This PhD thesis focuses on the study of the α4β2 nicotinic acetylcholine receptors (nAChRs). This dissertation is divided into two parts. The first part is centred on the design, synthesis, and biological evaluation of compounds selective for the orthosteric and unorthodox binding sites of the α4β2 nAChRs. In the second part, attention is paid to the study of a photoactivatable α4β2 nAChRs full agonist. Part 1 The first goal of Part 1 was the mutational studies of Ser108 residue of the hydrophilic pocket of the β2(-) side of the α4β2 orthosteric binding site with the purpose of studying the importance of the hydroxyl group of serine residue and the steric encumbrance in the pocket. For this reason, binding experiments were performed on the heterologously expressed human α4β2 receptor and its three Ser108àLeu, Ser108àPhe and Ser108àAla mutants using the most promising partial agonists synthesized previously. The second goal was the synthesis and pharmacological evaluation of the naked and hydroxy/methoxy decorated benzofuran-based compound in order to study the implications of the flexibility reduction, abolishment of cycle stereogenic and of the one of the two oxygen in comparison with previously tested benzodioxane derivatives. Subsequently, the most promising benzodioxane compounds and the obtained benzofuran derivatives were evaluated for their affinity and activity on the two distinct α4β2 nAChR isoforms: (α4)2(β2)3 and (α4)3(β2)2. The third goal was the study of the in vivo effects of prolinol aryl ethers and pyrrolidinyl benzodioxanes full and partial agonists. These compounds were tested for their effects on zebrafish behaviour. In particular, full agonists were tested for their ability to improve spatial memory and attention, and partial agonist for its ability to block the rewarding effects of nicotine and therefore its potential use for smoking cessation. The fourth goal was the design, synthesis and pharmacological evaluation of selective agonists for the α4α4 unorthodox binding site of the α4β2 nAChR based on the structure of NS9283, an α4α4selective compound, described in the literature. The main aim was to identify new hit V compounds potentially selective at the (α4)3(β2)2 receptor isoform with a better pharmacological profile. Part 2 Regarding the second part of this thesis, it focuses on the synthesis of a photoactivatable α4β2 nAChR full agonist with the purpose to spatiotemporally control its release with light at the specific site of action. To follow this strategy, 8-bromo-7-hydroxy-2-methylquinoline (BHQ) and 8-cyano-7-hydroxy-2-methylquinoline (CyHQ) protecting groups were synthesized and used to cage a previously synthesized full agonist. Subsequently, its release after the irradiation with the UV light at 365 nm was followed using HPLC in order to obtain a full photo- chemical characterization of the caged compounds to get necessary information for in vitro/in vivo experiments.

ALPHA4BETA2 NEURONAL NICOTINIC ACETYLCHOLINE RECEPTOR LIGANDS: FROM SUBTYPE SELECTIVE TO STOICHIOMETRIC ISOPHORM SELECTIVE PARTIAL AGONISM / R. Appiani ; tutor: C. Bolchi ; co-supervisor: M. Pallavicini ; coordinatore: G. Aldini. - : . Dipartimento di Scienze Farmaceutiche, 2022. ((35. ciclo, Anno Accademico 2022.

ALPHA4BETA2 NEURONAL NICOTINIC ACETYLCHOLINE RECEPTOR LIGANDS: FROM SUBTYPE SELECTIVE TO STOICHIOMETRIC ISOPHORM SELECTIVE PARTIAL AGONISM.

R. Appiani
2023

Abstract

This PhD thesis focuses on the study of the α4β2 nicotinic acetylcholine receptors (nAChRs). This dissertation is divided into two parts. The first part is centred on the design, synthesis, and biological evaluation of compounds selective for the orthosteric and unorthodox binding sites of the α4β2 nAChRs. In the second part, attention is paid to the study of a photoactivatable α4β2 nAChRs full agonist. Part 1 The first goal of Part 1 was the mutational studies of Ser108 residue of the hydrophilic pocket of the β2(-) side of the α4β2 orthosteric binding site with the purpose of studying the importance of the hydroxyl group of serine residue and the steric encumbrance in the pocket. For this reason, binding experiments were performed on the heterologously expressed human α4β2 receptor and its three Ser108àLeu, Ser108àPhe and Ser108àAla mutants using the most promising partial agonists synthesized previously. The second goal was the synthesis and pharmacological evaluation of the naked and hydroxy/methoxy decorated benzofuran-based compound in order to study the implications of the flexibility reduction, abolishment of cycle stereogenic and of the one of the two oxygen in comparison with previously tested benzodioxane derivatives. Subsequently, the most promising benzodioxane compounds and the obtained benzofuran derivatives were evaluated for their affinity and activity on the two distinct α4β2 nAChR isoforms: (α4)2(β2)3 and (α4)3(β2)2. The third goal was the study of the in vivo effects of prolinol aryl ethers and pyrrolidinyl benzodioxanes full and partial agonists. These compounds were tested for their effects on zebrafish behaviour. In particular, full agonists were tested for their ability to improve spatial memory and attention, and partial agonist for its ability to block the rewarding effects of nicotine and therefore its potential use for smoking cessation. The fourth goal was the design, synthesis and pharmacological evaluation of selective agonists for the α4α4 unorthodox binding site of the α4β2 nAChR based on the structure of NS9283, an α4α4selective compound, described in the literature. The main aim was to identify new hit V compounds potentially selective at the (α4)3(β2)2 receptor isoform with a better pharmacological profile. Part 2 Regarding the second part of this thesis, it focuses on the synthesis of a photoactivatable α4β2 nAChR full agonist with the purpose to spatiotemporally control its release with light at the specific site of action. To follow this strategy, 8-bromo-7-hydroxy-2-methylquinoline (BHQ) and 8-cyano-7-hydroxy-2-methylquinoline (CyHQ) protecting groups were synthesized and used to cage a previously synthesized full agonist. Subsequently, its release after the irradiation with the UV light at 365 nm was followed using HPLC in order to obtain a full photo- chemical characterization of the caged compounds to get necessary information for in vitro/in vivo experiments.
BOLCHI, CRISTIANO
ALDINI, GIANCARLO
Settore CHIM/08 - Chimica Farmaceutica
ALPHA4BETA2 NEURONAL NICOTINIC ACETYLCHOLINE RECEPTOR LIGANDS: FROM SUBTYPE SELECTIVE TO STOICHIOMETRIC ISOPHORM SELECTIVE PARTIAL AGONISM / R. Appiani ; tutor: C. Bolchi ; co-supervisor: M. Pallavicini ; coordinatore: G. Aldini. - : . Dipartimento di Scienze Farmaceutiche, 2022. ((35. ciclo, Anno Accademico 2022.
Doctoral Thesis
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/950273
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