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Event: 279

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

Thyroperoxidase, Inhibition

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
Thyroperoxidase, 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
Cell term
thyroid follicular cell

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
Organ term
thyroid follicle

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
Process Object Action
iodide peroxidase activity thyroid peroxidase decreased

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
AOP Name Role of event in AOP Point of Contact Author Status OECD Status
TPO Inhibition and Altered Neurodevelopment MolecularInitiatingEvent Evgeniia Kazymova (send email) Open for citation & comment TFHA/WNT Endorsed
Thyroid peroxidase- follicular adenoma/carcinoma MolecularInitiatingEvent Brendan Ferreri-Hanberry (send email) Under Development: Contributions and Comments Welcome
TPOi anterior swim bladder MolecularInitiatingEvent Evgeniia Kazymova (send email) Open for adoption EAGMST Under Review
TPO inhib alters metamorphosis MolecularInitiatingEvent Brendan Ferreri-Hanberry (send email) Under Development: Contributions and Comments Welcome
TPO inhibition and impaired fertility MolecularInitiatingEvent Cataia Ives (send email) Open for comment. Do not cite Under Development
TPOi eye structure MolecularInitiatingEvent Allie Always (send email) Under development: Not open for comment. Do not cite

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
rat Rattus norvegicus High NCBI
humans Homo sapiens High NCBI
pigs Sus scrofa High NCBI
Xenopus laevis Xenopus laevis High NCBI
chicken Gallus gallus High NCBI
zebrafish Danio rerio High NCBI
fathead minnow Pimephales promelas High NCBI
mouse Mus musculus 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 High

Sex Applicability

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

Thyroperoxidase (TPO) is a heme-containing apical membrane protein within the follicular lumen of thyrocytes that acts as the enzymatic catalyst for thyroid hormone (TH) synthesis. TPO catalyzes several reactions in the thyroid gland, including: the oxidation of iodide; nonspecific iodination of tyrosyl residues of thyroglobulin (Tg); and, the coupling of iodotyrosyls to produce Tg-bound monoiodotyrosine (MIT) and diiodotyrosine (DIT) (Divi et al., 1997; Kessler et al., 2008; Ruf et al., 2006; Taurog et al., 1996). The outcome of TPO inhibition is decreased synthesis of thyroxine (T4) and triiodothyronine (T3), a decrease in release of these hormones from the gland into circulation, and unless compensated, a consequent decrease in systemic concentrations of T4, and possibly T3. The primary product of TPO-catalyzed TH synthesis is T4 (Taurog et al., 1996; Zoeller et al., 2007) that would be peripherally or centrally deiodinated to T3.

It is important to note that TPO is a complex enzyme and that has two catalytic cycles and is capable of iodinating multiple species (Divi et al., 1997). Alterations in all of these events are not covered by some of the commonly used assays that measure “TPO inhibition” (e.g., guicacol and AmplexUltraRed, see below).  Therefore, in the context of this AOP we are using TPO inhibiton not in the classical sense, but instead to refer to the empirical data derived from the assays commonly used assays to investigate environmental chemicals.

Figure 1 below illustrates the enzymatic and nonenzymatic reactions mediated by TPO that result in the synthesis of thyroxine (T4) .

Inhibition of TPO can be reversible, with transient interaction between the enzyme and the chemical, or irreversible, whereby suicide substrates permanently inactivate the enzyme. Reversible and irreversible TPO inhibition may be determined by the chemical structure, may be concentration dependent, or may be influenced by other conditions, including the availability of iodine (Doerge and Chang, 2002).

The ontogeny of TPO has been determined using both direct and indirect evidence.  Available evidence suggests the 11th to 12th fetal week as the beginning of functional TPO in humans. In rodents, TPO function begins late in the second fetal week, with the first evidence of T4 secretion on gestational day 17 (Remy et al., 1980). Thyroid-specific genes appear in the thyroid gland according to a specific temporal pattern; thyroglobulin (Tg), TPO (Tpo), and TSH receptor (Tshr) genes are expressed by gestational day 14 in rats, and the sodium iodide symporter, NIS (Nis), is expressed by gestational day 16 in rats. Maturation to adult function is thought to occur within a few weeks after parturition in rats and mice, and within the first few months in neonatal humans (Santisteban and Bernal, 2005).  Tg is first detected in human fetuses starting at 5th week of gestation and rises throughout gestation (Thorpe-Beeston et al., 1992), but iodine trapping and T4 production does not occur until around 10-12 weeks.  Also, the dimerization of Tg, a characteristic of adult TH storage, is not found until much later in human gestation (Pintar, 2000). In rats, Tg immunoreactivity does not appear until day 15 of gestation (Fukiishi et al., 1982; Brown et al., 2000). The vast majority of research and knowledge on Tg is from mammals, although genomic orthologs are known for a variety of other species (Holzer et al., 2016).  It is important to note that prior to the onset of fetal thyroid function, TH are still required by the developing fetus which until that time relies solely on maternal sources. Chemical-induced TPO inhibition can affect synthesis in the maternal gland and in the fetal gland.

