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Event: 1887

Key Event Title

The KE title should describe a discrete biological change that can be measured. It should generally define the biological object or process being measured and whether it is increased, decreased, or otherwise definably altered relative to a control state. For example “enzyme activity, decreased”, “hormone concentration, increased”, or “growth rate, decreased”, where the specific enzyme or hormone being measured is defined. More help

Inhibition of CYP26B1 activity

Short name
The KE short name should be a reasonable abbreviation of the KE title and is used in labelling this object throughout the AOP-Wiki. The short name should be less than 80 characters in length. More help
CYP26B1, inhibition

Biological Context

Structured terms, selected from a drop-down menu, are used to identify the level of biological organization for each KE. Note, KEs should be defined within a particular level of biological organization. Only KERs should be used to transition from one level of organization to another. Selection of the level of biological organization defines which structured terms will be available to select when defining the Event Components (below). More help
Level of Biological Organization
Molecular

Cell term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help

Organ term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help

Key Event Components

Further information on Event Components and Biological Context may be viewed on the attached pdf.Because one of the aims of the AOP-KB is to facilitate de facto construction of AOP networks through the use of shared KE and KER elements, authors are also asked to define their KEs using a set of structured ontology terms (Event Components). In the absence of structured terms, the same KE can readily be defined using a number of synonymous titles (read by a computer as character strings). In order to make these synonymous KEs more machine-readable, KEs should also be defined by one or more “event components” consisting of a biological process, object, and action with each term originating from one of 22 biological ontologies (Ives, et al., 2017; See List). Biological process describes dynamics of the underlying biological system (e.g., receptor signalling). The biological object is the subject of the perturbation (e.g., a specific biological receptor that is activated or inhibited). Action represents the direction of perturbation of this system (generally increased or decreased; e.g., ‘decreased’ in the case of a receptor that is inhibited to indicate a decrease in the signalling by that receptor).Note that when editing Event Components, clicking an existing Event Component from the Suggestions menu will autopopulate these fields, along with their source ID and description. To clear any fields before submitting the event component, use the 'Clear process,' 'Clear object,' or 'Clear action' buttons. If a desired term does not exist, a new term request may be made via Term Requests. Event components may not be edited; to edit an event component, remove the existing event component and create a new one using the terms that you wish to add. More help

Key Event Overview

AOPs Including This Key Event

All of the AOPs that are linked to this KE will automatically be listed in this subsection. This table can be particularly useful for derivation of AOP networks including the KE. Clicking on the name of the AOP will bring you to the individual page for that AOP. More help

Stressors

This is a structured field used to identify specific agents (generally chemicals) that can trigger the KE. Stressors identified in this field will be linked to the KE in a machine-readable manner, such that, for example, a stressor search would identify this as an event the stressor can trigger. NOTE: intermediate or downstream KEs in one AOP may function as MIEs in other AOPs, meaning that stressor information may be added to the KE description, even if it is a downstream KE in the pathway currently under development.Information concerning the stressors that may trigger an MIE can be defined using a combination of structured and unstructured (free-text) fields. For example, structured fields may be used to indicate specific chemicals for which there is evidence of an interaction relevant to this MIE. By linking the KE description to a structured chemical name, it will be increasingly possible to link the MIE to other sources of chemical data and information, enhancing searchability and inter-operability among different data-sources and knowledgebases. The free-text section “Evidence for perturbation of this MIE by stressor” can be used both to identify the supporting evidence for specific stressors triggering the MIE as well as to define broad chemical categories or other properties that classify the stressors able to trigger the MIE for which specific structured terms may not exist. More help

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) can be selected from an ontology. In many cases, individual species identified in these structured fields will be those for which the strongest evidence used in constructing the AOP was available in relation to this KE. More help
Term Scientific Term Evidence Link
Human, rat, mouse Human, rat, mouse Moderate NCBI

Life Stages

The structured ontology terms for life-stage are more comprehensive than those for taxa, but may still require further description/development and explanation in the free text section. More help
Life stage Evidence
All life stages Moderate

Sex Applicability

The authors must select from one of the following: Male, female, mixed, asexual, third gender, hermaphrodite, or unspecific. More help
Term Evidence
Male High
Female High

