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Relationship: 1268

Title

A descriptive phrase which clearly defines the two KEs being considered and the sequential relationship between them (i.e., which is upstream, and which is downstream). More help

Activation, Constitutive androstane receptor leads to Altered expression of hepatic CAR-dependent genes

Upstream event
The causing Key Event (KE) in a Key Event Relationship (KER). More help
Downstream event
The responding Key Event (KE) in a Key Event Relationship (KER). More help

Key Event Relationship Overview

The utility of AOPs for regulatory application is defined, to a large extent, by the confidence and precision with which they facilitate extrapolation of data measured at low levels of biological organisation to predicted outcomes at higher levels of organisation and the extent to which they can link biological effect measurements to their specific causes. Within the AOP framework, the predictive relationships that facilitate extrapolation are represented by the KERs. Consequently, the overall WoE for an AOP is a reflection in part, of the level of confidence in the underlying series of KERs it encompasses. Therefore, describing the KERs in an AOP involves assembling and organising the types of information and evidence that defines the scientific basis for inferring the probable change in, or state of, a downstream KE from the known or measured state of an upstream KE. More help

AOPs Referencing Relationship

AOP Name Adjacency Weight of Evidence Quantitative Understanding Point of Contact Author Status OECD Status
Constitutive androstane receptor activation leading to hepatocellular adenomas and carcinomas in the mouse and the rat adjacent High Moderate Brendan Ferreri-Hanberry (send email) Open for citation & comment EAGMST Under Review

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) that help to define the biological applicability domain of the KER.In general, this will be dictated by the more restrictive of the two KEs being linked together by the KER.  More help
Term Scientific Term Evidence Link
rat Rattus norvegicus High NCBI
mouse Mus musculus High NCBI
human Homo sapiens Moderate NCBI

Sex Applicability

An indication of the the relevant sex for this KER. More help
Sex Evidence
Mixed High

Life Stage Applicability

An indication of the the relevant life stage(s) for this KER.  More help
Term Evidence
All life stages High

Key Event Relationship Description

Provides a concise overview of the information given below as well as addressing details that aren’t inherent in the description of the KEs themselves. More help

Activation of CAR by an endogenous or foreign substance leads to translocation of the CAR-RXR heterodimer into the nucleus, and this dimer binds to DNA recognition elements in the regulatory region of CAR-responsive genes. CAR activation thus alters gene expression and upregulates xenobiotic-metabolizing enzymes such as CYP2B, CYP2C, CYP3A, sulfotransferases, UDP-glucuronyltransferases and glutathione transferases, as well as xenobiotic transporters such as Mrp2 and Mrp4 (Omiecinski et al., 2011a). In addition, CAR alters genes involved in lipid homeostasis, glucose utilization and energy metabolism. In rats and mice, the expression of additional genes involved in cell proliferation and apoptosis control are altered; Gadd45beta and Cdc20 are examples of genes that function in this way and are upregulated in mice within hours of treatment with a CAR activator (Peffer et al., 2018aPeffer et al., 2018bTojima et al., 2012).

Evidence Collection Strategy

Include a description of the approach for identification and assembly of the evidence base for the KER.  For evidence identification, include, for example, a description of the sources and dates of information consulted including expert knowledge, databases searched and associated search terms/strings.  Include also a description of study screening criteria and methodology, study quality assessment considerations, the data extraction strategy and links to any repositories/databases of relevant references.Tabular summaries and links to relevant supporting documentation are encouraged, wherever possible. More help

Evidence Supporting this KER

Addresses the scientific evidence supporting KERs in an AOP setting the stage for overall assessment of the AOP. More help

For the MIE of CAR activation, direct measurement of this step after exposure to a stressor in vivo is difficult to attain, because of the complex sequence of intermediate steps that follow. Nuclear translocation of a fluorescently tagged CAR molecule following activation by a stressor such as phenobarbital has been observed in vitro in hepatocytes, and occurs fairly rapidly (i.e. within a few hours) (Chen et al., 2010). More commonly, indirect measurements of downstream effects, and absence of those effects in CAR-null mice or rats, are employed to demonstrate that a particular molecule or other stressor can activate CAR (Huang et al., 2005Peffer et al., 2018bPeffer et al., 2007Yamamoto et al., 2004).

