Comparison of VASP-phosphorylation assay to light-transmission aggregometry in assessing inhibition of the platelet ADP P2Y12 receptor

被引:46
作者
Pampuch, Agnieszka [1 ]
Cerletti, Chiara [1 ]
de Gaetano, Giovanni [1 ]
机构
[1] Catholic Univ, Res Labs, John Paul II Ctr High Technol Res & Educ Biomed S, Lab Cell Biol & Pharmacol Thrombosis, I-86100 Campobasso, Italy
关键词
VASP phosphorylation; platelet aggregation; P2Y(12) ADP receptor antagonist; cangrelor;
D O I
10.1160/TH06-09-0491
中图分类号
R5 [内科学];
学科分类号
1002 ; 100201 ;
摘要
There is need to improve platelet function testing to monitor the response to antiplatelet drugs. We compared flow-cytometric analysis of intraplatelet vasodilator-stimulated phosphoprotein phosphorylation (VASP-P) to light-transmission aggregometry for the detection of drug-induced in-vitro inhibition of the platelet P2Y(12) ADP receptor on 22 healthy subjects (10 males, 12 females, 28.5 +/- 6.6 years). The platelet reactivity index (PRI) of VASP was calculated both from mean fluorescence intensity (MFI) and percent of fluorescence-positive platelets in the presence of PGE(1) with or without ADP (10 mu M). Platelet aggregation was induced by ADP (1.25, 2.5 and 5 mu M). Cangrelor, a competitive inhibitor of the P2Y(12) receptor, preincubated 5 minutes, induced a concentration-dependent inhibition of platelet AIDP-receptor function in both tests. Indeed PRI (%) based on either MFI or percent platelets gated were highly correlated with each other (r = 0.97, p < 0.0001) and with aggregation induced by ADP. The IC50 of cangrelor against each of the three ADP concentrations used in aggregometry increased from 5.8 3.4 nM to 23.1 +/- 4.0 nM and to 98 25 nM, respectively. The IC50 of cangrelor based on VASP-P was within the same range (25.5 +/- 7.7 nM). No correlation was observed between IC50 values of cangrelor and ADP concentrations giving 50% effect (EC50) in the absence of the drug. However, at 10 nM cangrelor seven subjects could be identified by the VASP-P assay as "low responders" to the drug (PRI > 50%), and six of them also had an aggregation response to 5 mu M ADP > 50%. These six subjects showed the lowest ADP EC50 values in the absence of the drug, possibly reflecting high sensitivity of their platelet P2Y(12) receptors to ADP. In conclusion, both the VASP-P assay and light-transmission aggregometry detect in a comparable way in-vitro pharmacological inhibition of the platelet P2Y(12) ADP receptor and its individual variability.
引用
收藏
页码:767 / 773
页数:7
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