and PCB-126, are shown for the stop-exposure experiments and only two are not obscured by the other responses. The numerical ESA50 values are the half-maximal level of sustained AHR activation similar to an EC50 and described in the text.</div></div></div> The quantitative KER linking the sustained AHR activation to Hepatotoxicity/Hepatopathy is shown in Fig. 4 at the left. Indirect key event relationships. Sustained AHR activation was identified as the MIE and determined as the product of fractional AHR activation measured by CYP1A1 induction and time in weeks. The frequency of histopathology and tumors observed in rats in the NTP bioassays was plotted against sustained AHR activation. A) Relationships between the MIE and aspects of Hepatotoxicity/Hepatopathy and Cellular Proliferation / Hyperplasia; all response shown as filled circles). B) Relationships between the MIE and the two tumor types representing the adverse outcome (AO) (responses shown as an X). The Hill coefficients, BMD21 and TDV21 values are shown in the table below. The values shown as open circles in the two plots in B show tumor responses in animals in the stop-exposure group that were exposed to the maximum DLC concentration for 31 weeks. In each plot in B, three values, one each for TCDD, 4-PeCDF and PCB-126, are shown for the stop-exposure experiments and only two are not obscured by the other responses. The numerical ESA50 values are the half-maximal level of sustained AHR activation similar to an EC50 and described in the text.Using methods described in Simon et al. (2014), the transitional dose values of the sustained AHR activation index based on a 21% response level are shown in the table below Here, the transitional dose value is the projection from the 21% response level to the background response level using the slope of the dose-response at this same 21% response level (Sand et al., 2006; Simon et al., 2014). While not definitive thresholds, transitional dose values based on the 21% response level can be used as an approximation of a threshold. As noted, toxic hepatopathy includes a constellation of effects and the transitional dose value has the lowest value of the three effects representing the Hepatotoxicity/Hepatopathy and the Cellular Proliferation / Hyperplasia. Hill Model parameters and Transitional Dose Values (TDV) for Hepatotoxicity/Hepatopathy, Cellular Proliferation / Hyperplasia and the occurrence of Hepatocellular and Bile Duct Tumors based on the Quantitative Measure of the MIE or Sustained AHR Activation in ppb-weeks <table border="1"> <tr> <td> Endpoint </td> <td> Hill Coeff. </td> <td> Kd in SAA units </td> <td> BMD21 in SAA units </td> <td> TDV21 in SAA units </td></tr> <tr> <td colspan="5" align="center"> AO </td></tr> <tr> <td align="center"> Cholangiocarcinoma </td> <td align="center"> 13.48 </td> <td align="center"> 106.4 </td> <td align="center"> 96.4 </td> <td align="center"> 94.5 </td></tr> <tr> <td align="center"> Hepatoellular Adenoma </td> <td align="center"> 7.44 </td> <td align="center"> 128.2 </td> <td align="center"> 107.3 </td> <td align="center"> 103.4 </td></tr></table> ESA50 (Fig. 4) is similar to an EC50 - it is the effective level of sustained AHR activation necessary to achieve a 50% response. As described, the level of sustained AHR activation can be calculated by multiplying the fractional level of AHR activation measured by CYP1A1 induction by the number of weeks of dosing. Bile duct hyperplasia occurs at a higher level of sustained AHR activation and oval cell hyperplasia at an even higher level (Fig. 5A, middle and bottom plots). The dose-response for oval cell hyperplasia is much steeper than the other two histopathological effects that comprise KE#3. Although speculative, this higher level of sustained AHR activation needed for bile duct hyperplasia may be the reason why Kociba et al. (1978) failed to observe bile duct tumors whereas they were observed in NTP (2006a). Possibly, the distinction between the two studies may be that dosage regimen (diet vs. gavage) or changes in the Sprague-Dawley strain over time. In the rats used in Kociba et al. (1978), the degree of sustained AHR activation needed for promotion of bile duct tumors may not have been achieved.

Not Specified

This relationship occurs in rodents and possibly other animals but not in humans.

#<Reference::ActiveRecord_Associations_CollectionProxy:0x00005eb8a11e76d0> AOPWiki 2016-11-29T18:41:35

2016-11-29T20:41:31

<upstream-id>75aac401-7b65-462e-b716-964ecc9d1195</upstream-id> <downstream-id>aa51ef0d-fcd7-4f27-9e87-6230fb555362</downstream-id>

#<Reference::ActiveRecord_Associations_CollectionProxy:0x00005eb8a19c78a0> AOPWiki 2016-11-29T18:41:36

2016-12-02T11:47:53

<upstream-id>aa51ef0d-fcd7-4f27-9e87-6230fb555362</upstream-id> <downstream-id>96acf976-c1e9-4ca9-af06-697bed1db181</downstream-id>

#<Reference::ActiveRecord_Associations_CollectionProxy:0x00005eb8a19fb100> AOPWiki 2016-11-29T18:41:36

2016-12-02T11:49:01

<upstream-id>96acf976-c1e9-4ca9-af06-697bed1db181</upstream-id> <downstream-id>387be3d0-8933-48dc-b673-faa0ecee7669</downstream-id>

#<Reference::ActiveRecord_Associations_CollectionProxy:0x00005eb8a1a43838> AOPWiki 2016-11-29T18:41:36

2016-12-02T11:49:53

Sustained AhR Activation leading to Rodent Liver Tumours

Allie Always

Richard A Becker, American Chemical Council (ACC) on behalf of the Business Industry Advisory Committee (BIAC) email:Rick_Becker@americanchemistry.com Contributing authors to the development of this AOP are: Ted Simon (Ted Simon LLC), Robert Budinsky, (The Dow Chemical Company), Grace Patlewicz, (DuPont), Craig Rowlands, (The Dow Chemical Company).

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EAGMST Under Review

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An Adverse Outcome Pathway (AOP) represents the existing knowledge of a biological pathway leading from initial molecular interactions of a toxicant and progressing through a series of key events (KEs), culminating with an apical adverse outcome (AO) that has to be of regulatory relevance. An AOP based on the mode of action (MOA) of rodent liver tumor promotion by dioxin-like compounds (DLCs) has been developed and the weight of evidence (WoE) of key event relationships (KERs) evaluated using evolved Bradford Hill considerations. Dioxins and DLCs are potent aryl hydrocarbon receptor (AHR) ligands that cause a range of species-specific adverse outcomes. The occurrence of KEs is necessary for inducing downstream biological responses and KEs may occur at the molecular, cellular, tissue and organ levels. The common convention is that an AOP begins with the toxicant interaction with a biological response element; for this AOP, this initial event is binding of a DLC ligand to the AHR. Data from mechanistic studies, lifetime bioassays and approximately thirty initiation-promotion studies have established a number of substances, including dioxin-like chemicals and indole-3-carbinol from brassica vegetables, as rat liver tumor promoters. Such studies clearly show that sustained AHR activation, weeks or months in duration, is necessary to induce rodent liver tumor promotion; hence, sustained AHR activation is deemed the molecular initiating event (MIE). After this MIE, subsequent KEs are 1) changes in cellular growth homeostasis likely associated with expression changes in a number of genes and observed as development of hepatic foci and decreases in apoptosis within foci; 2) extensive liver toxicity observed as the constellation of effects called toxic hepatopathy; 3) cellular proliferation and hyperplasia in several hepatic cell types. This progression of KEs culminates in the AO, the development of hepatocellular adenomas and carcinomas and cholangiolar carcinomas. A rich data set provides both qualitative and quantitative knowledge of the progression of this AOP through KEs and the KERs. Thus, the WoE for this AOP is judged to be strong. Species-specific effects of dioxins and DLCs are well known -- humans are less responsive than rodents and rodent species differ in sensitivity between strains. Consequently, application of this AOP to evaluate potential human health risks must take these differences into account. Please also see Becker, R.A., Patlewicz, G., Simon, T.W., Rowlands, J.C., Budinsky, R.A. 2015. The adverse outcome pathway for rodent liver tumor promotion by sustained activation of the aryl hydrocarbon receptor. Regul. Toxicol. Pharmacol. 73, 172-190: PMID: 26145830. The file is open access.

For many years, EPA used a cancer slope factor of 1.5 E+05 per mg/kg/d based on the Kociba et al. (1978) bioassay. Today, the toxicity critierion for TCDD and other persistent AHR ligands is based on purported reproductive and developmental effects in humans.

