CHARACTERIZATION OF MOLECULAR-INTERACTIONS BETWEEN INTERCELLULAR-ADHESION MOLECULE-1 AND LEUKOCYTE FUNCTION-ASSOCIATED ANTIGEN-1

The ability of soluble ICAM-1 (sICAM-1) to inhibit ICAM-1/LFA-1 adhesi on events has been reported previously by numerous investigators. sICA M-1 has been demonstrated to inhibit various in vitro assays at concen trations ranging from 2 nM to greater than 40 mu M. Given the hypothes is that circulating ICAM-1 modulates immune functions, the ability of sICAM-1 to inhibit cellular functions may have significant ramificatio ns. Considering the potential clinical importance of the interaction b etween ICAM-1 and its receptor, LFA-1, it is necessary to understand t his receptor-ligand interaction at a molecular level. In this study, d irect binding experiments were utilized to determine the affinity betw een biotinylated monomeric sICAM-1 and immobilized LFA-1 (approximatel y 130 nM). Competitive binding experiments with unlabeled sICAM-1 and a truncated form of sICAM-1 (D1D2) yielded similar affinities. The spe cificity of this interaction was characterized using mAbs directed aga inst sICAM-1 or LFA-1. This assay system was extended to include multi meric species using nonblocking mAbs directed against domains D4 and D 5 of sICAM-1. Dimerizing sICAM-1 with a mAb alpha D4 or alpha D5 incre ased the affinity for immobilized LFA-1 by two orders of magnitude (si milar to 4 nM), an effect presumably due to avidity. These results ind icate that while the monomeric sICAM-1/LFA-1 interaction may involve o nly a moderate binding affinity, multimeric ICAM-1 present on a cell s urface may bind cell surface-immobilized LFA-1 with very high avidity. These sICAM-1/LFA-1 molecular assays should be useful in defining the efficacy of potential antagonists.