B7-1 and B7-2 Provide Secondary Signals that Regulate T Cell Activation
B7-1, B7-2 and T Cell Activation
CD28 and CTLA-4 are T cell-expressed receptors that bind to the B7 family proteins, B7-1/CD80 and B7-2/CD86, which are primarily expressed on antigen-presenting cells (APCs). While CD28 is constitutively expressed on naïve T cells and provides a co-stimulatory signal upon binding to these ligands, CTLA-4 is up-regulated on T cells following T cell activation and delivers a co-inhibitory signal following ligand-binding. CTLA-4 binds to both B7-1/CD80 and B7-2/CD86 with a higher affinity than CD28, and as a result, the T cell response is down-regulated following up-regulation of CTLA-4.1
CTLA-4 is also constitutively expressed on regulatory T cells and has been shown to remove B7-1/CD80 and B7-2/CD86 from APCs by trans-endocytosis, suggesting another mechanism by which CTLA-4 may down-regulate the T cell response.2, 3 In addition, CTLA-4 induces the production of indoleamine 2,3-dioxygenase (IDO) by dendritic cells (DCs), leading to degradation of the amino acid tryptophan and the production of immunosuppressive metabolites such as kynurenine.4, 5 Both the depletion of tryptophan and the production of kynurenine inhibit T cell activation and proliferation, indicating that CTLA-4 may also indirectly regulate T cell activity by acting on DCs.
Up-regulated and sustained expression of CTLA-4 along with other inhibitory receptors is associated with T cell exhaustion in cancer.6-8 T cell exhaustion is a dysfunctional state that is characterized by reduced T cell proliferation, decreased cytokine production, and reduced cytotoxicity, which leads to a down-regulation of the T cell-mediated anti-tumor immune response. As a result, blockade of CTLA-4 may not only improve anti-tumor immune responses by suppressing CTLA-4-mediated T cell co-inhibitory signaling and inhibiting the suppressive activity of regulatory T cells, but also by partially restoring the functions of exhausted T cells. To date, CTLA-4, along with PD-1, and PD-L1 are the most common proteins being targeted for cancer immunotherapy.8
B7-1/CD80 and B7-2/CD86 Regulate T Cell Activation by Binding to CD28 or CTLA-4
B7-1/CD80 and B7-2/CD86 mediate T cell co-stimulatory and co-inhibitory effects by binding to either CD28 or CTLA-4. Following recognition of the antigenic peptide/major histocompatibility complex (MHC) by the T cell receptor (TCR), binding of B7-1/CD80 or B7-2/CD86 to CD28 provides a co-stimulatory signal to promote T cell activation. Following T cell activation, the co-inhibitory receptor, CTLA-4, is up-regulated and binds with higher affinity to B7-1/CD80 and B7-2/CD86 than CD28, resulting in a down-regulation of T cell activity. In cancer, tumor-associated macrophages express B7-1/CD80 and B7-2/CD86, which allows them to suppress the functions of activated T cells through CTLA-4. Up-regulated expression of CTLA-4 and other inhibitory receptors is also associated with tumor-specific T cell exhaustion. This dysfunctional T cell state is characterized by a decrease in proliferative capacity and reduced T cell effector functions that prevent effective tumor elimination.
Analysis of the Binding Properties of R&D Systems B7-1 and CTLA-4 Proteins by SPR
Affinity Measurements and Binding Kinetics of the CTLA-4:B7-1 Interaction by Surface Plasmon Resonance. Sensorgram data of captured Avi-tag Biotinylated Recombinant Human CTLA-4 Fc Chimera (R&D Systems, Catalog # AVI7268) binding to Recombinant Human B7-1 Fc Chimera (R&D Systems, Catalog # 10133-B1). Briefly, Avi-tag Biotinylated Recombinant Human CTLA-4 Fc Chimera was captured at a low coupling density to the active flow cell via the Avi-tag biotin. Recombinant Human B7-1 Fc Chimera at a concentration range between 82 pM and 21 nM was flowed over both active and uncoupled reference flow cells at each concentration. Kinetic sensorgrams were fit to a 1:1 binding model to determine the binding kinetics and affinity, with an interaction affinity of KD=0.2511 nM. The corresponding overlaid kinetic fits with the residual plot shown below. The corresponding steady state affinity fit is shown at the bottom. The experiment was performed on a Biacore T200, GE Healthcare.
Assessment of the Bioactivity of R&D Systems Recombinant Human CTLA-4 Protein
CTLA-4 Inhibits IL-2 Secretion by Jurkat Cells. Jurkat human acute T cell leukemia cells were stimulated with Recombinant Human B7-1/CD80 Fc Chimera (R&D Systems, Catalog # 140-B1; 1 μg/mL) and then treated with the indicated concentrations of Recombinant Human CTLA-4 Fc Chimera (R&D Systems, Catalog # 7268-CT). IL-2 secretion was measured in cell culture supernatants using the Human IL-2 Quantikine® ELISA Kit (R&D Systems, Catalog # D2050). The ED50 for this effect is 0.03-0.15 μg/mL.
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T cell activation occurs following recognition of the antigenic peptide/major histocompatibility complex (MHC) by the T cell receptor (TCR) and a second antigen-independent co-stimulatory signal. Together, these two events result in the initiation of intracellular signaling pathways that lead to T cell proliferation, differentiation, and cytokine production. T cell activation is a critical part of the adaptive immune response as it helps to regulate the immune response to address specific microbial pathogens, leads to the elimination of infected or damaged cells, and promotes the activities of other immune cell types. The process of T cell activation, however, also must be precisely regulated to ensure overall immune homeostasis. Without this regulation, abnormal or excessive T cell activation could lead to inflammatory and autoimmune diseases. Recent reports suggest that the signals that tirgger T cell activation also promote the initiation of inhibitory pathways that are necessary to eventually terminate the T cell response.
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Grohmann, U. et al. (2002) CTLA-4-Ig regulates tryptophan catabolism in vivo. Nat. Immunol. 3:1097. PMID: 12368911.
Munn, D.H. et al. (2004) Ligation of B7-1/B7-2 by human CD4+ T cells triggers indoleamine 2,3-dioxygenase activity in dendritic cells. J. Immunol. 172:4100. PMID: 15034022.
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Seidel, J.A. et al. (2018) Anti-PD-1 and anti-CTLA-4 therapies in cancer: mechanisms of action, efficacy, and limitations. Front. Oncol. 8:86. PMID: 296444214.