SNAPSHOT

1201: Binding, Immunophilins

Short Name: Binding, Immunophilins

AOPs Including This Key Event

Biological Organization

Taxonomic Applicability

Life Stage Applicability

Sex Applicability

FKBP is found in a wide variety of organisms, from prokaryotes to multicellular organisms. (Siekierka et al. 1989a) Multiple subfamilies of FKBP have been reported, with at least eight types having been found in mammals. FKBP12 is reported to be expressed in B-cells, Langerhans cells, and mast cells as well as in T-cells of humans, mice, and other mammalian species.

Cyclophilins have been found in mammals, plants, insects, fungi, and bacteria. They are structurally conserved throughout evolution and all have PPIase activity. (Wang P et al. 2005)

980: Inhibition, Calcineurin Activity

Short Name: Inhibition, Calcineurin Activity

AOPs Including This Key Event

Stressors

Biological Organization

Taxonomic Applicability

Life Stage Applicability

Sex Applicability

Calcineurin is broadly distributed T-cells, B‑cells, and throughout the body. The structure of CnA and CnB is highly conserved from yeasts to humans. Also highly conserved are the amino acid sequences of the catalytic and regulatory domains of calcineurin A isoforms from different organisms. (Kincaid. 1996)

979: Interference, nuclear localization of NFAT

Short Name: Interference, nuclear localization of NFAT

AOPs Including This Key Event

Biological Organization

Taxonomic Applicability

Life Stage Applicability

Sex Applicability

NFAT expresses in B cells, mast cells, neutrophil granulocytes, dendritic cells, macrophages, and natural killer cells as well as T cells from humans, rodents, and other mammalian species. (Rao et al. 1997)

981: Reduction, NFAT complex formation

Short Name: Reduction, NFAT complex formation

AOPs Including This Key Event

Biological Organization

Taxonomic Applicability

Life Stage Applicability

Sex Applicability

FK506-induced interference with NFAT/AP-1 complex formation at the promoter site of the IL-2 gene might be in common among mammalian T cells including humans and rodents (Flanagan et al. 1991).

1202: Suppression, IL-2 and IL-4 production

Short Name: Suppression, IL-2 and IL-4 production

AOPs Including This Key Event

Biological Organization

Taxonomic Applicability

Life Stage Applicability

Calcineurin inhibitors suppress production of IL-2, IL-3, IL-4, IL-5, IFN-γ, 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, calcineurin inhibitors suppress production of IL-2, IL-3, IL-4, IL-5, IL-6, IL-10, TNF-α, IFN-γ, and GM-CSF, as induced by CD3/PMA stimulation, in human PBMC. (Dumont et al. 1998) Calcineurin inhibitors exhibit suppression of IL-2 production induced from mixed lymphocyte reactions in mice and humans. (Kino, T et al. 1987a)

984: Impairment, T-cell dependent antibody response

Short Name: Impairment, T-cell dependent antibody response

AOPs Including This Key Event

Biological Organization

Taxonomic Applicability

Life Stage Applicability

Sex Applicability

In vitro experiments showed that treatment with FK506 of peripheral blood mononuclear cells from blood-bank donors suppressed the production of immunoglobulin (Ig) M and G antibodies specific to T-cell dependent antigens. (Heidt et al, 2009) Also, in human PBMC cultures, FK506 suppressed the production of IgM antibodies in the presence of T-cell activation. (Sakuma et al. 2001b) Oral administration of FK506 to mice for 4 days impaired the response of plaque forming cells (PFC) in splenocyte after intravenous immunization with sheep erythrocytes. (Kino et al. 1987) 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)

Binding, Immunophilins leads to Inhibition, Calcineurin Activity

AOPs Referencing Relationship

Evidence Supporting Applicability of this Relationship

FKBP is found in a wide variety of organisms, from prokaryotes to multicellular organisms (Siekierka et al. 1989). Multiple subfamilies of FKBP have been reported, with at least eight types having been found in mammals. FKBP12 is reported to be expressed in B-cells, Langerhans cells, and mast cells as well as in T-cells of humans, mice, and other mammalian species.

