Chimeric Antithrombin Peptide

We investigated the properties of an artificial chimeric peptide that contains an Arg-Gly-Asp (RGD)tripeptide, the versatile cell recognition signal of extracellular matrix protein components, coupled to a carboxyl-terminal fragment of the highly specific a-thrombin hibitor, hirudin (residues 53-64): WG-SANGDFEEIPEEYL (RGD-hir~din'~-'~). Hir ~ d i n ~ ~ ' ~ and RGD-hir~din'~-'~ inhibited the fibrinogen clotting activity of a-thrombin and prolonged the activated partial thromboplastin time of human plasma. In addition, both peptides afforded total protection to thrombin from trypsinolysis. Neither h i r~din '~'~ nor R G D h i r ~ d i n ~ ~ ' ~ dramatically interfered with the thrombin-antithrombin inhibition reaction either in the absence or presence of added heparin. a-Thrombin-induced platelet aggregation was effectively inhibit d by and RGDhir~din'~'~. Unlike h i r ~ d i n ~ ~ ' ~ , R G D h i r ~ d i n ~ ~ ' ~ in solution inhibited integrin-mediated endothelial cell and fibroblast cell attachment to polystyrene wells in the presence of fetal bovine serum. Collectively, our results demonstrate that RGD-hir~din'~-'~ has anticoagulant/antiplatelet aggregation activity attributable to its hirudin sequence and integrin-directed cell attachment activity due to its RGD site. Our results suggest that this chimeric motif may serve as a prototype for a new class of anticoagulants where an integrin-specific sequence utargets" the peptide to a cell (ultimately through the platelet integrin trapped amid a thrombus with ensuing proteinase inhibition.