To the extent possible under law, AOP-Wiki has waived all copyright and related or neighboring rights to KE:1202

Event: 1202

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

Suppression, IL-2 and IL-4 production

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
Suppression, IL-2 and IL-4 production

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

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
Organ term
immune system

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
interleukin-2 production interleukin-2 decreased
interleukin-4 production interleukin-4 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
Immunosuppression KeyEvent Takumi Ohishi (send email) Open for comment. Do not cite WPHA/WNT Endorsed

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
Homo sapiens Homo sapiens High NCBI
Mus musculus Mus musculus High NCBI
cynomolgus monkey Macaca fascicularis High 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
Unspecific 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

Production of T cell cytokines including Interleukin (IL)-2 and IL-4 is regulated by nuclear factor of activated T cells (NFAT)/ activator protein-1 (AP-1) complexes. Activated NFAT/AP-1 complex that bind at the site of the IL-2 and IL-4 promoters, thereby induces transcription of IL-2 and IL-4 (Jain et al. 1993).  For IL-2, NFAT proteins are necessary for IL-2 gene expression and cooperation of NFAT with AP-1 is required for IL-2 gene transcription. For IL-4, At least five different NFAT sites have been described in the IL-4 promoter with at least three of them being composite sites binding NFAT and AP-1 (Macián et al. 2001).

IL-2 binds to IL-2 receptor (IL-2R) and acts on T cells. CD25 is one of IL-2R. Basiliximab (Simulect) is known as anti-CD25 antibody. Basiliximab binds to IL-2R and blocks IL-2 signaling. Clinical transplantation study of basiliximab reveals decreases in rejections. On the other hand, basiliximab inhibits the activation of antigen specific T cells (Novartis Pharma 2016). 

Calcineurin inhibitors (CNIs) such as FK506 and cyclosporin A (CsA) hinder the formation of the functional NFAT/AP-1 complexes by interfering with NFAT nuclear localization (Flanagan et al. 1991).  Reduced binding of NFAT/AP-1 complexes at the promoter site of the IL-2 gene lowers the transcription of the mRNA of IL-2 and the following cytokine production.

Transcription of IL-4 is also inhibited by CNIs in the same manner as IL-2 (Dumont et al. 1998).

In CD3/ phorbol 12-myristate-13-acetate (PMA)-activated human T cells, FK506 suppressed production of IL-2, IL-4, and Interferon (IFN)-γ at the concentrations of 1.2 to 12.5 nM after 22 to 24 hours culture as well as inhibited expression of IL-2, IL-4, and IFN-γ mRNA at 10 nM (Dumont et al. 1998).

Treatment with CsA completely eliminated detectable IL-2 release from 3A9 T cells co-cultured with antigen-bearing Ch27 B cells with an IC25 and IC50 for IL-2 production of 1.19 nM and 1.99 nM. Treatment with other immunosuppressant compounds (dexamethasone, azathioprine, methotrexate, benzo(a)pyrene and urethane) also resulted in decreased IL-2 release from stimulated 3A9 T cells at non-cytotoxic concentrations. Urethane, a weakly immunosuppressive chemical, was least potent in the assay, with an IC25 and IC50 for IL-2 secretion of 4.24 mM and 13.26 mM (D.M. Lehmann. et al. 2018).

In male CD-1 mice, chronic psychosocial stress (types of social outcome occurred: residents becoming subordinates) reduced IL-2 release in response to keyhole limpet hemocyanine (KLH) (Alessandro, B. et al. 2003).

In female B6C3F1 mice, 1,2:5,6-dibenzanthracene exposure reduced production of IL-2 in spleen cell culture supernatants after in vitro stimulation with Concanavalin A or lipopolysaccharide (Donna, C. et al. 2010).

Treatment with CsA at 50 mg/kg BID via oral gavage or 2C1.1 (a fully human anti-ORAI1 monoclonal antibody) at 25 mg/kg single IV resulted in reduction of IL-2, IL-4, IL-5, and IL-17 cytokine production from PMA/ionomycin stimulation of whole blood in the cynomolgus monkey (Kevin, G. et al. 2014).

CNIs is considered to increase carcinogenicity through the suppression of IL-2 and IL-4 production.

  • Renal transplant patients on immunosuppressive therapy were found to develop cancer within 10 years after surgery (Luster, M.I. et al. 1993).

In experimental animal studies, carcinogenicity of FK506 was reported as follows.

  • In mice subjected to topical application testing, in which 100 μL of FK506 ointment was applied once daily for two years to roughly 40% of the total body area, an increased incidence of lymphoma was found in mice of the 0.1% ointment group showing high blood concentrations of the drug (Maruho Co., Ltd 2014).
  • In hairless albino mice, virtually all of which developed skin tumors after a 40-week exposure to ultraviolet light, application of a 1% FK506 ointment reduced the time to outbreak of the skin tumors. (Maruho Co., Ltd 2014).

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). ?

Quantitation of cytokine content was done on appropriately diluted samples, run in duplicate, using Sandwich Enzyme-Linked ImmunoSorbent Assay (ELISA) kits to test matched Antibody pairs with biotin-horseradish peroxidase-streptavidin detection and 3,3',5,5'-tetramethylbenzidine substrate. ELISA plates were scanned in a Molecular Devices UVmax plate reader (Menlo Park, CA), using SOFT max software (Molecular Devices) (Dumont et al. 1998).

Ex vivo whole blood stimulated cytokine (IL-2, IL-4, IL-5, and IL-17) production assay in the supernatants were determined using an electrochemiluminescent immunoassay from Meso Scale Discovery (MSD; Gaithersburg, MD) (Kevin, G. et al. 2014).