Cyclophilins have been found in mammals, plants, insects, fungi, and bacteria. They are structurally conserved throughout evolution and all have PPIase activity. (Wang P et al. 2005)

Calcineurin is broadly distributed throughout the body, including T- and B-cells, and the structure of CnA and CnB is highly conserved from yeasts to humans. Also highly conserved are the amino acid sequences of the catalytic and regulatory domains of calcineurin A isoforms from different organisms. (Kincaid. 1996)

How Does This Key Event Relationship Work

Immunophilins are a general class of proteins that exhibit peptidyl-propyl isomerase (PPIase) activity, such as FKBP (FK506-binding protein) or cyclophilin. (Barik. 2006) FKBP and cyclophilin bind with calcineurin-inhibitors FK506 and cyclosporin A to form complexes, which inhibit calcineurin activity. (Barik. 2006)

While FKBP12, FKBP12.6, FKBP13, and FKBP52 are all part of the FK506-binding FKBP family, FKBP12 has a significant involvement in the mechanism of action for FK506-induced immunosuppression. (Siekierka et al. 1989, Kang et al. 2008)

FKBP12 is a 12-kDa protein localized in cytoplasm and has been isolated from Jurkat T-cells as a receptor that binds with the calcineurin inhibitor FK506. (Bram et al. 1993) FKBP12 has an FK506-binding domain (FKBD) that comprises 108 amino acids, and is expressed in T‑cells, B‑cells, Langerhans cells, and mast cells (Siekierka et al. 1990, Panhans-Gross et al. 2001, Hultsch et al. 1991).

Cyclophilin and FKBP both exhibit PPIase activity, but no structural similarities have been found between them. Additionally, while immunophilin complexes formed with either substance do inhibit CN phosphatase activity, the PPlase activity and the inhibition of activity that they indicate are unrelated to CN regulation.

Calcineurin is a heterodimer that comprises a catalytic subunit (CnA) and a Ca-binding regulatory subunit (CnB). CnA handles phosphatase activity as well as calmodulin binding, and CnB regulates intracellular calcium and CnA. (Klee et al. 1988, Zhang et al. 1996) CnA is a 59kDa protein with a serine-threonine phosphatase domain. A FK506-FKBP complex binds directly to CnA in the cell, causing steric hindrance of substrate binding to calcineurin, which in turn inhibits phosphatase activity of calcineurin. (Schreiber and Crabtree 1992, Liu et al. 1993, Bierer et al. 1993, Bram et al. 1993, Rao et al. 1997, Liu et al. 1991) Cyclophilin-CsA complexes also function in the same manner, binding directly to CnA in the cell, which in turn inhibits CN phosphatase activity.

Weight of Evidence

Biological Plausibility

Immunophilins are a general class of proteins that exhibit PPlase activity, but modification of these functions are unrelated to inhibition of CN activity, which is thought to arise in the molecular structure of the complexes.

FKBP12 is expressed in several types of immune cells, where it functions as an accessory molecule to classes of intracellular calcium channels, namely IP3 receptors and ryanodine receptors (RyRs) (Schreiber and Crabtree 1992, Cameron et al. 1997). In addition, FKBP12 binds to transforming growth factor-β receptor 1 (TGF-βR1) and acts as a natural ligand to TGF-β.

Empirical Support for Linkage

There is no evidence of any relationship between FKBP12 KO and the immune system in the FKBP12 knockout mouse model.

Uncertainties or Inconsistencies

Not specified.

Quantitative Understanding of the Linkage

In the phosphatase assay for calcineurin, addition of 1 molar equivalent of FKBP-FK506 complex inhibited over 80% of the calcineurin protein phosphatase activity. (mixed with 300 nM calmodulin-calcineurin and 30 µM FK506) (Liu et al. 1992) Dose-response analysis of the effects of FK506 on calcineurin-mediated phosphatase activity in KiSVMC4W cells showed that increased expression of FKBP12 resulted in a greater than ten-fold increase in the sensitivity of the KiSV-MC4W cells containing human FKBP12 cDNA to FK506-mediated inhibition of calcineurin phosphatase activity, as indicted by an IC50 value of ~3 nM (Fruman et al.1995). The phosphatase assay showed that FK506 inhibition of calcineurin activity was concentration-dependent and that IC50 values for calcineurin inhibition were approximately 0.5 nM for FK 506 (Fruman et al.1992).

