The important role of ligand–receptor binding kinetics in drug design and discovery is increasingly recognized by the drug research community. Over the past decade, accumulating evidence has shown... Show moreThe important role of ligand–receptor binding kinetics in drug design and discovery is increasingly recognized by the drug research community. Over the past decade, accumulating evidence has shown that optimizing the ligand’s dissociation rate constant can lead to desirable duration of in vivo target occupancy and, hence, improved pharmacodynamic properties. However, the association rate constant as a pharmacological principle remains less investigated, whereas it can play an equally important role in the selection of drug candidates. This review provides a compilation and discussion of otherwise scarce and dispersed information on this topic, bringing to light the importance of drug−target association in kinetics-directed drug design and discovery. Show less
Ligand-receptor binding kinetics is receiving increasing attention in the drug research community. The Motulsky and Mahan model, a one-state model, offers a method for measuring the binding... Show moreLigand-receptor binding kinetics is receiving increasing attention in the drug research community. The Motulsky and Mahan model, a one-state model, offers a method for measuring the binding kinetics of an unlabelled ligand, with the assumption that the labelled ligand has no preference while binding to distinct states or conformations of a drug target. As such, the one-state model is not applicable if the radioligand displays biphasic binding kinetics to the receptor. receptor ligands. In addition, limitations of the model were investigated as well. H]-NECA was used. The model was further validated by good correlation between simulated results and the experimental data. The two-state model is sufficient to analyse the binding kinetics of an unlabelled ligand, when a radioligand shows biphasic association characteristics. We expect this two-state model to have general applicability for other targets as well. BACKGROUND AND PURPOSE EXPERIMENTAL APPROACH KEY RESULTS CONCLUSION Show less
A considerable number of approved drugs show non-equilibrium binding characteristics, emphasizing the potential role of drug residence times for in vivo efficacy. Therefore, a detailed... Show moreA considerable number of approved drugs show non-equilibrium binding characteristics, emphasizing the potential role of drug residence times for in vivo efficacy. Therefore, a detailed understanding of the kinetics of association and dissociation of a target-ligand complex might provide crucial insight into the molecular mechanism-of-action of a compound. This deeper understanding will help to improve decision making in drug discovery, thus leading to a better selection of interesting compounds to be profiled further. In this review, we highlight the contributions of the Kinetics for Drug Discovery (K4DD) Consortium, which targets major open questions related to binding kinetics in an industry-driven public-private partnership. Show less
The duration of action of adenosine A(2A) receptor (A2A) agonists is critical for their clinical efficacy, and we sought to better understand how this can be optimized. The in vitro temporal... Show moreThe duration of action of adenosine A(2A) receptor (A2A) agonists is critical for their clinical efficacy, and we sought to better understand how this can be optimized. The in vitro temporal response profiles of a panel of A2A agonists were studied using cAMP assays in recombinantly (CHO) and endogenously (SH-SY5Y) expressing cells. Some agonists (e.g., 3cd; UK-432,097) but not others (e.g., 3ac; CGS-21680) demonstrated sustained wash-resistant agonism, where residual receptor activation continued after washout. The ability of an antagonist to reverse pre-established agonist responses was used as a surrogate read-out for agonist dissociation kinetics, and together with radioligand binding studies suggested a role for slow off-rate in driving sustained effects. One compound, 3ch, showed particularly marked sustained effects, with a reversal t(1/2) > 6 hours and close to maximal effects that remained for at least 5 hours after washing. Based on the structure-activity relationship of these compounds, we suggest that lipophilic N6 and bulky C2 substituents can promote stable and long-lived binding events leading to sustained agonist responses, although a high compound logD is not necessary. This provides new insight into the binding interactions of these ligands and we anticipate that this information could facilitate the rational design of novel long-acting A2A agonists with improved clinical efficacy. Show less
Controlling the Dissociation of Ligands from the Adenosine A2A Receptor through Modulation of Salt Bridge StrengthElena Segala, Dong Guo, Robert K. Y. Cheng, Andrea Bortolato, Francesca Deflorian,...Show moreControlling the Dissociation of Ligands from the Adenosine A2A Receptor through Modulation of Salt Bridge StrengthElena Segala, Dong Guo, Robert K. Y. Cheng, Andrea Bortolato, Francesca Deflorian, Andrew S. Doré, James C. Errey, Laura H. Heitman, Adriaan P. IJzerman, Fiona H. Marshall, and Robert M. CookeHeptares Therapeutics Ltd, Biopark Broadwater Road, Welwyn Garden City AL7 3AX, U.K.Division of Medicinal Chemistry, Leiden Academic Centre for Drug Research (LACDR), Leiden University P.O. Box 9502, 2300 RA Leiden, the NetherlandsAbstractThe association and dissociation kinetics of ligands binding to proteins vary considerably, but the mechanisms behind this variability are poorly understood, limiting their utilization for drug discovery. This is particularly so for G protein-coupled receptors (GPCRs) where high resolution structural information is only beginning to emerge. Engineering the human A2A adenosine receptor has allowed structures to be solved in complex with the reference compound ZM241385 and four related ligands at high resolution. Differences between the structures are limited, with the most pronounced being the interaction of each ligand with a salt bridge on the extracellular side of the receptor. Mutagenesis experiments confirm the role of this salt bridge in controlling the dissociation kinetics of the ligands from the receptor, while molecular dynamics simulations demonstrate the ability of ligands to modulate salt bridge stability. These results shed light on a structural determinant of ligand dissociation kinetics and identify a means by which this property may be optimized. Show less