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The Bradford Hill considerations, which is an approach consistent with the U.S. Environmental Protection Agency’s Guidelines for Carcinogen Risk Assessment available from: <a class="external free" href="http://www.epa.gov/raf/publications/pdfs/CANCER_GUIDELINES_FINAL_3-25-05.PDF" rel="nofollow" target="_blank">http://www.epa.gov/raf/publications/pdfs/CANCER_GUIDELINES_FINAL_3-25-05.PDF</a> as well as the WHO/IPCS’s human relevance-mode-of-action framework provides the framework for putting the proposed AOP into a weight-of-evidence evaluation. These considerations include dose-response, temporality, strength, consistency, specificity and biological plausibility of the proposed association (the AOP in this case). Alternative AOP propositions must be accounted for and either ruled in or out as part of applying the Bradford Hill considerations. To finalise the Bradford Hill assessment, the AOP template requires an examination of the uncertainties, inconsistencies, data gaps, and the quantitative nature of the KE (as well as the AE and ModFs). The Bradford Hill (BH) considerations (dose-response, temporality, strength, consistency, specificity and biological plausibility of the proposed association) form the basis for evaluating weight of evidence within the U.S. Environmental Protection Agency's Guidelines for Carcinogen Risk Assessment, the WHO/IPCS human relevance-MOA framework, the key events/dose-response frame- work (KEDRF) (Dellarco and Fenner-Crisp, 2012; Fenner-Crisp, 2012; Julien et al., 2009; Meek et al., 2003; Meek, 2008; OECD, 2013; USEPA, 2005). The BH considerations have recently been updated and additionally tailored for AOPs by the OECD to facilitate evaluations of KEs and KERs as well as the overall AOP (Meek et al., 2013, 2014a, 2014b; OECD, 2014; Becker et al., 2015). In the tables below, we summarize the weight of evidence evaluation conducted using these AOP-tailored BH considerations of biological plausibility, essentiality and empirical evidence for the sustained AHR activation rodent liver tumor promotion AOP.

The defining question contained in the OECD AOP guidance (OECD, 2014) for evaluation of essentiality is “are downstream KEs and/or the AO prevented if an upstream KE is blocked?” Overall, the evidence in support of essentiality for sustained AHR activation, the MIE, is strong. There is direct evidence of essentiality from the stop- exposure group in the cancer bioassay; the 100 ng/kg/d dose of TCDD was stopped after 30 weeks and at the 2-year termination, no statistically significant increase in tumor frequency was observed (NTP, 2006a; NTP, 2006b; NTP, 2006c). This observation also in- dicates that the MIE of sustained AHR activation requires more than 30 weeks of continuous exposure and is consistent with the general onset of hepatopathy around the same time (Hailey et al., 2005). Additional support for essentiality comes from studies that show that KEs fail to occur when AHR activity is lost through mutation, polymorphism or knockdown (Gasiewicz et al., 2008). Further- more, the loss of AHR responsivity to ligand-activation has been confirmed in reduction and/or loss of ligand-mediated gene tran- scription and resistance to TCDD-induced toxicity (Harrill et al., 2013). Conversely, constitutive AHR activity in mice increased the incidence of tumors and hepatotoxicity (Andersson et al., 2002; Brunnberg et al., 2006; Chopra and Schrenk, 2011; Moennikes et al., 2004).   <table border="1"> <tbody> <tr> <td>Support for Essentiality of KEs</td> <td>Defining Question</td> <td>High (Strong)</td> <td>Moderate</td> <td>Weak</td> </tr> <tr> <td> </td> <td>Are downstream key events and/or the AO prevented if an upstream key event is blocked? [e.g., stop/reversibility studies, antagonism, knock out models, etc.)</td> <td>Multiple lines of experimental evidence illustrating essentiality for several of the key events</td> <td>There is at least one line of experimental evidence indicating essentiality of an important key event</td> <td>Indirect or no experimental evidence of the essentiality of any of the key events</td> </tr> <tr> <td>Pre-MIE: Binding of ligands to the AHR</td> <td colspan="4">Essentiality of the pre-MIE is Strong. Rationale: Binding to the AHR is a necessary element and downstream KEs do not occur in knock-out animals. </td> </tr> <tr> <td>MIE: Sustained AHR Activation</td> <td colspan="4">Essentiality of the MIE is Strong. Rationale: Extensive qualitative and quantitative information showing that downstream KEs occur in with increasing time and extent of continued AHR activation </td> </tr> <tr> <td>KE#1: Changes in Cellular Homeostasis and Inhibition of Apoptosis</td> <td colspan="4">Essentiality of the KE1 is Strong Rationale: Growth of Altered hepatic foci has been explored in many initiation-promotion studies </td> </tr> <tr> <td>KE#2: Hepatoxicity, Hepatopathy</td> <td colspan="4">Essentiality of the KE2 is Strong Rationale: The regenerative nature of the liver is such that the extensive hepatopathy induced by sustained AHR activation leads to a highly proliferative environment in the liver. </td> </tr> <tr> <td>KE#3: Alterations in Cellular Proliferation/Hyperplasia</td> <td colspan="4">Essentiality of the KE3 is Strong Rationale: Hyperplasia has been strongly linked to the induction of cancer in many systems. </td> </tr> </tbody> </table> <h3> </h3>

