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Event: 984

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

Impairment, T-cell dependent antibody response

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
Impairment, T-cell dependent antibody response

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

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
Immunosuppression increased

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 AdverseOutcome Takumi Ohishi (send email) Open for comment. Do not cite WPHA/WNT Endorsed


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
Rattus norvegicus Rattus norvegicus 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

Antibody production to T-cell–dependent antigens is established through the coordination of B cells, antigen-presenting cells as well as T-cell–derived cytokines, which stimulate B cells to proliferate and differentiate. T-cell–dependent antibody response (TDAR) might be altered if any of these cell populations is affected.

Interleukin (IL)-2 stimulates B cells to proliferate through surface IL-2 receptors. IL-4 stimulates B-cells to proliferate, to switch immunoglobulin classes, and to differentiate into plasma and memory cells. Suppressing the production of these B-cell–related cytokines appears to impaire TDAR, as seen in the result of FK506 treatment (Heidt et al, 2009).

IL-2 and IL-4 are produced and secreted by helper T cells and play important roles in the development of TDAR. IL-4 affects maturation and class switching of B cells as well as proliferation, both of which induces/enhances T cell dependent antibody production. IL-2 promotes differentiation of B cells through IL-2 stimulates differentiation of the activated T cell into T cell called Th2 cell. Therefore, suppressed production of IL-2 and IL-4 impairs TDAR (Alberts et al. 2008).

In male CD-1 mice, chronic psychosocial stress (types of social outcome occurred: residents becoming subordinates) decrease in anti- keyhole limpet hemocyanine (KLH) immunoglobulin (Ig)G. (Alessandro, B. et al. 2003).

In female B6C3F1 mice, 1,2:5,6-dibenzanthracene (DBA) exposure reduced total IgG antibody in spleen cell culture supernatants after in vitro stimulation with lipopolysaccharide (LPS) (Donna, C. et al. 2010).

Treatment with cyclosporin A (CsA) at 50 mg/kg BID via oral gavage in cynomolgus monkey resulted in reduction of serum sheep red blood cells (SRBC)-specific IgM and IgG (Kevin, G. et al. 2014).

After a 9-day culture of B cells and non-pre-activated T cell stimulation with FK506 or CsA, the levels of IgM and IgG in the culture supernatant were reduced at 0.3 and 1.0 ng/mL (0.37 and 1.24 nM) of FK506 or 50 and 100 ng/mL (41.6 and 83.2 nM) of CsA (Heidt et al, 2009).

After a 4-day culture of SKW6.4 cells (IL-6-dependent IgM-secreting human B-cell line) and anti-CD3/CD28 stimulated peripheral blood mononuclear cells (PBMC) culture supernatant with FK506 or CsA, the level of IgM in the culture supernatant was reduced at concentrations of 0.01 to 100 ng/mL (0.012 to 124 nM) of FK506 or 0.1 to 1000 ng/mL (0.083 to 83.2 nM) of CsA (Sakuma et al. 2001b).

Rats were treated with FK506 for over four weeks and immunized with KLH, after which serum concentration of anti-KLH IgM and IgG was reduced at the dose level of 3 mg/kg/day (Ulrich et al. 2004).

Mice were treated with FK506 or CsA for 4 days, and immunized with SRBC, after which antigen-specific plaque-forming splenocytes were reduced at dose levels of 3.2, 10, 32 and 100 mg/kg of FK506 or 32 and 100 mg/kg of CsA (Kino et al. 1987b).

As immunosuppression-derived adverse outcomes by calcineurin inhibition, FK506 and CsA increase the frequency and/or severity of infections and allergic reactions impaired TDAR deems to be one of the causative factors for these side effects . Some clinical trials of FK506 and CsA revealed these adverse effects as follows.

