Design and Synthesis of 5-HT ia Receptor Ligands A thesis presented to the University of London in partial fulfilment of the requirements for the degree of Doctor of Philosophy by Kit Yee Wong University College London Chemistry Department Christopher Ingold Laboratories 20 Gordon Street London WC1H OAJ April 1997 Attention is drawn to the fact that copyright of this thesis rests with its author. This copy of the thesis has been supplied on condition that anyone who consults it, is understood to recognise that the copyright rests with its author and that no quotation from the thesis and no information derived from it may be published without prior consent of the author. ProQuest Number: 10017478 All rights reserved INFORMATION TO ALL USERS The quality of this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a complete manuscript and there are missing pages, these will be noted. Also, if material had to be removed, a note will indicate the deletion. uest. ProQuest 10017478 Published by ProQuest LLC(2016). Copyright of the Dissertation is held by the Author. All rights reserved. This work is protected against unauthorized copying under Title 17, United States Code. Microform Edition © ProQuest LLC. ProQuest LLC 789 East Eisenhower Parkway P.O. Box 1346 Ann Arbor, Ml 48106-1346 To My Parents Abstract Serotonin (5-hydroxytryptamine, 5-HT) is a biogenic amine known to function through a number of different 5-HT receptor subtypes. The 5 -HT ia receptor subtype is implicated in conditions such as depression and anxiety. There are many examples of selective agonists for this receptor, although selective antagonists have only recently become available. Aryloxyalkylamine p- adrenergic antagonists, such as pindolol and propranolol, are one of the few classes of ligands that exhibit antagonist activity at 5 -HT ia receptors, although they are not selective. This thesis describes the synthesis and biological activity of a series of pindolol and propranolol analogues, with the aim of improving their 5 -H T ia receptor affinities and selectivities. Modifications to the terminal amine substituents of these compounds were made and the effect of 7-substitution in the naphthalene ring was explored. These analogues possess high 5-H T ia versus p-adrenergic receptor selectivities, although few compounds show significant 5-H T ia versus 5-HTi o receptor selectivity. In particular, the 7 -CONH2 and 7-C02M e substituted N,N- ethylbutylaminoethoxynaphthalenes were found to be potent agonists at the 5-HT-|a receptor, having IC50 values of 3.3 and 5.2 nM respectively. Substitution of different groups in the 7-position of the naphthalene ring was found to have a profound effect on the 5 -HT i a and 5-HT i d receptor affinities. Using the Hansch linear-free energy model, attempts were made to determine the relationships between the physicochemical nature of the substituents and their biological activities at these receptors. A multiple least squares linear regression procedure was used to generate several QSAR models. The models were analysed by statistical procedures to determine their validity and significance. Although the procedure did not yield a valid model for the 5 -HT ia receptor affinities, good correlations were obtained for the 5 -HT id receptor affinities. Acknowledgements I wish to thank my supervisor, Professor Robin Ganellin for giving me the opportunity to work on this project. I am extremely grateful for his constant support and encouragement throughout the course of this work. Thanks to Mr Wasyl Tertiuk and all the members of the group, past and present for their friendship and helpful discussions. To all my friends in the Chemistry Department, past and present, who have made the whole experience so enjoyable. Special thanks are due to Ashley (manager of the mini-cluster), Nicola, Tiz, Zena, Najeeb, Simon, Rodney, Stefan and Alvin for all the good times over the last three years. I am indebted to Dr Margaret Beer and her team at Merck Sharp and Dohme (Harlow) for carrying out the 5 -HT ia and 5-HT-| d receptor binding assays, and for organising the p-adrenergic receptor assays. I am extremely grateful for her endless patience, friendship and encouraging e-mails during the course of this study. I would like to express my gratitude to all the technical staff - Steve Corker, John Hill, Jill Maxwell, Don Shipp (Birkbeck), and Alan Stones for their efficient services. Thanks also to Chris Cooksey for his assistance with the NMR experiments, to Dr Derek Banthorpe for his entertaining and thought-provoking discussions and to Dr Mike Abraham for his helpful suggestions. Thanks to Peter Leighton for looking after (!) my overnight experiments. I would also like to thank Dr Clare Stanford for giving me the opportunity to undertake some pharmacological binding assays at UCL Pharmacology