mg» ., N. It ; u:- nan—"F...“ . . V, ._ x v? :3?! u 1 .“f‘ v "A I 1.! 1.". \37 ‘35! z: .b 3: .LM I am... a .- a v .55. J33... Lukas: i. rn... r593»: . E tiff. .. 33...?! ......va. ‘of vi. ; .. .sex 91...“... .353... {1‘ t3...- . .. \.. cl“ '1'? .mr. I 41L“... int... . a... ‘31 n 3. pi “G‘Hfi LIBRARY Michigan State University This is to certify that the dissertation entitled SYNTHESES OF a,a-DlSUBSTITUTED-a-AMINO ACIDS AND IMIDAZOLINES DERIVED FROM OXAZOL-5(4H)-ONES presented by Jason Scott Fisk has been accepted towards fulfillment of the requirements for the Doctoral degree in Chemistry K Jajorfifiofes‘sor’s‘ Signature $1 26 zmor‘ Date MSU is an Afiinnatr‘ve Action/Equal Opportunity Employer ..-.-.--o.-.----_.-.‘.-.-.-u-.-.—n-u-.-n-.---n--u—o--.--._.---c----—-_.-.—.--<-.------.--.--.-.-.-.-.--.. PLACE IN RETURN BOX to remove this checkout from your record. To AVOID FINES return on or before date due. MAY BE RECALLED with earlier due date if requested. DATE DUE DATE DUE DATE DUE 5/08 KrlProleocapres/CIRCIDateDuoindd SYNTHESES OF a,a-DISUBSTITUTED-a-AMINO ACIDS AND IMIDAZOLINES DERIVED FROM OXAZOL-5(4H)-ONES By Jason Scott Fisk A DISSERTATION Submitted to Michigan State University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Chemistry 2009 ABSTRACT SYNTHESES OF a,a—DISUBSTITUTED-a-AMINO ACIDS AND IMIDAZOLINES DERIVED FROM OXAZOL—5(4H)-ONES By Jason Scott Fisk The research presented in this dissertation focuses primarily on the development of new synthetic organic methodology utilizing oxazoI-5(4H)-ones for the purpose of producing biologically useful heterocyclic compounds. The initial chapter describes and illustrates the general reactivity patterns of oxazol-5(4H)-ones using examples from the current literature. The remainder of the chapters describes studies pertaining to the use of oxazoI-5(4H)-ones for synthesizing two different classes of molecules: a,a—disubstituted-a—amino acids and 2- imidazolines. The first study presented in this dissertation involves a brief structure activity relationship (SAR) investigation of a class of 2-imidazolines found to inhibit NF-KB mediated gene transcription. Previous studies within our research group indicated that oxazol-5(4H)-ones undergo Lewis acid promoted [3+2] cycloadditions with imines to diastereoselectively afford highly substituted 2- imidazolines. Select members of this class of 2-imidazolines were previously found to be relatively potent inhibitors of NF-KB mediated gene transcription. In a collaborative effort to optimize this class of compounds for their ability to inhibit NF-KB mediated gene transcription, our research group conducted a SAR study. Small libraries of 2—imidazolines were synthesized from oxazol-5(4H)-ones and subsequently evaluated for their ability to inhibit NF-KB mediated gene transcription in both human cervical epithelial (HeLa) cells and human whole blood. Included in this dissertation are the results of this structure activity relationship study, along with a description of the synthetic procedures used to prepare the compounds. The second study presented in this dissertation describes the development of a novel alkylation reaction of oxazol-5(4H)-ones and its use towards the syntheses of a,a-disubstituted-a-amino acids. The reaction is best described as an intermolecular ene-type reaction of oxazol-5(4H)-ones with enol ethers. This dissertation describes the initial discovery of the reaction along with the exploration of the reaction’s substrate scope and mechanism. Using this chemistry, oxazol-5(4H)—ones were alkylated using enol ethers and subsequently derivatized to afford a variety of a,a—disubstituted-a—amino esters. In addition, investigations using Bronsted acid catalysis for improving the overall diastemoselectivity of these reactions are discussed. This dissertation is dedicated to my late grandmothers: Rose Fisk and Jane Byers. The completion of these studies would not have been possible without the love, guidance and inspiration they provided me throughout my life. iv ACKNOWLEDGEMENTS The successful completion of the requirements for a research doctorate degree in the field of science can be an overwhelming experience involving both harsh disappointments as well as breathtaking triumphs. Often for one to effectively accomplish this task, one must call upon others for help in overcoming the physical and mental challenges at hand. During the course of these studies, I was fortunate enough to have such a support group. Without their help, I truly believe that I would have been successfully at completing this dissertation. Therefore, I would like to take the time to formally thank these people. To begin with, I would like to thank my graduate advisor, Professor Jetze J. Tepe, for the opportunity to study in his research group. The research experience Professor Tepe provided me not only allowed me improve my expertise as a synthetic chemist, but also helped me to grow overall as a scientist. His style of graduate student advisement was very complimentary to the manner in which I prefer to learn and advance. He allowed me to study relatively independently, thus providing me the opportunity to direct my research in a manner which I found fulfilling. He listened to my thoughts regarding my studies, and only provided criticism when absolutely needed. He also allowed me to learn from my mistakes, never getting too upset as I made them. Secondly, I would like thank my advisory committee members: Professor William Wulff, Professor Babak Borhan, and Professor Aaron Odom. From an outsider's perspective, it would appear that I did not take full advantage of my advisory committee as I did not all too often seek their advice or guidance. However, I learned a great deal from them by just merely observing the manner in which they conducted themselves as advisors and scientists. They are truly professionals and do a brilliant job of relaying the knowledge they have learned from their own experiences to their students and other colleagues. As I continue to establish my career, these gentlemen will continue to be role models that I will aspire to become. Last but certainly not least, I would like to thank my family and friends for their love and support during my dissertation studies. It is not the chemistry or long nights in the lab that I will remember most, but rather the interaction I had with my friends and family. Their love and support during this time meant more to me than they will ever know. I can only hope that the relationships that l established during these studies will continue for the rest of my life. Of these people, I would like to especially thank my wife, Professor Jaime Curtis-Fisk, for her love and support during these studies. She stood by me during both the good and the bad days of graduate school. I love her very dearly, and I look fonrvard to life with her for many years to come. vi TABLE OF CONTENTS LIST OF TABLES .............................................................................. x LIST OF FIGURES ............................................................................ xii LIST OF SCHEMES .......................................................................... xvi LIST OF SYMBOLS AND ABBREVIATIONS .......................................... xxi CHAPTER I OXAZOL-5(4H)-ONES AS TEMPLATES FOR GENERATING DIVERSE LIBRARIES OF BIOLOGICALLY INTERESTING COMPOUNDS ............................................................. 1 A. The need for chemical diversity in the drug discovery process ........... 1 B. Diversity oriented synthesis ....................................................... 4 C. Introduction to oxazol-5(4H)-ones ................................................ 5 D. Oxazol-5(4H)-ones as templates in diversity oriented synthesis ........ 8 E. Oxazol-5(4H)—one transformations associated with their acidity .......... 11 1. Alkylation of oxazol-5(4H)-ones .......................................... 12 2. Acylation of oxazol-5(4H)—ones .......................................... 13 3. Allylation of oxazol-5(4H)-ones ........................................... 15 4. Arylation of oxazoI-5(4H)-ones ........................................... 16 F. Oxazol-5(4H)-one transformations associated with their electrophilicity ......................................................................... 17 G. Cycloaddition reactions of oxazol-5(4H)-ones ................................ 23 1. [2+2] cycloadditions utilizing oxazol-5(4H)-ones .................... 25 2. [3+2] cycloadditions utilizing oxazol-5(4H)-ones .................... 28 H. Current work ........................................................................... 38 l. References ............................................................................. 41 CHAPTER II INHIBITION OF NF-KB MEDIATED GENE TRANSCRIPTION BY 2- IMIDAZOLINES DERIVED FROM OXAZOL-5(4H)-ONES ......................... 48 A. Synthesis of 2-imidazolines via mfinchnonefimine cycloadditions ......................................................................... 48 B. 2-imidazolines as inhibitors of NF-KB mediated gene transcription ........................................................................... 52 C. NF- KB mediated gene transcription ............................................. 54 D. Development of a new class of 2-imidazoline based NF-KB vii inhibitors ................................................................................ 56 E. Structure activity relationship study of trans-Z-imidazolines .............. 62 F. Structure activity relationship investigation of R1 ............................. 65 G. Structure activity relationship investigation of R2 ............................ 66 H. Structure activity relationship investigation of R3 ............................ 68 I. Structure activity relationship investigation of R4 ............................ 70 J. Experimental ........................................................................... 73 1. General .......................................................................... 73 2. Materials ........................................................................ 73 3. Compound synthesis and characterization ............................. 74 4. Cell culture ..................................................................... 100 5. NF- KB- Luc reporter assay ................................................. 101 6. Human whole blood IL-1B challenge .................................... 116 7. General procedure (whole blood assay) ................................ 116 K. References ............................................................................. 133 CHAPTER III INTERMOLECULAR REACTIONS OF OXAZOL-5(4H)-ONES WITH ALKENES: FROM CYCLOADDITION TO ALKYLATION REACTIONS ........................ 142 A. Synthesis of A1-pyrrolines via mflnchnonelalkene cycloadditions ...... 142 B. Proposed cycloaddition of munchnones with enol ethers ................. 145 C. Attempted cycloadditions using 4-methyl-2-phenyl-5(4H)-oxazolone and enol ethers ....................................................................... 150 D. Reversing the electronics of the reaction ...................................... 151 E. Discovery of a novel alkylation reaction of oxazol-5(4H)-ones ............ 153 F. Comparison to similar reactions found in literature .......................... 156 1. Conia—ene cyclization ....................................................... 156 2. The ortho-alkylation of phenols using alkenes ...................... 158 G. Solvent screening and product isolation ....................................... 160 H. Scope of enol ether ................................................................. 162 I. Scope of oxazol-5(4H)—one ........................................................ 167 J. Mechanistic investigation .......................................................... 169 K. In situ oxazol—5(4H)-one—formation .............................................. 177 L. Experimental ........................................................................... 182 1. General methods .............................................................. 182 2. Materials ......................................................................... 182 3. Procedures for synthesis of oxazoI-5(4H)-ones ....................... 183 4. Procedures for synthesis of enol ethers ................................ 190 5. Alkylation reactions of oxazol-5(4H)-ones and enol ethers ............................................................................ 197 6. General procedures for the alkylation reaction ........................ 198 M. References ............................................................................ 243 viii CHAPTER IV SYNTHESIS OF TERT-ALKY L AMINO HYDROXY CARBOXYLIC ESTERS VIA AN INTERMOLECULAR ALKYLATION REACTION OF OXAZOL-5(4H)-ONES Chiral Bronsted acid catalyzed ene—type reactions of oxazol- 5(4H)-ones and enol ethers as reported by Terada and co-workers....275 USING ENOL ET HERS ..................................................................... 252 A. Introduction to a,a—disubstituted a-amino acids .............................. 252 B. Synthesis of a,a—disubstituted a—amino acids using oxazoI-5(4H)-ones .................................................................. 253 C. Significance of ten-alkyl amino hydroxy carboxylic acids .................. 255 D. Synthesis of tert-alkyl amino hydroxy carboxylic acid using oxazol-5(4H)-ones ................................................................... 256 E. Intermolecular ene—type alkylations of oxazol-5(4H)-ones for the synthesis of tert-alkyl amino hydroxy carboxylic acids ..................... 260 F. Improving the diastereoselectivity using Lewis acids ....................... 261 G. Optimization of reaction conditions for improving diastereoselectivity .................................................................. 263 H. Other phosphoric acids ............................................................. 266 l. Various enol ethers .................................................................. 269 J. Various oxazol-5(4H)-ones ........................................................ 271 K. Application of the alkylation methodology towards the synthesis of Salinosporamide A .................................................................. 273 L. M . Experimental .......................................................................... 279 1. General information ......................................................... 279 2. Materials ......................................................................... 280 3. General procedure for the synthesis of oxazol-5(4H)—ones lV-27 to IV-29 .................................................................. 280 4. Synthesis of other enol ethers ............................................ 281 5. Synthesis of phosphoric acid derivatives .............................. 283 6. Synthesis of ted-alkyl amino hydroxy carboxylic esters ............. 287 N. References ............................................................................. 305 LIST OF TABLES CHAPTER II Table "-1. Evaluation of the degradation products produced from imidazoline "-4 for their ability to inhibit NF-KB signaling pathways ............. 60 Table "-2. Structure activity relationship study of R1 ................................. 66 Table "-3. Structure activity relationship study of R2 ................................. 68 Table "-4. Structure activity relationship study of R3 ................................. 70 Table "-5. Structure activity relationship study of R4 ................................. 72 CHAPTER III Table III-1. Reaction of 4-methyl-2-phenyl-5(4H)-oxazolone III-1 with butyl vinyl ether III-2 in the presence of Lewis acids ................................... 151 Table Ill-2. Reaction of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone III-5 with tart-butyl vinyl ether Ill-6 in the presence of various Lewis acids ........... 155 Table Ill-3. Screening of various solvents for the reaction of 2-phenyl-4- carbmethoxy-5(4H)-oxazolone III-5 and felt-butyl vinyl ether III-6 ................ 161 Table Ill-4. Reaction of 2-phenyl-4-carbmethoxy4(5H)-oxazolone III-5 with enol ethers of varying substitution pattern ............................................... 163 Table Ill-5. Reaction of 2-phenyl-4—carbmethoxy-4(5H)-oxazolone III-5 with enol ethers having varying protecting groups .................................... 166 Table Ill-6. Alkylation of various amino acids with 2,3-dihydrofuran III-9 utilizing oxazol-5(4H)-one intermediates ................................................ 179 Table Ill-7. The alkylation of 2-(methoxycarbonyI)-2-(benzamido)acetic acid III-4 using various enol ethers via oxazol-5(4H)-one intermediates ........ 180 CHAPTER IV Table IV-1. Reaction of 2-phenyl-4-carbmethoxy—5(4H)-oxazolone III-5 with tart-butyl vinyl ether III-2 followed by reduction with sodium borohydride .................................................................................... 264 Table N—2. Screening of various Lewis acids in the reaction of 2-phenyI-4-carbmethoxy-5(4H)-oxazolone III-5 with tart-butyl vinyl ether III-6 ........................................................................................ 265 Table lV-3. Screening of other diaryl phosphoric acid derivatives in the reaction of 2-phenyI-4wcarbmethoxy-5-oxazolone III-5 and ten-butyl vinyl ether III-6 ................................................................................. 267 Table IV-4. Reaction of various enol ethers with 2-phenyl-4- carbmethoxy-S-oxazolone III-5 under the optimized reaction conditions ....................................................................................... 270 Table IV-5. The reaction of various oxazol-5-(4H)-ones with tart-butyl vinyl ether III-6 under the optimized reaction conditions ............................ 271 xi LIST OF FIGURES CHAPTER I Figure I-1. Convergent synthetic pathway of target oriented synthesis... 2 Figure I-2. Comparison of TOS and DOS in terms of structural diversity. 3 Figure l-3. Divergent pathway of diversity oriented synthesis .................. 5 Figure I-4. Various isomeric forms of oxazolones .............................. 6 Figure l-5. Numbering of oxazol-5(4H)-one ring system according to the Hantzsch-erdman rules .............................................................. 8 CHAPTER II Figure "-1. Origin of diastereoselectivity using 2-aryloxazol-5(4H)-ones ...... 52 Figure "-2. Small molecule inhibitors of NF-KB mediated gene transcription .................................................................................... 53 Figure "-3. Mechanistic activation of the transcription factor NF-KB ............. 56 Figure "-4. Structure activity relationship of imidazoline Il-10 ..................... 63 Figure "-5. Methods for analyzing NF-KB inhibition .................................. 64 Figure "-6. Dose response curve of imidazoline lI-5 ................................. 102 Figure "-7. Dose response curve of imidazoline lI-8 ................................. 103 Figure "-8. Dose response curve of imidazole II-7 .................................... 104 Figure "-9. Dose response curve of imidazoline II-19 ............................... 105 Figure "-10. Dose response curve of imidazoline II-20 .............................. 106 Figure "-11. Dose response curve of imidazoline Il-21 .............................. 107 Figure "-12. Dose response curve of imidazoline II-22 .............................. 108 Figure "-13. Dose response curve of imidazoline II-23 .............................. 109 xii Figure "-14. Figure "-15. Figure "-16. Figure "-17. Figure "-18. Figure "-1 9. Figure "-20. Figure "—21. Figure "-22. Figure "-23. Figure "-24. Figure "-25. Figure "-26. Figure "-27. Figure “-28. Figure "-29. Figure "-30. Figure "-31. Figure "—32. Figure "-33. Figure "-34. Figure "-35. Dose response curve of imidazoline ll-24 .............................. 110 Dose response curve of imidazoline Il-25 .............................. 111 Dose response curve of imidazolineII-28 .............................. 112 Dose response curve of imidazoline Il-27 .............................. 113 Dose response curve of imidazoline Il-35 .............................. 114 Dose response curve of imidazoline II-36 .............................. 115 NMR spectra of imidazoline "-5 117 NMR spectra of imidazoline "-8 118 NMR spectra of imidazoline II-10 ............................................ 119 NMR spectra of imidazoline II-19...... 120 NMR spectra of imidazoline II-20 ............................................ 121 NMR spectra of imidazoline Il-21 ............................................ 122 NMR spectra of imidazoline Il-22 NMR spectra of imidazoline II-23 NMR spectra of imidazoline Il-24............... NMR spectra of imidazoline ll-25 NMR spectra of imidazoline II-26 NMR spectra of imidazoline Il-27...... NMR spectra of imidazoline II-34 NMR spectra of imidazoline II-35 NMR spectra of imidazoline "46 NMR spectra of imidazoline II-37 xiii 123 124 125 126 127 128 129 130 131 132 CHAPTER III Figure III-1. Naturally occurring AI-pyrrolines .......................................... 143 Figure Ill-2. Sustmann’s classification of 1,3-dipolar cycloaddition reactions ......................................................................................... 148 Figure III-3. Frontier molecular orbital explanation of the cycloaddition between munchnones and alkenes ...................................................... 152 Figure Ill-4. HMQC spectra of amino malonate III-15 ................................ 172 Figure Ill-5. 1H NMR spectra of the product mixture obtained from treatment of oxazoI-5(4H)-one Ill-5 with enol ether III-38 containing 50% deuterium ....................................................................................... 173 Figure Ill-6. 1H NMR spectra of the product mixture obtained from treatment of oxazoI-5(4H)-one III-5 with enol ether III-38 containing 80% deuterium ....................................................................................... 174 Figure III-7. NMR spectra of compound Ill-5 ........................................... 222 Figure III-8. NMR spectra of compound Ill-44 .......................................... 223 Figure III-9. NMR spectra of compound Ill-8 ........................................... 224 Figure Ill-10. NMR spectra of compound Ill-13 ........................................ 225 Figure Ill-11. NMR spectra of compound Ill-14 ........................................ 226 Figure Ill-12. NMR spectra of compound III-15 ........................................ 227 Figure III-13. NMR spectra of compound III-16 ........................................ 228 Figure III-14. NMR spectra of compound III-17 ........................................ 229 Figure Ill-15. NMR spectra of compound III-19 ........................................ 230 Figure Ill-16. NMR spectra of compound III-20 ........................................ 231 Figure III-17. NMR spectra of compound III-26 ........................................ 232 Figure III-18. NMR spectra of compound Ill-27 ........................................ 233 Figure Ill-19. NMR spectra of compound III-29 ........................................ 234 xiv Figure III-20. NMR spectra of compound III-33b ...................................... 235 Figure Ill-21. NMR spectra of compound III-36 ........................................ 236 Figure Ill-22. NMR spectra of compound III-37b ....................................... 237 Figure III-23. NMR spectra of compound III-39 ........................................ 238 Figure III-24. NMR spectra of compound III-41 ........................................ 239 Figure Ill-25. NMR spectra of compound Ill-46 ........................................ 240 Figure Ill-26. NMR spectra of compound Ill-49 ........................................ 241 Figure III-27. NMR spectra of compound Ill-50 ........................................ 242 CHAPTER N Figure lV-1. General structures of both a-amino acids and a,a-disubstituted cr-amino acids (quaternary a—amino acids) ............................................. 253 Figure IV-2. Naturally occurring tert-alkyl amino hydroxy carboxylic acids ..... 256 Figure lV-3. Crystal structure of compound IV-1A .................................... 266 Figure IV-4. NMR spectra of compound IV-1A ........................................ 296 Figure IV—5. NMR spectra of compound lV-1B ........................................ 297 Figure IV—6. NMR spectra of compound IV-22A ....................................... 298 Figure lV-7. NMR spectra of compound lV-228 ....................................... 299 Figure lV—8. NMR spectra of compound IV-23A ....................................... 300 Figure lV-9. NMR spectra of compound IV-23B ....................................... 301 Figure lV-10. NMR spectra of compound IV-24A ..................................... 302 Figure IV-11. NMR spectra of compound IV-248 ..................................... 303 Figure IV-12. NMR spectra of compound IV-25 ....................................... 304 XV LIST OF SCHEMES CHAPTER I Scheme l-1. Erlenmeyer and Mohr syntheses of oxazoI-5(4H)-ones ........... 7 Scheme l-2. Synthesis of oxazol-5(4H)-ones .......................................... 9 Scheme I-3. Pluripotent reactivity of oxazol-5(4H)-ones ............................ 10 Scheme l-4. Oxazol-5(4H)-ones as building blocks to create diverse libraries of compounds ........................................................................ 11 Scheme l-5. Alkylation of oxazol-5(4H)-ones with alkyl halides .................. 12 Scheme I-6. Michael addition of oxazol-5(4H)-ones to a,B-unsaturated aldehydes ....................................................................................... 13 Scheme l-7. Steglich rearrangement of O-acyl oxazoles ........................... 14 Scheme l-8. F u’s enantioselective Steglich rearrangement ....................... 15 Scheme l-9. Palladium catalyzed allylation of oxazol-5(4H)-ones using allenes ................................................................................... 16 Scheme l-10. Arylation of oxazol-5(4H)-ones ........................................... 17 Scheme l-11. Dynamic kinetic resolution of oxazol-5(4H)-ones .................. 18 Scheme l-12. Fu’s DKR of oxazol-5(4H)—ones ......................................... 19 Scheme I-13. Thiourea catalyzed DKR of oxazol-5(4H)-ones .................... 20 Scheme l-14. Synthesis of oxazoles from oxazoI-5(4H)-ones .................... 21 Scheme l-15. Synthesis of a marine alkaloid from the tunicate Dendrodoa grossulan'a ........................................................................ 22 Scheme I-16. Equilibrium of oxazoI-5(4H)-ones with their munchnone and amidoketene isomers ................................................. 24 Scheme I-17. Methods for increasing the concentration of the mlinchnone/amidoketene tautomers ..................................................... 25 xvi Scheme I-18. Staudinger reactions of oxazoI-5(4H)—ones ......................... 26 Scheme l-19. Diastereoselective Staudinger reaction of mtinchnones ....... 27 Scheme l-20. Synthesis of B-lactams via palladium catalyzed amidoketene formation ....................................................................... 28 Scheme l-21. Synthesis of pyrroles from mtimchnones ............................ 29 Scheme I-22. Synthesis of pyrroles via [3+2] cycloadditions of miinchnones ................................................................................... 30 Scheme l-23. Regioselective synthesis of pyrroles via [3+2] cycloadditions of a,8-unsaturated benzofuran-3(2H)-ones and munchnones ................... 31 Scheme l-24. Synthesis of imidazoles via [3+2] cycloadditions of munchnones ................................................................................... 32 Scheme I-25. Synthesis of imidazoles using solid supports ....................... 33 Scheme l-26. Synthesis of AZ-pyrrolines from munchnones ...................... 34 Scheme I-27. Reaction of oxazoI-5(4H)-ones with two equivalents of alkene to afford bicyclic heterocyclic scaffolds ...................................... 35 Scheme l-28. Maryanoff’s isolated primary cycloadduct ............................ 36 Scheme I-29. Amdtsen’s synthesis of 2-imidazoline carboxylates .............. 37 Scheme l-30. Lewis acid mediated cycloadditions of oxazoI-5(4H)-ones ....... 38 Scheme l-31. Research presented in this dissertation ............................... 39 CHAPTER II Scheme "-1. Traditional cycloadditions of mtinchnonesl amidoketenes with imines .................................................................. 50 Scheme "-2. Lewis acid mediated synthesis of imidazolines ..................... 51 Scheme "-3. Synthesis of the NF-KB inhibitor imidazoline Il-4 ................... 54 Scheme "-4. Deearboxylation of imidazoline II-4 ..................................... 58 xvii Scheme "-5. Derivatization of imidazoline II-4 ........................................ 61 CHAPTER III Scheme Ill-1. Formation of AZ-pyrrolines from N-alkylated munchnones ..... 144 Scheme III-2. Isolation of a primary cycloadduct from an intermolecular cycloaddition of a mUnchnone ........................................ 145 Scheme Ill-3. Lewis acid mediated cycloaddition of oxazoI-5(4H)-ones with electron deficient alkenes ................................... 145 Scheme III-4. Proposed cycloaddition of oxazoI-5(4H)-ones with enol ethers ...................................................................................... 146 Scheme Ill-5. Retrosynthetic analysis of a proposed synthesis of Lactacystin using oxazol-5(4H)-ones ..................................................... 147 Scheme Ill-6. Johnson's 1,3-dipolar cycloaddition between azomethine ylides and enol ethers ........................................................................ 149 Scheme Ill-7. Austin’s endo—selectllle 1,3-dipolar cycloaddition of isomtinchnones and enol ethers ............................................................. 150 Scheme Ill-8. Reaction of 2-pheyl-4-carbmethoxy—5(4H)-oxazolone III-5 with tart-butyl vinyl ether Ill-6 to form the quaternary substituted oxazolone Ill-7 .................................................................................. 154 Scheme III-9. Retrosynthetic route to the synthesis of Lactacystin using an alkylation reaction of an oxazoI-5(4H)-one by an enol ether as the key step ..................................................................................... 156 Scheme III-10. General thermal Conia-ene cyclization ............................. 157 Scheme Ill-11. Comparison of the Conia-ene cyclization to the intermolecular alkylation of oxazol-5(4H)-ones using enol ethers ............... 158 Scheme Ill-12. The thermal ortho-alkylation of phenols using enol ethers.... 159 Scheme III-13. Intermolecular alkylation of 2-phenyl-4—carbmethoxy- 5(4H)-oxazolone III-5 with tort-butyl vinyl ether III-6 followed by methanolysis .................................................................................... 162 Scheme Ill-14. Reactions of oxazol-5(4H)—ones with 2-methoxypropene III-18 ................................................................................................................ 164 xviii Scheme III-15. Reaction of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone III-5 with 1-butoxyethyne III-29 followed by methanolysis ................................. 167 Scheme Ill-16. Inten'nolecular ene-type alkylation reaction of oxazol- 5(4H)-ones III-5 and III-32 using ten-butyl vinyl ether Ill-6 .......................... 168 Scheme III-17. Reaction of 4-aryloxazol-5(4H)-ones and 4-alkyloxazol- 5(4H)—ones with tart-butyl vinyl ether Ill-6 ................................................ 169 Scheme Ill-18. Mechanistic investigation using the reaction of 2-phenyl-4wcarbmethoxy-5(4H)-oxazolone Ill-5 and 5—deutero-3,4- dihydro-2H-pyran III-38 ...................................................................... 170 Scheme Ill-19. Reaction of 2-phenyI-4-carbmethoxy-5(4H)-oxazolone Ill-5 with 2-deuterobutoxy ethyne Ill-40 ........................................................ 175 Scheme III-20. Comparison of the reactivity of 2-phenyl-4-carbmethyoxy- 5(4H)-oxazolone III-5 and 2-ethyl-4-carbmethoxy—5(4H)-oxazolone III-44 ...... 176 Scheme Ill-21. Current mechanistic understanding of the alkylation of oxazol-5(4H)—ones with enol ethers ....................................................... 177 Scheme Ill-22. Conducting the intermolecular alkylation reaction of oxazoI-5(4H)-ones with enol ethers while generating the starting oxazol-5(4H)-one in silu ...................................................................... 178 Scheme Ill-23. Potential hydrogen bonding interaction between the oxazol-5(4H)-one scaffold and the urea byproduct ................................... 181 CHAPTER IV Scheme lV-1. Synthesis of quaternary amino acids from quaternary oxazolones ...................................................................... 254 Scheme lV-2 Alkylation of oxazol-5(4H)-ones with alkyl halides ................ 254 Scheme IV-3. Hartwig’s palladium catalyzed arylation of oxazol-5(4H)-ones ............................................................................ 255 Scheme lV-4. Fu’s enantioselectilVe Steglich rearrangement ................... 258 Scheme IV-5. Allylic alkylation of oxazol-5(4H)-ones using alkoxy allenes... 259 xix Scheme IV-6. Synthesis of Sphinglofungin F starting from oxazol-5-(4H)-one IlV-1 ...................................................................... 260 Scheme lV-7. Proposed synthesis of tert-alkyl amino hydroxy carboxylic acids .............................................................................................. 261 Scheme IV-8. Proposed Lewis acid catalysis for improving the stereoselectivity of the alkylation reactions between oxazoI-5(4H)-ones and enol ethers ................................................................................ 262 Scheme lV-9. Reaction of 2-phenyl-4-carbmethoxy—5(4H)-oxazolone III-5 and ted-butyl vinyl ether III-6 in the presence of catalysts IV-14 and IV-15 ........................................................................................ 268 Scheme IV-10. Comparison of the reactivity between 2-aryloxazol- 5(4H)—ones and 2-alkyloxazol-5(4H)-ones ............................................... 273 Scheme lV—1 1. Corey’s synthesis of Salinosporamide A ........................... 274 Scheme lV-12. Synthesis of Corey’s intermediate lV-34 starting from lV-30A ............................................................................................ 275 Scheme IV-13. Enantioselective ene-type reaction of oxazoI-5(4H)-ones and enol ethers ................................................................................ 277 Scheme lV—14. Terada’s enantioselective ene-type reaction of oxazole-5(4H)-ones and enol ethers ..................................................... 278 KEY TO SYMBOLS AND ABBREVIATIONS Ac - Acetyl ALLN — N-acetyI-Ieucinyl-Ieucinyl-norieucinal Ar - Arvl Bn - Benzyl CSA - Camphor sulphonic acid DCE — Dichloroethane DCC - Dicyclohexylcarbodiimide DIBALH - Diisobutylaluminum hydride DKR - Dynamic kinetic resolution DMAP - 4-dimethylamino pyridine DMEM - Dulbecco’s modified eagle’s medium DMF - Dimethyl forrnamide DMSO — Dimethyl sulfoxide DNA - Deoxyribonucleic acid DOS — Diversity oriented synthesis EA — Elemental analysis EC5o- Half maximal effective concentration EDCI - Ethyldimethylaminopropyl carbodiimide ELISA — Enzyme linked immunosorbent assay ESI - Electrospray mass spectrometry Et - Ethyl EWG - Electron withdrawing group FAB — Fast atom bombardment FDA - Food and drug administration FMO - Frontier molecular orbital HeLa - Human cervical epithelial HOMO — Highest occupied molecular orbital HPLC — High pressure liquid chromatography HRMS - High resolution mass spectrometry leo- Half maximal inhibitory concentration I-KB - Inhibitory kappa B IL— Interleukin IR - Infrared LA - Lewis acid LRMS - Low resolution mass spectrometry Luc — Luciferase LUMO - Lowest unoccupied molecular orbital Me - Methyl MS - Mass spectrometry NF-KB - Nuclear transcription factor kappa B NMR - Nuclear magnetic resonance Ph - Phenyl RA - Rheumatoid arthritis R&D — Research and development SAR - Structure activity relationship tBu — tart-butyl TFA - Trifiuoroacetic acid TFAA - Trifluoroacetic anhydride TfOH — Triflic acid THF - Tetrahydrofuran TIPS — Triisopropylsilyl TLC — Thin layer chromatography TMS - Trimethylsilyl TNF-a — Tumor necrosis factor alpha TOS — Target oriented synthesis Ts - Tosyl UV — Ultraviolet xxiii CHAPTER I OXAZOL-5(4H)-ONES AS TEMPLATES FOR GENERATING DIVERSE LIBRARIES OF BIOLOGICALLY INTERESTING COMPOUNDS A. The need for chemical diversity in the drug discovery process Modern drug discovery involves screening libraries of small molecules for their ability to affect preselected biological targets including proteins and/or biological pathways.1 The completion of the human genome project has presented researchers with many more potential drug targets that were previously unknown. However, during recent years there has been a steady decline in the number of prescription drugs approved by the Food and Drug Administration (FDA).2'3 Furthermore, relatively few of the new prescription pharmaceuticals approved by the FDA could be classified as significant improvements over existing drugs.2'3 Several factors have contributed to the decline in the approval of new pharmaceuticals including the increased expense of researching and developing novel drug candidates. A second and perhaps more controllable reason for the decline in new drug approvals lies in the structural diversity of the small molecule libraries being screened for drug leads.” Although pharmaceutical companies are constantly combing nature for new drug leads, most small molecule collections come from commercial suppliers or previous medicinal chemistry projects. These libraries tend to be somewhat focused and lack structural diversity. However, the proteins that carry out biological processes essential for life are generally complex macromolecules containing high levels of structural diversity. This suggests that the molecular libraries we screen for drug leads should contain a complimentary amount of structural diversity. Synthetic organic chemists rely on three main approaches for synthesizing small molecules. The first approach is target oriented synthesis (T OS), which relies primarily on nature to discover potential synthetic targets. Synthetic targets may be identified by screening natural product extracts or proposed by means of spectroscopic analysis of target proteins. The second approach that synthetic organic chemists rely on is medicinal or combinatorial chemistry. This involves the structural optimization of an identified drug candidate through the synthesis of analogues. Both TOS and medicinal/combinatorial approaches involve the synthesis of small molecules to perturb a predetermined biological target. Synthetic routes using both approaches are generally linear and/or convergent (Figure l-1). They are planned in a reverse—synthetic order where complex molecules are transformed into simple and smaller precursors by mentally performing chemical reactions in reverse order. Target Oriented Synthesis O—-*O\ I“. >4 (:3 ——» :J/ ‘ Target Figure I-1. Convergent synthetic pathway of target oriented synthesis. Both TOS and medicinal/combinatorial chemistry approaches tend not to efficiently generate libraries of structurally diverse compounds. The goal of both approaches is to access very focused portions of chemical space (Figure I-2, A)?"4 Although synthetic'intermediates are generated along the synthetic pathway, many of the structures resemble that of the previous. Molecules with very similar structures quite often possess similar biological profiles. More recently, synthetic chemists have begun to utilize a third approach for generating small molecule libraries referred to as diversity oriented synthesis (DOS). In constrast to TOS and medicinal/combinatorial chemistry approaches, the goal of DOS is to create small molecule libraries comprised of compounds encompassing broad areas of chemical space (Figure l-2, B). The combination of DOS along with TOS and medicinal chemistry approaches will hopefully lead to the identification of new drug leads and ultimately to the treatment of disease. Descriptor 1 A Descriptor 2 Descriptor 3 V Descriptor 2 Descriptor 3 Figure I-2. Comparison of TOS and DOS in terms of structural diversity. 8. Diversity oriented synthesis During recent years, diversity oriented synthesis has become increasingly important to the development of new pharmaceuticals.3'4 New synthetic methods are allowing for efficient and rapid production of libraries of small yet complex molecules of biological importance. Screening of these libraries leads not only to identification of new drug candidates, but also to simultaneous identification of therapeutic protein targets with their small molecule regulators. In contrast to the convergent synthetic pathways used in T08 and medicinal/combinatorial chemistry, DOS rapidly produces libraries of compounds in a divergent manner (Figure I-3). DOS approaches aim to access areas of poorly populated or even vacant chemical space. There typically is no specified target in diversity oriented synthesis, thus retro-synthetic analysis is not applicable to planning. Instead, researchers must utilize more of a “forward- synthetic analysis” when proposing to use DOS. Chemists are to imagine generating complex and diverse molecules from simple precursors. DOS approaches rely on diversity-generating reactions, which are defined as the transformation of similar compounds into diverse libraries of molecules. Synthetic pathways should be no longer than three to five steps and therefore should avoid protection group manipulation when possible. To generate the highest levels of structural diversity, one must utilize reactions that generate molecules containing diversity in terms of core structure, stereochemistry, and functional group appendages. Diversity Oriented Synthesis \ o < 9, / O O _. Q < :1 >Multiple \ Q Targets [I] <2 0 J Figure I-3. Divergent pathway of diversity oriented synthesis. C. Introduction to oxazol-5(4H)-ones Oxazolones have proven to be very useful substrates in the field of synthetic organic chemistry.5 They exist in five different isomeric forms as illustrated in Figure I-4. Each isomeric form of oxazolone has unique attributes quite different from the others, making the class of molecules appealing for various applications. As a part of our research program focused on creating highly diverse libraries of small heterocyclic compounds for the purpose of discovering novel biologically active compounds, we have focused on the use of oxazol- 5(4H)—ones (also called azlactones) due to their availability and chemical versatility.