Key Event Description

A description of the biological state being observed or measured, the biological compartment in which it is measured, and its general role in the biology should be provided. For example, the biological state being measured could be the activity of an enzyme, the expression of a gene or abundance of an mRNA transcript, the concentration of a hormone or protein, neuronal activity, heart rate, etc. The biological compartment may be a particular cell type, tissue, organ, fluid (e.g., plasma, cerebrospinal fluid), etc. The role in the biology could describe the reaction that an enzyme catalyses and the role of that reaction within a given metabolic pathway; the protein that a gene or mRNA transcript codes for and the function of that protein; the function of a hormone in a given target tissue, physiological function of an organ, etc. Careful attention should be taken to avoid reference to other KEs, KERs or AOPs. Only describe this KE as a single isolated measurable event/state. This will ensure that the KE is modular and can be used by other AOPs, thereby facilitating construction of AOP networks. More help

CYP26B1 and its function

Numerous physiological processes as well as fetal development is dependent on correct levels of retinoic acid (RA). RA homeostasis is tightly regulated and involves, amongst many other factors, breakdown by the Cytochrome P450 enzymes of family 26 (Kedishvili, 2013; White et al, 1997; White et al, 1996). The family consists of the enzymes CYP26A1, CYP26B1, and CYP26C1 (Isoherranen & Zhong, 2019). These RA-metabolizing enzymes convert all-trans-RA to more polar metabolites, including 4-hydroxy-RA and 4-oxo-RA (White et al, 1996; White et al, 2000).

CYP26B1 is particularly important for the clearance of RA in the fetal mouse testis, with knockout models having established its critical role in normal testis development in preventing premature meiotic initiation of male germ cells (Bowles et al, 2018; Bowles et al, 2006; Li et al, 2009; MacLean et al, 2007). CYP26B1 is necessary for steroidogenesis and male reproductive tract formation (Bowles et al, 2018), and may also play a role in maintaining normal spermatogenesis in the adult male mouse (Hogarth et al, 2015). CYP26B1 has been shown to be expressed in the postnatal mouse ovary, where it is proposed to regulate granulosa cell proliferation (Kipp et al, 2011).

In addition to reproductive development, CYP26B1 is important for other aspects of embryonic development. The expression of CYP26B1 is initiated at embryonic day 8 in the mouse hindbrain and is expressed in the limb buds from the beginning of their outgrowth as well as later in many different tissues during organ development including the genital tubercle, craniofacial areas, and spinal cord (Abu-Abed et al, 2002; MacLean et al, 2001; Ross & Zolfaghari, 2011). Consistent with this, Cyp26b1-knockout mice display various developmental defects including severe limb malformations and pups born alive die right after birth due to respiratory distress (Yashiro et al, 2004)

Besides from the important role CYP26B1 plays during fetal development, CYP26B1 is also expressed in multiple tissues in the adult human including adipose tissue, bladder, blood vessels, brain, kidney, adrenals, liver, lung, pancreas, intestines, testis, uterus, and skin (The-Human-Protein-Atlas, 2021; Topletz et al, 2012).

CYP26B1 inhibition as Key Event

The main function of CYP26B1 is to inactivate all-trans RA. The major primary metabolites formed from RA by CYP26B1 are 4-OH-RA and 18-OH-RA (Topletz et al, 2012) and CYP26B1 shows preference for the following substrates: all-trans-RA > 9-cis-RA > 13-cis-RA (Topletz et al, 2012; White et al, 2000). CYP26B1 is regulated by a range of molecular factors but also by RA itself (Isoherranen & Zhong, 2019; White et al, 2000).

How It Is Measured or Detected

One of the primary considerations in evaluating AOPs is the relevance and reliability of the methods with which the KEs can be measured. The aim of this section of the KE description is not to provide detailed protocols, but rather to capture, in a sentence or two, per method, the type(s) of measurements that can be employed to evaluate the KE and the relative level of scientific confidence in those measurements. Methods that can be used to detect or measure the biological state represented in the KE should be briefly described and/or cited. These can range from citation of specific validated test guidelines, citation of specific methods published in the peer reviewed literature, or outlines of a general protocol or approach (e.g., a protein may be measured by ELISA).Key considerations regarding scientific confidence in the measurement approach include whether the assay is fit for purpose, whether it provides a direct or indirect measure of the biological state in question, whether it is repeatable and reproducible, and the extent to which it is accepted in the scientific and/or regulatory community. Information can be obtained from the OECD Test Guidelines website and the EURL ECVAM Database Service on Alternative Methods to Animal Experimentation (DB-ALM). ?