References

 

1. Barik, S. (2006). Immunophilins: for the love of proteins. Cellular and Molecular Life Sciences 63(24): 2889-900.
2. Bierer, B.E., Holländer, G., Fruman, D. and Burakoff, S.J. (1993). Cyclosporin A and FK506: molecular mechanisms of immunosuppression and probes for transplantation biology. Current opinion in immunology 5 (5): 763-73.
3. Bram, R.J., Hung, D.T., Martin, P.K., Schreiber, S.L. and Crabtree, G.R. (1993). Identification of the immunophilins capable of mediating inhibition of signal transduction by cyclosporin A and FK506: roles of calcimeurin binding and cellular location. Molecular and cellular biology 13 (8): 4760-9.
4. Cameron, A.M., Nucifora, F.C. Jr., Fung, E.T., Livingston, D.J., Aldape, R.A., Ross,C.A. and Snyder, S.H. (1997). FKBP12 binds the inositol 1,4,5-trisphosphate receptor at leucine-proline (1400-1401) and anchors calcineurin to this FK506-like domain. The Journal of biological chemistry 272 (44): 27582-8.
5. Fruman, D.,A., Klee, C. B., Bierer, B. E. and Burakoff, S. J. (1992). Calcineurin phosphatase activity in T lymphocytes is inhibited by FK 506 and cyclosporin A. Proceedings of the National Academy of Sciences of the United States of America. 89(9):3686-90.
6. Fruman, D. A., Bierer, B. E., Benes, J. E., Burakoff, S. J., Austen, K. F. and Katz, H. R. (1995). The complex of FK506-binding protein 12 and FK506 inhibits calcineurin phosphatase activity and IgE activation-induced cytokine transcripts, but not exocytosis, in mouse mast cells. Journal of Immunology.154(4):1846-51.
7. Hultsch, T., Albers, M. W., Schreiber, S.L. and Hohman, R. J. (1991). Immunophilin ligands demonstrate common features of signal transduction leading to exocytosis or transcription. Proceedings of the national academic science of the United States of America. 14: 6229-6233.
8. Kang, C. B., Hong, Y., Dhe-Paganon, S. and Yoon, H. S. (2008). FKBP family proteins: immunophilins with versatile biological functions. Neurosignals. 16: 318-325.
9. Kincaid, R..L. (1993). Calmodulin-dependent protein phosphatases from microorganisms to man. A study in structural conservatism and biological diversity. Adv Second Messenger Phosphoprotein Res. 27:1-23.
10. Klee, C. B., Draetta, G. F. and Hubbard, M. J. (1988). Calcineurin. Advances in enzymology and related areas of molecular biology. 61:149-200.
11. Liu, J., Farmer, J. D. Jr., Lane, W. S., Friedman, J., Weissman, I. and Schreiber, S. L. (1991). Calcineurin is a common target of cyclophilin-cyclosporin A and FKBP-FK506 complexes. Cell. 66(4): 807-815.
12. Liu, J., Albers, M. W., Wandless, T. J., Luan, S., Alberg, D. G., Belshaw, P. J., Cohen, P., MacKintosh, C., Klee, C. B. and Schreiber, S.L.. (1992). Inhibition of T cell signaling by immunophilin-ligand complexes correlates with loss of calcineurin phosphatase activity. Biochemistry. 31(16):3896-901.
13. Liu, J. (1993). FK506 and cyclosporin, molecular probes for studying intracellular signal transduction. Immunology today. 14(6): 290-305.
14. Panhans-Gross, A., Novak, N., Kraft, S. and Bieber, T. (2001). Human epidermal Langerhans' cells are targets for the immunosuppressive macrolide tacrolimus (FK506). Journal of Allergy and Clinical Immunology 107(2): 345-52.
15. Rao, A., Luo, C. and Hogan, PG. (1997). Transcription factors of the NFAT family: regulation and function. Annual Review of Immunology 15: 707-47.
16. Schreiber, SL. and Crabtree, GR. (1992). The mechanism of action of cyclosporin A and FK506. Immunology Today 13(4): 136-42.
17. Siekierka, JJ., Hung, SH., Poe, M., Lin, CS. and Sigal, NH. (1989). A cytosolic binding protein for the immunosuppressant FK506 has peptidyl-prolyl isomerase activity but is distinct from cyclophilin. Nature 341(6244): 755-57.
18. Siekierka, JJ., Wiederrecht, G., Greulich, H., Boulton, D., Hung, SH., Cryan, J., Hodges, PJ. and Sigal, NH. (1990). The cytosolic-binding protein for the immunosuppressant FK-506 is both a ubiquitous and highly conserved peptidyl-prolyl cis-trans isomerase. Journal of Biological Chemistry 265(34): 21011-5.
19. Zhang, B.W., Zimmer, G., Chen, J., Ladd, D., Li, E., Alt, F.W., Wiederrecht, G., Cryan, J., O'Neill, E.A., Seidman, C.E., Abbas, A.K. and Seidman, J.G. (1996). T cell responses in calcineurin A alpha-deficient mice. Journal of experimental medicine 183(2): 413-20.
20. Wang, P. and Heitman, J. (2005) The cyclophilins. Genome Biology 6 (7):226.