This section provides brief descriptions of the essentiality of each of the KEs and the biological plausibility, empirical support and any uncertainties or inconsistencies in the key event relationships (KERs). These descriptions are consistent with the table presented in the AOP User’s Guide from OECD. Whilst the overall confidence in the AOP as a result of these evaluations is high, the challenge is to apply this knowledge for a regulatory purpose. An additional section is included below on the Application of the AOP. This provides a discussion of the possible regulatory uses of this AOP. <h4>Empirical Evidence</h4> The OECD AOP guidance (OECD, 2014) for evaluation of empirical evidence focuses on dose-response, temporality and incidence concordance. Defining questions include: “Does the empirical evidence support that a change in KEup leads to an appropriate change in KEdown? Does KEup occur at lower doses and earlier time points than KEdown and is the incidence of KEup > than that for KEdown?” Highly consistent dose-response relationships (KERs) along the sequence of KEs in this AOP exhibit dose- and time-concordance. This consistency in the concordance of both temporality and incidence supports a weight of evidence determination of high for the empirical evidence underpinning this AOP. When the KEs and associative events are placed in sequential order based on dose and temporality, the dose-response slopes from Hill dose-response model fits of the data increase in value. Thus, the later KEs occur with a steeper slope than early KEs and the number of KEs observed increases as a function of dose and time (Simon et al., 2009; Budinsky et al., 2014). Induction of xenobiotic metabolizing enzymes is one the earliest and most sensitive responses to AHR activation (Budinsky et al., 2010; Silkworth et al., 2005). Xenobiotic metabolizing enzyme induction reflects acute transcriptional and proteomic changes that are more aligned with the concept of a pre-MIE and thus provides an associative event for AHR activation. Since measurable enzyme induction persists for at least one year, we used the AUC of a biomarker for this enzyme induction as a measure of sustained AHR activation. Both Hill equation coefficients and half maximal concentrations increase with increasing values of sustained AHR activation and reflect dose-dependent transitions as KEs occur at the various levels of biological organization (Simon et al., 2009; Budinsky et al., 2014). The dose-response temporality table above depicts the KEs increasing in frequency in both dose and time (Meek et al., 2013; Simon et al., 2014). This table is an essential requirement of the human relevance MOA framework and is recommended in the OECD AOP guidance (OECD, 2014). Here, the value of the sustained AHR activation index has been provided for each dose-time combination. One can easily see the increase in sustained AHR activation due to the increase in dose going down the table and the increase in duration going across the table. The need for AHR activation for a sustained period of time, i.e. temporal concordance, is supported by the stop-exposure group in the TCDD cancer bioassay, which showed that when the administration of 100 ng/kg/d TCDD was stopped after 30 weeks, a statistically significant increase in tumor frequency was not observed (NTP, 2006a). This observation also indicates that the AHR activation needs to be sustained for more than 30 weeks for KE#2 to occur (Hailey et al., 2005). <h3> </h3> <h4>Concordance of dose-response relationships</h4> <h4>Dose-Time Concordance Table</h4> Empirical evidence: application of the dose and temporal concordance AOP weight of evidence considerations for Key Events (KEs) at dose/time combination. This table is based on NTP (2006a), Teeguarden et al. (1999) and Maronpot et al. (1993). The dose in the left most column shows the range of average liver concentration (ng/kg) from 14 weeks to 2 years from NTP (2006a). The number in parentheses is the administered gavage dose in ng/kg/d. The numerical value of the sustained AHR activation index (ppb-weeks) is shown for each dose/time combination; the calculation of this value is described on the MIE page. <table border="1"> <tbody> <tr> <td>Dose</td> <td colspan="4">Increasing Time --></td> </tr> <tr> <td> </td> <td>Weeks to Months (14 wk) </td> <td>Months (31 wk) </td> <td>1 yr (53 wk) </td> <td>2 yr (104 wk) </td> </tr> <tr> <td>5-20 (0.1)**</td> <td>0.1 ppb-wk</td> <td>0.5 ppb-wk</td> <td>1 ppb-wk</td> <td>2.4 ppb-wk</td> </tr> <tr> <td>100-200 (1)**</td> <td>1.7 ppb-wk</td> <td>4.7 ppb-wk</td> <td>8.8 ppb-wk</td> <td>19 ppb-wk</td> </tr> <tr> <td>450-650 (3)**</td> <td>4.3 ppb-wk</td> <td>11 ppb-wk</td> <td>19 ppb-wk</td> <td>40 ppb-wk</td> </tr> <tr> <td>1500-2000 (10)</td> <td>MIE = 8.3 ppb-wk Apoptosis Inhibition (KE#1) </td> <td>MIE = 19 ppb-wk Apoptosis Inhibition (KE#1)(presumed) </td> <td>MIE = 34 ppb-wk</td> <td>MIE = 69 ppb-wk Toxic Hepatopathy (KE#2) Hyperplasia/Proliferation (KE#3) </td> </tr> <tr> <td>3500-4200 (22)</td> <td>MIE = 11 ppb-wk Apoptosis Inhibition (KE#1)(presumed) </td> <td>MIE = 24 ppb-wk Apoptosis Inhibition (KE#1)(presumed) </td> <td>MIE = 42 ppb-wk</td> <td>MIE = 83 ppb-wk Toxic Hepatopathy (KE#2) Hyperplasia/Proliferation (KE#3) </td> </tr> <tr> <td>7000-8000 (46)</td> <td>MIE = 12 ppb-wk Apoptosis Inhibition (KE#1)(presumed) </td> <td>MIE = 27 ppb-wk Apoptosis Inhibition (KE#1)(presumed) </td> <td>MIE = 47 ppb-wk Toxic Hepatopathy (KE#2) </td> <td>MIE = 93 ppb-wk Toxic Hepatopathy (KE#2) Hyperplasia/Proliferation (KE#3) Cholangiocarcinomas (AO) </td> </tr> <tr> <td>15000-17000 (100)</td> <td>MIE = 13 ppb-wk Apoptosis Inhibition (KE#1) </td> <td>MIE = 29 ppb-wk Apoptosis Inhibition (KE#1) Toxic Hepatopathy (KE#2) </td> <td>MIE = 50 ppb-wk Toxic Hepatopathy (KE#2) Hyperplasia/Proliferation (KE#3) </td> <td>MIE = 98 ppb-wk Toxic Hepatopathy (KE#2) Hyperplasia/Proliferation (KE#3) Cholangiocarcinomas (AO) Hepatic Adenomas (AO) </td> </tr> </tbody> </table> <ul> <li> <ul> <li>these dose levels are insufficient to induce the degree of sustained AHR activation necessary to exceed the threshold of homeostasis/adaptation; exceeding this threshold is required to trigger the MIE.</li> </ul> </li> </ul> When the KEs and associative events are placed in sequential order based on dose and temporality, the dose-response slopes from Hill dose-response model fits of the data increase in value. Thus, the later KEs occur with a steeper slope than early KEs and the number of KEs observed increases as a function of dose and time (Table 2) (Simon et al., 2009; Budinsky et al., 2014). Induction of xenobiotic metabolizing enzymes is one the earliest and most sensitive responses to AHR activation (Budinsky et al., 2010; Silkworth et al., 2005). Xenobiotic metabolizing enzyme in- duction reflects acute transcriptional and proteomic changes that are more aligned with the concept of a pre-MIE and thus provides an associative event for AHR activation. Since measurable enzyme induction persists for at least one year, we used the AUC of a biomarker for this enzyme induction as a measure of sustained AHR activation. Both Hill equation coefficients and half maximal concentrations increase with increasing values of sustained AHR activation and reflect dose-dependent transitions as KEs occur at the various levels of biological organization (Simon et al., 2009; Budinsky et al., 2014). The table shown above is the dose-response temporality table and depicts the KEs increasing in both dose and time (Meek et al., 2013; Simon et al., 2014). This table is an essential requirement of the human relevance MOA framework and is recommended in the OECD AOP guidance (OECD, 2014). Here, the value of the sustained AHR activation index has been provided for each dose-time combination. One can easily see the increase in sustained AHR activation due to the increase in dose going down the table and the increase in duration going across the table. The need for AHR activation for a sustained period of time, i.e. temporal concordance, is supported by the stop-exposure group in the TCDD cancer bioassay, which showed that when the administration of 100 ng/kg/d TCDD was stopped after 30 weeks, a statistically significant increase in tumor frequency was not observed (NTP, 2006a). This observation also indicates that the AHR activation needs to be sustained for more than 30 weeks for KE#2 to occur (Hailey et al., 2005). <h4>Support for the Biological Plausibility of the KERs</h4> The OECD AOP guidance (OECD, 2014) for evaluation of biological plausibility of an AOP provides this defining question for evaluating biological plausibility: “is there a mechanistic (i.e., structural or functional) relationship between KEup and KEdown consistent with established biological knowledge?” Under the OECD guidance, a high degree of confidence is afforded when there is an established mechanistic basis and “extensive understanding of the KER based on extensive previous documentation and broad acceptance.” For biological plausibility, for this AOP, the WoE for each KER is judged to be strong, as is the WoE for the overall AOP. All the elements in this AOP are strongly associated with the biological steps and elements of carcinogenesis (Hanahan and Weinberg, 2011). First, there is extensive body of mechanistic evi- dence in support the biological plausibility of this MOA (see recent review by Budinsky et al., 2014). Further, the relationships between sustained AHR activation and 1) decreased intrafocal apoptosis (KE#1); 2) increased cell proliferation (KE#2); 3) toxic hepatopathy (KE#3); and 4) eventual tumor formation (AO) are evident from a surfeit of published studies (e.g., Fig. 4). Moreover, overall consistency with knowledge of the pathogenesis of liver tumor promo- tion is supported by replication of events related to tumor promotion across different laboratories and the multiple lines of evidence for sustained AHR activation acting as a mechanism of liver tumor promotion. Thus, the AOP is well supported by the KEs, consistent with the biology of carcinogenesis and the events of tumor promotion (Dietrich and Kaina, 2010; Gasiewicz et al., 2008; Roberts et al., 1997). The unique sensitivity of the female rat response suggests a possible role for estrogen as a modulating factor in the tumorigenic MOA. Estrogen is an established co-promoter of tumorigenesis and thus may play a role in the MOA (Graham et al., 1988; Hiraku et al., 2001; Lucier et al., 1991; Vickers and Lucier, 1996; Vickers et al., 1989). Crosstalk between the AHR pathway and the estrogen receptor pathway may also be a contributing factor (Matthews and Gustafsson, 2006). Such receptor mediated cross talk is consistent with the sustained AHR MOA.   <table border="1"> <tbody> <tr> <td>Support for Biological Plausibility of KERs</td> <td>Defining Question</td> <td>High (Strong)</td> <td>Moderate</td> <td>Weak</td> </tr> <tr> <td> </td> <td>a) Is there a mechanistic (i.e., structural or functional) relationship between KEup and KEdown consistent with established biological knowledge?</td> <td>Extensive understanding of the KER based on extensive previous documentation and broad acceptance (e.g., mutation leading to tumors) -Established mechanistic basis </td> <td>The KER is plausible but scientific understanding is not completely established.</td> <td>Only limited or indirect evidence for KER (i.e., based on empirical support, only (See 3.)</td> </tr> <tr> <td>Binding of Ligands to the AHR leading to sustained activation of the AHR</td> <td colspan="4">Biological Plausibility of the pre-MIE => MIE is Strong Rationale: Long-established knowledge of and extensive research on dioxin-like chemicals and other AHR ligands. </td> </tr> <tr> <td>Sustained AHR Activation directly leading to Changes in Cellular Homeostasis and Inhibition of Apoptosis</td> <td colspan="4">Biological Plausibility of MIE => KE1 is Strong. Rationale: Direct empirical evidence showing continued application of AHR activators leads to growth alteration of altered hepatic foci. </td> </tr> <tr> <td>Sustained AHR Activation indirectly leading to Hepatoxicity and Hepatopathy</td> <td colspan="4">Biological Plausibility of MIE => KE2 is Strong Rationale: Long established knowledge : Empirical data from two-year bioassays using AHR activation and sustained administration </td> </tr> <tr> <td>Sustained AHR Activation indirectly leading to Alterations in Cellular Proliferation and Hyperplasia</td> <td colspan="4">Biological Plausibility of MIE => KE3 is Strong Rationale: Empirical data from two-year bioassays using AHR activation and sustained administration </td> </tr> <tr> <td>Sustained AHR Activation indirectly leading to Hepatocellular Adenomas and Cholangiocarcinomas</td> <td colspan="4">Biological Plausibility of MIE => AO is Strong Rationale: Empirical data from two-year bioassays using AHR activation and sustained administration </td> </tr> </tbody> </table> <h3> </h3> <h4>Uncertainties, Inconsistencies and Conflicting Evidence for KERs and the AOP</h4> The evidence supporting the KERs and AOP is strong. Alternative MOA(s) or KEs and KE elements can be examined to help ascertain the confidence in the MOA considered most likely (Boobis et al., 2006, 2008, 2009; Cohen et al., 2003; Cohen et al., 2004; Julien et al., 2009; Meek et al., 2003; Meek, 2008; Seed et al., 2005; Sonich-Mullin et al., 2001; USEPA, 2005). One such alternative MOA would be that DLCs act to produce liver tumors in rodents by a mutagenic mechanism. However, there is substantial evidence that DLCs are neither mutagenic nor genotoxic compounds and thus do not act by a mutagenic MOA (Bock and Kohle, 2005; Dragan and Schrenk, 2000; Knerr et al., 2006; Poland and Glover, 1979; Randerath et al., 1990; Schwarz et al., 2000; Turteltaub et al., 1990; Wassom et al., 1977; Whysner and Williams, 1996). Effects on gap junctions or induction of oxidative stress are two other potential mechanisms. TCDD disrupts normal gap junction activity and intercellular communication in rat primary hepatocytes and WB-344 cells (Andrysík et al., 2013; Bager et al., 1997; Herrmann et al., 2002; Weiss et al., 2008). Further research is needed to understand the contribution of this mechanism to DLC- induced rodent liver tumor formation. Oxidative stress appears less likely as an alternative MOA and may be a late-occurring associative event due to continued high activity of phase 1 mixed function oxidases and accompanying cytotoxicity. In summary, the WoE in support of sustained AHR activation leading to changes in cellular growth homeostasis and eventually promotion of liver tumors in rodents is strong. The WoE supporting the alternative MOAs is much weaker. <table border="1"> <tbody> <tr> <td>Uncertainty and Conflicting Evidence for KERs and AOP</td> <td>Defining Question</td> <td>High (Strong)</td> <td>Moderate</td> <td>Weak</td> </tr> <tr> <td> </td> <td>Are there inconsistencies in empirical support across taxa, species which don’t align with appropriate pattern for hypothesized KERs and AOP? Are there significant knowledge gaps or uncertainties with regard to the relationship between the KEs and overall AOP? </td> <td>No (or very few) knowledge gaps or inconsistent / conflicting lines of evidence.)</td> <td>Some inconsistent evidence but which can be explained by factors such as experimental design, technical considerations, differences among laboratories, etc.)</td> <td>Contradictory evidence in for which no plausible explanation is known.</td> </tr> <tr> <td>Binding of Ligands to the AHR leading to sustained activation of the AHR</td> <td colspan="4">Inconsistencies / Uncertainties of Pre-MIE => MIE is Strong Rationale: Highly certain. While a large number of ligands bind to and activate the AHR, only the biologically persistent ligands, such as dioxin-like chemicals produce the MIE, sustained AHR activation </td> </tr> <tr> <td>Sustained AHR Activation directly leading to Changes in Cellular Homeostasis and Inhibition of Apoptosis</td> <td colspan="4">Inconsistencies / Uncertainties of MIE => KE1 is Strong. Rationale: A large number of initiation-promotion studies with TCDD or other dioxin-like chemicals documented changes in both cell proliferation and inhibition of apoptosis within alterered hepatic foci. </td> </tr> <tr> <td>Sustained AHR Activation indirectly leading to Hepatoxicity and Hepatopathy</td> <td colspan="4">Inconsistencies / Uncertainties of MIE => KE2 is 'Strong Rationale: Although the exact mechanism is not known, sustained AHR activation often leads to increased concentrations of ROS that may be a factor in generating cytotoxicity. Hepatopathy is common effect of sustained adiministration of AHR activators. </td> </tr> <tr> <td>Sustained AHR Activation indirectly leading to Alterations in Cellular Proliferation and Hyperplasia</td> <td colspan="4">Inconsistencies / Uncertainties of MIE => KE3 is Strong Rationale: All the biologically persistent AHR activators such as DLCs damage the liver to a sufficient extent that a proliferative/regenerative environment is created in the organ. </td> </tr> <tr> <td>Sustained AHR Activation indirectly leading to Hepatocellular Adenomas and Cholangiocarcinomas</td> <td colspan="4">Inconsistencies / Uncertainties of MIE => AO is Strong Rationale: These tumors are outcomes of the AHF growth in KE#1 and the increased proliferation in KE#3. Both are induced by the MIE, sustained AHR activation. A large number of bioassays have documented the fact that persistent AHR ligands produce liver tumors in rodents. </td> </tr> </tbody> </table> <h2> </h2>

