DEVELOPMENT OF A SMALL RGD PEPTIDE FIBRINOGEN RECEPTOR ANTAGONIST WITH POTENT ANTIAGGREGATORY ACTIVITY INVITRO

The development of potent antithrombotic agents from the fibrinogen platelet receptor binding sequences Fg-alpha 572-575-Arg-Gly-Asp-Ser- and Fg-gamma 400-411-HHLGGAKQAGDV, believed to be a cryptic RGD-type sequence, is described. The tetrapeptide Ac-RGDS-NH2 itself is capable of inhibiting platelet aggregation in vitro at high concentrations, IC50 91.3 +/- 0.1-mu-M [in vitro antiaggregatory activity employing dog platelet rich plasma (PRP)/ADP], due to low platelet fibrinogen receptor affinity, K(i) 2.9 +/- 1.9-mu-M (purified, reconstituted human platelet GPIIb/IIIa), relative to fibrinogen, K(i) 38.0 +/- 6.0 nM. The peptide is also unstable to plasma, suffering total loss of in vitro activity upon incubation in PRP for 3 h (T1/2 90 min). Only modest improvements in potency were achieved with linear analogues of Ac-RGDS-NH2, while dramatic results were achieved with cyclic analogues, culminating in the cyclic disulfide Ac-cyclo-S,S-[Cys-(N-alpha-Me)Arg-Gly-Asp-Pen]-NH2 (SK&F 106760) with improved plasma stability (100% activity after 3 h), affinity (K(i) 58 +/- 20 nM purified human receptor), and potency (IC50 0.36 +/- 0.4-mu-M dog PRP/ADP). The affinity of this peptide is 2 orders of magnitude greater than that of Ac-RGDS-NH2. The affinity of the analogue is also comparable to fibrinogen. This peptide constitutes a first potent small peptide entry into the class of novel antithrombotic agents called fibrinogen receptor antagonists.