THEORETICAL STUDIES OF STEROID HORMONES AND RELATED COMPOUNDS Michael Hugh Charlton A Thesis Submitted for the Degree of PhD at the University of St Andrews 1992 Full metadata for this item is available in St Andrews Research Repository at: http://research-repository.st-andrews.ac.uk/ Please use this identifier to cite or link to this item: http://hdl.handle.net/10023/14449 This item is protected by original copyright Theoretical Studies of Steroid Hormones and Related Compounds. A Thesis Presented for the Degree of Doctor of Philosophy in the Faculty of Science of the University of St. Andrews. by Michael Hugh Charlton, B.Sc. (Hons). St. Leonard's College September 1991 ProQuest Number: 10167149 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 10167149 Published by ProQuest LLC(2017). 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 Abstract. A theoretical study of steroidal inhibitors of the enzymes Glucose-6-Phosphate Dehydrogenase and Aromatase is presented. Both enzyme systems are of interest in the study of cancer, the latter being the final step in the biosynthesis of oestrogens which are involved in certain types of breast cancer. Two levels of theory are employed in the study, namely, Ab Initio and Semi Empirical methods. Structures and charges have been calculated using the MOPAC and GAUSSIAN programs and these have been used to model the efficacy of various inhibitors. The major tool in comparing these steroids has been the molecular electrostatic potential ( MEP ). Maps of the MEP and an analysis of the similarity between the MEP s of different molecules have led both to a method of assessing the activities of steroids as enzyme inhibitors and requirements for the electronic structure of the steroid binding sites within these enzymes. A molecular graphics display program has been developed to facilitate this work. It has been designed to make full use of the facilities available. The quality of the resulting display has improved greatly on what was previously available and has been of value in studies of large molecular systems. The program is written in VAX FORTRAN and uses the Graphics Kernel System (GKS ) to produce graphical output and should be reasonably easy to transfer to other systems. Finally, to determine whether PM3 really is a significant advance on AMI, a comparison of the two semi empirical methods is presented. The calculated properties of steroid hormones are compared to those of both Ab Initio calculations and experimental determinations, allowing the quality of the semi empirical predictions to be assessed. Declarations, I, Michael Hugh Charlton, hereby certify that this thesis has been composed by myself, that it is a record of my own work, and that it has not been accepted in partial or complete fulfilment of any other degree or professional qualification. Michael Charlton. I was admitted to the Faculty of Science at the University of St. Andrews under Ordinance General No. 12 on 1st October 1988 and as a candidate for the degree of PhD on the 1st October 1989. sep-^f Michael Charlton. I hereby certify that the candidate has fulfilled the conditions of the Resolution and Regulations appropriate to the degree of PhD. 3o /^/ Colin Thomson. iii Copyrightt In submitting this thesis to the University of St. Andrews, I understand that I am giving permission for it to be made available for use in accordance with the regulations of the University Library for the time being in force, subject to any copyright vested in the work not being affected thereby. I also understand that the title and abstract will be published, and that a copy of the work may be made and supplied to any bona fide library or research worker. Michael Charlton. AcknQwlcdgs.msjo.ts.! I would like to thank Colin Thomson for the opportunity to carry out this project and for his advice, the S.E.R.C. for financial support and the N.F.C.R., A.I.C.R. and Digital for their support of the group. I have also had useful discussions with other members of staff, and would like to thank David Calvert, Chris Glidewell, Gordon Harris, Kenneth Harris, and Gordon Woolley. Thanks are also due to John Walton, whose question sparked off the idea for the Geometry Optimisation study. Other members of the department deserving mention are Andrew Leech for a helpful discussion, and Jim Bews for his skill in keeping the Macintoshes under control. The past and present members of the group have been invaluable, and I would like to mention Paola, Piera, Derek, Donatella and John, but particularly my great friends Andrew and Julie. Audrey and Catherine deserve a mention, just for listening to my ideas and pretending that I made sense. The encouragement and financial assistance of my parents has kept me going both mentally and physically, and I should like to thank them both. Finally, the greatest thanks must go to Sarah, whose help with typing, references, spelling and grammar have been invaluable. She has had to become a minor expert in Theoretical Chemistry to understand me, and a disaster relief worker to clear up the trail of destruction following in the wake of this thesis. iv Contents Chapter One Introduction. 