Inhibition, Calcineurin Activity leads to Interference, nuclear localization of NFAT

AOPs Referencing Relationship

Evidence Supporting Applicability of this Relationship

NFAT expresses in B cells, mast cells, neutrophil granulocytes, dendritic cells, macrophages, and natural killer cells as well as T cells from humans, rodents, and other mammalian species. (Rao et al. 1997)

How Does This Key Event Relationship Work

The nuclear factor of activated T cells (NFAT) is a substrate of calcineurin (Rao et al. 1997). When calcineurin activates through stimulus from outside of the cell, it binds directly to the N‑terminal of NFAT in cytoplasm, after which dephosphorylation of SP motifs exposes NLS and covers NES, thereby promoting nuclear localization of NFAT. (Matsuda and Koyasu 2000, Zhu and McKeon 1999). When T-cell activation takes place, T-cell receptor (TCR)-mediated stimulus increases the intracellular concentration of calcium and activates CnB, which subsequently induces CnA phosphatase activation, leading to dephosphorylation of NFAT followed by nuclear localization.

When CN activity is inhibited by the binding of immunophilin complexes, dephosphorylation does not occur in NFAT, thereby interfering with nuclear localization.

Weight of Evidence

Biological Plausibility

A NFAT has an N-terminal with a plurality of SP motifs rich in serine and proline, which are controlled by means of phosphorylation and dephosphorylation. There is a nuclear localization signal (NLS) held between these SP regions as well as a nuclear export signal (NES) in the N-terminal adjacent to the SP motifs. (Beals et al. 1997, Zhu and McKeon 1999, Serfling et al. 2000) SP motifs ordinarily phosphorylate, which covers the NLS and leaves NES exposed, so NFAT localizes in cytoplasm. Immunophilin complexes such as FK506-FKBP12 inhibit calcineurin phosphatase activation, thereby interfering with NFAT nuclear localization.

Empirical Support for Linkage

Nothing in particular

Uncertainties or Inconsistencies

Nothing in particular

Quantitative Understanding of the Linkage

Interference with translocation of NFAT to the nucleus is detected using a gel mobility shift assay to test nuclear extracts and cytoplasmic extracts. (Flanagan et al. 1991) NFAT translocation has been found to be regulated by the concentration of CNI additives.

References

1. Beals, C.R., Clipstone, N.A., Ho, S.N. and Crabtree, G.R. (1997). Nuclear localization of NF-ATc by a calcineurin-dependent, cyclosporin-sensitive intramolecular interaction. Genes & development 11 (7): 824-34.
2.  
3. Bhattacharyya, S., Deb, J., Patra, A.K., Thuy Pham, D.A., Chen, W., Vaeth, M., Berberich-Siebelt, F., Klein-Hessling, S., Lamperti, E.D., Reifenberg, K., Jellusova, J., Schweizer, A., Nitschke, L., Leich, E., Rosenwald, A., Brunner, C., Engelmann, S., Bommhardt, U., Avots, A., Müller, M.R., Kondo, E. and Serfling, E. (2011). NFATc1 affects mouse splenic B cell function by controlling the calcineurin-NFAT signaling network. The Journal of experimental medicine 208 (4): 823-39.
4.  
5. Matsuda, S., Koyasu, S. (2000). A second target of cyclosporin A and FK506. Tanpakushitsu kakusan koso. 45(11): 1823-1831.
6.  
7. Rao, A., Luo, C., and Hogan, PG. (1997). Transcription factors of the NFAT family: regulation and function. Annual Review of Immunology 15: 707-47.
8.  
9. Serfling, E., Berberich-Siebelt, F., Chuvpilo, S., Jankevics, E., Klein-Hessling, S., Twardzik, T., and Avots, A., (2000). The role of NF-AT transcription factors in T cell activation and differentiation. Biochimica et Biophysica Act 1498 (1): 1-18.
10.  
11. Zhu, J. and McKeon, F. (1999). NF-AT activation requires suppression of Crm1-dependent export by calcineurin. Nature. 398(6724): 256-60.

