Purification and characterisation of soluble intercellular adhesion molecule-1 and its effect on cell-mediated cytolysis of bladder tumour cells.

Intercellular adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein and a major ligand for the integrin leukocyte function-associated antigen1 (LFA-1). The interaction between ICAM-1 and LFA-1 plays an important role in many lymphocyte activities during inflammatory and immunological reactions. Recently, a soluble form of ICAM-1 (SICAM-1) has been detected in normal serum [ 13 and we have previously demonstrated the secretion of SICAM-1 by cytokine stimulated bladder tumour cell lines as well as its detection in the urine of patients receiving bacillus Calmette-Guerin (BCG) immunotherapy for bladder cancer 121. Reports of a functional role for SICAM-1 are conflicting. While some have suggested that SICAM-1 can inhibit cell-cell adhesion events there are several which have failed to demonstrate this. In an attempt to elucidate a role for SICAM-1 we studied whether highly purified SICAM-1 could compete with membrane bound ICAM-1 for lymphocyte binding, thus allowing tumour cells to evade immunodetection. This was approached by isolating and characterising SICAM-1 from a variety of starting materials and determining their effect on the lymphokine-activated killer (LAK) cell-mediated cytolysis of bladder tumour cells in vitm. Soluble ICAM-1 was isolated by immunoaffinity chromatography from normal human serum, bladder tumour cell culture supernatants and the urine of patients receiving BCG immunotherapy for bladder cancer on two different anti-ICAM-1 monoclonal antibody (MAb) columns. Figure 1 shows that the SICAM-1 isolated from all three sources resolved as a single diffuse band of molecular mass 80 to 90 kDa under both nonreducing and reducing conditions characteristic of a heavily glycosylated glycoprotein and that this was slightly higher than the mass found for recombinant SICAM-1. The molecular mass range obtained is in the expected size range for monomeric SICAM-1 and is in agreement with previous results [1,31. Monomeric SICAM-1 idated from serum retained its ability to inhibit the binding of an anti-ICAM-1 MAb to ICAM-1 positive target cells. In contrast, physiological concentrations of monomeric ICAM1 isolated from serum and urine failed to inhibit the LAK cell mediated cytolysis of four bladder tumour cell lines (Table 1). However, sensitivity to lysis could be effectively blocked by the addition of antibodies to both ICAM-1 and LFA-1 demonstrating the importance of this adhesion pathway (Table 1). It would therefore appear unlikely that the concentrations of SICAM-1 found in the urine following BCG immunotherapy could antagonise LFAl/ICAM1 mediated events in vivo in the bladder.