Although endothelial cell retraction is required before tumor cell invasion, its molecular mechanism still remains obscure. We previously demonstrated that conditioned medium (CM) derived from a human pan-creatic cancer cell line, PSN-1, induced endothelial cell retraction and facilitated tumor cell invasion. To investigate the molecular change of events in the transduction of extracellular signals during endothelial cell retraction, we examined the effect of the CM derived from PSN-1 cells on the tyrosine phosphorylation in endothelial cells. Immunoblot analyses revealed that the PSN-1 CM decreased tyrosine phosphorylation of a 120-130 kD protein, and induced the concomitant down-regulation of focal adhesion kinase, pp125 FAK, dur-ing endothelial cell retraction in time- and dose-dependent fashions. These changes preceded endothelial cell retraction and were reversible after removal of the CM. Further quantitative densitometric analyses demon-strated that the extent of decrea se in tyrosine phosphorylated 120-130 kD protein during the endothelial cell retraction was likely to be proportional to that of the down-regulation of pp125 FAK. A tyrosine phosphory-lated 120-130 kD protein immunoprecipitated by anti-phosphotyrosine antibody immunoreacted with anti-pp125 FAK antibody. These results suggested that decreased amount of a tyrosine phosphorylated 120-130 kD protein probably due to the down-regulation of pp125 FAK might be associated with the signal transduction pathway in the endothelial cells during their retraction. Furthermore, these findings were also observed in the CM from another four human cancer cell lines, suggesting the down-regulation of pp125 FAK in endothelial cells during tumor cell invasion.