°'7 R1 0 R1 0 R1 0 O O O O t Y Y Y R2 R2 0 R1 R1 oxazol-5(4H)-one oxazol-5(2H)-one oxazol-4(5H)-one oxazol-2(3H)-one oxazol-2(5H)-one or or or azlactone psuedoxazolone isoxazolone Figure l-4. Various isomeric forms of oxazolones. Oxazol-5(4H)-ones or azlactones are generally divided into two different classes: saturated and unsaturated (Scheme M). The first unsaturated oxazol-5(4H)-one was synthesized by Plbchl more than a century ago in 1883 via a condensation of benzaldehyde with hippuric acid in the presence of acetic anhydride.8 However, it was Erlenmeyer who established the first correct structure of oxazol-5(4H)-ones naming them “azlactones” in 1900.9’10 Erlenmeyer was also one of the first scientists to explore the use of other aldehydes in the reaction and to establish the use of unsaturated oxazol- 5(4H)-ones as precursors to new amino acid derivatives. The first saturated oxazol-5(4H)-one was not synthesized until 1908 by Mohr and co-workers, as it is believed that researchers before him failed to appreciate the ease at which saturated oxazol-5(4H)-ones can hydrolyze in the presence of moisture.11 Since that time, new and more efficient methods for producing oxazol-5(4H)-ones have been developed, some of which will be described in the following section. Erlenmeyer Azlactone Synthesis 0 Ph 0 /IL /\ + )l ACZO Y0 O Unsaturated Ph lNI COzH Ph N \ oxazol-5(4H)-one Ph Mohr Synthesis 0 R2 A 0 R1 0 )L A °2 F 0 Saturated R1 hl 002H oxazol-5(4H)-one R2 Scheme I-1. Erlenmeyer and Mohr syntheses of oxazol-5(4H)-ones. After the early work of Plbchl, Erlenmeyer, and Mohr, the chemistry of oxazol-5(4H)-ones remained fairly unexplored and was mainly limited to using oxazol-5(4H)-ones to make amino acid derivatives. It was not until the 1940’s, at which time the structure of penicillin was incorrectly thought to be an oxazol-5(4H)-one,12 that the chemical potential of oxazol-5(4H)-ones as synthetic intermediates was realized. Although it was eventually determined that the structure of penicillin actually contained a beta lactam ring system instead of an oxazol-5(4H)-one ring, the information obtained from these studies led way to the future development of new chemistry using the oxazol- 5(4H)-one scaffold. A variety of methods appear in the literature for naming oxazol-5(4H)- ones.”13 The ring is generally numbered according to the Hantzsch-Wildman rules giving priority to the oxygen atom and numbering the ring in the direction of the nitrogen atom as shown in Figure I-5.'3 One method refers to the oxazol-5(4H)-one substrate as an amino acid derivative.10 For example, the oxazoI-5(4H)-one derived from N-benzoyl alanine would be refered to as benzoyl alanine azlactone. A second method for naming oxazoI-5(4H)-ones describes the substrate as a dihydrooxazole.13 The oxazol-5(4H)-one derived from N-benzoyl alanine would then be referred to as 5-keto-4-methyI-2-phenyl -4,5-dihydrooxazole. This dissertation will primarily use a third system which consists of naming the scaffold as an oxazolone derivative. The compound is given the parent name of oxazol-5(4H)-one and the substituents are described in correspondence to their position on the ring. Using this system the oxazolone prepared from N-benzoyl alanine would be described as 4-methyl- \FO lso N34 2-phenyl-5(4H)-oxazolone. R1 R2 Figure l-5. Numbering of the oxazol-5(4H)-one ring system according to the Hantzsch-Wildman rules. D. OxazoI-5(4H)-ones as templates in diversity oriented synthesis Oxazol-5(4H)-ones have multiple features that make them attractive for use as building blocks in diversity oriented synthesis.7 To begin with, oxazol-5(4H)- ones are easily synthesized from the cyclodehydration of N-acyl-o-amino acids (Scheme l-2). A variety of N-acyl-a-amino acids are available for purchase from commercial suppliers. Furthermore, they are also easily synthesized either under Schotten-Baumann conditions or from the acylation of d-amino esters followed by hydrolysis.“ As seen in the pioneering studies of Erlenmeyer and Mohr, N-acyl-a-amino acids are traditionally converted into oxazol-5(4H)-ones by refluxing them in acetic anhydride.9"° These methods are often met with product isolation struggles directly associated with the problematic removal of the acid byproducts. Presently, oxazoI-5(4H)-ones are routinely synthesized in high purity under much milder conditions allowing for them to be subsequently used without the need for further purification. These methods generally consist of using relatively more reactive dehydrating reagents such as activated anhydrides (e.g. trifluoroacetic anhydride) or carbodiimides (e.g. occ or EDCI).15 QOH R O . 1 O )L’i-T‘ Dehydrating: Y 0 R1 N N H R2 R2 Reagent Scheme I-2. Synthesis of oxazol-5(4H)-ones. In addition to their ready accessibility, oxazol-5(4H)-ones contain numerous reactive sites allowing for a diverse set of possible transformations. The pluripotent reactivity of oxazol-5(4H)-ones allows for the rapid generation of a wide range of compounds making oxazol-5(4H)-ones ideal starting materials for DOS.16 The acidic nature of the proton(s) found at the C-4 position (pKa ~ 9)17 of the oxazol-5(4H)-one scaffold allows for the easy formation of an oxazole enolate, which can react with a range of electrophiles to form both O-alkylated and C-alkylated products (Scheme l-3, A). Alternatively, the use of Lewis acids with the oxazoI-5(4H)-ones results in the formation of either the 1,3-dipole B (also known as a munchnone) or the reactive ketene intermediate C (Scheme l- 3), each yielding the possibility of synthesizing novel heterocyclic compounds via cycloaddition reactions.18 Additionally, the oxazol-5(4H)-one ring contains a relatively electrophilic carbonyl susceptible to reaction with a wide range of nucleophiles including alcohols, amines, and hydrides to form various types of protected amino acids (Scheme l-3, D). R1 0 R2 T0 8 )L R; O A Bas:\ Audeophile D R1YO l Nfo R2 lM-X (Lewis acid) 0 H-X + / = + HX R B 2 M c Scheme I-3. Pluripotent reactivity of oxazoI-5(4H)-ones. This diverse reactivity of oxazoI-5(4H)-ones makes them excellent substrates for synthesizing a wide variety of useful and biologically interesting molecules (Scheme l-4). Highly substituted heterocyclic scaffolds can be directly accessed from oxazol-5(4H)-ones relatively easily and in a stereoselective manner. Furthermore, natural and unnatural amino acids can also be easily isolated in enantiopure form using oxazol-5(4H)-one intermediates. The remainder of this chapter will aim to illustrate how the pluripotent reactivity of oxazoI-5(4H)-ones allows for a wide range of 10 transformations, which in turn leads to the preparation of diverse libraries of biologically interesting compounds. R4 R1/ R OR2H HN/ 3 00 MR4 JV )L R3 R2 0 \ / H R2 3 9“ {R‘YC’ ‘ “4 R‘Y" M“) “1 NI / R3 *— R ——> I 'R3 2 \. R2 k Oxazolone R2 COZH / \ R4 R, 0 R1 Scheme I-4. Oxazol-5(4H)-ones as building blocks to create diverse libraries of compounds. E. Oxazol-5(4H)-one transformations associated with their acidity As compared to their acyclic N-acyl-a-amino acid precursors, oxazol-5(4H)-ones are considerably more acidic.”19 Their relatively high acidity in combination with their cyclic/less sterically encumbering structures allows for a variety of transformations generally difficult to perform with acyclic a-amino acids. These transformations include the alkylation, acylation, allylation, and arylation of oxazoI-5(4H)-ones, each of which potentially results in the formation of quaternary substituted oxazolones. Subsequent nucleophilic ring opening of 11 quaternary substituted oxazolones produces a variety of interesting molecules, including novel a,a—disubstituted a-amino acids. 1. Alkylation of oxazoI-5(4H)-ones. One of the more traditional methods for synthesizing a,a-disubstituted a-amino acids involves the alkylation of oxazol- 5(4H)-ones. Treatment of oxazoI-5(4H)-ones with mild bases such as triethylamine or Ht'rnig’s base results in the deprotonation of the oxazol-5(4H)— one and formation of an aromatic oxazole enolate.20 Formation of these oxazole enolates while in the presence of highly reactive electrophiles leads to the formation of the desired quaternary substituted oxazolones (Scheme l-5).21 These reactions often suffer from the formation of undesired side products primarily due to competitive O-alkylation of the enolate intermediates. Recent developments optimizing the reaction conditions have allowed for the use of a wider range of electrophiles,21 although O-alkylation still remains a problem with many substrates. R1 0 R1 0 R1 R e O 1 0 Base \Il/ R3-X 1r 0 —' NJg‘O _’ 122:0 + iii/X‘OR" R2 R2 R2 R3 R2 Oxazole Quaternary Aromatic Enolate Oxazolone Oxazole Scheme I-5. Alkylation of oxazol-5(4H)—ones with alkyl halides. A variety of more regioselective methods have appeared in the literature as of late for the alkylation of oxazol-5(4H)-ones. One of the more recent methods for alkylating oxazol-5(4H)-ones was reported by Jorgensen and oo- 12 workers in 2008.22 The authors reported an organocatalytic enantioselective Michael addition of oxazoI-5(4H)-ones to a,B—unsaturated aldehydes (Scheme l- 6). Although these reactions generally proceed with low to moderate diastereoselectivity, they do produce two new stereogenic centers in relatively high yields and moderate to excellent enantioselectivity. Ar R1 0 R1 N Ar \— + / A? "‘2: 3W Ar = 3.5(CF3I206H3 Ra R2 R2 Toluene, r.t. \ 0 83-96% e.e. 0-82% d.e. 38-88% yield Scheme I-6. Michael addition of oxazol-5(4H)-ones to (LB-unsaturated aldehydes. 2. Acylation of oxazoI-5(4H)-ones. The traditional method for synthesizing 4-acyl substituted oxazol-5(4H)-ones is the Steglich rearrangement. The reaction was first discovered by Steglich and co-workers in 1970.23 The authors reported a nucleophilic base (e.g. DMAP) catalyzed rearrangement of O-acylated oxazoles to form C-4 acylated oxazolones (Scheme l-7). The initial reaction of DMAP with the O-acylated oxazole is believed to be reversible, while formation of the product appears to be an irreversible process (Scheme l-7).2‘ Since the initial discovery of the reaction, a variety of chiral nucleophiles have been developed for catalyzing the reaction to make new enantiomerically enriched oxazolone scaffolds.25 13 >43 DMAP R1170 l'" R1 0 _ e 0 R2 16> R3 DMAP _ J Scheme l-7. Steglich rearrangement of O-acyl oxazoles. The first asymmetric Steglich rearrangment of oxazoI-5(4H)-ones was reported in 1998 by Fu and coworkers.”26 The authors reported the use of a chiral ferrocene-fused DMAP derivative, PPY*, for promoting the asymmetric acyl migration (Scheme l-8). Alkyl substituents at the 2 position of the oxazol- 5(4H)-one scaffold generally provide lower enantioselectivity than aryl and heteroaryl substituents at that same position. The reaction tolerates a variety of substituents at the C-4 position of the oxazol-5(4H)-one resulting in high enantioselectivity (Scheme l-8). Choosing the appropriate migrating acyl group can enhance the stereoselectivity of the reaction. The use of benzyl substituted acyl groups tend to result in higher stereoselectivity than most aliphatic groups.24 14 M90 60: Y} :>—oen 2 mol%( ) PPY; : 17° 0 0°C, 2-6 hrs 0 tert-amyl alcohol RBnO a 88-92% ee 93-95% yield ‘N l Scheme I-8. F u’s enantioselective Steglich rearrangement. 3. Allylation of oxazoI-5(4H)-ones. A third transformation resulting in the formation of quaternary oxazolones is the allylation of oxazoI-5(4H)-ones.27 A variety of transition-metal catalyzed methods have been reported for the allylation of oxazol-5(4H)-ones.27'29 One of the more recent methods for the allylation of oxazol—5(4H)-ones was reported by Trost and co—workers in 2003. The authors reported a palladium-catalyzed addition of oxazol-5(4H)—ones to electron rich allenes (Scheme l-9).29 Their methodology avoids any regioselectivity problems by electronically biasing one end of the allene with an electron rich alkoxy group. Overall the reaction works very well, generating two new stereogenic centers including a quaternary center at the C-4 position of the oxazolone ring. Oxazol-5(4H)-ones with aliphatic substitutions at the C-4 position provide the best results affording the products in moderate yields (67- 87%) with excellent enantiomeric excesses (90-94%) (Scheme l-9). Furthermore, the diastereoselectivity observed in these reactions was also 15 reported to be relatively high, usually occurring in approximately a 20:1 ratio (Scheme l-9). o O l=d(oiiCl=3)2 (2 mol%) 0 BnO H OBn , R\(lko 1 (6 m0|%) > M ==/ KO‘Bu (2 mol%) —‘< \ R\\‘ O N— _ ph Hippuric Acid (20 mol %) NT< H2CI2, Ft. Ph ‘ 67-87% yield 85-94% ee 0 , 0 Up to 20:1 dr NH HN PththP 1 Scheme I-9. Palladium catalyzed allylation of oxazol-5(4H).ones using allenes. 4. Arylation of oxazoI-5(4H)-ones. The arylation of oxazol-5(4H)-ones at the 04 position results in the formation of novel quaternary aryl-glycine derivatives?“1 In 2003, Hartwig and co-workers reported the first palladium catalyzed arylation of oxazol-5(4H)-ones for the synthesis of quaternary amino acids (Scheme MO).30 The reaction involves the coupling of the sp2 carbon of arenes with the enolate of oxazol-5(4H)-ones. Their catalyst system consisted of using Pd(OAc)2 along with the sterically hindered electron rich ligand Ad2P(t- Bu) (Scheme MO). The reaction provides reasonable yields with a wide range of aryl bromide substrates. The use of electron rich or electron neutral aryl groups provides the best results (75-85%), whereas electron poor aryl groups tend to afford slightly lower yields (~60%). Aryl groups with the potential to undergo Heck reactions, such as 4-bromostyrene, underwent the desired 16 coupling reaction in good yields (75%). A wide range of oxazol-5(4H)-one substrates consisting of both aliphatic and aromatic substituents undergo the desired coupling reaction in good yields. ArBr Pd(OAc)2 Ph\(o o Ad2P(t-Bu)> Ph \«0 0 gr choa ”I Toluene Ar R R 58-85% yield Scheme l-10. Arylation of oxazol-5(4H)-ones. F. OxazoI-5(4H)-one transfonnations associated with their electrophilicity The oxazol—5(4H)-one scaffold contains a highly electrophilic carbonyl that readily undergoes a variety of transformations including hydrolysis, alcoholysis, aminolysis, hydride reduction, and Friedel-Crafts reactions to generate both a- amino acids as well as novel heterocyclic compounds.16 Early studies involving oxazol—5(4H)-ones include their participation as intermediates during peptide coupling reactions. These studies indicated that oxazol-5(4H)-one intermediates are primarily responsible for the racemization of amino acid residues when peptides are coupled with nucleophiles using reagents such as DCC.12 More recent work has taken advantage of not only the ability of oxazol— 5(4H)-ones to rapidly epimerize, but also their highly electrophilic character to generate q-amino acid derivatives. The dynamic kinetic resolution of oxazol- 5(4H)-ones has proven to be very useful for the preparation of enantiomerically pure q-amino acids (Scheme l-11).32 During the process, a chiral catalyst (small molecule or enzyme) preferentially activates one of the 17 two enantiomers of the oxazol-5(4H)-one racemate towards irreversible alcoholysis, thus forming an amino ester product. Simultaneously, the unreactive enantiomer undergoes epimerization to the more reactive enantiomer, which subsequently undergoes alcoholysis in the presence of the catalyst theoretically allowing for complete conversion of the racemate to the desired stereoisomer (Scheme l-11).3'2 R1 0 R1 0 R1 0 112:0 _. DOH , 11220 R‘ H R2 if R2 Chiral Catalyst Chiral Catalyst R30H R3OH 0 R2 0 R2 “infirm, R1/ILu/Hr0R3 O 0 Scheme I-11. Dynamic kinetic resolution of oxazol-5(4H)—ones. The dynamic kinetic resolution of oxazol-5(4H)-ones has been reported using both enzymatic33 and small molecule catalyst systems.“35 One of the first succesful examples using a small molecule to catalyze the dynamic kinetic resolution of oxazol-5(4H)-ones was demonstrated by Fu and co- workers in 1998 (Scheme l-12).3’5 The authors described the use of the chiral ferrocene-fused DMAP derivative 2 to catalyze the methanolysis of oxazol- 5(4H)-one substrates resulting in enantiomerically enriched amino acids?6 Examples using both 4-aryl-oxazol-5(4H)-ones and 4-alkyl-oxazol-5(4H)-ones provided a-amino esters in excellent yields (>90%) with moderate 18 enantioselectivity (44-61% e.e.) (Scheme l-12). The authors also reported the stereochemical outcome of the reaction to be solvent dependent with toluene furnishing the highest level of enantioselectivity. Adding steric bulk to the alcohol nucleophile increases the enantioselectivity of the reaction, albeit with highly increased reaction times. 0 R Ph 0 0 Hr o 5 mol/oz > )L We N MeOH Ph ii R 1 toluene, rt. 0 0% PhCOzH ’ ‘ 93-98% yield MezN 44-61% ee CF? Me Fe Me Me’b‘Me Me 2 L Scheme l-12. Fu’s DKR of oxazoI-5(4H)-ones. More recent work in this area was published by Berkessel and co- workers in 2005 utilizing urea and thiourea bifunctional organocatalysts to promote the dynamic kinetic resolution of oxazol-5(4H)-ones.34 The Lewis acidic urea moiety of the catalyst activates the oxazol-5(4H)-one carbonyl via hydrogen bonding, while a tethered Lewis basic portion of the catalyst presumably directs the approach of the nucleophile by means of a second hydrogen bonding interaction (Scheme l-13). These catalysts work well with a wide range of oxazol-5(4H)-ones providing enantiomeric excesses up to 91%. Analogous to Fu’s studies, the use of smaller primary alcohols results in higher conversion rates than bulkier alcohols with allyl alcohols affording the 19 highest yields. To complement the theory that complexation of the catalyst to the substrate occurs via hydrogen bonding, solvents capable of accepting hydrogen bonds (e.g. THF) provide little or no stereoselectivity. 5mol%3 'R‘ R1 0 R1 0 H‘N al lalcohol 1.5 _ 112:0 fly 1 eq)_ 112—0'“ S R2 toluene, r.t. I-I R2 H’N *Chlral F'is Me {Bu s Bn’N u N ‘ o NMe2 3 i F32 - O\/\ ., my \ 77-94% yield 78-91%ee Scheme l-13. Thiourea catalyzed DKR of oxazoI-5(4H)-ones. The electrophilic nature of oxazoI-5(4H)-ones not only allows for their use in the synthesis of a-amino acids, but also permits access to a variety of heterocycles including oxazoles, hydantoins, thiohydantoins, pyridones, pyrimidinones and many more.7 In an effort to develop a new route to highly substituted oxazoles, our group recently published a protocol consisting of a one pot Friedel-Crafts / Robinson-Gabriel synthesis for producing 2,4,5- ).3‘3 Previous trisubstituted oxazoles from oxazol-5(4H)-ones (Scheme l-14 reports have established the Robinson-Gabriel cyclodehydration of 2- acylamino ketones as one of the most versatile routes for producing oxazoles. Furthermore, 2-acylamino ketones can be readily prepared from oxazol-5(4H)- 20 ones via Friedel-Crafts reactions.36 The two reactions can be carried out in tandem utilizing a combination of aluminum chloride and trifluoromethanesulfonic acid, resulting in the formation of oxazoles directly from oxazol-5(4H)-ones. The reaction works well for a wide variety of oxazol- 5(4H)-one substrates including both aromatic and alkyl substituted oxazol- 5(4H)-ones. As anticipated, the reaction affords the highest yields with either electron neutral or electron rich arenes. Electron deficient arenes provide very little or no product formation. Presumably the oxazol-5(4H)-one is initially activated by aluminum chloride promoting the FriedeI-Crafts reaction providing an 2-acylamino ketone intermediate (Scheme M4). The ketone carbonyl of the intermediate is then protonated by the trifluoromethanesulfonic acid, activating the substrate towards cyclization and subsequent dehydration to the corresponding oxazole. R3 R3 0 R ““601” O - )L 493 N AlCl3 R1 fl R2 0 l TfOH R3 R3 1 7— 11W N N R2 H20 L R2 _ 55-81% yield Scheme l-14. Synthesis of oxazoles from oxazol-5(4H)-ones. 21 More recently, our research group utilized the electrophilic nature of the oxazol-5(4H)-one scaffold to synthesize a marine alkaloid from the tunicate Dendrodoa grossulan’a (Scheme MS)” The synthesis utilizes two key rearrangements to afford the final compound in rapid and efficient fashion (12 steps). The first rearrangement consists of the cyclodehydration of thiourea 4 to afford an oxazole intermediate, which subsequently undergoes a Claissen rearrangement to produce a quaternary substituted oxazolone.’""39 The oxazolone intermediate is then treated with sodium methoxide facilitating the second rearrangement to yield a quaternary hydantoin.39 The synthesis is completed in five additional steps affording the natural product 5 in 13% overall yield. ii i ° 8 \ N\ Ts 4 Oxazole Intermediate 0 H N N EtO C’ 0 o 2 WT .— HN NaOMe N \ "l “l Ts Ts Hydantoin Oxazolone Scheme l-15. Synthesis of a marine alkaloid from the tunicate Dendmdoa grossulan'a. 22 G. Cycloaddition reacb'ons of oxazol-5(4H)-ones Cycloaddition reactions of oxazol-5(4H)-ones have been utilized to generate a wide variety of heterocyclic scaffolds.7"°'“ Oxazol-5(4H)-ones exist in equilibrium with both their mesoionic (also referred to as a miinchnone) and amidoketene isomers (Scheme l-16).“2 The relative concentration of each isomeric form of the molecule is highly dependent on reaction conditions and the substitution pattern of the oxazol-5(4H)-one scaffold. Milnchnones are best described as cyclic l aromatic azomethine ylides. Analogous to their acyclic azomethine ylide counterparts, miinchnones undergo [3+2] cycloadditions while in the presence of a range of dipolarphiles to afford a variety of highly substituted heterocyclic scaffolds.“'“ Hypothetically, mt'lnchnones exist in equilibrium with a low concentration of their respective amidoketene isomer.“45 Although the amidoketene isomers of oxazol-5(4H)-ones have not been observed spectroscopically to date, certain chemical transformations provide plausible evidence for their existence. For example, treatment of oxazol-5(4H)-ones with imines at elevated temperatures results in the formation of highly substituted [3- lactams.“1 The B—lactam products are presumed to arise via a [2+2] cycloaddition of the amidoketene isomer of the oxazol-5(4H)—one with the imine. 23 O 0 R1 0 V. .___- t/ «i ._._-. 11 ll RAN 1 R2 R2 H R2 OxazoI-5(4H)-one Hunchnone Amidoketene Scheme I-16. Equilibrium of oxazol-5(4H)-ones with their mi'lnchnone and amidoketene isomers. Previous studies by Huisgen and co-workers demonstrated that a moderate equilibrium concentration of the miinchnone tautomer is necessary to promote 1,3—dipolar cycloadditions of oxazol-5(4H)-ones.46 The concentration of munchnone tautomer can be increased in a variety of ways including the N- alkylation of the oxazoI-5(4H)-ones (Scheme l-17). Huisgen and co-workers first described the synthesis and cycloaddition chemistry of N-alkylated m0nchnones.“"6"7 N-alkyl mitnchnones were generated via the cyclodehydration of N-alkyl, N-acyl amino acids. More recently, Amdtsen and co-workers reported a transition metal catalyzed synthesis of N-alkylated mt'lnchnones (Scheme l-17).“"49 The desired m0nchnone species was generated utilizing a palladium catalyzed coupling of an imine. acid chloride and carbon monoxide. In addition, Merlic and co-workers reported acylamino carbene complexes to readily undergo carbon monoxide insertion, thus generating N-alkylated munchnones (Scheme l-17).5o Alternatively, the concentration of the mt'lnchnone tautomer may be increased while in the presence of Lewis acids (Scheme l-17).5"53 Recently, our research group described mild conversions of oxazol-5(4H)-ones to dihydroheterocyclic scaffolds 24 via Lewis acid promoted [3+2] cycloadditions. This work will be described in more detail in the following chapters of this dissertation. Formation of N-alkyl WinchnonelAmldoketenes R3. N O CO 0 /Ik /IL )\ Dehydrating iii Pd(0) R2 H R1 N R2 / I + r Reagent R3 (9 /U\N R2 CO 0 R3 ' NEt‘Pr R3 2 JL N-alkyl N-alkyl Cl R1 Munchnone Amldoketene Fonnatlon of Lewis Acid Coordlnated Mtinchnones CO insertation R1 0 M-X (Lewis Acid) 0 Cr(CO)s {8:0 ..___. R\NIFO + HX . M 6') 6 R1 fill R2 R R2 R3 Lewls Acid Coordinated Miinchnone Scheme l-17. Methods for increasing the concentration of the munchnone/amidoketene tautomers. 1. [2+2] cycloadditions utilizing oxazol-5(4H)-ones The development of novel and efficient routes to synthesize B-lactams is an area of significant interest primarily due to their antibiotic properties. Previous studies by Staudinger and co-workers established that ketenes undergo themal [2+2] cycloadditions with imines to form highly substituted [3- lactams.“5"‘7 Oxazol-5(4H)-ones participate in Staudinger-type cycloaddition reactions with imines via their amidoketene isomer (Scheme l-18).‘2'£54 The first Staudinger reaction of oxazol-5(4H)-ones was demonstrated by Huisgen and co-workers in 1971.55 The authors proposed munchnones to exist in 25 equilibrium with a low concentration of amidoketene, which in turn undergoes a [2+2] cycloaddition while in the presence of an imine to afford the observed B—Iactam product (Scheme I-18). N’Ra - o T l o ,R .l / 0 -————- )L ——-—-‘ 2 )L R2 H369 P“ '1‘ P“ P“ '7‘ Ph R1 Ph CH3 CH3 Scheme I-18. Staudinger reactions of oxazol-5(4H)-ones. Recenty, Cremonesi and co-workers reported the use of bicyclic mt'mchnones for the diastereoselective synthesis of B—lactams (Scheme l- 19).56 The authors illustrated the ability to control the stereochemical outcome of the reaction through alteration of the imine N-substituent (Scheme l-19, R2). The reaction afforded primarily the cis—B-lactam product (with respect to the sulfur and phenyl groups) when performed with imines substituted with electron withdrawing moieties. Conversely, high yields of trans-B-lactams were selectively obtained utilizing imines substituted with electron donating groups.56 The authors speculate that both products arise from the initial attack of the imine to the least-hindered side of the amidoketene intermediate (Scheme l-19). Electron withdrawing groups destabilize the initial iminium intermediate, thus leading to the cis-product. On the other hand, electron donating substituents stabilize the iminium intermediate allotting for double bond isomerization, which consequentially gives rise to the thermodynamically favored trans-product.56 26 ,COR1 N ,COR1 90R1 40 RZ‘NA Ph {Noe (”H0 H N N QY NEta, CH2CI2 / N® H _ S TBflUX \I/ sR2 fjh ‘Rz 20-91% yield Ph - 015 '1 36-94% d.e. ll .- R, o ,COR1 OR Y O (N N’C 1 O G O {11‘ 851° -—» Ce 3 Ph N9 Ph - N. Y R2 H R2 H trans Scheme l-19. Diastereoselective Staudinger reaction of munchnones. More recently, Amdsten and co-workers reported a multicomponent palladium-catalyzed synthesis of amidoketenes, which were subsequently treated with imines to afford B-lactams in moderate to high yields (Scheme l- 20).57 Their overall reaction sequence couples four reagents to efficiently provide B—lactams in a single step. The amidoketene species is generated in situ starting from an imine, acid chloride and carbon monoxide as shown below in Scheme l-20. The formation of the amidoketene species is initiated by an oxidative addition of an N-acyliminium salt to Pd(0). The resulting palladacyle then undergoes carbon monoxide insertion followed by B—hydride elimination to form the needed amidoketene intermediate. The amidoketene intermediate then undergoes a Staudinger-like [2+2] cycloaddition with an imine to afford the final B-Iactam product. For selective B-lactam formation, the authors reported the requirement of base to eliminate HCI from the reaction. Insufficient removal of HCI leads to lower yields of product presumably due to 27 competing [3+2] cycloaddition pathways. These reactions can be performed either using two equivalents of the same imine or with sequential addition of two different imines. ° “ i313 + —> / GD R1JLCI R~°"N4‘\R2 R10 Psz 0.9 “EM co 0 R o N 3 H R2 O+Pd CO /ll\ R2 .— + l. Pd(0) JV” R1 N [2+2] R1 N /H\R2 R1 N l R2 H R l R2 R3 3 R3 Cl O/Pdio HCI R1/u\ H l R2 R3 Scheme l-20. Synthesis of B-Iactams via palladium catalyzed amidoketene formation. 2. [3+2] cycloadditions utilizing oxazol-5(4H)-ones A wide variety of heteroaromatic and dihydro-heterocyclic scaffolds are accessible utilizing oxazol-5(4H)-ones by means of their 1,3-dipolar munchnone tautomer. Analagous to acyclic azomethine ylides, munchnones undergo cycloadditions with a variety of dipolarphiles.“44 For example, the thermal 1,3-dipolar cycloaddition of mt'lnchnones with electron deficient alkynes directly results in the formation of highly substituted pyrroles (Scheme l-21).18 Mechanistically, these reactions proceed through the initial formation of a bicyclic primary cycloadduct, which subsequentaly undergoes aromatization via the loss of carbon dioxide. This area of research was 28 pioneered by Huisgen and co-workers starting in 1970.46 The authors proposed that a moderate concentration of the mesoionic mtinchnone tautomer is essential for promoting the 1,3-dipolar cycloaddition reactions of oxazol-5(4H)-ones. The authors reported the preparation of a wide range of pyrroles with diversity at every substituent of a pyrrole scaffold utilizing N- alkylated munchnones (Scheme l-21).46 N-alkylation of oxazoI-5(4H)-ones locks them into their mfinchnone form, allowing for the formation of a variety of heteroaromatic molecules including pyrroles.‘“"5°'58 O R 1&0 ...___. T}? “3 = R4. H’e R2 R2 Oxazol-5(4H)—one Mllnchnone R3 R3 R1 / -co R1 HN / R4 ‘——-2 Wm H 9 R2 R2 C02 Pyrrole Scheme l-21. Synthesis of pyrroles from miinchnones. Similarly, pyrroles may also be synthesized from miinchnones through their reaction with alkyne equivalents. The regioselective cycloaddition using vinyl phosphonium salts with N-alkyl mfinchnones has been reported by Clerici and co-workers (Scheme l-22).‘o Miinchnones were refluxed (THF/DMF solvent mixture) in the presence of vinyl phosphonium salts resulting in high 29 yields of the desired pyrroles by way of the decarboxylation of the primary cycloadducts followed by elimination of PPh3_ The regioselectivity observed in the reaction is driven by the strong electrostatic interaction between the phosphonium species and the enolate of the miinchnone. Additionally, the regioselective synthesis of pyrroles has been demonstrated in the reaction of miinchnones with electron deficient vinyl-chlorinated alkenes (Scheme l-22)."’9 Aromatization of the primary cycloadducts to the final pyrrole products is accomplished via decarboxylation of the primary cycloadduct followed by subsequent elimination of the chlorine atom. '32 R1\<\:‘IR3 R4 9 R4 %9 Br 35-68% yield R2 0 e N EDIE/2‘0 R4——: R5 ‘ WWW, R{ T R3 R5 R4 60-98% yield F3C R3 R2 R1 \N/ R3 R300 CF3 33-89% yield Scheme l-22. Synthesis of pyrroles via [3+2] cycloadditions of mfinchnones. 30 More recently, Park and co-workers reported a regioselective synthesis of pyrroles utilizing 1,3—dipolar cycloadditions of o,B-unsaturated benzofuran- 3(2H)-ones and mlinchnones (Scheme l-23). Their method involves the use of AgOAc to generate the needed miinchnone species in situ, which readily undergoes a [3+2] cycloaddition with an o,[3-unsaturated benzofuran-3(2H)— one. Subsequent spontaneous decarboxylation of the primary cycloadduct results in the formation of the desired tetrasubstituted pyrroles in high yields with regioselectivities greater than 99:1. Benzofuranone derivatives more electron withdrawing in nature produced the highest yields and regioselectivities, while benzofuranone derivatives containing more electron donating substituents resulted in decreased reaction rates and regioselectivity. R F n) R q 1Y 1 R AgOAc (5N3 3%0 THF Microwave R2 0 O -COz R3 R1 / O OH HN / R2 84-94% Yields Scheme l-23. Regioselective synthesis of pyrroles via [3+2] cycloadditions of o,B-unsaturated benzofuran-3(2H)-ones and miinchnones. 31 The construction of the imidazole scaffold has also been accomplished starting from oxazol-5(4H)-ones through the reaction of their manchnone tautomer with nitriles and imines. Early work by Huisgen and co—workers demonstrated the ability of munchnones to undergo thermal cycloadditions with electron deficient nitriles (Scheme l-24).‘30 Similar to the reactions of mfinchnones with alkynes, these reactions proceed by initially forming a bicyclic—cycloadduct, which undergoes decarboxylation resulting in the formation of the imidazole products. The reaction of N-alkyl munchnones with various imines also results in the formation of highly substituted imidazoles. Consonni and co-workers demonstrated that the reaction of munchnones with N-(phenylmethylene)benzenesulphonamides afford imidazoles in moderate yields (Scheme l-24).61 Aromatization occurs by way of the decarboxylation of the intermediate bicyclic cycloadduct followed by expulsion of benzenesulphinic acid. '32 '32 R1\«N/ R3 = ®NI f0 —:N R1\«N/ R3 N—Z/ Rz/ NI R4 R3 R4 20-65 yield% 41 -71 % yield Scheme I-24. Synthesis of imidazoles via [3+2] cycloadditions of mflnchnones. The 1,3-dipolar cycloaddition reaction of muchnones with imines often results in low yields of imidazole product due to the formation of a variety of side products including B-lactams and dimerized miinchnone. Recent approaches to increase the yield of such cycloadditions include the use of 32 solid supports to prevent the dimerization of the miinchnone starting material. Bilodeau and co-workers demonstrated the benefit of this approach by using N-acyl o-amino acids bound to solid support.62 The N-acyl o-amino acids were cyclodehydrated using EDCI affording resin bound miinchnones, which subsequentially underwent the desired 1,3-dipolar cycloaddition reaction with N-tosyl imines to yield polymer-linked imidazoles (Scheme l-25). Liberation from the imidazole product from the solid support resin by heating in acetic acid afforded the desired imidazoles in high yields. OMe R OMe R1 1 /\ o rag/K0 E00 0 )QN c5 CHZClz 6 R2 Scheme l-25. Synthesis of imidazoles using solid supports. 53 - 99% yield Although relatively few examples have been reported, dihydro- heterocyclic scaffolds are accessible utilizing 1,3-dipolar cycloaddition reactions of miinchnones.1"'5"~"3'53'65 Traditional methods for conducting oxazol-5(4H)-one cycloadditions (e.g reflux in acetic anhydride) often lead to heteroaromatic products or complex mixtures of products containing various isomeric mixtures of dihydro-heterocyclic molecules (Scheme l-26). Studies by Gotthardt, Huisgen, and Schaefer illustrated that Az-pyrrolines could be isolated in cycloadditions of mt'inchnones with electron deficient alkenes by decreasing the temperature of the reactions (Scheme l-26).66 MiJnchnones were heated at relatively low temperatures (50-100 °C) in xylenes while in the 33 presence of alkenes resulting in primarily the isolation of Az-pyrrolines with small amounts of pyrrole. Unfortunately, the decarboxylation of the primary cycloadducts of these reactions could not be controlled and ultimately led to mixtures of 152-pyrroline regioisomers. R1 0 mm R5 R1 _002 R1 elf" 9,; R. ell R. e / R2, R2 R3 CO? R2/ R4 R4 R1 R1 / R + R / N 5 / N / 5 R2 R2 R3 R3 Az-Pyrrollnee Scheme I-26. Synthesis of Az-pyrrolines from mlJnchnones. Studies by Huisgen and co-workers explored the mechanism by which the primary cycloadducts obtained from miinchnone cycloadditions with alkenes decarboxylate and form pyrroline and pyrrole products.“'66 Their studies indicated that the initial loss of carbon dioxide leads to the formation of a relatively reactive cyclic azomethine ylide (Scheme l-27). Evidence of the cyclic azomethine intermediate was illustrated through the addition of a second equivalent of the dipolarphile, which readily prompted a second [3+2] cycloaddition affording a new azabicyclo-[2,2,1]-heptane product.”66 Both the pyrroline and pyrrole products are believed to derive from the newly formed azomethine ylide intermediate. Protonation of the cyclic azomethine ylide either from a 1,2 prototropic shift (R2 = H) or an external proton source (R2 = alkyl) leads to the formation of A1-pyrrolines and Az-pyrrolines respectively (Scheme l-27). Subsequent oxidation of the resulting pyrroline species ultimately leads to the formation of aromatic pyrrole products. R F32 R4 1Y0 FMNR" R, N R1 R‘ R ,N 9 HM / 5 2 R3 R5 R3 R5 R3 \ R \ R Pym'” R4er'\/ 5 WM 5 I ' R, ' _ R, " R, R, R, I R 602 R, | R R, I R R, / R (:DN (5" (EN 5 N 5 0' ,N 5 R2 co R2 R2 _ R3 2 R3 4 Ra Ra Prime Azomethlne R2 = H R2 = Alkyl Cycload uct Yllde ”Win” Scheme l-27. Reaction of oxazol-5(4H)-ones with two equivalents of alkene to afford bicyclic heterocyclic scaffolds. Relatively few examples regarding the isolation of primary cycloaddition adducts from intermolecular cycloaddition reactions of miJnchnones have been reported. Padwa and co-workers previously disclosed the isolation of primary cycloaddition adducts from intramolecular cycloadditions of mi'inchnones with 35 terminal alkenes.18 Likewise, Maryanoff and co-workers described the isolation of a A1-pyrroline-5-carboxylic acid in the intermolecular cycloaddition of a miinchnone with 1,2-dicyanobutene (Scheme l-28).63 In each of the above cases, decarboxylation is presumably prevented due to the steric constraints of the cycloadduct products. These studies not only provided support for previously proposed reaction mechanisms, but provided the first indications that the cycloadditions of miinchnones with alkenes proceed with exo-stereochemistry. Me Yi" <9 9N / 0 H No H' NC CN ; Me N ", 002H Cl 50%kmbmd CI yew Scheme l-28. Maryanoff’s isolated primary cycloadduct. As stated earlier, it is thought that the relatively harsh reaction conditions traditionally employed in miinchnone cycloaddition reactions (refluxing in acetic anhydride) helps to facilitate the decarboxylation of the primary cycloadduct thus generating decarboxylated heterocyclic products. In 2001, Amdtsen and co-workers reported a method in which an imine, acid chloride and carbon monoxide could be coupled using palladium catalysis to generate mfinchnones under much milder reaction conditions.‘57 Exposure of the generated mi’inchnone species to a second equivalent of imine resulted in the isolation of carboxylate subsituted imidazolines (Scheme l-29). The authors attributed their ability to isolate the primary cycloadducts to their mildly acidic reaction conditions. Elimination of the HCI generated during 36 these reaction results predominately in the formation of undesired B-lactam byproducts, illustrating perhaps the active dipolarphile may indeed be the HCl salt of the imine substrate. These reactions were diastereoselective generating imidazoline products as single diastereomers with the stereochemistry illustrated in Scheme I-29 as determined by X-ray crystallography. O R /U\c co (1 atm) F 1 i R 1 l Pd2(dba)3 (5 mol%) 1 3 + bipy (10 mom) RW’O Ra‘N’ R2 R N H t | O t: \H/ R2 A R9 e e Scheme I-29. Arndtsen's synthesis of 2-imidazoline carboxylates. Recently, Lewis acids have been demostrated to increase the concentration miinchnone tautomers under much milder conditions allowing for the isolation of various dihydro—heterocyclic scaffolds. In 2004, we reported a silver acetate mediated cycloaddition of oxazoI-5(4H)-ones with electron deficient alkenes.53 These reactions stereoselectively proceeded in moderate to high yields affording A1-pyrrolines (Scheme l-30). The reactions also proceeded with exo-selectivity as seen in Maryanoff’s studies.53 An enantioselective version of this chemistry was later reported by Toste and co- workers in which the authors utilized cationic gold to catalyze the reaction.” In addition, we previously reported a silicon mediated 1,3-dipolar cycloaddition of oxazol-5(4H)-ones with imines to afford highly substituted imidazolines.52 The stereochemical outcome of the reaction is highly dependent on the 37 substitution pattern of the oxazol-5(4H)-one scaffold. The above methodology involving Lewis acid mediated cycloadditions of oxazol-5(4H)-ones with both imines and alkenes will be described in more detail in the following chapters of this dissertation. R3 R1YN83 R3 /N§/R4 R\“/O Rim/R3 R1 “4 ‘ "R?" = .l «R4 TMSCI AgOAc : 2002H R} COZH 60-90% yield 59-95% yield Scheme I-30. Lewis acid mediated cycloadditions of oxazol-5(4H)-ones. H. Current work Since their initial discovery, oxazol-5(4H)-ones have emerged as an important class of compounds for synthesizing biologically interesting targets. As demonstrated throughout this chapter, the oxazol-5(4H)—one scaffold contains numerous reactive sites allowing for a large diversity of transformations. The reseach presented during the remainder of this dissertation illustrates the potential of using oxazol-5(4H)-ones in diversity oriented synthesis drug discovery programs. The following chapters include both the development of new chemistry using oxazol-5(4H)-ones to rapidly create small libraries. of novel molecules, as well as the evaluation of the molecular libraries for biological function. The first project presented in this dissertation involves the synthesis and evaluation of small libraries of 2-imidazolines for their ability to inhibit NF- KB mediated gene transcription (Scheme l-31). Previously we reported that oxazol-5(4H)-ones undergo diastereoselective [3+2] cycloadditions with 38 imines while in the presence of Lewis acids to afford highly substituted 2- imidazolines.""'52 Furthermore, this class of imidazolines were found to be potent inhibitors of NF-KB mediated gene transcription.“ Due to the potential 44,69 we therapeutic value of inhibiting NF-KB mediated gene transcription, undertook a structure activity relationship study of this class of compounds with the hopes of optimizing the compounds as potential drug candidates. This project was done in collaboration with Dr. Daljinder Kahlon and Theresa Lansdell. The data obtained from the structure activity relationship study as well as the synthesis of the compounds is presented in detail in the following chapter. R5 R R1 0 . 1 0 OR R‘YN R5’NVR‘ HT 0 =/ 3 Y o .- R‘ R‘Z C02R3 R2 2 0R3 Inhibitors of NF—xB Key intermediates for mediated gene transcription the synthesis of proteasome inhibitors Scheme l-31. Research presented in this dissertation. The second research project presented in this disseration involves the development of a new method for the alkylation of oxazol-5(4H)-ones.16 We discovered oxazol-5(4H)-ones to undergo ene-type alkylation reactions with enol ethers to generate quaternary substituted oxazolones (Scheme l-31).16 These quaternary substituted oxazol-5(4H)-ones represent pivotal intermediates for synthesizing a variety of biologically interesting molecules including ten-alkyl amino hydroxy carboxylic acids. The last two chapters of 39 this dissertation describe our current findings on this new methodology including the scope and mechanism of the reaction. Furthermore, this dissertation also includes the use of Bronsted acid catalysis for improving the stereoselectivity of the reaction.70 Also included is the use of this chemistry in synthesizing a class of molecules known to be potent inhibitors of the 208 proteasome. 4O l. References (1) (2) (3) (4) (5) (6) (7) (8) (9) (10) (11) Ellman, J. A.; Desi n, synthesis, and evaluation of small-molecule libraries. Acc. Chem. es. 1996, 29, 132-143. Drews, J.; Drug discovery: A historical perspective. Science. 2000, 287, 1960-1964. Spring, D. R.; Diversity-oriented synthesis; a challenge for synthetic chemists. Org. Bio. Chem. 2003, 1, 3867-3870. (a) Burke, M. D.; Berger, E. M.; Schreiber, S. L.; Generating diverse skeletons of small molecules combinatorially. Science. 2003, 302, 613- 618; (b) Burke, M. D.; Berger, E. M.; Schreiber, S. L.; A synthesis strategy yielding skeletally diverse small molecules combinatorially. J. Am. Chem. Soc. 2004, 126, 14095-14104; (c) Burke, M. D.; Schreiber, S. L.; A planning strategy for diversity-oriented synthesis. Angew. Chem. Int. Ed. 2004, 43, 46-58; Schreiber, S. L.; Target—oriented and diversity-oriented organic synthesis in drug discovery. Science. 2000, 287, 1964-1969. Filler, R.; Roa, A. Y.; New Developments in the Chemistry of Oxazolones. Adv. Hetero. Chem. 1977, 21, 175. Carter, H. E.; Azlactones. Org. React. 1946, 3, 198-239. Mukerjee, A. K.; Azlactones - Retrospect and Prospect. Hetemcycles. 1967, 26, 1077-1097. Plochl; Ber. Chem. 1883, 16, 2815. (a) Erlenmeyer, E.; Ann. Chim. 1893, 275, 1; (b) Erlenmeyer, E.; Halsey; Ber. Chem. 1897, 30, 2981; (c) Erlenmeyer, E.; Halsey; Ann. Chim. 1899, 307, 138; (d) Erlenmeyer, E.; Kunlin, J.; Ann. Chim. 1901, 316, 145; (e) Erlenmeyer, E.; Kunlin, J.; A synthesis of the upsilon-naphtoacid and of the naphtalin. Ber. Chem. 1902, 35, 384-386; (0 Erlenmeyer, E.; Matter, 0.; The azlactones from the cinnimic aldehyde resp cuminol and hippuric acid. Ann.Chim. 1904, 337, 271-282; (g) Erlenmeyer, E.; Stadlin, W.; The azlactones from furfural and resp salicylic aldehydes and hippuric acids. Ann. Chim. 1904, 337, 283-293; (h) Erlenmeyer, E.; Wittenberg, F.; The condensation of m-Oxybebzene aldehydes resp anise aldehydes and hippuric acids arising from azlactones. Ann. Chim. 1904, 337, 294-301. Erlenmeyer, E.; Information on alpha-amido acids. Ber. Chem. 1900, 33, 2036-2041. (a) Mohr, E.; Announcement on lactone-similar anhydrids of acylated amino acids. Joumal Fur Praktische Chemie-Leipzig 1910. 82, 60—64; (b) Mohr, E.; Report on the lacto like anhydride acylised amino acid. Joumal Fur Praktische Chemie-Leipzig 1910. 81, 473-500; (c) Mohr, E.; Gels, T.; Benzglsamgto-iso butanoic acid-lactimones. Ber Dtsch Chem 693 1908. 41, 7 -79 . 41 (12) (13) (14) (15) (16) (17) (18) (19) (20) (21) Clarke, H. T.; Johnson, J. R.; Robinson, R., The Chemistry of Penicillin. in Oxazoles and Oxazolones, Princeton University Press, Princeton, NJ, 1949; 688-848. Anon ous; Definitive Rules for Nomenclature of Organic Chemistry. J. Am. hem. Soc. 1960, 82, 5545-5574. Georg, G. I.; Boge, T. C.; Cheruvallath, Z. S.; Harriman, G. C. B.; Hepperle, M; Park, H, Schotten-Baumann Acylation of N- -Debenzoyltaxol - an Efficient Route to N-Acyl Taxol Analogs and Their Biological Evaluation. Bioorg. Med. Chem. Lett. 1994, 4, 335-338. (a) Bergman, J.; Lidgren, 6.; Reaction of Tryptophan with Trifluoroacetic- Anhydride. Tetrahedron Lett. 1989, 30, 4597-4600; (b) Chen, F. M. F.; Kuroda, K.; Benoiton, N. L.; Simple Preparation of 5—Oxo—4,5—Dihydro-1,3- Oxazoles (Oxazolones). Synthesis. 1979, 230-232. Fisk, J. S.; Tepe, J. J.; Intermolecular Ene Reactions Utilizing Oxazolones and Enol Ethers. J. Am. Chem. Soc. 2007, 129, 3058-3059. Goodman, M.; Levine, L.; Peptide Synthesis VIa Active Esters . 