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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 (αLβ2) 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 αL 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 αL I domain have not been described. Here, we report the identification and characterization of a human antibody AL‐57, which binds to the αL 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‐αL 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.