While binding of ligand to the AHR is identified as a pre-MIE, sustained AHR activation by persistent ligands such as DLCs is linked qualitatively and quantitatively to both downstream KEs and the AO. These linkages notwithstanding, additional work is needed to develop and evaluate such a prediction model before the MIE of sustained AHR activation can be used in a quantitative prediction model of the AO. Any prediction model based on this AOP needs to consider the unique aspects of the AHR and its response to DLCs, including the involvement of initiated or partially differentiated stem cells, and such a model would need evaluation/validation for its intended use (Cox et al., 2014; Patlewicz et al., 2015).

The OECD guidance for AOP development (OECD, 2013) suggests a number of potential uses for AOPs. These include 1) category formation for read-across, 2) integrated approaches for testing and assessment 3) development or refinement of test methods such as OECD test guidelines and 4) hazard identification (classification/ labeling) and risk assessment. The use of a specific AOP for any one or more of these applications depends on scientific confidence in the AOP for each specific use. An AOP can offer practical utility in certain applications even if confidence is not sufficient to quantitatively predict the AO from the MIE. Application of the sustained AHR activation AOP for several applications was described in brief in Patlewicz et al. (2015); Below the application of and confidence in this AOP is discussed in more detail. <h4>Which Key Events can be used to predict the AO?</h4> To date, no quantitative models have been developed to predict the adverse outcome from AHR activation by ligand binding. With the exception of DLCs, PCDDs, PCDFs and co-planar PCBs, this predictive capability is highly uncertain for the plethora of AHR ligands. For example, indole 3-carbinol is an AHR ligand occurring in cruciferous vegetables and acts as a cancer chemopreventive agent. In the stomach, indole 3-carbinol forms 3,3- diinolylmethane, a potent AHR ligand that has shown promise for preventing tumor reoccurrence in humans (Banerjee et al., 2011). Dietary administration of indole-3-carbinol for 23 weeks inhibited tumor formation in rats initiated with diethylnitrosamine (Tanaka et al., 1990). However, also in diethylnitrosamine-initiated rats, prolonged indole 3-carbinol administration (>26 weeks) increased the progression of altered hepatic foci to hepatocellular adenomas (Yamamoto et al., 2013). A recent NTP 2-year cancer bioassay failed to demonstrate indole 3-carbinol as a liver tumor promoter in fe- male rats (NTP, 2014). Furthermore, endogenous AHR ligands and naturally occurring exogenous ligands occurring in foods have cancer-preventive properties and likely contribute to a relatively high level of AHR activation activity in human blood (Connor et al., 2008; Navarro et al., 2009, 2011; Peterson et al., 2009; Wincent et al., 2009). These naturally occurring and endogenous ligands induce both their own metabolism and that of other AHR ligands through increased induction of xenobiotic metabolizing enzymes. The transient metabolic increase may be one aspect of the pre- ventive effect against sustained AHR activation that would lead to KEs related to liver tumor promotion. Neither binding of ligand to the AHR nor short-term transcriptional changes and cellular responses are sufficient to produce liver tumors in rats. Strains of rats that show resistance towards the toxic and carcinogenic effects of DLCs express different genomic profiles outside of the conserved core battery response (Boutros et al., 2011; Yao et al., 2012). Acute genomic changes do not appear to be pre- dictive for the cancer endpoint (Fielden et al., 2011; Ovando et al., 2010). AHR activation-induced transcriptional changes occur within hours of ligand activation; yet the subsequent KEs and AO require months of sustained AHR activation for tumors to occur. Hence, the distinction between short-term and sustained activation of the AHR is an important one and AHR activation must be sus- tained for more than 30% of the rodent lifespan to result in tumor promotion. While binding of ligand to the AHR is identified as a pre-MIE, sustained AHR activation by persistent ligands such as DLCs is linked qualitatively and quantitatively to both downstream KEs and the AO. These linkages notwithstanding, additional work is needed to develop and evaluate such a prediction model before the MIE of sustained AHR activation can be used in a quantitative prediction model of the AO. Any prediction model based on this AOP needs to consider the unique aspects of the AHR and its response to DLCs, including the involvement of initiated or partially differentiated stem cells, and such a model would need evaluation/validation for its intended use (Cox et al., 2014; Patlewicz et al., 2015). <h4>Using this AOP for grouping chemicals into chemical categories for read-across</h4> Without some measure of sustained activation, the use of pre- MIEs for any purpose other than preliminary screening is problematic as a predictive criteria for liver tumor promotion. Evidence clearly shows it is the combination of sustained AHR activation and the subsequent biological changes involving complex parenchymal and non-parenchymal cell interactions that underlie the hepatotoxicity, the increase in cell proliferation and the apical tumor response. Hence, the MIE is defined as sustained AHR activation, and not simply AHR activation. In addition, the promiscuity of the AHR and the species- and strain-specificity of the initial genomic responses suggest that category development may prove a challenge (Denison, 2011; Dere, 2011). <h4>Using this AOP for integrated approaches to testing and assessment (IATA)</h4> The most straightforward use of this AOP within an integrated testing and assessment approach for hazard evaluation would be to determine the potential for a substance to activate the AHR in a sustained manner with long-term changes in gene transcription involving multiple cell types, which leads to increased liver cell proliferation. An IATA decision-tree approach, for illustrative purposes has been already presented by Patlewicz et al. (2015). The initial steps in the IATA focus on evaluating molecular and cellular events related to AHR binding and transcriptional activation using rapid and cost effective in silico or in vitro assays. Compounds found to be inactive in such assays would not proceed forward into further testing. At the present time, there is insufficient understanding to permit the use transcription profiling as a metric of sustained AHR activation to quantitatively predict development of rat liver foci and liver tumors. Therefore, the IATA proposes that substances found to be active in the AHR mechanistic assays be subjected to a decision framework for further evaluating the potential to act as rodent liver tumor promoter. For example, a subchronic study, utilizing an appropriate dosing regimen, may be able to rule-in or rule-out the substance's ability to trigger the critical histological components of hepatopathy (Hailey et al., 2005). Or a rodent liver initiation- promotion assay could be considered, though interpretation can be challenging (Tanaka et al., 1990; Yamamoto et al., 2013; NTP, 2014). Patlewicz et al. (2015) also illustrate how exposure information can be used in conjunction with the AOP to inform IATA decisions. <h4>Using this AOP to inform test method development or refinement</h4> An IATA consisting of a suite of in vitro and in vivo (e.g., sub- chronic) assays to predict hepatopathy, a complex histological response, may be needed to differentiate AHR ligands with and without liver tumor promotion potential. Theoretically, it may be plausible to consider using a combination of AHR-binding, AHR- transcriptional activation and rat liver initiation-promotion assays to develop a prediction model for sustained AHR activation- induced rat liver tumors. Therefore, within the OECD test guidelines program, it may be worthwhile to consider developing performance criteria that could be applied to judge the scientific quality and reliability of in vitro AHR-binding and transactivation assays, liver stem cell assays, as well as a validated test guideline for a rat liver tumor (hepatic foci) initiation-promotion assay.