4 . Racemization + Ring-Opening Reactions of Optically Active Oxazolones. J. Am. Chem. Soc. 1964, 86, 2918-2922. Padwa, A.; Pearson, W. H., Synthetic Applications of 1,3-Dipolar Cycloaddition Chemistry Toward Heterocycles and Natural Products. 1rst Ed. John Wiley and Sons, Hoboken, NJ, 2002, 682-747. Dejersey, J.; Willadse.P; Zemer, B.; Oxazolinone lnterrnediates in Hydrolysis of Activated N-Acylamino Acid Esters . Relevance of Oxazolinones to Mechanism of Action of Serine Proteinases. Biochemistry. 1969, 8, 1959. Kubel, B.; Gruber, P.; Humaus, R.; Steglich, W.; Reactions of Oxazolin-5- One Anions .8. Alpha-Substituted Alpha-Amino-Acids \fIa Alkylation of Oxazolin-5-Ones. Chem. Ber.l1979, 112, 128-137. (a) Obrecht, D.; Altorfer, M.; Lehmann, C.; Schonholzer, P.; Muller, K.; An efficient strategy to orthogonally protected (R)- and (S)-alpha- methyl(alkyl)serine-containing peptides via a novel azlactone/oxazoline interconversion reaction. J. Org. Chem. 1996, 61, 4080-4086; (b) Obrecht, D.; Bohdal, U.; Broger, C.; Bur, D.; Lehmann, C.; Ruffieux, R.; Schonholzer, P.; Spiegler, C.; Muller, K.; L-Phenylalanine Cyclohexylamide - a Simple and Convenient Auxiliary for the Synthesis of Optically Pure Alpha,AIpha-Disubstituted (R)-Amino and (S)-Amino Acids. Helv. Chim. Acta. 1995, 78, 563-580; (c) Obrecht, D.; Bohdal, U.; Ruffieux, R.; Muller, K.; A Reinvestigation of the Alpha-Alkylation of 4- Monosubstituted 2-Phenyloxazol-5(4H)-Ones (Azlactones) - a General Entry into Highly Functionalized Alpha, Alpha-Disubstituted Alpha-Amino- Acids. Helv. Chim. Acta. 1994, 77, 1423-1429; (d) Obrecht, D.; Spiegler, C.; Schonholzer, P.; Muller, K.; Heimgartner, H.; Stierli, F .; A New General App proach to Enantiomerically Pure Cyclic and Open-Chain (R)-Alpha, Apha-Disubstituted and (S)-Alpha, AIpha-Disubstituted Alpha-Amino- Acids. Helv. Chim. Acta. 1992, 75, 1666-1696. 42 (22) (23) (24) (25) (26) (27) (23) (29) (30) Cabrera, S.; Reyes, E.; Aleman, J.; Milelli, A.; Kobbelgaard, S.; Jorgensen, K. A.; Organocatalytic asymmetric synthesis of alpha, alpha- disubstituted alpha-amino acids and derivatives. J. Am. Chem. Soc. 2008, 130, 12031-12037. Steglich, W.; Hofle, 6.; Simple Synthesis of AcyI-Oxazolin-S-Ones from 5- Acyloxy-Oxazoles 2. lnforrnation on Hypemucleophilic Acylation Catalysts. Tetrahedron Lett. 1970, 4727-4730. Ruble, J. C.; Fu, G. C.; Enantioselective constmction of quaternary stereocenters: Rearrangements of O-acylated azlactones catal ed by a planar-chiral derivative of 4-(pyrrolidino)pyridine. J. Am. Chem. 00. 1998, 120, 11532-11533. (a) Nguyen, H. V.; Butler, D. C. D.; Richards, C. J.; A metallocene- pyrrolidinopyridine nucleophilic catalyst for asymmetric sgnthesis. Org. Lett. 2006, 8, 769-772; (b) Seitzberg, J. G.; Dissing, .; Sotofte, .; Norrby, P. O.; Johannsen, M.; Synthesis and application of a l- ferrocenyl(pseudo-biarylic) complexes. Part 5. Design and synthesis 0 a new type of ferrocene-based planar chiral DMAP analogues. A new catalyst system for asymmetric nucleophilic catalysis. J. Org. Chem. 2005, 70, 8332-8337; (c) Shaw, S. A.; Aleman, P.; Christy, J.; Kampf, J. W.; Va, P.; Vedejs, E.; Enantioselective TADMAP-catalyzed carboxyl migration reactions for the synthesis of stereogenic quaternary carbon. J. Am. Chem. Soc. 2006, 128, 925-934; (d) Shaw, S. A.; Aleman, P.; Vedejs, E.; Development of chiral nucleophilic pyridine catalysts: Applications in aggg‘eggrgguatemary carbon synthesis. J. Am. Chem. Soc. 2003. 125, Fu, G. C.; Enantioselective nucleophilic catalysis with "planar-chiral" heterocycles. Acc. Chem. Res. 2000, 33, 412-420. Trost, B. M.; Ariza, X.; Enantioselective allylations of azlactones with ugsgmmetrical acyclic allyl esters. J. Am. Chem. Soc. 1999, 121, 10727- 7 . gt) Kawatsura, M.; ldeda, D.; Tamiko, l.; Komatsu, Y.; Uenishi, J.; alladium-Catalyzed Regio-and Diastereoselective Allylic Alkylation with Azlactones Using Triphenylarsine. Synlett. 2006, 15, 2435-2438; (b) Trost, B. M.; Ariza, X.; Catalytic asymmetric alkylation of nucleophiles: Asymmetric synthesis of alpha-alkylated amino acids. Angew. Chem. Int. Ed. 1997, 36, 2635-2637; (c) Trost, B. M.; Dogra, K.; Synthesis of novel quaternary amino acids using molybdenum-catalyzed asymmetric allylic alkylation. J. Am. Chem. Soc. 2002. 124, 7256-7257. Trost, B. M.; Jakel, C.; Plietker, B.; Palladium-catalyzed asymmetric addition of pronucleophiles to allenes. J. Am. Chem. Soc. 2003, 125, 4438-4439. Liu, X. X.; Hartwig, J. F.; Palladium-catalyzed alpha-arylation of azlactones to form quaternary amino acid derivatives. Org. Lett. 2003, 5, 1915-1918. 43 (31) (32) (33) (34) (35) (36) (37) (38) (39) (40) (a) Morgan, J.; Pinhey, J. T.; Reaction of Or anolead Triaoetates with 4- EthoxycarbonyI-2-Methyloxazol-5-One - the ynthesis of Alpha-Aryl and Alpha-Vinyl N-Acetylglycine Ethyl-Esters and Their Enzymatic Resolution. Tetrahedron Lett. 1994, 35, 9625-9628; (b) Morgan, J.; Pinhey, J. T.; Sherry, C. J.; Reaction of organolead triacetates with 4-ethoxycarbonyI-2- methyl-4,5-dihydro-1,3—oxazoI-5-one. The synthesis of alpha-aryl— and alpha-vinyl—N-acetylgl cines and their ethyl esters and their enzymic resolution. J. Chem. 00. Perkin Tran. 1. 1997, 813—619. ggi/ssier, H.; Dynamic kinetic resolution. Tetrahedron. 2003, 59, 8291- (a) Brown, S. A.; Parker, M. C.; Turner, N. J.; Dynamic kinetic resolution: synthesis of optically active alpha-amino acid derivatives. Tetrahedron: Asymm. 2000, 11, 1687-1690; (b) Crich, J. 2.; Brieva, R.; Marquart, P.; Gu, R. L.; Flemming, S.; Sih, C. J.; Enzymatic Asymmetric-Synthesis of Alpha-Amino-Acids - Enantioselective Cleavage of 4-Substituted Oxazolin- 5-ones and Thiazolin-5-ones. J. Org. Chem. 1993, 58, 3252-3258; (c) Daffe, V.; Fastrez, J.; Enantiomeric Enrichment in the Hydrolysis of Oxazolones Catalyzed b Cyclodextrins or Proteolytic-Enzymes. J. Am. Chem. Soc. 1980, 102, 3 01-3605. Berkessel, A.; Mukherjee, S.; Cleemann, F.; Muller, T. N.; Lex, J.; Second- generation organocatalysts for the highly enantioselective dynamic kinetic resolution of azlactones. Chem. Comm. 2005, 1898-1900. Lian , J.; Ruble, J. C.; Fu, G. 0.; Dynamic kinetic resolutions catalyzed by a p anar—chiral derivative of DMAP: Enantioselective synthesis of protected alpha-amino acids from racemic azlactones. J. Org. Chem. 1998, 63, 3154-3155. Keni, M.; Tepe, J. J.; One-pot Friedel-CraftsIRobinson-Gabriel synthesis of2 1ograzoles using oxazolone templates. J. Org. Chem. 2005, 70, 4211- 4 . Hupp, C. D.; Tepe, J. J.; Total synthesis of a marine alkaloid from the tunicate Dendrodoa grossularia. Org. Lett. 2008, 10, 3737-3739. a) Castelhano, A. L.; Home, 8.; Taylor, G. J.; Billedeau, R.; Krantz, A.; ynthesis of Alpha-Amino-Acids with Beta,Gamma-Unsaturated Side- Chains. Tetrahedron. 1988, 44, 5451-5466; (b) Holladay, M. W.; Nadzan, A. M.; Synthesis of Alpha-Benzyl Gamma—Lactam, Alpha-Benzyl Delta— Lactam, and Alpha-Benzylproline Derivatives as Confonnationally Restricted Analogs of Phenylalaninamide. J. Org. Chem. 1991, 56, 3900- 3905. HUpp, C.; Tepe, J. J.; 1-EthyI-3-(3-dimethylaminopropyl)carbodiimide Hydrochloride-Mediated Oxazole Rearrangement: Gaining Access to a Unique Marine Alkaloid Scaffold. J. Org. Chem. 2009, 74, 3406-3413. Clerici, F.; Gelmi, M. L.; Trimarco, P.; 5(4H)-oxazolones. Part XI. Cycloaddition reaction of oxazolones and munchnones to triphenylvinylphosphonium salts as synthetic equivalents of alkynes. Tetrahedron. 1998, 54, 5763-5774. 44 (41) (42) (43) (44) (45) (45) (47) (43) (49) (50) (51) (52) Fisk, J. S.; Mosey, R. A.; Tepe, J. J.; The diverse chemistry of oxazoI-5- (4H)—ones. Chem. Soc. Rev. 2007, 36, 1432-1440. Fu, N. Y.; Allen, A. D.; Kobayashi, S.; TidwelI, T. T.; Vukovic, S.; Arumugam, S.; Popik, V. V.; Mishima, M.; Amino substituted bisketenes: Generation, structure, and reactivity. J. Org. Chem. 2007, 72, 1951-1956. Coldham, |.; Hufton, R.; Intramolecular dipolar cycloaddition reactions of azomethine ylides. Chem. Rev. 2005, 105, 2765-2809. Pandey, G.; Banerjee, P.; Gadre, S. R.; Construction of enantiopure pyrrolidine ring system via asymmetric [3+2] cycloaddition of azomethine ylides. Chem. Rev. 2006, 106, 4484-4517. Huisgen, R.; Funke, E.; Schaefer, F. C.; Knorr, R.; Possible Valence Tautomerism of a Mesoionic Oxazol-5-One with an Acylaminoketene. Angew. Chem. Int. Ed. 1967, 6, 367. Huisgen, R.; GotthardH; Bayer, H. O.; Schaefer, F. C.; 1,3—Dipolar Cycloadditions .58. A Convenient Synthesis of N-Substituted Pyrroles from Mesoionic Oxazolones and Alkynes. Chem. Ber. 1970, 103, 2611. Huisgen, R.; Funke, E.; 1,3—C Ioadditions of Mesoionic Oxazolones to Carbonyl Compounds. Angew. hem. Int. Ed. 1967, 6, 365. (a) Dhawan, R.; Amdtsen, B. A.; Palladium-catalyzed multicomponent coupling of alkynes, imines, and acid chlorides: A direct and modular approach to pyrrole synthesis. J. Am. Chem. Soc. 2004, 126, 468-469; (b) St. Cyr, D. J.; Martin, N.; Amdtsen, B. A.; Direct synthesis of pyrroles from imines, alkynes, and acid chlorides: An isocyanide-mediated reaction. Org. Lett. 2007, 9, 449-452. (a) Siamaki, A. R.; Amdtsen, B. A.; A direct, one step synthesis of imidazoles from imines and acid chlorides: A palladium catalyzed multicomponent coupling approach. J. Am. Chem. Soc. 2006, 128, 6050- 6051; (b) Siamaki, A. R.; Black, D. A.; Amdtsen, B. A.; Palladium- catalyzed carbonylative cross-coupling with imines: A multicomponent synthesis of imidazolones. J. Org. Chem. 2008, 73, 1135-1138. Merlic, C. A.; Baur, A.; Aldrich, C. C.; Acylamino chromium carbene complexes: Direct carbonyl insertion, formation of munchnones, and trapping with dipolarophiles. J. Am. Chem. Soc. 2000, 122, 7398-7399. (a) Peddibhotla, S.; Jayakumar, S.; Tepe, J. J.; Highly diastereoselective multicomponent synthesis of unsymmetrical imidazolines. Org. Lett. 2002, 4, 3533-3535; (b) Shanna, V.; Tepe, J. I.; Diastereochemical diversity of imidazoline scaffolds via substrate controlled TMSCI mediated cycloaddition of azlactones. Org. Lett. 2005, 7, 5091-5094. Peddibhotla, S.; Tepe, J. J.; Multicomponent synthesis of highly substituted imidazolines via a silicon mediated 1,3-deolar cycloaddition. Synthesis. 2003, 1433-1440. 45 (53) (54) (55) (55) (57) (58) (59) (50) (51) (62) (63) (64) (55) Peddibhotla, S.; Tepe, J. J.; Stereoselective synthesis of highly substituted Delta(1)-pyrrolines: exo-selective 1,3-dipolar cycloaddition reactions with azlactones. J. Am. Chem. Soc. 2004, 126, 12776-12777. Cossio, F. P.; Ugalde, J. M.; Lopez, X.; Lecea, B.; Palomo, C.; A Semiempirical Theoretical-Study on the Formation of Beta-Lactams from Ketenes and Imines. J. Am. Chem. Soc. 1993, 115, 995-1004. Funke, E.; Huisgen, R.; Ketenoid Reactivity of a Mesoionic Oxazol-5-One. Chem. Ber. 1971, 104, 3222. Cremonesi, G.; DaIIa Croce, P.; La Rosa, 0.; [2+2] Cycloaddition reactions of imines with cyclic ketenes: Synthesis of 1,3-thiazolidine derived spiro- beta-lactams and their transformations. Helv. Chim. Acta. 2005, 88, 1580- 588. Dhawan, R.; Dghaym, R. D.; St Cyr, D. J.; Amdtsen, B. A.; Direct, palladium-catalyzed, multicomponent synthesis of beta-Iactams from iggfiggs, acid chloride, and carbon monoxide. Org. Lett. 2006, 8, 3927- Hershenson, F. M.; Pavia, M. R.; Synthesis of N-Substituted Pyrroles from Azlactones Via 1,3-Oxazolium 5-Oxides. Synthesis. 1988, 999-1001. Okano, T.; Uekawa, T.; Morishima, N.; Eguchi, 8.; Synthesis of Beta- (T rifluoromethyl)Pyrroles We the Cycloaddition of Munchnones to Eéecgrog-Ggeficient Trifluoromethylated Olefins. J. Org. Chem. 1991, 56, 5 5 -5 . Brunn, E.; Funke, E.; Gotthard.H; Huisgen, R.; 1.3-Dipolar Cycloadditions .61. Cycloadditions of N-Substituted Oxazolium-5-Olates to Nitriles, Nitro, Nitroso and A20 Compounds. Chem. Ber. 1971, 104, 1562. Consonni, R.; Croce, P. D.; Ferraccioli, R.; Larosa, C.; A New Approach to Imidazole Derivatives. J. Chem. Res. 1991, 188-189. Bilodeau, M. T.; Cunningham, A. M.; Solid-supported synthesis of imidazoles: A strategy for direct resin-attachment to the imidazole core. J. Org. Chem. 1998, 63, 2800-2801. Maryanoff, C. A.; Karash, C. B.; Turchi, I. J.; Corey, E. R.; Maryanoff, B. E.; Characterization of a Stable Carboxylic-Acid lnterrnediate from 1,3- Dipolar Cyclo-Addition of a Munchnone with 1,2-Dicyanocyclobutene. J. Org. Chem. 1989, 54, 3790-3792. Maryanoff, C. A.; Turchi, l. J.; Mechanism and Stereochemical Implications of the Reaction of an Oxazolium-S-Oxide with 1,2- Dicyanocyclobutene - an Am1 Study. Heterocycles. 1993, 35, 649-657. Melhado, A. D.; Luparia, M.; Toste, F. D.; Au(I)-catalyzed enantioselective 1,3-dipolar cycloadditions of Munchnones with electron-deficient Alkenes. J. Am. Chem. Soc. 2007, 129, 12638. 46 (55) (67) (68) (69) (70) Gotthardt, H.; Huisgen, R.; Schaefer, F. C.; Delta-2-Pyrroline Aus Mesoionischen Oxazolen Und Olefinen. Tetrahedron Lett. 1964, 487-491. Dghaym, R. D.; Dhawan, R.; Amdtsen, B. A.; The use of carbon monoxide and imines as peptide derivative synthons: A facile palladium-catalyzed synthesis of alpha-amino acid derived imidazolines. Angew. Chem. Int. Ed. 2001, 40, 3228. (a) Sharma, V.; Hupp, C. D.; Tepe, J. J.; Enhancement of chemotherapeutic efficacy by small molecule inhibition of NF- kappaB and checkpoint kinases. Curr.. Med. Chem. 2007, 14, 1061-74; (b) Shanna, V.; Peddibhotla, S.; Tepe, J. J.; Sensitization of Cancer Cells to DNA 9D1angaging Agents by lmidazolines. J. Am. Chem. Soc. 2006, 128, 9137- 4 . Kan'n, M.; Yamamoto, Y.; Wang, O. M.; The lKK NF-kappa B system: a treasure trove for drug development. Nat. Rev. Drug Discov. 2004, 3, 17- 26. Mosey, R. A.; Fisk, J. S.; Friebe, T. L.; Tepe, J. J.; Synthesis of tert-Alkyl amino hydroxy carboxylic esters via an intermolecular ene—type reaction of oxazolones and enol ethers. Org. Lett. 2008. 10, 825-828. 47 CHAPTER II INHIBITION OF NF-KB MEDIATED GENE TRANSCRIPTION BY 2- IMIDAZOLINES DERIVED FROM OXAZOL-5(4H)-ONES A. Synthesis of 2-imidazolines via miinchnonelimine cycloadditions. The imidazoline scaffold is found in a variety of molecules exhibiting biologically interesting properties.‘ lmidazoline derivatives have been illustrated to exhibit a wide range of biological activity including anti-inflammatory, anti-nociceptive, immuno-modulating, antioxidant activities and many more.2'3 Furthermore, molecules containing imidazoline cores have been shown to possess anti-cancer relevant properties.‘°""5 In addition to exhibiting a wide range of biological activity, imidazolines have also proven to be useful substrates in synthetic organic chemistry. Imidazolines are convenient building blocks for the synthesis of a variety of biologically interesting molecules including azapenams, dioxocyclams, and diazapinones.6 They also have been illustrated to be valuable substrates in asymmetric catalysis both as chiral catalysts and as chiral auxiliaries. Enantiopure imidazolines serve as useful intermediates for the synthesis of various chiral ligands including 1,2-diamines.7 Additionally, imidazolines have served as precursors to chiral catalysts such as N-heterocyclic carbenes."'9 As a result of their biological and chemical significance, new methods for synthesizing highly substituted 2-imidazolines are still of high interest. Traditional 48 methods for their preparation include the condensation of 1,2-diamines to construct aminals, which are subsequently reduced to afford the desired imidazolines.10 Interestingly, relatively few examples have been reported using oxazol-5(4H)-ones to synthesize 2-imidazolines."‘13 Cycloadditions of imines with the 1,3-mesoionic tautomer of oxazol-5(4H)-ones, milnchnones, present researchers with an ideal opportunity to synthesize 2-imidazolines with high levels of structural diversity.9'14 Unfortunately, cycloaddition reactions of mIJnchnones with imines are generally plagued by the formation of undesired side products.9'“'16 Highly substituted B-lactams are often produced in high yields upon heating oxazol-5(4H)-ones in the presence of imines.17 Previous studies have indicated that B—lactams are produced from oxazol-5(4H)-ones via Staudinger-like [2+2] cycloadditions of their amidoketene isomer (Scheme "-1).18 Investigations by Huisgen and co-workers established that N-alkylation of munchnones increases the concentration of the mesoionic tautomer required to facilitate the desired [3+2] cycloaddition. To this end, Consonni and coworkers successfully illustrated that N-alkyl mtinchnones undergo [3+2] cycloadditions with N-(phenylmethylene)benzenesuIphonamides to afford fully substituted imidazoles (Scheme "-1).19 The authors propose that aromatization occurs through the decarboxylation of the intermediate cycloadduct followed by expulsion of benzenesulphinic acid. 49 R1 0 e R J / O KAN’SOZPh 1 ’N Rz/N ; /N\/e—R4 3 [3+2] R2 R3 Milnehnone O O A R5 0 'R“ x .. ~' 1 :2 R, N R3 ' é [2+2] R1 w R3 R5 2 R2 Amldoketene Scheme "-1. Traditional cycloadditions of miinchnoneslamidoketenes with imines. Recently, we pioneered the use of Lewis acids as mild and effective reagents for promoting [3+2] cycloaddition reactions of oxazol-5(4H)- ‘2"3'2° Lewis acids coordinate to oxazol-5(4H)-ones increasing the ones. equilibrium concentration of their mesoionic milnchnone tautomers. The relatively mild conditions at which the munchnones are generated prevent the decarboxylation of primary cycloadducts, thus allowing for the isolation of dihydro-heterocycles such as 2-imidazolines. We demonstrated that imidazolines with four point diversity can be diastereoselectively produced via a silicon mediated 1,3-dipolarcycloadditions of oxazoI-5(4H)-ones with imines (Scheme "-2).13 OxazoI-5(4H)-ones were treated with TMSCI while in the presence of imines affording highly substituted 2-imidazoline products. 50 _ e' .. _ R - R’NVR4 eCI H4129 R4 eCI R1 0 R /N~\~/R4 ?/ R Y O 3 > R1YO 1 O TMSCI (1 eq) (9,} 0 R2 20'2 ms’ TMS’ reflux R2 R2 _ R3 _ ‘Y" ‘Y“ R NLZMR‘ or lez—R‘ 1%0 _ ’H 3' N \- R2 cozH R2 COZH TMS, R 69 Syn-lmidazoline. Anti-lmidazoline. __ 2 _ 60-90% yields Scheme "-2. Lewis acid mediated synthesis of imidazolines. Interestingly, varying the substitution pattern of the oxazoI-5(4H)-one precursors allows for the selective generation of either anti-imidazolines or syn-imidazolines (with respect to the R2 and R4 substituents, Scheme "-2). The diastereoselectivity of these reactions appears to be the result of a combination of steric and electronic factors. The reaction of 2-aryloxazol- 5(4H)-ones with imines results predominantly in the formation of anti- imidazolines (Figure "-1). The high diastereoselectivity is attributed to a steric interaction between the bulky silyl moiety and the R4 substituent of the imine causing a preferential endo approach of the imine.13 Complete reversal of diastereoselectivity was observed utilizing 2-alkyl-4-aronxazoI-5(4H)-ones and imines substituted with aryl R4 substituents.21 The syn diastereoselectivity appears to result from either n-stacking or edge to face interactions between 51 the aryl R4 and R2 substituents. An increase in syn selectivity was noted when using rr-donating R2 groups and tr-accepting R4 groups, supporting the rationale for reversal of diastereoselectivity. Little or no diastereoselectivity is observed in reactions involving 2,4—diaIkyloxazoI-5(4H)-ones with imines. R3 R3’N r IMR4 1N ‘ “\H r \'(:0\r/O r, O\:/O vs AT‘K \(2'6 49 ”\ng ‘SihéD R2 Endo-Favored Exo-Disfavored Figure "-1. Origin of diastereoselectivity using 2-aronxazol-5(4H)—ones. B. 2-imidazolines as inhibitors of NF-xB mediated gene transcription. NF-KB is a mammalian transcription factor responsible for the regulation of more than 150 genes?"23 The inhibition of the NF-KB signaling pathway has been a focus of intense academic and industrial research as a target for development novel pharmaceuticals.24 Since NF-KB is a pivotal regulator of a number of genes associated with immune, inflammatory and anti-apoptotic responses, it is recognized as an attractive target for controlling various disease states such as cancer and arthritis."""'26 A variety of small molecule inhibitors of NF-KB mediated gene transcription have been developed, some of which have gone through or are currently undergoing clinical trials (Figure "-3). Perhaps the most common method for impeding NF-KB mediated gene transcription is via the inhibition of the proteolytic activity of the 268 proteasome.27 The most successful example of a 268 proteasome inhibitor is Bortezomib (also known as Velkade or PS- 52 341).”29 Currently, Bortezomib is clinically used as a treatment of multiple myeloma.29'3° Other known 26$ proteasome inhibitors include MG-132,31 N- acetyI-leucinyl-Ieucinyl-norleucinal (ALLN)32 and the natural products Lactacystin33 and Salinosporamide A.34 Other small molecule inhibitors of NF-KB mediated gene transcription include general kinase inhibitors (e.g. hymenialdisines),35'37 specific lKK inhibitors (e.g. PS-1145 and EMS-345541), inhibitors of I-KB ubiquitination (e.g. Ro106-9920)38 and many more. M64 32 ALLN Bortezomib Figure "-2. Small molecule inhibitors of NF-KB mediated gene transcription. In 2002, we reported the oxazoI-5(4H)-one derived imidazoline "-4 to be a potent inhibitor of NF-KB mediated gene transcription via the inhibition of I-KB degradation (Scheme "-3).”39 lmidazoline ll-4 inhibited NF-KB mediated gene transcription in human cervical epithelial (HeLa) cells activated by TNF-a with an ECso = 0.95 (M. In addition, imidazoline "-4 was found not to induce apoptosis as a single agent, but did increase the efficacy of several chemotherapeutic reagents (e.g. camptothecin and cis-platin) illustrating its potential value in the 53 treatment of cancer. The synthesis of imidazoline "-4 is outlined in Scheme "-3 below.'2'13 Cyclodehydration of N-benzoyl phenyl glycine with TFAA afforded oxazoI—5(4H)-one "-2 in 92% yield. OxazoI-5(4H)-one "-2 was subsequently treated with TMSCI while in the presence of N-benzylidene-1- phenylmethanamine to afford imidazoline "-3 as the hydrochloride salt. Treatment of imidazoline "-3 with saturated sodium bicarbonate afforded imidazoline "-4 as a white crystalline solid. Ph 0 f2H >PhYO Ph V N V Ph 6)th >Ph Pit/[Ltd| Ph 9235;. ZYQA’L Id Nfo TMSCI CIC+DN '9 CHZCIz H reflux P“ CO?” ",1 ".2 68% Yield ".3 Sat NaH003 Solution F‘h> PI‘I \j/ N I N jz-Pn Ph“ c0211 "-4 Scheme "-3. Synthesis of the NF-KB inhibitor imidazoline "-4. C. NF-kB mediated gene transcription As stated earlier, NF-KB is a transcription factor responsible for the regulation of a wide variety of genes.”23 Genes regulated by NF-KB include those related to 41 42 stress,"0 inflammatory stimuli, activation of immune cell function, cellular proliferation, apoptosis,43 and oncogenesis.“ Specific proteins regulated by NF- KB include various cytokines (IL-1, lL-2, TNF-o, and IL-6), chemokines (IL-8 and 54 RANTES), cell adhesion molecules (ICAM 1, VCAM-1, and E-selectin), growth factors, cyclin D1, cyclooxygenase (COX-2), matrix metalloproteinase (MMP—9) as well as many others.4548 The misregulation of NF-KB mediated gene transcription is associated with a variety of diseases including rheumatoid arthritis?“ inflammatory bowel disease,50 and cancer.51 For example, the deregulation of TNF-a expression has been related to many diseases including Crohn’s disease, rheumatoid arthritis (RA), multiple sclerosis and Alzheimer’s disease.52 Increased levels of TNF-a, IL-6 and IL-18 have been found in primary fibroblast-like synoviocytes from patients with rheumatoid arthritis and osteoarthritis.2‘"“'53 Furthermore, the levels of NF-KB-DNA binding are much greater in patients with rheumatoid arthritis, which is consistent with the observed increased levels of pro-inflammatory cytokine production.“ Structurally, NF-KB is a multi-subunit complex consisting of various members of the Rel family of transcription factors including NF-KB1 (p105/p50), NF-KB2 (p100/p52), ReIA (p65), ReIB, and c-Rel.2"“'55 These subunits can exist as a variety of heterodimers and homodimers and are used to control the selectivity of certain DNA control elements”:56 In most unstimulated mammalian cells, NF-KB exists primarily as a p50lp50 homodimer or as a p50/p65 heterodimer. While in its inactive form, NF—KB exists in the cytoplasm as a complex with its inhibitory protein, I-KB (Figure "-3). The NF-KB signaling pathway is initiated by a variety of extracellular stimuli including antineoplastic agents, viruses, phorbol esters, oxidative stress, and cytokines (e.g. TNF-a and IL-18).57 Upon activation of the NF-KB pathway, lKK kinases phosphorylate I-KB 55 on serines 32 and 36.58 This is followed by subsequent ubiquitinylation and degradation of I-KB by the 26S proteosome.“47 Upon proteolytic degradation of l-KB, NF-KB is released allowing for its translocation into the nucleus.59 Once inside the nucleus, it binds to various DNA control elements thus initiating gene transcription.“'°° 1;? Chemotheroputic $3 lKK «a.» >_. 4%" Kincses M, . % Serine . . ""65 43% Kinose TranscnptIon (TNF—a, lL—IB) g K. m Ubiquifinilyation . figgiiiilitiiitiii e443 V Proteosome ”(3 (agree ' Figure "-3. Mechanistic activation of the transcription factor NF-KB. D. Development of a new class of 2-imidazoline based NF-KB inhibitors The inhibition of NF-KB mediated gene transcription is recognized as an attractive method for potentially treating a wide variety of disease states.61 To this end, the development of novel small molecule regulators of NF-KB mediated gene transcription is of great value. As previously mentioned, prior studies in our research group found imidazoline "-4 to be a relatively potent inhibitor of NF-KB mediated gene transcription.”39 In addition, imidazoline “-4 represents a new 56 class of molecule for the inhibition of NF-KB signaling pathways. As a part of our diversity oriented synthesis research program aimed at the development of new biologically interesting compounds, we set out to optimize imidazoline "-4 for its ability to inhibit the NF-KB mediated gene transcription. While imidazoline "-4 was demonstrated to be a potent inhibitor of NF-KB mediated gene transcription, questions regarding the structural integrity of the compound made imidazoline "-4 an unlikely candidate for pharmaceutical use. lmidazoline "-4 is a primary cycloadduct obtained from a [3+2] cycloaddition reaction of a miinchnone with an imine. Early studies regarding mtinchnone cycloaddition chemistry illustrated that the primary cycloaddition adducts generated from these reactions readily decarboxylate and aromatize to afford a cts.‘5"°’7'62 In comparison to earlier methods variety of hetero-aromatic produ (refluxing in acetic anhydride), the Lewis acid mediated cycloaddition used to synthesize imidazoline "-4 was conducted under relatively milder 12.13.15.2o.37.62 The mild reaction conditions (reflux in dichloromethane) conditions. utilized in the synthesis is more than likely responsible for preventing the decarboxylation/aromatization of the product imidazoline "-4. Although new reaction methodology allowed for the synthesis and isolation of imidazoline "-4, the tendency of this class of primary cycloadducts to decarboxylate led us to investigate the structural stability of the compound. Exposure of imidazoline "-4 to slightly elevated temperatures did in fact lead to decarboxylation and generation of imidazolines "-5 and "-6 and imidazole "-7 (Scheme "-4). The decarboxylation of imidazoline "-4 is believed to take place 57 as outlined in Scheme "-4.63 Upon being exposed to slightly elevated temperatures, imidazoline II-4 decarboxylates generating an azomethine ylide intermediate. The azomethine ylide intermediate then undergoes a 1,2- prototopic shift affording imidazolines "-5 and "-6. While imidazole "-7 is likely produced from the oxidation of both imidazolines "-5 and "-6,64 only imidazoline "-6 has been demonstrated to undergo aromatization in our laboratories. However, it should be noted that 4,5-diaryl 2-imidazolines, such as imidazolines “-5 and "-6, have previously been demonstrated to exist in equilibrium under a variety of conditions“)66 This leads to the possibility that imidazoline "-5 may potentially aromatize to imidazole "-7 by equilibrating to imidazoline "-6 followed by subsequent aromatization. Ph V Ph _ 7 rah> N a N WNI/ Ph Ply/N Ph THF 6911/ Ph reflux H’ 9 Ph COZH HPh C02 ph II-4 — _ Ph /\ PhW/EZg-Pn <—— ”Hie—P" -——= PhY‘Z—Ph "-7 "-53.1 Ph Scheme "-4. Decarboxylation of imidazoline “-4. The propensity of imidazoline "-4 to decarboxylate under relatively mild reaction conditions prompted us to question if imidazoline "-4 was the molecule 58 responsible for inhibiting NF-KB signaling or if it was merely a precursor to the actual reactive species.9 To help answer this question, degradation products II- 5, "-6, and "-7 were isolated and evaluated for their ability to inhibit NF-KB mediated gene transcription (Table "-1). Compounds "-5, "-6, and "-7 were evaluated for their ability to inhibit NF-KB mediated gene transcription in HeLa cells activated by TNF-a using a Iuciferase-based reporter assay (See experimental section). In addition, the three compounds were further evaluated for their ability to inhibit lL-6 production in human whole blood stimulated by IL- 18. Both assays illustrated that all three degradation products were less active than their precursor, imidazoline "-4, for inhibiting NF-KB mediated gene transcription. Of the degradation products, the trans-aryl 2-imidazoline ll-5 illustrated the best inhibitory properties inhibiting NF-KB mediated luciferin production with an ECso value of 4.6 (M and lL-6 production with an leo value equal to 2.5 (M (T able "-1). The cis-aryl 2-imidazoline “-6 was found to be slightly less effective in the Iuciferase reporter assay affording an E050 value of 11.0 (M, while interestingly provided very similar results for inhibiting lL-6 production with an leo equal to 2.4 pM (Table "-1). The fully aromatized imidazole "-7 exhibited no ability to inhibit NF-KB mediated gene transcription in the luciferase based reporter assay, and thus was not analyzed using the human whole blood assay (Table "-1). These results seem to indicate that parent molecule imidazoline "-4 is most likely the molecular species responsible for the nanomolar activity observed in our previous studies, although current studies are still ongoing to further validate this conclusion. 59 Compound Commund Inhibition of Inhibition of IL-6 Number Structure HoLgcliz-(tB'lLuc In ”13:81:?“ Ph '8" "-4 FIE—Ph 0.95 >20 Ian“c COZH Bn Ph ,3 11.5 112—9h 4.6 2.5 i=n Bn Ph N "-6 \INI/ Ph 1 1.0 2.4 Ph "-7 11:24:" >20 — Ph Table "-1. Evaluation of the degradation products produced from imidazoline "-4 for their ability to inhibit NF-KB signaling pathways. Simultaneous to the structural integrity studies regarding imidazoline “-4, other members of our research group focused on its derivatization with aspirations of discovering a compound illustrating increased stability while maintaining a similar biological profile. To this end, Dr. Daljinder Kahlon synthesized imidazoline derivatives "-8, "-9, and "-10 starting from imidazoline "-4.9 Esterifimtion of imidazoline "-4 using TMSCHN2 followed by subsequent reduction with LiAlH4 afforded alcohol substituted imidazoline "-8. Primary amide substituted imidazoline "-9 was synthesized via an EDCI mediated coupling of 60 imidazoline "-4 with (NH4)2COa. Finally, imidazoline "-4 was treated with (COCI)2 producing an acid chloride intermediate, which was subsequently treated with ethanol affording the ethyl ester substituted imidazoline lI-10. Compounds "-8, "-9, and "-10 were next evaluated for their ability to inhibit NF-KB mediated gene transcription in HeLa cells using the luciferase based reporter assay. Both imidazolines "-8 and "-9 were devoid of any ability to inhibit NF-KB mediated Iuciferase production in HeLa cells, while imidazoline "-10 illustrated relatively good activity with an ECso value of 2.5 uM. Ian> PhYN I N_ P" £050=>2onM Ph‘“ NH2 o "-9 EDCI, HOBt DIPEA, (NH4)2003 THF Ian> Ph 1. TMSCHN2 Y" Ph 4 Benzene/MeOH PhYN Ph 1. (00002 = PhYN Ph Ph THF IHCOZH 2.30.1 Ph“. COzEt "-8 "-10 ECso=>20 uM E050=2.5p.M Scheme "-5. Derivatization of imidazoline "-4. Upon discovering the ethyl ester substituted imidazoline “-10 to be a potent inhibitor of NF-KB mediated gene transcription, Dr. Kahlon and Dr. Daniel Jones further evaluated imidazoline "-10 for its structural stability.9 Small molecules containing ester moieties are often susceptible to hydrolysis by blood 61 and cellular esterases. To this end, imidazoline "-10 was evaluated for its ability to resist hydrolysis in human whole blood to ensure that imidazoline "-10 was in fact the active compound and not just a pro-drug of imidazoline II-4. Incubation of imidazoline "-10 in human whole blood for 24 hours at 37°C afforded no degradation of the compound as determined by LC/MS analysis providing plausible evidence that imidazoline "-10 was not a pro-drug for imidazoline "-4. E. Structure activity relationship study of trans-24midazolines Upon discovering imidazoline "-10 to be a relatively potent inhibitor of NF-KB mediated gene transcription, we next sought to structurally optimize the molecule for its ability to inhibit NF-KB mediated gene transcription.‘37 Structural optimization of this novel class of molecules would not only possibly lead to more effective inhibitors of NF-KB signaling, but may also provide further insight into their inhibitory mechanism. In a collaborative effort to structurally optimize these compounds for their ability to inhibit NF-KB mediated gene transcription, Dr. Daljinder Kahlon and I synthesized a variety of analogues of imidazoline "-10. We systematically synthesized a series of analogues of "-10, varying four different positions of the imidazoline scaffold (Figure "-4, R1 - R4). The compounds were synthesized via Lewis acid mediated 1,3-dipolar cycloaddition reactions of oxazol-5(4H)-ones with imines as described previously.‘2"3'21 62 PhY\2—Ph l:> “Rig—R3 Ph 002151 "-10 Figure "-4. Structure activity relationship study of imidazoline “-10. The imidazoline analogues were then evaluated for their ability to inhibit NF-KB mediated gene transcription using a Iuciferase based reporter assay in human cervical epithelial (HeLa) cells with a stably transfected NF-KB-IUC gene. The cells were treated in the presence or absence various concentrations of the imidazolines. The proteasome inhibitor MG-132 and DMSO (vehicle) were used as positive and negative controls, respectively. Cells were pretreated for 30 minutes with the imidazolines/controls followed by TNF-d stimulation. Treatment with the cytokine TNF-a, initiated the NF-KB signaling pathway, thus leading to the degradation of the inhibitory protein '46 (Figure "-5). Upon being released into the cytoplasm, NF-KB was then translocated into the nucleus where it bound to DNA and initiated the transcription of various genes including those responsible for the production of the enzyme Iuciferase. Luciferase production was evaluated after 8 hours. All samples were normalized to the TNF-o activation control. Treatment of the HeLa/NF-KB-Iuc cells with imidazolines without any TNF-d activation did not induce a significant amounts of luciferase activity, indicating that the imidazolines did not stimulate the NF-KB pathway. 63 I II I I I II I I I III; I_ I$333111:ijI,I§I,I;I:£IZIZI;I,I IIIgI:I:II{I}:l:l:i:t:|:i:€t}:l:l:fl:t:t:l:lEl: 333311331; OI. Translocation Nucleus ‘ Luciferase Gene . . a“ ‘ I - Transcription or <— 5 . , . ,. Responder NF-KB a. Gene Binding ,si' Figure "-5. Methods for analyzing NF-KB inhibition. Nuclear l The compounds were further evaluated for their ability to inhibit production of the cytokine lL-6 in human whole blood by Theresa Lansdell. Human whole blood was activated with IL-1B inducing the production of cyctokine, IL-6, via NF- KB mediated gene transcription (Figure "-5). Plasma was harvested and lL-1B induced lL-6 production was measured 22 hours after stimulation using a human lL-6 ELISA (R&D Systems). The circulating IL-6 levels in lL—1B stimulated samples were significantly higher than in unstimulated or the vehicle treated blood. Pretreatment of the blood for 2 hours with the imidazolines, followed by IL- 18 stimulation resulted in a strong dose dependent inhibition of IL-6 production, as compared to the no-drug treated control. The inhibitory properties of all the imidazolines evaluated in this study are summarized in the following sections. F. Structure activity relationship investigation of R1. The structural optimization studies of the oxazol-5(4H)-one derived imidazoline II- 10 was initiated by first examining the R1 substituent of the scaffold. These derivatives were readily synthesized utilizing oxazoI-5(4H)-ones derived from phenyl glycine, alanine, tryptophan, and phenyl-alanine. Due to the increased stability of imidazoline "-10 over "-4, all intermediate imidazolines were esterified with TMSCHN2 to provide compounds "-11 through "-15 (Table "-2). Pre- treatment of the HeLa NF-KB—IUC cells with imidazolines "-11 through "-15 followed by TNF-a stimulation resulted in a dose dependent decrease in Iuciferase production (Table "-2). lmidazolines "-11 and "-15 were the most effective at inhibiting NF-KB mediated luciferin production with ECso values of 7.2 (1M and 5.9 (M respectively (T able "-2). Compounds "-11 to "-15 were also evaluated for their ability to inhibit NF—KB mediated lL-6 production in lL-1B stimulated human blood. Human whole blood was incubated with the imidazolines "-11 to "-15, for 2 hours and then activated with lL-1B inducing an NF-KB mediated cytokine response. Similar to the HeLa NF-KB-luc assays, compounds "-11 and "-15 were found to be the most potent with leo values of 65 3.0 (M and 4.0 (M, respectively for the inhibition of lL-6 production. Since compound "-11 was found to be slightly more potent than compound "-15, the remainder of the compounds found in this study were derived from 2,4—diphenyl- 5(4H)-oxazolone "-2. Ph) PhYN I N\2—-r>n R; 002MB Inhibition of Inhibition of IL-6 Compound R1 HeLa NF-kB Luc In human blood 5060 Ill") 'cso (IN) H. "-11 g 7.5 3.0 "-12 H303”:- ~20 6.3 "-13 \l/i“ 18.0 6.6 “‘a "-14 d 11.2 16.2 N H “a, "-15 OJ 5.9 4.0 G. Structure activity relationship investigation of R2. Table "-2. Structure activity relationship study of R1. The next series of derivatives synthesized were designed to evaluate the ester functional group of imidazoline "-10. Previously, we found that the ester or carboxylic acid moieties found in imidazolines "-10 and "-4 respectively to be 66 critical for their activity?“ lmidazolines containing other functional groups (e.g. amides and alcohols) were found to be less active than their respective ester or carboxylic acid derivatives.9 Therefore, ester substituted imidazolines "-16 through "-18 were synthesized via the esterification of imidazoline "-4. All esters (ll-10, "-11, "-16 to lI-18) were found to be excellent NF-KB inhibitors. Consistent with our previous studies, the primary amide substituted imidazoline "-9 was devoid of any significant activity in both assays (Table "-3). Interestingly, complete removal of the R2 substituent did not result in complete loss of inhibitory function (Table "-3, compound "-5) although a reduction in potency was noted (ECso = 5.5 uM and IC50 = 2.5 (M). The ethyl ester substituted imidazoline "-10 provided the best results inhibiting NF-KB mediated luciferin and lL-6 production with an E050 value of 2.5 (M and ICso value of 0.8 uM respectively. Due to its potency and stability against hydrolysis by esterases,9 the ethyl ester substituted imidazoline "-10 was further evaluated through functionalization of its R3 and R4 moieties (Tables "-4 and "-5). 67 Ph ) Ph N 118.1% Ph‘“~ R2 j compound a, "marsh. 'I'I"l'.’.'.?.:'.‘.%l&f lice (11M) 'Csomll) lI-11 002Me 7.5 3.0 "-10 C02Et 2.5 0.8 "-1 6 C02"Pr 2.3 1 .9 "-17 COZ‘Pr 3.5 2.0 "-18 C028n 3.5 6.0 "-9 CONH2 >20 18.1 "-6 H 5.5 2.5 Table "-3. Structure activity relationship study of R2. H. Structure activity relationship study of R2. We next examined the R3 position of the ethyl ester substituted imidazoline "-10. A series of imidazolines (II-19 to II-26) containing various aromatic R3 substituents were synthesized and subsequently evaluated for their ability to inhibit NF-KB mediated gene transcription (Table "-4). Minor structural changes had significant affects on overall potency of the imidazolines analyzed. Substitution of this moiety with hetero-aromatic substituents seemed to cause a 68 decrease in activity. The 4-pyridino-substituted imidazoline “-19 was devoid of activity in our HeLa NF-KB-IUC assay and showed relatively weak activity (Table "-4, IC50 = 12.4 (M) in our whole blood assay. Furthermore, the 2-furyl substituted imidazoline "-27 only exhibited moderate activity towards inhibiting Iuciferase production (ECso = 7.8 pM), while illustrating a relatively good capability to inhibit lL—6 production (le0 = 1.9 uM). On the other hand, the para- chloro substituted imidazoline "-24 proved to be the most potent analogue in this series inhibiting NF-KB mediated luciferin and IL-6 production with an E050 value of 1.9 pM and I050 value of 0.3 (M respectively. As seen previously, the two assays corresponded well in terms of their relative potencies. The only possible exception was the aniline substituted imidazoline "-21, which illustrated relatively weak activity in the HeLa NF-KB-Iuc assay (Table "-4, EC50 = 10 (M) but showed excellent activity (Table "-4, IC50 = 0.5 (M) in our whole blood assay. Overall, this data indicated that increasing the Iipophilic nature of the R3 substituent increased the overall inhibitory activity of the compounds. 69 Ph Ph N Ph“; c0251 J \II/ R, 7 N W... a. mister... W... 506001") 1cm (pm) "-10 §——© 2.5 08 "-19 /__\N >20 12.4 "'20 E‘Ol‘wz 3.4 0.5 "'21 i‘QNHz 10.4 0.5 "'22 i-Q‘OMe 3.6 1.3 "'23 EQF 5.0 1.2 "'24 E—Q—CI 1.5 0.3 "'25 g—Q—Cfi 1.4 0.8 "'26 g0 Br 1 .5 __ "'27 E—(tll 7-5 1.9 Table "-4. Structure activity relationship study of R3. I. Structure activity relationship study of R4. We next synthesized and analyzed imidazolines "-28 to "-37 to analyze the structural activity relationship of the fourth domain (Table "-4, R4 substituent). This substituent proved to be the most sensitive to derivatization. Substitution of 70 the R4 group of imidazoline "-10 by benzyl groups containing Iipophilic substituents increased its activity as indicated by the compounds "-28 to "-33. The para-bromobenzyl imidazoline "-32 provided the best results in this set of compounds inhibiting NF-KB mediated luciferin and lL-6 production with an E050 value equal to 1.6 pM and I050 value of 0.5 (M respectively. Debenzylation of imidazoline Il-10 afforded imidazoline "-37, which was found to be inactive in both assays. Interestingly, replacement of the benzyl moiety found in imidazoline "-10 with electron deficient substituents almost completely abrogated any inhibitory activity. For example, replacement of the benzyl substituent with an acyl group (Table "-5, compound II-36) resulting in complete loss of activity. In addition, tosyl and benzoyl substituted imidazolines "-34 and "-35 provided no inhibition of NF-KB luciferin production and only moderate efficacy in our whole blood assay (Table "-5, ICso values of 6.6 (1M and 5.9 (M respectively). 71 .R4 PhYN | N\2—Ph Ph“ c0213 Inhibition of Inhibition of IL-6 Compound R. HeLa NF-kB Luc in human blood EC” (pH) |Geo In”) "-10 /—© 2.5 0.8 my "-28 /—©—OMe 5.5 1.4 ”We "-29 f—Q-Me 2.5 4.6 '42., "-30 /—-©—F 4.7 1.2 View "-31 f—Q—Cl 4.2 1.6 "We “-32 /—©—Br 1.6 0.5 “44, "-33 f—Q—ca 6.7 1.5 we, o "-34 >—© 20 6.6 "In 0.x "-35 /S\\ Me >20 5.9 w». o o "-36 >20 >20 we, "-37 H >20 >20 '14.. Table "-5. Structure activity relationship study of R4. 72 J. Experimental 1. General. Reactions were carried out in flame-dried glassware under nitrogen atmosphere. All reactions were magnetically stirred and monitored by TLC with 0.25 pm pre- coated silica gel plates using W light to visualize the compounds. Column chromatography was carried out on Silica Gel 60 (230-400 mesh) supplied by EM Science. Yields refer to chromatographically and spectroscopically pure compounds unless otherwise noted. Infrared spectra were recorded on a Nicolet lR/42 spectrometer. 1H and 13C NMR spectra were recorded on a Varian Unity Plus-500 spectrometer. Chemical shifts are reported in ppm relative to tetramethylsilane (0.00 ppm) and CHCI3 (7.26 ppm for 1H NMR) and CDCI3 (77.0 ppm 13C NMR) as the internal standards. The following abbreviations are used to denote the multiplicities: s = singlet, d = doublet, t = triplet, q = quartet and m = multiplet. Gas chromatography I low resolution mass spectra were recorded on a Hewlet-Packard 5890 Series II gas chromatograph connected to a TRIO-1 El mass spectrometer. HRMS were obtained at the Mass Spectrometry Facility of Michigan State University with a JEOL JMS HX-110 mass spectrometer. Melting points were obtained using an Electrotherrnal® capillary melting point apparatus and are uncorrected. 