Antibacterial Agents: 1,4-Disubstituted 1,2,3-Triazole Analogs of the Oxazolidinone A dissertation presented to the faculty of the College of Arts and Sciences of Ohio University In partial fulfillment of the requirements for the degree Doctor of Philosophy George Acquaah-Harrison June 2010 © 2010 George Acquaah-Harrison. All Rights Reserved. 2 This dissertation titled Antibacterial Agents: 1,4-Disubstituted 1,2,3-Triazole Analogs of the Oxazolidinone by GEORGE ACQUAAH-HARRISON has been approved for the Department of Chemistry and Biochemistry and the College of Arts and Sciences by Stephen C. Bergmeier Professor of Chemistry and Biochemistry Benjamin M. Ogles Dean, College of Arts and Sciences 3 ABSTRACT ACQUAAH-HARRISON, GEORGE, Ph.D., June 2010, Chemistry and Biochemistry Antibacterial Agents: 1,4-Disubstituted 1,2,3-Triazole Analogs of the Oxazolidinone ) (235 pp.) Director of Dissertation: Stephen C. Bergmeier The rational design, development and synthesis of structurally diverse small molecule that target RNA is immensely important in antibacterial therapy. Utilizing rational design approach to drug discovery, two lead 4,5-disubstituted 2-oxazolidinone compounds that bind to the highly conserved region of bacterial RNA with high affinity and specificity had been previously identified. This biological target called the T box antiterminator system regulates the expression of many genes including aminoacyl synthethase genes and is found predominantly in Gram-positive pathogens. But, owing to the moderate solubilities of these leads, the focus was directed to improving the solubility challenges without compromising biological activity. To address the solubility challenges with the intent of enhancing or maintaining biological activity, a library of one hundred eight 1,4-disubstituted 1,2,3-triazole compounds that encompasses the diversity elements of the oxazolidinones were developed. This library, which entailed the bioisosteric replacement of the oxazolidinone scaffold was afforded in high yield and purity by employing the regioselective copper(I)-catalyzed azide/alkyne cycloaddition reaction. Three lead compounds, GHB-7, GHB-9 and GHB-16 with enhanced biological activity were identified that rendered them important candidates for structural activity 4 relationship studies (SAR). Besides the SAR studies, few 1,5-regioisomers were prepared to investigate the effect of regioselectivity on biological activity. By embarking on empirical observations, thirty-two structurally relevant analogs were prepared for the SAR and other structural elaboration studies. While biological evaluation is currently ongoing, the preliminary data of the analogs, GHB-144, GHB- 151, GHB-153, GHB-156 and GHB-157 coupled with the enhanced biological activity of GHB-7 relative to the lead oxazolidinone compounds re-inforce the plausibility of finding new 1,4-substituted 1,2,3-triazole compounds with improved antibacterial activity. Approved: _____________________________________________________________ Stephen C. Bergmeier Professor of Chemistry and Biochemistry 5 ACKNOWLEDGMENTS I would like to express my sincere gratitude to my academic advisor Dr. Stephen C. Bergmeier for his mentorship, guidance, insightful discussions and encouragements during my study. I would also like to thank the members of my dissertation committee Dr. Mark C. McMills, Dr. Glen Jackson and Dr. Xiaozhuo Chen for accepting my invitation to serve on my committee. Special thanks go to the faculty of the Department of Chemistry and Biochemistry, especially Dr Jared Butcher, Dr. Klaus Himmeldirk and Dr. Kumar Pichumani as well as the administrative staff of the department of Chemistry and biochemistry for their suggestions and support during my study. I am thankful to the members of Dr. Bergmeier’s research group both past (Dr. Pulipaka Aravinda, Dr. Junfeng Huang, Dr. Ahbigit Nayek, Nova Emerald and Dr. Iwona Maciagiewicz) and present (Dr. Crina M. Orac, Weihe Zhang, Fang Fang, Susann Krake, Gregg Wells and Dennis Roberts) for their suggestions and constructive criticisms as well as promoting a conducive laboratory atmosphere. I am also thankful to Shu Zhou in Dr. Hines laboratory for testing all triazole analogs. I am greatly indebted to my family: Harris Osafo-Acquaah, Mary F. Kusi- Appouh, Franklin Effah, Victoria Dankwa & family, Gina Peters & family, Kwabena Appiah Peprah, Mary Appiah Peprah and the rest of Peprah family as well as Dr. Isaac Blankson and Joseph Blankson for their incessant support, advice and prayers during my doctoral studies. 6 Finally to my wife, Vivian Acquaah-Harrison and three wonderful daughters, Ama Afrakomah Acquaah-Harrison, Nana-Afia Acquaah-Harrison and Nana-Akua Acquaah-Harrison, thank you for recognizing the importance of education. Your love, Charisma, encouragement, patience and motivation is what energized me through this study. 7 Dedicated to my wife Vivian A. Acquaah-Harrison and Three wonderful daughters Ama Afrakomah Acquaah-Harrison, Nana-Afia Antwiwaa Acquaah-Harrison and Nana-Akua Osafoaa Acquaah-Harrisonson 8 TABLE OF CONTENTS Page Abstract................................................................................................................................3 Acknowledgments................................................................................................................5 Dedication ...........................................................................................................................7 List of Tables .......................................................................................................................9 List of Figures....................................................................................................................10 List of Schemes..................................................................................................................12 Chapter 1.Introduction......................................................................................................15 1.1 Statement of purpose ........................................................................................... 15 1.2 Introduction.......................................................................................................... 15 Chapter 2.Background......................................................................................................23 2.1 History of antibiotics ........................................................................................... 23 2.1.1 Natural antibiotics......................................................................................... 24 2.1.2 Synthetic antibiotics...................................................................................... 32 2.1.3 Antibiotics: problems and challenges ........................................................... 35 2.2 Antibacterial drug discovery................................................................................ 37 2.3 RNA domain as a novel target for antimicrobial therapy.................................... 38 2.3.1 T box antiterminator RNA System ............................................................... 40 2.4 Identification of lead compound.......................................................................... 42 2.4.1 Lead compound modification/optimization.................................................. 44 2.5 Oxazolidinone compounds .................................................................................. 46 2.5.1 Oxazolidinone library generation ................................................................. 47 2.6 Significance of research....................................................................................... 51 Chapter 3.Synthesis and Preliminary Results of 1,4-Disubstituted 1,2,3-Triazole Library52 3.1 Introduction.......................................................................................................... 52 3.2 Retrosynthetic analysis of the triazole library ..................................................... 56 3.3 Synthesis of azide components............................................................................ 57 3.4 Synthesis of alkyne components.......................................................................... 65 3.5 Determination of reaction condition for 1,2,3-triazole library synthesis............. 68 9 3.5.1 Model studies ................................................................................................ 71 3.5.2 Determination of workup conditions ............................................................ 72 3.5.3 Generation of 1,4-disubstituted 1,2,3-triazole library................................... 74 3.6 Biological evaluation of 1,4-disubstituted 1,2,3-triazole compounds ................. 81 3.6.1 Fluorescence resonance energy transfer assay.............................................. 81 3.6.2 Antibacterial Assay....................................................................................... 87 3.7 Synthesis of 1,5-disubstituted 1,2,3-triazoles analogs......................................... 89 3.7.1 Biological evaluation of 1,5-analogs ............................................................ 94 3.8 Conclusion ........................................................................................................... 95 Chapter 4.Structure Activity Studies and Structural Elaborations of the 1,2,3-Triazole Ring 97 4.1 Introduction.......................................................................................................... 97 4.2 Plan