Cutting Edge: Monovalency of CD28 Maintains the Antigen Dependence of T Cell Costimulatory Responses

Abstract

CD28 and CTLA-4 are the major costimulatory receptors on naive T cells. But it is not clear why CD28 is monovalent whereas CTLA-4 is bivalent for their shared ligands CD80/86. We generated bivalent CD28 constructs by fusing the extracellular domains of CTLA-4 or CD80 with the intracellular domains of CD28. Bivalent or monovalent CD28 constructs were ligated with recombinant ligands with or without TCR coligation. Monovalent CD28 ligation did not induce responses unless the TCR was coligated. By contrast, bivalent CD28 ligation induced responses in the absence of TCR engagement. To extend these findings to primary cells, we used novel superagonistic and conventional CD28 Abs. Superagonistic Ab D665, but not conventional Ab E18, predominantly ligates CD28 bivalently at low CD28/Ab ratios and induces Ag-independent T cell proliferation. Monovalency of CD28 for its natural ligands is thus essential to provide costimulation without inducing responses in the absence of TCR engagement.

Antibody C Region Influences TGN1412-like Functional Activity In Vitro

“…It was proposed that binding of a superagonistic mAb to the C99D loop of CD28 in the "parallel" conformation transmits a greater perturbation of the cytoplasmic tails of CD28 and hence increased interaction with signaling molecules than binding of conventional, agonistic anti-CD28 mAb to the "relaxed" form of the CD28 homodimer. Although the CD28 homodimer is functionally monovalent for its natural ligand (44), other studies have shown that T cell activation in the absence of TCR engagement only occurs after bivalent ligation (45). Furthermore, bivalent ligation of recombinant CD28 homodimer occurred more efficiently with a superagonistic mAb than with a conventional mAb.…”

Proliferative signals mediated by CD28 superagonists require the exchange factor Vav1 but not phosphoinositide 3‐kinase in primary peripheral T cells

“…Crystallographic evidence and studies of binding stoichiometry suggest that CD28SA similarly induce the formation of a receptor lattice structure [13,16]. These results suggest that CD28SA induces a physiologically relevant, although dysregulated signalling pathway at the extremity of CD28 co-stimulatory signalling, in the same way through which ligation with CD3 antibodies or high-avidity lymphocytic choriomeningitis virus antigens induces co-stimulation-independent signalling at the extremity of the TCR pathway [2,17].…”

“…We have previously generated two functionally distinct types of CD28 antibodies [13][14][15]. Conventional CD28 antibodies recognise an epitope adjacent to the CD80/86-binding site, and like natural ligands, induce proliferative responses only when combined with TCR stimulation.…”

“…Conventional CD28 antibodies recognise an epitope adjacent to the CD80/86-binding site, and like natural ligands, induce proliferative responses only when combined with TCR stimulation. By contrast, CD28SA recognise a membrane proximal epitope and activate T cells without the need for TCR ligation [13,14]. Mitogenic CD28 signals can also be induced after stimulation with the recombinant natural ligand CD80 [13].…”

“…By contrast, CD28SA recognise a membrane proximal epitope and activate T cells without the need for TCR ligation [13,14]. Mitogenic CD28 signals can also be induced after stimulation with the recombinant natural ligand CD80 [13]. However, activation by recombinant CD80 only occurs if the physiologically monovalent CD28 extracellular region is replaced with a bivalent receptor such as the extracellular domains of CTLA-4.…”