Ticagrelor inverse agonist activity at the P2Y₁₂ receptor is non-reversible versus its endogenous agonist adenosine 5'-diphosphate

Background and Purpose: Ticagrelor is labelled as a reversible, direct-acting platelet P2Y(12) receptor (P2Y(12)R) antagonist that is indicated clinically for the prevention of thrombotic events in patients with acute coronary syndrome (ACS). As with many antiplatelet drugs, ticagrelor therapy increases bleeding risk in patients, which may require platelet transfusion in emergency situations. The aim of this study was to further examine the reversibility of ticagrelor at the P2Y(12)R. Experimental Approach: Studies were performed in human platelets, with P2Y(12)Rstimulated GTPase activity and platelet aggregation assessed. Cell-based bioluminescence resonance energy transfer (BRET) assays were undertaken to assess G protein-subunit activation downstream of P2Y(12)R activation. Key Results: Initial studies revealed that a range of P2Y(12)R ligands, including ticagrelor, displayed inverse agonist activity at P2Y(12)R. Only ticagrelor was resistant to washout and, in human platelet and cell-based assays, washing failed to reverse ticagrelor-dependent inhibition of ADP-stimulated P2Y(12)R function. The P2Y(12)R agonist 2MeSADP, which was also resistant to washout, was able to effectively compete with ticagrelor. In silico docking revealed that ticagrelor and 2MeSADP penetrated more deeply into the orthosteric binding pocket of the P2Y(12)R than other P2Y(12)R ligands. Conclusion and Implications: Ticagrelor binding to P2Y(12)R is prolonged and more akin to that of an irreversible antagonist, especially versus the endogenous P2Y(12)R agonist ADP. This study highlights the potential clinical need for novel ticagrelor reversal strategies in patients with spontaneous major bleeding, and for bleeding associated with urgent invasive procedures.