Abraham, K., Geusau, A., Tosun, Y., Helge, H., Bauer, S., Brockmoller, J., 2002. Severe 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) intoxication: insights into the measurement of hepatic cytochrome P450 1A2 induction. Clin. Pharmacol. Ther. 72, 163-174. Andersen M.E., Preston R.J., Maier A., Willis A.M., Patterson J. 2014. Dose-response approaches for nuclear receptor-mediated modes of action for liver carcinogenicity: Results of a workshop, Crit. Rev. Toxicol. 44, 50-63. Andrysík, Z., Prochazkova, J., Kabatkova, M., Umannova, L., Simeckova, P., Kohoutek, J., Kozubík, A., Machala, M., Vondracek, J., 2013. Aryl hydrocarbon receptor-mediated disruption of contact inhibition is associated with connexin43 downregulation and inhibition of gap junctional intercellular communication. Arch. Toxicol. 87, 491-503. Ankley G.T., Bennett R.S., Erickson R.J., Hoff D.J., Hornung M.W., Johnson R.D., Mount D.R., Nichols J.W., Russom C.L., Schmieder P.K., Serrrano J.A., Tietge J.E., Villeneuve D.L. 2010. Adverse outcome pathways: a conceptual framework to support ecotoxicology research and risk assessment, Environ. Toxicol. Chem. 29, 730-741. Bager, Y., Lindebro, M.C., Martel, P., Chaumontet, C., W€arngård, L., 1997. Altered function, localization and phosphorylation of gap junctions in rat liver epithelial, IAR 20, cells after treatment with PCBs or TCDD. Environ. Toxicol. Pharmacol. 3, 257- 266. Banerjee, S., Kong, D., Wang, Z., Bao, B., Hillman, G.G., Sarkar, F.H., 2011. Attenuation of multi-targeted proliferation-linked signaling by 3,30 -diindolylmethane (DIM): from bench to clinic. Mutat. Res. 728, 47-66. Becker, R.A., Patlewicz, G., Simon, T.W., Rowlands, J.C., Budinsky, R.A. 2015. The adverse outcome pathway for rodent liver tumor promotion by sustained activation of the aryl hydrocarbon receptor. Regul. Toxicol. Pharmacol. 73, 172-190. Black, M.B., Budinsky, R.A., Dombkowski, A., Lecluyse, E.L., Ferguson, S.S., Thomas, R.S., Rowlands, J.C., 2012. Cross-species comparisons of transcriptomic alterations in human and rat primary hepatocytes exposed to 2,3,7,8- tetrachlorodibenzo-p-dioxin. Toxicol. Sci. 127, 199-215. Boobis, A.R., Cohen, S.M., Dellarco, V., McGregor, D., Meek, M.E.B., Vickers, C., Willcocks, D., Farland, W., 2006. IPCS framework for analyzing the relevance of a cancer mode of action for humans. Crit. Rev. Toxicol. 36, 781-792. Boobis, A.R., Daston, G.P., Preston, R.J., Olin, S.S., 2009. Application of key events analysis to chemical carcinogens and noncarcinogens. Crit. Rev. Food Sci. Nutr. 49, 690-707. Boobis, A.R., Doe, J.E., Heinrich-Hirsch, B., Meek, M.E.B., Munn, S., Ruchirawat, M., Schlatter, J., Seed, J., Vickers, C., 2008. IPCS framework for analyzing the rele- vance of a noncancer mode of action for humans. Crit. Rev. Toxicol. 38, 87-96. Boutros, P.C., Yao, C.Q., Watson, J.D., Wu, A.H., Moffat, I.D., Prokopec, S.D., Smith, A.B., Okey, A.B., Pohjanvirta, R., 2011. Hepatic transcriptomic responses to TCDD in dioxin-sensitive and dioxin-resistant rats during the onset of toxicity. Toxicol. Appl. Pharmacol. 251, 119-129. Coenraads, P.J., Olie, K., Tang, N.J., 1999. Blood lipid concentrations of dioxins and dibenzofurans causing chloracne. Br. J. Dermatol. 141, 694-697. Connor, K.T., Harris, M.A., Edwards, M.R., Budinsky, R.A., Clark, G.C., Chu, A.C., Finley, B.L., Rowlands, J.C., 2008. AH receptor agonist activity in human blood measured with a cell-based bioassay: evidence for naturally occurring AH re- ceptor ligands in vivo. J. Expo. Sci. Environ. Epidemiol. 18, 369-380. Becker et al. 2015. Increasing Scientific Confidence in Adverse Outcome Pathways: Application of Tailored Bradford-Hill Considerations for Evaluating Weight of Evidence, Regul. Toxicol. Pharmacol. 72, 514-37. Beebe L.E., Fornwald L.W., Diwan B.A., Anver M.R., Anderson L.M. 1995. Promotion of N-nitrosodiethylamine-initiated hepatocellular tumors and hepatoblastomas by 2,3,7,8-tetrachlorodibenzo-p-dioxin or Aroclor 1254 in C57BL/6, DBA/2, and B6D2F1 mice, Cancer. Res. 55, 4875-4880. Beischlag, T.V., Luis Morales, J., Hollingshead, B.D., Perdew, G.H., 2008. The aryl hydrocarbon receptor complex and the control of gene expression. Crit. Rev. Eukaryot. Gene. Expr. 18, 207-250. Black, M.B., Budinsky, R.A., Dombkowski, A., Lecluyse, E.L., Ferguson, S.S., Thomas, R.S., Rowlands, J.C., 2012. Cross-species comparisons of transcriptomic alterations in human and rat primary hepatocytes exposed to 2,3,7,8- tetrachlorodibenzo-p-dioxin. Toxicol. Sci. 127, 199-215. Bock K.W., Kohle C. 2005. Ah receptor- and TCDD-mediated liver tumor promotion: clonal selection and expansion of cells evading growth arrest and apoptosis, Biochem. Pharmacol. 69, 1403-1408. Buchmann A., Stinchcombe S., Korner W., Hagenmaier H., Bock K.W. 1994. Effects of 2,3,7,8-tetrachloro- and 1,2,3,4,6,7,8-heptachlorodibenzo-p-dioxin on the proliferation of preneoplastic liver cells in the rat, Carcinogenesis. 15, 1143-1150. Budinsky, R.A., LeCluyse, E.L., Ferguson, S.S., Rowlands, J.C., Simon, T., 2010. Human and rat primary hepatocyte CYP1A1 and 1A2 induction with 2,3,7,8- tetrachlorodibenzo-p-dioxin, 2,3,7,8-tetrachlorodibenzofuran, and 2,3,4,7,8- pentachlorodibenzofuran. Toxicol. Sci. 118, 224-235. Budinsky R.A., Schrenk D., Simon T., Van den Berg M., Reichard J.F., Silkworth J.B., Aylward L.L., Brix A., Gasiewicz T., Kaminski N., Perdew G., Starr T.B., Walker N.J., Rowlands J.C. 2014. Mode of action and dose-response framework analysis for receptor-mediated toxicity: The aryl hydrocarbon receptor as a case study, Crit. Rev. Toxicol. 44, 83-119. Carlson, E.A., McCulloch, C., Koganti, A., Goodwin, S.B., Sutter, T.R., Silkworth, J.B., 2009. Divergent transcriptomic responses to aryl hydrocarbon receptor agonists between rat and human primary hepatocytes. Toxicol. Sci. 112, 257-272. Carmichael N., Bausen M., Boobis A.R., Cohen S.M., Embry M., Fruijtier-Pölloth C., Greim H., Lewis R., Bette Meek M.E., Mellor H., Vickers C., Doe J. 2011. Using mode of action information to improve regulatory decision-making: an ECETOC/ILSI RF/HESI workshop overview, Crit. Rev. Toxicol. 41, 175-186. Cohen, S. M., Ellwein, L. B., 1990. Cell proliferation in carcinogenesis. Science. 249, 1007-11. Cohen, S.M., Klaunig, J., Meek, M.E., Hill, R.N., Pastoor, T., Lehman-McKeeman, L., Bucher, J., Longfellow, D.G., Seed, J., Dellarco, V., Fenner-Crisp, P., Patton, D., 2004. Evaluating the human relevance of chemically induced animal tumors. Toxicol. Sci. 78, 181-186. Cohen, S.M., Meek, M.E., Klaunig, J.E., Patton, D.E., Fenner-Crisp, P.A., 2003. The human relevance of information on carcinogenic modes of action: overview. Crit. Rev. Toxicol. 33, 581-589. Cohen, S. M., Arnold, L. L., 2011. Chemical carcinogenesis. Toxicol Sci. 120 Suppl 1, S76-92. Connor, K.T., Aylward, L.L., 2006. Human response to dioxin: aryl hydrocarbon re- ceptor (AhR) molecular structure, function, and dose-response data for enzyme induction indicate an impaired human AhR. J. Toxicol. Environ. Health B Crit. Rev. 9, 147-171. Connor, K.T., Harris, M.A., Edwards, M.R., Budinsky, R.A., Clark, G.C., Chu, A.C., Finley, B.L., Rowlands, J.C., 2008. AH receptor agonist activity in human blood measured with a cell-based bioassay: evidence for naturally occurring AH re- ceptor ligands in vivo. J. Expo. Sci. Environ. Epidemiol. 