A binding kinetics study of human adenosine A3 receptor agonists

Abstract

The human adenosine A (hA) receptor has been suggested as a viable drug target in inflammatory diseases and in cancer. So far, a number of selective hA receptor agonists (e.g. IB-MECA and 2-Cl-IB-MECA) inducing anti-inflammatory or anticancer effects are under clinical investigation. Drug-target binding kinetics is increasingly recognized as another pharmacological parameter, next to affinity, for compound triage in the early phases of drug discovery. However, such a kinetics-driven analysis has not yet been performed for the hA receptor. In this study, we first validated a competition association assay for adenosine A receptor agonists to determine the target interaction kinetics. Affinities and Kinetic Rate Index (KRI) values of 11 ribofurano and 10 methanocarba nucleosides were determined in radioligand binding assays. Afterwards, 15 analogues were further selected (KRI <0.70 or KRI >1.35) for full kinetics characterization. The structure-kinetics relationships (SKR) were derived and longer residence times were associated with methanocarba and enlarged adenine N and C2 substitutions. In addition, from a k-k-K kinetic map we divided the agonists into three subgroups. A residence time "cliff" was observed, which might be relevant to (N)-methanocarba derivatives' rigid C2-arylalkynyl substitutions. Our findings provide substantial evidence that, next to affinity, additional knowledge of binding kinetics is useful for developing and selecting new hAR agonists in the early phase of the drug discovery process.

Exploring the Effect of Cyclization of Histamine H1 Receptor Antagonists on Ligand Binding Kinetics

“…These results are in line with recent findings for adenosine A 3 receptor antagonists, 38 whereas a series of A 3 agonists showed a better correlation between the affinity and the association rate. 39 A recent study exploring the binding kinetics of histamine H 3 R reference ligands showed a better correlation between the affinity and the association, 40 illustrating that the relationships between affinity and binding kinetics vary with receptors and compounds (series dependent). For the compounds in Table 1 , all tricyclic ligands have a lower dissociation rate k off (longer RT) than the non-tricyclic ligands.…”

Binding kinetics of ligands acting at GPCRs

“…Studies into the kinetic parameters of agonists binding to the muscarinic M 3 receptor and the adenosine A 1 receptor have been carried out in the presence of GTP to promote G protein uncoupling (Sykes et al, 2009; Xia et al, 2016). Kinetic parameters of an agonist (NECA) binding to the adenosine A 2A receptor were found to be unchanged in the presence or absence of GTP (Guo et al, 2012) whereas for the adenosine A 3 receptor it was found that the presence of GTP did not change the association rate constant for agonists but did have an effect on the dissociation rate constants (Xia et al, 2018). Determining kinetic rates in the presence and absence of GTP is likely to become increasingly more important as fluorescent tracers with agonist-like properties become more widely available for the study of unlabelled compound kinetics (Klein-Herenbrink et al, 2016; Sykes et al, 2017).…”

Purinergic Signaling: Impact of GPCR Structures on Rational Drug Design

“…Signaling bias and receptor residence time have been examined for rigidified A 3 AR agonists [79] . A rigid C2 extension, which we proposed to outwardly displace TM2, is a source of signaling bias in addition to increasing A 3 AR vs A 1 /A 2A AR affinity.…”