Adenosine is an endogenous ligand which exerts its action by activating adenosine receptors (ARs), while its circulating levels are controlled via a variety of mechanism and proteins, amongst... Show moreAdenosine is an endogenous ligand which exerts its action by activating adenosine receptors (ARs), while its circulating levels are controlled via a variety of mechanism and proteins, amongst others the equilibrative nucleoside transporters (ENTs). Distortion of the adenosinergic tone is implicated in a multitude of pathophysiological conditions, thus numerous drug discovery efforts have been made to develop drugs targeting ARs and ENTs. Yet, there is only a limited number of drugs targeting these proteins on the market. Therefore, there is a pivotal need to develop novel concepts that allow us to increase our understanding of the mechanism of action at a molecular level, as well as physiologically relevant assays to evaluate drug candidates in early stages of drug discovery. Hence, this thesis focuses on exploring the concept of binding kinetics for two adenosinergic targets, i.e. the A2BAR and ENT1 (SCL29A1), as well as to develop novel kinetic binding and label-free functional assays. Show less
This thesis provides novel insights in the molecular mechanism of action of antagonists for the chemokine receptor CCR2. CCR2 belongs to the protein family of G protein-coupled receptors (GPCRs).... Show moreThis thesis provides novel insights in the molecular mechanism of action of antagonists for the chemokine receptor CCR2. CCR2 belongs to the protein family of G protein-coupled receptors (GPCRs). It is involved in several inflammatory diseases and therefore many small molecule antagonists targeting this receptor have been developed over the years. Unfortunately all clinical candidates tested so far appeared to lack efficacy in man, which stresses the need for a better understanding of their mechanism of action. This thesis revealed three separate binding pockets throughout the transmembrane receptor domain via which CCR2 can be pharmacologically modulated. Different routes towards insurmountable antagonism of CCR2 were described, either via noncompetitive or via long residence time antagonists. These results may allow a more rational design of future antagonists, and are equally important to understand the outcomes of studies with existing CCR2 antagonists. In concert with the currently expanding insight in the structure and signalling capacities of GPCRs, the data presented in this thesis allow to better fine-tune the pharmacological modulation of the chemokine receptor CCR2, and GPCRs in general Show less
