Regulation of melanoma cell adhesion stabilization to fibronectin

Tumour cell arrest and the formation of stable adhesive interactions between tumour cells and endothelial cells or underlying matrix in the microvasculature are crucial steps in the metastatic process. We have developed a sensitive hydrodynamic adhesion assay to investigate the regulation of melanoma cell adhesion stabilization to the extracellular matrix protein fibronectin. Modulators of human MeWo melanoma Ca2+ concentration and stores, including ionomycin, thapsigargin, dantrolene and caffeine, inhibited cell adhesion stabilization to fibronectin; however, removal of Ca2+ from the extracellular medium did not affect stabilization. The calmodulin inhibitor W-7 and the protein kinase C inhibitor chelerythrine also blocked MeWo adhesion stabilization to fibronectin, as did the tyrosine kinase inhibitor genistein and the cytoskeletal inhibitor cytochalasin D. Manipulation of MeWo cell intracellular CAMP levels had no effect of adhesion stabilization to fibronectin, nor did treatment of cells with phorbol ester, pertussis toxin or cholera toxin. Drug treatments that inhibited adhesion stabilization also had significant effects on the actin cytoskeleton organization of the melanoma cells. This study suggests a role for calcium, calmodulin, protein kinase C and tyrosine kinases in the intracellular regulation of MeWo adhesive stabilization.