  • In clinical trials of renal transplantation using FK506 or CsA, opportunistic infections such as candida, cytomegalovirus and herpes simplex virus were reported (Ekberg et al. 2007).
  • In recipients of liver transplants treated with FK506 or CsA, opportunistic infections such as cytomegalovirus, hepatitis C virus, hepatitis B and herpes simplex virus were reported (Fung et al. 1991).
  • Cardiac transplant patients treated with cyclosporin developed pulmonary infections within the first year after surgery (Luster, M.I. et al. 1993).
  • In patients of X-linked autoimmune enteropathy treated with CsA or FK506, serum levels of IgE developed extremely high during the immunosuppressive therapy (Kawamura et al. 1997).
  • Renal transplant recipients treated with belatacepy/mycophenolate (MMF)/predonisone or FK506/MMF/prednisone showed significantly lower the geometric mean hemagglutination inhibition titer against influenza vaccine, hemagglutination-specific IgG and isotype IgG1 antibodies, and IgG-antibody secreting cells response (Gangappa et al. 2019).

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). ?

TDAR could be examined in vivo and in vitro.

In vivo studies of antigen-specific antibodies are usually performed by measuring serum antibody levels with Enzyme-Linked ImmunoSorbent Assay (ELISA) or with a plaque-forming cell (PFC) assay.

  • Rats were repeatedly administered FK506 orally for 4 weeks and immunized with KLH, after which the serum was examined for T-cell–dependent, antigen-specific, IgM and IgG levels using a Sandwich ELISA kit (Ulrich et al. 2004).
  • Mice were repeatedly administered calcineurin inhibitors (CNIs) including FK506 and CsA orally for 4 days and immunized with SRBC, after which spleen cells were examined using a PFC assay (Kino et al. 1987).
  • Cynomolgus monkeys received 50 mg/kg CsA twice a day via oral gavage (10 h apart) for 23 days and were immunized with SRBC, after which the serum was examined for Anti-SRBC IgM and IgG levels using an ELISA specific for SRBC antigen (Kevin, G. et al. 2014).
  • Mice were exposed a single pharyngeal aspiration of DBA, after which supernatants of splenocytes cultured for 24 h in the presence of LPS and assayed using a mouse IgM or IgG matched pairs antibody kit (Bethyl Laboratories, Montgomery, TX) (Donna, C. et al. 2010).

For in vitro studies, total IgM and IgG levels in culture supernatant are often measured after polyclonal T-cell activation rather than measuring antigen stimulation in immune cell cultures.

  • T cells and B cells isolated from human peripheral blood mononuclear cells (PBMC) were co-cultured with a CNIs for nine days in the presence of polyclonal–T-cell stimulation, after which supernatants were tested for immunoglobulin IgM and IgG levels using a Sandwich ELISA kit. Treatment with FK506 or CsA reduced the levels of IgM and IgG at the concentrations of 0.3 and 1.0 ng/mL or 50 and 100 ng/mL (Heidt et al, 2009).
  • SKW6.4 cells (IL-6-dependent IgM-secreting human B-cell line) were cultured with anti-CD3/CD28 antibody-stimulated PBMC culture supernatant. After culturing for four days, IgM produced in the culture supernatants was measured using an ELISA kit. FK506 or CsA reduced the levels of IgM at the concentrations of 0.01 to 100 ng/mL or 0.1 to 1000 ng/mL (Sakuma et al. 2001b).
  • In order to examine class switching, T cells derived from human PBMCs were cultured with CNIs, and cytokine mRNA levels of Interferon-gamma, IL-2, IL-4, IL-5, IL-10, IL-13, and other B-cell–stimulatory cytokines produced in T cells were measured by quantitative PCR (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 induced impairment of TDAR is demonstrated with rodent studies. That is, oral administration of FK506 or CsA to mice for 4 days impaired the response of PFC in splenocytes after intravenous immunization with sheep erythrocytes (Kino et al. 1987). Likewise, oral administration of FK506 to rats over a four-week period reduced production of both anti-KLH-IgG and IgM antibodies after subcutaneous immunization with KLH (Ulrich et al. 2004). Moreover, Treatment with CsA at 50 mg/kg BID via oral gavage in cynomolgus monkey resulted in reduction of serum SRBC-specific IgM and IgG (Kevin, G. et al. 2014). As for humans, in vitro experiments showed that treatment with FK506 or CsA of peripheral blood mononuclear cells from blood-bank donors suppressed the production of IgM and IgG antibodies specific to T-cell–dependent antigens. (Heidt et al, 2009) Also, in SKW6.4 cells (IL-6–dependent, IgM-secreting, human B-cell line) cultures, FK506 or CsA suppressed the production of IgM antibodies in the presence of T-cell activation. (Sakuma et al. 2001b)  Considering that FKF506 and CsA reduce T cell-derived cytokines including IL-2 and IL-4, these findings strongly suggest that impairment of TDAR following reduced production of such cytokines occurs at least in common among humans monkey and rodents.

