We have previously reported that amidinonaphthol derivatives, which have been developed as synthetic serine protease inhibitors, inhibited the binding of adhesive proteins, such as fibrinogen and fibronectin, to ADP-stimulated platelets in a competitive manner. Because this effect was similar to those of Arg-Gly-Asp (RGD) peptides, we examined the effect of amidinonaphthol derivatives on the chemical cross-linking of RGD-peptides to stimulated platelets. The radiolabeled peptides including RGD-sequence (RGDSPASSKP and KYGRGDS) were coupled to platelets by subsequent addition of chemical cross-linking agent. Platelet membrane glycoprotein IIb-IIIa (GPIIb-IIIa) became radiolabeled with the RGD peptide, and stimulation with ADP increased the extent of cross-linking. Cross-linking of the labeled peptides to ADP-stimulated platelets was inhibited by excess of nonlabeled RGD peptides, an amino acid sequence of corresponding to the carboxyl terminus of γ-chain of fibrinogen, fibrinogen and fibronectin, but not by Gly-Arg-Gly-Glu-Ser-Pro (GRGESP).
The cross-linking reaction was inhibited by addition of amidinonaphthol derivatives, such as nafamostat mesilate or FUT-6258, but less effectively by gabexate mesilate, which does not have amidinonaphthol in the structure. The inhibitory effect of nafamostat mesilate was dose-dependent, and 50% inhibition was obtained at the concentration of 6×10-5M.
This result suggested that amidinonaphthol derivatives inhibited the binding of adhesive proteins to platelets by the blockade for RGD peptide binding sites on GPIIb-IIIa.