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 approved OECD or EPA guideline study protocols for measurement of TPO inhibition.. However, there is an OECD scoping document on identification of chemicals that modulate TH signaling that provides details on a TPO assay (OECD, 2017). 

From the early 1960's, microsomal fractions prepared from porcine thyroid glands and isolated porcine follicles were used as a source of TPO for inhibition experiments (Taurog, 2005). Limited information has been published using microsomes from human goiter samples (Vickers et al., 2012) and rat thyroid glands (Paul et al., 2013; 2014; Paul-Friedman et al., 2016).

TPO activity has been measured for decades via indirect assessment by kinetic measurement of the oxidation of guaiacol (Chang & Doerge 2000; Hornung et al., 2010; Schmutzler et al., 2007).  This method is a low-throughput assay due to the very rapid kinetics of the guaiacol oxidation reaction. More recently, higher-throughput methods using commercial fluorescent and luminescent substrates with rodent, porcine, and human microsomal TPO have been developed (Vickers et al., 2012; Paul et al., 2013; 2014; Kaczur et al., 1997). This assay substitutes a pre-fluorescent substrate (Amplex UltraRed) for guaiacol, that when incubated with a source of peroxidase and excess hydrogen peroxidase, results in a stable fluorescent product proportional to TPO activity (Vickers et al., 2012).  The stability of the fluorescent reaction product allows this assay to be used in a higher throughput format (Paul-Friedman et al., 2016). This approach is appropriate for high-throughput screening but does not elucidate the specific mechanism by which a chemical may inhibit TPO (Paul-Friedman et al., 2016), and as with most in vitro assays, is subject to various sources of assay interference (Thorne et al., 2010).

HPLC has been used to measure of the activity of TPO via formation of the precursors monoiodotyrosine (MIT), diiodotyrosine (DIT), and both T3 and T4, in a reaction mixture containing TPO, or a surrogate enzyme such as lactoperoxidase (Divi & Doerge 1994). The tools and reagents for this method are all available. However, HPLC or other analytical chemistry techniques make this a low throughput assay, depending on the level of automation. A primary advantage of this in vitro method is that it directly informs hypotheses regarding the specific mechanism by which a chemical may impact thyroid hormone synthesis in vitro.  

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

TPO inhibition is a MIE conserved across taxa, with supporting data from experimental models and human clinical testing. This conservation is likely a function of the high degree of protein sequence similarity in the catalytic domain of mammalian peroxidases (Taurog, 1999). Ample data available for human, rat, and porcine TPO inhibition demonstrate qualitative concordance across these species (Schmultzer et al., 2007; Paul et al., 2013; Hornung et al., 2010). A comparison of rat TPO and pig TPO, bovine lactoperoxidase, and human TPO inhibition by genistein demonstrated good qualitative and quantitative (40–66%) inhibition across species, as indicated by quantification of MIT and DIT production (Doerge and Chang, 2002). Ealey et al. (1984) demonstrated peroxidase activity in guinea pig thyroid tissue using 3,3'-diaminobenzidine tetrahydrochloride (DAB) as a substrate that is oxidized by the peroxidase to form a brown insoluble reaction product. Formation of this reaction product was inhibited by 3-amino-1,2,4-triazole and the TPO inhibitor, methimazole (MMI). A comparative analysis of this action of MMI between rat- and human-derived TPO indicates concordance of qualitative response. Data also suggest an increased quantitative sensitivity to MMI in rat compared to human (Vickers et al., 2012). Paul et al. (2013) tested 12 chemicals using the guaiacol assay using both porcine and rat thyroid microsomes. The authors concluded that there was an excellent qualitative concordance between rat and porcine TPO inhibition, as all chemicals that inhibited TPO in porcine thyroid microsomes also inhibited TPO in rat thyroid microsomes when tested within the same concentration range. In addition, these authors noted a qualitative concordance that ranged from 1.5 to 50-fold differences estimated by relative potency. Similary, Takayama et al. (1986) found a very large species difference in potency for sulfamonomethoxine between cynomologus monkeys and rats.