1 A The Occurrence of Cancer. 2 B Cell Growth and Cancer. 3 C Carcinogenesis. 4 D Steroid Hormones and Cancer. 6 E The Use of Theoretical Methods and Computational Chemistry Software. 10 F Molecular Graphics. 12 G Equipment, Program Details and Methods. 13 Chapter Two Theoretical Methods and Computational Chemistry Software. 16 A Classical Mechanics. 17 B The Schrodinger Wave Equation. 17 C Properties of the Wave Function. 20 D Hartree - Fock Molecular Orbital Theory. 21 E The Basis Set Expansions: The Lineal’ Combination of Atomic Orbitals Method. 25 F Types of Basis Set. 28 G Direct SCF. 32 H Limitations of the Hartree - Fock Method. 32 I Semi Empirical Methods. 34 J Molecular Geometries. 38 K Properties Calculated from the Hartree - Fock SCF Wavefunction. 43 L Software. 49 M Electrostatic Potential Software. 51 N Molecular Graphics. 53 O Other Programs Written as Part of this Research. 53 V Chapter Three Design and Implementation of a Molecular Graphics Display Program. 56 A Introduction. 57 B Details of the Program. 58 C Data Type Conversion and File Manipulation. 59 D Graphical Display. 65 E Lists. 69 F Chopping Out Sections of Molecules. 71 G Geometry Comparison. 72 H Other Functions of the Graphics Program. 75 I Conclusion. 75 J Description of Photographs. 76 Chapter Four A Discussion of Electrostatic Potential and Geometry Optimisation. 84 A Introduction. 85 B Rationale Behind the use of MEP in Biological Studies. 85 C Uses of the MEP. 86 D Calculation of the Potential Maps. 88 E Dependency of the MEP upon the Accuracy of the Wavefunction. 89 F Calculation of Potential Fitted Charges. 90 G Study of Geometry Optimisation Using MOPAC. 97 Chapter Five A Theoretical Investigation of DHEA and Other G6PDH Inhibitors. 121 A Introduction. 122 B DHEA as an Inhibitor of G6PDH. 123 C Quantum Mechanical Study of Steroidal Inhibition of G6PDH. 125 D Rationalisation of the Activities of the Steroids as Inhibitors of G6PDH. 142 E ASP Study of the Androstane Series. 157 F Other Similarity Work on the Androstane Series. 164 G Final Conclusions on the Androstane Electrostatic Potential Studies. 171 H Electrostatic Potential Work on the Pregnane Series. 172 vi I ASP Work for the Pregnane Series. 175 J Comment on the Accuracy of the Experimental Data. 179 K Summary of the Work on Steroidal Inhibitors of G6PDH. 180 Chapter Six Studies of Inhibition of Oestrogen Biosynthesis. 195 A Introduction. 196 B Aromatase Inhibitors Studied in this Chapter. 200 C Semi Empirical Calculations. 203 D Structure of the 17 P-side Chain. 204 E Conformation of the Steroid A-ring. 212 F Rationalisation of the Competitive Inhibition Using Theoretical Techniques. 215 G Analysis of the Electrostatic Potential Maps for the Aromatase Inhibitors. 225 H Theoretical Model of the Enzyme Inactivation. 227 I Conclusions. 236 Chapter Seven A Comparison of AMI and PM3 with Ab Initio and Experimental Results. 240 A Introduction. 241 B Comparison of AMI and PM3 by evaluation of the Ab Initio energy for the Optimised Geometry. 245 C Comparison of Semi Empirical and Ab Initio Structures. 253 D Comparison of Experimental and Calculated Dipole Moments. 264 E Quality of Calculated Ionisation Potentials. 271 F Assessment of Calculated Atomic Charges and Chemical Shifts. 275 G Comparison of Calculated Structures and X-Ray Geometries. 289 H Final Comments. 296 Chapter Eight Conclusion. 298 References. 303 J Vll Appendices. Appendix One 313 Appendix Two 314 Appendix Three 325 Chapter One Introduction. 2 The topic of this thesis is a study, using the methods of theoretical chemistry, of some of the processes involved in cancer. This chapter gives a short outline of the disease, and briefly indicates how steroid hormones (a field of particular interest in this work) are involved. It also describes how computational chemistry can be used to study biological systems, and gives an outline of how it will be applied to the study of the mechanism of action of steroids in the chapters of this thesis.