Department, and for her invaluable discussions. Thanks to Ming for the loan of his computer in the first two years and for buying and installing the memory into my own computer. Thanks to Kit Junior for all the letters and phone calls during the writing-up period. Finally I am indebted to my mum and dad for their love, encouragement and support throughout my life, and it is to them that I dedicate this thesis. Contents Abstract 3 Acknowledgements 4 Contents 5 Glossary 9 1. Introduction 10 1.1 Discovery of Serotonin 10 1.2 Distribution of 5-HT 10 1.3 Biosynthesis and Catabolism of 5-HT 11 1.4 5-HT Receptors 11 1.4.1 Historical 11 1.4.2 Cloning of 5-HT Receptors 13 1.4.3 Structure of 5-HT Receptors 13 1.5 Present Classification of 5-HT Receptors 14 1.5.1 5-HT-i Receptors 15 1.5.2 5 -HT2 Receptors 21 1.5.3 5 -HT3 Receptor 24 1.5.4 5 -HT4 Receptor 25 1.5.5 5-hts Receptors 27 1.5.6 5-hte Receptor 27 1.5.7 5-hty Receptor 28 1.6 The 5 -HT1A Receptor 28 1.6.1 Molecular Biology 28 1.6.2 5 -HT1A Receptor Distribution and Function 29 1.6.3 Structure of the 5 -HT ia Receptor 29 1.6.4 5-HTiA Receptor Ligands 30 1.7 Aryloxyalkylamines as 5-HTi a Receptor Ligands 35 1.8 Structure-Activity Analysis 35 1.8.1 Benzyloxypropanoiamines 36 1.8.2 Pindolol Derivatives 38 1.8.3 Propranolol Derivatives 42 2. Selection of Compounds 47 2.1 Project Aims 47 2.2 Pindolol Series 47 2.3 Propranolol Series 49 3. Synthesis of Aryloxyalkylamines 53 3.1 Synthetic Strategy for Unsubstituted Indolyloxyalkylamines 53 3.1.1 Synthesis of 4-Hydroxyindole 54 3.1.2 Synthesis of Side Chain Intermediates 56 3.1.3 Synthesis of Target Pindolol Derivatives 57 3.2 Attempted Synthesis of 2-Substituted Pindolol Analogues 58 3.3 Synthesis of Propranolol Analogues 61 3.4 Synthetic Strategy for Type A Compounds 63 3.4.1 Synthesis of 4-Acetylphenylbutanoic acid 65 3.4.2 Oxidation of 4-Acetylphenylbutanoic acid 67 3.4.3 Cyclisation of 4-Carboxyphenylbutanoic acid 69 3.4.4 Dehydrogenation of 7-Carboxymethyl-1-tetralone 71 3.4.5 Formation of Aminoalkyl Side Chain Propranolol Analogues 75 3.4.6 Derivatisation of the Ester Function of UCL 1693 79 3.4.7 Formation of 7-Carboxymethyl-Propranolol 83 3.5 Synthetic Strategy for Type B Compounds 86 3.5.1 Friedel-Crafts Acylation of Substituted Benzenes 88 3.5.2 Clemmensen Reduction of Keto Acids 89 3.5.3 Synthesis of Substituted Tetralones (X = Me, 'Pr, OMe, F) 90 3.5.4 Synthesis of 7-Chloro-1-tetralone 90 3.5.5 Formation of 7-Substituted-1-naphthols (X = Me, 'Pr, OMe, F) 91 3.5.6 Synthesis of 7-Chloro-1-naphthol 93 3.5.7 Formation of Target 1,7-Naphthalene Compounds 97 4. Pharmacological Methods 100 4.1 Receptor-Ligand Interactions 100 4.2 Radioligand Binding Studies 102 4.3 Functional Studies 103 4.4 Pharmacological Protocols 104 5. Pharmacological Results and Discussion 109 5.1 Indolyloxyalkylamine Analogues 109 5.1.1 5-HT 1A Receptor Binding and Functional Studies 109 5.1.2 5-HT 1A versus 5-HTi q Receptor Selectivity 111 5.1.3 p-Adrenergic Receptor Binding 112 5.1.4 Summary 113 5.2 Naphthyloxyalkylamine Analogues 113 5.2.1 5-HT ia Receptor Binding and Functional Studies 113 5.2.2 5-HT 1A versus 5-HT-| d Receptor Selectivity 119 5.2.3 p-Adrenergic Receptor Binding 122 5.2.4 Summary 126 5.2.5 Functional Activity of Propranolol Derivatives 127 5.3 QSAR Studies 129 5.3.1 Introduction 129 5.3.2 Hansch Linear Free-Energy Model 130 5.3.3 QSAR Procedure 133 5.3.4 Limitations of QSAR Studies 136 5.3.5 Regression Procedure 137 5.3.6 Experimental 138 5.3.7 Results 139 5.4 Conclusion and Future Perspectives 144 6. Experimental 147 References 216 Appendix - Selected NMR Spectra 226 Glossary agonist a drug that can interact with receptors and initiate a drug response antagonist a drug that opposes the effect of another by physiological or chemical action, or by a competitive mechanism for the same receptor sites a str (IR) asymmetric stretch br s broad singlet CDCI3 deuterated chloroform CD3OD deuterated methanol d doublet dd doublet of doublets ddd doublet of doublets of doublets dg-DMSO deuterated dimethylsulfoxide, (CD 3 )2 SO dt doublet of triplets HPLC high performance liquid chromatography IC50 median inhibitory constant (concentration at which an antagonist exerts its half maximal effect) IR infrared Ki inhibitory constant calculated from experimentally obtained IC 50 values using the Cheng-Prusoff equation, Kj = IC 5 o/(1 + L/Kq) (L = final concentration of radioligand; Kq = radioligand equilibrium dissociation constant) m(IR) medium absorption m (NMR) multiplet MS mass spectrum NMR nuclear magnetic resonance NOE nuclear Overhauser enhancement q quartet QSAR quantitative structure-activity relationship quint quintet s(IR) strong absorption s (NMR) singlet sept septet s str (IR) symmetric stretch t triplet ‘t’ pseudotriplet td triplet of doublets w(IR) weak absorption Introduction 1.