Interference, nuclear localization of NFAT leads to Reduction, NFAT complex formation

AOPs Referencing Relationship

Evidence Supporting Applicability of this Relationship

NFAT expresses in B cells, mast cells, neutrophil granulocytes, dendritic cells, macrophages, and natural killer cells as well as T cells from humans, rodents, and other mammalian species. (Rao et al. 1997)

How Does This Key Event Relationship Work

T-cell receptor (TCR)-mediated stimulus increases the intracellular concentration of calcium and activates CnB, which subsequently induces CnA phosphatase activation, leading to dephosphorylation of NFAT followed by nuclear localization (Matsuda and Koyasu 2000, Zhu and McKeon 1999).

Nuclear localization of NFAT results in the NFAT binding with AP‑1 at the IL-2 promoter region, (Schreiber and Crabtree 1992; Jain et al. 1992) and induces transcription of IL-2. (Jain et al. 1993) In addition to IL-2, NFAT localized in the nucleus of T cells also binds to the promoter region of the other classes of cytokines including IL-4 and IL-13.

Weight of Evidence

Biological Plausibility

Interference with the nuclear localization of NFAT in T cells leads to a reduction in the formation of NFAT complexes, thereby suppressing transcription of IL‑2, IL‑4, and a number of other cytokines.

In contrast to T-cells, B-cell receptor (BCR)-mediated increases in intracellular concentration of calcium in B cells leads to NFAT nuclear localization, thereby producing some classes of cytokines in the same manner as T-cells (Bhattacharyya et al.2011). However, there has been no report of any evidence that CNI acts directly on B cells to effect antibody production.

Empirical Support for Linkage

The following phenotypes have been observed in NFAT knockout mice: moderate hyperproliferation with splenomegaly, reduced proliferative responses by T cells, mild hyperactivation of peripheral T cells. (Macian. 2005)

This can be understood as an indication that NFAT has no direct relation to the division of T cells and that NFAT is related to the production of some albeit not all classes of cytokines. Additionally, it is conceivable that CNI suppresses some induction of transcription related to NFAT.

Uncertainties or Inconsistencies

Nothing especially

Quantitative Understanding of the Linkage

inhibitor concentrations up to 1 μM (1000 nM). Higher concentrations induced cellular toxicity and resulted in cell death. Dose-dependent interference of nuclear NFAT1 translocation per calcineurin inhibition was also observed in CD4+ T cells from healthy donors, again at maximal concentrations of 1 μM. (Maguire et al. 2013).

Nevertheless, there are no reports of direct evidence that interference with the binding of NFAT–AP-1 complexes at the site of cytokine promoters is dependent on CNI dosage.

References

1. Matsuda, S., Koyasu, S. (2000). A second target of cyclosporin A and FK506. Tanpakushitsu kakusan koso. 45(11): 1823-31.
2. Zhu, J. and McKeon, F. (1999). NF-AT activation requires suppression of Crm1-dependent export by calcineurin. Nature. 398(6724): 256-60.
3. Schreiber, SL., and Crabtree, GR. (1992). The mechanism of action of cyclosporin A and FK506. Immunology Today 13(4): 136-42.
4. 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): 801-4.
5. 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): 837-48.
6. Matsuda S, Shibasaki F, Takehana K, Mori H, Nishida E, Koyasu S. (2000) Two distinct action mechanisms of immunophilin–ligand complexes for the blockade of T-cell activation. EMBO Rep. 1(5):428-34.
7. Macian, F. (2005). NFAT proteins: key regulators of T-cell development and function. Nat Rev Immunol. 5(6): 472-84.
8. Maguire O, Tornatore KM, O'Loughlin KL, Venuto RC and Minderman H. (2013) Nuclear translocation of nuclear factor of activated T cells (NFAT) as a quantitative pharmacodynamic parameter for tacrolimus. Cytometry A. 83(12):1096-104.
9. Rao, A., Luo, C., and Hogan, PG. (1997). Transcription factors of the NFAT family: regulation and function. Annual Review of Immunology 15: 707-47.