Evidence for Perturbation by Stressor

Overview for Molecular Initiating Event

When a specific MIE can be defined (i.e., the molecular target and nature of interaction is known), in addition to describing the biological state associated with the MIE, how it can be measured, and its taxonomic, life stage, and sex applicability, it is useful to list stressors known to trigger the MIE and provide evidence supporting that initiation. This will often be a list of prototypical compounds demonstrated to interact with the target molecule in the manner detailed in the MIE description to initiate a given pathway (e.g., 2,3,7,8-TCDD as a prototypical AhR agonist; 17α-ethynyl estradiol as a prototypical ER agonist). Depending on the information available, this could also refer to chemical categories (i.e., groups of chemicals with defined structural features known to trigger the MIE). Known stressors should be included in the MIE description, but it is not expected to include a comprehensive list. Rather initially, stressors identified will be exemplary and the stressor list will be expanded over time. For more information on MIE, please see pages 32-33 in the User Handbook.

There is a wealth of information on the inhibition of TPO by drugs such as MMI and PTU, as well as environmental xenobiotics. In the landmark paper on thyroid disruption by environmental chemicals, Brucker-Davis (1998) identified environmental chemicals that depressed TH synthesis by inhibiting TPO. Hurley (1998) listed TPO as a major target for thyroid tumor inducing pesticides. More recent work has tested over 1000 chemicals using a high-throughput screening assay (Paul-Friedman 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

Brown RS, Shalhoub V, Coulter S, Alex S, Joris I, De Vito W, Lian J, Stein GS.  Developmental regulation of thyrotropin receptor gene expression in the fetal and neonatal rat thyroid: relation to thyroid morphology and to thyroid-specific gene expression.  Endocrinology. 2000 Jan;141(1):340-5.

Brucker-Davis F. 1998. Effects of environmental synthetic chemicals on thyroid function. Thyroid 8:827-856.

Chang, H. C. and D. R. Doerge (2000) Dietary genistein inactivates rat thyroid peroxidase in vivo without an apparent hypothyroid effect. Toxicol Appl Pharmacol. 168:244-252.

Divi, R. L., & Doerge, D. R. (1994). Mechanism-based inactivation of lactoperoxidase and thyroid peroxidase by resorcinol derivatives. Biochemistry 33(32), 9668–9674.

Divi, R. L., Chang, H. C., & Doerge, D. R. (1997). Anti-Thyroid Isoflavones from Soybean. Biochem. Pharmacol.  54(10), 1087–1096.

Doerge DR, Chang HC. Inactivation of thyroid peroxidase by soy isoflavones, in vitro and in vivo. J Chromatogr B Analy Technol Biomed Life Sci. 2002 Sep 25;777(1-2):269-79.

Ealey PA, Henderson B, Loveridge N.A quantitative study of peroxidase activity in unfixed tissue sections of the guinea-pig thyroid gland. Histochem J. 1984 Feb;16(2):111-22.

Fukiishi Y, Harauchi T, Yoshizaki T, Hasegawa Y, Eguchi Y.  Ontogeny of thyroid peroxidase activity in perinatal rats. Acta Endocrinol (Copenh). 1982 101(3):397-402.

Holzer G, Morishita Y, Fini JB, Lorin T, Gillet B, Hughes S, Tohmé M, Deléage G, Demeneix B, Arvan P, Laudet V. Thyroglobulin Represents a Novel Molecular Architecture of Vertebrates. J Biol Chem. 2016 Jun 16.

Hornung, M. W., Degitz, S. J., Korte, L. M., Olson, J. M., Kosian, P. a, Linnum, A. L., & Tietge, J. E. (2010). Inhibition of thyroid hormone release from cultured amphibian thyroid glands by methimazole, 6-propylthiouracil, and perchlorate. Toxicol Sci 118(1), 42–51.

Hurley PM. 1998. Mode of carcinogenic action of pesticides inducing thyroid follicular cell tumors in rodents. Environ Health Perspect 106:437-445.

Kaczur, V., Vereb, G., Molnár, I., Krajczár, G., Kiss, E., Farid, N. R., & Balázs, C. (1997). Effect of anti-thyroid peroxidase (TPO) antibodies on TPO activity measured by chemiluminescence assay. Clin. Chem 43(8 Pt 1), 1392–6.