2. Materials. Reagents and solvents were purchased from commercial suppliers and used without further purification. Anhydrous methylene chloride and benzene were 73 dispensed from a delivery system which passes the solvents through a column packed with dry neutral alumina. Anhydrous TMSCI required for these reactions was distilled from calcium hydride. 3. Compound synthesis and characterization. lmidazolines "-11 to “-15, "-16 to "-18, and “-28 to "-33 were all prepared using procedures previously reported.9"2'13'67 lmidazolines "-8 and "-9 were also prepared using reported procedures.9 These compounds were prepared and fully characterized by Dr. Daljinder Kahlon. Imidazole "-7 was an observed byproduct from the decarboxylation of imidazoline lI-4. Imidazole "-7 had been previously prepared in our laboratories by Christopher Hupp. His spectroscopic data was used to confirm the structure of imidazole "-7. For further details regarding either the synthesis or characterization of these compounds, please see the supporting information of the following publication: Kahlon, D. K.; Lansdell, T. A.; Fisk, J. S.; Tepe, J. J.; Structural-activity relationship study of highly-functionalized imidazolines as potent inhibitors of nuclear transcription factor-kappaB mediated lL-6 production. Bioorg. Med. Chem. 2009, 17, 3093-3103.67 0 COZH Ph N Ph H "-1 2-benzamido-2-phenylacetic acid (II-1): A solution of 2-phenylglycine (12.0 g, 79.4 mmol) in 150 mL of 1M NaOH solution was treated dropwise with benzoyl chloride (12.3 g, 87.3 mmol) at 0 °C. The solution was stirred overnight while being allowed to warm to room temperature. The solution was then washed with 74 EtOAc (1x100mL), cooled to 0 °C. and acidified with 2M HCI. The aqueous solution was then washed again with EtOAc (3x100mL). The combined EtOAc washes were dried over magnesium sulfate and concentrated in vacuo. The resulting crude solid was recrystallized using EtOAc/Hexanes to afford 14.5 g (71% yield) of the title compound as a white crystalline solid. (mp. = 178 °C — 179 °C) 1H NMR (500 MHz, DMSO-dis): 0 5.61 (d, J = 7.5 Hz, 1H), 7.34 — 7.39 (m, 3H), 7.44 - 7.53 (m, 5H), 7.92 (d, J = 8.0 Hz, 2H), 9.02 (d, J = 7.5 Hz, 1H), 12.92 (bs, 1H); "‘0 NMR (125 MHz, 0111150416): 5 56.82, 127.69, 127.88, 128.15, 128.18, 128.39, 131.45, 133.75, 137.11, 166.30, 171.92; IR (neat): 3306 cm“, 3054 cm“, 2923 cm", 1734 cm", 1653 cm"; LRMS (El): m/z calcd for C15H13NO3, 255.3; fOUfld, 255.4. Ph "-2 2,4-diphenyl-5(4H)-oxazolone (ll-35): A solution of N-benzoyl-2-phenylglycine "-34 (10.0 g, 39.2 mmol) in 250 mL of anhydrous dichloromethane was treated with trifluoroacetic anhydride (8.6 g, 41.1 mmol) at room temperature for 12 hours. The solution was then washed with saturated NaHCO2, (3x100 mL) and brine (1x100mL), dried over sodium sulfate, and concentrated in vacuo resulting ‘ in 8.5 g (92% yield) of the title compound as a yellow solid (mp. = 80 °C — 81 °C). The title compound was used as is without further purification. 1H NMR (500 MHz, CDCI3) (T MS): 5 5.53 (s, 1H), 7.33 - 7.44 (m, 5H), 7.50 - 7.53 (m, 2H), 7.59 — 7.63 (m, 1H), 8.07 — 8.09 (m, 2H); ”'0 NMR + DEPT (125 MHz, 75 coch) (TMS): 5 68.02, 125.50, 126.82, 128.13, 128.75, 128.90, 129.01, 133.15, 133.29, 162.74, 176.06; IR (KBr): 3063 cm", 3032 cm", 1827 cm", 1649 cm"; LRMS(EI)I (TI/Z CBICd fOl' C15H11N02, 237.3; found, 236.9. Ph> gig—ion |Io‘h ll00211 dl-(4S,SS)-1 benzyl-2,4,5-triphenyI-4,5-dihydro-1 H-imidazole-4-carboxylic acid hydrochloride (II-3): A solution of benzyl amine (3.2 g, 29.5 mmol) and benzaldehyde (3.1 g, 29.5 mmol) in 200 mL of anhydrous benzene was refluxed under nitrogen for 2 hours and then concentrated in vacuo. The resulting residue was redissolved into 300 mL of anhydrous dichloromethane. Then 2,4-diphenyl- 5(4H)-oxazolone "-2 (7.0 g, 29.5 mmol) and chlorotrimethylsilane (4.2 g, 38.4 mmol) were added and the mixture was refluxed under nitrogen for 12 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing 9.4g (68% yield) of the title compound as a white solid precipitate. The spectra matches that of previously published literature?” (m.p. = 154 °c - 155 °C) 1H NMR (500 MHz, DMSO-dB): 5 4.18 (d, J = 16 Hz, 1H), 4.86 (d, J = 16 Hz, 1H), 5.59 (s, 1H), 6.71 (d, J = 7.5 Hz, 2H), 7.08-7.11 (m, 2H), 7.17-7.20 (m, 1H), 7.45-7.59 (m, 8H), 7.72-7.77 (m, 4H), 7.82- 7.85 (m, 1H), 7.97 (d, J = 7.5 Hz, 2H), 12.86 (bs, 1H), 13.99 (bs, 1H); 13C NMR + DEPT (125 MHz, DMSO-dis): 5 48.67 (-CH2), 73.86 (-CH), 75.91 (quaternary —C), 121.39 (aromatic -CH), 125.74 (aromatic -CH), 127.21 (aromatic -CH), 128.31 76 (aromatic -CH), 128.67 (aromatic -CH), 128.89 (aromatic —CH), 129.05 (aromatic -CH), 129.11 (aromatic —CH), 129.45 (aromatic —CH), 129.55 (aromatic —CH), 129.95 (aromatic -CH), 132.80 (quaternary aromatic -C), 133.17 (quaternary aromatic —C), 134.26 (quaternary aromatic —C), 139.18 (quaternary aromatic —C), 165.66, 167.64; LRMS (El): m/z calcd for C29H25N2O2CI, 469.0; found, 387.8 (-CO2). Ph 7 Ph N 11:84," Ph“: COZH lI-4 dl-(4S,SS)-1 -benzyI-2,4,5-tri phenyl-4,5-dihyd ro-1 H-imidazoIe-4-carboxylic acid (II-4): A solution of imidazoline "-3 (5.0 g, 10.7 mmol) in 100 mL of dichloromethane was washed with saturated sodium bicarbonate solution (2 X 100 mL) and brine solution (1 X 100 mL). The solution was then dried over sodium sulfate and concentrated in vacuo. The resulting crude solid was purified via crystallization (dichloromethane l hexanes) to afford 3.8 g (83% yield) of the title compound as a white crystalline solid. The spectra matches that as previously published literature.9 (m.p. = 116 °c - 120 °C) 1H NMR (500 MHz, CDCI3): 5 3.77 (d, J = 16 Hz, 1H), 4.59 (d, J = 16 Hz, 1H), 4.92 (s, 1H), 6.59 (d, J = 7.5 Hz, 2H), 7.05-7.08 (m, 1H), 7.26-7.32 (m, 5H), 7.36-7.39 (m, 5H), 7.46-7.49 (m, 1H), 7.49-7.52 (m, 2H), 7.86 (d, J = 7.5 Hz, 1H), 9.18 (bs, 1H); 13C NMR 4- DEPT (125 MHz, CDCI3): 5 48.35 (-CH2), 75.58 (-CH), 79.04 (quaternary —C), 122.86 (quaternary aromatic -C), 125.73 (aromatic —CH),, 126.70 (aromatic — 77 CH), 127.36 (aromatic —CH), 127.91 (aromatic —CH), 128.18 (aromatic —CH), 128.85 (aromatic —CH), 128.91 (aromatic -CH), 128.97 (aromatic —CH),129.27 (aromatic —CH), 132.93 (aromatic -CH), 133.74 (quaternary aromatic —C), 136.01 (quaternary aromatic —C), 143.15 (quaternary aromatic —C), 164.70, 167.96; IR (neat): 3061 cm", 2700 cm“, 1633 cm“, 1550 cm", 1340 cm" Ph ”w? N|\;)—Ph Ph "-5 dI-(4S,5$)-1-benzyl-2,4,5-triphenyl-4,5-dihydro-1H-imidazole (II-5): The title compound was isolated in small amounts via a decarboxylation of imidazoline ll- 4. A solution of imidazoline "-4 (0.7 g, 1.62 mmol) in 100 mL of THF was refluxed for 5 hours. The solution was then concentrated in vacuo resulting in a crude mixture of imidazolines "-5 and "-6 and imidazole "-7 in a 2:1:1 ratio respectively. The crude residue was then purified via column chromatography (10% MeOH / 90% CH2CI2) affording 112 mg (18% yield) of the title compound as a white solid. (mp. = 78 °C — 80 °C) 1H NMR (500 MHz, CDCI3): 5 3.99 (d, J = 16 Hz, 1H), 4.41 (d, J = 8.5 Hz, 1H), 4.77 (d, J = 15.5 Hz, 1H), 5.07 (d, J = 8.5 Hz, 1H), 7.00-7.01 (m, 2H), 7.18-7.23 (m, 2H), 7.24-7.30 (m, 4H), 7.31-7.33 (m, 2H), 7.34-7.39 (m, 3H), 7.42-7.45 (m, 2H), 7.53-7.55 (m, 3H), 7.88-7.90 (m, 2H); 13C NMR + DEPT (125 MHz, CDCI3): 5 49.63, (-CH2), 72.57 (-CH), 77.95 (-CH), 126.72 (aromatic —CH), 126.96 (aromatic —CH), 127.13 (aromatic —CH), 127.42 (aromatic —CH), 127.72 (aromatic -CH), 127.94 (aromatic —CH), 128.34 78 (aromatic -CH), 128.42 (aromatic -CH), 128.58 (aromatic -CH), 128.65 (aromatic -CH), 128.81 (aromatic -CH), 130.07 (aromatic —CH), 131.34 (aromatic quaternary —C), 136.42 (aromatic quaternary -C),141.82 (aromatic quaternary -C), 143.90 (aromatic quaternary —C), 165.91; IR (neat): 3063 cm“, 3030 cm", 1595 cm"; HRMS (ESI): m/z calcd for C23H24N2 [M-I-H], 369.2016; found, 389.2020. Ph dl-(4R,58)-2,4,5-triphenyl-4,5-dihydro-1H-imidazole: This compound was made according to a reported literature procedure.65 A solution of benzaldehyde (10.0 g, 94.2 mmol) and hexamethyldisalizane (18.2 9, 113.0 mmol) was treated with benzoic acid (57.5 mg, 0.5 mmol) and heated to 120 °C for 22 hours. The solution was then dissolved into 50 mL of toluene and washed with sat. NaHCO:, (2 x 50 mL), water (1 x 50 mL), and brine (1 x 50 mL). The solution was then dried over magnesium sulfate and concentrated in vacuo. The resulting residue was purified via column chromatography (10% MeOH, 90% CH2CI2) affording 5.8 g (62% yield) of the title compound as a white solid. (mp. = 123 ° - 124 °C) 1H NMR (500 MHz, CDCI3): 5 4.90 (bs, 1H), 5.43 (s, 2H), 6.95-7.15 (m, 10H), 7.47- 7.50 (m, 2H), 7.51-7.60 (m, 1H), 7.97-8.10 (m, 2H); 130 NMR (125 MHz, cock.) (T MS): 5 70.70, 126.68, 127.21, 127.45, 127.53, 128.54, 130.05, 130.93, 138.98; IR (neat): 3385 cm", 3165 cm", 3028 cm", 1599 cm"; HRMS (ESI): m/z calcd for C21H13N2 [M-l-H], 299.1548; found, 299.1541. 79 ._...—._ ___ F Ph N Y\8—Ph Ph "-6 dl-(4R,SS)-1benzyl-2,4,5triphenyl-4,5-dihydro-1H-imidazole (ll-6): A solution of dI-(4R,58)-2,4,5-triphenyI-4,5-dihydro-1H-imidazole (0.3 g, 1.01 mmol) and benzyl bromide (0.2 g, 1.06 mmol) in 20 mL of anhydrous benzene was treated with triethyl amine (0.2 g, 2.02 mmol). The solution was refluxed for 15 hours and then washed with saturated NaHCO3 (2x20mL) and brine (1x20mL). The solution was then dried over sodium sulfate and concentrated in vacuo. The resulting crude residue was purified via column chromatography (10% MeOH I 90% CH2CI2) affording 77 mg (20% yield) of the title compound as a yellow oil. 1H NMR (500 MHz, CDCla): 5 3.85 (d, J = 15.5 Hz, 1H), 4.77 (d, J = 16 Hz, 1H), 4.92 (d, J = 11.5 Hz, 1H), 5.55 (d, J = 11 Hz, 1H), 6.90-6.96 (m, 5H), 6.97-7.00 (m, 4H), 7.02-7.09 (m, 3H), 7.25-7.29 (m, 3H), 7.50-7.53 (m, 3H), 7.81-7.83 (m, 2H); 130 NMR + DEPT (125 MHz, CDCIa): 5 48.97 (-CH2), 68.38 (CH), 72.94 (- CH), 126.23 (aromatic -CH), 127.07 (aromatic —CH), 127.30 (aromatic -CH), 127.52 (aromatic -CH), 127.80 (aromatic -CH), 127.86 (aromatic —CH), 127.93 (aromatic -CH), 128.07 (aromatic —CH), 128.54 (aromatic -CH), 128.56 (aromatic —CH), 128.72 (aromatic —CH), 130.18 (aromatic —CH), 131.17 (aromatic quaternary -C), 136.64 (aromatic quaternary —C), 136.82 (aromatic quaternary —C), 139.32 (aromatic quaternary -C), 167.15; IR (neat): 3030 cm", 80 2924 cm", 1595 cm"; HRMS (ESI): m/z calcd for 024124112 [M+H], 389.2016; found, 389.2017. Ph 7 Ph N Whiz—oh on“: com 11.10 (4S,SS)-ethyl-1 -benzyl-2,4,5-triphenyI-4,5-dihydro-1 H-imidazoIe-AI— carboxylate (ll-10): A solution of dl-(4S,5$)-1-benzyI-2,4,5-triphenyl-4,5- dihydro-1H-imidazole—4-carboxylic acid hydrochloride "-3 (4.0 g, 8.53 mmol) and oxalyl chloride (3.25 g, 25.6 mmol) in 200 mL of anhydrous dichloromethane was treated with 200 uL of DMF at 0°C. The solution was stirred for 3 hours and then concentrated in vacuo. The resulting yellow solid was cooled to 0 °C and treated with 125 mL of ethanol and stirred for 4 hours. The solution was concentrated in vacuo and then dissolved into 100 mL of dichloromethane before being washed with saturated NaHCO2, (3 x 100 mL) and brine (1 x 100 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude residue was purified via silica gel column chromatography (40% EtOAc l 60% Hexanes) affording 3.6 g (92% yield) of the title compound as a white solid (mp. = 87 ° - 89 °C). 1H NMR (500 MHz, CDCI3): 5 0.82 (t, J = 7 Hz, 3H), 3.61 (dq, J1 = 11 Hz, J2 = 7.5 Hz, 1H), 3.73 (dq, J; = 11 Hz, J2 = 7.5 Hz, 1H), 3.85 (d, J = 16 Hz, 1H), 4.64 (d, J = 16 Hz, 1H), 4.94 (s, 1H), 6.74 (d, J = 7.5 Hz, 2H), 7.04- 7.07 (m, 2H), 7.09-7.12 (m, 1H), 7.25-7.30 (m, 1H), 7.32-7.40 (m, 7H), 7.45-7.48 (m, 3H), 7.73-7.77 (m, 4H); ”C NMR + DEPT (125 MHz, CDCI3): 5 13.46 (- 81 CH3), 48.61 (-CH2), 60.95 (-CH2), 73.78 (-CH), 82.89 (quaternary —C), 126.80 (aromatic —CH), 127.12 (aromatic —CH), 127.30 (aromatic —CH), 127.34 (aromatic —CH), 127.97 (aromatic -CH), 128.15 (aromatic -CH), 128.27 (aromatic —CH), 128.37 (aromatic -CH), 128.44 (aromatic —CH), 128.55 (aromatic —CH), 128.82 (aromatic -CH), 130.26 (aromatic -CH), 130.65 (quaternary aromatic -C), 136.67 (quaternary aromatic -C), 137.95 (quaternary aromatic -C), 144.15 (quaternary aromatic —C), 165.39, 170.78; IR (neat): 3063 cm", 2960 cm", 1732 cm", 1595 cm"; HRMS (ESI): m/z calcd for Cg1H28N2O2 [M+H], 461.2229; found, 461.2225. Ph Ph > T30 N \ / on“ c0251 11.19 dl-(4S,5$)-ethyI-1 -benzyI-2,4~diphenyl-5-(pyridin-4-yI)-4,5-dihydro-1 H- imidazoIe-4-carboxylate (ll-19): A solution of benzyl amine (0.2 g, 2.11 mmol) and 4-pyiidylcarboxaldehyde (0.2 g, 2.11 mmol) in 50 mL of anhydrous benzene was reflux under nitrogen for 12 hours and then concentrated in vacuo. The resulting residue was redissolved into 50 mL of anhydrous dichloromethane. Then 2,4-diphenyI-5(4H)-oxazolone "-2 (0.5 g, 2.11 mmol) and chlorotrimethylsilane (0.3 g, 2.74 mmol) were added and the mixture was refluxed under nitrogen for 12 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing a white solid precipitate (0.4 g) which was isolated via filtration. The white solid was then 82 dissolved into 50 mL of dichloromethane, cooled to 0 °C. and treated with oxalyl chloride (0.3 g, 2.68 mmol) and DMF (30 uL). The solution was stirred for 2 hours, concentrated in vacuo, and redissolved into 20 mL of EtOH before being left to stir overnight. The solution was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaHCOa (3 x 50 mL) and brine (1 x 50 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude residue was purified via column chromatography (100% EtOAc) to afford 220 mg (23% yield) of the title compound as a clear oil. 1H NMR (500 MHz, CDCI3): 5 0.83 (t, J = 7.5 Hz, 3H), 3.66 (dq, J; = 11 Hz, J2 = 7 Hz, 1H), 3.75 (dq, J1 = 11 Hz, J2 = 7.5 Hz, 1H), 3.83 (d, J = 15.5 Hz, 1H), 4.62 (d, J = 15.5 Hz, 1H), 4.86 (s, 1 H), 6.74 (d, J = 7.5 Hz, 2H), 7.05-7.08 (m, 2H), 7.12-7.14 (m, 1H), 7.28-7.37 (m, 5H), 7.49-7.52 (m, 3H), 7.71-7.72 (m, 2H), 7.79-7.81 (m, 2H), 8.63 (d, J = 6 Hz, 2H); 130 NMR + DEPT (125 MHz, CDCI3): 5 13.39 (-CH3), 49.24 (-CH2), 61.20 (-CH2), 72.62 (-CH), 83.06 (quaternary -C), 122.95 (aromatic —CH), 126.49 (aromatic -CH), 127.14 (aromatic -CH), 127.56 (aromatic -CH), 127.61 (aromatic —CH), 128.11 (aromatic —CH), 128.43 (aromatic -CH), 128.65 (aromatic -CH), 128.77 (aromatic -CH), 129.93 (aromatic quaternary —C), 130.60 (aromatic —CH), 135.86 (quaternary aromatic -C), 143.34 (aromatic quaternary —C), 147.28 (aromatic quaternary -C), 149.93 (aromatic -CH), 165.71, 170.23; IR (neat): 3063 cm", 2982 cm", 1734 cm", 1597 cm"; HRMS (ESI): m/z calcd for C30H27N3O2 [M+H], 462.2162; found, 462.2177. 83 Ph 7 two Ph‘c‘ 002Et "-20 Ph dl-(4S,5$)-ethyI-1 -benzyl-5-(4—nitrophenyl)-2,4-diphenyl-4,5-dihyd ro-1 H- imidazole-4-carboxylate (II-20): A solution of benzyl amine (0.2 g, 2.11 mmol) and 4-nitrobenzaldehyde (0.3 g, 2.11 mmol) in 50 mL of anhydrous benzene was reflux under nitrogen for 12 hours and then concentrated in vacuo. The resulting residue was redissolved into 50 mL of anhydrous dichloromethane. Then 2,4- diphenyl-5(4H)-oxazolone "-2 (0.5 g, 2.11 mmol) and chlorotrimethylsilane (0.3 g, 2.74 mmol) were added and the mixture was refluxed under nitrogen for 24 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing a white solid precipitate (0.6 g) which was isolated via filtration. The white solid was then dissolved into 25 mL of dichloromethane, cooled to 0 °C. and treated with oxalyl chloride (0.5 g, 3.56 mmol) and DMF (30 pL). The solution was stirred for 2 hours, concentrated in vacuo, and redissolved into 25 mL of EtOH before being left to stir overnight. The solution was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaH003 (3 x 50 mL) and brine (1 x 50 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude solid was purified via column chromatography (50% EtOAc, 50% hexanes) affording 540 mg (51% yield) of the title compound as a white solid. (mp. = 173 °c -174 °C) ‘H NMR (500 MHz, cock): 5 0.63 (t, J = 7.5 Hz, 3H), 3.64 (dq, J1 = 11 Hz, J2 = 7 Hz, 1H), 3.74 (dq, J1 = 11 Hz, J2 = 7 Hz, 1H), 3.81 (d, J = 16 Hz, 1H), 4.60 (d, J = 16 Hz, 1H), 4.93 (s, 1H), 6.71 (d, J = 7.5 Hz, 2H), 7.03-7.10 (m, 2H), 7.10-7.13 (m, 1H), 7.27-7.35 (m, 3H), 7.48-7.51 (m, 3H), 7.54 (d, J = 8.5 Hz, 2H), 7.67-7.69 (m, 2H), 7.77-7.80 (m, 2H), 8.21 (d, J = 9.5 Hz, 2H); ”C NMR + DEPT (125 MHz, CDCI3): 5 15.59 (-CH3), 49.46 (-CH2), 61.30 (-CH2), 73.16 (-CH), 83.32 (quaternary —C), 123.61 (aromatic —CH), 126.49 (aromatic -CH), 127.25 (aromatic -CH), 127.69 (aromatic —CH), 127.75 (aromatic -CH), 128.24 (aromatic —CH), 128.53 (aromatic -CH), 128.76 (aromatic -CH), 128.87 (aromatic —CH), 128.96 (aromatic —CH), 129.98 (aromatic —-CH), 130.74 (aromatic —CH), 135.83 (quaternary aromatic -C), 143.45 (quaternary aromatic —C), 146.09 (quaternary aromatic -C), 147.76 (quaternary aromatic -C), 165.76, 170.42; IR (neat): 3063 cm", 2982 cm", 1732 cm", 1597 cm", 1350 cm"; HRMS (ESI): m/z calcd for CaIH27N304 [Mi-H], 506.2080; found, 506.2076. Ph 7 iW~~2 Ph“° COzEt 11.21 Ph dl-(4S,5$)-ethyI-5-(4-aminophenyI)-1 -benzyI-2,4-diphenyl-4,5-dihydro-1 H- imidazole-4-carboxylate (II-21): A solution of dl-(4S,5S)-ethyI-1-benzyl-5-(4- nitrophenyl)-2,4-diphenyI-4,5-dihydro-1H-imidazole-4-carboxylate "-20 (0.1 g, 0.2 mmol) H20 (36 mg, 2.0 mmol) in 10 mL of ethanol was treated with SnCI2'2H2O (0.3 g, 1.2 mmol). The solution was heated to reflux for 2 hours and cooled to 85 room temperature before being poured over ice (~50 g). The pH of the resulting aqueous solution was adjusted (pH = 8) using NaHCO:, powder. The solution was then washed with EtOAc (3x50 mL). The combined EtOAc washes were dried over sodium sulfate and concentrated in vacuo. The resulting residue was purified via column chromatography (EtOAc) to afford 57 mg (60% yield) of the title compound as a white solid. (mp. = 60 °c - 62 °C) 1H NMR (500 MHz, CDCI3): 5 0.91 (t, J = 7.5 Hz, 3H), 3.68-3.82 (m, 4H) 3.86 (d, J = 16 Hz, 1H), 4.60 (d, J = 15.5 Hz, 1H), 4.86 (s, 1H), 6.68 (d, J = 8.5 Hz, 2H), 6.78 (d, J = 7.5 Hz, 2H), 7.08-7.16 (m, 3H), 7.18 (d, J = 8 Hz, 2H), 7.27—7.30 (m, 1H), 7.35-7.36 (m, 2H), 7.47-7.50 (m, 3H), 7.75-7.78 (m, 4H); ”C NMR + DEPT (125 MHz, CDCI3): 513.57 (-CH3), 48.24 (-CH2), 60.86 (-CH2), 73.48 (-CH), 82.51 (quaternary -C), 114.84 (aromatic —CH), 126.78 (aromatic -CH), 127.07 (aromatic —CH), 127.16 (aromatic —CH), 127.19 (aromatic —CH), 127.86 (aromatic —CH), 128.29 (aromatic -CH), 128.46 (aromatic —CH), 128.70 (aromatic —CH), 129.18 (aromatic -CH), 130.09 (aromatic —CH), 130.84 (quaternary aromatic —C), 136.88 (quaternary aromatic —C), 144.24 (quaternary aromatic -C), 146.51 (quaternary aromatic -C), 165.19, 170.96; IR (KBr): 3460 cm", 3373 cm", 3063 cm", 1732 cm", 1614 cm"; HRMS (ESI): m/z calcd for C31H29N302 [Mi-H], 476.2338; found, 476.2332. 86 KM} COzEt lI-22 dl-(4S,SS)-ethyl-1 -benzyl -5-(4—methoxyphenyI)-2,4-diphenyl4,5-dihydro-1 H- imidazole-4-carboxylate (II-22): A solution of benzyl amine (0.2 g, 1.91 mmol) and p-anisaldehyde (0.3 g, 1.91 mmol) in 50 mL of anhydrous benzene was reflux under nitrogen for 3 hours and then concentrated in vacuo. The resulting residue was redissolved into 50 mL of anhydrous dichloromethane. Then 2,4- diphenyl-5(4H)-oxazolone "-2 (0.5 g, 1.91 mmol) and chlorotrimethylsilane (0.3 g, 2.48 mmol) were added and the mixture was refluxed under nitrogen for 12 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing a white solid precipitate (450 mg) which was isolated via filtration. The crude solid was then dissolved into 20 mL of EtOH, treated with 1 mL of concentrated H2804, and refluxed for 42 hours. The solution (was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaH003 (3 x 50 mL) and brine (1 x 50 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude solid was purified via column chromatography (50% EtOAc l 50% Hexanes) to afford 336 mg (36% yield) of the title compound as an off-white solid. (mp. = 143 °C — 144 °C) 1H NMR (500 MHz, CDCI3): 5 0.85 (t, J = 7 Hz, 3H), 3.64 (dq, J1 = 10.5 Hz, J2 = 7.5 Hz, 1H), 3.74 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.80 (d, J = 16 Hz, 1H), 3.81 (s, 3H), 4.57 (d, J = 15.5 Hz, 1H), 4.86 (s, 1H), 87 6.74 (d, J = 7.5 Hz, 2H), 6.89 (d, J = 8.5 Hz, 2H), 7.04-709 (m, 2H), 7.09—7.20 (m, 1H), 7.25-7.33 (m, 5H), 7.44-7.47 (m, 3H), 7.72-7.74 (m, 4H); 130 NMR + DEPT (125 MHz, CDCI3): 5 13.58 (-CH3), 48.45 (-CH2), 55.25 (-CH3), 60.94 (-CH2), 73.33 (CH), 82.86 (quaternary —C), 113.82 (aromatic —CH), 126.79 (aromatic — CH), 127.12 (aromatic -CH), 127.27 (aromatic -CH), 127.30 (aromatic —CH), 127.94 (aromatic —CH), 128.37 (aromatic -CH), 128.53 (aromatic -CH), 128.77 (aromatic -CH), 129.34 (aromatic —CH), 129.76 (aromatic —CH), 130.20 (aromatic -CH), 130.76 (aromatic quaternary —C), 136.76 (aromatic quaternary - C), 144.17 (aromatic quaternary -C), 159.59 (aromatic quaternary -C), 165.30, 170.90; IR (neat): 3032 cm", 2934 cm", 1732 cm", 1595 cm"; HRMS (ESI): m calcd for C32H30N202 [M+H], 491.2335; found, 491.2332. Ph 7 ‘62:} N . F Ph“ COzEt "-23 Ph dl-(4S,5$)-ethyI-1-benzyl-5-(4-fluorophenyl)-2,4-diphenyI-4,5-dihydro-1H- lmidazole-4-carboxylate (II-23): A solution of benzyl amine (0.2 g, 2.11 mmol) and 4-fluorobenzaldehyde (0.3 g, 2.11 mmol) in 50 mL of anhydrous benzene was reflux under nitrogen for 12 hours and then concentrated in vacuo. The resulting residue was redissolved into 50 mL of anhydrous dichloromethane. Then 2,4—diphenyI-5(4H)-oxazolone "-2 (0.5 g, 2.11 mmol) and chlorotrimethylsilane (0.3 g, 2.74 mmol) were added and the mixture was refluxed under nitrogen for 12 hours. The solution was concentrated in vacuo 88 and the resulting residue was resuspended in EtOAc producing a white solid precipitate (216 mg) which was isolated via filtration. The crude solid was then dissolved into 15 mL of EtOH, treated with 1 mL of concentrated H2804, and refluxed for 48 hours. The solution was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaHCO2, (3 x 50 mL) and brine (1 x 50 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude solid was purified via column chromatography (50% EtOAc l 50% Hexanes) to afford 124 mg (12% yield) of the title compound as a white solid. (mp. = 96 °C - 97 °C) 1H NMR (500 MHz, CDCI3): 5 0.84 (t, J = 7.5 Hz, 3H), 3.67 (dq, J; = 10.5 Hz, J2 = 7.5 Hz, 1H), 3.76 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.84 (d, J = 16 Hz, 1H), 4.63 (d, J = 15.5 Hz, 1H), 4.92 (s, 1H), 6.75-6.78 (m, 2H), 7.06-7.12 (m, 4H), 7.12-7.16 (m, 1H), 7.28-7.42 (m, 5H), 7.48-7.54 (m, 3H), 7.72-7.80 (m, 4H); 13c NMR + DEPT (125 MHz, CDCI3): 5 13.53 (-CH3), 48.73 (-CH2), 61.04 (-CH2), 73.01 (-CH), 82.78 (quaternary —C), 115.35 (d, J = 21.6 Hz, aromatic -CH), 126.68 (aromatic —CH), 127.11 (aromatic —CH), 127.39 (aromatic —CH), 127.43 (aromatic —CH), 128.01 (aromatic —CH), 128.40 (aromatic -CH), 128.59 (aromatic —CH), 128.77 (aromatic -CH), 129.73 (d, J = 8 Hz, aromatic -CH), 130.37 (aromatic —CH), 130.43 (aromatic -CH), 133.77 (d, J = 3 Hz, quaternary aromatic -C), 136.40 (quaternary aromatic —C), 143.90 (quaternary aromatic —C), 162.61 (d, J = 245 Hz, quaternary aromatic —C), 165.39, 170.70; IR (neat): 3063 cm“, 2982 cm", 1732 cm", 1597 cm“; HRMS (ESI): m/z calcd for 031H27N2O2F [M+H], 479.2135; found, 479.2130. 89 Ph) Ph Y N | Ph“ 6023 "-24 dl-(4S,5S)-ethyl-1 -benzyl-5-(4-chlorophenyI)-2,4—diphenyl-4,5-dihydro-1 H- imldazole-4-carboxylate (II-24): A solution of benzyl amine (2.7 g, 25.3 mmol) and 4-chlorobenzaldehyde (3.6 g, 25.3 mmol) in 250 mL of anhydrous benzene was reflux under nitrogen for 24 hours and then concentrated in vacuo. The resulting residue was redissolved into 250 mL of anhydrous dichloromethane. Then 2,4-diphenyl-5(4H)-oxazolone "-2 (6.0 g, 25.3 mmol) and chlorotrimethylsilane (3.6 g, 32.9 mmol) were added and the mixture was refluxed under nitrogen for 18 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing a white solid precipitate (7.9 g) which was isolated via filtration. The white solid was then dissolved into 250 mL of dichloromethane, cooled to 0 °C, and treated with oxalyl chloride (6.0 g, 47.1 mmol) and DMF (300 (IL). The solution was stirred for 3 hours, concentrated in vacuo, and redissolved into 250 mL of EtOH before being left to stir overnight. The solution was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaHCOa (3 x 200 mL) and brine (1 x 200 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude solid was recrystallized using EtOAc/Hexanes to afford 6.23 g (50% yield) of the title compound as a white crystalline solid. (mp. = 165 °c — 166 °C) 1H NMR (500 MHz, 00013) (TMS): 5 90 0.86 (t, J = 7 Hz, 3H), 3.63 (dq, J1 = 11 Hz, J2 = 7 Hz, 1H), 3.73 (dq, J; = 10.5 Hz, J2 = 7 Hz, 1H), 3.80 (d, J = 15.5 Hz, 1H), 4.61 (d, J = 15.5 Hz, 1H), 4.87 (s, 1H), 6.74 (d, J = 7 Hz, 2H), 7.05—7.08 (m, 2H), 7.11-7.12 (m, 1H), 7.26-7.29 (m, 1H), 7.31-7.36 (m, 6H), 7.47-7.49 (m, 3H), 7.70-7.72 (m, 2H), 7.75-7.77 (m, 2H); 130 NMR + DEPT (125 MHz, CDCI3) (T MS): 5 13.55 (-CH3), 48.83 (-CH2), 61.12 (- CH2), 73.10 (-CH), 82.87 (quaternary -C), 126.68 (aromatic —CH), 127.17 (aromatic —CH), 127.46 (aromatic —CH), 127.49 (aromatic -CH), 128.06 (aromatic -CH), 128.45 (aromatic —CH), 128.63 (aromatic -CH), 128.65 (aromatic -CH), 128.81 (aromatic —CH), 129.49 (aromatic -CH), 130.41 (quatemary aromatic —C), 130.43 ((aromatic —CH), 134.08 (quaternary aromatic —C), 136.35 (quaternary aromatic -C), 136.63 (quaternary aromatic -C),143.84 (quaternary aromatic -C), 165.48, 170.65; IR (KBr): 3063 cm", 2980 cm“, 1732 cm", 1595 cm"; HRMS (ESI): an calcd for C31H270IN2O2 [Mi-H], 495.1833; found, 495.1834. TWO; Ph‘s COzEt II-25 dl-(4S,5S)-ethyl-1benzyl-2,44iphenyl-5-(4-(trifluoromethyl)phenyl)-4,5- dihydro-1H-imidazole-4-carboxylate (II-25): A solution of benzyl amine (0.5 g, 2.11 mmol) and 4-trifluoromethyI-benzaldehyde (0.4 g, 2.11 mmol) in 50 mL of anhydrous benzene was reflux under nitrogen for 2 hours and then concentrated in vacuo. The resulting residue was redissolved into 50 mL of anhydrous 91 dichloromethane. Then 2,4-diphenyl-5(4H)-oxazolone "-2 (0.5 g, 2.11 mmol) and chlorotrimethylsilane (0.3 g, 2.74 mmol) were added and the mixture was refluxed under nitrogen for 24 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing a white solid precipitate (0.7 g) which was isolated via filtration. The crude solid was then dissolved into 15 mL of EtOH, treated with 1 mL of concentrated H2SO4, and refluxed for 64 hours. The solution was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaHC03 (3 x 50 mL) and brine (1 x 50 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude solid was purified via column chromatography (40% EtOAc l 60% Hexanes) to afford 159 mg (14% yield) of the title compound as a white crystalline solid. (mp. = 155 °C — 156 °C) 1H NMR (500 MHz, CDCI3) (T MS): 5 0.78 (t, J = 7 Hz, 3H), 3.63 (dq, J1 = 11 Hz, J2 = 7 Hz, 1H), 3.73 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.82 (d, J = 16 Hz, 1H), 4.62 (d, J = 15.5 Hz, 1H), 4.94 (s, 1H), 6.73 (d, J = 7.5 Hz, 2H), 7.04-7.10 (m, 2H), 7.11-7.13 (m, 1H), 7.27-7.32 (m, 1H), 7.32-7.36 (m, 2H), 7.48-7.50 (m, 3H), 7.52 (d, J = 7.5 Hz, 2H), 7.64 (d, J = 8 Hz, 2H), 7.71-7.73 (m, 2H), 7.78-7.80 (m, 2H); “C NMR + DEPT (125 MHz, CDCI3) (TMS): 5 13.36 (-CH3), 49.05 (-CH2), 61.13 (-CH2), 73.21 (-CH), 83.07 (quaternary —C), 125.35 (q, J = 3.6 Hz, aromatic —CH), 123.97 (q, J = 270 Hz, quaternary -C), 126.62 (aromatic —CH), 127.16 (aromatic —CH), 127.51 (aromatic —CH), 127.56 (aromatic —CH), 128.10 (aromatic —CH), 128.44 (aromatic —CH), 128.65 (aromatic -CH), 128.83 (aromatic -CH), 130.46 (q, J = 32 Hz, quaternary —C), 130.53 (aromatic —CH), 92 136.13 (aromatic quaternary —C), 142.44 (aromatic quaternary —C), 143.68 (aromatic quaternary -C), 165.59, 170.49; IR (neat): 3065 cm", 2962 cm", 1734 cm", 1597 cm"; HRMS (ESI): m/z calcd for Cg2H27N2O2F [M+H], 529.2103; found, 529.21 10. Ph W HQ Phi COzEt ll-26 (4S,5$)-ethyI-1 -benzyI-5-(4-bromophenyI)-2,4—diphenyl-4,5-dihyd ro-1 H- imidazoleA—carboxylate (ll-26): A solution of benzyl amine (0.2 g, 2.11 mmol) and 4-bromobenzaldehyde (0.4 g, 2.11 mmol) in 50 mL of anhydrous benzene was reflux under nitrogen for 3 hours and then concentrated in vacuo. The resulting residue was redissolved into 50 mL of anhydrous dichloromethane. Then 2,4-diphenyI-5(4H)-oxazolone "-2 (0.5 g, 2.11 mmol) and chlorotrimethylsilane (0.3 g, 2.74 mmol) were added and the mixture was refluxed under nitrogen for 16 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing a white solid precipitate (0.7 g) which was isolated via filtration. The crude solid was then dissolved into 50 mL of EtOH, treated with 1 mL of concentrated H2804, and refluxed for 24 hours. The solution was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaH003 (3 x 20 mL) and brine (1 x 20 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude residue was purified via 93 column chromatography (40% EtOAc I 60% Hexanes) to afford 201 mg (18% yield) of the title compound as a white solid. (mp. = 186 °C - 188 °C) 1H NMR (500 MHz) (CDCI3): 5 0.87 (t, J = 7 Hz, 3H), 3.69 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.77 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.82 (d, J = 16 Hz, 1H), 4.62 (d, J = 16 Hz, 1H), 4.87 (s, 1H), 6.75 (d, J = 7.5 Hz, 2H), 7.07-7.10 (m, 2H), 7.13-7.16 (m, 1H), 7.27—7.31 (m, 3H), 7.33-7.36 (m, 2H), 7.49-7.53 (m, 5H), 7.72-7.74 (m, 2H), 7.76-7.79 (m, 2H); 13C NMR + DEPT (125 MHz) (CDCI3): 5 13.55 (-CH3), 48.8 (—CH2), 61.13 (-CH2), 73.14 (-CH), 82.83 (quaternary —C), 122.20 (quaternary aromatic —C), 126.66 (aromatic —CH), 127.15 (aromatic -CH), 127.45 (aromatic -CH), 127.49 (aromatic -CH), 128.05 (aromatic —CH), 128.44 (aromatic -CH), 128.63 (aromatic -CH), 128.80 (aromatic —CH), 129.80 (aromatic -CH), 130.37 (quaternary aromatic —C), 130.43 (aromatic —CH), 131.60 (aromatic -CH), 136.31 (quaternary aromatic —C), 137.15 (quaternary aromatic —C), 143.80 (quaternary aromatic —C), 165.48, 170.63; IR (neat): 3100 cm“, 2981 cm“, 1730 cm", 1595 cm"; HRMS (ESI): m/z calcd for CaIH23N2O2Br [M+H], 539.1334; found, 539.1338. dI-(4S,5R)-ethyl-1 -benzyl-5-(furan-2-yI)-2,4-diphenyl-4,5-dihyd ro-1 H- imidazole-4-carboxylate (ll-27): A solution of benzyl amine (0.2 g, 2.11 mmol) and 2-furylaldehyde (0.2 g, 2.11 mmol) in 50 mL of anhydrous benzene was 94 reflux under nitrogen for 3 hours and then concentrated in vacuo. The resulting residue was redissolved into 50 mL of anhydrous dichloromethane. Then 2,4- diphenyl-5(4H)-oxazolone "-2 (0.5 g, 2.11 mmol) and chlorotrimethylsilane (0.3 g, 2.74 mmol) were added and the mixture was refluxed under nitrogen for 22 hours. The solution was concentrated in vacuo and the resulting residue was resuspended in EtOAc producing a white solid precipitate (0.5 g) which was isolated via filtration. The crude solid was then dissolved into 15 mL of EtOH, treated with 1 mL of concentrated H2804, and refluxed for 21 hours. The solution was concentrated in vacuo and then dissolved into dichloromethane before being washed with saturated NaHCO2, (3 x 20 mL) and brine (1 x 20 mL). The solution was then dried over sodium sulfate and then concentrated in vacuo. The resulting crude residue was purified via column chromatography (40% EtOAc I 60% Hexanes) to afford 238 mg (25% yield) of the title compound as a yellow oil. 1H NMR (500 MHz) (CDCI3): 5 1.01 (t, J = 7 Hz, 3H), 3.86 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.94 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.88 (d, J = 15.5 Hz, 1H), 4.54 (d, J = 15.5 Hz, 1H), 5.01 (s, 1H), 6.36-6.39 (m, 2H), 6.81-6.83 (m, 2H), 7.08- 7.13 (m, 2H), 7.25-7.28 (m, 1H), 7.31-7.34 (m, 2H), 7.43-7.46 (m, 4H), 7.71-7.75 (m, 4H); 130 NMR + DEPT (125 MHz) (CDCI3): 5 13.72 (~CH3), 49.01 (-CH2), 61.27 (-CH2), 67.97 (-CH), 81.21 (quaternary —C), 109.30, (aromatic -CH), 110.50 (aromatic —CH), 126.55 (aromatic -CH), 127.08 (aromatic -CH), 127.30 (aromatic —CH), 127.43 (aromatic —CH), 128.03 (aromatic -CH), 128.39 (aromatic -CH), 128.46 (aromatic -CH), 128.68 (aromatic -CH), 130.17 (aromatic —CH), 130.65 (quaternary aromatic -C), 136.51 (quaternary aromatic - 95 C), 142.51 (aromatic -CH), 143.34 (quaternary aromatic —C), 151.41 (quaternary aromatic -C), 165.47, 170.75; IR (neat): 3063 cm", 2980 cm", 1734 cm", 1597 cm“; HRMS (ESI): mIz calcd for C29H26N203 [M+H], 451.2016; found, 451.2005. 0 yPh N F\Z—Ph Ph“: COzEt "-34 Ph (4S,SS)-ethyl-1 -benzoyl-2,4,5-tri phenyl-4,5-dihydro-1 H-imidazole-4- carboxylate (II-34): A solution of dI-(4S,58)-ethyl-2,4,5-triphenyI-4,5-dihydro- 1H-imidazoIe-4-carboxylate "-37 (100 mg, 0.27 mmol) and triethylamine (30.4 mg, 0.3 mmol) in 20 mL of anhydrous dichloromethane was treated with benzoyl chloride (45.0 mg, 0.32 mmol) and DMAP (~20 mg). The solution was stirred at room temperature for 24 hours and then washed with 2M HCI solution (2 x 20mL), saturated NaHCO3 solution (2 x 20mL), and brine solution (1 x 20mL). The solution was then dried over sodium sulfate and concentrated in vacuo. The resulting residue was purified via silica gel column chromatography using silica gel (30% ethyl acetate I 70% hexane as eluant) to afford the product as a white solid (94 mg, 73% yield). (mp 61 °- 63 °C); 1H NMR (500 MHz, CDCI3): 5 0.82 (t, 3H, J = 6.5 Hz), 3.71 (dq, J1 = 11 Hz, J2 = 7.5 Hz, 1H), 3.79 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1 H), 5.91 (s, 1H), 7.02-7.06 (m, 4H), 7.18-7.24 (m, 3H), 7.28-7.31 (m, 1H), 7.38-7.40 (m, 4H), 745-749 (m, 4H), 7.65-7.67 (m, 2H), 7.85-7.87 (m, 2H); 130 NMR (125 MHz, CDCI3 + DEPT): 5 131.41 (-CH3), 61.44 (-CH2), 74.49 (- CH), 82.86 (quaternary —C), 126.51 (aromatic —CH), 127.54 (aromatic —CH), 96 127.77 (aromatic -CH), 128.04 (aromatic -CH), 128.32 (aromatic —CH), 128.35 (aromatic —CH), 128.57 (aromatic —CH), 128.59 (aromatic —CH), 128.61 (aromatic —CH), 128.90 (aromatic -CH), 130.62 (aromatic quaternary -C), 130.80 (aromatic -CH), 131.27 (aromatic —CH), 134.62 (aromatic quaternary - C), 137.95 (aromatic quaternary -C), 140.59 (aromatic quaternary —C), 161.12, 166.76, 169.41; IR (neat): 3061 cm“, 2982 cm", 1749 cm", 1721 cm", 1599 cm‘ 1; HRMS (ESI): m/z calcd for 031H27N2O2, [M+H], 475.2022; found, 475.2028 \0 %Y\Z-Ph Ph‘ 002Et Otis dl-(4S,5$)-ethyI-2,4,5-triphenyl-1 -tosyI-4,5-dihyd ro-1 H-imidazole-d- carboxylate (ll-35): A solution of dI-(4S,5S)-ethyl-2,4,5-triphenyI-4,5-dihydro— 1H-imidazoIe-4-carboxylate "-37 (155 mg, 0.42 mmol) and triethyl amine (42.5 mg, 0.42 mmol) in 20 mL of anhydrous dichloromethane was treated with tosyl chloride (87.4 mg, 0.46 mmol) and DMAP (~10mg). The solution was stirred at room temperature for 22 hours and then washed with 2M HCI solution (2x20mL), saturated NaH003 (2x20mL), and brine (1x20mL). The solution was then dried over sodium sulfate and concentrated in vacuo. The resulting residue was purified via precipitation (CH2Cl2ll-Iexanes) to afford 160 mg (73% yield) of the title compound as a white solid. (mp. = 150 °- 151 °C) 1H NMR (500 MHz) 97 (00013): 5 0.71 (t, J = 7 Hz, 3H), 2.24 (s, 3H), 3.49 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 3.57 (dq, J1 = 10.5 Hz, .12 = 7 Hz, 1H), 5.78 (s, 1H), 6.79 (d, J = 7.5 Hz, 2H), 6.96 (d, J = 8.5 Hz, 2H), 7.23-7.36 (m, 3H), 7.37-7.44 (m, 3H), 7.49-7.56 (m, 4H), 7.57-7.60 (m, 2H), 7.89-7.91 (m, 2H); 13c NMR (125 MHz) (c0613): 5 13.29, 21.43, 61.59, 75.16, 83.26, 126.67, 127.05, 127.34, 127.47, 127.79, 128.11, 128.54, 126.61, 129.20, 129.61, 130.25, 131.58.134.74, 136.53, 142.20,144.08, 160.07, 169.04; IR (neat): 3065 cm", 3034 cm", 1736 cm", 1599 cm“; HRMS (ESI): m/z calcd for C31H28N204S [M+H], 525.1646; found, 525.1659. H1\“/;2—Ph Ph c0251 dl-(4S,5$)-ethyl-1 -acetyI-2,4,5-triphenyl-4,5-dihydro-1 H-imidazole-4- carboxylate (ll-36): A solution of dl-(4S,5S)-ethyl-2,4,5—triphenyl-4,5-dihydro— 1H-imidazole-4-carboxylate "-37 (100 mg, 0.3 mmol), acetic anhydride (32.7 mg, 0.32 mmol), EtaN (30.1 mg, 0.3 mmol) and 20 mL of anhydrous dichloromethane was treated with DMAP (~ 10 mg). The solution was stirred for 48 hours and then washed with 2M HCI (2 x 20 mL) and brine (1 x 20 mL). The solution was then dried over sodium sulfate and concentrated in vacuo. The resulting crude residue was purified via column chromatography (30% EtOAc I 70% Hexanes) to afford 93 mg (84% yield) of the title compound as a white solid. 1H NMR (500 MHz, CDCla): 6 0.75 (t, J = 7 Hz, 3H), 1.72 (s, 3H), 3.62 (dq, J1 = 10.5 Hz, J2 = 7.5 Hz, 1H), 3.70 (dq, J1 = 10.5 Hz, J2 = 7 Hz, 1H), 5.88 (s, 1H), 7.30-7.38 (m, 98 4H), 7.40—7.49 (m, 4H), 7.50-7.53 (m, 1H), 7.73 (d, J = 7 Hz, 2H), 7.78-7.80 (m, 2H); 13C NMR + DEPT (125 MHz, CDCI3): 6 13.35 (-CH3), 24.84 (-CH3), 61.41 (-CH2), 72.77 (-CH), 82.42 (quaternary -C), 126.47 (aromatic —CH), 127.36 (aromatic —CH), 127.71 (aromatic -CH), 128.24 (aromatic -CH), 128.51 (aromatic -CH), 128.57 (aromatic -CH), 128.64 (aromatic —CH), 131.13 (aromatic -CH), 131.36 (quaternary aromatic -C), 138.07 (quaternary aromatic — C), 141.04 (quaternary aromatic —C), 160.01, 167.43, 169.31; IR (neat): 3065 cm", 2982 cm", 1736 cm", 1664 cm", 1624 cm"; HRMS (ESI): m/z calcd for C23H24N203 [M+H], 413.1865; found, 413.1906. Ph',;,| F\2—Ph Pr?“ COzEt "-37 dl-(4S,58)-othyl-2,4,5-triphenyl-4,54!ihydro-1H-imidazole-4—carboxylato (ll- 37): A solution of dl-(4S,5$)-ethyl-1-benzy|-2,4,5-tripheny|-4,5-dihydro-1H- imidazole-4-carboxylate "-10 (790 mg, 1.72 mmol), 10 mL of cyclohexene and 50 mL of anhydrous THF was treated with 300 mg of 10% PdIC. The solution was stirred under reflux for 24 hours and then filtered through oelite. The resulting solution was concentrated in vacuo resulting in a yellowish crude solid. The solid was recrystallized (EtOAc/Hexanes) to afford 553 mg (87% yield) of the product as a crystalline white solid (mp. = 140 °C - 142 °C). 1H NMR (500 MHz, CDCI3): 6 0.75 (t, J = 7 Hz, 3H), 3.46-3.52 (m, 1H), 3.67 (dq, J1 = 11 Hz, J2 = 7.5, 1H), 5.53 (bs, 1H), 6.13 (bs, 1H), 7.25-7.34 (m, 6H), 7.37-7.40 (m, 2H), 7.42-7.45 (m, 99 2) 2H), 7.48-7.51 (m, 1H), 7.76 (d, J = 6 Hz, 2H), 6.00 (d, J = 7.5 Hz, 2H); 13c NMR (125 MHz, c0613): 5 13.23, 29.60, 61.61, 63.95, 126.43, 127.69, 127.72, 127.81, 127.93, 126.19, 126.30, 128.47, 129.61, 131.16, 140.04, 143.25, 162.83, 171.33; IR (neat): 3366 cm", 3063 cm", 2982 cm", 1730 cm", 1599 cm“; HRMS (ESI): m/z calcd for C24H22N202 [M+H], 371.1760; found, 371.1761. 4. Cell culture. The human cell line HeLa-NF-KB-luc was purchased from Panomics Inc (Fremont, CA). The cells were maintained in Dulbecco’s Modified Eagle’s medium (DMEM, Gibco lnvitrogen, Fredrick, MD) containing 4.5 glL glucose, 3.7 gIL bicarbonate, and supplemented with 5% fetal bovine serum, 100 UlmL penicillin, 100 pglmL streptomycin, 1 mM sodium pyruvate, 0.2 mM L-glutamine and 100 uglmL of hygromycin B (Roche). The human cell line HeLa was purchased from ATCC (Rockville, MD). The cells were maintained in DMEM media containing 4.5 glL glucose, 3.7 g/L bicarbonate, and supplemented with 10% fetal bovine serum, 100 U/mL penicillin, 100 mg/mL streptomycin, 1 mM sodium pyruvate, and 0.2 uM L-glutamine. Cells were cultured at 37 °C, 5% 002 atmosphere, 97% relative humidity and were routinely passaged by trypsin-EDTA (Life technologies, Gran Island NY) treatment to maintain a cell density between 2x105to1x106. 100 5. NF-KB-Luc reporter assay. HeLa NF-KB-luc cells (~5.0 x 105 cells/mL) were seeded into a 96 well white opaque plate using DMEM medium supplemented with 5% fetal bovine serum, 100 UlmL penicillin, 100 pg/mL streptomycin, 1 mM sodium pyruvate, 0.2 mM L- glutamine and 100 uglmL of hygromycin B. The cells were incubated for approximately 24 hours (37 °C, 5% CO2 atmosphere, 97% relative humidity). The media was then replaced with DMEM medium supplemented with 100 U/mL penicillin and 100 uglmL streptomycin. Cell cultures were pretreated with vehicle (1% DMSO), 50 pM MG-132, or imidazoline (final concentrations of 20, 10, 5, 1, 0.5, 0.1, 0.05 uM) for 30 minutes at 37 °C, 5% CO2 atmosphere with 97% relative humidity. Then TNF—a was added to a final concentration of 25 nglmL and the samples were further incubated 8 hours. The plate was then equilibrated back to room temperature and treated with 100 pL of Steady-Glo assay reagent in each well. The contents of the plate were gently stirred for 5 minutes and the luminescence of each well was measured using a Veritas microplate Iuminometer. All reported data are the average of two independent experiments. The data was normalized to TNF-a activation and the ECso values were calculated using the equation for the sigmodial curve for variable slope. lmidazolines "-8 to "-18, "-28 to “-34 and "-37 were all evaluated by Dr. Daljinder Kahlon for their ability to inhibit NF-KB mediated gene transcription using the luciferase based reporter assay in human cervical epithelial (HeLa) cells. The data obtained for the remainder of the imidazolines found within this chapter are as follows: 101 Ph N 1? Ph Ph % Fold lnductlon a) O "-5 I T I I I I I -1.5 -1.0 -0.5 0.0 0.5 1 .0 1.5 to: [II-5 (“Mil Figure "-6. Dose response curve of imidazoline "-5. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (Top—Bottom)l(1+10"((LogE050-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGEC50 0.6615 HILLSLOPE -1 .573 E050 4.587 Std. Error LOGEC50 0.06644 HILLSLOPE 0.2199 95% Confidence Intervals LOGEC50 0.5167 to 0.8063 HILLSLOPE -2.052 to -1.094 EC50 3.287 to 6.402 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.9053 Weighted Sum of Squares (1N2) 0.4776 Absolute Sum of Squares 1680 Sy.x 1 1.83 102 120- 110‘ 1004 90. 80- 70- 60- 504 40. Ph Ph ,2 lie—Pr: Ph 30- ll-6 20. 10 I T I 1 I I -1.5 -1.0 -0.5 0.0 0.5 1.0 1.5 to: [II-6 (NM)! % Fold lnductlon Figure "-7. Dose response curve of imidazoline “-6. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op—Bottom)/( 1 +1 0*((LogEC$0-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGECSO 1.041 HILLSLOPE -2.323 EC50 10.98 Std. Error LOGEC50 0.02614 HILLSLOPE 0.2636 95% Confidence Intervals LOGEC50 0.9836 to 1.098 HILLSLOPE -2.898 to -1.749 EC50 9.630 to 12.52 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.9063 Weighted Sum of Squares (1IY’) 0.1684 Absolute Sum of Squares 1170 Sy.x 9.876 103 300- 9256. p. 220°: 1 1 Ph ,2 g 150‘ I Y/ Ph — 100: I T N g ‘ I‘I Ph u. 50- 11-7 c‘ t 1 -1.5 -1'.0 -0'.5 0:0 075 1.0 1.5 WSW-”PM” Figure "-8. Dose response curve of imidazole "-7. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)l( 1 +1 0"((LogE050-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 1.442 HILLSLOPE 4.380 E050 27.67 Std. Error LOGE050 1.271 HILLSLOPE 38.93 95% Confidence Intervals LOGE050 -1.328 to 4.212 HILLSLOPE -89.20 to 80.44 E050 0.04703 to 16276 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) -0.7661 Weighted Sum of Squares (1N2) 2.353 Absolute Sum of Squares 83736 Sy.x 83.53 104 Ph ‘1 90 N \ / 1311“" 00251 11.19 cub-I GOO-3 GOO l-l-l r—H l—-I— Ph % Fold lnductlon O N O O l-I-I cT I V I V i W -1.5 -1.0 -0.5 0.0 0.5 1.0 1.5 to: [ll-1901M” Figure "-9. Dose response curve of imidazoline "-19. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)/( 1 +1 0*((LogE050-X)*HillSIope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 -16000000 HILLSLOPE 4.577e-008 E050 0.0 Std. Error LOGE050 6.437e+013 HILLSLOPE 0.1846 95% Confidence Intervals LOGE050 -140300000000000 to 1.403e+014 HILLSLOPE -0.4022 to 0.4022 E050 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) -0.04894 Weighted Sum of Squares (1IY’) 0.6642 Absolute Sum of Squares 5107 Sy.x 20.63 105 1 25- I g 100- ' ' Ph :3 > 0 Ph N g 75" Wm» . \c 2 50d Ph‘ 0028 0 IL "-20 33 25.. C I I I U fl U -1 .5 -1 .0 -0.5 0.0 0.5 1 .0 1 .5 l-OB [ll-2001M)! Figure "-10. Dose response curve of imidazoline "-20. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)/(1 +10*((LogE050-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 0.5274 HILLSLOPE -1.103 E050 3.368 Std. Error LOGE050 0.09375 HILLSLOPE 0.2016 95% Confidence Intervals LOGE050 0.3231 to 0.7317 HILLSLOPE -1.542 to -0.6633 E050 2.104 to 5.391 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.8888 Weighted Sum of Squares (1/Y’) 0.5685 Absolute Sum of Squares 2622 Sy.x 14.78 106 Ph N T2092 Ph“ 6025: 11.21 °/o Fold lnductlon m 0 ‘28 1 100 4—7 T Ph 90 ) 80 t } I l I r -1.5 -1.0 05 0.0 0.5 1.0 1.5 to: [II-21 (HM)! Figure "-11. Dose response curve of imidazoline "-21. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)l(1+1 0*((LogE050-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 1.018 HILLSLOPE -3.997 E050 10.42 Std. Error LOGE050 0.04253 HILLSLOPE 0.6514 95% Confidence Intervals LOGE050 0.9251 to 1.110 HILLSLOPE -5.417 to -2.578 E050 8.416 to 12.89 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.7016 Weighted Sum of Squares (1/Y‘) 0.7730 Absolute Sum of Squares 6579 Sy.x 23.42 107 1 25- 5 1001 E Ph g I Ph ,2 'E 75‘ i i D—me " N .- 2 50- Ph“ c0251 ,2 "-22 3 254 c I I I I I -1.5 -1.0 -o.5 0.0 0.5 1.0 1.5 to: [II-22 IIIMII Figure "-12. Dose response curve of imidazoline "-22. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)l(1 +1 0"((LogE050-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 0.5606 HILLSLOPE -2.827 EC50 3.636 Std. Error LOGE050 0.04631 HILLSLOPE 0.2267 95% Confidence Intervals LOGE050 0.4597 to 0.6615 HILLSLOPE -3.321 to -2.333 E050 2.882 to 4.587 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.8824 Weighted Sum of Squares (1le) 0.9896 Absolute Sum of Squares 2657 Sy.x 14.88 108 Ph 7 PhYN I ?—-< >— N . F Ph“ CO2Et "-23 111% l % Fold lnductlon N O I I I I I I -1.5 -1.0 -0.5 0.0 0.5 1.0 1.5 to: [II-23 (“Mil Figure "-13. Dose response curve of imidazoline "-23. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)l(1 +1 0"((LogE050-X)*HIIISlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGEC50 0.6955 HILLSLOPE -3.501 E050 4.960 Std. Error LOGE050 0.02404 HILLSLOPE 0.1814 95% Confidence Intervals LOGEC50 0.6431 to 0.7479 HILLSLOPE -3.896 to -3.106 EC50 4.396 to 5.596 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.9216 Weighted Sum of Squares (1N2) 0.3893 Absolute Sum of Squares 2316 Sy.x 13.89 109 110- 100- 901 80- 701 60- 50- 40- 30- 204 10- c I I r I I -1.5 -1.0 -0.5 0.0 0.5 1.0 1.5 we [ll-2401M” Ph W‘flfi N . I Ph“ coza "-24 Ph % Fold lnductlon Figure "-14. Dose response curve of imidazoline "-24. Equation Equation:SigmoidaI dose-response (variable slope) Y=Bottom + (T op-Bottom)l(1 +1 0"((LogE050—X)*HiIISlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGEC50 0.1865 HILLSLOPE -1 .372 E050 1.536 Std. Error LOGE050 0.1216 HILLSLOPE 0.2049 95% Confidence Intervals LOGEC50 -0.07849 to 0.4515 HILLSLOPE -1.818 to -0.9252 E050 0.8347 to 2.828 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.9176 Weighted Sum of Squares (1IY’) 1.296 Absolute Sum of Squares 1784 Sy.x 12.19 110 1 25- I c . rm) .2 1” ‘5 I PhYN g 754 ' “@615, 5 Ph‘c CO2Et 2;, 5°“ II. "a 32 254 c I I I I I I -1.5 -1.0 -0.5 0.0 0.5 1.0 1.5 Log III-25 luMll Figure "-15. Dose response curve of imidazoline II-25. Equaflon Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)l(1 +1 0"((LogE050-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 0.1328 HILLSLOPE -1.114 E050 1.358 Std. Error LOGE050 0.08592 HILLSLOPE 0.1 163 95% Confidence Intervals LOGE050 -0.05440 to 0.3200 HILLSLOPE -1.367 to -0.8605 EC50 0.8823 to 2.090 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.9193 Weighted Sum of Squares (1/Y’) 0.5104 Absolute Sum of Squares 2147 Sy.x 13.37 111 120- 1 10' 100‘ 90- 80- 70- 60- 50- 4o. 30- 20- 10- 31.5 4'0 05 0:0 0:5 1:0 1:5 Log [II-26 (uMll Br % Fold Induction Figure "-16. Dose response curve of imidazoline lI-26. Equafion Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (Top-Bottom)l(1+10"((LogEC50-X)’HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 0.1806 HILLSLOPE -0.6560 E050 1.516 Std. Error LOGE050 0.2083 HILLSLOPE 0.1962 95% Confidence Intervals LOGE050 -0.2733 to 0.6346 HILLSLOPE -1.084 to -0.2285 E050 0.5329 to 4.311 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.6390 Weighted Sum of Squares (1N2) 2.023 Absolute Sum of Squares 5723 Sy.x 21.84 112 Ph‘c c0251 ".27 =1. \ % Fold lnductlon O O cI I r I I I .1.5 -1.o -0.5 0.0 0.5 1.0 1.5 Log [II-27 (uM)] Figure “-17. Dose response curve of imidazoline "-27. Equation Equation:SigmoidaI dose-response (variable slope) Y=Bottom + (T op—Bottom)l( 1 +1 0"((LogE050-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 0.8822 HILLSLOPE 4.538 E050 7.625 Std. Error LOGE050 0.01939 HILLSLOPE 0.2486 95% Confidence Intervals LOGE050 0.8400 to 0.9245 HILLSLOPE -5.080 to -3.997 E050 6.918 to 8.404 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.9361 Weighted Sum of Squares (1IY’) 0.2804 Absolute Sum of Squares 1504 Sy.x 1 1.20 113 1509 . I i 2 i =S~ 1: 1‘0 — PhWI’N 2 I ”jg-Pb 3 50. Pb“. CO2Et :2 1135 0 . I r I I j -1.5 -1.0 -0.5 0.0 0.5 1.0 1.5 L0: [II-35 IuMII Figure "-18. Dose response curve of imidazoline "-35. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)l(1 +1 0"((LogE050-X)*HillS|ope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 1.562 HILLSLOPE -1 .288 EC50 36.47 Std. Error LOGE050 0.4649 HILLSLOPE 1 .505 95% Confidence Intervals LOGE050 0.5489 to 2.575 HILLSLOPE -4.567 to 1.992 E050 3.539 to 375.9 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) -0.004981 Weighted Sum of Squares (1/Y’) 0.9391 Absolute Sum of Squares 15189 Sy.x 35.58 114 l-I-i-I Ii: 1. o 1g: i E ‘- Tm Y 80 \2—Ph N 1311“" c0251 use PI'I 4. % Fold lnductlon c» O 0I I I I I I I -1.5 -1.0 -0.5 0.0 0.5 1.0 1.5 Log [II-36 (pM)] Figure "-19. Dose response curve of imidazoline "-36. Equation Equation:Sigmoidal dose-response (variable slope) Y=Bottom + (T op-Bottom)l( 1 +1 0*((LogEC50-X)*HillSlope)) ;X is the logarithm of concentration. Y is the response ;Y starts at Bottom and goes to Top with a sigmoid shape. ;This is identical to the "four parameter logistic equation" Sigmoidal dose-response (variable slope) Best-fit values BOTTOM 0.0 TOP 100.0 LOGE050 2.225 HILLSLOPE -0.8564 E050 167.8 Std. Error LOGE050 1.824 HILLSLOPE 1 .448 95% Confidence Intervals LOGE050 -1 .749 to 6.198 HILLSLOPE -4.010 to 2.298 E050 0.01783 to 1.579e+006 Goodness of Fit Degrees of Freedom 12 R2 (unweighted) 0.06122 Weighted Sum of Squares (1IY’) 0.3488 Absolute Sum of Squares 3321 Sy.x 16.64 115 6. Human whole blood lL-1B challenge. All experiments utilizing human whole blood were conducted by Theresa Lansdell. For further information regarding specific experimental details, please see the following publication: Kahlon, D. K.; Lansdell, T. A.; Fisk, J. S.; Tepe, J. J.; Structural-activity relationship study of highly-functionalized imidazolines as potent inhibitors of nuclear transcription factor-kappaB mediated IL-6 production. Bioorg. Med. Chem. 2009, 17, 3093-3103. 7. General procedure (whole blood assay). After obtaining the appropriate approval for de-identified human cell lines, human whole blood was obtained through the Jasper Research Clinic, Kalamazoo, MI, from a single healthy, fasted human volunteer and was collected in glass citrated tubes by venipuncture. Only samples with a white blood count falling within the normal range (4800-10,800 white blood cells per liter) were used. To support the viability of white blood cells, blood was diluted 1:10 in RPMI-1640 media supplemented with 10% fetal bovine serum, 100 U/mL penicillin, and 100 pglmL streptomycin. Aliquots of diluted blood (1 mL) were preincubated with vehicle (0.1% DMSO, final concentration) or imidazoline (final concentrations were 10, 3, 1, 0.3 and 0.1 (M) for 2 hours at 37° C, 5% CO2. lL-1B (Roche) was added to a final concentration of 200 UlmL and the samples were further incubated for 18 hours at 37 °C, 5% 002. At the end of the incubation period, the blood samples were centrifuged at 3000 RPM for 5 minutes. 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J? 88... .8800 9...... .2 $7 2%.... ova: 130 .8... 85.8825. .8 9.8% 8.22 .8... 288.“. 8 8. 8 8 8 8 88 8. 8 8 8.. 8. 88. 8.. 8. 8.8 ._ . 8.8. 8.8 8.. 8.~ 8.8 8... 8.8 8.8 8.. 8.8 8.8 8.8. 8... 8.8. J314 1‘“ 11 88... .8800 .9... frvh z 8.. x O 131 fin... 05.0328... .0 9.0on ”:22 .mn... 2:2...— U... o or on on 9. cm on on on ca 0 2. cm... on. at. on. o ..N - - 1 _! - 1 1 —4 fl 4 1 m6- md m... 9N m.» mé m... m6 m.» md ad ad m... p»....—>L#.-bb—._ppb—...—.p..rP>FrFP_.—..FL..-.._.FF_. 132 .8.. 8.00 .9... gig, IZ gm K. References (1) (2) (3) (4) (5) (6) (7) (8) Penn, R. J.; Riebsomer, J. L.; The Chemistry of the 2-lmidazolines and lmidazolidines. Chem. Rev. 1954, 54, 593-613. Acharya, A. N.; Ostresh, J. M.; Houghten, R. A.; A novel approach for solid-phase synthesis of substituted imidazolines and bis-imidazolines. J. Org. Chem. 2001, 66, 8673-8676. Concellon, J. M.; Riego, E.; Suarez, J. R.; Garcia-Granda, S.; Diaz, M. R.; Synthesis of enantiopure imidazolines through a ritter reaction of 2—(1- aminoalkyl)aziridines with nitriles. Org. Lett. 2004, 6, 4499-4501. Chem, J. W.; Liaw, Y. C.; Chen, C. S.; Rong, J. 6.; Huang, C. L.; Chan, C. H.; Wang, A. H. J.; Studies on 1,2,4-Benzothiadiazine 1,1-Dioxides-Vli and Quinazolinones-lv - Synthesis of Novel Built-in Hydroxyguanidine Tricycles as Potential Anticancer Agents. Heterocycles. 1993, 36, 1091- 1103. Vassilev, L. T.; Vu, B. T.; Graves, B.; Carvajal, D.; Podlaski, F.; Filipovic, Z.; Kong, N.; Kammlott, U.; Lukacs, C.; Klein, C.; Fotouhi, N.; Liu, E. A.; In vivo activation of the p53 pathway by small-molecule antagonists of MDM2. Science. 2004, 303, 844-848. (a) Bon, R. 8.; Hong, C. G.; Bouma, M. J.; Schmitz, R. F.; de Kanter, F. J. J.; Lutz, M.; Spek, A. L.; Orru, R. V. A.; Novel multicomponent reaction for the combinatorial synthesis of 2-imidazolines. Org. Lett. 2003, 5, 3759- 3762; (b) Gentili, F.; Bousquet, P.; Brasili, L.; Dontenwill, M.; Feldman, J.; Ghelfi, F.; Giannella, M.; Piergentili, A.; Quaglia, W.; Pigini, M.; lmidazoline binding sites (IBS) profile modulation: Key role of the bridge in determining l-1-IBS or l-2-IBS selectivity within a series of 2- phenoxymethylimidazoline analogues. J. Med. Chem. 2003, 46, 2169- 2176. _ (a) Dai, L. X.; Lin, Y. R.; Hou, X. L.; Zhou, Y. 6.; Stereoselective reactions with imines. Pure App. Chem. 1999, 71, 1033-1040; (b) Lucet, 0.; Le Gall, T.; Mioskowski, C.; The chemistry of vicinal diamines. Angew. Chem. Int. Ed. 1998, 37, 2581-2627; (c) Zhou, X. T.; Lin, Y. R.; Dai, L. X.; Sun, J.; Xia, L. J.; Tang, M. H.; A catalytic enantioselective access to optically active 2-imidazoline from N-sulfonylimines and isocyanoacetates. J. Org. Chem. 1999, 64, 1331-1334. (a) Betschart, C.; Hegedus, L. 8.; Synthesis of Azapenams, Diazepinones, and Dioxocyclams We the Photolytic Reaction of Chromium Alkoxycarbene Complexes with lmidazolines. J. Am. Chem. Soc. 1992, 114, 5010-5017; (b) Boland, N. A.; Casey, M.; Hynes, S. J.; Matthews, J. 133 (9) (10) (11) (12) (13) (14) W.; Smyth, M. P.; A novel general route for the preparation of enantiopure imidazolines. J. Org. Chem. 2002, 67, 3919-3922; (c) Botteghi, C.; Schionato, A.; Chelucci, G.; Brunner, H.; Kurzinger, A.; Obennann, U.; Asymmetric Catalysis .46. Enantioselective Hydrosilylation of Ketones with [Rh(Cod)Cl]2 and Optically-Active Nitrogen Ligands. J. Organomet. Chem. 1989, 370, 17-31. Kahlon, D. K.; Lansdell, T. A.; Fisk, J. S.; Hupp, C. D.; Friebe, T. L.; Hovde, S.; Jones, A. D.; Dyer, R. D.; Henry, R. W.; Tepe, J. J.; NF- kappaB mediated inhibition of cytokine production by imidazoline scaffolds. J. Med. Chem. 2009, 52, 1302-1309. (a) Fujioka, H.; Murai, K.; Kubo, 0.; Ohba, Y.; Kita, Y.; One-pot synthesis of imidazolines from aldehydes: detailed study about solvents and substrates. Tetrahedron. 2007, 63, 638-643; (b) Fujioka, H.; Murai, K.; Ohba, Y.; Hiramatsu, A.; Kita, Y.; A mild and efficient one-pot synthesis of 2-dihydroimidazoles from aldehydes. Tetrahedron Lett. 2005, 46, 2197- 2199; (c) lshihara, M.; Togo, H.; An efficient preparation of 2-imidazolines and imidazoles from aldehydes with molecular iodine and (diacetoxyiodo)benzene. Synlett. 2006, 227-230; (d) lshihara, M.; Togo, H.; Facile preparation of 2-imidazolines from aldehydes with ten-butyl hypochlorite. Synthesis. 2007, 1939-1942; (e) lshihara, M.; Togo, H.; Direct oxidative conversion of aldehydes and alcohols to 2-imidazolines and 2-oxazolines using molecular iodine. Tetrahedron. 2007, 63, 1474- 1480. Dghaym, R. D.; Dhawan, R.; Amdtsen, B. A.; The use of carbon monoxide and imines as peptide derivative synthons: A facile palladium-catalyzed synthesis of alpha-amino acid derived imidazolines. Angew. Chem. Int. Ed. 2001, 40, 3228. Peddibhotla, S.; Jayakumar, S.; Tepe, J. J.; Highly diastereoselective multicomponent synthesis of unsymmetrical imidazolines. Org. Lett. 2002, 4, 3533-3535. Peddibhotla, S.; Tepe, J. J.; Multicomponent synthesis of highly substituted imidazolines via a silicon mediated 1,3-dipolar cycloaddition. Synthesis. 2003, 1433-1440. (a) Mukeljee, A. K.; Azlactones - Retrospect and Prospect. Heterocycles. 1987, 26, 1077-1097; (b) Padwa, A.; Pearson, W. H., Synthetic Applications of 1,3—Dipolar Cycloaddition Chemistry Toward Heterocycles and Natural Products, 1rst edn, 2002, John VWley and Sons, Hoboken, NJ p 682-747. 134 (15) (16) (17) (13) (19) (20) (21) (22) (23) (24) (25) Huisgen, R.; Gotthard.H; Bayer, H. 0.; Schaefer, F. C.; 1,3-Dipolar Cycloadditions .56. A Convenient Synthesis of N-Substituted Pyrroles from Mesoionic Oxazolones and Alkynes. Chem. Ber. 1970, 103, 2611. Gotthardt, H.; Huisgen, R.; Schaefer, F. C.; Delta-2-Pyrroline Aus Mesoionischen Oxazolen Und Olefinen. Tetrahedron Lett. 1964, 487-491. (a) Cossio, F. P.; Ugalde, J. M.; Lopez, X.; Lecea, B.; Palomo, C.; A Semiempirical Theoretical-Study on the Formation of Beta-Lactams from Ketenes and Imines. J. Am. Chem. Soc. 1993, 115, 995-1004; (b) Funke, E.; Huisgen, R.; Ketenoid Reactivity of a Mesoionic Oxazol-S-One. Chem. Ber. 1971, 104, 3222; (c) Huisgen, R.; Funke, E.; Schaefer, F. C.; Knorr, R.; Possible Valence Tautomerism of a Mesoionic Oxazol-5-One with an Acylaminoketene. Angew. Chem. Int. Ed. 1967, 6, 367. Fu, N. Y.; Allen, A. D.; Kobayashi, S.; Tidwell, T. T.; Vukovic, S.; Arumugam, S.; Popik, V. V.; Mishima, M.; Amino substituted bisketenes: Generation, structure, and reactivity. J. Org. Chem. 2007, 72, 1951-1956. Consonni, R.; Croce, P. D.; Ferraccioli, R.; Larosa, C.; A New Approach to Imidazole Derivatives. J. Chem. Res. Synop. 1991, 188-189. Peddibhotla, S.; Tepe, J. J.; Stereoselective synthesis of highly substituted Delta(1)-pyrrolines: exo-selective 1,3-dipolar cycloaddition reactions with azlactones. J. Am. Chem. Soc. 2004, 126, 12776-12777. Shanna, V.; Tepe, J. J.; Diastereochemical diversity of imidazoline scaffolds via substrate controlled TMSCI mediated cycloaddition of azlactones. Org. Lett. 2005, 7, 5091-5094. Perkins, N. D.; The RellNF-kappa B family: friend and foe. Trends Biochem. Sci. 2000, 25, 434-40. Baldwin, A. 8., Jr.; The NF-kappa B and I kappa B proteins: new discoveries and insights. Annu. Rev. Immunol. 1996, 14, 649-83. (a) Karin, M.; Yamamoto, Y.; Wang, Q. M.; The lKK NF-kappa B system: a treasure trove for drug development. Nat. Rev. Drug Discov. 2004, 3, 17-26; (b) Pande, V.; Ramos, M. J.; NF-kappaB in human disease: current inhibitors and prospects for de novo structure based design of inhibitors. Curr. Med. Chem. 2005, 12, 357-74. (a) Kim, H. J.; Hawke, N.; Baldwin, A. S.; NF-kappa B and lKK as therapeutic targets in cancer. Cell Death and Differentiation 2006, 13, 738- 747; (b) Wahl, C.; Liptay, S.; Adler, G.; Schmid, R. M.; Sulfasalazine: a potent and specific inhibitor of nuclear factor kappa B. J. Clin. Invest. 135 (26) (27) (23) (29) (30) (31) (32) (33) (34) 1998, 101, 1163-1174; (c) Yamamoto, Y.; Gaynor, R. 8; Therapeutic potential of inhibition of the NF-kappa B pathway in the treatment of inflammation and cancer. J. Clin. Invest. 2001, 107, 135-142. Makarov, S. S.; NF-kappa B in rheumatoid arthritis: a pivotal regulator of inflammation, hyperplasia, and tissue destruction. Arthritis Res. 2001, 3, 200-206. Bon’ssenko, L.; Groll, M.; 20S proteasome and its inhibitors: Crystallographic knowledge for drug development. Chem. Rev. 2007, 107, 687-717. (a) Adams, J.; Elliott, P. J.; New agents in cancer clinical trials. Oncogene 2000, 19, 6687-6692; Feinman, R.; Gangurde, P.; Miller, 8.; Barton, 8.; Harrison, L. E.; Adams, J.; Elliott, P. J.; Siegel, D. S.; Proteasome inhibitor PS341 inhibits constitutive NF-kappa B activation and bypasses the anti- apototic bcl-2 signal in human mutiple myeloma cells. Blood. 2001, 98, 640A. Stinchcombe, T. E.; Mitchell, 8. S.; Depcik-Smith, N.; Adams, J.; Elliott, P.; Shea, T. C.; Orlowski, R. Z.; PS-341 is active in multiple myeloma: Preliminary report of a phase I trial of the proteasome inhibitor PS-341 in patients with hematologic malignancies. Blood. 2000, 96, 516A. Kane, R. C.; Farrell, A. T.; Sridhara, R.; Pazdur, R.; United States Food and Drug Administration approval summary: Bortezomib for the treatment of progressive multiple myeloma after one prior therapy. Clin. Cancer Res. 2006, 12, 2955-2960. Shirley, R. B.; Kaddour-Djebbar, l.; Patel, D. M.; Lakshmikanthan, V.; Lewis, R. W.; Kumar, M. V.; Combination of proteasomal inhibitors lactacystin and MG132 induced synergistic apoptosis in prostate cancer cells. Neoplasia. 2005, 7,1104-1111. Schow, S. R.; Joly, A.; N-acetyl-leucinyl-leucinyl-norleucinal inhibits lipopolysaccharide-induced NF-kappa B activation and prevents TNF and lL-6 synthesis in vivo. Cell. Immunol. 1997, 175, 199—202. Maryanoff, C. A.; Karash, C. B.; Turchi, I. J.; Corey, E. R.; Maryanoff, B. E.; Characterization of a Stable Carboxylic-Acid lnterrnediate from 1,3- Dipolar Cyclo-Addition of a Munchnone with 1,2-Dicyanocyclobutene. J. Org. Chem. 1989, 54, 3790-3792. Feling, R. H.; Buchanan, G. 0.; Mincer, T. J.; Kauffman, C. A.; Jensen, P. R.; Fenical, W.; Salinosporamide A: A highly cytotoxic proteasome 136 (35) (36) (37) (38) (39) (40) (41) (42) (43) (44) inhibitor from a novel microbial source, a marine bacterium of the new genus Salinospora. Angew. Chem. Int. Ed. 2003, 42, 355. Roshak, A.; Jackson, J. R.; ChabotFletcher, M.; Marshall, L. A.; Inhibition of NF kappa B-mediated interleukin-1 beta-stimulated prostaglandin E-2 formation by the marine natural product hymenialdisine. J. Pharm. Exp. Ther. 1997, 283, 955-961. Shanna, V.; Lansdell, T. A.; Peddibhotla, S.; Tepe, J. J.; Sensitization of tumor cells toward chemotherapy: Enhancing the efficacy of camptothecin with imidazolines. Chem. Biol. 2004, 11, 1689-1699. Dhawan, R.; Amdtsen, B. A.; Palladium-catalyzed multicomponent coupling of alkynes, imines, and acid chlorides: A direct and modular approach to pyrrole synthesis. J. Am. Chem. Soc. 2004, 126, 468-469. Swinney, D. C.; Xu, Y. Z.; Scarafia, L. E.; Lee, |.; Mak, A. Y.; Gan, Q. F.; Ramesha, C. S.; Mulkins, M. A.; Dunn, J.; So, 0. Y.; Biegel, T.; Dinh, M.; Volkel, P.; Barnett, J.; Dalrymple, S. A.; Lee, S.; Huber, M.; A small molecule ubiquitination inhibitor blocks NF-kappa B-dependent cytokine expression in cells and rats. J. Biol. Chem. 2002, 277, 23573-23581. Shanna, V.; Peddibhotla, S.; Tepe, J. J.; Sensitization of cancer cells to DNA damaging agents by imidazolines. J. Am. Chem. Soc. 2006, 128, 9137-9143. Piva, R.; Belardo, G.; Santoro, M. G.; NF-kappaB: A Stress-Regulated Switch for Cell Survival. Antioxid. Redox. Signal. 2006, 8, 478-86. D'Acquisto, F.; May, M. J.; Ghosh, 8.; Inhibition of Nuclear Factor Kappa B (NF-B):: An Emerging Theme in Anti-Inflammatory Therapies. Mol. Interv. 2002, 2, 22-35. Baeuerle, P. A.; Henkel, T.; Function and activation of NF-kappa B in the immune system. Annu. Rev. Immunol. 1994, 12, 141-79. Wang, C. Y.; Mayo, M. W.; Baldwin, A. S., Jr.; TNF- and cancer therapy- induced apoptosis: potentiation by inhibition of NF-kappaB. Science. 1996, 274, 784-7. (a) Mayo, M. W.; Baldwin, A. S.; The transcription factor NF-kappaB: control of oncogenesis and cancer therapy resistance. Biochim. Biophys. Acta. 2000, 1470, M55-62; (b) Greten, F. R.; Karin, M.; The lKK/NF- kappaB activation pathway-a target for prevention and treatment of cancer. Cancer Lett. 2004, 206, 193-9; (c) Wang, C. Y.; Cusack, J. C., Jr.; Liu, R.; Baldwin, A. 3., Jr.; Control of inducible chemoresistance: 137 (45) (46) (47) (48) (49) (50) (51) enhanced anti-tumor therapy through increased apoptosis by inhibition of NF-kappaB. Nat. Med. 1999, 5, 412-7; (d) Luo, J. L.; Kamata, H.; Karin, M.; lKK/NF-kappaB signaling: balancing life and death-a new approach to cancer therapy. J. Clin. Invest. 2005, 115, 2625-32; (e) de Martin, R.; Schmid, J. A.; Hofer-Warbinek, R.; The NF-kappaB/Rel family of transcription factors in oncogenic transformation and apoptosis. Mutat. Res. 1999, 437, 231-243. Perkins, N. D.; Achieving transcriptional specificity with NF-kappa B. Int. J. Biochem. Cell Biol. 1997, 29, 1433-48. (a) Baud, V.; Liu, Z. G.; Bennett, 8.; Suzuki, N.; Xia, Y.; Karin, M.; Signaling by proinflammatory cytokines: oligomerization of TRAF2 and TRAF6 is sufficient for JNK and lKK activation and target gene induction via an amino-tenninal effector domain. Genes Dev. 1999, 13, 1297-308; (b) Luo, J. L.; Kamata, H.; Karin, M.; The anti-death machinery in lKK/NF- kappaB signaling. J. Clin. Immunol. 2005, 25, 541-50; (c) Barnes, P. J.; Karin, M.; Nuclear factor-kappaB: a pivotal transcription factor in chronic inflammatory diseases. N. Engl. J. Med. 1997, 336, 1066-71. Chen, F. M. F.; Kuroda, K.; Benoiton, N. L.; Simple Preparation of 5-Oxo- 4,5-Dihydro-1,3-Oxazoles (Oxazolones). Synthesis. 1979, 230-232. Senftleben, U.; Karin, M.; The lKK/NF-kappaB pathway. Crit. Care Med. 2002, 30, $18-$26. Okazaki, Y.; Sawada, T.; Nagatani, K.; Komagata, Y.; lnoue, T.; Muto, S.; Itai, A.; Yamamoto, K.; Effect of nuclear factor-kappaB inhibition on rheumatoid fibroblast-like synoviocytes and collagen induced arthritis. J. Rheumatol. 2005, 32, 1440-1447. (a) Ardizzone, S.; Bianchi Porro, G.; Biologic therapy for inflammatory bowel disease. Drugs. 2005, 65, 2253-86; (b) Boone, D. L.; Lee, E. G.; Libby, 8; Gibson, P. J.; Chien, M.; Chan, F.; Madonia, M.; Burkett, P. R.; Ma, A.; Recent advances in understanding NF-kappaB regulation. Inflamm. Bowel Dis. 2002, 8, 201-212; (c) Schreiber, S.; Nikolaus, S.; Hampe, J.; Activation of nuclear factor kappa B inflammatory bowel disease. Gut. 1998, 42, 477-484. (a) Shanna, V.; Hupp, C. D.; Tepe, J. J.; Enhancement of chemotherapeutic efficacy by small molecule inhibition of NF-kappaB and checkpoint kinases. Curr. Med. Chem. 2007, 14, 1061-1074; (b) Haefner, B.; Targeting NF-kappaB in anticancer adjunctive chemotherapy. Cancer Treat. Res. 2006, 130, 219-245. 138 (52) (53) (54) (55) (56) (57) Balkwill, F.; Foxwell, B.; Brennan, F.; TNF is here to stay! Immunol. Today. 2000, 21, 470-1. (a) Moreland, L. W.; Inhibitors of tumor necrosis factor for rheumatoid arthritis. J. Rheumatol. 1999, 26 Suppl 57, 7-15; (b) Miagkov, A. V.; Kovalenko, D. V.; Brown, C. E.; Didsbury, J. R.; Cogswell, J. P.; Stimpson, S. A.; Baldwin, A. S.; Makarov, S. S.; NF-kappaB activation provides the potential link between inflammation and hyperplasia in the arthritic joint. Proc. Natl. Acad. Sci. U. S. A. 1998, 95, 13859-64; (c) Jimi, E.; Aoki, K.; Saito, H.; D'Acquisto, F.; May, M. J.; Nakamura, |.; Sudo, T.; Kojima, T.; Okamoto, F.; Fukushima, H.; Okabe, K.; Ohya, K.; Ghosh, S.; Selective inhibition of NF-kappa B blocks osteoclastogenesis and prevents inflammatory bone destruction in vivo. Nat. Med. 2004, 10, 617-624; (d) Miossec, P.; Cytokines in rheumatoid arthritis: is it all TNF-alpha? Cell Mol. Biol. 2001, 47, 675-8. (a) Aupperle, K.; Bennett, 8; Han, Z.; Boyle, D.; Manning, A.; Firestein, G.; NF-kappa B regulation by l kappa B kinase-2 in rheumatoid arthritis synoviocytes. J. Immunol. 2001, 166, 2705-11; (b) Han, Z.; Boyle, D. L.; Manning, A. M.; Firestein, G. S.; AP-1 and NF-kappaB regulation in rheumatoid arthritis and murine collagen-induced arthritis. Autoimmunity. 1998, 28, 197-208. Hoffrnann, A.; Baltimore, 0.; Circuitry of nuclear factor kappa B signaling. Immunol. Rev. 2006, 210, 171 -186. Webster, G. A.; Perkins, N. D.; Transcriptional cross talk between NF- kappa B and p53. Mol. Cell. Biol. 1999, 19, 3485-3495. (a) Baldwin, A. S.; Control of oncogenesis and cancer therapy resistance by the transcription factor NF-kappa B. J. Clin. Invest. 2001, 107, 241-246; (b) Hideshima, T.; Chauhan, 0.; Richardson, P.; Mitsiades, C.; Mitsiades, N.; Hayashi, T.; Munshi, N.; Dang, L.; Castro, A.; Palombella, V.; Adams, J.; Anderson, K. C.; NF—kappa B as a therapeutic target in multiple myeloma. J. Biol. Chem. 2002, 277, 16639-16647; (c) Das, K. C.; White, C. W.; Activation of NF-kappa B by antineoplastic agents - Role of protein kinase C. J. Biol. Chem. 1997, 272, 14914-14920; (d) Bottero, V.; Busuttil, V.; Loubat, A.; Magne, N.; Fischel, J. L.; Milano, 6.; Peyron, J. F.; Activation of nuclear factor kappa B through the IKK complex by the topoisomerase poisons SN38 and doxorubicin: A brake to apoptosis in HeLa human carcinoma cells. Cancer Res. 2001, 61, 7785-7791; (e) Peddibhotla, S.; Jayakumar, S.; Tepe, J. J.; Highly diastereoselective multicomponent synthesis of unsymmetrical imidazolines. Org. Lett. 2002, 4, 3533-3535; (0 Liu, S. X.; Yu, Y. Z.; Zhang, M. H.; Wang, W. Y.; Cao, X. T.; The involvement of TNF-alpha-related apoptosis-inducing ligand in the enhanced cytotoxicity of IFN-beta-stimulated human dendritic cells to 139 (53) (59) (60) (61) (62) tumor cells. J. Immunol. 2001, 166, 5407-5415; (9) Kim, J. Y.; Lee, S.; Hwangbo, B.; Lee, C. T.; Kim, Y. M.; Han, S. K.; Shim, Y. S.; Yoo, C. G.; NF-kappa B activation is related to the resistance of lung cancer cells to TNF-alpha-induced apoptosis. Biochem. Biophys. Res. Comm. 2000, 273, 140-146; (h) Weldon, C. B.; Burow, M. E.; Rolfe, K. W.; Clayton, J. L.; Jaffe, B. M.; Beckman, B. S.; NF-kappa B-rnediated chemoresistance in breast cancer cells. Surgery. 2001, 130, 143-150; (i) Arlt, A.; Vomdamm, J.; Breitenbroich, M.; Folsch, U. R.; Kalthoff, H.; Schmidt, W. E.; Schafer, H.; Inhibition of NF-kappa B sensitizes human pancreatic carcinoma cells to apoptosis induced by etoposide (VP16) or doxorubicin. Oncogene. 2001, 20, 859-868. (a) Pickart, C. M.; Eddins, M. J.; Ubiquitin: structures, functions, mechanisms. Biochim. Biophys. Acta. 2004, 1695, 55-72; (b) Karin, M.; Ben-Neriah, Y.; Phosphorylation meets ubiqutination: The Control of NF- kB activity. Ann. Rev. Immunol. 2000, 18, 621-663. Lin, Y. C.; Brown, K.; Siebenlist, U.; Activation of Nf-Kappa-B Requires Proteolysis of the Inhibitor l-Kappa-Alpha - Signal-Induced Phosphorylation of l-Kappa-B-Alpha Alone Does Not Release Active Nf- Kappa-B. Proc. Natl. Acad. Sci. U. S. A. 1995, 92, 552-556. (a) Barnes, P. J.; Karin, M.; Nuclear factor-k8 - a pivotal transcription factor in chronic inflammatory diseases. N. Engl. J. Med. 1997, 336, 1066- 1071; (b) Baeuerle, P. A.; Baichwal, V. R.; NF-kappa B as a frequent target for immunosuppressive and anti-inflammatory molecules. Adv. in Immunol, 1997, 65, 111-137; (0) Siebenlist, U.; Franzoso, G.; Brown, K.; Structure, regulation and function of NF-kB. Ann. Rev. Cell Biol. 1994, 10, 405-455. Lee, J. I.; Burckart, G. J.; Nuclear factor kappa B: important transcription factor and therapeutic target. J. Clin. Pharrnacol. 1998, 38, 981-93. (3) Dhawan, R.; Dghaym, R. D.; Amdtsen, B. A.; The development of a catalytic synthesis of Munchnones: A simple four-component coupling approach to alpha-amino acid derivatives. J. Am. Chem. Soc. 2003, 125, 1474-1475; (b) Erba, E.; Gelmi, M. L.; Pocar, D.; Trimarco, P.; Nu- Triazolines .26. 1,2,5-Trisubstituted 3-Pyrrolecarbaldehydes from N- Substituted Oxazolium-5-Olates and 5-Amino-4,5-Dihydro-4-Methylene- Nu-Triazoles. Chem. Ber. 1986, 119, 1083-1089; (c) Huisgen, R.; Funke, E.; 1,3—Cyloadditions of Mesoionic Oxazolones to Carbonyl Compounds. Angew. Chem. Int. Ed. 1967, 6, 365; (d) Siamaki, A. R.; Amdtsen, B. A.; A direct, one step synthesis of imidazoles from imines and acid chlorides: A palladium catalyzed multicomponent coupling approach. J. Am. Chem. Soc. 2006, 128, 6050-6051; (e) St Cyr, D. J.; Martin, N.; Amdtsen, B. A.; 140 (63) (64) (65) (65) (67) (68) Direct synthesis of pyrroles from imines, alkynes, and acid chlorides: An isocyanide-mediated reaction. Org. Lett. 2007, 9, 449-452. Huisgen, R.; Gotthard.H; Bayer, H. 0.; 1,3—Dipolar Cycloadditions .55. DeIta1-Pyrrolines and 7-Azabicyclo 2,2,1lHeptane Derivatives from Azlactones and Activated Alkenes. Chem. Bar. 1970, 103, 2368. (a) Campos, M. E.; Jimenez, R.; Martinez, F.; Salgado, H.; 1H-lmidazole Preparation Via Permanganate Dehydrogenation of 2-lmidazolines. Heterocycles. 1995, 40, 841-849; (b) Hughey, J. L.; Knapp, S.; Schugar, H.; Dehydrogenation of 2-Imidazolines to Imidazoles with Barium Manganate. Synthesis. 1980, 489-490; (c) Mohammadpoor—Baltork, |.; Zolfigol, M. A.; Abdollahi-Alibeik, M.; Novel, mild and chemoselective dehydrogenation of 2-imidazolines with trichloroisocyanuric acid. Synlett. 2004, 2803-2805; (d) Mohammadpoor-Baltork, I.; Zolfigol, M. A.; Abdollahi-Alibeik, M.; Novel and chemoselective dehydrogenation of 2- substituted imidazolines with potassium permanganate supported on silica gel. Tetrahedron Lett. 2004, 45, 8687-8690; (e) Parik, P.; Senauerova, S.; Liskova, V.; Handlir, K.; Ludwig, M.; Study of synthesis of 2-(2- alkoxyphenyl)-1H-imidazoles. Comparison of oxidative aromatization reactions of imidazolines. J Hetemcycl. Chem. 2006, 43, 835-841. Braddock, D. C.; Redmond, J. M.; Hermitage, S. A.; White, A. J. P.; A convenient preparation of enantiomerically pure (+)-(1R,2R)—and (-)- (1S,2$)-1,2-diamino-1,2-diphenylethanes. Adv. Synthesis Cat. 2006, 348, 911-916. (a) Matsuura, T.; Ito, Y.; Photochemical Cis-Trans lsomerization of 2,4,5- Triphenylimidazolines. Bull. Chem. Soc. Jap. 1975, 48, 3669-3674; (b) Mistryukov, E. A.; One-pot synthesis of rac-1,2—diphenylethylene-1,2- diamine. Russ. Chem. Bull. 2002, 51, 2308-2309. Kahlon, D. K.; Lansdell, T. A.; Fisk, J. S.; Tepe, J. J.; Structural-activity relationship study of highly-functionalized imidazolines as potent inhibitors of nuclear transcription factor-kappaB mediated lL-6 production. Bioorg. Med. Chem. 2009, 17, 3093-3103. (a) Sharrna, V.; Lansdell, T. A.; Peddibhotla, S.; Tepe, J. J.; Sensitization of Tumor Cells toward Chemotheraphy: Enhancing the Efficacy of Camptothecin with lmidazolines. Chem. Biol. 2004, 11, 1689-1699; (b) Shanna, V.; Peddibhotla, S.; Tepe, J. J.; Sensitization of Cancer Cells to DNA Damaging Agents by lmidazolines. J. Am. Chem. Soc. 2006, 128, 9137-9143. 141 CHAPTER III INTERMOLECULAR REACTIONS OF OXAZOL-5(4H)-ONES WITH ALKENES: FROM CYCLOADDITION TO ALKYLATION REACTIONS A. Synthesis of A1-pyrrolines via miinchnonelalkene cycloadditions Highly substituted A1-pyrroline scaffolds can be found in the structure of numerous biologically interesting natural products including myosmine, amathaspiramide E, broussonetine U1, veracintine, and many more (Figure "I- 1).1 Molecules containing A1-pyrroline scaffolds have been shown to exhibit a wide range of biological activity including anti-viral, anti-tumor and immunoactivity.2 In addition to exhibiting a wide range of biological activity, A‘- pyrrolines have also exhibited value in terms of being both biosynthetic3 and synthetic intermediates.4 Their structures contain up to three contiguous stereogenic centers and one prochiral center masked as a cyclic imine. Due to their wide range of biological activity and their synthetic usefulness, novel methods for constructing highly substituted A1-pyrrolines are still of great interest to the scientific community. 142 \ N I / N Myosmlne Amathesplramide E Figure Ill-1. Naturally occurring A1-pyrrolines. Oxazol-5(4H)-ones have been shown to be useful materials for generating a variety of nitrogen containing heterocycles by way of 1,3—dipolar cycloadditions of their munchnone tautomers.“ Until recently though, cycloaddition reactions of munchnones had not been thought of as a general method for synthesizing A'- pyrrolines. Early investigations by Gotthardt, Huisgen, and Schaefer illustrated that thermal cycloadditions of N-alkylated mtinchnones with alkenes result in the formation of a variety of products including pyrrolines and pyrroles (Scheme Ill-1)."'9 Although current mechanistic evidence suggests that these reactions proceed through the initial formation of A‘-pyrroline cycloadducts, generally only Az-pyrrolines along with pyrroles were isolated in early studies. The formation of the Az-pyrroline products are thought to come about from the decarboxylation of the primary cycloadduct followed by double bond lsomerization (Scheme lll-1).9'1o 143 ll _ R4 .. o o Fifi/\R R5 R2 R4 R2.< 5 R \ 2 O _. l @N e o ®N R5 1 R / 9 R1 3 R1/N R R1 R3 CO? L. 3 4 -COZ R4 P R4 _ R R5 .915 - NI N R (W 9R I R11 3 R1 3 J Scheme III-1. Formation of Az-pyrrolines from N-alkylated munchnones. Relatively few examples exist in the literature where primary cycloadducts from the cycloadditions of munchnones with alkenes were isolated.‘°'“ In 1989, Maryanoff and co-workers reported the isolation and characterization of a stable A1-pyrroline-5-carboxylic acid resulting from the cycloaddition of a mt‘mchnone with 1,2-dicyanocyclobutene (Scheme Ill-2).11 In contrast to previous studies of acyclic azomethine ylides cycloadditions, only the exo-cycloadduct was observed in the reaction.15 The decarboxylation of the primary cycloadduct is presumably hindered by the structural constraints of the fuse-bicyclic product. In follow up studies, the authors reported the decarboxylation of the primary cycloadduct under elevated temperatures (reflux in decalin). 144 OCH Me ,ILOZ @1138 FL H 110°C 50% Isolated CI Cl YIBId Scheme Ill-2. Isolation of a primary cycloadduct from an intermolecular cycloaddition of a mIJnchnone. Recently, milder methods for generating miinchnones using Lewis acids have been reported allowing for the isolation of the 131-pyrroline primary cycloaddition adducts from a wider range of substrates.“14 In 2004, we reported the use of AgOAc to promote flie diastereoselective synthesis of A1-pyrrolines starting from oxazol-5(4H)-ones and electron deficient alkenes (Scheme Ill-3).14 The reactions proceeded with moderate to good yields of product formation with high diastereoselectivity. As seen in Maryanoff’s studies (Scheme Ill-2), these reactions produced primarily exo—cycloadducts.‘°'11 R O R EWG 1 EWG 1 118:0 ¢\E\NG | "'EWG = N R2 AgOAc R; 0021'. Oxazol-SMI-lrones Pyrrollnes Scheme Ill-3. Lewis acid mediated cycloaddition of oxazol-5(4H)-ones with electron deficient alkenes. 8. Proposed cycloaddition of miinchnones with enol ethers To further extend the classes of heterocyclic scaffolds accessible by way of cycloadditions of mflnchnones with alkenes, we proposed to develop a mild 145 method to promote a cycloaddition reaction between oxazol-5(4H)-ones and enol ethers (Scheme Ill-4). As seen in our previous studies, we envisioned utilizing Lewis acids to generate munchnones while in the presence of the electron rich enol ethers.7"2'14 The development of such methodology would greatly benefit our diversity oriented synthesis program aimed at discovering new heterocyclic molecules for the treatment of disease by allowing access to novel A1-pyrroline scaffolds?”16 Furthermore, we envisioned using the pyrroline products as templates to access other highly substituted heterocyclic molecules. For example, proper substitution of the substituent at the 2-position (R1) of the A‘- pyrroline scaffold would allow for hydrolysis of the molecule to generate novel pyrrolidines (Scheme Ill-4).17 R1 R3 R3 0 R / R1 0 Y o 3 mom I N? ........... .> N 0R4 ________ .5 ”N 0R4 Lewis Acid COZH R COZH R2 R2 2 Scheme Ill-4. Proposed cycloaddition of oxazol-5(4H)-ones with enol ethers. The development of such methodology would allow ready accessibility to a range of biologically interesting molecules, including the natural product Lactacystin (Scheme Ill-5). Lactacystin is a pyrrolidinone-based secondary metabolite first isolated by Omura and co-workers in 1991 from the culture broth of Streptomyces sp. OM-6519.""19 Lactacystin exhibits remarkably selective and potent irreversible inhibition of the mammalian 20$ proteasome.20 It inhibits the 208 proteasome by covalently acylating the enzyme’s N-terrninal threonine residue via its corresponding B-lactone analogue, Omuralide.20 Multiple 146 syntheses along with several structure-activity relationship studies of Lactacystin have been reported.21 Even though many elegant syntheses of Lactacystin have been reported, new routes to the molecule may provide additional insight into its biological mechanism and access to more active analogues. R1 0 NHAc 3.1" : ii CO H 0 R2 R40 R2 2 (+)-Lactacyetin Scheme Ill-5. Retrosynthetic analysis of a proposed synthesis of Lactacystin using oxazol-5(4H)-ones. To the best of our knowledge, to date there are no reports of cycloadditions occurring between miinchnones derived from oxazoI-5(4H)-ones and electron rich alkenes (e.g. enol ethers). This may be in part due to the energy gap between the frontier molecular orbitals (FMOs) of the dipole and alkene involved.22 The transition states of concerted 1,3-dipolar cycloaddition reactions are governed by FMO interactions. The highest occupied molecular orbital (HOMO) of the dipole may interact with the lowest unoccupied molecular orbital (LUMO) of the alkene.22 Conversely, the HOMO of the alkene may also interact with the LUMO of the dipole.22 Sustmann and co-workers classified 1,3- dipolar cycloaddition reactions into three different categories (Types I to Ill, Figure III-2).23 In Type I 1,3-dipolar cycloadditions, the dominant FMO interaction occurs between the HOMO of the dipole and the LUMO of the alkene, whereas 147 Type in 1,3-dipolar cycloadditions proceed via a dominant interaction .of the alkene HOMO and the LUMO of the dipole. In Type II 1,3-dipolar cycloaddition reactions, the relative energies of the FMOs of the dipole and alkene are energetically similar allowing for either HOMO/LUMO interactions to occur. A ,/ .H. +1. Alkene Dipole Alkene Dipole er wmu wmm Figure Ill-2. Sustmann’s classification of 1,3-dipolar cycloaddition reactions. Traditionally azomethine ylides, such as mt'Jnchnones, are considered to be electron rich 1,3-dipoles.15 They are classified as having relatively high energy FMOs causing them to typically participate in Type I cycloadditions where the dominant FMO interaction occurs between the HOMO of the azomethine ylide and the LUMO of the dipolarphile (Figure Ill-2).15 Azomethine ylides are known to readily undergo cycloadditions with electron deficient alkenes, presumably due to a narrow HOMO/LUMO energy gap.15 Since the FMOs associated with electron rich alkenes (e.g. enol ethers) tend to be higher in energy than their electron deficient counterparts, their ability to participate in 148 Type I 1,3-dipolar cycloaddition reactions with azomethine ylides may be dismal due to a large energy difference between the FMOs involved. Recent reports in the literature illustrate precedence that 1,3-dipolar cycloaddition reactions between munchnones and enol ethers may be feasible though. In 2004, Johnson and co-workers reported a Lewis acid promoted carbon to carbon bond cleavage of aziridines to form azomethine ylides, which subsequently underwent [3 + 2] cycloaddition reactions with enol ethers to form highly substituted pyrrolidines (Scheme Ill-6).“ These 1,3-dipolar cycloaddition reactions occurred with moderate to good yields but relatively poor diastereoselectivity. They proposed their Lewis acid coordinated azomethine ylide intermediates to be extremely electron poor thus allowing it, according to the Sustrnann classification of 1,3-dipolar cycloadditions, to undergo a Type III cycloaddition.23 Ph co Et 0M8 an N'Ph ZnCIz Ph 9N 2 2‘ Me P“ N COZEt Ph cozEt T V ‘9 mcoza oluene \ ,Zn—CI c‘ OMe coza r.t. EtOzc 0 )3, Me . 69% yield d.r. =1.4:1 Scheme Ill-6. Johnson’s 1,3-dipolar cycloaddition between azomethine ylides and enol ethers. In addition, Austin and co-workers published a diastereoselective cycloaddition between vinyl ethers and isomlinchnones (Scheme III-7). The reactions occur in very high yields (82% to >98%) and with near complete diastereoselectivity.25'2° For every dipole evaluated in the study, the endo orientation of the alkoxy group of the enol ether was maintained. Furthermore, 149 the authors have also reported the use of chiral auxiliaries for conducting these reactions enantioselective with d.e.’s up to 95%.26 lsomt‘inchnones also readily undergo 1,3—dipolar cycloadditions with electron deficient alkenes illustrating that these dipoles probably proceed through a Type II cycloaddition according to the Sustrnann classification of 1,3—dipolar cycloadditions.23 O \ O R‘\®( o R" T R333 (’0 IN 0 O NH R2 0 1 Yields = 82-98% Only endo adducts observed Scheme Ill-7. Austin’s endo-selectlive 1,3-dipolar cycloaddition of isomtinchnones and enol ethers. C. Attempted cycloadditions using 2-phenyI-4-methyI-5(4H)-oxazolone and enol others We initiated our studies on the 1,3-dipolar cycloaddition between oxazol-5(4H)- ones and enol ethers by evaluating a variety of Lewis acids for their ability to promote the 1,3—dipolar cycloaddition between 2-phenyl-4-methyl—5(4H)- oxazolone Ill-1 and n-butyl vinyl ether Ill-2. Based on our previous studies involving Lewis acid promoted cycloaddition reactions of oxazol-5(4H)-ones, we chose TMSCI and AgOAc to begin our investigation (T able Ill-1). A solution of 2- phenyl-4-methyI-5(4H)-oxazolone III-1 (1 equivalent) and n-butyl vinyl ether Ill-2 (3 equivalents) was treated with either TMSCI or AgOAc (3 equivalents). The solutions were stirred for 48 hours at either room temperature or refluxing in THF and then analyzed for pyrroline formation. Unfortunately, no desired cycloaddition products were observed in any of the reactions. Most of these 150 reactions resulted in the recovery of the starting oxazol-5(4H)-one, although some decomposition of the starting enol ether could also be observed. O"Bu O =/ O" - Ph Bu Ph \« 0 III 2 \ N Lewis Acid N . COZH Sovlent 5 III-1 Entry Lewis Acid Solvent Temp.(°C) Yield No No No No Table Ill-1. Reaction of 4-methyl-2-phenyl-5(4H)-oxazolone III-1 with butyl vinyl ether III-2 in the presence of Lewis acids. D. Reversing fl'ie electronics of the reaction One possible explanation as to the failure of our initial cycloaddition attempts using oxazol-5(4H)-ones and enol ethers may lie in the energy gap between the frontier molecular orbitals involved.” As stated earlier, munchnones are considered to be electron rich 1,3-dipoles.15 They are classified as having relatively high energy FMOs causing them to typically participate in Type I cycloadditions with electron deficient alkenes where the dominant FMO interaction occurs between the HOMO of the azomethine ylide and the LUMO of the alkene (Figure Ill-3, A).15 Since the frontier molecular orbitals associated 151 with electron rich alkenes (e.g. enol ethers) tend to be relatively higher in energy, their ability to participate in 1,3-dipolar cycloadditions with azomethine ylides may be dismal due to a large energy difference between the FMOs involved (Figure III-3, B). A Energy it :1: :1: Di e .fi. 0' pol ipole Electron +i— Electron Electr ATECh All?" on . Poor ene Dipole ene Alkene A B C Figure Ill-3. Frontier molecular orbital explanation of the cycloaddition between mtlnchnones and alkenes. We hypothesized that stabilization of the mt‘mchnone species involved would lower the energy levels of its corresponding FMOs, potentially allowing it to undergo either a Type II or Type III cycloaddition with enol ethers (Figure III-3, C). We envisioned two alterations to our reaction conditions that could help facilitate the reaction. One modification would be to change the substitution pattern of the oxazol-5(4H)-one in an attempt to stabilize the dipole. For example, placing an electron withdrawing substituent at the 4-position of the oxazol-5(4H)-one scaffold would help to stabilize the anionic portion of the dipole. A second possible alteration to our initial reaction conditions for promoting the 152 cycloaddition reaction would be to screen a wider variety of Lewis acids. The strength of the Lewis acid involved in the formation of the miJnchnone species could play a major role in its stability.” Furthermore, coordination of the Lewis acid to the enol ether reactant may also help to promote these cycloadditions. E. Discovery of a novel alkylation reaction of oxazol-5(4H)-ones The first modification we made to our reaction conditions was to position an electron withdrawing substituent at the 4-positlon of the oxazol-5(4H)-one scaffold. The ester substituted oxazoI-5(4H)-one, 2-phenyl-4-carbmethoxy- 5(4H)-oxazolone Ill-5, was synthesized according to a known literature procedure.” Treatment of 2-phenyI-4-carbmethoxy-5(4H)-oxazolone III-5 with a variety of Lewis acids while in the presence of 3 equivalents of tart-butyl vinyl ether Ill-6 once again resulted in no cycloaddition product. In contrast to our earlier studies though, these reactions did not result in the recovery of the starting oxazol-5(4H)-one. Instead, we observed the formation of a diastereomeric mixture of quaternary substituted oxazolone products III-7 as illustrated in Scheme Ill-8.28 153 Ph \I’O PhYO __/ . Lewis Acid I o III-6 I t3 Nfo ; ,N G) _+ N . O U LA 8 e0 COZH cozule C02Me M 2 Ill-5 , | Ph Y0 . I ~C MeO cf 2 O‘Bu L Ill-7 Scheme III-8. Reaction of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone Ill-5 with tart-butyl vinyl ether III-6 to form the quaternary substituted oxazolone Ill-7. The quaternary substituted oxazolone Ill-7 was formed in every reaction of 2-phenyI-4-carbmethoxy-5(4H)-oxazolone Ill-5 with tart-butyl vinyl ether Ill-6 independent of the Lewis acid used. Each reaction afforded a relatively high yield of product formation with ZnClz producing the highest yield at 98% (Table III-2, entry 2). Interestingly, the diastereomeric ratio of the product mixture was approximately the same irrespective of the Lewis acid screened bringing into question the role of the Lewis acid catalyst. This prompted us to conduct the experiment of reacting 2-phenyl-4-carbmethoxy-5(4H)-oxazolone III-5 with tert- butyl vinyl ether Ill-6 without the use of any catalyst. To our delight, the same quaternary oxazolone product III-7 was formed in quantitative yield and in the same diastereomeric ratio indicating that the use of Lewis acids was not required in the reaction (Scheme Ill-2, entry 4). 