Identification and characterization of a human monoclonal antagonistic antibody AL-57 that preferentially binds the high-affinity form of lymphocyte function-associated antigen-1

LFA-1 (α_Lβ₂) mediates cell-cell and cell-extracellular matrix adhesions essential for immune and inflammatory responses. One critical mechanism regulating LFA-1 activity is the conformational change of the ligand-binding α_L I domain from low-affinity (LA), closed form, to the high-affinity (HA), open form. Most known integrin antagonists bind both forms. Antagonists specific for the HA α_L I domain have not been described. Here, we report the identification and characterization of a human antibody AL-57, which binds to the α_L I domain in a HA but not LA conformation. AL-57 was discovered by selection from a human Fab-displaying library using a locked-open HA I domain as target. AL-57 Fab-phage bound HA I domain-expressing K562 cells (HA cells) in a Mg²⁺-dependent manner. AL-57 IgG also bound HA cells and PBMCs, activated by Mg²⁺/EGTA, PMA, or DTT. The binding profile of AL-57 IgG on PBMCs was the same as that of ICAM-1, the main ligand of LFA-1. In contrast, an anti-α_L murine mAb MHM24 did not distinguish between the HA and LA forms. Moreover, AL-57 IgG blocked ICAM-1 binding to HA cells with a potency greater than MHM24. It also inhibited ICAM-1 binding to PBMCs, blocked adhesion of HA cells to keratinocytes, and inhibited PHA-induced lymphocyte proliferation with potencies comparable with MHM24. These results indicate that specifically targeting the HA I domain is sufficient to inhibit LFA-1-mediated, adhesive functions. AL-57 represents a therapeutic candidate for treatment of inflammatory and autoimmune diseases.