Kessler, J., Obinger, C., Eales, G., 2008. Factors influencing the study of peroxidase- generated iodine species and implications for thyroglobulin synthesis. Thyroid 18, 769–774.

OECD (2017) New Scoping Document on in vitro and ex vivo Assays for the Identification of Modulators of Thyroid Hormone Signalling. Series on Testing and Assessment. No. 207.  ISSN: 20777876 (online) http://dx.doi.org/10.1787/20777876

Paul KB, Hedge JM, Macherla C, Filer DL, Burgess E, Simmons SO, Crofton KM, Hornung MW. Cross-species analysis of thyroperoxidase inhibition by xenobiotics demonstrates conservation of response between pig and rat. Toxicology. 2013. 312:97-107

Paul, K.B., Hedge, J.M., Rotroff, D.M., Hornung, M.W., Crofton, K.M., Simmons, S.O. 2014. Development of a thyroperoxidase inhibition assay for high-throughput screening. Chem.  Res. Toxicol. 27(3), 387-399.

Paul-Friedman K, Watt ED, Hornung MW, Hedge JM, Judson RS, Crofton KM, Houck KA, Simmons SO. 2016. Tiered High-Throughput Screening Approach to Identify Thyroperoxidase Inhibitors Within the ToxCast Phase I and II Chemical Libraries.  Toxicol Sci. 151:160-80.

Pintar, J.E. (2000) Normal development of the hypothalamic-pituitary-thyroid axis. In. Werner & Ingbar’s The Thyroid. (8th ed), Braverman. L.E. and Utiger, R.D. (eds) Lippincott Williams and Wilkins, Philadelphia.

Remy L, Michel-Bechet M, Athouel-Haon AM, Magre S. Critical study of endogenous peroxidase activity: its role in the morphofunctional setting of the thyroid follicle in the rat fetus. Acta Histochem. 1980;67(2):159-72.

Ruf, J., & Carayon, P. (2006). Structural and functional aspects of thyroid peroxidase. Archives of Biochemistry and Biophysics, 445(2), 269–77.

Santisteban P, Bernal J. Thyroid development and effect on the nervous system. Rev Endocr Metab Disord. 2005 Aug;6(3):217-28.

Schmutzler, C., Bacinski, A., Gotthardt, I., Huhne, K., Ambrugger, P., Klammer, H., Schlecht, C., Hoang-Vu, C., Gruters, A., Wuttke, W., Jarry, H., Kohrle, J., 2007a. The ultraviolet filter benzophenone 2 interferes with the thyroid hormone axis in rats and is a potent in vitro inhibitor of human recombinant thyroid peroxidase. Endocrinology 148, 2835–2844.

Taurog A. 2005. Hormone synthesis. In: Werner and Ingbar’s The Thyroid: A Fundamental and Clinical Text (Braverman LE, Utiger RD, eds). Philadelphia:Lippincott, Williams and Wilkins, 47–81

Taurog, a, Dorris, M. L., & Doerge, D. R. (1996). Mechanism of simultaneous iodination and coupling catalyzed by thyroid peroxidase. Archives of Biochemistry and Biophysics, Taurog A. Molecular evolution of thyroid peroxidase. Biochimie. 1999 May;81(5):557-62

Takayama S, Aihara K, Onodera T, Akimoto T. Antithyroid effects of propylthiouracil and sulfamonomethoxine in rats and monkeys. Toxicol Appl Pharmacol. 1986 Feb;82(2):191-9.

Thorne N, Auld DS, Inglese J.  Apparent activity in high-throughput screening: origins of compound-dependent assay interference. Curr Opin Chem Biol. 2010 Jun;14(3):315-24.

Thorpe-Beeston JG, Nicolaides KH, McGregor AM. Fetal thyroid function. Thyroid. 1992 Fall;2(3):207-17. Review.

Vickers AE, Heale J, Sinclair JR, Morris S, Rowe JM, Fisher RL. Thyroid organotypic rat and human cultures used to investigate drug effects on thyroid function, hormone synthesis and release pathways. Toxicol Appl Pharmacol. 2012 Apr 1;260(1):81-8.

Zoeller, R. T., Tan, S. W., & Tyl, R. W. (2007). General background on the hypothalamic-pituitary-thyroid (HPT) axis. Critical Reviews in Toxicology, 37(1-2), 11–53.