18, 369-380. Cox, L.A., Popken, D., Marty, M.S., Rowlands, J.C., Patlewicz, G., Goyak, K.O., Becker, R.A., 2014. Developing scientific confidence in HTS-derived prediction models: lessons learned from an endocrine case study. Regul. Toxicol. Pharmacol. 69, 443-450. Della Porta G., Dragani T.A., Sozzi G. 1987. Carcinogenic effects of infantile and long-term 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment in the mouse, Tumori. 73, 99-107. Dellarco V., Fenner-Crisp P.A. 2012. Mode of Action: Moving toward a More Relevant and Efficient Assessment Paradigm, J. Nutr. 142, 2192S-2198S. Denison M.S., Nagy S.R. 2003. Activation of the aryl hydrocarbon receptor by structurally diverse exogenous and endogenous chemicals, Annu. Rev. Pharmacol. Toxicol. 43, 309-334. Denison M.S., Soshilov A.A., He G., DeGroot D.E., Zhao B. 2011. Exactly the same but different: promiscuity and diversity in the molecular mechanisms of action of the aryl hydrocarbon (dioxin) receptor, Toxicol. Sci. 124, 1-22. Dere, E., Lee, A.W., Burgoon, L.D., Zacharewski, T.R., 2011. Differences in TCDD- elicited gene expression profiles in human HepG2, mouse Hepa1c1c7 and rat H4IIE hepatoma cells. BMC Genomics 12, 193. Dragan Y.P., Xu X.H., Goldsworthy T.L., Campbell H.A., Maronpot R.R., Pitot H.C. 1992. Characterization of the promotion of altered hepatic foci by 2,3,7,8-tetrachlorodibenzo-p-dioxin in the female rat, Carcinogenesis. 13, 1389-1395. Dragan, Y.P., Schrenk, D., 2000. Animal studies addressing the carcinogenicity of TCDD (or related compounds) with an emphasis on tumour promotion. Food. Addit. Contam. 17, 289-302. Fenner-Crisp P.A. 2012. Application of the International Life Sciences Institute Key Events Dose-Response Framework to food contaminants, J. Nutr. 142, 2199S-2206S. Fielden, M.R., Adai, A., Dunn, R.T., Olaharski, A., Searfoss, G., Sina, J., Aubrecht, J., Boitier, E., Nioi, P., Auerbach, S., Jacobson-Kram, D., Raghavan, N., Yang, Y., Kincaid, A., Sherlock, J., Chen, S.-J., Car, B., Predictive Safety Testing Consortium, Carcinogenicity Working Group, 2011. Development and evaluation of a genomic signature for the prediction and mechanistic assessment of non- genotoxic hepatocarcinogens in the rat. Toxicol. Sci. 124, 54-74. Flaveny, C.A., Murray, I.A., Perdew, G.H., 2010. Differential gene regulation by the human and mouse aryl hydrocarbon receptor. Toxicol. Sci. 114, 217-225. Gasiewicz T.A., Henry E.C., Collins L.L. 2008. Expression and activity of aryl hydrocarbon receptors in development and cancer, Crit. Rev. Eukaryot. Gene. Expr. 18, 279-321. Goodman D.G., Sauer R.M. 1992. Hepatotoxicity and carcinogenicity in female Sprague-Dawley rats treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD): a pathology working group reevaluation, Regul. Toxicol. Pharmacol. 15, 245-252. Guzelian, P., Quattrochi, L., Karch, N., Aylward, L., Kaley, R., 2006. Does dioxin exert toxic effects in humans at or near current background body levels?: An evidence-based conclusion. Hum. Exp. Toxicol. 25, 99-105. Hailey J.R., Walker N.J., Sells D.M., Brix A.E., Jokinen M.P., Nyska A. 2005. Classification of proliferative hepatocellular lesions in harlan sprague-dawley rats chronically exposed to dioxin-like compounds, Toxicol. Pathol. 33, 165-174. Haarmann-Stemmann, T., Bothe, H., Kohli, A., Sydlik, U., Abel, J., Fritsche, E., 2007. Analysis of the transcriptional regulation and molecular function of the aryl hydrocarbon receptor repressor in human cell lines. Drug. Metab. Dispos. 35, 2262-2269. Herrmann S., Seidelin M., Bisgaard H.C., Vang O. 2002. Indolo[3,2-b]carbazole inhibits gap junctional intercellular communication in rat primary hepatocytes and acts as a potential tumor promoter, Carcinogenesis. 23, 1861-1868. Hill A.B. 1965. The Environment and Disease: Association or Causation? Proc. R. Soc. Med. 58, 295-300. Hu Q., He G., Zhao J., Soshilov A., Denison M.S., Zhang A., Yin H., Fraccalvieri D., Bonati L., Xie Q., Zhao B. 2013. Ginsenosides are novel naturally-occurring aryl hydrocarbon receptor ligands, PLoS. One. 8, e66258. Julien E., Boobis A.R., Olin S.S., Ilsi Research Foundation Threshold Working Group 2009. The Key Events Dose-Response Framework: a cross-disciplinary mode-of-action based approach to examining dose-response and thresholds, Crit. Rev. Food. Sci. Nutr. 49, 682-689. Kim, S., Dere, E., Burgoon, L. D., Chang, C. C., Zacharewski, T. R., 2009. Comparative analysis of AhR-mediated TCDD-elicited gene expression in human liver adult stem cells. Toxicol Sci. 112, 229-44. Knerr S., Schaefer J., Both S., Mally A., Dekant W., Schrenk D. 2006. 2,3,7,8-Tetrachlorodibenzo-p-dioxin induced cytochrome P450s alter the formation of reactive oxygen species in liver cells, Mol. Nutr. Food. Res. 50, 378-384. Kociba R.J., Keyes D.G., Beyer J.E., Carreon R.M., Wade C.E., Dittenber D.A., Kalnins R.P., Frauson L.E., Park C.N., Barnard S.D., Hummel R.A., Humiston C.G. 1978. Results of a two-year chronic toxicity and oncogenicity study of 2,3,7,8-tetrachlorodibenzo-p-dioxin in rats, Toxicol.Appl.Pharmacol.. 46, 279-303. Lambert, G.H., Needham, L.L., Turner, W., Lai, T.J., Patterson, D.G.J., Guo, Y.L., 2006. Induced CYP1A2 activity as a phenotypic biomarker in humans highly exposed to certain PCBs/PCDFs. Environ. Sci. Technol. 40, 6176-6180. Maronpot R.R., Foley J.F., Takahashi K., Goldsworthy T., Clark G., Tritscher A., Portier C., Lucier G. 1993. Dose response for TCDD promotion of hepatocarcinogenesis in rats initiated with DEN: histologic, biochemical, and cell proliferation endpoints 8, Environ. Health. Perspect.. 101, 634-642. Matthews, J., Gustafsson, J.A., 2006. Estrogen receptor and aryl hydrocarbon receptor signaling pathways. Nucl. Recept. Signal 4, e016. Matthews, J., Wihlen, B., Thomsen, J., Gustafsson, J.A., 2005. Aryl hydrocarbon receptor-mediated transcription: ligand-dependent recruitment of estrogen receptor alpha to 2,3,7,8-tetrachlorodibenzo-p-dioxin-responsive promoters. Mol. Cell Biol. 25, 5317-5328. Meek, M.E., 2008. Recent developments in frameworks to consider human rele- vance of hypothesized modes of action for tumours in animals. Environ. Mol. Mutagen 49, 110-116. Meek, M.E., Bucher, J.R., Cohen, S.M., Dellarco, V., Hill, R.N., Lehman- McKeeman, L.D., Longfellow, D.G., Pastoor, T., Seed, J., Patton, D.E., 2003. A framework for human relevance analysis of information on carcinogenic modes of action. Crit. Rev. Toxicol. 33, 591-653. Meek M.E., Bolger M., Bus J.S., Christopher J., Conolly R.B., Lewis R.J., Paolini G.M., Schoeny R., Haber L.T., Rosenstein A.B., Dourson M.L. 2013. A framework for fit-for-purpose dose response assessment, Regul. Toxicol. Pharmacol. 66, 234-240. Meek M.E.B., Palermo C.M., Bachman A.N., North C.M., Jeffrey Lewis R. 2014a. Mode of action human relevance (species concordance) framework: Evolution of the Bradford Hill considerations and comparative analysis of weight of evidence, J. Appl. Toxicol. 34, 595-606. Meek M.E., Boobis A., Cote I., Dellarco V., Fotakis G., Munn S., Seed J., Vickers C. 2014b. New developments in the evolution and application of the WHO/IPCS framework on mode of action/species concordance analysis, J. Appl. Toxicol. 