Deciphering biased inverse agonism of cangrelor and ticagrelor at P2Y 12 receptor

P2Y 12 receptor (P2Y 12 -R) is one of the major targets for drug inhibiting platelet aggregation in the treatment/prevention of arterial thrombosis. However, the clinical use of P2Y 12 -R antagonists faces some limitations, such as a delayed onset of action (clopidogrel) or adverse effect profile (ticagrelor, cangrelor), justifying the development of a new generation of P2Y 12 -R antagonists with a better clinical benefit–risk balance. Although the recent concept of biased agonism offers the possibility to alleviate undesirable adverse effects while preserving therapeutic outcomes, it has never been explored at P2Y 12 -R. For the first time, using highly sensitive BRET2-based probes, we accurately delineated biased ligand efficacy at P2Y 12 -R in living HEK293T cells on G protein activation and downstream effectors. We demonstrated that P2Y 12 -R displayed constitutive Gi/o-dependent signaling that is impaired by the R122C mutation, previously associated with a bleeding disorder. More importantly, we reported the biased inverse agonist efficacy of cangrelor and ticagrelor that could underlie their clinical efficacy. Our study points out that constitutive P2Y 12 -R signaling is a normal feature of the receptor that might be essential for platelets to respond faster to a vessel injury. From a therapeutic standpoint, our data suggest that the beneficial advantages of antiplatelet drugs might be more related to inverse agonism at P2Y 12 -R than to antagonism of ADP-mediated signaling. In the future, deciphering P2Y 12 -R constitutive activity should allow the discovery of more selective biased P2Y 12 -R blockers demonstrating therapeutic advantages over classical antiplatelet drugs by improving therapeutic outcomes and concomitantly relieving undesirable adverse effects.