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Medicinal chemistry techniques to gain selectivity at G protein-coupled receptors targeting the CNS: M₄ muscarinic acetylcholine receptor positive allosteric modulators, dopamine D₂ receptor biased ligands, M₁/D₂ merged ligands and D₂ fluorescently labelled ligands

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posted on 01.03.2017, 01:05 authored by Szabo, Monika
The study of G protein-coupled receptors (GPCRs) and their role in central nervous system (CNS) disorders and/or disease states is integral to drug discovery. For schizophrenia, a favourable polypharmacology profile is useful to effectively treat all symptom domains of the disorder. However, GPCRs can be difficult to gain subtype selectivity in order to avoid binding to receptors in the same family and to various off-target receptors. This thesis explores multiple medicinal chemistry approaches to achieve subtype selectivity or pathway specificity, and tools to screen for new compounds and/or scaffolds. Chapter 2 investigates the use of positive allosteric modulators (PAMs) for the M₄ muscarinic acetylcholine receptor (mAChR). A comprehensive structure activity relationship (SAR) study around the PAM, LY2033298, was conducted, which investigates different linkage points, halogen replacements and different substitution combinations on the thienopyridine scaffold. The compounds are evaluated via the use of an operational model of allosterism to determine values of functional affinity (Kʙ), cooperativity (αβ) and intrinsic agonism (τʙ) for all compounds. These parameters allowed the elucidation of the molecular determinants of allostery that may be important for certain functional changes. Chapter 3 explores the concept of biased agonism as an approach to gain pathway-specific selectivity at the dopamine D₂ receptor (D₂R). The determinants of efficacy, affinity and bias for three privileged structures for the D₂R were explored by focusing on changes to linker length and incorporation of a heterocyclic unit. By quantifying bias at two signalling pathways (cAMP and pERK1/2), distinct bias patterns were observed associated with the substitution of certain phenylpiperazine structures. Subtle structural changes to the heterocycle resulted in significant effects on bias that over-ruled the effect of the phenylpiperazine substitution pattern. As such the series of compounds may represent useful tools to gain further insight into the contribution of biased agonism to antipsychotic efficacy. Chapter 4 uses a “designed multiple ligand” approach, to rationally synthesise compounds with a favourable polypharmacology profile. Specifically, D₂/5-HT₂ᴀ activity has been implicated as useful for antipsychotic efficacy and activity at M1 mAChRs is highlighted as an important target for the cognitive deficits in key CNS disorders. The hybridisation process makes use of substituted piperazine or pipridine privileged structures with D₂ or D₂/5-HT₂A activity in combination with the putative M1 mAChR allosteric agonist LuAE51090. The focused library of compounds was profiled in both radioligand binding assays in addition to functional assays at the M₁ mAChR, D₂R and 5-HT2AR. From this we identified a compound which retained activity at all three receptors and therefore represents an ideal starting point for further optimisation. Chapter 5 explores the use of fluorescently labelled ligands as pharmacological tools for the D₂R of which there is a notable lack thereof in the literature. A series of ligands based on two clinical compounds, clozapine and ropinirole, in addition to a negative allosteric modulator, SB269652, and its high-affinity variant 2-MPP-SB269652, were selected to create a series of fluorescently labelled ligands. The ligands were tethered to a spectrum of fluorophores (BODIPY 630/650-X, Cy5, Cy3, FITC and LRB) as each of the fluorophores varied in spectral properties and chemical scaffolds. Three of the four series maintained functionality at the D₂R, with the derivatives of 2-MPP-SB269652 maintaining high binding affinities. Three novel fluorescent ligands demonstrated rather specific cell membrane binding and very weak non-specific binding. These fluorescent ligands represent useful tools to be adapted towards multiple applications for the D2R and a starting point for further optimisation. Finally, chapter 6 provides a brief summary of the outcomes from this thesis, as well as directions for future prospects.


Campus location


Principal supervisor

Ben Capuano

Year of Award


Department, School or Centre

Medicinal Chemistry and Drug Discovery Biology


Doctor of Philosophy

Degree Type



Faculty of Pharmacy and Pharmaceutical Sciences