154 Ph -——/ P“ 0 \fo 0 III-8 ; Y o N . . r N \8: Lewrs ACld 002Me WEE” MeOZC‘; O‘Bu Ill-5 Ill-7 Entry Lewis Acid Time D.R. Yield 1 AgOAc 24 hrs 1.3:1 73% 2 ZnCl2 36 hrs 1.2:1 98% 3 11(03u)4 10 min 1.21 70% 4 none 1 hr 1.2:1 99% Table III-2. Reaction of 2-phenyI-4-carbmethoxy-5(4H)-oxazolone Ill-5 with tert- butyl vinyl ether Ill-6 in the presence of various Lewis acids. The alkylation of 2-phenyI-4-carbmethoxy-5(4H)-oxazolone Ill-5 using tert- butyl vinyl ether Ill-6 presented us with an interesting synthetic opportunity. First of all, it presented perhaps a novel method for the alkylation of oxazoI-5(4H)- ones to form quaternary oxazolone substrates. Quaternary oxazolones are useful intermediates for the synthesis of a variety of substrates including biologically interesting d,a-disubstituted a-amino acids.‘°'29 Secondly, dependent on the scope of the reaction, we envisioned utilizing this new alkylation reaction as a key step towards the total synthesis of the Lactacystin family of molecules (Scheme Ill-9). Alkylation of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone III-5 with a higher substituted enol ether would directly result in almost the entire carbon skeleton of Lactacystin. Subsequent hydride reduction of the quaternary 155 oxazolone intermediate“O followed by oxidation would produce an amino ester intermediate ideally substituted to complete the synthesis (Scheme Ill-9). , 0 H020 I O \KILNH Aldol o: "- . C)OH Omuralide Ene-like \ Reaction 0; N: + PO’Y <1: 0 COZMO Scheme III-9. Retrosynthetic route to the synthesis of Lactacystin using an alkylation reaction of an oxazoI-5(4H)-one by an enol ether as the key step. F. Comparison to similar reactions found in literature 1. Conia-ene cyclization. Although this particular reaction appears to be novel, similar types of reactions are often referred to as ene-reactions. Ene reactions represent an atom efficient and powerful reaction for the formation of carbon- carbon bonds.31 One example of such a reaction is the Conia-ene reaction, a reaction that potentially serves as an alternative to enolate alkylations."’2 Traditionally, the Conia-ene reaction is thought of as an intramolecular ene reaction of unsaturated ketones and aldehydes, in which the carbonyl serves as the ene component via its enol tautomer (Scheme Ill-10).32 156 ll 9~ 0 / R3 / R3 3 R3 _ _l Scheme III-10. General thermal Conia-ene cyclization. Overall the Conia-ene reaction has not received as much attention in the literature as other types of ene reactions. This may be in part due to the fact that the reaction generally needs to be conducted at very high temperatures to overcome the large activation energy barrier of the reaction.33 Metal catalyzed versions of the reaction allow for lower temperatures, although enolate generation,”4 strong acid,35 or photochemical activation“5 are usually required. Recent reports have demonstrated that using catalysts such as gold,” nickel, and indiuma’8 can effectively promote Conia-type ene cyclizations under much milder conditions. More recently the first enantioselective Conia-ene cyclization reaction was reported using a Pd(ll) l Yb(IIl) dual catalyst system.39 Intermolecular versions of this type of ene reaction would greatly enhance the utility of ene reactions as an enolate alkylation alternative.40 The intermolecular alkylation of oxazoI-5(4H)-ones using enol ethers represents a possible advancement towards the development of intermolecular Conia-ene reactions.28 The reaction occurs under very mild conditions without the use of any catalyst. We hypothesized that oxazol-5(4H)-ones would be ideal substrates for the development of an intermolecular ene reaction of this nature based on the ease of formation of the aromatic enol tautomer (Scheme Ill-11). The overall transformation closely resembles that of the Conia-ene reaction, although differs 157 being an intermolecular reaction and also utilizes enol ethers as the enophile rather than alkenes and alkynes. Conia-Ena Cyclization 1 \J _. / / Intermolecular Alkyation F _ R1 0 o R 0 o-H * R 0 o \«f , 1% / V( _. 1W N N '\ / N R2 R2 1/ R2 OR OxazoI-5(4H)-one 0R3 3 Concerted? Scheme Ill-1 1. Comparison of the Conia-ene cyclization to the intermolecular alkylation of oxazol-5(4H)-ones using enol ethers. 2. The ortho-alkylation of phenols using alkenes. A second reaction illustrating similarity to the intermolecular alkylation of oxazol-5(4H)-ones using enol ethers is the ortho-alkylation of phenols using alkenes.“42 Both reactions involve the C-alkylation of substrates exhibiting high enolic character by alkenes. The ortho-alkylation of phenols using alkenes has drawn much attention from researchers due to the industrial applications of alkylated phenols.43 These reactions generally provide reaction mixtures consisting of not only ortho and para alkylated products, but also tend to produce O-alkylated products. Mechanistically, it is believed that these reactions initially produce high levels of O-alkylated intermediate followed by a series of ionic rearrangements eventually providing the final C-alkylated products.“2 These reactions may occur without the need for any catalyst, although high temperatures (260 °C to 425 °C) are 158 generally required. A variety of both homogeneous and heterogeneous catalyst systems have been developed to help both decrease the required temperature of the reactions along with improve the overall regioselectivity.41 1'- 8 1 0 .0 OH U 0 O O - O . OHO O Scheme Ill-12. The thermal ortho-alkylation of phenols using enol ethers. Pinhey and co-workers have reported the thermal ortho-alkylation of phenols using enol ethers (Scheme Ill-12).44 As compared to earlier reports using unactivated alkenes, the ortho alkylation reaction of phenol by enol ethers occurs under less extreme conditions (~150 °C). The product mixtures obtained in these studies only consisted of O-alkylated and ortho-alkylated products. The authors proposed these reactions to prowed via the initial formation of the O- alkylated ether product, which subsequently dissociates allowing for the formation of the more thermodynamically stable ortho-alkylated phenol product. The absence of para-alkylated product combined with the observation that the electronic nature of the phenol ring had negligible effects on its outcome led the 159 authors to propose that the ortho-alkylated product was produced via an ene- type mechanism. G. Solvent screening and product isolation We initiated our studies of optimizing the intermolecular alkylation of oxazoI-5(4H)-ones with enol ethers by screening various solvents. We treated 2- phenyI-4-carbmethoxy-5(4H)-oxazolone III-5 with tart—butyl vinyl ether III-6 in a variety of solvents. Reaction of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone III-5 with tort-butyl vinyl ether Ill-6 provided the desired alkylated oxazolone product in near quantitative yields using most solvents at room temperature (T able Ill-3). Less polar solvents, such as benzene, provided the highest levels of diastereoselectivity with a diastereomeric ratio of approximately 1.7 to 1 (Table Ill-3, entry 1), although little or no diastereoselectivity was observed for most solvents. More polar solvents tended to help facilitate the reaction at a faster rate with CH2C|2 producing the most rapid result at 1 hour (Table III-3, entry 2). In addition, solvents containing lone pairs of electrons tended to have longer reaction times as compared to solvents of similar polarity lacking lone electrons. For most solvents, no degradation of the enol ether was observed except in the case of DMSO where very little product was formed largely due to enol ether decomposition (Table III-3, entry 6). 160 O‘Bu =/ Ph Ph 1’" .... W10 ”f0 1.3 guiv N C Solvent 5 MeO C COzMe rt 2 0,3” III-5 III-7 Entry Solvent Tlrne d.r. Yield(%) 1 Benzene 36 hours 1.7 to 1 99 2 CH2CI2 1 hour 1.2 to 1 99 3 THF 19 hours 1.1to1 99 4 1,4-dioxane 19 hours 1.1 to 1 99 5 CH3CN 3 hours 1 to 1 99 6 DMSO-d6 1 day 1 to1 Low Table Ill-3. Screening of various solvents for the reaction of 2-phenyl-4- carbmethoxy-5(4H)-oxazolone III-5 and tart-butyl vinyl ether Ill-6. The quaternary oxazolone product III-7 proved to be moisture sensitive making characterization and separation of the two diastereomeric products relatively difficult. Therefore following the alkylation, the reaction mixture was treated with methanol to provide the more stable quaternary substituted amino malonate derivative Ill-8 (Scheme III-13). Treatment of Ill-7 with methanol overnight at room temperature produced amino malonate Ill-8 in near quantitative yield. It should be noted that we also found that treating quaternary oxazolone intermediate III-7 with sodium methoxide also provided Ill-8 in similar yields but at a much faster rate (~1 hour). 161 0 Ill-Iiu Y O‘Bu H CIW B ’ . COzMe rzt 2 MeOZC Bu NaOMe MezOZC“ 002Me 99% overall yield III-5 III-7 Ill-8 Scheme Ill-13. lnterrnolecular alkylation of 2-phenyI-4-carbmethoxy-5(4H)- oxazolone Ill-5 with tart-butyl vinyl ether III-6 followed by methanolysis. H. Scope of enol ether The methyl ester substituted oxazolone III-5 was subsequently evaluated for its reactivity with a range of substituted enol ethers. Enol ethers containing different substitution patterns were reacted with 2-phenyl-4-carbmethoxy-5(4H)- oxazolone III-5 to form quaternary oxazolone adducts, which were then analyzed after methanolysis to form the more stable amino malonate. Unsubstituted enol ethers (tart-butyl enol ether III-6 and n-butyl vinyl ether Ill-2) provided the alkylated oxazolones in approximately one hour and in near quantitative yields (Table Ill-4, entries 1 and 2). Both 2,3-dihydropyran Ill-9 and 3,4-dihydro-(2H)-pyran Ill-10 also provided the desired products in good yields, albeit with longer reaction times (Table Ill-4, entries 3 and 4, 2 and 22 hours respectively). A decrease in reactivity was noted for the higher substituted enol ethers III-11 and III-12 (I’ able III-4, entries 5 and 6), which needed to be refluxed in CH2CI2 for 48 hours to facilitate product formation. 162 o H Y o 1 Enol Ethe 1.3 N ) r( e9; 32,an 2 2) MeOH, rt Ill-5 III-8 and Ill-13 to Ill-17 Entry Enol Ether R Tlme (hrs) Temp(°C) %Yleld O‘Bu 1 do‘Bu 1 RT. 99 “a, Ill-6 Ill-8 O"Bu 2 =/0"Bu 1 RT. 98 ’111‘ Ill-2 Ill-13 3 < :0 p 2 R T 99 ‘a III-9 Ill-14 4 < :0 D 24 RT. 99 "' “a Ill-10 III-15 5 08" CB" 48 4o 68 >’4 m/‘Y Ill-11 Ill-16 o 3 _° )3 48 4o 81 a“ Ill-12 III-17 Table III-4. Reaction of 2-phenyl-4-carbmethoxy-5(4H)—oxazolone Ill-5 with enol ethers of varying substitution pattern. Enol ethers containing alkyl substituents geminal to the alkoxy group (such as 2-methoxypropene Ill-18) produced products less stable as compared to other enol ethers. For instance, treatment of 2-phenyl-4-carbmethoxy-5(4H)- oxazolone Ill-5 with 2-methoxypropene Ill-18 provided the desired alkylated 163 oxazolone product Ill-19 within minutes at room temperature (Scheme Ill-14). Interestingly, Ill-19 proved to be considerably unstable and reverted back to the starting materials under either methanolysis conditions or vacuum. Even though the products produced are fairly unstable, it is possible to obtain o—amino esters using 2-methoxypropene Ill-18 utilizing less acidic oxazol—5(4H)—ones. For example, the alkylated oxazolone obtained from the reaction of me less acidic 2,4-diphenyl-5(4H)-oxazolone "-2 with 2-methoxypropene Ill-18 smoothly converted to the desired quaternary a—amino ester derivative Ill-20 when treated with sodium methoxide. £6 Ph 0 Ph 0 H 118:0 Ill-18 Y o ”90* H, BZ,N OMe N ”‘2th 0’ MeOZC cone 002Me l'. ' M8020 OMe NaOMe Ill-5 Ill-19 Never Isolated Observed in >95% yield 118:0 Ill-18 WNI’ o NaOMe Bz’NXkOMe CHth'Z Ph 002Me Ph r. - Ph OMe "-2 Ill-20 76% overall yield Scheme Ill-14. Reactions of oxazol-5(4H)—ones with 2-methoxypropene Ill-18. We also investigated the replacement of the alkyl protection group of the enol ether some other traditional protection groups. Less substituted silyl enol ethers Ill-21 and Ill-22 were less reactive than their alkyl enol ether counterparts, but did provide the desired products in reasonable amounts with 164 yields of 83% and 48% respectively. Disappointingly, the higher substituted silyl enol ether Ill-23 provided no product formation when reacted with 2-phenyI-4- carbmethoxy-5(4H)-oxazolone Ill-5. The donating nature of the protection group of the enol ether appears to be critical to the success of these reactions. Replacement of the alkyl or silyl moiety with an electron withdrawing group, such as an acetate, completely abrogates the reaction resulting in isolation of only starting materials (Table Ill-5, entries 4 and 5). This may be in part due to the need to stabilize positive charge accumulation on the carbon adjacent to the oxygen atom during the reaction. 165 N Y}o 1) Enol Ether(1.3 eq) H ,N R 32‘ CHZC'Z M902Cc- 002MB COzMe 2) NaOMe, rt Ill-6 Ill-28 and Ill-27 Entry Enol Ether R Time (Hrs) Temp(°C) %Yleld OTIPS 1 __ 0T|PS 24 4o 83 “'1. Ill-21 Ill-28 2 OTIPS OTlPS 24 RT. 48 '11" Ill-22 Ill-27 3 OTIPS 0“” 24 4o 0 > Q\( Ill-23 4 _/OAc OA" 24 4o 0 1., Ill-24 OAC fig: 40 o 5 =< 4% 24 Ill-25 We also briefly investigated the use of alkoxy alkynes in these Treatment of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone Ill-5 with 1- Table III-5. Reaction of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone Ill-5 with enol ethers having varying protecting groups. 166 intermolecular ene-type alkylation reactions of oxazol-5(4H)-ones (Scheme III- butoxyethyne Ill-29 in dichloromethane at room temperature for 24 hours resulted in a 86% yield of alkylated oxazolone product Ill-30. Treatment of quaternary oxazolone Ill-30 with sodium methoxide unexpectedly resulted in the formation unsaturated amino ester Ill-31. Presumably Ill-31 is formed by way of a demrboxylation reaction followed by double bond isomerization. Future work in this area would entail the investigation of internally substituted alkoxy alkynes. Ph : OnBU Ph Y0 o Ill-29 Y0 o MeOH H 0'3” N = N ——x——- Bz,N CH2C|2 or COzMe r.t.. Meozc NaOMe MeOZC COzMe 86% yield 0"Bu III-5 Ill-30 Never Isolated NaOMe 88% yield 0 COzMe Ph N \ OnB“ H “H1 Scheme Ill-15. Reaction of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone Ill-5 with 1-butoxyethyne Ill-29 followed by methanolysis. l. Scope of oxazoI-5(4H)-one In order to further expand the scope of this reaction, the nature of the substituent at the R2 position of the oxazol-5(4H)-one was explored (Scheme Ill-16). Various oxazol-5(4H)-ones were reacted with tert-butyl vinyl ether Ill-6 and evaluated for quaternary oxazolone product formation. The data supports the hypothesis that increased acidity and enol character of the oxazol-5(4H)-one is helpful for the induction of this ene-type alkylation reaction. Substituents that stabilize the aromatic enol tautomer of the oxazol-5(4H)-one appear to promote the reaction 167 much more readily than those that do not. For example, acylated oxazol-5(4H)- ones Ill-5 and Ill-32 reacted readily to provide the ene—products in excellent yields at room temperature (Scheme Ill-16). th/o o flo‘Bu . P“ Y0 ”\8: Ill-6 (1.3 equrv.) N‘é R2 CH2C|2 : R; O‘BU Ill-5 R2 = COzMe m. Ill-7923A: awe Y "I-32 R2 = "L33 R2 = COCH3 COCH3 98% yield Scheme Ill-16. lnterrnolecular ene-type alkylation reaction of oxazol-5(4H)-ones Ill-5 and Ill-32 using tart-butyl vinyl ether Ill-6. Aryl substituted oxazol-5(4H)-ones also provided alkylated products in excellent yields, albeit at higher temperatures (Scheme Ill-17). Treatment of 2,4— diphenyl—5(4H)-oxazolone "-2 with tort-butyl vinyl ether Ill-6 at room temperature in dichloromethane resulted in only trace amounts of product formation at room temperature, while refluxing 4-aryloxazol-5(4H)-ones "-2 and Ill-35 with tart-butyl vinyl ether Ill-6 in toluene resulted in high yields of quaternary oxazolone products Ill-36 and Ill-37 respectively. Unfortunately, oxazol-5(4H)-ones containing alkyl substituents have not produced any desired products to date and generally result in the isolation of the starting materials (Scheme Ill-17). 168 P“ \n/o _JO‘Bu P" Y0 o — - o ”‘8: "H (3.0 equiv.) Ni/ R2 Toluene R2 reflux 03” "-2 R2 = Ph Ill-36 R2 = Ph 99% yield Ill-35 R2 = 1- Ill-37 R2 = 1-Naphthyl Naphthyl 98% yield R2 = M8 Ill-1 R2 = Me No Reaction Scheme Ill-17. Reaction of 4-aryloxazol-5(4H)-ones and 4-alkyloxazol-5(4H)- ones with tart-butyl vinyl ether Ill-6. J. Mechanistic investigation Several experiments were conducted in attempts to gain further insight about the mechanistic nature of these intermolecular ene-type alkylation reactions of oxazol-5(4H)-ones and enol ethers. We hypothesized that these reactions are likely to proceed either through a concerted mechanism or through a stepwise mechanism involving the formation of an oxonium ion intermediate. In an attempt to determine which mechanism these reactions proceed through, we proposed to conduct a deuterium labeling study utilizing a deuterium labeled enol ether. Our first deuterium labeling study involved the treatment of 2-phenyl-4- carbmethoxy-5(4H)-oxazolone Ill-5 with 5-deutero—3,4—dihydro-2H-pyran Ill-38 (Scheme Ill-18). Protonation of the enol ether by the acidic oxazol-5(4H)-one followed by condensation on to the resulting oxonium ion (path A) would result in a mixture of diastereomers, where as a concerted reaction (path 8) would be stereospecific and result in the formation of one single diastereomer. 169 Ph T 0 ° + L) cone D Path A ill-5 lll<38 Path B 6b _Ph 0 o- 1 Ph 0 (89 | :4ka \ / N \_/ D Meozc / COZMG O 2 J Ph 0 Ph 0 r . t o M8020 0 Me02C 0 D D“' MeOH MeOH M8020 CO M MeOZC CO M 2 e 2 e BZ\N O BZ\N O H H D D“' Mixture of Diastereomers Single Diastereomer Ill-39 Scheme Ill-18. Mechanistic investigation using the reaction of 2-phenyl-4- carbmethoxy-5(4H)-oxazolone Ill-5 and 5—deutero-3,4-dihydro-2H-pyran Ill-38. We utilizing a variety of NMR techniques to identify the diastereomeric ratio of the product mixture obtained from the treatment of 2-phenyl-4- carbmethoxy-5(4H)—oxazolone Ill-5 with 5-deutero-3,4-dihydro-2H-pyran Ill-38. Since the chemical shifts of the two possible product diastereomers were found 170 to be very similar, we relied on the 1H NMR integration values of Ha and Hg, (Figure Ill-4) to determine the diastereomeric ratio of the product mixture. A product mixture consisting of a diastereomeric mixture would result in equal deuterium incorporation at both H1. and Hb. On the other hand, if the reaction proceeded through a concerted mechanism, deuterium incorporation would only be observed at Hb. Prior to conducting the experiment, we conducted an HMQC experiment utilizing unlabeled quaternary amino malonate Ill-15 to identify the chemical shifts of both H, and H, (Figure Ill-4). Upon analyzing the data obtained from the HMQC experiment, the chemical shifts of H. and H, were observed to be at 1.32 ppm and 2.14 ppm respectively. 171 MeOZC 002Me 255' '27'5' '2'7'5 '255 55.5 25.5 25.5 221.5 24.5 25.5 235 2'25 225 Figure Ill-4. HMQC spectra of amino malonate III-15. Upon determining the chemical shifts of both Ha and Hb, we next sought to analyze the product mixtures obtained from treatment of 2—phenyl-4- carbmethoxy-5(4H)-oxazolone Ill-5 with the deuterium labeled enol ether Ill-38. In our first attempt, we conducted the experiment utilizing a batch of 5-deutero— 3,4-dihydro-2H-pyran Ill-38 containing only 50% deuterium (Figure Ill-5). Treatment of 2-phenyl-4-carbmethoxy—5(4H)-oxazolone III-5 with the deuterium labeled enol ether Ill-38 provided the product III-39 in near quantitative yields after methanol work-up. Upon analyzing the product mixture by 1H NMR, we found that of the signal related to H2. contained 35% deuterium incorporation and the signal from H, contained approximately 15% deuterium incorporation. This 172 initial result was inconclusive indicating the actual mechanism of the reaction beMen 2-phenyl+cerbmethoxy~5(4H)-oxazolone Ill-5 and 5—deutero—3,4- dihydro-2H-pyran Ill-38 to lie somewhere in between being totally concerted and l r / { l l . 8 d T ‘9. . “Q 'T9r'r'T*1fir'r"—1'rfirfir‘r‘r'rfrfir'rfior-rvrvr' 2.10 1.95 1.80 1.65 1.50 1.35 1.20 Figure Ill-5. 1H NMR spectra of the product mixture obtained from treatment of oxazol-5(4H)-one Ill-5 with enol ether Ill-38 containing 50% deuterium. Since the initial experiment between oxazol-5(4H)-one Ill-5 and enol ether ill-38 was inconclusive, the experiment was repeated in an attempt to gain more concrete evidence as to the mechanism of these ene-type reactions of oxazol- 5(4H)-ones with enol ethers. The second experiment was conducting utilizing a fresh batch of 5-deutero-3,4-dihydro-2H-pyran Ill-38 which contained approximately 80% deuterium (Figure Ill-6). Once again, treatment of 2-phenyl- 173 4-carbmethoxy—5(4H)-oxazolone Ill-5 with the deuterium labeled enol ether Ill-38 provided the product Ill-39 in near quantitative yields after methanol work-up. Analysis of the 1H NMR spectra provided plausible evidence of a stepwise/unconcerted mechanism with the signals from both H5. and Hi, illustrating equal amounts of deuterium incorporation. This suggests that the oxazol-5(4H)—one initially protonates the enol ether producing an oxonium ion intermediate, which is then trapped by the corresponding oxazole enolate. O In ' 1 ' to: 1 1 r - r ' r v r f I "-. r v T * r ' T r T f r 1 1 fl f' r ' r 'd1fiv—r ' 2.20 2.05 1.90 1.75 1.60 1.45 1.30 Figure Ill-6. 1H NMR spectra of the product mixture obtained from treatment of oxazol-5(4H)-one Ill-5 with enol ether III-38 containing 80% deuterium. A third deuterium labeling study was conducted to confirm the final results from the first study which indicated that these reactions proceed through a stepwise protonation type mechanism.5 For the third experiment, we synthesized 174 the deuterium labeled alkoxy alkyne Ill-40 and treated it with the 2-phenyl-4- carbmethoxy-5(4H)-oxazolone ill-5 (Scheme Ill-19). The reaction was conducted in both dichloromethane and benzene to determine if the nature of solvent could affect the reaction mechanism. In each case the reaction resulted in the formation of the of the alkylated oxazolone Ill-41 as a 1:1 mixture of diastereomers with yields of 96% and 76% respectively, confirming our prior results that these ene-type intermolecular alkylation reactions of oxazol-5(4H)- ones proceed through a stepwise process. D : OBu Ph \l/O Ph Y0 N CH2CI2 N co Me 3"“ MeO c / D 2 Benzfne 2 OBu r. 1:1 Mixture of Dlastereomers Ill-5 Ill-41 Scheme Ill-19. Reaction of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone Ill-5 with 2-deuterobutoxy ethyne Ill-40. Another piece of mechanistic insight was gained while evaluating the reactivity of 2-alkyloxazol-5(4H)-ones as compared to that of 2-aryloxazol-5(4H)- ones (Scheme Ill-20).5 To compare the reactivity of 2-aryloxazol-5(4H)-ones to that of 2-alkyloxazol-5(4H)-ones, we treated 2-ethyI-4-carbmethoxy-5(4H)- oxazolone Ill-44 with tart-butyl vinyl ether Ill-6. Interestingly, analysis of the crude reaction mixture revealed not only the formation of the desired C-alkylated product Ill-45, but also the presence of its O-alkylated regioisomer Ill-46. Furthermore, upon standing we observed the conversion of the O-alkylated oxazole intermediate III-46 to the desired C-alkylated products Ill-45 along with 175 some degradation of the enol ether component of the reaction. This observation infers that these ene-type alkylation reactions of oxazol-5(4H)-ones may proceed through an O to C migration, although this was never observed for any of the 2- aryloxazol-5(4H)-ones. O‘Bu Ph 0 =/ Ph 0 Ph 0 >__Ot3u Y o Ill-6 : \IT 0 \n’ o N CH Cl N + N / 2 2 \" O‘Bu 002Me r-t- M6020 002Me Ill-5 Ill-7 Full conversion to Not Observed C alkylated product Et _JOtB" Et Et 0 — o o >—O’Bu Y o Ill-6 : \II/ 0 Y / o N CH 0,2 N . + N 2 “ O‘Bu COzMe r.t. ””902C COzMe III-44 Ill-45 III-46 1:1 mixture 0 and C alkylated Products CDCI3 ~1 Week Et 0 \r N . O Meozc‘ O‘BU Ill-45 Convention to c alkylated product Scheme Ill-20. Comparison of the reactivity of 2-phenyl-4wcarbmethoxy-5(4H)- oxazolone Ill-5 and 2-ethyI-4-carbmethoxy-5(4H)-oxazolone Ill-44. 176 Our current understanding of the mechanism of these reactions is as summarized in Scheme Ill-21. We propose that initially the oxazol-5(4H)—one protonates the enol ether producing an oxonium ion. The resulting oxazole enolate then traps the oxonium ion forming either an O-alkylated oxazole or a C- alkylated quaternary oxazolone depending upon the steric and/or electronic nature of the starting enol ether and oxazoL5(4H)—one. Oxazole formation via 0- acylation appears to be a reversible process and may lead to C-alkylated product over time. The stability of the final C-alkylated products appears to be highly dependent on both the oxazol—5(4H)-one and enol ether used. Further studies are being conducted to further advance our knowledge of these reactions. R1 iéz 2155. R‘Y" Scheme Ill-21. Current mechanistic understanding of the alkylation of oxazol- 5(4H)-ones with enol ethers. K. In situ oxazol-5(4H)-one formation Oxazol-5(4H)-ones are moisture sensitive compounds and have the potential to undergo rapid hydrolysis to their corresponding acyclic N-acyl a—amino acids.“5 The rate at which each particular oxazoI-5(4H)-one undergoes hydrolytic ring opening is heavily dependent on its substitution pattern from both an electronic 177 and steric standpoint. Present day dehydrating reagents have allowed for the mild and efficient syntheses of oxazol-5(4H)—ones without the formation of undesired side products.46 This allows researchers to think about generating and utilizing oxazol-5(4H)-ones without the need to isolate or purify them. To this extent, we were hopeful to simplify our reactions utilizing oxazol- 5(4H)-ones and enol ethers by generating our starting oxazol-5(4H)-ones in situ. If successful this would allow us to generate products from highly hydroscopic oxazol-5(4H)-ones without the need for isolating the starting oxazol-5(4H)-one. Additionally, choosing the proper dehydrating reagent would allow the oxazol- 5(4H)-one synthesis to be conducted in the same reaction vessel as the ene-type alkylation reaction, thus saving time towards making the desired quaternary d- amino acid derivatives (Scheme Ill-22). R R1 i iozH _—.E°°' 1Y0 0 ———/0R3 W70 0 R1 E R2 N N R R2 l. 2 _l 0R3 Not Isolated Scheme Ill-22. Conducting the intermolecular alkylation reaction of oxazol-5(4H)- ones with enol ethers while generating the starting oxazol-5(4H)—one in situ. To begin our study, we chose EDCI as the cyclodehydrating reagent fearing that the use of other reagents, such as TFAA, would result in the formation of highly acidic side products detrimental to the alkylation chemistry (Table Ill-6). A solution of 2-(methoxycarbonyl)-2-(benzamido)acetic acid Ill-4 was treated with 1.1 equivalents of EDCI while in the presence of 2,3- dihydrofuran Ill-9. To our delight, this reaction resulted in the clean formation of 178 the desired alkylated oxazolone product as a mixture of diastereomers. The oxazolone product was then treated with methanol and stirred overnight to afford the product malonate Ill-14 resulting in an overall (3 steps) 91% yield of product after purification (T able Ill-6, Entry 1). We next turned our attention to other N- acyl d-amino acids whose corresponding oxazol-5(4H)-ones had previously in our hands been difficult to isolate. Both 2-(methoxycarbonyl)—2- (isobutyramido)acetic acid ill-48 and 2-(methoxycarbonyl)-2— (propionamido)acetic acid Ill-43 produced relatively high yields of product when reacted in the presence of both EDCI and 2,3—dihydrofuran Ill-9 (Table Ill-6, entries Ill-50 and Ill-49 respectively). o \ o 1 002MB 0 COzH ) Ill-9 /|L R2 R N R 500' s R‘ N O 1 H 2 CHzclz 2) MeOH Ill-4, Ill-48, Ill-43 lll-14,lll-49, Ill-50 Entry R1 R2 Temp. Yield(%) Ill-14 Ph cone RT. 91 Ill-49 iPr 002515 RT. 92 Ill-50 Et cone RT. 77 Table Ill-6. Alkylation of various amino acids with 2,3-dihydrofuran Ill-9 utilizing oxazol-5(4H)-one intermediates. Unfortunately our method for alkylating N-acyl o-amino acids with enol ethers utilizing oxazol-5(4H)-one intermediates was not amendable for the use with a wide range of enol ethers (I' able Ill-7). Reaction of 2-(methoxycarbonyl)- 179 2-(benzamido)acetic acid Ill-4 with 3,4—dihydro-2H-pyran III-10 in the presence of EDCI provided the desired alkylated product Ill-15 in 88% yield after methanolysis. All attempts using acyclic enol ethers with this procedure have been unsuccessful thus far. Both tart-butyl vinyl ether Ill-6 and n-butyl vinyl ether Ill-2 provided relatively low yields of product formation with yields of 15% and 17% respectively. The higher substituted enol ether Ill-11 did not provide any product formation (Table Ill-7, entry 2). O COZH 1)Enol Ether (1.3 eq) 0 COzMe CO Me Ph N COzMe CHZClz, rt PhJLN)Y2 2 H 2) MeOH, rt H Ill-4 ill-8 to Ill-16 Entry Enol Ether R %Yield 3‘ o 1 < O U 88 Ill-10 Ill-15 OBn 2 >=/ “AS/k o OBn Ill-11 Ill-16 OtBU (5r, OtBU 3 =/ \r 15 Ill-6 Ill-8 O"Bu O"Bu 4 =/ fir 17 III-2 Ill-13 Table Ill-7. The alkylation of 2-(methoxycarbonyl)-2-(benzamido)acetic acid Ill-4 using various enol ethers via oxazol-5(4H)-one intermediates. Hydrogen bonding of the urea side product may be to blame for the lack of product formation using acyclic enol ethers (Scheme Ill-23). Both ureas and 180 thioureas have been previously reported to increase the acidity of oxazol-5(4H)- ones by hydrogen bonding with the oxazol-5(4H)-one scaffold.47 Upon formation of the oxonium ion intermediate, it is likely that the urea by product from EDCI stabilizes the resulting oxazole enolate. Stabilization of the oxazole enolate by the urea byproduct could likely lead to the decomposition of the oxonium ion intermediate, thus leading to low product formation. O O £\NJLN}. .sNJLN/s H H F H H ' Ph Y O 1” 0“ Ph YO ”,e\‘\‘ I, \\ I . . NI / O‘H OR N / 0 @OR Decomposrtron _ , /, —-—> .o oxonlum COzMe COzMe intermedlate Scheme Ill-23. Potential hydrogen bonding interaction between the oxazol- 5(4H)-one scaffold and the urea byproduct. 181 L. Experimental 1 . General methods. Reactions were carried out in flame-dried glassware under nitrogen atmosphere. Commercial solvents and reagents were used as received. Anhydrous methylene chloride was dispensed from a delivery system which passes the solvents through a column packed with dry neutral alumina. All reactions were magnetically stirred and monitored by TLC with 0.25 pm pre-coated silica gel plates. Column chromatography was carried out on Silica Gel 60 (230-400 mesh) supplied by EM Science. Yields refer to chromatographically and spectroscopically pure compounds unless otherwise noted. Infrared spectra were recorded on a Nicolet lR/42 spectrometer. 1H and 13C NMR spectra were recorded on a Varian lnova-3OO spectrometer, a Varian Gemini-300 spectrometer and a Varian Unity Plus-500 spectrometer. Chemical shifts are reported relative to the residue peaks of the solvent CDCI3 (7.24 ppm for 1H and 77.0 ppm for 13C). HRMS were obtained at the Mass Spectrometry Facility of Michigan State University with a JEOL JMS HX-11O mass spectrometer. Gas chromatography I low resolution mass spectra were recorded on a Hewlet- Packard 5890 Series II gas chromatograph connected to a TRIO-1 El mass spectrometer. 2. Materials. Reagents and solvents were purchased from commercial suppliers and used without further purification. Anhydrous methylene chloride, benzene, acetonitrile, 182 tetrahydrofuran, and 1,4—dioxane were dispensed from a delivery system which passes the solvents through a column packed with dry neutral alumina. Anhydrous toluene and triethyl amine were distilled from calcium hydride. Trifluoroacetic anhydride and EDCI were purchased from Sigma Aldrich, checked for purity and used without further purification. 3. Procedures for synthesis of oxazol-5(4H)-ones. PhYO Q“) Ill-1 2-phenyl+methyl-5(4H)-oxazolone (Ill-1): A solution of N-benzoyl alanine (0.15 g, 0.78 mmol) in 20 mL of anhydrous dichloromethane was treated with trifluoroacetic anhydride (0.20 g, 0.93 mmol). The solution was stirred overnight and then washed with saturated sodium bicarbonate (3x50mL) and brine (1x50mL). The solution was then dried over sodium sulfate and concentrated in vacuo resulting in 135 mg (99% yield) of the title compound as a clear oil. 1H NMR (300 MHz, CDCI3) (T MS): 6 1.56 (d, J = 7.5 Hz, 3H), 4.43 (q, J = 7.5 Hz, 1H), 7.47-7.50 (m, 2H), 7.53-7.58 (m, 1H), 7.96-7.98 (m, 2H); 13C NMR (75 MHz, CDCIg) (T MS): 6 16.82, 60.99, 125.87, 127.83, 128.74, 132.70, 161.54, 178.87; IR (KBr): 2986 cm", 2937 cm", 1829 cm", 1653 cm"; LRMS (El): m/z calcd for CrngNOz, 175.2; found, 175.0. 183 O COzMe Ph N COzMe H Ill-3 Dimethyl-Z-(benzamido)malonate (Ill-3): A solution of dimethylaminomalonate hydrochloride (6.0 g, 32.7 mmol), triethylamine (9.9 g, 98.1 mmol) and anhydrous CHZCI2 (75 ml) was treated dropwise with benzoyl chloride (4.6 g, 32.7 mmol) in a flame dried round bottom flask. The solution was stirred at room temperature under nitrogen atmosphere for 5 hours. The solution was then washed exhaustively with 2M HCI solution and dried over magnesium sulfate. The solvent was removed under vacuum and the resulting crude solid was recrystallized using EtOAc l Hexanes to yield 6.4 g (77% yield) of the title compound as a white crystalline solid. 1H NMR (300 MHz) (CDCI3): 6 3.84 (s, 6H), 5.38 (d, J = 6.9 Hz, 1H), 7.12 (d, J = 6.9 Hz, 1H), 7.41-7.46 (m, 2H), 7.50- 7.55 (m, 1H), 7.81-7.84 (m, 2H); 130 NMR (75 MHz) (cools): 53.55, 100.52, 127.27, 128.66, 132.20, 132.81, 166.79; IR (neat): 3333 cm", 3009 cm“, 2957 cm", 1746 cm"; LRMS(EI): m/z calcd for C12H13N05 251.2; found, 251.1. 0 COZH Ph ” COzMe Ill-4 2-(methoxycarbonyl)-2-(benzamido)acetic acid (Ill-4): A solution of dimethyl- 2-(benzamido)malonate Ill-3 (2.0 g, 7.96 mmol) and 50 ml of methanol was cooled to 0 °C and treated dropwise with LiOH'H20 (0.33 g, 7.96 mmol) in 50 ml 184 of H20 over approximately 30 minutes. The solution was stirred over night while being allowed to warm to room temperature. The following morning the methanol was removed under vacuum and the resulting aqueous solution was first washed once with diethyl ether and then acidified using 2M HCI. The solution was washed three more times with diethyl ether and the combined ether layers were dried over magnesium sulfate. The solvent was removed under vacuum and the resulting crude solid was recrystallized in diethyl ether to yield the product as a white crystalline solid (1.75 g, 93% yield). 1H NMR (300 MHz) (DMSO): 6 7.75 (s, 3H), 5.28 (d, J = 7.5 Hz, 1H), 7.48-7.54 (m, 2H), 7.56-7.62 (m, 1H), 7.93-7.96 (m, 2H), 9.20 (d, J = 7.5 Hz, 1H), 13.53 (bs, 1H); 13C NMR (75 MHz) (CDCI3): 52.62, 56.56, 127.65, 128.37, 131.83, 133.01, 166.37, 167.59, 167.70. LRMS(EI): m/z calcd for C11H11NO5 237.2; found, 193.2 (-C02). 2-phenyl-4-carbomethoxy-5(4H)-oxazolone (ill-5): A solution of 2- (methoxycarbonyl)-2-(benzamido)acetic acid Ill-4 (2.00 g, 8.44 mmol) and 40 mL of anhydrous diethyl ether was treated with TFAA (2.13 g, 10.13 mmol). The solution was stirred for 3 hours during which time a yellow solid precipitated out of the solution. The yellow solid was filtered off and washed with cold diethyl ether. The yellow solid was then further dried under high vacuum yielding 1.77 g (96 % yield) of the title compound (m.p. = 169 °C - 170 °C). 1H NMR (500 MHz) 185 (CDClalC5D5N): 6 3.18 (s, 3H), 6.57-6.60 (m, 1H), 6.66-6.69 (m, 2H), 7.36-7.37 (m, 2H), 14.08 (bs, 1H); “C NMR + DEPT (125 MHz) (CDCI3/CstN): 6 48.95 (- COZCH3), 97.68 (quaternary C), 123.74 (aromatic —CH), 127.05 (aromatic -CH), 127.10 (aromatic —CH), 127.50 (quaternary C), 144.85 (quatemary C), 164.10 (quaternary C), 187.24 (quaternary C); IR (KBr): 2970 cm", 1777 cm“, 1830 cm' 1, 1495 cm", 1458 cm"; HRMS (FAB): m/z calcd for C11H10NO4 [M+H], 220.0609; found, 220.0609. Ill-32 (E)-4-(1-hydroxyethylidene)-2-phenyloxazol-5(4H)-one (Ill-32): A solution of sodium hippurate (12.5 g, 62.13 mmol) and acetic anhydride (25.4 9, 248.5 mmol) was heated to reflux for 20 minutes. The solution was then cooled and dissolved into 150 mL of diethyl ether. The solution was washed with water (2x50 mL) and concentrated in vacuo. The solution was then suspended in a minimal amount of diethyl ether producing a red solid which was isolated via filtration. The solid was then dried under vacuum affording 1.2 g (9.5% yield) of the title compound as a red solid. (mp. = 190 °C — 191°C) 1H NMR (500 MHz, CDCI3IPyridine-d5): 6 2.21 (s, 2H), 6.99-7.05 (m, 3H), 7.47 (d, J = 7.5 Hz, 2H), 7.85 (bs, 1H); 130 NMR (125 MHz, CDCIaleridine-ds): 6 18.47, 112.88, 125.75, 126.52, 127.83, 130.03, 152.87, 167.19, 171.69; IR (KBr): 3010 cm", 1734 cm“, 186 1701 cm", 1882 cm"; HRMS (FAB): m/z calcd for c.1H9No3 [M+H], 204.0881; found, 204.0853. 0 C02H ”A” O "1534 2-benzamido-2-(naphthalen-1-yl)acetic acid (Ill-34): A solution of 1- naphthylglycine (2.0 g, 9.93 mmol) in 30 mL of 1M NaOH was treated dropwise with benzoyl chloride (1.5 g, 10.9 mmol) at 0 °C. The solution was stirred overnight while being allowed to warm to room temperature. The solution was then washed once with EtOAc and the acidified with 2M HCI. The aqueous solution was washed again with EtOAc (3x50 mL) and the combined EtOAc washes were dried over sodium sulfate and concentrated in vacuo. The resulting crude solid was recrystallized affording 2.9 g (97% yield) of the title compound as a white solid. (mp. = 210 °C - 212 °C) 1H NMR (300 MHz, CDCI3): 6 6.30 (d, J = 7.5 Hz, 1H), 7.18-7.41 (m, 6H), 7.44-7.47 (d, J = 7 Hz, 1H), 7.65-7.68 (m, 5H), 8.12 (d, J = 8.5 Hz, 1H), 13.10 (bs, 1H); ”C NMR (125 MHz, CDCI3): 6 53.32, 123.04, 124.76, 125.27, 125.40, 126.19, 126.80, 127.82, 128.21, 128.45, 130.75, 131.08, 132.79, 133.22, 133.37, 188.17, 172.38; IR (neat): 3431 cm", 3059 cm' 1, 1724 cm“, 1851 cm"; LRMS (El): m/z calcd for C19H15N03, 305.3; found, 305.3. 187 Ill-35 2-phenyl-4-(1-naphthyl)oxazol-5(4H)-one (Ill-35): Using the general procedure, cyclodehydration of N—benzoyl-naphthylglycine Ill-34 (0.2 g, 0.66 mmol) with trifluoroacetic anhydride (0.15 g, 0.73 mmol) resulted in 0.19 g of product as a yellow solid in a 98% yield. 1H NMR (500 MHz) (CDCI3): 6 6.29 (1H, s), 7.41 (2H, m), 7.54 (3H, m), 7.63 (2H, m), 7.90 (2H, m), 8.12 (2H, dd, J1 = 5 Hz, J2 = 7 Hz), 8.18 (1 H, d, J = 8.5 Hz); 130 NMR (125 MHz) (CDCI3): 65.7, 123.6, 124.8, 125.2, 125.5, 126.2, 126.9, 128.1, 128.8, 128.9, 129.3, 129.7, 130.9, 133.2, 134.1, 183.0, 175.8; lR (neat): 3081 cm", 1828 cm", 1853 cm"; EI(LRMS) (m/z): 287. 0 (302088 N 002Me H Ill-42 Dimethyl-2-(propionamido)malonalle (III-42): A solution of dimethyl amino malonate hydrochloride (3.0 g, 16.3 mmol) and triethyl amine (5.0 g, 49.0 mmol) in 40 mL of anhydrous dichloromethane was treated dropwise with propionyl chloride (1.5 g, 16.3 mmol) at room temperature for 12 hours. The solution was then washed with 2M HCI (3x50mL) and dried over sodium sulfate. The solution was next concentrated in vacuo and the resulting crude solid was recrystallized 188 (EtOAc l Hexanes ) to afford 2.6 g (77% yield) of the title compound as a white crystalline solid. (mp. = 121°C -122 °C) 1H NMR (300 MHz) (DMSO): 8 0.97 (t, J = 7.5 Hz, 3H), 2.20 (q, J = 7.5 Hz, 2H), 3.89 (s, 6H), 5.12 (d, J = 7.5 Hz, 1H), 8.73 (d, J = 7.5 Hz, 1H); 13C NMR (75 MHz) (DMSO): 9.50, 27.88, 52.81, 55.75, 187.02, 173.28. IR (neat): 3298 cm", 2938 cm", 1740 cm", 1849 cm“, 1539 cm’ 1; HRMS (FAB): m/z calcd for C3H13NO5 [M+H], 204.0872; found, 204.0889. 0 C02H N 002Me H Ill-43 3-methoxy-3-oxo-2-propionamidopropanoic acid (III-43): A solution of dimethyl—Z—(propionamido)malonate Ill-42 (2.0 g, 9.84 mmol) in 50 mL of methanol was treated dropwise with a solution of LiOH'H20 (0.4 g, 9.84 mmol) in 50 mL of water at 0 °C. The solution was stirred overnight while being allowed to warm to room temperature. The methanol was then removed under vacuum and the resulting aqueous solution was cooled to 0 °C. The solution was then acidified with 2M HCI (pH = 2) and washed with diethyl ether (3x50mL). The combined diethyl ether washes were dried over magnesium sulfate and concentrated in vacuo. The resulting crude solid was purified via recrystallization to afford 1.6 g (84% yield) of the title compound as a white crystalline solid. 1 H NMR (300 MHz, DMSO-d“): 6 0.96 (t, J = 7.5 Hz, 3H), 2.19 (q, J = 7.5 Hz, 2H), 3.68 (s, 3H), 5.00 (d, J = 7.5 Hz, 1H), 8.58 (d, J = 7.5 Hz, 1H), 13.47 (bs, 1H); 13C NMR (75 MHz, DMSO-d“): 6 9.62, 27.73, 52.62, 56.10, 167.73, 173.29; 189 ET" | N O COzMe Ill-44 2-ethyl-4—carbomethoxy-5(4H)—oxazolone (Ill-44): Using the general procedure, TFAA (0.38 g, 1.80 mmol) was added to a suspension of 2- (methoxycarbonyl)-2-(propionamido)acetic acid (0.15 g, 0.80 mmol) in diethyl ether (3 mL), and water (16.5 mg, 0.92 mmol) was added once the reaction had been cooled. Collection of the precipitate afforded 0.11 g of the title compound (80% yield). The title compound is moisture sensitive and relatively unstable only allowing for partial characterization. 1H NMR (500 MHz) (CDCldCstN): 6 1.24 (t, J = 7.6 Hz, 3 H), 2.67 (q, J = 7.6 Hz, 2 H), 3.82 (s, 3 H), 15.83 (s, 1 H); 13C NMR + DEPT (125 MHz) (CDCI31C5D5N): 6 8.5 (~CH3), 19.8 (-CH2), 48.0 (-CH3), 181.8 (quaternary -C); IR (KBr): 3040 cm", 1789 cm", 1629 cm", 1495 cm", 1289 cm“. 4. Procedures for synthesis of enol efliers: The following enol ethers used in these studies were purchased and used as received: Ill-2 (n-butyl vinyl ether), Ill-6 (tart-butyl vinyl ether), Ill-9 (2,3- dihydrofuran), Ill-10 (3,4—dihydro—(2H)-pyran), Ill-18 (2-methoxypropene), Ill-24 (vinyl acetate) and Ill-25 (prop-1-en-2-yl acetate). All other enol ethers used in these studies were prepared as follows: 190 FM : OBn Ill-1 1 ((2-methylprop-1-enyloxy)methyl)benzene (Ill-11): A solution of AICl3 (8.7 g, 65.1 mmol) and triethylamine (13.2 9, 130.3 mmol) in 100 mL of anhydrous diethyl ether was stirred at room temperature for 2 hours. The solution was then treated with (2-methylpropane-1,1-diyi)bis(oxy)bis(methylene)dibenzene (8.8 g, 32.6 mmol) and then refluxed for 24 hours. The solution was then washed with 10% NaOH solution (2x100 mL) and dried over sodium sulfate. The solution was concentrated in vacuo and the resulting residue was purified via column Chromatography (10% diethyl ether/ 90% hexanes) to afford 4.95 g (94% yield) of the title compound as a clear oil. 1H NMR (500 MHz, CDCI3): 6 1.53 (s, 3H), 1.64 (s, 3H), 4.72 (s, 2H), 5.87 (s, 1H), 7.27-7.36 (m, 5H); 13C NMR (125 MHz, CDCI3): 6 15.09, 19.53, 73.27, 111.22, 127.30, 127.65, 128.37, 138.09, 139.70; LRMS (El): m/z calcd for C11H14O, 162.2; found, 162.1. )3. Ill-12 4-methyl-2,3-dihydrofuran (Ill-12): A solution of 3-methyldihydrofuran-2(3H)— one (0.5 g, 5.14 mmol) in 20 mL of THF was treated with DIBALH (5.65 mmol) at -78 °C. The solution was stirred for 5 hours and then quenched with 15 mL of MeOH producing a white solid. The white solid was removed via filtration. The filtrate was then concentrated in vacuo affording a crude residue. The residue was then diluted with 3 mL of quinoline and treated with 10 mg of TsOH. A 191 distillation head was placed on the flask and the solution was heated. A crude mixture of the title compound and water distilled over into a flask containing 5 mL of 2M NaOH solution which was cooled to -78 °C during the distillation. The crude mixture was then washed once with diethyl ether. The diethyl ether layer was dried using magnesium sulfate and concentrated under vacuum very carefully affording 63 mg (15% yield) of the title compound as a Clear oil. The spectroscopic data was in complete agreement with the literature data.“8 OTIPS Ill-21 Triisopropyl(vinyloxy)silane (Ill-21): Acetaidehyde (1.08 g, 24.5 mmol) was added dropwise to a solution consisting of tn'isopropylsilyl trifluoromethane suifonate (3.0 g, 9.79 mmol), triethylamine (1.49 g, 14.7 mmol) and 30 ml of anhydrous CH20|2 at 0 °C. The solution was stirred for three hours before being washed once with diluted HCI and dried over magnesium sulfate. The solvent was removed under vacuum and the resulting crude oil was purified via column chromatography (pentane) to yield 1.5 g of the desired enol ether as a Clear oil (77% yield). 1H NMR (300 MHz), CDCI3: 6 1.06 (d, J = 5.4 Hz, 18H), 1.08-1.20 (m, 3H), 4.06 (d, J = 6.3 Hz, 1H), 4.43 (d, J = 13.2 Hz, 1H), 6.50 (dd, J1 = 5.4 Hz, J2 = 13.2 Hz, 1H); 13C NMR (75 MHz) CDCI3: 12.00, 17.89, 93.98, 148.92; IR (neat): 2945 cm", 2888 cm"; LRMS(EI): m calcd for C11H24OSi 200.4; found, 200.1. 