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Competitive mode and site of interaction of ticagrelor at the human platelet P 2 Y 12 ‐receptor
Author(s) -
Hoffmann K.,
Lutz D. A.,
Straßburger J.,
Baqi Y.,
Müller C. E.,
Kügelgen I.
Publication year - 2014
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1111/jth.12719
Subject(s) - ticagrelor , p2y12 , agonist , receptor , chemistry , chinese hamster ovary cell , pharmacology , biology , biochemistry , aspirin , clopidogrel
Summary Background The G‐protein‐coupled P2Y 12 ‐receptor plays a crucial role in platelet aggregation. Recently, ticagrelor was licensed as the first perorally active and reversible P2Y 12 ‐receptor antagonist. Objective The present study investigated the site and the antagonistic mode of action of ticagrelor at wild‐type or mutant human P2Y 12 ‐receptors. Methods Recombinant wild‐type or mutant human P2Y 12 ‐receptors were stably expressed in Chinese hamster ovary Flp‐In cells. Receptor function was assessed by quantification of ADP‐ and 2‐methylthio‐ADP‐mediated inhibition of forskolin‐induced cellular cAMP production either using a [ 3 H] cAMP ‐radioaffinity assay or a cAMP response element‐driven luciferase reporter gene assay. Results The natural agonist ADP inhibited forskolin‐induced cAMP formation at the wild‐type P2Y 12 ‐receptor with a lower potency (EC 50 209 n m ) than the synthetic agonist 2‐methylthio‐ADP (EC 50 1.0 n m ). Ticagrelor shifted the concentration‐response curves of both agonists in a parallel and surmountable manner to the right. Increasing concentrations of ticagrelor caused increasing shifts. Schild‐plot analysis revealed pA 2 values of 8.85 for ticagrelor against ADP, and 8.69 against 2‐methylthio‐ADP, and slopes of the regression lines not different from unity. In cells expressing a recombinant C194A 5.43 ‐mutant P2Y 12 ‐receptor construct, ticagrelor lost antagonistic potency when tested against ADP or 2‐methylthio‐ADP. Conclusions The experiments reveal a surmountable and competitive mode of antagonism of ticagrelor at P2Y 12 ‐receptors activated by either the natural agonist ADP or the synthetic agonist 2‐methylthio‐ ADP . Cys194 5.43 is likely to be involved in the interaction of ticagrelor with ADP and 2‐methylthio‐ ADP . The data give new insights into the site and mode of action of ticagrelor at the human P2Y 12 ‐receptor.