Total RNA was extracted using RNeasy mini kit (Qiagen, Chatsworth, CA) and quantitated by absorbance at 260 nm. Cytokine mRNAs were detected using a RiboQuant MultiProbe RPA system (PharMingen, San Diego, CA). Riboprobes were 32P-labeled and hybridized overnight with 10 to 30 mg of the RNA samples. The hybridized RNA was treated with RNase and purified according to the RiboQuant protocol. The samples were then electrophoresed in 6% polyacrylamide-Tris-borate-EDTA-urea gels using the Seqi -Gen GT Nucleic Acid Electrophoresis Cell (Bio-Rad, Hercules, CA), or minigels (Novex, San Diego, CA). The gels were dried, exposed and quantitated in a PhosphorImager (Molecular Dynamics, Sunnyvale, CA) using the ImageQuant software (Dumont et al. 1998).

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

CNIs suppress production of IL-2, IL-3, IL-4, IL-5, IFN-γ, Granulocyte Macrophage colony-stimulating Factor (GM-CSF), and other cytokines, as induced by CD2/CD3 or CD3/CD26 stimulation, in human peripheral blood mononuclear cells (PBMC) (Sakuma et al. 2001a). Also, CNIs (FK506 and CsA) suppress production of IL-2, IL-3, IL-4, IL-5, IL-6, IL-10, Tumor Necrosis Factor-α, IFN-γ, and GM-CSF, as induced by CD3/PMA stimulation, in human PBMC (Dumont et al. 1998).

CNIs (FK506 and CsA) exhibit suppression of IL-2 production induced from mixed lymphocyte reactions in mice and humans (Kino, T et al. 1987a).

Treatment with CsA or 2C1.1 resulted in reduction of IL-2, IL-4, IL-5, and IL-17 cytokine production from PMA/ionomycin stimulation of whole blood in the cynomolgus monkey (Kevin, G. et al. 2014).

These facts indicate that Calcineurin-NFAT system-mediated suppression of cytokines is commonly found in humans, monkey and mice.

Evidence for Perturbation by Stressor

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
  1. Dumont, F.J., Staruch, M.J., Fischer, P., DaSilva, C. and Camacho, R. (1998). Inhibition of T cell activation by pharmacologic disruption of the MEK1/ERK MAP kinase or calcineurin signaling pathways results in differential modulation of cytokine production. Journal of immunology 160 (6): 2579-89.
  2. Flanagan, W.M., Corthésy, B., Bram, R.J. and Crabtree, G.R. (1991). Nuclear association of a T-cell transcription factor blocked by FK-506 and cyclosporin A. Nature 352 (6338): 803-7.
  3. Jain, J., McCaffrey, P. G., Valge-Archer, V. E. and Rao, A. (1992). Nuclear factor of activated T cells contains Fos and Jun. Nature. 356(6372): 801-804.
  4. Jain, J., Miner, Z. and Rao, A. (1993). Analysis of the preexisting and nuclear forms of nuclear factor of activated T cells. Journal of immunology. 151(2): 837-848.
  5. Kino, T., Hatanaka, H., Miyata, S., Inamura, N., Nishiyama, M., Yajima, T., Goto, T., Okuhara, M., Kohsaka, M. and Aoki, H. (1987a). FK-506, a novel immunosuppressant isolated from a Streptomyces. II. Immunosuppressive effect of FK-506 in vitro. Journal of antibiotics. 40(9): 1256-1265.
  6. Macián, F., López-Rodríguez, C. and Rao, A. (2001). Partners in transcription: NFAT and AP-1. Oncogene. 20(19): 2476-89.
  7. Novartis Pharma K.K. (2016). Drug interview form Simulect i.v. injection 20 mg. 10th edition.
  8. Sakuma, S., Higashi, Y., Sato, N., Sasakawa, T., Sengoku, T., Ohkubo, Y., Amaya, T., and Goto, T. (2001a). Tacrolimus suppressed the production of cytokines involved in atopic dermatitis by direct stimulation of human PBMC system. (Comparison with steroids). International Immunopharmacology 1(6): 1219-26.
  9. Schreiber, SL., and Crabtree, GR. (1992). The mechanism of action of cyclosporin A and FK506. Immunology Today 13(4): 136-42.
  10. Luster, M.I., and Rosenthal, G.J. (1993). Environmental Health Perspectives. 100: 219-36.
  11. Maruho Co.,Ltd. (2014) Drug interview form Protopic ointment 0.1% Revised 16th edition.
  12. Alessandro B, Paola S, Alberto E. Paneraic, Tiziana P,Paola Palanzaa and Stefano P(2003). Chronic psychosocial stress-induced down-regulation of immunity depends upon individual factors Journal of Neuroimmunology 141: 58–64
  13. Donna C. S, Matthew J. S and Kimber L. W Jr. (2010) Systemic immunosuppression following a single pharyngeal aspiration of 1,2:5,6-dibenzanthracene in female B6C3F1 mice, Journal of Immunotoxicology, 7:3, 219-231
  14. Kevin G, Hossein S, Raju S, Valerie A, Anna K, Ming Z, Fen-Fen L, Hung Q. N, Lei Z, John K. S, Min W and Helen J. M(2015) Inhibition of CRAC with a human anti-ORAI1 monoclonal antibody inhibits T-cell-derived cytokine production but fails to inhibit a T-cell-dependent antibody response in the cynomolgus monkey, Journal of Immunotoxicology, 12:2, 164-173,
  15. D.M. Lehmann, W.C. Williams.(2018) Development and Utilization of a Unique In Vitro Antigen Presentation Co-culture Model for Detection of Immunomodulating Substances. Toxicol In Vitro.53: 20–28.