Reduction, NFAT complex formation leads to Suppression, IL-2 and IL-4 production

AOPs Referencing Relationship

Evidence Supporting Applicability of this Relationship

In addition to data on suppression of cytokine production by CNI in rodents, the following is known from the use of human cells. That is to say, FK506 inhibited expression of both IL-2 and mRNA in human anti-CD3/PMA-activated cells. (Dumont et al. 1998) Also, FK506 suppresses expression of IL-2R (CD25) and costimulatory molecules CD80 (B7.1)/CD40 in human Langerhans cells. (Panhans-Gross A et al. 2001)

How Does This Key Event Relationship Work

NFAT–AP-1 complexes appear to bind to promoter or enhancer regions of cytokine genes in T cells. There are reports that production of IL-2 after activation of T cells is suppressed by calcineurin inhibitor treatment in vitro and in vivo as the result of interference to nuclear translocation of NFAT. (Dumont et al. 1998) Production in T cells of IL‑4 and other classes of cytokines is also suppressed in the same manner as IL-2.

Weight of Evidence

Biological Plausibility

In addition to NFAT–AP-1 complexes, NFAT forms complexes at the site of IL-3 and IL-4 enhancers with avian musculoaponeurotic fibrosarcoma oncogene homolog (MAF), early growth response 1 (EGR1), early growth response 4 (EGR4), interferon-regulatory factor 4 (IRF4), octamer-binding transcription factor (OCT), and other transcriptional partners to induce transcription of a variety of cytokines. (Macian 2005) The production of cytokine induced by these transcriptional partners also suppressed by CNI.

FK506 suppresses expression of IL-2R (CD25) and costimulatory molecules CD80 (B7.1)/CD40 in Langerhans cells. (Panhans-Gross A et al. 2001)

In human NK cells, FK506 suppresses IL-2 responsive proliferation and cytokine production as well as lowers cytotoxicity directed toward K562 tumor cells. (Kim et al. 2010) FK506 suppresses IL-2 production of NKT cell line DN32.D3 induced by stimulus from phorbol 12-myristate 13-acetate (PMA)/calcium-ionophore. (van Dieren et al. 2010)

The relationship between the above mechanisms and NFAT is unclear.

Empirical Support for Linkage

There is no evidence of a relationship between FKBP12 KO and the immune system in the FKBP12 knockout mouse model.

The following phenotypes are observed in NFAT knockout mice: moderate hyperproliferation with splenomegaly; moderately enhanced B- and T-cell responses, with bias towards Th2-cell responses; decreased IFN-γ production in response to TCR ligation; reduced proliferative responses by T cells; impaired repopulation of the thymus and lymphoid organs; impaired Th2-cell responses and IL-4 production; grossly impaired T-cell effector functions, with profound defects in cytokine production and cytolytic activity; B-cell hyperactivity; impaired development of CD4 and CD8 single-positive cells, with increased apoptosis of double-positive thymocytes; and mild hyperactivation of peripheral T cells. (Macian, 2005)

Uncertainties or Inconsistencies

Nothing in particular

Quantitative Understanding of the Linkage

FK506 suppressed, in a dose-dependent (1.2 to 12.5 nM) manner, production of IL-2, IL-4, and IFN-γ by human T cells stimulated with anti-CD3 mAb in the presence of PMA, as well as inhibited, also in a dose-dependent (10 nM) manner, expression of IL-2, IL-4, and IFN-γ mRNA in anti-CD3/PMA-activated cells. (Dumont et al. 1998)

In addition to the above quantitative expressions of cytokine production in T cells, it is possible to measure quantitatively expression of IL‑2 receptors and IL‑2 production in other immune cells per CNI. That is to say, FK506 suppresses, in a dose-dependent (0.1 to 1000 nM) manner, expression of IL-2R (CD25), as well as suppresses, also in a dose-dependent (10 nM) manner, expression of costimulatory molecules CD80 (B7.1)/CD40 in Langerhans cells. Thus, FK506 is roughly 100 times more potent than β-Methasone valerate at inhibiting LC stimulatory function. (Panhans-Gross A et al. 2001)