34, 1-18. Navarro, S.L., Chen, Y., Li, L., Li, S.S., Chang, J.-L., Schwarz, Y., King, I.B., Potter, J.D., Bigler, J., Lampe, J.W., 2011. UGT1A6 and UGT2B15 polymorphisms and acet- aminophen conjugation in response to a randomized, controlled diet of select fruits and vegetables. Drug. Metab. Dispos. 39, 1650-1657. Navarro, S.L., Peterson, S., Chen, C., Makar, K.W., Schwarz, Y., King, I.B., Li, S.S., Li, L., Kestin, M., Lampe, J.W., 2009. Cruciferous vegetable feeding alters UGT1A1 activity: diet- and genotype-dependent changes in serum bilirubin in a controlled feeding trial. Cancer Prev. Res.. (Phila) 2, 345-352. NTP 1980. Bioassay of a Mixture of 1,2,3,6,7,8-Hexachlorodibenzo-p-dioxin and 1,2,3,7,8,9-Hexachlorodibenzo-p-dioxin (Gavage) for Possible Carcinogenicity (CAS No. 57653-85-7,CAS No. 19408-74-3), Natl. Toxicol. Program. Tech. Rep. Ser. 198, 1-187. NTP 1982a. Carcinogenesis Bioassay of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (CAS No. 1746-01-6) in Osborne-Mendel Rats and B6C3F1 Mice (Gavage Study), Natl. Toxicol. Program.Tech.Rep.Ser.. 209, 1-195. NTP 1982b. Carcinogenesis Bioassay of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (CAS No. 1746-01-6) in Swiss-Webster Mice (Dermal Study), Natl. Toxicol. Program.Tech.Rep.Ser.. 201, 1-113. NTP 2006a. NTP technical report on the toxicology and carcinogenesis studies of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (CAS No. 1746-01-6) in female Harlan Sprague-Dawley rats (Gavage Studies), Natl. Toxicol. Program.Tech.Rep.Ser.. 4-232. NTP 2006b. NTP Toxicology and Carcinogenesis Studies of 2,3,4,7,8-Pentachlorodibenzofuran (PeCDF) (CAS No. 57117-31-4) in Female Harlan Sprague-Dawley Rats (Gavage Studies), Natl. Toxicol. Program.Tech.Rep.Ser.. 1-198. NTP 2006c. NTP toxicology and carcinogenesis studies of 3,3',4,4',5-pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) in female Harlan Sprague-Dawley rats (Gavage Studies), Natl. Toxicol. Program.Tech.Rep.Ser.. 4-246. NTP 2006d. NTP technical report on the toxicology and carcinogenesis studies of 2,2',4,4',5,5'-hexachlorobiphenyl (PCB 153) (CAS No. 35065-27-1) in female Harlan Sprague-Dawley rats (Gavage studies), Natl. Toxicol. Program.Tech.Rep.Ser.. 4-168. NTP 2006e. NTP Toxicology and Carcinogenesis Studies of a Binary Mixture of 3,3',4,4',5-Pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) and 2,2',4,4',5,5'-Hexachlorobiphenyl (PCB 153) (CAS No. 35065-27-1) in Female Harlan Sprague-Dawley Rats (Gavage Studies), Natl. Toxicol. Program.Tech.Rep.Ser.. 1-258. NTP 2006f. NTP Toxicology and Carcinogenesis Studies of a Mixture of 2,3,7,8-Tetrachlorodibenzo-p-Dioxin (TCDD) (CAS No. 1746-01-6), 2,3,4,7,8-Pentachlorodibenzofuran (PeCDF) (CAS No. 57117-31-4), and 3,3',4,4',5-Pentachlorobiphenyl (PCB 126) (CAS No. 57465-28-8) in Female Harlan Sprague-Dawley Rats (Gavage Studies), Natl. Toxicol. Program.Tech.Rep.Ser.. 1-180. NTP 2010. Toxicology and carcinogenesis studies of 2,3',4,4',5-pentachlorobiphenyl (PCB 118) (CAS No. 31508-00-6) in female harlan Sprague-Dawley rats (gavage studies), Natl. Toxicol. Program. Tech. Rep. Ser. 1-174. NTP, 2014. NTP toxicology studies of indole-3-carbinol (CAS No. 700-06-01) in F344/N rats and B6C3F1/N mice and toxicology and carcinogenesis studies of indole-3-carbinol in Harlan Sprague-Dawley rats and B6C3F1/N mice (Gavage studies). Natl. Toxicol. Program Tech. Rep. Ser. 1-258. Nguyen L.P., Bradfield C.A. 2008. The search for endogenous activators of the aryl hydrocarbon receptor, Chem. Res. Toxicol. 21, 102-116. Okey A.B. 2007. An Aryl Hydrocarbon Receptor Odyssey to the Shores of Toxicology: The Deichmann Lecture, International Congress of Toxicology-XI, Toxicol. Sci. 98, 5-38. Organisation for Economic Cooperation and Development. 2013. Guidance Document on Developing and Assessing Adverse Outcome Pathways. ENV/JM/MONO(2013)6. Paris. At <a class="external free" href="http://www.oecd.org/officialdocuments/publicdisplaydocumentpdf/?cote=env/jm/mono(2013)6&doclanguage=en" rel="nofollow" target="_blank">http://www.oecd.org/officialdocuments/publicdisplaydocumentpdf/?cote=env/jm/mono(2013)6&doclanguage=en</a> Organisation for Economic Cooperation and Development. 2014. Users' Handbook Supplement to the Guidance Document for Developing and Asessing AOPs. Supplement to ENV/JM/MONO(2013)6. Paris. Organisation for Economic Cooperation and Development. 2015. Adverse Outcome Pathways, Molecular Screening and Toxicogenomics at <a class="external free" href="http://www.oecd.org/chemicalsafety/testing/adverse-outcome-pathways-molecular-screening-and-toxicogenomics.htm" rel="nofollow" target="_blank">http://www.oecd.org/chemicalsafety/testing/adverse-outcome-pathways-molecular-screening-and-toxicogenomics.htm</a>. Accessed March 17, 2015. Ovando, B.J., Ellison, C.A., Vezina, C.M., Olson, J.R., 2010. Toxicogenomic analysis of exposure to TCDD, PCB126 and PCB153: identification of genomic biomarkers of exposure to AhR ligands, BMC. Genomics 11, 583. <a class="external free" href="http://dx.doi.org/10.1186/" rel="nofollow" target="_blank">http://dx.doi.org/10.1186/</a> 1471-2164-11-583, 1471-2164-11-583 [pii]. Patlewicz G., Simon T., Goyak K., Phillips R.D., Rowlands J.C., Seidel S., Becker R.A. 2013. Use and validation of HT/HC assays to support 21st century toxicity evaluations, Regul. Toxicol. Pharmacol. 65, 259-268. Patlewicz, G., Simon, T. W., Rowlands, J. C., Budinsky, R. A., Becker, R. A., 2015. Proposing a scientific confidence framework to help support the application of adverse outcome pathways for regulatory purposes. Regul Toxicol Pharmacol. 71, 463-477. Peterson, S., Schwarz, Y., Li, S.S., Li, L., King, I.B., Chen, C., Eaton, D.L., Potter, J.D., Lampe, J.W., 2009. CYP1A2, GSTM1, and GSTT1 polymorphisms and diet effects on CYP1A2 activity in a crossover feeding trial. Cancer Epidemiol. Biomarkers Prev. 18, 3118-3125. Petkov P.I., Rowlands J.C., Budinsky R., Zhao B., Denison M.S., Mekenyan O. 2010. Mechanism-based common reactivity pattern (COREPA) modelling of aryl hydrocarbon receptor binding affinity, SAR. QSAR. Environ. Res. 21, 187-214. Pintilie D.G., Shupe T.D., Oh S.-H., Salganik S.V., Darwiche H., Petersen B.E. 2010. Hepatic stellate cells' involvement in progenitor-mediated liver regeneration, Lab. Invest. 90, 1199-1208. Poland, A., Glover, E., 1979. An estimate of the maximum in vivo covalent binding of 2,3,7,8-tetrachlorodibenzo-p-dioxin to rat liver protein, ribosomal RNA, and DNA. Cancer. Res. 39, 3341-3344. Randerath, K., Putman, K.L., Randerath, E., Zacharewski, T., Harris, M., Safe, S., 1990. Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on I-compounds in hepatic DNA of Sprague-Dawley rats: sex-specific effects and structure-activity relationships. Toxicol. Appl. Pharmacol. 103, 271-280. Schrenk, D., Schmitz, H.J., Bohnenberger, S., Wagner, B., Worner, W., 2004. Tumor promoters as inhibitors of apoptosis in rat hepatocytes. Toxicol. Lett. 149, 43-50. Schwarz, M., Buchmann, A., Stinchcombe, S., Kalkuhl, A., Bock, K., 2000. Ah receptor ligands and tumor promotion: survival of neoplastic cells. Toxicol. Lett. 112e113, 69-77. Seed, J., Carney, E.W., Corley, R.A., Crofton, K.M., DeSesso, J.M., Foster, P.M., Kavlock, R., Kimmel, G., Klaunig, J., Meek, M.E., Preston, R.J., Slikker, W.J., Tabacova, S., Williams, G.M., Wiltse, J., Zoeller, R.T., Fenner-Crisp, P., Patton, D.E., 2005. Overview: using mode of action and life stage information to evaluate the human relevance of animal toxicity data. Crit. Rev. Toxicol. 