There are no OECD validated assays for measuring CYP26B1 inhibition.

CYP26b1 mRNA and protein levels can be measured using various probes, antibodies as well as ELISA kits that are commercially available.

Enzyme activity can be measured in different types of assays including using microsomes measuring conversion of RA to metabolites (Van Wauwe et al, 1988).

Domain of Applicability

This free text section should be used to elaborate on the scientific basis for the indicated domains of applicability and the WoE calls (if provided). While structured terms may be selected to define the taxonomic, life stage and sex applicability (see structured applicability terms, above) of the KE, the structured terms may not adequately reflect or capture the overall biological applicability domain (particularly with regard to taxa). Likewise, the structured terms do not provide an explanation or rationale for the selection. The free-text section on evidence for taxonomic, life stage, and sex applicability can be used to elaborate on why the specific structured terms were selected, and provide supporting references and background information.  More help

CYP26B1 is highly evolutionary conserved with e.g. a human to mouse sequence homology of 93% (Thatcher & Isoherranen, 2009).

This KE is applicable for both sexes, across developmental stages into adulthood, in numerous cells and tissues and across taxa.

Evidence for Perturbation by Stressor

Ketoconazole

Ketoconazole: Using tazarotenic acid as a substrate in vitro incubations and metabolite production measured by LC-MS/MS showed an IC50 of 190 nM (Foti et al, 2016). Using hamster liver microsomes ketoconazole decreased RA metabolite formation and in vivo, oral ketoconazole administration suppressed metabolite formation after RA dosing in rats (Van Wauwe et al, 1988).

Clotrimazole

Clotrimazole: Using tazarotenic acid as a substrate in vitro incubations and metabolite production measured by LC-MS/MS showed an IC50 of 50 nM (Foti et al, 2016).

Itraconazole

Itraconazole: Using tazarotenic acid as a substrate in vitro incubations and metabolite production measured by LC-MS/MS showed an IC50 of 160 nM (Foti et al, 2016).

Fluconazole

Fluconazole: mRNA induction in mouse embryos after in utero exposure (Tiboni et al, 2009) and using tazarotenic acid as a substrate in vitro incubations and metabolite production measured by LC-MS/MS showed an IC50 of 19.8 µM (Foti et al, 2016).

Candesartan cilexetil

Candesartan Cilexetil: Using tazarotenic acid as a substrate in vitro incubations and metabolite production measured by LC-MS/MS showed an IC50 of 270 nM (Foti et al, 2016).

Montelukast

Montelukast: Using tazarotenic acid as a substrate in vitro incubations and metabolite production measured by LC-MS/MS showed an IC50 of 610 nM (Foti et al, 2016).

Repaglinide

Repaglinide: Using tazarotenic acid as a substrate in vitro incubations and metabolite production measured by LC-MS/MS showed an IC50 of 610 nM (Foti et al, 2016).

References

List of the literature that was cited for this KE description. Ideally, the list of references, should conform, to the extent possible, with the OECD Style Guide (https://www.oecd.org/about/publishing/OECD-Style-Guide-Third-Edition.pdf) (OECD, 2015). More help

Abu-Abed S, MacLean G, Fraulob V, Chambon P, Petkovich M, Dollé P (2002) Differential expression of the retinoic acid-metabolizing enzymes CYP26A1 and CYP26B1 during murine organogenesis. Mech Dev 110: 173-177

Bowles J, Feng CW, Inseson J, Miles K, Spiller CM, Harley VR, Sinclair AH, Koopman P (2018) Retinoic Acid Antagonizes Testis Development in Mice. Cell Rep 24: 1330-1341

Bowles J, Knight D, Smith C, Wilhelm D, Richman J, Mamiya S, Yashiro K, Chawengsaksophak K, Wilson MJ, Rossant J, Hamada H, Koopman P (2006) Retinoid signaling determines germ cell fate in mice. Science 312: 596-600

Foti RS, Diaz P, Douguet D (2016) Comparison of the ligand binding site of CYP2C8 with CYP26A1 and CYP26B1: a structural basis for the identification of new inhibitors of the retinoic acid hydroxylases. J Enzyme Inhib Med Chem 31: 148-161