192 OTI PS Ill-22 Triisopropyl(prop-1-en-2-yloxy)silane (Ill-22): Acetone (1.42 g, 24.5 mmol) was added dropwise to a solution consisting of triisopropylsilyl trifluoromethane suifonate (3.0 g, 9.79 mmol), triethylamine (1.49 g, 14.7 mmol) and 30 ml of anhydrous CH2Cl2 at 0 °C. The solution was stirred for four hours before being washed once with brine and dried over magnesium sulfate. The solvent was removed under vacuum and the resulting crude oil was purified via column chromatography (pentane) to yield 1.77 g of the desired enol ether as a clear oil (88% yield). 1H NMR (500 MHz), CDCl3: 6 1.05 to 1.09 (m, 18H), 1.12 to 1.21 (m, 3H), 1.79 (s, 3H), 4.0 (s, 1H), 4.03 (s, 1H); 13C NMR (125 MHz) CDCI3: 12.59, 17.95, 22.74, 90.48, 158.37; IR (neat): 2945 cm", 2889 cm", 1278 cm", 1053 cm"; LRMS(EI): m/z calcd for C12H260$i 214.4; found, 214.2. : OTIPS Ill-23 triisopropyl(2-methylprop-1-enyloxy)silane (ill-23): A solution of isobutyryl aldehyde (0.5 g, 6.9 mmol) and triethylamine (1.1 g, 10.4 mmol) in 25 mL of anhydrous benzene was treated with triisopropylsilyl trifluroromethane suifonate (2.3 g, 7.62 mmol) at room temperature. After 17 hours of stirring the solution was diluted with dichloromethane (25 mL) and washed twice with 2M HCI (2x50mL). The solution was then dried over magnesium sulfate and concentrated in vacuo. The crude mixture was purified via column 193 chromatography (hexanes) to afford 1.0 g (63% yield) of the title compound as a clear oil. 1H NMR (300 MHz, CDCI3): 6 1.05—1.21 (m, 21H), 1.52 (s, 3H), 1.62 (s, 3H), 6.14 (s, 1H); 13C NMR (75 MHz, CDCI3): 6 12.01, 14.67, 17.78, 19.24, 112.38, 134.02; LRMS (El): m/z calcd for C13H2308i, 228.2; found, 228.1. 1-(2-bromovinyloxy)butane (III-28): A solution of n-butyl vinyl ether (20.0 g, 200 mmol) in 50 ml of dichloromethane was treated with a solution of bromine (31.9 g, 200 mmol) in 50 ml of dichloromethane dropwise over 1 hour at -78 °C. Then triethylamine (40.4 g, 400 mmol) was added dropwise to the reaction mixture. The solution was stirred for an additional hour at -78 °C before being allowed to warm to room temperature. The solvent was removed under vacuum and replaced with pentane. The solution was filtered to remove the resulting precipitate and the solvent was removed again under vacuum. Distillation of the crude red residue resulted in 18.7 g (52% yield) of product as a 15:1 (cisztrans) mixture of diastereomers according to H1 NMR. 1-((Z)-2-bromovinyloxy)butane: 1H NMR (500 MHz), CDCI3: 6 0.92 (t, J = 7 Hz, 3H), 1.39 (sextet, J = 7 Hz, 2H), 1.64 (p, J = 7 Hz, 2H), 3.88 (t, J = 7 Hz, 2H), 5.06 (d, J = 4 Hz, 1H), 6.56 (d, J = 4 Hz, 1H); 13C NMR (125 MHz) CDCI3: 13.71, 18.83, 31.71, 73.18, 81.91, 147.82; 1-((E)-2-bromovinyloxy)butane: 1H NMR (500 MHz), CDCI3: 6 0.92 (t, J = 7 Hz, 3H), 1.39 (sextet, J = 7 Hz, 2H), 1.64 (p, J = 7 Hz, 2H), 3.68 (t, J = 7 Hz, 2H), 5.33 (d, J = 12 Hz, 1H), 8.73 (d, J = 12 Hz, 1H); 13C NMR (125 MHz) CDCI3: 194 13.66, 18.98, 31.06, 69.60, 82.50, 150.63. HRMS (FAB): m/z calcd for C6H1208r [M+H], 179.0072; found, 179.0162. E—OBu III-29 1-(ethynyioxy)butane (Ill-29): A solution of diethylamine (5.92 g, 80.9 mmol) in 75 ml of THF was treated with a 2M solution of n-BuLi in pentane at 0 °C. The solution was stirred for 10 mins and then 1-(2-bromovinyloxy)butane Ill-28 (5.0 g, g 27.9 mmol) was added dropwise over 10 mins. The solution was stirred for 30 mins at 0 °C at which time the volatiles were removed at reduced pressure. The resulting lithium salts were cooled to -78 °C and 75 ml of brine solution was added as quickly as possible making sure to continually swirl the flask contents to reduce freezing rate. The aqueous solution was then extracted with decahydronaphthalene (3 x 30 ml) and the combined decahydronaphthalene . fractions were dried over magnesium sulfate. The organic solution was then filtered and the product was purified by vacuum distillation resulting in 1.41 g (51% yield) of product as a clear oil. 1H NMR (500 MHz), CDCI3: 6 0.92 (t, J = 7.5 Hz, 3H), 1.40 (s, J = 7.5 Hz, 2H), 1.49 (s, 1H), 1.71 (p, J = 7 Hz, 2H), 4.05 (t, J = 6.5 Hz, 2H); 13C NMR (125 MHz) CDCI3: 13.53, 18.50, 26.00, 30.59, 78.70, 91.23. 195 ,f) Ill-38 5-deutero-3,4-dihydro-2H-pyran (Ill-38): A solution of 3,4—dihydro-(2H)—pyran (15.0 9, 178.3 mmol) and 24 mL of MeOD was treated with 15 mg of TsOH. The solution was refluxed for 12 hours and concentrated in vacuo. The resulting residue was redissolved into 24 mL of MeOD and refluxed for an additional 12 hours. Then 10mL of quinoline was added and a distillation head was then placed on the flask. The solution was heated distilling over a crude mixture of MeOH and the title compound. The crude mixture was purified via an additional distillation affording 3.3 g (21% yield, 70% deuterium incorporation) of the title compound as a clear oil. (b.p. = 87 °C) 1H NMR (500 MHz, CDCI3): 6 1.83 (p, J = 6.5 Hz, 2H), 1.92 (t, J = 6.5 Hz, 2H), 3.93 (t, J = 6 Hz, 2H), 4.63-4.65 (m, 0.3H, (70% 0)), 8.32 (d, J = 4.5 Hz, 1H); 13C NMR + DEPT (125 MHz, CDCI3): 8 19.38, 22.70, 65.74, 100.41 (t, J = 24.6 Hz), 144.02; HRMS (FAB): m/z calcd for C5H30D [M+H], 86.0716; found, 86.0715. D I: OBu Ill-40 2-deutero-1-butoxyethyne (III-40): A solution of diethylamine (5.92 g, 80.9 mmol) in 75 ml of THF was treated with 32.1 mL of a 2M solution of n-BuLi in pentane at 0 °C. The solution was stirred for 10 minutes and then 1-(2- bromovinyloxy)butane Ill-28 (5.0 g, 27.9 mmol) was added dropwise over 10 minutes. The solution was stirred for 30 minutes at 0 °C after which time the 196 volatiles were removed at reduced pressure. The resulting lithium salts were cooled to -78 °C and 80 ml of 020 was added as quickly as possible making sure to continually swirl the flask contents to reduce freezing rate. The aqueous solution was then extracted with decahydronaphthalene (3 x 30 ml) and the combined decahydronaphthalene fractions were dried over magnesium sulfate. The organic solution was then filtered and the product was purified by vacuum distillation resulting in 1.17 g (42% yield) of the title compound with 78% deuterium incorporation as a clear oil. 1H NMR (500 MHz)(CDC|3): 6 0.92 (t, J = 7.5 Hz, 3H), 1.40 (sextet, J = 7.5 Hz, 2H), 1.49 (s, 0.22H), 1.71 (p, J = 7 Hz, 2H), 4.05 (t, J = 6.5 Hz, 2H); 13C NMR (125 MHz) (CDCI3): 13.53, 18.50, 26.00, 30.59, 78.70, 90.9 (t, J = 9.1 Hz); HRMS (FAB): m/z calcd for CanOD [M + H], 100.0872; found, 100.0872. 5. Alkylation reactions of oxazol-5(4H)-ones and enol ethers. Ph 0 Ph 0 31¢) T o N 5 M80 C" MeO C‘ - 2 O‘Bu 2 ill-7a Ill-7b Methyl-4-(1-tert-butoxyethyI)-5-oxo-2-phenyl4.5-dihydrooxazoIe-4- carboxylate (Ill-7): A solution of 4—carbmethoxy—2-phenyl-5(4H)—oxazolone (55 mg, 0.25 mmol) Ill-5 in 20 mL of dichloromethane was treated with tert-butyl vinyl ether ill-6 at room temperature for 1 hour. The solution was then concentrated in vacuo affording 80 mg (99% yield) of the title compound as a clear oil. Ill-7a: 1H NMR (500 MHz, CDCI3): 6 1.10 (s, 9H), 1.37 (d, J = 6 Hz, 3H), 3.77 (s, 3H), 4.62 197 (q, J = 8 Hz, 1H), 7.44-7.47 (m, 2H), 7.54-7.57 (m, 1H), 8.03-8.05 (m, 2H); 13C NMR + DEPT (125 MHz, CDCI3): 6 18.92 (-CH3), 28.50 (-CH3), 53.43 (-CH3), 70.79 (-CH), 75.21 (quaternary —C), 81.98 (quaternary —C), 125.46 (aromatic quaternary —C), 128.33 (aromatic —CH), 128.65 (aromatic -CH), 132.99 (aromatic -CH), 163.22, 165.24, 173.20; III-7b: 1H NMR (500 MHz, CDCI3): 6 1.04 (s, 9H), 1.39 (d, J = 6.5 Hz, 3H), 3.78 (s, 3H), 4.42 (q, J = 6.5 Hz, 1H), 7.44- 7.47 (m, 2H), 7.54~7.57 (m, 1H), 7.99-8.01 (m, 2H); 13C NMR + DEPT (125 MHz, CDCl3): 6 17.23 (-CH3), 28.46 (-CH3), 53.29 (-CH3), 70.31 (-CH), 74.99 (quaternary —C), 80.49 (quaternary —C), 125.34 (aromatic quaternary -C), 128.14 (aromatic -CH), 128.79 (aromatic -CH), 133.05 (aromatic -CH), 163.15, 165.64, 171.79; IR (neat): 2978 cm“, 1830 cm", 1751 cm", 1653 cm“; 6. General procedure for alkylation reaction: A solution of oxazolone (0.5 mmol) and enol ether (0.6 mmol to 1.5 mmol) in dry dichloromethane was stirred under nitrogen atmosphere in a flame dried flask for the requisite amount of time as monitored by TLC. The solvent was removed under vacuum and replaced with MeOH. Completion of oxazolone ring opening was monitored by TLC. The solvent was removed under vacuum and the product was isolated if necessary by column chromatography on silica gel with an ethyl acetate I hexanes mixture. 198 BzHN O‘Bu Ill-8 Dimethyl-241-tert-butoxyethyl)-2-(benzamido)malonate (Ill-8): Using the general procedure, a solution of 2-phenyl-4-carbmethoxy-5(4H)—oxazolone (0.1 g, 0.46 mmol) and tert-butyl vinyl ether (0.06 g, 0.59 mmol) in 20 mL of anhydrous CHZCI2 was stirred at room temperature for 30 min. The resulting ene adduct was stirred in 20 mL MeOH overnight, after which 0.16 g of malonate ill-8 was obtained (99 % yield) as a Clear oil. 1H NMR (300 MHz), CDCI3: 6 1.10 (s, 9H), 1.36 (d, J = 6 Hz, 3H), 3.73 (s, 3H), 3.77 (s, 3H), 4.58 (q, J = 6, 1H), 7.23 (bs, 1H), 7.38-7.58 (m, 3H), 7.78-7.82 (m, 2H); 13C NMR + DEPT (75 MHz) CDCI3: 19.23 (-CH3), 28.84 (-CH3), 52.74 (-C02CH3), 53.20 (-C02CH3), 70.59 (quaternary C), 71.52 (-CH), 74.43 (quaternary C), 127.20 (aromatic CH), 128.56 (aromatic CH), 131.79 (aromatic CH), 133.75 (aromatic quaternary C), 166.41, 166.62, 168.14; IR (cm’1): 3430, 2979, 1748, 1676; HRMS (FAB): m/z calcd for C13H26N05 [M + H], 352.1757; found, 352.1760. BzHN O"Bu Ill-13 Dimethyl-2-(benzamido)-2-(1-butoxyethyl)malonate (III-13): Using the general procedure, a solution of 2-phenyI-4-carbmethoxy-5(4H)-oxazolone (0.1 g, 0.46 mmol) and n-butyl vinyl ether (0.06 g, 0.59 mmol) in 20 mL of anhydrous 199 CH2CI2 was stirred at room temperature for 30 min. The resulting ene adduct was stirred in 20 mL MeOH overnight, after which 0.16 g of malonate Ill-13 was obtained (98 % yield) as a clear oil. 1H NMR (300 MHz), CDCI3: 6 0.85 (t, J = 7 Hz, 3H), 1.22-1.34 (m, 2H), 1.32 (d, J = 6 Hz, 3H), 1.37-1.48 (m, 2H), 3.16-3.24 (m, 1H), 3.51-3.59 (m, 1H), 3.73 (s, 3H), 3.81 (s, 3H), 4.33 (q, J = 6 Hz, 1H), 7.32 (bs, 1H), 7.40-7.54 (m, 3H), 7.80-7.83 (m, 2H); 13C NMR + DEPT (75 MHz) CDCI3: 13.76 (-CH3), 14.78 (-CH3), 19.21 (-CH2), 31.84 (—CH2), 52.83 (- COZCH3), 53.40 (-C02CH3), 68.95 (-CH2), 69.90 (quaternary C), 78.09 (-CH), 127.20 (aromatic CH), 128.55 (aromatic CH), 131.87 (aromatic CH), 133.44 (aromatic quaternary C), 166.33, 188.44, 168.08; IR (cm'1): 3424,2957, 1747, 1676; HRMS (FAB): m/z calcd for C13H25N06 [M + H], 352.1759; found, 352.1760. M9020 COzMe Bsz O Ill-14 Dimethyl-Z-(benzamido)-2-(tetrahydrofuran-Z-yl)malonate (ill-14): Using the general procedure, a solution of 2-phenyl-4-carbmethoxy-5(4H)-oxazolone (0.1 g, 0.46 mmol) and 2,3-dihydrofuran (0.04 g, 0.59 mmol) in 20 mL of anhydrous CH2CI2 was stirred at room temperature for 2 hr. The resulting ene adduct was stirred in 20 mL MeOH overnight, after which 0.14 g of malonate III-14 was obtained (99 % yield) as a clear oil. 1H NMR (300 MHz), CDCI3: 6 1.70-1.91 (m, 2H), 2.07-2.22 (m, 1H), 2.19-2.32 (m, 1H), 3.67-3.82 (m, 2H), 3.76 (s, 3H), 3.84 (s, 3H), 4.72 (t, J = 7 Hz, 1H), 7.38 (bs, 1H), 7.38-7.54 (m, 3H), 7.80-7.84 (m, 200 2H); 130 NMR + DEPT (75 MHz) CDCI3: 26.01 (-CH2), 27.40 (-CH2), 52.94 (- C02CH3), 53.77 (-COZCH3), 68.54 (quaternary C), 68.86 (-CH2), 81.47 (-CH), 127.18 (aromatic CH), 128.63 (aromatic CH), 132.02 (aromatic CH), 133.27 (aromatic quaternary C), 166.43, 166.53, 168.08; IR (cm“): 3414, 2955, 1743, 1670; HRMS (FAB): m/z calcd for C16H20N05 [M + H], 322.1290; found, 322.1290. 3.1.340 o III-15 Dimethyl-2-(benzamido)-2-(tetrahydropyran-2-yl)malonate (Ill-15): Using the general procedure, a solution of 2-phenyl-4—carbmethoxy-5(4H)-oxazolone (0.1 g, 0.46 mmol) and 3,4-dihydro-2H-pyran (0.05 g, 0.59 mmol) in 20 mL of anhydrous CH2CI2 was stirred at room temperature for 22 hr. The resulting ene adduct was stirred in 20 mL MeOH overnight yielding 0.14 g of malonate Ill-15 (99 % yield) as a clear colorless oil after silica gel chromatography (20% ethyl acetate / 80% hexanes). 1H NMR (300 MHz), CDCI3: 8 1.26-1.64 (m, 4H), 1.80- 1.86 (m, 1H), 2.10-2.18 (m, 1H), 3.38-3.48 (m, 1H), 3.73 (s, 3H), 3.82 (s, 3H), 3.90-3.96 (m, 1H), 4.24 (dd, J = 12 Hz, J = 2 Hz, 1H), 7.39 (bs, 1H), 7.40-7.54 (m, 3H), 7.82-7.86 (m, 2H); 13C NMR + DEPT (75 MHz) CDCI3: 22.97 (-CH2), 25.71 (-CH2), 26.78 (-CH2), 52.82 (-COZCH3), 53.69 (-COZCH3), 69.14 (quaternary C), 69.42 (-CH2), 80.99 (-CH), 127.22 (aromatic CH), 128.54 (aromatic CH), 131.90 (aromatic CH), 133.33 (aromatic quaternary C), 165.99, 201 166.41, 168.16; IR (cm'1): 3422, 2953, 1745, 1674; HRMS (FAB): m/Z calcd for CnHngOe [M + H], 338.1445; found, 338.1447. OBn BzHN Ill-16 Dimethyl-Z-(benzamido)-2-(1-benzyloxy-2-methylpropyl)malonatle (Ill-16): Using the general procedure, a solution of 2-phenyl-4-carbmethoxy-5(4H)- oxazolone (0.1 g, 0.46 mmol) and 1-benzyloxy-2-methylpropene (0.096 g, 0.59 mmol) in 20 mL of anhydrous CHZClz was heated to reflux for 48 hours. The resulting ene adduct was stirred in 20 mL MeOH overnight yielding 0.13 g of malonate Ill-16 (68 % yield) as a clear colorless oil after silica gel chromatography (20% ethyl acetate / 80% hexanes). 1H NMR (300 MHz), CDCI3: 6 0.92 (d, J = 7 Hz, 3H), 1.15 (d, J = 7 Hz, 3H), 2.37-2.45 (m, 1H), 3.70 (s, 3H), 3.77 (s, 3H), 4.44 (d, J = 2 Hz, 1H), 4.59 (d, J = 11 Hz, 1H), 4.68 (d, J = 11 Hz, 1H), 7.24-7.51 (m, 9H), 7.77-7.80 (m, 2H); 13C NMR + DEPT (75 MHz) CDCI3: 16.23 (-CH3), 23.87 (-CH3), 29.78 (-CH), 53.08 (-COZCH3), 53.39 (- CO2CH3), 70.09 (quaternary C), 75.31 (-CH2), 84.97 (-CH), 127.14 (aromatic CH), 127.42 (aromatic CH), 127.51 (aromatic CH), 128.25 (aromatic CH), 128.49 (aromatic CH), 128.63 (aromatic CH), 131.90 (aromatic CH), 133.44 (aromatic quaternary C), 138.26 (aromatic quaternary C), 166.41, 167.22, 168.35; IR (cm' 1): 3418, 2955, 1743, 1672; HRMS (FAB): m/z calcd for ngHngOr, [M + H], 414.1919; found, 414.1916. 202 e02C M COzMe BZHN 0 60/0 65H .- Me Ill-17 Dimethyl-2-(benzamido)-2-(tetrahydro-3-methylfuran-2-yl)malonate (III-17): Using the general procedure, a solution of 2-phenyl-4-carbmethoxy—5(4H)- oxazolone (0.05 g, 0.23 mmol) and 2,3—dihydro-4-methyifuran (0.058 g, 0.68 f mmol) in 20 mL of anhydrous CH2CI2 was heated to reflux for 24 hours. The it; resulting ene adduct was stirred in 20 mL MeOH overnight yielding 0.06 g of malonate Ill-17 (81 % yield) as a white solid after silica gel Chromatography (40% ethyl acetate I 60% hexanes). 1H NMR (300 MHz), CDCI3: 6 1.19 (d, J = 7 Hz, 3H), 1.48-1.56 (m, 1H), 1.80-1.92 (m, 1H), 2.70-2.80 (m, 1H), 3.72-3.84 (m, 1H), 3.77 (s, 3H), 3.82-3.88 (m, 1H), 3.84 (s, 3H), 3.41 (d, J = 5 Hz, 1H), 7.37 (bs, 1H), 7.41-7.52 (m, 3H), 7.80-8.83 (m, 2H); 13C NMR + DEPT (75 MHz) CDCI3: 19.77 (~CH3), 34.65 (-CH2), 34.77 (-CH), 52.91 (-COZCH3), 53.83 (-C02CH3), 68.02 (-CH2), 68.94 (quaternary C), 88.13 (-CH), 127.18 (aromatic CH), 128.68 (aromatic CH), 132.06 (aromatic CH), 133.28 (aromatic quaternary C), 166.50, 168.10; IR (cm"): 3414, 2955, 1745, 1672; HRMS (FAB): m/z calcd for C17H22N03 [M + H], 336.1449; found, 336.1447. 203 0 O \ N cone Pl'l OMe Ill-19 MedlyI-4.5-dihydro-4-(2-methoxypropan-2-yIH-oxo-2-phenyloxazoIe-4- carboxylate (Ill-19): A solution of 2-phenyl-4—carbmethoxy-5(4H)—oxazolone (0.2 g, 0.91 mmol) and 2-methoxy propene (0.09 g, 1.19 mmol) in 20 mL of anhydrous CHZClz was stirred at room temperature for 30 min. The resulting ene adduct III-19 was Characterized by NMR. All attempts at ring opening the oxazolone with methanol were unsuccessful due to the reversibility of this reaction. According to the NMR spectra the initial ene reaction resulted in a yield of greater than 95%. 1H NMR (300 MHz), CDCI3: 6 1.52 (s, 3H), 1.55 (s, 3H), 3.12 (s, 3H), 3.76 (s, 3H), 7.40 (m, 3H), 7.99 (m, 2H); 13C NMR (75 MHz) CDCI3: 18.66, 20.39, 49.59, 53.22, 79.81, 82.57, 125.49, 128.30, 128.68, 133.00. 163.13, 165.01, 172.57; IR (cm'1): 2953, 1825, 1755, 1651, 1066. H OMe 82’ N Ph COzMe Ill-20 methyl 2-benzamido—O‘Bu N / cone Ill-46 Methyl-5-(1-tert-butoxyethoxy)-2-ethyloxazole-4-carboxylate (Ill-46): The title compound is an observed intermediate when 2-ethyl-4-carbmethoxy-5(4H)- oxazolone is treated with tart-butyl vinyl ether in anhydrous CHZClz. Since the compound is not produced in pure form during the reaction of 2-ethyl-4- carbmethoxy—5(4H)-oxazolone with ten-butyl vinyl ether, the following method was utilized in order to produce the compound in a more pure form for the purpose of characterization: A solution of 2-(methoxycarbonyl)-2- (propionamido)acetic acid (50 mg, 0.26 mmol), tort-butyl vinyl ether (39.1 mg, 0.39) and 10 mL of anhydrous CH2CI2 was treated with 1-ethyl-3-(3- dimethylaminopropyl) carbodiimide hydrochloride (55.6 mg, 0.29 mmol). The solution was stirred for 20 minutes and then washed once with 10 mL of water. The solution was then dried over sodium sulfate and concentrated in vacuo to yield 36 mg (51% yield) of the title compound as a white solid. The compound is fairly unstable so only partial characterization was able to be obtained. 1H NMR (500 MHz) (CDCI3): 6 1.19 (s, 9H), 1.30 (t, J = 7.5 Hz, 3H), 1.54 (d, J = 6 Hz, 3H), 2933.11 (m, 2H), 3.79 (s, 3H), 7.08 (q, J = 8 Hz, 1H); 13C NMR + DEPT (125 MHz) (CDCl3) 6: 9.92 (-CH3), 21.65 (-CH2), 23.45 (-CH3), 27.60 (-CH3), 50.99 (-CO2CH3), 77.70 (quaternary C), 80.82 (-CH), 154.31 (quaternary C), 160.62 (quaternary C), 162.43 (quaternary C). 217 o C02Me N cone H Ill-47 Dimethyl-Z-(isobutyramido)malonate (Ill-47): A solution of dimethylaminomalonate hydrochloride (3.0 g, 16.3 mmol), triethylamine (4.96 g, 49.0 mmol) and anhydrous CH2CI2 (40 ml) was treated dropwise with isobutyrl chloride (1.74 g, 16.3 mmol) in a flame dried round bottom flask. The solution was stirred at room temperature under nitrogen atmosphere for 5 hours. The solution was then washed exhaustively with 2M HCI solution and dried over magnesium sulfate. The solvent was removed under vacuum and the resulting crude solid was recrystallized using EtOAc/Hexanes to yield 2.85 g (81% yield) of the title compound as a white crystalline solid. 1H NMR (300 MHz) (CDCI3): 8 1.14 (d, J = 6.9 Hz, 6H), 2.46 (septet, J = 6.9 Hz, 1H), 3.78 (s, 3H), 5.16, (d, J = 7.2 Hz, 1H), 8.45 (d, J = 7.2 Hz, 1H); 13C NMR (75 MHz) (CDCI3): 19.20, 35.02, 53.35, 55.93, 188.85, 178.83; IR (neat): 3298 cm", 2972 cm", 1755 cm“, 1847 cm"; LRMS(EI): ancalcd for C9H15N05 217.2; found, 217.1. 0 COZH N 002MB H Ill-48 2-(methoxycarbonyI)-2-(isobutyramido)acetic acid (III-48): A solution of dimethyl-2-(isobutyramido)malonate (2.0 g, 9.2 mmol) and 50 ml of methanol was added to a round bottom flask and cooled to 0 °C. A solution consisting of 218 LiOH'HzO (0.39 g, 9.2 mmol) in 50 ml of H20 was then added to the flask dropwise over the course of approximately 30 minutes. The solution was stirred over night while being allowed to warm to room temperature. The following morning the methanol was removed under vacuum and the resulting aqueous solution was washed once with diethyl ether. The solution was then acidified using 2M HCI and washed three more times with diethyl ether. The combined ether layers were dried over magnesium sulfate and the solvent was removed under vacuum. The resulting crude solid was recrystallized in diethyl ether to yield the title compound as a white crystalline solid (0.92 g, 49% yield). 1H NMR (300 MHz) (CDCI3): 6 0.97 (d, J = 6.9 Hz, 6H), 2.58 (septet, J = 6.9 Hz, 1H), 3.68 (s, 3H), 4.99 (d, J = 7.5 Hz, 1H), 8.51 (d, J = 7.5 Hz, 1H), 13.44 (bs, 1H); 130 NMR (75 MHz) (DMSO): 19.32, 33.11, 52.57, 56.00, 167.70, 167.73, 176.51. HRMS (FAB): m/z calcd for C3H12N05 [M+H], 226.0691; found, 226.0675. 0 COzMe \'/IL )828‘3 N H 0 III-49 Dimethyl-Z-(isobutyramido)-2-(tetrahydrofuran-Z-yl)malonate (Ill-49): A solution of 2-(methoxycarbonyl)-2-(isobutyramido)acetic acid (0.1 g, 0.49 mmol), EDCI (0.1 g, 0.54 mmol), 2,3—dihydrofuran (0.05 g, 0.64 mmol) and anhydrous CH2CI2 was stirred at room temperature under nitrogen atmosphere for 18 hours. The solution was then washed with brine twice and dried over magnesium sulfate. The solvent was removed under vacuum and replaced with 20 ml of 219 MeOH. After 48 hours of stirring at room temperature the solvent was removed to yield 0.13 g of the title compound (92% yield) as a clear oil. 1H NMR (500 MHz), CDCI3: 8 1.12 (d, J = 7 Hz, 3H), 1.13 (d, J = 7 Hz, 3H), 1.70—1.80 (m, 1H), 1.78-1.86 (m, 1H), 1.94-2.21 (m, 1H), 2.11-2.20 (m, 1H), 2.44 (septet, J = 7 Hz, 1H), 3.66-3.80 (m, 2H), 3.70 (s, 3H), 3.78 (s, 3H), 4.59 (t, J = 7.5 Hz, 1H), 8.84 (bs, 1H); 13C NMR + DEPT (125 MHz) CDCI3: 19.47 (-CH3), 19.70 (-CH3), 28.28 (-CH2), 27.44 (-CH2), 35.55 (-CH), 53.06 (-Co2CH3), 53.90 (-C02CH3), 88.31 (quaternary C), 89.08 (-CH2), 81.58 (-CH), 166.73 (-COZCH3), 188.37 (- C02CH3), 176.64 (-CONH); IR (neat): 3378 cm", 2918 cm", 1741 cm", 1874 cm"; LRMS(EI): m/zcalcd for C13H21Noe 287.3; found, 288.1. 0 COzMe \/IL COZMe N H O Ill-50 Dimethyl-Z-(propionamido)-2-(tetrahydrofuran-Z-yl)malonate (Ill-50): A solution of 2-(methoxycarbonyl)-2-(propionamido)acetic acid (0.1 g, 0.53 mmol), EDCI (0.1 g, 0.58 mmol), 2,3-dihydrofuran (0.05 g, 0.69 mmol) and anhydrous CHZCIz was stirred at room temperature under nitrogen atmosphere for 24 hours. The solution was then washed with brine and dried over magnesium sulfate. The solvent was removed under vacuum and replaced with 20 ml of MeOH. After 48 hours of stirring at room temperature the solvent was removed to yield 0.11 g of the title compound (77% yield) as a clear oil. 1H NMR (500 MHz), CDCI3: 6 1.13 (t, J = 7.5 Hz, 3H), 1.71-1.80 (m, 1H), 1.79-1.87 (m, 1H), 1.97-2.04 (m, 1H), 2.13- 220 2.20 (m, 1H) 2.28 (qd, J1 = 7.5 Hz, J2 = 2.5 Hz, 2H), 3.65-3.73 (m, 1H), 3.71 (s, 3H), 3.73-3.79 (m, 1H), 3.78 (s, 3H), 4.59 (t, J = 7.5 Hz, 1H), 6.62 (bs, 1H); 13C NMR + DEPT (125 MHz) CDCI3: 9.57 (-CH3), 25.98 (-CH2), 27.23 (-CH2), 29.34 (-CH2), 52.85 (-C02CH3), 53.63 (-C02CH3), 68.13 (quaternary C), 88.78 (- CH2), 81.28 (-CH), 188.47 (-cozCH3), 188.00 (-C02CH3), 173.20 (~CONH); IR (Neat): 3376 cm", 2955 cm", 1745 cm", 1682 cm"; LRMS(EI): m/z calcd for C12H19N06 273.3; found, 273.7. 221 LC) 0 F or — P cm on _ 0% rr om - co 6.... 6:33:38 .0 «human «.22 K... 2.5.“. o2 cm on - om h .— P _ (- oww _ j j P In J on? on? our owN — . _l . 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WV 9m m.m 6.0 m6 OK Wu oé m6 0d Wm de m2: — p L r L b b b — + P P —L F L p p b p — b — P h L! FF P F - p n n m p p u p p h p P h h p p b h b 1 4: fl « ¢ fl _ 3 .> azuoo M. References (1) (2) (3) (a) Hughes, C. C.; Trauner, D.; The total synthesis of (-)-amathaspiramide F. Angew. Chem. Int. Ed. 2002, 41, 4556; (b) Kleinsasser, N. H.; Wallner, B. C.; Haneus, U. A.; Zwickenpflug, W.; Richter, E.; Genotoxic effects of myosmine in human lymphocytes and upper aerodigestive tract epithelial cells. Toxicology. 2003, 192, 171-177; (c) Ohkura, H.; Berbasov, D. 0.; Soloshonok, V. A.; Simple and highly diastereoselective synthesis of trifluoromethyl-containing myosmines via reaction between 2- (aminomethyl)pyridine and 1 ,1 ,1 ,5,5,5-hexafluoro-2,4-pentanedione. Tetrahedron Lett. 2003. 44, 2417-2420; (d) Tomko, J .; Brazdova, V.; Voticky, Z.; Veratrum Alkaloids .22. Veracintine - Novel Type of Veratrum Alkaloid with a Pyrroline Ring. Tetrahedron Lett. 1971, 3041; (e) Tsukamoto, D.; Shibano, M.; Kusano, 6.; Studies on the constituents of Broussonetia species X. Six new alkaloids from Broussonetia kazinoki SIEB. Chem. Pharm. Bull. 2001, 49, 1487-1491; (f) Tsukamoto, D.; Shibano, M.; Okamoto, R.; Kusano, 6.; Studies on the constituents of Broussonetia species Vlll. Four new pyrrolidine alkaloids, broussonetines R, S, T, and V and a new pyrroline alkaloid, broussonetine U, from Broussonetia kazinoki Sieb. Chem. Phann. Bull. 2001, 49, 492-496; (g) Usubillaga, A.; label, V.; Watson, W. H.; The Revised Structure of Solamaladine, 3-Beta-Hydroxy-22-(4-Methyl-1-Pyrrolin-2-YI)-23,24-Dinor- 5-Alpha-Cholane-4,22-Dione. Acta Crystallogr. Sect. B. 1982, 38, 966- 969. (a) Birouk, M.; Harraga, S.; Panouseperrin, J.; Robelt, J. F.; Damelincourt, M.; Theobald, F.; Mercier, R.; Panouse, J. J.; Aryl and Ethoxycarbonyl Derivatives of Pyrroles, 2H-Pyrroles and 3,4-Dihydropyrroles and Their lmmunoactivity of Human Lymphocytes-T. Eur. J. Med. Chem. 1991, 26, 91-99; (b) Fuska, J.; Fuskova, A.; Vassova, A.; Voticky, 2.; New Substances with Cyto-Toxic and Anti-Tumor Effects .4. lnvitro Effect of Some Veratrum Alkaloids and Their Derivatives on Leukemia P388 Cells. Neoplasma. 1981, 28, 709-714; (c) Moya, P.; Cantin, A.; Castillo, M. A.; Primo, J.; Miranda, M. A.; Primo-Yufera, E.; Isolation, structural assignment, and synthesis of N-(2-methyl-3-oxodecanoyI)-2-pyrroline, a new natural product from Penicillium brevicompactum with in vivo anti- juvenile hormone activity. J. Org. Chem. 1998, 63, 8530-8535. (a) Kuo, M. S.; Yurek, D. A.; Coats, J. H.; Chung, S. T.; Li, G. P.; Isolation and Identification of 3-Propylidene—Delta(1)-Pyrroline-5-Carboxylic Acid, a Biosynthetic Precursor of Lincomycin. J. Antibiot. 1992, 45, 1773-1777; (b) Luesch, H.; Hoffmann, D.; Hevel, J. M.; Becker, J. E.; Golakoti, T.; Moore, R. E.; Biosynthesis of 4-methylproline in cyanobacteria: Cloning of nosE and nosF genes and biochemical characterization of the encoded dehydrogenase and reductase activities. J. Org. Chem. 2003, 68, 83-91; (c) Miltyk, W.; Palka, J. A.; Potential role of pyrroline 5-carboxylate in 243 tr:- (4) (5) (6) regulation of collagen biosynthesis in cultured human skin fibroblasts. Comp. Biochem. Physiol. A. 2000, 125, 265-271; (d) Stapon, A.; Li, R. F.; Townsend, C. A.; Carbapenem biosynthesis: Confirmation of stereochemical assignments and the role of CarC in the ring stereoinversion process from L-proline. J. Am. Chem. Soc. 2003, 125, 8486-8493; (e) Stocking, E. M.; Martinez, R. A.; Silks, L. A.; Sanz- Cervera, J. F.; Williams, R. M.; Studies on the biosynthesis of paraherquamide: Concerning the mechanism of the oxidative cyclization of l-isoleucine to beta-methylproline. J. Am. Chem. Soc. 2001, 123, 3391- 3392. (a) Ballini, R.; Marcantoni, E.; Petrini, M.; A Nitrone-Based Approach to the Enantioselective Total Synthesis of (-)-Anisomycin. J. Org. Chem. 1992, 57, 1316-1318; (b) Dannhardt, 6.; Kiefer, W.; 1-pyrrolines (3,4- dihydro-ZH-pyrroles) as a template for new drugs. Arch. Pharm. 2001, 334, 183-188; (c) Fehn, 8.; Burger, K.; An efficient, stereoselective synthesis of (-)-bulgecinine from (S)-aspartic acid. Tetrahedron: Asymmetry. 1997, 8, 2001-2005; (d) Goti, A.; Cicchi, S.; Mannucci, V.; Cardona, F.; Guama, F.; Merino, P.; Tejero, T.; Iterative organometallic addition to chiral hydroxylated cyclic nitrones: Highly stereoselective syntheses of alpha, alpha'- and alpha, alpha-substituted hydroxypyrrolidines. Org. Lett. 2003, 5, 4235-4238; (e) Lombardo, M.; Fabbroni, S.; Trombini, C.; Entropy-controlled selectivity in the vinylation of a cyclic chiral nitrone. An efficient route to enantiopure polyhydroxylated pyrrolidines. J. Org. Chem. 2001, 66, 1264-1268; (0 Okue, M.; Kobayashi, H.; Shin-ya, K.; Furihata, K.; Hayakawa, Y.; Seto, H.; Watanabe, H.; Kitahara, T.; Synthesis of the proposed structure and revision of stereochemistry of kaitocephalin. Tetrahedron Lett. 2002, 43, 857-860; (g) Shvekhgeimer, M. G. A.; Methods of synthesis and chemical transformations of 3,4-2H-dihydropyrroles (Delta(1)-pyrrolines) (Review). Khimiya Geterotsiklicheskikh Soedinenii 2003, 483-529; (h) Sun, P.; Sun, C. X.; Weinreb, S. M.; A new total synthesis of the marine tunicate alkaloid lepadiformine. Org. Lett. 2001, 3, 3507-3510; (i) Watanabe, H.; Okue, M.; Kobayashi, H.; Kitahara, T.; The first synthesis of kaitocephalin based on the structure revision. Tetrahedron Lett. 2002, 43, 861-864. Fisk, J. S.; Mosey, R. A.; Tepe, J. J.; The diverse chemistry of oxazol-5- (4H)-ones. Chem. Soc. Rev. 2007, 36, 1432-1440. (a) Mukerjee, A. K.; Azlactones - Retrospect and Prospect. Heterocycles. 1987, 26, 1077-1097; (b) Padwa, A.; Pearson, W. H., Synthetic Applications of 1,3-Dipolar Cycloaddition Chemistry Toward Heterocycles and Natural Products, 1rst Ed. 2002, John Wiley and Sons, Hoboken, NJ, p 682-747. 244 (7) (8) (9) (10) (11) (12) (13) (a) Peddibhotla, S.; Jayakumar, S.; Tepe, J. J.; Highly diastereoselective multicomponent synthesis of unsymmetrical imidazolines. Org. Lett. 2002, 4, 3533-3535; (b) Peddibhotla, S.; Tepe, J. J.; Multicomponent synthesis of highly substituted imidazolines via a silicon mediated 1.3-dipolar cycloaddition. Synthesis. 2003, 1433-1440; (c) Sharma, V.; Tepe, J. J.; Diastereochemical dlllersity of imidazoline scaffolds via substrate controlled TMSCI mediated cycloaddition of azlactones. Org. Lett. 2005, 7, 5091-5094. (a) Gotthard.H; Huisgen, R.; 1,3-Dipolar Cycloadditions .57. Preparation of Delta-2-Pyrrolines from N-Substituted Oxazolium 5-Oxides and Olefinic Dipolarophiles. Chem. Bar. 1970, 103, 2625; (b) Huisgen, R.; Bayer, H. O.; Schaefer, F. C.; Gotthardt, H.; New Type of Mesoionic Aromatic Compound + Its 1,3-Dipolar Cycloaddition Reactions with Acetylene Derivatives. Angew. Chem. Int. Ed. 1964, 3, 136; (c) Huisgen, R.; Gotthard.H; Bayer, H. O.; 1,3-Dipolar Cycloadditions .55. Delta-1- Pyrrolines and 7-Azabicyclo-2,2,1-Heptane Derivatives from Azlactones and Activated Alkenes. Chem. Bar. 1970, 103, 2368; (d) Huisgen, R.; Gotthard.H; Bayer, H. O.; Schaefer, F. C.; 1,3-Dipolar Cycloadditions .56. A Convenient Synthesis of N-Substituted Pyrroles from Mesoionic Oxazolones and Alkynes. Chem. Bar. 1970. 103, 2611; (e) Huisgen, R.; Gotthardt, H.; Bayer, H. O.; 1.3-Dipolare Cycloadditionen Der Azlactone an Cc-Doppelbindungen. Tetrahedron Lett. 1964, 481-485. Gotthard.H; Huisgen, R.; Bayer, H. 0.; 1,3-Dipolar Cycloaddition Reactions .53. Question of 1,3-Dipolar Nature of Delta-2-Oxazolin-5-Ones. J. Am. Chem. Soc. 1970, 92,4340. Maryanoff, C. A.; Turchi, I. J.; Mechanism and Stereochemical Implications of the Reaction of an Oxazolium-5-Oxide with 1,2- Dicyanocyclobutene - an Am1 Study. Hetemcycles. 1993, 35, 649-657. Maryanoff, C. A.; Karash, C. B.; Turchi, I. J.; Corey, E. R.; Maryanoff, B. E.; Characterization of a Stable Carboxylic-Acid Inten'nediate from 1,3- Dipolar Cyclo-Addition of a Munchnone with 1,2-Dicyanocyclobutene. J. Org. Chem. 1989, 54, 3790-3792. Melhado, A. D.; Luparia, M.; Toste, F. D.; Au(I)-catalyzed enantioselective 1,3-dipolar cycloadditions of Munchnones with electron-deficient Alkenes. J. Am. Chem. Soc. 2007, 129, 12638. (a) Myers, J. A.; Wilkerson, W. W.; Council, S. L.; 1,3-Dipolar Addition of an Oxazolium 5-Oxide to Cyclopentadienequinone and to Anthracenequinone. J. Org. Chem. 1975, 40, 2875-2877; (b) Padwa, A.; Gingrich, H. L.; Lim, R.; Regiochemistry of Intramolecular Munchnone 245 (14) (15) (16) (17) (13) (19) (20) CycIo-Additions - Preparative and Mechanistic Implications. J. Org. Chem. 1982, 47, 2447-2456. Peddibhotla, S.; Tepe, J. J.; Stereoselective synthesis of highly substituted Delta(1)—pyrrolines: exo-selective 1,3—dipolar cycloaddition reactions with azlactones. J. Am. Chem. Soc. 2004, 126, 12776-12777. (a) Coldham, l.; Hufton, R.; Intramolecular dipolar cycloaddition reactions of azomethine ylides. Chem. Rev. 2005, 105, 2765-2809; (b) Pandey, G.; Banerjee, P.; Gadre, S. R.; Construction of enantiopure pyrrolidine ring system via asymmetric [3+2]—cycloaddition of azomethine ylides. Chem. Rev. 2006, 106, 4484-4517. (a) Arya, P.; Joseph, R.; Gan, Z. H.; Rakic, 8.; Exploring new chemical space by stereocontrolled diversity-oriented synthesis. Chem. Biol. 2005, 12, 163-180; (b) Burke, M. D.; Berger, E. M.; Schreiber, S. L.; Generating diverse skeletons of small molecules combinatorially. Science. 2003, 302, 613-618; (c) Burke, M. D.; Berger, E. M.; Schreiber, S. L.; A synthesis strategy yielding skeletally diverse small molecules combinatorially. J. Am. Chem. Soc. 2004, 126, 14095-14104; (d) Burke, M. D.; Schreiber, S. L.; A planning strategy for diversity-oriented synthesis. Angew. Chem. Int. Ed. 2004, 43, 46-58; (e) Schreiber, S. L.; Target-oriented and diversity- oriented organic synthesis in drug discovery. Science. 2000, 287, 1964- 1969; (f) Spring, D. R.; Diversity-oriented synthesis; a challenge for synthetic chemists. Org. Biomol. Chem. 2003, 1, 3867-3870. Alvarezlbarra, C.; Csaky, A. G.; deSiIanes, I. L.; Quiroga, M. L.; Diastereoselective synthesis of alpha alpha-disubstituted gamma- carboxypyroglutamates via Sm(Ill)-azomethine ylide cycloadditions. J. Org. Chem. 1997, 62, 479—484. Omura, S.; Fujimoto, T.; Otoguro, K.; Matsuzaki, K.; Moriguchi, R.; Tanaka, H.; Sasaki, Y.; Lactacystin, a Novel Microbial Metabolite, Induces Neuritogenesis of Neuroblastoma-Cells. J. Antibiot. 1991, 44, 113-116. Omura, S.; Matsuzaki, K.; Fujimoto, T.; Kosuge, K.; Furuya, T.; Fujita, S.; Nakagawa, A.; Structure of Lactacystin, a New Microbial Metabolite Which Induces Differentiation of Neuroblastoma-Cells. J. Antibiot. 1991, 44, 117- 1 18. (a) Dick, L. R.; Cruikshank, A. A.; Destree, A. T.; Grenier, L.; McCormack, T. A.; Melandri, F. D.; Nunes, S. L.; Palombella, V. J.; Parent, L. A.; Plamondon, L.; Stein, R. L.; Mechanistic studies on the inactivation of the proteasome by lactacystin in cultured cells. J. Biol. Chem. 1997, 272, 182- 188; (b) Dick, L. R.; Cruikshank, A. A.; Grenier, L.; Melandri, F. D.; Nunes, S. L.; Stein, R. L.; Mechanistic studies on the inactivation of the 246 (21) (22) (23) proteasome by lactacystin A central role for clasto-Iactacystin beta- lactone. J. Biol. Chem. 1996, 271, 7273-7276; (c) Fenteany, G.; Standaert, R. F.; Lane, W. S.; Choi, 8.; Corey, E. J.; Schreiber, S. L.; Inhibition of Proteasome Activities and Subunit-Specific Amino-Terminal Threonine Modification by Lactacystin. Science. 1995, 268, 726-731. (a) Balskus, E. P.; Jacobsen, E. N.; alpha,beta-unsaturated beta-silyl imide substrates for catalytic, enantioselective conjugate additions: A total synthesis of (+)-Iactacystin and the discovery of a new proteasome inhibitor. J. Am. Chem. Soc. 2006, 128, 6810-6812; (b) Brennan, C. J.; Pattenden, G.; Rescourio, 6.; Formal synthesis of (+)-lactacystin based on a novel radical cyclisation of an alpha-ethynyl substituted serine. Tetrahedron Lett. 2003, 44, 8757-8760; (0) Corey, E. J.; Li, W. D. 2.; Total synthesis and biological activity of lactacystin, omuralide and analogs. Chem. Pharm. Bull. 1999, 47, 1-10; (d) Donohoe, T. J.; Sintim, H.; Sisangia, L.; Harling, J. D.; An efficient synthesis of lactacystin beta- Iactone. Angew. Chem. Int. Ed. 2004, 43, 2293-2296; (e) Fukuda, N.; Sasaki, K.; Sastry, T. V. R. S.; Kanai, M.; Shibasaki, M.; Catalytic asymmetric total synthesis of (+)-Iactacystin. J. Org. Chem. 2006, 71, 1220-1225; (f) GilIey, C. B.; Buller, M. J.; Kobayashi, Y.; New entry to convertible isocyanides for the ugi reaction and its application to the stereocontrolled formal total synthesis of the proteasome inhibitor Omuralide. Org. Lett. 2007, 9, 3631-3634; (9) Green, M. P.; Prodger, J. C.; Hayes, C. J.; An enantioselective formal synthesis of the proteasome inhibitor (+)-Iactacystin. Tetrahedron Lett. 2002, 43, 6609-6611; (h) Hayes, C. J.; Sherlock, A. E.; Selby, M. D.; Enantioselective total syntheses of (-)- clasto-Iactacystin beta-lactone and 7-epi-(-)-cIasto-Iactacystin beta- lactone. Org. Biomol. Chem. 2006, 4, 193-195; (i) Masse, C. E.; Morgan, A. J.; Adams, J.; Panek, J. S.; Syntheses and biological evaluation of (+)- Iactacystin and analogs. Eur. J. Org. Chem. 2000, 2513-2528; (j) Ooi, H.; lshibashi, N.; Iwabuchi, Y.; lshihara, J.; Hatakeyama, S.; A concise route to (+)-lactacystin. J. Org. Chem. 2004, 69, 7765-7768; (k) Yoon, C. H.; Flanigan, D. L.; Yoo, K. S.; Jung, K. W.; Stereogenic evolution of clasto- lactacystin beta-lactone from L-serine. Eur. J. Org. Chem. 2007, 37-39. Gothelf, K. V.; Jorgensen, K. A.; Asymmetric 1.3-dipolar cycloaddition reactions. Chem. Rev. 1998, 98, 863-909. (a) Houk, K. N.; Frontier Molecular-Orbital Theory of Cycloaddition Reactions. Acc. Chem. Res. 1975, 8, 361-369; (b) Houk, K. N.; Sims, J.; Watts, C. R.; Luskus, L. J.; Origin of Reactivity, Regioselectivity, and Periselectivity in 1,3-Dipolar Cycloadditions. J. Am. Chem. Soc. 1973, 95, 7301-7315; (c) Sustrnann, R.; Simple Model for Substituent Effects in Cycloaddition Reactions .1. 1,3-Dipolar Cycloadditions. Tetrahedron Lett. 1971, 2717. 247 (24) (25) (25) (27) (28) (29) (30) (31) Pohlhaus, P. D.; Bowman, R. K.; Johnson, J. S.; Lewis acid-promoted carbon-carbon bond cleavage of aziridines: Divergent cycloaddition pathways of the derived ylides. J. Am. Chem. Soc. 2004, 126, 2294-2295. Savinov, S. N.; Austin, D. J.; The diastereoselective cycloaddition of vinyl ethers with isomunchnones. Chem. Comm. 1999, 1813-1814. Savinov, S. N.; Austin, D. J.; Modular evolution of a chiral auxiliary for the 1.3-dipolar cycloaddition of isomunchnones with vinyl ethers. Org. Lett. 2002, 4, 1415-1418. (a) Koen, M. J.; Morgan, J.; Pinhey, J. T.; Arylation of 4-Ethoxycarbonyl-2- Phenyloxazol-S-One by Aryllead Triaoetates - a Convenient Route to Alpha-Arylglycines. J. Chem. Soc., Perkin Trans. 1 1993, 2383-2384; (b) Morgan, J.; Pinhey, J. T.; Sheny, C. J.; Reaction of organolead triacetates with 4-ethoxycarbonyI-2-methyl-4,5-dihydro-1 ,3-oxazol-5-one. The synthesis of alpha-aryl- and alpha-vinyl-N-acetylglycines and their ethyl esters and their enzymic resolution. J. Chem. Soc., Perkin Trans. 1 1997, 613—619. Fisk, J. S.; Tepe, J. J.; lnterrnolecular Ene Reactions utilizing Oxazolones and Enol Ether. J. Am. Chem. Soc. 2007, 129, 3058-3059. (a) Avenoza, A.; Busto, J. H.; Cativiela, C.; Peregrina, J. M.; Reactivity of (Z)-4-arylidene-5(4H)-oxazolones: [4+2] cycloaddition versus [4+3] cycloaddition/nucleophilic trapping. Tetrahedron Lett. 2002, 43, 4167- 4170; (b) Ohfune, Y.; Shinada, T.; Enantio- and diastereoselective construction of alpha,alpha-disubstituted alpha-amino acids for the synthesis of biologically active compounds. Eur. J. Org. Chem. 2005, 5127-5143; (c) Vogt, H.; Brase, 8.; Recent approaches towards the asymmetric synthesis of alpha,alpha-disubstituted alpha-amino acids. Org. Biomol. Chem. 2007, 5, 406-430. (a) Garcia, J.; Mata, E. G.; Tice, C. M.; Hormann, R. E.; Nicolas, E.; Albericio, F.; Michelotti, E. L.; Evaluation of solution and solid-phase approaches to the synthesis of libraries of alpha,alpha-disubstituted-alpha- acylaminoketones. J. Comb. Chem. 2005, 7, 843-863; (b) Ruble, J. C.; Fu, G. C.; Enantioselective construction of quaternary stereocenters: Rearrangements of O-acylated azlactones catalyzed by a planar—chiral derivative of 4-(pyrrolidino)pyridine. J. Am. Chem. Soc. 1998, 120, 11532- 11533; (c) Tice, C. M.; Hormann, R. E.; Thompson, C. S.; Friz, J. L.; Cavanaugh, C. K.; Michelotti, E. L.; Garcia, J.; Nicolas, E.; Albericio, F.; Synthesis and SAR of alpha-acylaminoketone ligands for control of gene expression. Bioorg. Med. Chem. Lett. 2003, 13, 475-478. Hoffrnann, H. M.; Ene Reaction. Angew. Chem. Int. Ed. 1969, 8, 556. 248 (32) (33) (34) (35) (36) (37) Conia, J. M.; Leperchec, P.; Thermal Cyclization of Unsaturated Carbonyl- Compounds. Synthesis. 1975, 1-19. Paderes, G. D.; Jorgensen, W. L.; Computer-Assisted Mechanistic Evaluation of Organic-Reactions .20. Ene and Retro-Ene Chemistry. J. Org. Chem. 1992, 57, 1904-1916. (a) Balme, G.; Bouyssi, D.; Faure, R.; Gore, J.; Vanhemelryck, 8.; Formation of Cyclopentanes from Delta-Ethylene Malonates by Catalysis in Palladium(0) - Stereochemistry and Mechanism. Tetrahedron. 1992, 48, 3891-3902; (b) Bouyssi, D.; Monteiro, N.; Balme, G.; Intramolecular carbocupration reaction of unactivated alkynes bearing a stabilized nucleophile: Application to the synthesis of iridoid monoterpenes. Tetrahedron Lett. 1999, 40, 1297-1300; (c) Kitagawa, 0.; Suzuki, T.; lnoue, T.; Watanabe, Y.; Taguchi, T.; Carbocyclization reaction of active methine compounds with unactivated alkenyl or alkynyl groups mediated by TiCI4-Et3N. J. Org. Chem. 1998, 63, 9470-9475; (d) McDonald, F. E.; Olson, T. C.; Group VI metal-promoted endo-carbocyclizations via alkyne- derived metal vinylidene carbenes. Tetrahedron Lett. 1997, 38, 7691- 7692. Boaventura, M. A.; Drouin, J.; Conia, J. M.; Doubly Catalyzed Cyclization of Epsilon-Acetylenic Carbonyl-Compounds. Synthesis. 1983, 801-804. (a) Cruciani, P.; Aubert, C.; Malacria, M.; Studies on Diastereoselectivity of the CobaIt(l) Catalyzed Cycloisomerization of Substituted Epsilon- Acetylenic Beta-Ketoester. Tetrahedron Lett. 1994, 35, 6677-6680; (b) Cruciani, P.; Stammler, R.; Aubert, C.; Malacria, M.; New cobalt-catalyzed cycloisomerization of epsilon-acetylenic beta-keto esters. Application to a powerful cyclization reactions cascade. J. Org. Chem. 1996, 61, 2699- 2708; (c) Renaud, J. L.; Aubert, C.; Malacria, M.; Cobalt-mediated cycloisomerization of delta-substituted epsilon-acetylenic beta-ketoesters construction of angular triquinane by a sequence ene/Pauson-Khand reactions. Tetrahedron. 1999, 55, 5113-5128; (d) Renaud, J. L.; Petit, M.; Aubert, C.; Malacria, M.; Synthetic usefulness of the cobalt(l)-mediated ene type reaction for the diastereoselective construction of bicyclo[n.3.0]derivatives. Synlett. 1997, 931. (a) Kennedy-Smith, J. J.; Staben, S. T.; Toste, F. D.; Gold(I)-catalyzed conia-ene reaction of beta-ketoesters with alkynes. J. Am. Chem. Soc. 2004, 126, 4526-4527; (b) Staben, S. T.; Kennedy-Smith, J. J.; Toste, F. 0.; Gold (I)-catalyzed 5-endo—dig carbocyclization of acetylenic dicarbonyl compounds. Angew. Chem. Int. Ed. 2004, 43, 5350-5352. 249 (38) (39) (40) (41) (42) (43) (44) (45) (46) (47) Takahashi, K.; Midori, M.; Kawano, K.; lshihara, J.; Hatakeyama, 8.; Entry to heterocycles based on indium-catalyzed Conia—ene reactions: Asymmetric synthesis of (-)-salinosporamide A. Angew. Chem. Int. Ed. 2008, 47, 6244-6246. Corkey, B. K.; Toste, F. 0.; Catalytic enantioselective Conia-ene reaction. J. Am. Chem. Soc. 2005, 127, 17168-17169. (3) Endo, K.; Hatakeyama, T.; Nakamura, M.; Nakamura, E.; Indium- catalyzed 2-alkenylation of 1,3-dicarbonyl compounds with unactivated alkynes. J. Am. Chem. Soc. 2007, 129, 5264-5271; (b) Nguyen, R. V.; Yao, X. 0.; Bohle, D. S.; Li, C. J.; Gold- and silver-catalyzed highly regioselective addition of active methylenes to dienes, triene, and cyclic enol ethers. Org. Lett. 2005, 7, 673-675. (a) Kolka, A. J.; Napolitano, J. P.; Ecke, G. G.; The Ortho-Alkylation of Phenols. J. Org. Chem. 1956, 21, 712-713; (b) Kolka, A. J.; Napolitano, J. P.; Filbey, A. H.; Ecke, G. G.; The Ortho-Alkylation of Phenols. J. Org. Chem. 1957, 22, 642-646; (b) Yadav, G. D.; Kumar, R; Alkylation of phenol with cyclohexene over solid acids: Insight in selectivity of 0- versus C-alkylation. Appl. Catal. 2005, 286, 61-70. Ma, Q. S.; Chakraborty, D.; Faglioni, F.; Muller, R. P.; Goddard, W. A.; Harris, T.; Campbell, C.; Tang, Y. C.; Alkylation of phenol: A mechanistic view. J. Phys. Chem. A. 2006, 1 10, 2246-2252. Weissemel, K. A., H.; Industrial Organic Chemistry. 3'‘1 Ed., VCH Weinheim: 1997, p. 358. Pinhey, J. T.; Xuan, P. T.; The Thermal Ortho-Substitution of Phenols by Vinyl Ethers. Aust. J. Chem. 1988, 41, 69-80. Clarke, H. T.; Johnson, J. R.; Robinson, R., The Chemistry of Penicillin, in Oxazoles and Oxazolones, Princeton University Press, Princetone, NJ: 1949; p 688-848. (a) Bergman, J.; Lidgren, G.; Reaction of Tryptophan with Trifluoroacetic— Anhydride. Tetrahedron Lett. 1989, 30, 4597-4600; (b) Chen, F. M. F.; Benoiton, N. L.; 4-AlkyI-5(4H)-Oxazolones from Mixed Anhydrides of N- Forrnylamino Acids. Int. J. Pept. Protein Res. 1991, 38, 285-286; (b) Chen, F. M. F.; Kuroda, K.; Benoiton, N. L.; Simple Preparation of 5-Oxo- 4,5-Dihydro-1,3-Oxazoles (Oxazolones). Synthesis. 1979, 230-232. (a) Berkessel, A.; Cleemann, F.; Mukherjee, S.; Muller, T. N.; Lex, J.; Highly efficient dynamic kinetic resolution of azlactones by urea-based bifunctional organocatalysts. Angew. Chem. Int. Ed. 2005, 44, 807-811; 250 (48) (b) Berkessel, A.; Mukherjee, S.; Cleemann, F.; Muller, T. N.; Lex, J.; Second-generation organocatalysts for the highly enantioselective dynamic kinetic resolution of azlactones. Chem. Comm. 2005, 1898-1900. Collins, D. J.; James, A. M.; Preparation of 2-(3-Bromo-1-Methylpropyl)- 1,3-Dioxolan and the Corresponding Chloride from 2-Methylbutyrolactone. Aust. J. Chem. 1989, 42, 223-228. 