Evidence for Perturbation by Stressor

Regulatory Significance of the Adverse Outcome

An AO is a specialised KE that represents the end (an adverse outcome of regulatory significance) of an AOP. For KEs that are designated as an AO, one additional field of information (regulatory significance of the AO) should be completed, to the extent feasible. If the KE is being described is not an AO, simply indicate “not an AO” in this section.A key criterion for defining an AO is its relevance for regulatory decision-making (i.e., it corresponds to an accepted protection goal or common apical endpoint in an established regulatory guideline study). For example, in humans this may constitute increased risk of disease-related pathology in a particular organ or organ system in an individual or in either the entire or a specified subset of the population. In wildlife, this will most often be an outcome of demographic significance that has meaning in terms of estimates of population sustainability. Given this consideration, in addition to describing the biological state associated with the AO, how it can be measured, and its taxonomic, life stage, and sex applicability, it is useful to describe regulatory examples using this AO. More help

The ICH S8 guideline, which covers immunosuppression of small molecule drugs, determines the need for immunotoxicity studies by comprehensively evaluating the findings of pharmacology, changes in the immune system in repeated-dose toxicity studies, and other factors using a Weight of Evidence approach. If there is concern about immunotoxicity, the presence or absence of immunotoxicity should be determined using an in vivo test system capable of assessing the functional changes of predicted immunotoxic target cells. If immunotoxicity is observed, additional studies including in vitro assays or clinical evaluation should be considered to assess the risk of immunotoxicity in humans. Because TDAR involves many immune cell populations, including T cells, B cells, and antigen-presenting cells, evaluation of TDAR is recommended when there is concern about immunotoxicity but the immunotoxic target cells are unclear. The S8 guidelines list KLH, SRBC, and tetanus toxin as antigens for TDAR.

The draft FDA immunotoxicity testing guidance (2020) covers immunosuppressive and immunostimulatory drugs and biologics; evaluating immunosuppressive drugs in the draft FDA guidance is similar to that in the S8 guideline, with in vivo TDAR assays recommended when toxic target cells are unknown. The draft guidance states that TDAR assays using KLH as an antigen have been established in mice, rats, dogs, minipigs, and cynomolgus monkeys, but the use of SRBC and tetanus toxin as antigens is also acceptable.

For the assessment for pesticides, US EPA OPPTS 870.7800 immunotoxicity testing guideline recommends TDAR using SRBC. The REACH guideline does not provide for immunotoxicity testing, but it provides triggers for conducting immunotoxicity testing.

The WHO/IPSS Immunotoxicity Risk assessment Guidance (2012) describes a strategy for assessing five categories of immunotoxicity risks, including immunosuppression. For risk assessment of immunosuppression, it calls for identification of immunosuppression risks, prediction of pathogenesis that may occur, and consideration of safety margins based on the WoE approach from human findings, infection resistance tests, immune function tests, general immune system assays, histopathological findings and organ weights in general toxicity studies, and hematological data.

The evaluation of immunotoxicity in F1 animals in the OECD Guidelines for Extended First Generation Reproductive and Developmental Toxicity Studies (TG443) requires that PFC and ELSA assays to measure primary IgM antibody production by TDAR using T-cell dependent antigens (SRBC, KLH, etc.) be performed. Furthermore, if changes are observed, the significance of the changes should be examined by comprehensively evaluating other data.