Molecular Basis of Ligand Dissociation from the Adenosine A2A ReceptorDong Guo,Albert C. Pan,Ron O. Dror,Tamara Mocking,Rongfang Liu,Laura H. Heitman,David E. ShawandAdriaan P. IJzermanMole... Show moreMolecular Basis of Ligand Dissociation from the Adenosine A2A ReceptorDong Guo,Albert C. Pan,Ron O. Dror,Tamara Mocking,Rongfang Liu,Laura H. Heitman,David E. ShawandAdriaan P. IJzermanMolecular Pharmacology May 2016, 89 (5) 485-491; DOI: https://doi.org/10.1124/mol.115.102657AbstractHow drugs dissociate from their targets is largely unknown. We investigated the molecular basis of this process in the adenosine A2A receptor (A2AR), a prototypical G protein–coupled receptor (GPCR). Through kinetic radioligand binding experiments, we characterized mutant receptors selected based on molecular dynamic simulations of the antagonist ZM241385 dissociating from the A2AR. We discovered mutations that dramatically altered the ligand’s dissociation rate despite only marginally influencing its binding affinity, demonstrating that even receptor features with little contribution to affinity may prove critical to the dissociation process. Our results also suggest that ZM241385 follows a multistep dissociation pathway, consecutively interacting with distinct receptor regions, a mechanism that may also be common to many other GPCRs. Show less
Ligand–receptor binding kinetics is an emerging topic in the drug research community. Over the past years, medicinal chemistry approaches from a kinetic perspective have been increasingly applied... Show moreLigand–receptor binding kinetics is an emerging topic in the drug research community. Over the past years, medicinal chemistry approaches from a kinetic perspective have been increasingly applied to G protein-coupled receptors including the adenosine receptors (AR), which are involved in a plethora of physiological and pathological conditions. The study of ligand–AR binding kinetics offers room for detailed structure–kinetics relationships next to more traditional structure–activity relationships. Their combination may facilitate the triage of candidate compounds in hit-to-lead campaigns. Furthermore, kinetic studies also help in understanding AR allosterism. Allosteric modulation may yield a change in the activity and conformation of a receptor resulting from the binding of a compound at a site distinct from where the endogenous agonist adenosine binds. Hence, in this Review, we summarize available data and evidence for the binding kinetics of orthosteric and allosteric AR ligands. We hope this Review will raise awareness to consider the kinetic aspects of drug–target interactions on both ARs and other drug targets. Show less
Guo, D.; Dijksteel, G.S.; Duijl, T. van; Heezen, M.; Heitman, L.H.; IJzerman, A.P. 2016
Classical evaluation of target selectivity is usually undertaken by measuring the binding affinity of lead compounds against a number of potential targets under equilibrium conditions, without... Show moreClassical evaluation of target selectivity is usually undertaken by measuring the binding affinity of lead compounds against a number of potential targets under equilibrium conditions, without considering the kinetics of the ligand-receptor interaction. In the present study we propose a combined strategy including both equilibrium- and kinetics-based selectivity profiling. The adenosine receptor (AR) was chosen as a prototypical drug target. Six in-house AR antagonists were evaluated in a radioligand displacement assay for their affinity and in a competition association assay for their binding kinetics on three AR subtypes. One of the compounds with a promising kinetic selectivity profile was also examined in a [(35)S]-GTPγS binding assay for functional activity. We found that XAC and LUF5964 were kinetically more selective for the A1R and A3R, respectively, although they are non-selective in terms of their affinity. In comparison, LUF5967 displayed a strong equilibrium-based selectivity for the A1R over the A2AR, yet its kinetic selectivity thereon was less pronounced. In a GTPγS assay, LUF5964 exhibited insurmountable antagonism on the A3R while having a surmountable effect on the A1R, consistent with its kinetic selectivity profile. This study provides evidence that equilibrium and kinetic selectivity profiling can both be important in the early phases of the drug discovery process. Our proposed combinational strategy could be considered for future medicinal chemistry efforts and aid the design and discovery of different or even better leads for clinical applications.KEYWORDS: Adenosine receptors; Binding kinetics; Binding selectivity profiling; GPCR; Kinetic selectivity profiling; Residence time Show less
Traditionally structure-activity/affinity relationships (SAR) have dominated research in medicinal chemistry. However, structure-kinetics relationships (SKR) can be very informative too. In this... Show moreTraditionally structure-activity/affinity relationships (SAR) have dominated research in medicinal chemistry. However, structure-kinetics relationships (SKR) can be very informative too. In this viewpoint we explore the molecular determinants of binding kinetics and discuss challenges for future binding kinetics studies. A scheme for future kinetics-directed drug design and discovery is also proposed. Show less
Ligand-receptor binding kinetics is increasingly recognized to play a pivotal role in the early phase of drug design and discovery. In this thesis ligand-receptor binding kinetics, particularly... Show moreLigand-receptor binding kinetics is increasingly recognized to play a pivotal role in the early phase of drug design and discovery. In this thesis ligand-receptor binding kinetics, particularly residence time, at the adenosine A1 and A2A receptors was extensively investigated. Several case studies at these two prototypical GPCRs demonstrate that binding kinetics as an emerging paradigm can provide additional information of drug-target interactions at the molecular level. This thesis will contribute to stipulating the importance of kientics-based drug design and discovery in the future. Show less