Hogarth CA, Evans E, Onken J, Kent T, D. M, Petkovich M, Griswold MD (2015) CYP26 Enzymes Are Necessary Within the Postnatal Seminiferous Epithelium for Normal Murine Spermatogenesis. Biol Reprod 93: 19

Isoherranen N, Zhong G (2019) Biochemical and physiological importance of the CYP26 retinoic acid hydroxylases. Pharmacol Ther 204: 107400

Kedishvili NY (2013) Enzymology of retinoic acid biosynthesis and degradation. J Lipid Res 54: 1744-1760

Kipp JL, Golebiowski A, Rodriguez G, Demczuk M, Kilen SM, Mayo KE (2011) Gene expression profiling reveals Cyp26b1 to be an activin regulated gene involved in ovarian granulosa cell proliferation. Endocrinology 152: 303-312

Li H, MacLean G, Cameron D, Clagett-Dame M, Petkovich M (2009) Cyp26b1 expression in murine Sertoli cells is required to maintain male germ cells in an undifferentiated state during embryogenesis. PLoS One 4: e7501

MacLean G, Abu-Abed S, Dollé P, Tahayato A, Chambon P, Petkovich M (2001) Cloning of a novel retinoic-acid metabolizing cytochrome P450, Cyp26B1, and comparative expression analysis with Cyp26A1 during early murine development. Mech Dev 107: 195-201

MacLean G, Li H, Metzger D, Chambon P, Petkovich M (2007) Apoptotic extinction of germ cells in testes of Cyp26b1 knockout mice. Endocrinology 148: 4560-4567

Nelson CH, Buttrick BR, Isoherranen N (2013) Therapeutic potential of the inhibition of the retinoic acid hydroxylases CYP26A1 and CYP26B1 by xenobiotics. Curr Top Med Chem 13: 1402-1428

Ross AC, Zolfaghari R (2011) Cytochrome P450s in the regulation of cellular retinoic acid metabolism. Annu Rev Nutr 31: 65-87

Thatcher JE, Isoherranen N (2009) The role of CYP26 enzymes in retinoic acid clearance. Expert Opin Drug Metab Toxicol 5: 875-886

The-Human-Protein-Atlas. (2021) Tissue expression of CYP26B1.

Tiboni GM, Marotta F, Carletti E (2009) Fluconazole alters CYP26 gene expression in mouse embryos. Reprod Toxicol 27: 199-202

Topletz AR, Thatcher JE, Zelter A, Lutz JD, Tay S, Nelson WL, Isoherranen N (2012) Comparison of the function and expression of CYP26A1 and CYP26B1, the two retinoic acid hydroxylases. Biochem Pharmacol 83: 149-163

Van Wauwe JP, Coene MC, Goossens J, Van Nijen G, Lauwers W (1988) Ketoconazole inhibits the in vitro and in vivo metabolism of all-trans-retinoic acid. J Pharmacol Exp Ther 245: 718-722

White JA, Beckett-Jones B, Guo YD, Dilworth FJ, Bonasoro J, Jones G, Petkovich M (1997) cDNA cloning of human retinoic acid-metabolizing enzyme (hP450RAI) identifies a novel family of cytochromes P450. J Biol Chem 272: 18538-18541

White JA, Guo YD, Baetz K, Beckett-Jones B, Bonasoro J, Hsu KE, Dilworth FJ, Jones G, Petkovich M (1996) Identification of the retinoic acid-inducible all-trans-retinoic acid 4-hydroxylase. J Biol Chem 271: 29922-29927

White JA, Ramshaw H, Taimi M, Stangle W, Zhang A, Everingham S, Creighton S, Tam SP, Jones G, Petkovich M (2000) Identification of the human cytochrome P450, P450RAI-2, which is predominantly expressed in the adult cerebellum and is responsible for all-trans-retinoic acid metabolism. Proc Natl Acad Sci U S A 97: 6403-6408

Yashiro K, Zhao X, Uehara M, Yamashita K, Nishijima M, Nishino J, Saijoh Y, Sakai Y, Hamada H (2004) Regulation of retinoic acid distribution is required for proximodistal patterning and outgrowth of the developing mouse limb. Dev Cell 6: 411-422