Abstract Number: PB1460
Meeting: ISTH 2020 Congress
Background: Ticks puncture human and animals´ skin and secrete saliva into the bloodstream of the host, containing anticoagulant proteins. We studied disagregin, a potent platelet-inhibiting protein derived from the salivary glands of Ornithodoros moubata, an African soft tick. Conventional αIIbβ3 antagonists contain an RGD-sequence for integrin binding. In contrast, disagregin contains an RED-sequence, hypothesizing a differential mode of inhibitory action.
Aims: We aimed to compare the inhibitory effects of native disagregin and its RGD-mutant on platelet activation and to unravel the molecular basis of disagregin-αIIbβ3 integrin interactions.
Methods: Disagregin (MW: 6952 Da) was synthesized by solid-phase peptide synthesis using tert-butyloxycarbonyl(Boc)-chemistry and native chemical ligation. Effects of both disagregins on platelet aggregation were assessed by light transmission aggregometry in human platelet-rich plasma. Whole-blood thrombus formation was investigated by perfusing blood over collagen type I with and without tissue factor (500 pM) at high wall-shear rate (1000s-1) in the presence of disagregin, disagregin-RGD or eptifibatide, a clinically approved αIIbβ3 integrin inhibitor. Thrombi formed on microspots were assessed by 9 parameters, reflecting platelet adhesion, thrombus formation and fibrin formation. Parameters were analysed by Friedman tests.
Results: Disagregin showed dose-dependent inhibition of collagen- and adenosinediphosphate (ADP)-induced classical platelet aggregation with IC50 values of 65 and 100 nM, respectively. Multiparameter assessment of thrombus formation showed suppressed platelet adhesion and aggregate formation in presence of disagregin or disagregin-RGD, while eptifibatide treatment allowed platelets to form small aggregates.
Time to fibrin formation was delayed when adding disagregin or disagregin-RGD (p< 0.01) or eptifibatide (p< 0.05). However, surface covered by fibrin was lower in presence of disagregin and disagregin-RGD (p< 0.01), while addition of eptifibatide did not affect fibrin surface area coverage.
Conclusions: Both αIIbβ3-blocking disagregins have potent inhibitory effects on platelet adhesion, thrombus formation and fibrin formation in a dose-dependent manner. Eptifibatide was less effective, at concentrations fully blocking classical platelet aggregation.
To cite this abstract in AMA style:van den Kerkhof DL, Nagy M, Heemskerk JWM, Hackeng TM, Dijkgraaf I. Inhibition of Platelet Adhesion, Thrombus Formation and Fibrin Formation by a Potent αIIbβ3 Integrin Inhibitor [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/inhibition-of-platelet-adhesion-thrombus-formation-and-fibrin-formation-by-a-potent-%ce%b1iib%ce%b23-integrin-inhibitor/. Accessed September 24, 2023.
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