The outcomes of immunosuppression are susceptibility to infection and tumorigenesis, and the FDA guidance requires that immunosuppressive drugs be evaluated for carcinogenic risk using WoE approach based on the results of carcinogenicity and immunotoxicity studies. Meanwhile, the ICH S1B(R1) Draft Step 2 Guidelines for Carcinogenicity Testing calls for evaluation of carcinogenicity by WoE approach instead of rat carcinogenicity testing, because rodent carcinogenicity test models are less capable of detecting carcinogenicity. On the other hand, it is difficult to define susceptibility to infection as a measurable AO with a clear mechanism, because immune responses vary among pathogens. In fact, many immunotoxicity guidelines require that the risk of immunotoxicity be identified and assessed by evaluating immune functions.

In AOP154, it was difficult to define susceptibility to infection as an AO for the AOP154, so TDAR, which is recommended as an indicator of immunosuppresoin in many guidelines, was used as an AO. It is expected that several AOPs with TDARs as AOs will be developed, and based on these AOPs, it may be possible to develop an IATA to assess the risk of immunotoxicity characterized by TDARs.


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 ( (OECD, 2015). More help
  1. Alberts, B., Johnson, A., Lewis, L., Raff, M., Roberts, K. and Walter, P. (2008). Molecular Biology of the Cell. 5th ed., Garland Science, New York. 1539-1601
  2. Heidt, S., Roelen, D. L., Eijsink, C., Eikmans, M., van Kooten, C., Claas, F. H. and Mulder, A. (2010). Calcineurin inhibitors affect B cell antibody responses indirectly by interfering with T cell help. Clinical and experimental immunology. 159(2): 199-207.
  3. Sakuma, S., Kato, Y., Nishigaki, F., Magari, K., Miyata, S., Ohkubo, Y., and Goto, T. (2001b). Effects of FK506 and other immunosuppressive anti-rheumatic agents on T cell activation mediated IL-6 and IgM production in vitro. International Immunopharmacology 1(4): 749-57.
  4. Kino, T., Hatanaka, H., Hashimoto, M., Nishiyama, M., Goto, T., Okuhara, M., Kohsaka, M., Aoki, H. and Imanaka, H. (1987). FK-506, a novel immunosuppressant isolated from a Streptomyces. I. Fermentation, isolation, and physico-chemical and biological characteristics. Journal of antibiotics. 40(9): 1249-1255.
  5. Ulrich, P., Paul, G., Perentes, E., Mahl, A., and Roman D. (2004). Validation of immune function testing during a 4-week oral toxicity study with FK506. Toxicology Letters 149(1-3): 123-31.
  6. 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.
  7. Ekberg, H., Tedesco-Silva, H., Demirbas, A., Vítko, S., Nashan, B., Gürkan, A., Margreiter, R., Hugo, C., Grinyó, J.M., Frei, U., Vanrenterghem, Y., Daloze, P. and Halloran, P.F.; ELITE-Symphony Study. (2007). Reduced exposure to calcineurin inhibitors in renal transplantation. The New England journal of medicine 357 (25): 2562-75.
  8. Fung, J., Abu-Elmagd, K., Jain, A., Gordon, R., Tzakis, A., Todo, S., Takaya, S., Alessiani, M., Demetris, A., Bronster, O., Martin, M., Mieles, L., Selby, R., Reyes, J., Doyle, H., Stieber, A., Casavilla, A. and Starzl, T. (1991). A randomized trial of primary liver transplantation under immunosuppression with FK 506 vs cyclosporine. Transplantation proceedings 23 (6): 2977-83.
  9. Luster, M.I., and Rosenthal, G.J. (1993). Environmental Health Perspectives. 100: 219-36.
  10. 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
  11. 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
  12. 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,
  13. Gangappa S, Wrammert J, Wang D, Li ZN, Liepkalns JS, Cao W, Chen J, Levine MZ, Stevens J, Sambhara S, Begley B, Mehta A, Pearson TC, Ahmed R, Larsen CP. (2019) Kinetics of antibody response to influenza vaccination in renal transplant recipients. Transpl Immunol. 53:51-60.
  14. Kawamura N, Furuta H, Tame A, Kobayashi I, Ariga T, Okano M, Sakiyama Y. (1997) Extremely high serum level of IgE during immunosuppressive therapy: paradoxical effect of cyclosporine A and tacrolimus. Int Arch Allergy Immunol. 112(4):422-4.