Blockade of T cell activation using a surface-linked single-chain antibodyto CTLA-4 (CD152)

CTLA-4 (CD152) engagement can down-regulate T cell activation and promote t he induction of immune tolerance. However, the strategy of attenuating T ce ll activation by engaging CTLA-4 has been limited by sharing of its natural ligands with the costimulatory protein CD28, In the present study, a CTLA- 4-specific single-chain Ab (scFv) was developed and expressed on the cell s urface to promote selective engagement of this regulatory molecule. Transfe ctants expressing anti-CTLA-4 scFv at their surface bound soluble CTLA-4 bu t not soluble CD28, Coexpression of anti-CTLA-4 scFv with anti-CD3 epsilon and anti-CD28 scFvs on artificial APCs reduced the proliferation and IL-2 p roduction by resting and preactivated bulk T cells as well as CD4(+) and CD 8(+) T cell subsets. Importantly, expression of anti-CTLA-4 scFv on the sam e cell surface as the TCR ligand,vas essential for the inhibitory effects o f CTLA-4-specific ligation, CTLA-4-mediated inhibition of tyrosine phosphor ylation of components of the proximal TCR signaling apparatus was similarly dependent on coexpression of TCR and CTLA-4 ligands on the same surface. T hese findings support a predominant role for CTLA-4 function in the modific ation of the proximal TCR signal. Using T cells from DO11.10 and 2C TCR tra nsgenic mice, negative regulatory effects of selective CTLA-4 ligation were also demonstrated during the stimulation of Ag-specific CD4(+) and CD8(+) T cells by MHC/peptide complexes. Together these studies demonstrate that s elective ligation of CTLA-4 using a membrane-bound scFv results in attenuat ed T cell responses only when coengaged with the TCR during T cell/APC inte raction and define an approach to harnessing the immunomodulatory potential of CTLA-4-specific ligation.