Background: Although many G-protein-coupled receptors (GPCRs) show varying degrees of constitutive activity, a detailed molecular understanding of this phenomena is lacking.  Recent studies have revealed that the platelet expressed P2Y₁₂ receptor (P2Y₁₂R) displays a high degree of constitutive activity and that ticagrelor, a clinical antiplatelet drug, is an inverse agonist at this receptor (Aungraheeta et al., 2016).    

Aims: Use of molecular dynamic simulations (MDs) alongside bioluminescence resonance energy transfer (BRET) assays to further our understanding of the molecular determinants underlying GPCR constitutive activity.

Methods: 1µs MDs of several P2Y12-ligand receptor complexes, employing the ff14SB forcefield. Residues believed to be important for regulating activity were mutated and transiently transfected into HEK293 cells where receptor/G protein coupling was assessed by BRET.

Results: MDs revealed that ticagrelor binds to a region of the receptor similar to that of AZD1283 and 2MeSADP, but not ADP. Ticagrelor interacts with transmembrane domains (TM) 3 and 5-7. ADP sits in an alternative region contacting TM1-3, TM5 and TM7. Principal component analysis reveals that ticagrelor induces movements in TM5 resulting in a shift at the intracellular end, towards TM3. Experimental mutation of C194 to an alanine produced a 64% decrease in ticagrelor inverse agonism.

Conclusions: The orthosteric cavity of the P2Y₁₂R can be divided into two pockets with 2MeS-ADP, AZD1283 and ticagrelor binding in a distinct pocket to ADP. Ticagrelor induces a distinct conformation in TM5 bringing it into closer proximity with TM3. This likely occludes G-protein binding and in part defines the ability of ticagrelor to act as an inverse agonist.
Aungraheeta, R., Conibear, A., Butler, M., Kelly, E., Nylander, S., Mumford, A. and Mundell, S. (2016). Inverse agonism at the P2Y12 receptor and ENT1 transporter blockade contribute to platelet inhibition by ticagrelor. Blood, 128(23), pp.2717-2728. 

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ISTH Congress Abstracts - https://abstracts.isth.org/abstract/defining-the-molecular-features-of-inverse-agonism-insights-from-the-p2y12-receptor-and-the-antiplatelet-drug-ticagrelor/