References

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. Kim, T., Kim, N. and Kang, H. J. (2010). FK506 causes cellular and functional defects in human natural killer cells. Journal of leukocyte biology. 88:1089-1097.
3. Macian, F. (2005) NFAT proteins: key regulators of T-cell development and function. Nat Rev Immunol. 5(6): 472-84.
4. Panhans-Gross, A., Novak, N., Kraft, S., and Bieber, T. (2001). Human epidermal Langerhans’ cells are targets for the immunosuppressive macrolide tacrolimus (FK506). Journal of Allergy and Clinical Immunology 107(2): 345-52.
5. van Dieren, J.M., Lambers, M.E.H., Kuipers, E.J., Samsom, J.N., van der Woude, C.J. and Nieuwenhuis, E.E.S. (2010). Local immune regulation of mucosal inflammation by tacrolimus. Digestive diseases and sciences 55(9): 2514-19.
6. Zhang, BW., Zimmer, G., Chen, J., Ladd, D., Li, E., Alt, FW., Wiederrecht, G., Cryan, J., O'Neill, EA., Seidman, CE., Abbas, AK., Seidman, JG. (1996). T cell responses in calcineurin A alpha-deficient mice. J Exp Med. 183(2): 413-20.

 

Suppression, IL-2 and IL-4 production leads to Impairment, T-cell dependent antibody response

AOPs Referencing Relationship

Evidence Supporting Applicability of this Relationship

The effects of FK506 on production of IL-2, IL-4, and other B-cell stimulating cytokines as well as production of T-cell dependent antibodies have been demonstrated in vitro and in vivo using mice and rats. Also, FK506 contributes to the regulation of cytokine secretion in normal human T cells. (Dumont et al. 1998) FK506 strongly suppressed humoral immunity (antibody production) with a marginal decrease in the number of spleen cells. FK506 showed higher activity than CS on PFC response in mice. (Kino, T et al. 1987a, 1987b)

How Does This Key Event Relationship Work

Experiments have shown that IL-2 mRNA expression in T cells stimulated with anti-CD3/ anti-CD28 antibodies decrease after treatment with FK506. (Dumont et al. 1998). Additionally other experiments have shown that mRNA levels of IL-2, IL-4, and other B cell stimulatory cytokines decreases in T cells stimulated with anti-CD3/anti-CD28 antibodies after treatment with FK506. These results suggest that FK506 is a potent inhibitor of T cell-dependent antibody production. (Heidt, S. et al. 2009) Thus, after treatment with FK506, production of IL‑2, IL‑4, and other cytokines decreases in T cells, reducing stimulation of B cells as well as proliferation, activation, and class switching, and leading to impairment of TDAR.

Weight of Evidence

Biological Plausibility

In humoral immunity, calcineurin inhibitors (CNI) do not affect B cells directly but rather indirectly through T cells. (Heidt et al, 2009) CNI suppress cytokine mRNA expression levels in T cells that stimulate proliferation of B cells as well as B cell activation and class switching induced by IL-2 and IL-4. IL-2 stimulates B cells to proliferate through the surface IL-2 receptors. IL-4 stimulates B cells to proliferate, to induce class switching, and to differentiate into plasma and memory cells. Calcineurin inhibitors inhibit transcription of IL-2, IL‑4, and other cytokines to suppress production of T-cell dependent antibodies. In tests examining antibody production in blood samples obtained from blood-bank donors, PBMC treated with FK506 suppressed the production of immunoglobulin (Ig) M and G antibodies to T-cell dependent antigens. (Heidt et al, 2009)

Empirical Support for Linkage

FK506 is capable of inhibiting immunoglobulin production when B cells are cultured with non-pre-activated T cells, but FK506 fails to inhibit immunoglobulin levels when pre-activated T cells are used to stimulate B cells. The inhibition of B cell response by FK506 appears due solely to inhibition of T helper cells. (S.Heidt et al. 2009)

Quantitative Understanding of the Linkage

FK506 suppressed, in a dose-dependent (1.2 to 12.5 nM) manner, production of IL-2, IL-4, and IFN-γ in human T cells stimulated with anti-CD3 mAb in the presence of PMA, and inhibited, in a dose-dependent (10 nM) manner, expression of IL-2, IL-4, and IFN-γ mRNA expression in anti-CD3/PMA-activated cells. (Dumont et al. 1998).

References

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. 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. Boussiotis, V.A., Nadler, L.M., Strominger, J.L. and Goldfeld, A.E. (1994). Tumor necrosis factor alpha is an autocrine growth factor for normal human B cells. Proceedings of the National Academy of Sciences of the United States of America 91 (15): 7007-11.
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