Development of a small RGD peptide fibrinogen receptor antagonist with potent antiaggregatory activity in vitro

Abstract

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 microM [in vitro antiaggregatory activity employing dog platelet rich plasma (PRP)/ADP], due to low platelet fibrinogen receptor affinity, Ki 2.9 +/- 1.9 microM (purified, reconstituted human platelet GPIIb/IIIa), relative to fibrinogen, Ki 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 (Ki 58 +/- 20 nM purified human receptor), and potency (IC50 0.36 +/- 0.4 microM 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.

Destabilizing loop swaps in the CDRs of an immunoglobulin VL domain

“…Data points are the mean values of three or four independent determinations. The lack of significant temperature dependence to the inhibitory action of the peptide antagonist SKF#106760 (Samanen et al, 1991) and the snake venom disintegrin echistatin (Can et al, 1988) show that the temperature dependence for RGD23 and RGD34 derives from a thermal effect on these REI mutants and not on the receptor. The wild-type REI VL exhibits no binding at any temperature (Lee et al, 1993).…”

Binding Properties of a Peptide Derived from β-Lactamase Inhibitory Protein

“…Binding is mediated by ␤-turn regions within fibrinogen that contain the sequence Arg-Gly-Asp (RGD). A cyclic peptide with the sequence RGDS flanked by cyclic disulfide analogues binds to GPIIb/IIIa and is a potent inhibitor of platelet aggregation (41).…”

Synthesis, Conformational Analysis, and Biological Activity of Proposed Vitronectin Antagonists

“…According to CD, Mamb constrained each peptide into a ßll' turn. These new analogs exhibited higher binding affinities and selectivities for the cqibß3 receptor as well as increased metabolic stability over the most potent compound known at the time, SK&F 106760, which contains a disulfide bridge resulting from the coupling of cysteine and penicillamine side chains [40]. Thus, a conformational change in at least this particular turn mimetic appears to accompany a change in receptor specificity [41].…”