Abstract
Background and Purpose Biased agonism describes the ability of ligands to differentially regulate multiple signalling pathways when coupled to a single receptor. Signalling is affected by rapid agonist-induced receptor internalisation. Hence, the conventional use of equilibrium models may not be optimal, because (i) receptor numbers vary with time and, in addition, (ii) some pathways may show non-monotonic profiles over time.
Experimental Approach Data were available from internalisation, cAMP inhibition and phosphorylation of ERK (pERK) of the cannabinoid-1 (CB1) receptor using a concentration series of six CB1 ligands (CP55,940, WIN55,212-2, anandamide, 2-arachidonylglycerol, Delta(9)-tetrahydrocannabinol and BAY59,3074). The joint kinetic model of CB1 signalling was developed to simultaneously describe the time-dependent activities in three signalling pathways. Based on the insights from the kinetic model, fingerprint profiles of CB1 ligand bias were constructed and visualised.
Key Results A joint kinetic model was able to capture the signalling profiles across all pathways for the CB1 receptor simultaneously for a system that was not at equilibrium. WIN55,212-2 had a similar pattern as 2-arachidonylglycerol (reference). The other agonists displayed bias towards internalisation compared to cAMP inhibition. However, only Delta(9)-tetrahydrocannabinol and BAY59,3074 demonstrated bias in the pERK-cAMP pathway comparison. Furthermore, all the agonists exhibited little preference between internalisation and pERK.
Conclusion and Implications This is the first joint kinetic assessment of biased agonism at a GPCR (e.g. CB1 receptor) under non-equilibrium conditions. Kinetic modelling is a natural method to handle time-varying data when traditional equilibria are not present and enables quantification of ligand bias.