Following the activation of CAR by stressors such as phenobarbital, TCPOBOP or metofluthrin in rats or mice, the in vivo and in vitro data are strong that demonstrate a characteristic pattern of changes in CAR-responsive genes. The data for these example molecules are described further within this KER.

Biological Plausibility
Addresses the biological rationale for a connection between KEupstream and KEdownstream.  This field can also incorporate additional mechanistic details that help inform the relationship between KEs, this is useful when it is not practical/pragmatic to represent these details as separate KEs due to the difficulty or relative infrequency with which it is likely to be measured.   More help

It is highly plausible that activation of CAR would produce changes in expression of specific genes, since transcription of genes specifically associated with CAR response elements is how this nuclear receptor achieves its biological effects (Tien and Negishi, 2006). In mice, rat and human hepatocytes, CAR activation produces changes in genes related to CYP2B and CYP3A enzymes, certain sulfotransferase and glucuronyltransferase enzymes, hepatic transport proteins and genes related to lipid and glucose metabolism (Maglich et al., 2003Omiecinski et al., 2011bOmiecinski et al., 2011aTien and Negishi, 2006). In mice and rats, CAR activation has been shown to also achieve alterations in genes that give an overall pathway change associated with increased progression through the cell cycle such as Gadd45b and Cdc20 (Deguchi et al., 2009Geter et al., 2014Ross et al., 2010Tojima et al., 2012).

Uncertainties and Inconsistencies
Addresses inconsistencies or uncertainties in the relationship including the identification of experimental details that may explain apparent deviations from the expected patterns of concordance. More help

In general, CAR activators show very consistent, large fold-increases for the characteristic expression of Cyp2b isoforms across in vivo studies in multiple species and with many different molecules. While certain genes related to a pro-proliferative effect appear to be CAR-mediated and reproducibly impacted in multiple studies (Currie et al., 2014Deguchi et al., 2009Geter et al., 2014Peffer et al., 2007Tojima et al., 2012), there are examples where changes in a specific gene was not observed. For example, Ross et al. (2010) tested 80 mg/kg/day (ip dosing) phenobarbital for 4 days in WT C57BL/6J mice, and they observed a 15.8-fold increase in Cdc20, but did not see an increased expression of Gadd45b. Mapping of a specific genes’ changes following activation of CAR by a particular CAR activator may be affected by the species, strain, dose level and time point examined, as well as the other non-CAR effects of that molecule. Examining for a significant pathway change is likely to be a more reliable measure of this Key Event Relationship (Oshida et al., 2015a), but this is also somewhat dependent on the experimental design, the species and duration of treatment, and the pathway analysis tools.

Known modulating factors

This table captures specific information on the MF, its properties, how it affects the KER and respective references.1.) What is the modulating factor? Name the factor for which solid evidence exists that it influences this KER. Examples: age, sex, genotype, diet 2.) Details of this modulating factor. Specify which features of this MF are relevant for this KER. Examples: a specific age range or a specific biological age (defined by...); a specific gene mutation or variant, a specific nutrient (deficit or surplus); a sex-specific homone; a certain threshold value (e.g. serum levels of a chemical above...) 3.) Description of how this modulating factor affects this KER. Describe the provable modification of the KER (also quantitatively, if known). Examples: increase or decrease of the magnitude of effect (by a factor of...); change of the time-course of the effect (onset delay by...); alteration of the probability of the effect; increase or decrease of the sensitivity of the downstream effect (by a factor of...) 4.) Provision of supporting scientific evidence for an effect of this MF on this KER. Give a list of references.  More help

Activation of PXR (NR1I2), a related nuclear receptor to CAR (NR1I3), is a possible confounding factor that may be operative for certain substances. There is much cross-talk between CAR and PXR, and similar responsive genes, and a particular agent could produce a mixed set of gene expression response by activating both PXR and CAR (Tojima et al., 2012; Stanley et al., 2006).

