In Vitro Dose Response to Different GPIIb/IIIa-Antagonists: Inter-Laboratory Comparison of Various Platelet Function Tests

Abstract

Aims: The aim of this study was to assess the inter- and intra-laboratory variation of the concentration–response to the GPIIb/IIIa-antagonists abciximab and eptifibatide on platelet aggregometry and to compare results with flow cytometric tests as well as the rapid platelet function analyser (RPFA). Methods: In five different laboratory sites, blood from three to five healthy donors was spiked with abciximab or eptifibatide, followed by the assessment of: (1) aggregometry (anticoagulant: sodium citrate 3.18% or hirudin 5 μg/ml); (2) flow cytometry (fibrinogen binding or PAC1-expression), or (3) RPFA. Dose–response curves were established on the basis of a sigmoidal I max-model [ I=( I max* C γ )/(IC 50 γ + C γ )]. Results: For citrated blood, aggregation induced by 20 μM ADP was blocked up to 100% by both GPIIb/IIIa-antagonists, IC 50 values varied between 0.11–0.22 μg/ml for eptifibatide and 1.25–2.3 μg/ml for abciximab. I max of the response to 5 μg/ml collagen ranged from 46% to 100%, and IC 50 values varied between 0.28–0.34 μg/ml for eptifibatide and 2.3–3.8 μg/ml for abciximab. In hirudinized blood, IC 50 values for eptifibatide were 1.5- to 3-fold higher than those obtained with citrated plasma. Inhibition of PAC1-expression by abciximab (IC 50 0.84 μg/ml) showed results similar those of the RPFA (approx. 1.0 μg/ml); larger differences between PAC1 and RPFA results were observed for eptifibatide. Based on aggregometry, eptifibatide concentrations for 80% inhibition varied from 0.27 to 0.55 μg/ml, and were considerably less when the RPFA was taken as basis (0.15 or 0.22 μg/ml). A similar pattern was observed for abciximab. Conclusions: We found quite a low inter- and intra-laboratory variation in the in vitro pharmacodynamic characterization of GPIIb/IIIa-antagonists by aggregometry, making results of these tests obtained from different laboratories during clinical trials at least comparable. The RPFA exhibits a higher sensitivity to inhibitory GPIIb/IIIa-effects, in keeping with the “real” inhibition of the activated receptor (PAC1) as assessed with more elaborate flow cytometry.