35, 664-672. Silkworth, J.B., Koganti, A., Illouz, K., Possolo, A., Zhao, M., Hamilton, S.B., 2005. Comparison of TCDD and PCB CYP1A induction sensitivities in fresh hepato- cytes from human donors, sprague-dawley rats, and rhesus monkeys and HepG2 cells. Toxicol. Sci. 87, 508-519. Simon T.W., Simons S.S., Preston R.J., Boobis A.R., Cohen S.M., Doerrer N.G., Fenner-Crisp P.A., McMullin T.S., McQueen C.A., Rowlands J.C., RISK21 Dose-Response Subteam 2014. The use of mode of action information in risk assessment: Quantitative key events/dose-response framework for modeling the dose-response for key events, Crit. Rev. Toxicol. 44 Suppl 3, 17- 43. Sobus, J. R., Tan, Y.-M., Pleil, J. D., Sheldon, L. S., 2011. A biomonitoring framework to support exposure and risk assessments. Sci Total Environ. 409, 4875-84. Sonich-Mullin, C., Fielder, R., Wiltse, J., Baetcke, K., Dempsey, J., Fenner-Crisp, P., Grant, D., Hartley, M., Knaap, A., Kroese, D., Mangelsdorf, I., Meek, E., Rice, J.M., Younes, M., International Programme on Chemical Safety, 2001. IPCS conceptual framework for evaluating a mode of action for chemical carcinogenesis. Regul. Toxicol. Pharmacol. 34, 146-152. Stinchcombe S., Buchmann A., Bock K.W., Schwarz M. 1995. Inhibition of apoptosis during 2,3,7,8-tetrachlorodibenzo-p-dioxin-mediated tumour promotion in rat liver, Carcinogenesis. 16, 1271-1275. Sutter, C.H., Bodreddigari, S., Sutter, T.R., Carlson, E.A., Silkworth, J.B., 2010. Analysis of the CYP1A1 mRNA dose response in human keratinocytes indicates that relative potencies of dioxins, furans, and PCBs are species and congener specific. Toxicol. Sci. 118, 704-715. Tanaka M., Itoh T., Tanimizu N., Miyajima A. 2011. Liver stem/progenitor cells: their characteristics and regulatory mechanisms, J. Biochem. 149, 231-239. Tang, N.J., Liu, J., Coenraads, P.J., Dong, L., Zhao, L.J., Ma, S.W., Chen, X., Zhang, C.M., Ma, X.M., Wei, W.G., Zhang, P., Bai, Z.P., 2008. Expression of AhR, CYP1A1, GSTA1, c-fos and TGF-alpha in skin lesions from dioxin-exposed humans with chlor- acne. Toxicol. Lett. 177, 182-187. Teeguarden J.G., Dragan Y.P., Singh J., Vaughan J., Xu Y.H., Goldsworthy T., Pitot H.C. 1999. Quantitative analysis of dose- and time-dependent promotion of four phenotypes of altered hepatic foci by 2,3,7,8-tetrachlorodibenzo-p-dioxin in female Sprague-Dawley rats, Toxicol. Sci. 51, 211-223. Tomasetti C., Vogelstein B. 2015. Cancer etiology. Variation in cancer risk among tissues can be explained by the number of stem cell divisions, Science. 347, 78-81. Turteltaub, K.W., Felton, J.S., Gledhill, B.L., Vogel, J.S., Southon, J.R., Caffee, M.W., Finkel, R.C., Nelson, D.E., Proctor, I.D., Davis, J.C., 1990. Accelerator mass spectrometry in biomedical dosimetry: relationship between low-level exposure and covalent binding of heterocyclic amine carcinogens to DNA. Proc. Natl. Acad. Sci. U. S. A. 87, 5288-5292. United States Environmental Protection Agency (USEPA). 2005. Guidelines for Carcinogen Risk Assessment. Uno, S., Endo, K., Ishida, Y., Tateno, C., Makishima, M., Yoshizato, K., Nebert, D.W., 2009. CYP1A1 and CYP1A2 expression: comparing ‘humanized’ mouse lines and wild-type mice; comparing human and mouse hepatoma-derived cell lines. Toxicol. Appl. Pharmacol. 237, 119-126. Van den Berg M., Birnbaum L.S., Denison M., De V.M., Farland W., Feeley M., Fiedler H., Hakansson H., Hanberg A., Haws L., Rose M., Safe S., Schrenk D., Tohyama C., Tritscher A., Tuomisto J., Tysklind M., Walker N., Peterson R.E. 2006. The 2005 World Health Organization reevaluation of human and Mammalian toxic equivalency factors for dioxins and dioxin-like compounds, Toxicol.Sci.. 93, 223-241. Villeneuve, D. L., Crump, D., Garcia-Reyero, N., Hecker, M., Hutchinson, T. H., LaLone, C. A., Landesmann, B., Lettieri, T., Munn, S., Nepelska, M., Ottinger, M. A., Vergauwen, L., Whelan, M., 2014a. Adverse outcome pathway (AOP) development I: strategies and principles. Toxicol Sci. 142, 312-20. Villeneuve, D. L., Crump, D., Garcia-Reyero, N., Hecker, M., Hutchinson, T. H., LaLone, C. A., Landesmann, B., Lettieri, T., Munn, S., Nepelska, M., Ottinger, M. A., Vergauwen, L., Whelan, M., 2014b. Adverse outcome pathway development II: best practices. Toxicol Sci. 142, 321-30. Wang X., Foster M., Al-Dhalimy M., Lagasse E., Finegold M., Grompe M. 2003. The origin and liver repopulating capacity of murine oval cells, Proc. Natl. Acad. Sci. U. S. A. 100 Suppl 1, 11881-11888. Wang Y.-J., Chang H., Kuo Y.-C., Wang C.-K., Siao S.-H., Chang L.W., Lin P. 2011. Synergism between 2,3,7,8-tetrachlorodibenzo-p-dioxin and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone on lung tumor incidence in mice, J. Hazard. Mater. 186, 869-875. Wassom, J.S., Huff, J.E., Loprieno, N., 1977. A review of the genetic toxicology of chlorinated dibenzo-p-dioxins. Mutat. Res. 47, 141-160. Weiss, C., Faust, D., Schreck, I., Ruff, A., Farwerck, T., Melenberg, A., Schneider, S., Oesch-Bartlomowicz, B., Zatloukalova, J., Vondracek, J., Oesch, F., Dietrich, C., 2008. TCDD deregulates contact inhibition in rat liver oval cells via Ah receptor, JunD and cyclin A. Oncogene 27, 2198-2207. Westerink, W.M., Stevenson, J.C., Schoonen, W.G., 2008. Pharmacologic profiling of human and rat cytochrome P450 1A1 and 1A2 induction and competition. Arch. Toxicol. 82, 909-921. Whysner, J., Williams, G.M., 1996. 2,3,7,8-Tetrachlorodibenzo-p-dioxin mechanistic data and risk assessment: gene regulation, cytotoxicity, enhanced cell proliferation, and tumor promotion. Pharmacol. Ther. 71, 193-223, Wincent, E., Amini, N., Luecke, S., Glatt, H., Bergman, J., Crescenzi, C., Rannug, A., Rannug, U., 2009. The suggested physiologic aryl hydrocarbon receptor acti- vator and cytochrome P4501 substrate 6-formylindolo[3,2-b]carbazole is pre- sent in humans. J. Biol. Chem. 284, 2690-2696. Wolfle D., Becker E., Schmutte C. 1993. Growth stimulation of primary rat hepatocytes by 2,3,7,8-tetrachlorodibenzo-p-dioxin, Cell. Biol. Toxicol. 9, 15-31. Xu, L., Li, A. P., Kaminski, D. L., Ruh, M. F., 2000. 2,3,7,8 Tetrachlorodibenzo-p-dioxin induction of cytochrome P4501A in cultured rat and human hepatocytes 1. Chem Biol Interact. 124, 173-189 Yamamoto, R., Shimamoto, K., Ishii, Y., Kimura, M., Fujii, Y., Morita, R., Suzuki, K., Shibutani, M., Mitsumori, K., 2013. Involvement of PTEN/Akt signaling and oxidative stress on indole-3-carbinol (I3C)-induced hepatocarcinogenesis in rats. Exp. Toxicol. Pathol. 65, 845-852. Yao, C.Q., Prokopec, S.D., Watson, J.D., Pang, R., P'ng, C., Chong, L.C., Harding, N.J., Pohjanvirta, R., Okey, A.B., Boutros, P.C., 2012. Inter-strain heterogeneity in rat hepatic transcriptomic responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Toxicol. Appl. Pharmacol. 260, 135-145. Zhao, Y.-Y., Tao, F.-M., Zeng, E.Y., 2008. Theoretical study of the quantitative structure-activity relationships for the toxicity of dibenzo-p-dioxins. Chemosphere 73, 86-91. AOPWiki 2016-11-29T18:41:16

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