Response-response Relationship
Provides sources of data that define the response-response relationships between the KEs.  More help

Because the extent or degree of activation of CAR is not readily measurable with in vivo studies, the nature of the relationship between CAR activation and altered mRNA expression (e.g. linear, exponential, other) cannot be stated. In addition, the direction of change in certain CAR-responsive genes is variable, in that CAR activation causes an increase in expression of some genes and a decrease in expression of other genes. For example, in mice treated with TCPOBOP by ip injection, Tojima et al. (2012) showed large increases in expression of Cyp2b10 (151x), Cdc20 (37x) and Gadd45b (14x), plus decreases in expression of lipid-related genes Acsl5 (0.5x), Slc21a1 (0.2x) and Hmgcs2 (0.3x). The different fold-change values for specific genes also indicates that the magnitude of the gene expression differences can be very dependent on the properties of the individual gene, and whether it is normally active or quiescent prior to treatment with a CAR activator.

Time-scale
Information regarding the approximate time-scale of the changes in KEdownstream relative to changes in KEupstream (i.e., do effects on KEdownstream lag those on KEupstream by seconds, minutes, hours, or days?). More help

The onset of CAR activation (upstream KE) and the time scale of measurable differences in gene expression (downstream KE) are influenced by variables such as pharmacokinetics and route of administration of an administered stressor. However, it is established that exposure of the livers of mice to a CAR activator (TCPOBOP) by intraperitoneal injection, thus bypassing the intestinal transit and absorption processes, was able to achieve large changes in gene expression in the liver after 12 hours (Tojima et al., 2012). The cascade of normal biological events following CAR activation by a ligand (i.e. binding, nuclear translocation, altered gene expression, and post-transcriptional modifications of mRNA) likely takes several hours to be completed, after which a measurable difference vs. control in mRNA levels is detectable by RT-PCR or microarrays. After the initial perturbation by a stressor of CAR, the time course of changes in gene expression is indefinite. That is, some gene expression changes can continue to be observed for as long as the CAR activator is present. For example, in mice treated with phenobarbital for up to 32 weeks following initiation with a single dose of diethylnitrosamine, differences in gene expression of CAR-responsive genes such as Gadd45b and Cdc2 continued to be observed throughout the entire treatment period in both non-neoplastic liver tissue and in liver tumors of treated vs. untreated animals (Phillips et al., 2009a).

Known Feedforward/Feedback loops influencing this KER
Define whether there are known positive or negative feedback mechanisms involved and what is understood about their time-course and homeostatic limits. More help

In terms of this specific KER1 of activation of CAR (MIE) leading directly to altered expression of CAR-responsive genes (KE1), there are no known feedback loops that would affect this overall process.  As an example of this lack of feedback alteration, when mice were treated long-term with phenobarbital, they continued to show significant changes in expression of CAR-responsive genes up through 32 weeks of treatment (Phillips et al., 2009).

Domain of Applicability

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CAR receptors are present in the livers of virtually all mammalian species; however, there are important differences in protein sequence and thus ligand binding properties (Omiecinski et al., 2011bReschly and Krasowski, 2006). In reporter assays for mouse, rat, dog and human CAR, clear qualitative as well as quantitative differences in the ability of suspected CAR activators to activate CAR from the different species were demonstrated (Omiecinski et al., 2011b). In terms of the specific KER of CAR activation directly leading to altered gene expression specific to CAR activation, in vitro hepatocyte experiments indicate that human hepatocytes have only partial overlap with mice and rats in terms of the genes that are affected. In particular, genes that are related to CYP induction (e.g. Cyp2b isoforms) show increases in expression across mouse, rat and human if the CAR molecule for that species is activated, but the pro-proliferative gene pathways  have been shown to be activated only in mice and rats (Elcombe et al., 2014Hasmall and Roberts, 1999Hirose et al., 2009Lake, 2009).  For example, the total number of altered genes in livers of chimeric mice that reflected human hepatocytes (293) compared to livers of similarly treated CD-1 mice (846) was much lower, and only 10 differentially expressed genes (primarily CYP genes) were common to both species’ liver samples following treatment with phenobarbital at dose levels of 1000 – 2500 ppm in the diet (Yamada et al., 2014).

References

List of the literature that was cited for this KER description. More help

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