251 CHAPTER W SYNTHESIS OF TERT-ALKYL AMINO HYDROXY CARBOXYLIC ESTERS VIA AN INTERMOLECULAR ALKYLATION REACTION OF OXAZOL-5(4H)-ONES USING ENOL ETHERS A. Introduction to q,q-disubstituted a-amino acids Non-proteinogenic amino acids have proven to be valuable substrates for a wide range of applications within the fields of synthetic organic, bioorganic and medicinal chemistry.1 In particular, a,a-disubstituted a-amino acids (or quaternary a-amino acids) have received considerable attention from the scientific community as of late (Figure lV-1).2‘5 The additional alkyl substituent at the a-carbon of the amino acid changes its physical properties in multiple ways. For example, the additional substituent often helps to sterically constrain the free rotation of the residue’s side chain helping to cause unique folding when incorporated into peptides."4 Peptides containing quaternary a-amino acids also tend to have increased hydrophobicity, as well as an increased stability towards both chemical6 and metabolic7 decomposition. These unique physical properties make them intriguing tools for the design and study of peptides and proteins. In addition, a,cr-disubstituted a-amino acids derivatives can be found in nature either in their free form, or within the structures of many biologically interesting heterocyclic natural molecules)?"8 252 HzNYCOZH HZN COzH R R) R2 o-amino acid a,a-disubstituted a-amino acid Figure lV-1 . General structures of both a-amino acids and a,a-disubstituted a- amino acids (quaternary a-amino acids). B. Synthesis of a,a-disubstitut:ed a-amino acids using oxazol-5(4H)-ones The importance of a-amino acids containing quaternary carbons has caused a high interest in the development of new and efficient methods for their synthesis.2'5'° Classical methods for their synthesis include the alkylation of a- amino esters protected as imines9 and the Strecker reaction10 of ketimines. Oxazol—5(4H)-ones have been proven to be excellent substrates for synthesizing quaternary substituted a-amino acids derivatives.”12 Quaternary substituted amino acid derivatives can be directly accessed from the nucleophilic ring opening of quaternary substituted oxazol-5(4H)-ones (Scheme IV-1). Quaternary substituted oxazol-5(4H)-ones can be produced through a variety of methods.12 The relatively high acidity of the oxazoI-5(4H)-one a-protons allows for diverse transformations not traditionally seen when attempting to derivatize a-amino acids themselves.”14 In addition, their cyclic structure is less sterically encumbering as compared to their acylic a-amino acid counterparts further helping to assist in derivatization adjacent to the carbonyl. 253 I. H R1 0 R1YO R1\n/N\ leo O R} R3 R3 R3 R2 Quaternary Quaternary Amlno Acid Oxazolone °m01'514H)-0|16 Scheme lV-1 . Synthesis of quaternary amino acids from quaternary oxazolones. One of the earliest methods for generating alkylated oxazol-5(4H)-ones to construct a,a-disubstituted a-amino acids was reported by Steglich and co- workers in 1979.13 OxazoI-5(4H)—ones were suspended in polar aprotic solutions and treated with Hiinig’s base in the presence of highly reactive electrophiles (Scheme lV-2). The resulting quaternary oxazolones were in turn hydrolyzed to afford the desired a,a-disubstituted a-amino acids. When utilizing this chemistry, less reactive electrophiles tend to be problematic and lead to unwanted side products primarily due to competitive O-alkylation of the enolate intermediates. Recent developments optimizing the reaction conditions have allowed for the use of wider range of electrophiles,15 although O-alkylation still remains a problem with many substrates. R1 0 R1 0 R1 0 V0 Ra-X 822:0 + VORE’ N ——" N N Base R R2 R2 3 R2 Quaternary Aromatic Oxazolone Oxazole Scheme IV-2. Alkylation of oxazoI-5(4H)-ones with alkyl halides. Recent progress in oxazol-5(4H)-one chemistry has led to the development of novel methods utilizing transition metal catalysts as well as 254 organocatalysts to overcome the regioselectivity issues associated with the alkylation of oxazol-5(4H)-ones. For example, Trost and co-workers have reported multiple transformations utilizing transition metal catalysis for the allylic alkylation of oxazol-5(4H)-ones (See Chapter 1).16 Transition metals have also been utilized in the arylation of oxazol-5(4H)-ones to synthesis quaternary aryl glycine derivatives.17 In 2003, Hartwig and co-workers reported the first palladium catalyzed arylation of oxazol—5(4H)-ones for the synthesis of quaternary amino acids (Scheme lV-3).18 The reaction involves the coupling of the sp2 carbon of the aromatic enolate of oxazol-5(4H)-ones with aryl and vinyl bromides. The catalyst system consists of using Pd(OAc)2 along with the sterically hindered electron rich ligand Ad2P(t-Bu) (Scheme lV—3). ArBr Pd(OAc)2 O [EJI K2003 "IN R Toluene R 58-85% yield Scheme IV-3. Hartwig’s palladium catalyzed arylation of oxazol-5(4H)-ones. C. Significance of tart-alkyl amino hydroxy carboxylic acids Included within the class of quaternary substituted non-proteinogenic a-amino acids are amino hydroxy carboxylic acids whose structural features include a carbon core surrounded by alkyl, amino, carboxylic, and hydroxyl functional groups in various combinations?"8 Highly substituted amino hydroxy carboxylic acids are structural features present in numerous microorganism metabolites including the sphingofungins, lactacystins, salinosporamides, altemicidines, 255 Q‘s __J I oxazolomycins and many others (Figure lV-2).3"8 The diverse and potent biological activity of these molecules has stimulated many researchers to pursue ”2°21” and in-depth biological evaluations.23'2‘ A general their total syntheses method for rapidly and efficiently synthesizing the quaternary amino acid core found within this class of molecules would greatly enhance the rate at which their biological profile could be studied. 9H 9H cozn C6H13WNH2 ”_1 HO OLD/EM.“ COzH '3 H COZHH (-) Sphingofungln E R = OH (+) Sphingofungin F R a H Kaltocephalln O O JVSOzNHz 1 I\COZ|" . Me” OH HO s’fi/COzl-I ,i. .<‘0H NHAc CONHz (-) Altemlcldin (+) Lactacystin Salinosporamide A —OR1 PPY’ Arm/o Nfo N\:o s‘ 0 R2 R10 Ar 0 a 88-92% ee Wig—O 93-95% yield N / 0 R2 JLG) LR1O PPY" Scheme lV-4. Fu’s enantioselecitve Steglich rearrangement. A more recent method for alkylating of oxazol-5(4H)-ones to form tert- alkyl amino hydroxy carboxylic acid precursors was reported by Trost and coworkers involving a palladium-catalyzed addition of oxazol-5(4H)-ones to electron rich allenes (Scheme IV-5).28 The reaction overcomes regioselectivity problems generally associated with allene chemistry by substituting one end of the allene with an electron rich alkoxy group. The reaction works very well for with oxazoI-5(4H)-ones containing aliphatic substitutions at the C-4 position affording the highest yields (67—87%) along with excellent enantiomeric excesses (QC-94%) (Scheme lV-5). The diastereoselectivity was also reported to be high in these reactions usually occurring in about a 20:1 ratio. The stereoselectivity of the reaction is induced using their chiral cyclohexyldiamine derived ligand 1 . 258 0 ll Pd(OCCF3)2 (2 mol%) 0 "rag? 1 (6 mol%) > w =-=/OB KO‘Bu (2 mol%) Re‘ 0 4% __(Ph Hippuric Add (20 "‘0' %) CH2CI2, Lt. NH l-IN PththP Scheme IV-5. Allylic alkylation of oxazoI-5(4H)-ones using alkoxy allenes. 67-87% yield 85-94% ee Up to 20: 1 dr Furthermore, oxazol-5(4H)-ones have previously been used to synthesize molecules containing tart-alkyl amino hydroxy carboxylic acid cores. In 1998 Trost and Lee reported an asymmetric total synthesis of the naturally occurring tart-alkyl amino hydroxy carboxylic acid Sphingofungin F (Scheme lV-6).29 Sphingofungin F was first isolated by Merck in 1992 as an antifungal agent from the fermentation of Paecilomyces variotii.3° The compound has been found to be a potent inhibitor of the biosynthesis of sphingolipids due to its inhibitory activity against serine palmitoyltransferase.31 Trost and Lee utilized an asymmetric allylic alkylation of 2-phenyl-4-methyl-5(4H)-oxazolone Ill-1 with a gem-diacetate as their key step to synthesizing the natural product (Scheme IV-6). The reaction generated the tart-alkyl amino hydroxy carboxylic core in 70% yield with 89% as. The synthesis required only 15 linear steps and proceeded with an impressive 17% overall yield. 259 o 1 H $0 (.110ng anCI) GAG O N=J\ 5 2.7 TBDPSO Ill-1 Ph THF, {5°C 9 N‘:_< OAC 70% we” Ph 4. TBDPSONOAC 89% 86 J NH HN OH OH ”C6H13W\/\/\ HN ene-type :' x.- COzMe reaction M902C 0R2 Me02C\COH R1 Scheme IV-7. Proposed synthesis of tart-alkyl amino hydroxy carboxylic acids. F. Improving the diastereoselectivity using Lewis acids As reported in the previous chapter, the reaction of oxazol-5(4H)-ones with enol ethers results in the formation of quaternary substituted oxazolone intermediates as mixtures of diastereomers?” Our initial studies indicated that the ratio of the two product diastereomers is partially dependent upon the nature of the substrates and solvent being used, although it should be noted the diastereomeric ratio is generally approximately 50:50. In order to make this alkylation chemistry of oxazol-5(4H)-ones using enol ethers more appealing for use in synthetic organic chemistry, we sought to develop a method for improving the diastereoselectivity of the reaction. During our previous mechanistic studies, we determined these reactions likely involve the initial formation of an oxonium ion intermediate as a direct result of the protonation of the enol ether by the acidic oxazoI-5(4H)-one.3'2 The intermediate oxonium ion is then trapped by the oxazol-5(4H)-one substrate to form the new quaternary center (Chapter III). Based off these observations, we envisioned using a catalyst to either increase the acidity of the oxazol-5(4H)-one 261 or to directly initiate the formation of the oxonium ion species (Scheme lV-8). Either method would hopefully help to improve the overall stereoselectivity of the reaction by means of the complexation of the catalyst to the reacting species. Lewis acids have previously been used to complex with oxazol-5(4H)- ones to promote stereoselective transformations adjacent to their carbonyl. For example, previous reports have illustrated the use of Lewis acids to catalyze the dynamic kinetic resolutions of oxazoI-5(4H)-ones to synthesize enantiomerically pure a-amino acids.35 Furthermore, Lewis acids have been used to catalyze 35,37 In similar reactions such as the ortho-alkylation of phenols (Chapter III). addition, Lewis acids have been used to catalyze other carbon to carbon bond forming reactions utilizing enol ethers. For example, both cationic gold as well as trifluoroacetic acid has been used to catalyze the addition of enol ethers to 1,3- dicarbonyl containing substrates. 3° .—R1 0 - R1 0 .03 _H:>_<_, 10 on @ __. I o 2 F8123 R2 * 0R3 - X . R1 0 LA 0R3 ”R1 LA - R O 1 0 F/ Ox ——-=/ Y/ 0 ———> Y 0 LA: H N 6') a R2 LA R2 FOR3 R2 *OR3 Scheme lV-8. Proposed Lewis acid catalysis for improving the stereoselectivity of the alkylation reactions between oxazol-5(4H)-ones and enol ethers. 262 G. Optimization of reaction conditions for improving diastereoselectivity During our previous studies, we observed these ene-type alkylation reactions of oxazol-5(4H)-ones and enol ethers to proceed with little or no diastereoselectivity.32 In an effort to make this synthetic approach more broadly applicable, we initiated a study screening various reaction conditions with hopes to improve the overall diastereoselectivity of the reaction. We began our study by first attempting to optimize the solvent being used in these reactions. The reaction of 4-carbmethoxy-2-phenyl—5(4H)-oxazolone Ill- 5 and tart-butyl vinyl ether Ill-6 was performed in various solvents. The resulting quaternary oxazolone was reduced to the corresponding amino alcohol using sodium borohydride. The diastereomeric ratio of the product mixture was determined using integration values obtained from the 1H NMR spectra. Most solvents screened resulted little or no diastereoselectivity in the reaction of 4- carbmethoxy-2-phenyI-5(4H)-oxazolone Ill-5 and tart-butyl vinyl ether III-6. The use of more polar solvents tended to result in faster product formation with CH2CI2 producing the most rapid result at 1 hour (T able lV-1, entry 2). In addition, solvents containing lone pairs of electrons tended to have longer reaction times as compared to solvents of similar polarity lacking lone electrons. The use of highly polar solvents such as DMSO yielded very little product presumably due to enol ether decomposition (T able lV-1, entry 5). The use of less polar solvents such as benzene did provide improved stereoselectivity with a diastereomeric ratio of approximately 1.7 : 1, but also decreased the rate of reaction considerably (T able N—1, condition 4). 263 O‘Bu I o Ill-8 : 32’" . BZ,N)/‘\ co M Solvent. rt Meozc“ OH MeOZC "'—0H 2 e 2) NaBH4 lV-1A lV-1B III-5 A B Conditions Solvent Time (h) me as Yleld 1 CH3CN 3 50:50 90 2 CH2CI2 1 55:45 91 3 THF 19 52 :43 90 4 Benzene 36 67: 33 90 5 DMSO 24 50:50 Low Table lV-1. Reaction of 4-carbmethoxy-2-phenyl-5(4H)-oxazolone Ill-5 with tert- butyl vinyl ether Ill-6 followed by reduction with sodium borohydride. Upon examining the reaction solvent, next we turned our attention to exploring Lewis acid catalysis with the hope of increasing the reaction rate and diastereoselectivity. We initiated this study by screening a variety of Bronsted acids in the reaction of 4-carbmethoxy-Z-phenyI-5(4H)-oxazolone III-5 with tert- butyl vinyl ether Ill-6. To our delight, we found a substoichiometric amount (10 mol%) of diphenyl phosphate lV-3 to improve both the reaction rate and diastereoselectivity (Table lV-2, catalyst lV-3). Bronsted acids less acidic than diphenyl phosphate lV-3 also improved the diastereoselectivity of the reaction, but did little for increasing the reaction rate (Table IV-2, catalyst IV-2). The use of more acidic Brensted acids resulted in lower yields of desired product, presumably due to either enol ether or product decomposition (Table lV-2, catalysts lV-4 and lV-5). 264 1) ‘ u O‘Bu O‘B — u Y0 7'“ (1.5 eq.) H ; H ; N + N N \2: r 32’ . Bz/ >/\ Catalyst FC ,9— COzMe Benzene, it M9020 0“ M9020 OH Ph III-5 2) NaBH4 lV-1A IV-1B A 8 Catalyst Catalyst Time (hr) MB 96 Yield rv-2 3,5-Dinitrobenzoic acid 16 7o : 30 90 M3 Diphenyl phosphate 3 75 : 25 90 N4 CSA 1 74 : 26 83 M5 TFA 48 67 : 33 81 M8 Tl(O‘Pr)4 48 67: 33 61 M7 Yb(OTf)3 48 - : - o lV-8 TMSOTf 48 - : - o N-9 mom; 48 52 : 48 24 M10 Cu(OTf)2 48 — 1 - o Table lV-2. Screening of various Lewis acids in the reaction of 4-Cflbmethoxy-2- phenyl-5(4H)-oxazolone Ill-5 with tert-buyl vinyl ether III-6. Several additional oxaphilic and azaphilic Lewis acids were screened resulting in little or no product formation and low diastereoselectivity (Table lV-2, catalysts lV-6 through lV-10). The significant rate enhancement found when using Bronsted acids and not other Lewis acids suggests that the catalyst protonates the enol ether forming an oxonium ion, although coordination to the oxazol-5(4H)-one substrate cannot be dismissed. All of the conditions used in Table lV-1 and Table N-2 produced the same major diastereomer whose relative 265 stereochemistry was determined via crystal structure, which is depicted in Figure IV-3 below. 0" é, um , C/S‘ " C(15) v " «g, crrsr ,~. GM :3) CI6I / 15(4) Figure lV-3. Crystal structure of compound lV-1 A. H. Other phosphoric acids Upon determining diphenyl phosphate IV-3 to have the optimal acidity for both increasing the reaction rate as well as the reaction diastereoselectivity, we decided to screen a variety of other phosphoric acid derivatives. We synthesized a small library of phosphoric acid derivatives that both sterically and electronically differed from diphenyl phosphate lV-3. To compare the phosphoric acid derivatives lV-11 through IV-15, 4-Cflbmethoxy-2-phenyI-5(4H)-oxazolone Ill-5 was treated with tart-butyl vinyl ether Ill-6 in the presence of 10 mol% of each catalyst. Unfortunately, the majority of the phosphoric acid catalysts screened 266 did little for improving the reaction diastereoselectivity any more than using diphenyl phosphate lV-3. The use of bis(4-methoxyphenyl)phosphate lV-11 resulted in very similar yields and diastereoselectivity (Table lV-3, phosphoric acid lV-11) when compared to diphenyl phosphate lV-3. Substituting alkyl groups on the aryl rings either at the R1 or R2 resulted in a slight decrease in the diastereoselectivity of the reaction (T able IV-3, phosphoric acids lV-12 and IV- 13). _/O‘Bu Ph -- 1.5 sq. Ph Ph 0 O O Y. o ... ,_ Y.;. + i}... N Phosphoric Acid N ‘ ~,\O‘Bu N ‘ O‘Bu COzMe Benztene M902C7\| M802C7\r III-5 III-78 III-7b (R R3 szggm" R1 R2 R, (A: 8) Yield (96) 2 R2 R1 R1 rv-3 H H H 75: 25 99 0‘ ,9 \ ’ O lV-11 H H OMe 75: 25 99 '10 R 1 N42 Me H H 73: 27 99 R2 R2 R3 N-13 H tBu H 74:26 99 Phosphoric Acids IV-3 to ~43 Table IV-3. Screening of other diaryl phosphoric acid derivatives in the reaction of 4-carbmethoxy-2-phenyl-5(4H)-oxazolone Ill-5 and tart-butyl vinyl ether III-6. We also examined two additional phosphoric acid derivatives whose conformation differed in the fact that the two aryl groups of the acid were covalently linked together (Scheme lV-9, catalysts lV-14 and IV-15). Achiral 2,2’- 267 biphenylphosphoric acid IV-14 provided a similar diastereomeric ratio and yield as seen with diphenyl phosphate IV-3. Chiral BINOL derived phosphoric acid IV- 15 provided the highest diastereoselectivity of any of the catalysts investigated with a diastereomeric ratio of 79 to 21, but provided no observable enantioselectivity as determined by chiral HPLC (Scheme lV-9). Future work in this area may consist of the synthesis and evaluation of other BINOL derivatives for not only improving the diastereoselectivity of the reaction, but also hopefully inducing enantioselectivity during the transformation. _JO‘Bu Y o "I" - Y ~§=Q + Y §\=0 N Catalyst N ' .,.O‘Bu N ‘ O‘Bu cone Befiene Me02C Me02C Ill-5 III-7a Ill-7b Catalyst Time (h) AzB e.e. 91. Yield M14 22 76: 24 - 98 M15 18 79: 21 2 99 F or a \,I \I, O CO ~44 . IV-15 Scheme lV-9. Reaction of 4-carbmethoxy-2-phenyl-5(4H)-oxazolone Ill-5 and tart-butyl vinyl ether Ill-6 in the presence of catalysts IV-14 and lV-15. 268 l. Various enol ethers Utilizing the reaction conditions we optimized for improving the diastereoselectivity of these reactions, the scope of the reaction was further explored. Exposing 4-carbmethoxy-Z-phenyl-5(4H)-oxazolone III-5 to various vinyl enol ethers in the presence of 10% of diphenyl phosphate resulted in high yields of the desired products after sodium borohydride reduction (Table lV-4). The tart-butyl and benzyl protecting groups yielded better diastereoselectivity than the ethyl protecting group (Table lV-4, entries 1-3), indicating that enol ethers containing more sterically demanding oxygen protecting groups generally produce better diastereoselectivities. The less reactive higher substituted enol ethers resulted in lower yields of products and required the use of heat (T able IV- 4, entries 4-6) to produce the desired products in good yields. 'Both 1- butoxyethyne Ill-29 and 2-methoxypropene Ill-18 (T able lV-4, entries 7 and 8, respectively) also provided reasonable yields of products. 269 ph Y0 1) Enol Ether H Diphenyl- N R + | o 82 . 82 N ”52:35 MeOzCKOH M902 ox; COZM" 2) NaBH4 Ill-5 THF/H20 lV-‘Ia to N-26a IV-1b to N-26b A B Entry Substrate R Temp PC) A:B %Yield O‘Bu 1 O‘Bu 3 25 75: 25 90 Ill-8 rv-1 OBn OBn ? 2 :J EA 25 75 25 77 M16 M20 DB 3 OEt ; 25 67 33 85 M17 lV-21 9 4 < :0 - 50 38 62 57 — “it Ill-10 rv-22 OBn 93" 5 F__—/ = 50 69: 31 48 tea/V (5:1 trans to Cis) lV-18 M23 6 OMe 9M6 50 60 - 4o 33 >——/ .7117 . N-19 W- n 7 O"Bu O B" 25 - r — 67 '1." Ill-29 lV-25 8 OMe OMe 25 - - 62 “'11 Ill-18 IV-26 Table lV-4. Reaction of various enol ethers with 4-carbmethoxy-2-phenyI-5(4H)- oxazolone Ill-5 under the optimized reaction conditions. 270 J. Various oxazol-5(4H)-ones The role of the 2-position of the oxazol-5(4H)-one scaffold was investigated for its effect on reactivity and selectivity (T able IV-5). Several oxazol-5(4H)-ones with varying substitutions at the 2-position were prepared and were reacted with tert- butyl vinyl ether Ill-6 in the presence of diphenyl phosphate IV-3. Oxazol-5(4H)- ones with aryl substituents afforded high yields of product formation and erosion of diastereoselectivity was observed as electron deficiency increased (Table IV- 5, entries 1-3). Stereoselectivity was all but lost when oxazol-5(4H)-ones containing alkyl substituents were used (Table lV-5, entries 4 and 5). R 1) _JOtBu R o R o O _ if 0 "M HN 9‘8” H1? 913” Phosphate , :0 '-.,__ COzMe B an e, rt MeOzc OH M602C OH (n.5, m.“ 2) NaBH4 IV-1a to M33a lV-‘lb to lV-33b N-27 to N-29 A 8 Entry R A:B 96 Yield 1 Ph 75 : 25 90 Ill-5 2 4-MeO-Ph 74 : 26 88 lV-27 3 4-CF3-Ph 67 : 33 81 lV-28 4 Et 43 : 57 50 Ill-44 5 Bn 52 : 48 58 lV-29 Table lV-5. The reaction of various oxazol-5(4H)-ones with tart-butyl vinyl ether Ill-6 under the optimized reaction conditions. 271 bran-‘1 ‘i .. The decrease in both stereoselectivity and yield in the reactions involving 2-alkyI-oxazolones as compared to those involving 2-aryl-oxazolones may potentially be explained by a difference in mechanism. To investigate this phenomenon, we treated both 4—carbmethoxy-2-ethyl-5(4H)-oxazolone III-44 and 4-carbmethoxy-Z-phenyI-5(4H)—oxazolone Ill-5 with tart-butyl vinyl ether Ill-6 in the absence of catalyst (Scheme lV-10). Analysis of the crude reaction mixtures revealed an O-alkylated oxazole intermediate in the reaction involving 4- carbmethoxy-2-ethyl-5(4H)-oxazolone III-44 while the 4-carbmethoxy-2-phenyl- 5(4H)-oxazolone Ill-5 reaction produced only C-alkylated product. Upon standing, we observed the conversion of the O-alkylated intermediate to the C- alkylated product along with some degradation of the enol ether component. This observation infers that these ene-type reactions might proceed through an O to C migration, although this was never observed for any of the 2-aryl substituted oxazol-5(4H)-ones. 272 —__. N Et =/ Et 0:3 Et :5}. y}? u 4% O’Bu 002MB 002MB M8020 _Observed Intermediate III-44 III-45 III-45 O‘Bu — 1 PhYo =/ Ph 0 >_Ot3u PhYO I o Ill-6 WT / 0 l o N N N O‘Bu COzMe COzMe Me02C LNot detected by NMR‘ III-5 III-7 Scheme IV-10. Comparison of the reactivity between 2-aryl-oxazol-5(4H)-ones and 2-alkyloxazol-5(4H)-ones. K. Application of the alkylation methodology towards the synthesis of Salinosporamide A Salinosporamide A is a pyrrolidinone-based natural product isolated from the marine bacterium Salinospora in 2003 by Fenical and co-workers.39 Salinosporamide A exhibits remarkably selective and potent irreversible inhibition of the mammalian 26S proteasome, which is emerging as a novel target in anticancer therapy.“0 The compound structure and biological activity is similar to the naturally occurring metabolite Lactacystin (Chapter III). The structure of the natural product comprises of a highly functionalized fused bicyclic pyrrolidinone/B-lactone core, which is critical for its biological activity.”41 Many of the structural features necessary for the biological activity of Salinosporamide A have been identified through a combination of structure activity relationship 42,43 studies and x-ray crystallography. 273 The total synthesis of Salinosporamide A and other naturally related compounds has been completed by many research groups,“45 including that of E. J. Corey.46 Corey’s synthesis of Salinosporamide A consists of 17 linear steps starting from an N-acylated threonine methyl ester derivative (Scheme lV-11). During the synthesis, Corey and co-workers generate intermediate lV-34, which undergoes a Baylis-Hillman cyclization to yield the core pyrrolidinone scaffold of the natural product Salinosporamide A. MeO PMB H ' CO e QN .‘,COZM° SSteps Oj,N "II/228n118teps CI OWL /0 CH3 W-34 Salinosporamide A Scheme IV-1 1 . Corey’s synthesis of Salinosporamide A. To illustrate the potential of using ene-type alkylations of oxazol-5(4H)- ones and enol ethers towards the synthesis of compounds containing tart-alkyl amino hydroxy carboxylic acid cores, Robert A. Mosey of the Tape research lab synthesized Corey’s key intermediate IV-34 starting from compound lV-30A (Scheme lV-12).47 Mosey’s synthesis of compound lV-34 began with the cyclodehydration of lV-30A with MsCl to form oxazoline lV-35, which was subsequently reduced to amino alcohol lV-36 using sodium borocyanohydride. Protection of the primary alcohol found in lV-36 followed by deprotectlon of the tart-butyl group in amino ester lV-37 using aqueous phosphoric acids afforded amino alcohol IV-38.“3 Acylation of lV-38 with acrylyl chloride utilizing Corey’s 274 conditions followed by Dess Martin oxidation of the secondary alcohol produced Corey's key intermediate lV-34.“6 MeOzc reflux N-30A lV-35 N-36 92% 73% BnBr NaH DMF, r.t. MBCO M Fl’MBCO Me 2 e MBCOzMe 2 . H PO HN ill/OBn «lg—41— :Ni/OBn t 2: C) 4t :Q/OBn , r. . 2 2 ‘BuO N-34 98% £6.22 Scheme lV-12. Synthesis of Corey’s intermediate lV-34 starting from W-30A. L. Chiral Bransted acid catalyzed ene-type reactions of oxazol-5(4H)-ones and enol ethers as reported by Terada and co-workers. Chiral Bronsted acids have emerged as very useful catalysts for promoting a wide variety of enantioselective chemical transformations. This Class of organocatalysts is typically divided into two different classes: neutral and strong.37"5"9 Neutral chiral Bransted acids rely on their ability to hydrogen bond to Lewis basic heteroatom sites of the substrates involved, thus providing a chiral environment for the desired transformation to occur. These catalysts consist of chiral ureas, thioureas, alcohols, amides, and many more.37""5'49 On the contrary, 275 stronger chiral Bronsted acids are much more acidic allowing them to initially protonate the reactants producing a highly electrophilic substrate. Coordination of the chiral conjugate base of the catalyst to the newly formed electrophilic substrate establishes a chiral environment helping to afford the enantioselectivity observed in corresponding reactions. Found within this class of chiral Bransted acids are carboxylic acids, sulfonic acids, Lewis acid coordinated alcohols, ammonium salts and phosphoric acids.37"5'49 Given our results utilizing achiral phosphoric acids for increasing the diastereoselectivity of our ene-type reactions of oxazol-5(4H)—ones with enol ethers, we rationalized that chiral phosphoric acids may not only help increase the diastereoselectivity but also the enantioselectivity of these reactions. Protonation of the starting enol ether by the chiral phosphoric acid would result in the formation of an ion pair ion consisting of an oxonium ion stabilized by the chiral conjugate base of the phosphoric acid (Scheme lV-13). Studies by Terada and co-workers have demonstrated through a combination of DFT studies and deuterium labeling studies that oxonium ions are likely to be stabilized by phosphonates via hydrogen bonding interactions as illustrated in Scheme lV-13 below.‘ The acidic protons of the oxonium ion are proposed to interact with the anionic sites of the chiral conjugate base. Formation of the chiral oxonium I phosphonate ion pair results in the formation of a highly electrophilic center which would subsequently undergo an ene-type reaction while in the presence of the oxazol-5(4H)—one substrate. 276 H g + 0R3 L or J o’ I‘OH Ir ’ ‘ b0 CO‘Pl’O---H 0R3 o" T 06 _ _ _ H R1 le/O \||// 0 P/ N r N O - ’60 R R2 H '" 2 %A/H 0R3 R3 Scheme lV-13. Enantioselective ene-type reaction of oxazol-5(4H)-ones and enol ethers. Unfortunately, soon after our initial publication regarding the diastereoselective Bronsted acid catalysis of our ene-type methodology, Terada and co-workers reported the first enantioselective version of the chemistry.‘ They demonstrated the use of chiral BINOL phosphoric acids for promoting the enantioselective ene-type reaction of oxazol-5(4H)-ones with enol ethers (Scheme lV-14). Their methodology provided high yields of quaternary oxazolones with excellent diastereoselectivity and enantioselectivity. The products major diastereomer obtained in their reactions illustrated the same stereochemical relationship as we observed in our reactions. The quaternary oxazolones generated in the reactions were subsequently treated with sodium 277 methoxide to provide novel a,a-disubstituted a-amino acid derivatives with high levels of enantiomeric excess. Their studies indicated that aromatic substituents more electron-donating in nature at the An position helped to increase the stereoselectivity of the reaction. Enol ethers containing larger protecting groups (e.g. tart-butyl) exhibited higher diastereoselectivity than those with less sterically demanding protecting groups (e.g. n-butyl). Ar1 0 R2— 0R1 PAr1 O _ Al'1 0 be . mt“ % m r s __,...... a: . CH2Cl2 ‘ MeOH 2 Ar2 0°C AMI” Ar;(CO\2/Me 1 r 1 Yields up to 99% ‘ d.r. up to 33926 e.e. up to Scheme lV-14. Terada’s enantioselective ene-type reaction of oxazol-5(4H)- ones and enol ethers. 278 M. Experimental 1. General information. Reactions were carried out in flame-dried glassware under nitrogen atmosphere. All reactions were magnetically stirred and monitored by TLC with 0.25 pm pre- coated silica gel plates using either UV light or iodine to visualize the compounds. Column chromatography was carried out on Silica Gel 60 (230-400 mesh) supplied by EM Science. Yields refer to chromatographically and spectroscopically pure compounds unless otherwise noted. Infrared spectra were recorded on a Nicolet lRI42 spectrometer. 1H and 130 NMR spectra were recorded on a Varian Unity Plus-500 spectrometer. Chemiml shifts are reported relative to the residue peaks of the solvent (CDCI3: 7.24 ppm for 1H and 77.0 ppm for 13C) (DMSO-de: 2.49 ppm for 1H and 39.5 ppm for 13C). The following abbreviations are used to denote the multiplicities: s = singlet, d = doublet, t = triplet, q = quartet, p = pentet, and m = multiplet. Diastereomeric ratios were determined using the integration values obtained from 1H NMR. Gas chromatography / low resolution mass spectra were recorded on a Hewlet- Packard 5890 Series II gas chromatograph connected to a TRIO-1 EI mass spectrometer. HRMS were obtained at the Mass Spectrometry Facility of Michigan State University with a JEOL JMS HX-110 mass spectrometer. Elemental analysis data were obtained on a Perkin Elmer 2400 Series II CHNSIO analyzer. Melting points were obtained using an Electrothennal® capillary melting point apparatus and are uncorrected. 279 i‘ .‘ -. q 2. Materials. Reagents and solvents were purchased from commercial suppliers and used without further purification. Anhydrous methylene chloride, benzene, acetonitrile, and tetrahydrofuran were dispensed from a delivery system which passes the solvents through a column packed with dry neutral alumina. Ethyl vinyl ether IV- 17, tart-butyl vinyl ether III-6, 3,4-dihydro-2H-pyran Ill-10, 2-methoxy propene Ill- 18, trifluoroacetic anhydride, diphenyl phosphate lV-3, camphor sulfonic acid IV- 4, trifluoroacetic acid N-5, and 3,5—dinitrobenzoic acid lV-2 were all purchased from Sigma Aldrich, checked for purity and used without further purification. 3. General procedure for the synthesis of oxazol-5(4H)-ones lV-27 to lV-29: Oxazol—5(4H)—ones lV-27 through lV-29 were synthesized using a known literature procedure.‘ A suspension of carboxylic acid in anhydrous diethyl ether was treated with TFAA (2.2 equiv.) dropwise and then stirred for 1.5 hours. The reaction was then cooled to 0 °C and water (1.1 equiv.) was added dropwise, after which the ice bath was removed and the reaction stirred for an additional 1.5 hours. The reaction was cooled in a freezer for 30 minutes causing the oxazole-5(4H)-one product to precipitate out of solution which was collected via filtration and washed with cold diethyl ether. The oxazol-5(4H)-ones were then analyzed and used without further purification. 280 Us) COzMe COzMe 002Me IV-27 lV-28 lV-29 2-(4-methoxyphenyl)-4-carbomethoxy-5(4H)-oxazolone (IV-27), 2-(4- (trifluoromethyl)phenyl)-4-carbomethoxy-5(4H)-oxazolone (IV-28), and 2- benzyl-4—carbomethoxy-5(4l-l)-oxazolone (IV-29): Oxazol-5(4H)-ones lV-27 to lV-29 were all prepared according to a modified published procedure. The procedure used is very similar to the general procedure written above. These compounds were prepared and fully characterized by Robert A. Mosey. For further details regarding either the synthesis or characterization of these compounds, please see the PhD thesis of Robert A. Mosey or the supporting information of the following publication: Mosey, R. A.; Fisk, J. S.; Friebe, T. L.; Tepe, J. J. Org. Lett. 2008, 10, 825—828. 4. Synthesis of other enol others. The following enol ethers used in these studies were purchased and used as received: lV-6 (tart-butyl vinyl ether), lV-10 (3,4—dihydro—(2H)-pyran), lV-18 (2- methoxypropene), lV-17 (ethyl vinyl ether). Synthesis and characterization regarding Ill-28 (1-butoxyethyne) can be found in chapter Ill. Benzyl vinyl ether lV-16 was made Robert A. Mosey according to a literature procedure.50 All other enol ethers used in these studies were prepared as follows: 281 OBn (5: 1 mixture of diastereomers) lV-18 1-((prop-1-enyloxy)methyl)benzene (IV-18): This compound was made 5‘ A solution of aluminum chloride (3.12 9, according to a literature procedure. 23.40 mmol) in 60 mL of diethyl ether was treated with triethylamine (4.739, 46.79 mmol) dropwise over 10 minutes. The solution was stirred for 2 hours and then 1-((1-(benzyloxy)propoxy)methyl) benzene (3.00 g, 11.70 mmol) was added dropwise over 10 minutes. The solution was stirred under reflux for 24 hours after which it was washed with 10% NaOH (2 x 50 mL) and dried over magnesium sulfate. The solution was concentrated in vacuo and the resulting crude oil was purified via column chromatography (hexanes) yielding 0.51 g (29% yield) of the title compound as a yellowish oil (5 : 1 mixture of trans to cis isomers). The spectra matches that previously reported in the literature.52 Trans lsomer: 1H NMR (500 MHz) (CDCI3): 6 1.62 (dd, J1 = 1.5 Hz, J2 = 7 Hz, 3H), 4.43 (p, J = 7 Hz, 1H), 4.79 (s, 2H), 6.02 (dq, J1 = 7 Hz, J2 = 1.5 Hz, 1H), 7.28- 7.36 (m, 5H); 13c: NMR (125 MHz) (CDCl3): o 9.32, 73.48, 101.35, 127.26, 127.52, 128.44, 137.36, 145.17; Cis lsomer: 1H NMR (500 MHz) (CDCI3): 6 1.55 (dd, J1 = 1.5 Hz, J2 = 7 Hz, 3H), 4.70 (s, 2H), 4.86-4.92 (m, 1H), 6.30 (dq, J1 = 12.5 Hz, J2 = 1.5 Hz, 1H), 7.28-7.36 (m, 5H); 130 NMR (125 MHz) (CDCI3): 6 12.60, 71.09, 99.45, 127.52, 127.77, 128.43, 137.36, 146.26. 282 >_:/0Me lV-1 9 1-methoxy-2-methylprop-1-ene (IV-19): This compound was made according to literature procedure.53 A solution of 1,1-dimethoxy-2-methylpropane (16.2 g, 137 mmol) in 10 mL of quinoline was treated with a substoichiometric amount of toluene sulfonic acid (~75 mg). A simple distillation head was placed on the flask. The solution was heated and the desired product was distilled over (b.p. = 69 °C, lit b.p. = 70 °C) as it formed resulting in 8.5 g (72% yield) of the title compound as a clear oil. The spectra matches that previously reported in the literature. 1H NMR (500 MHz) (CDCla): 6 1.52 (s, 3H), 1.58 (s, 3H), 3.51 (s, 3H), 5.72 (s, 1H); 130 NMR (125 MHz) (opera): 5 14.78, 19.45, 59.15, 110.28, 141.40. LRMS(EI): m/z calcd for CsHmO 86.1 found, 86.2. 5. Synthesis of phosphoric acid derivatives. The following phosphoric acids used in these studies were purchased and used as received: lV-3 (diphenyl phosphate) and lV-15 ((R)—(-)-1,1’-binaphthyl-2,2’- diylhydrogenphosphate). All other phosphoric acids used in these studies were prepared as follows: 283 \\ lV-11 Bis(4-methoxyphenyl)phosphate (IV-11): A solution of 4-methoxy phenol (1.0 g, 8.06 mmol), triethyl amine (0.85 g, 8.46 mmol) and 20 mL of anhydrous CH2Cl2 was treated with POCI3 (0.62 g, 4.03 mmol) dropwise at 0°C over twenty minutes. The solution was stirred for 2 hours and then washed with 1M HCl (3 x 20 mL). It was dried over sodium sulfate and the solution was concentrated in vacuo. The resulting brown oil was dissolved into 10 mL of acetone and treated with 2 mL of water. After 2 hours of stirring, the solution was partitioned between CH2CI2 and water. Upon separation of the layers the CH2Cl2 phase was dried over sodium sulfate and concentrated in vacuo. The resulting solid was recrystallized using EtOAc and hexanes to yield 0.5 g (39% yield) of the title compound as a white crystalline solid. 1H NMR (500 MHz), CDCI3: 6 7.34 (s, 6H), 6.77 (d, J = 9 Hz, 4H), 7.04 (d, J = 9.5 Hz, 4H), 11.20 (bs, 1H); 13C NMR (125 MHz) CDCI3: 55.54, 55.62, 114.59, 114.60, 121.03, 121.07, 143.97, 0.. o ,9; 0 OH lV-12 144.03, 156.89, 156.90. Bis(2,6-dimethylphenyl)phosphate (IV-12): A solution of 2,6-dimethylphenol (0.98 g, 8.0 mmol), triethyl amine (0.85 g, 8.4 mmol) and 20 mL of anhydrous CH2Cl2 was treated with POCI3 (0.61 g, 4.0 mmol) dropwise at 0°C over twenty 284 minutes. The solution was stirred for 2 hours and then washed with 1M HCI (3 x 20 mL). It was dried over sodium sulfate and the solution was concentrated in vacuo. The resulting brown oil was dissolved into 10 mL of acetone and treated with 2 mL of water. After 2 hours of stirring, the solution was partitioned between CH2CI2 and water. Upon separation of the layers the CH2Cl2 phase was dried over sodium sulfate and concentrated in vacuo. The resulting solid was recrystallized using EtOAc and hexanes to yield 0.57 g (45% yield) of the title compound as a white crystalline solid. 1H NMR (500 MHz), CDCI3: 2.21 (s, 12H), 6.92-7.00 (m, 6H), 10.93 (bs, 1H); 130 NMR (125 MHz) CDCI3: 16.75, 16.70, 125.20, 125.21, 128.93, 128.94, 130.34, 130.37, 148.05, 148.12; LRMS(EI): m/z calcd for C16H1904P 306.3 found, 306.0. tBu lV-13 Bis(3,5-tert-butylphenyl)phosphate (IV-13): A solution of 3,5—di-terf-butyl phenol (1.65 g, 8.0 mmol), triethyl amine (0.85 g, 8.4 mmol) and 20 mL of anhydrous CH2CI2 was treated with POCI3 (0.61 g, 4.0 mmol) dropwise at 0°C over twenty minutes. The solution was stirred for 1 hour and then washed with 1M HCl (3 x 20 mL). It was dried over sodium sulfate and the solution was concentrated in vacuo. The resulting clear oil was dissolved into 10 mL of acetone and treated with 2 mL of water. After 12 hours of stirring, the solution was partitioned between CH2C|2 and water. Upon separation of the layers the 285 CH2CI2 phase was dried over sodium sulfate and concentrated in vacuo yielding the title compound as a clear oil. The product was used in subsequent steps without further purification. 1H NMR (500 MHz), CDCI3: 6 1.31 (s, 36H), 7.07 (s, 4H), 7.22 (s, 2H), 11.61 (bs, 1H); 13C NMR (125 MHz) CDCI3: 31.80, 35.22, 114.01, 119.50, 150.03, 152.31. C o. OOH N44 2,2’-biphenylphosphoric acid (IV-14): A solution of 2,2’biphenol (1.0 g, 5.37 mmol) and 8.7 mL of pyridine was treated dropwise with POCI3 (1.65 g, 10.8 mmol) at room temperature over 20 minutes. The solution was then stirred at room temperature for 4 hours before being cooled to 0 °C. The solution was then treated dropwise with 9 mL of water and stirred for an additional 30 minutes. The solution was then partitioned between CH2Cl2 and water. The CH2CI2 layer was washed with 1M HCI (3 x 50 mL), dried over sodium sulfate, and concentrated in vacuo to yield 0.3 g (23% yield) of the title compound as an off-white solid. The compound was used without further purification. 1H NMR (500 MHz), DMSO-d": 6 7.27 (d, J = 8 Hz, 2H), 7.38 (t, J = 7.5 Hz, 2H), 7.49 (t, J = 8 Hz, 2H), 7.61 (d, J = 3 Hz, 2H), 13.19 (bs, 1H); "’0 NMR (125 MHz) DMSO-ds: 121.43, 121.47, 125.89, 128.44, 129.74, 129.98, 148.16, 148.23; LRMS(EI): m/z calcd for C12H904P 248.2 found, 248.0. 286 6. Synthesis of tort-alkyl amino hydroxy carboxylic esters: General procedure for alkylation reaction. To a stirring suspension of oxazolone (0.5 mmol) in 20 mL of solvent were successively added enol ether (0.75 mmol) and diphenyl phosphate (0.05 mmol) under nitrogen atmosphere for the requisite amount of time as monitored by TLC. The reaction was washed successively with saturated NaHC03 solution and brine before being dried over M9804 and concentrated in vacuo. The crude reaction was diluted in THF and cooled to 0°C before NaBHz (1 mmol) and cold H2O were added. The reaction stirred at 0°C until complete by TLC. Saturated NH4Cl solution was added and the organic layer was extracted with CH2C|2 (x3). The combined organic extractions were dried (M9804) and concentrated. The crude reaction mixtures were purified via column chromatography on silica gel (diethyl ether I CH2CI2). Ph Yo H Q‘Bu H Q‘Bu I o O‘Bu N = + N 7 N + =/ ____’ Bz/ .‘ Bz/ )(\ 002m Meozc‘ OH MeO2C ’—OH Ill-5 Ill-6 N-1A lV-1B Major Minor Diastereomer Diastereomer Methyl-Z-benzamido-3-tert-butoxy-2-(hydroxymethyl)butarloate (IV-1): Using the general procedure, a suspension of 4-carbmethoxy-2—phenyl-5(4H)- oxazolone Ill-5 (0.10 g, 0.46 mmol), tort-butyl vinyl ether Ill-6 (69.11 mg, 0.69 mmol), diphenyl phosphate lV-3 (11.51 mg, 0.05 mmol) and 20 mL of anhydrous benzene was stirred at room temperature for 1 hour. After washings, the crude reaction intermediate was diluted in 2 mL THF and cooled to 0 °C before NaBH4 (34.8 mg, 0.92 mmol) and water (30 mL) were added. Purification via silica gel 287 chromatography (7% ether l 93% CH2CI2) afforded 0.14 g of the title compound (90% yield) as a 3:1 ratio of diastereomers. lV-1A (Major Diastereomer) (solid, mp. = 83 — 85 °C): 1H NMR (500 MHz) (CDCI3): 6 1.12 (s, 9H), 1.30 (d, J = 6.3 Hz, 3H), 3.79 (s, 3H), 3.95 (dd, J1 = 7.2 Hz, J2 = 11.6 Hz, 1H), 4.26 (dd, J1 = 6.3 Hz, J2 = 11.7 Hz, 1H), 4.37 (q, J = 6.3 Hz, 1H), 4.57 (t, J = 6.7 Hz, 1H), 7.41-7.46 (m, 3H), 7.50 (t, J = 7.4 Hz, 1H), 7.79 (d, J = 7.7 Hz, 2H). 13C NMR + DEPT (125 MHz) (CDCI3) 6: 17.9 (-CH3), 28.6 (- CH3), 52.6 (-CH3), 64.1 (—CH2), 69.8 (-CH), 72.1 (quaternary C), 75.2 (quaternary C), 127.1 (aromatic CH), 128.7 (aromatic CH), 131.8 (aromatic CH), 134.6 (aromatic quaternary C), 169.4 (quaternary C), 171.3 (quaternary C). IR (neat): 3412 cm", 3320 cm’1, 1736 cm", 1653 cm“, 1522 cm“, 1487 cm". MS (GCMS), calcd for CnH25N05 (M") — CH2OH: 292.15. Found: 290.9. Anal. Calcd. For C17H25N05: C, 63.14; H, 7.79; N, 4.33. Found: C, 63.04; H, 7.73; N, 4.55. lV-1B (Minor Diastereomer) (solid, mp. = 76 °c - 79 °C): 1H NMR (500 MHz) (CDCl3): 5 1.13 (d, J = 6.2 Hz, 3H), 1.16 (s, 9H), 3.74 (s, 3H), 3.83 (t, J = 11.8 Hz, 1H), 3.95 (dd, J1 = 2.8 Hz, J2 = 11.8 Hz, 1H), 4.27 (q, J = 6.2 Hz, 1H), 5.71 (dd, J1 = 2.8 Hz, J2 = 11.6 Hz, 1H), 7.26 (bs, 1H), 7.42 — 7.54 (m, 3H), 7.79 (dd, .l1 = 3.2 Hz, J2 = 5.3 Hz, 2H). 13C NMR (125 MHz) + DEPT (CDCI3) o: 19.1 (- CH3), 28.7 (-CH3), 52.7 (-CH3), 64.6 (-CH2), 68.6 (-CH), 70.0 (quaternary o), 74.8 (quaternary C), 127.0 (aromatic CH), 128.6 (aromatic CH), 131.8 (aromatic CH), 134.2 (aromatic quaternary C), 167.7 (quaternary C), 171.2 (quatemary C). IR (neat): 3405 cm", 3584 — 3156 cm", 1752 cm", 1669 cm", 1520 cm", 1487cm". 288 MS (GCMS), calcd for C17H25N05 (M’): 323.17. Found: 324.6. Anal. Calcd. For C17H25N05: C, 63.14; H, 7.79; N, 4.33. Found: C, 61.94; H, 7.94; N, 4.38. Ph Y0 o O H 9 H 9 N? + I __> Bz/N " + Bz/N ‘. Me02C ”—OH Meo2c‘ 0H CO2Me Ill-6 Ill-1o lV-22A N-ZZB Mq'or Minor Diastereomer Diastereomer Methyl-2-(benzamido)-2-(tetrahydro-2H-pyran-2-yl)-3-hydroxypropanoate (IV-22): Using the general procedure, a suspension of 4-carbmethoxy-2-phenyl- 5(4H)-oxazolone Ill-5 (0.1 g, 0.46 mmol), 3,4-dihydro-2H-pyran Ill-10 (58.0 mg, 0.69 mmol), and diphenyl phosphate lV-3 (11.5 mg, 0.05 mmol) in 20 mL of anhydrous benzene was stirred at 50 °C for 18 hours. After washings, the crude reaction intermediate was diluted in 2 mL THF and cooled to 0 °C before NaBH4 (34.8 mg, 0.92 mmol) and water (1 mL) were added. Purification via silica gel chromatography (20% ether l 80% CH2CI2) afforded 0.08 g of the title compound (57% yield) as a 1.6 :1 ratio of diastereomers. lV-22A (Major Diastereomer) (oil): 1H NMR (500 MHz) (CDCI3): 6 1.39-1.50 (m, 1H), 1.49-1.58 (m, 4H), 1.85-1.89 (m, 1H), 3.45 (td, J1 = 10 Hz, J2 = 3 Hz, 1H), 3.76 (s, 3H), 3.86 (t, J = 11.5 Hz, 1H), 3.93 (dd, J1 = 2 Hz, J2 = 11 Hz, 1H), 3.97- 4.00 (m, 1H), 4.02 (dd, J1 = 3 Hz, J2 = 12 Hz, 1H), 5.36 (dd, J1 = 3 Hz, J2 = 11 Hz, 1H), 7.07 (bs, 1H), 7.42-7.45 (m, 2H), 7.49-7.52 (m, 1H), 7.77-7.79 (m, 2H); “C NMR + DEPT (125 MHz) (cook): 5 22.56 (-CH2), 25.43 (-CH2), 25.78 (- CH2), 52.80 (-CO2CH3), 63.37 (-CH2), 68.94 (-CH2), 70.38 (quaternary C), 289 79.70 (-CH), 127.05 (aromatic -CH), 128.61 (aromatic -CH), 128.61 (aromatic — CH), 131.82 (aromatic -CH), 134.46 (quaternary aromatic C), 169.15 (quaternary 0), 171.05 (quaternary C); IR (neat): 3410 cm", 3360 cm", 2947 cm", 1743 cm‘ 1, 1657 cm"; HRMS (FAB): m/z calcd for C(3H22N05 [M + H], 308.1496; found, 308.1498. N-22B (Minor Diastereomer) (solid, mp. = 132 °C - 134 °C): 1H NMR (500 MHz) (CDCI3): 5 1.46-1.54 (m, 4H), 1.74-1.78 (m, 1H), 1.86-1.90 (m, 1H), 3.44 (t, J = 10Hz, 1H), 3.76 (s, 3H), 3.91 (d, J = 10.5 Hz, 1H), 4.01-4.07 (m, 2H), 4.19 (d, J = 11.5 Hz, 1H), 4.30 (t, J = 6.5 Hz, 1H), 7.30 (bs, 1H), 7.40-7.43 (m, 2H), 7.48-7.51 (m, 1H), 7.77-7.79 (m, 2H); “C NMR + DEPT (125 MHz) (CDCI3): 5 23.08 (-CH2), 25.68 (-CH2), 26.67 (-CH2), 52.64 (-COZCH3), 64.15 (-CH2), 67.92 (quaternary C), 69.47 (-CH2), 78.90 (-CH), 127.12 (aromatic —CH), 128.55 (aromatic -CH), 131.80 (aromatic -CH), 134.16 (quaternary aromatic C), 167.88 (quaternary C), 170.41 (quaternary C); IR (KBr): 3449 cm", 3358 cm", 2946 cm", 1743 cm", 1655 cm", 1527 cm"; HRMS (FAB): m/z calcd for 01.5H22No5 [M + H], 308.1495; found, 308.1498. \‘l/O QBn QBn O N + NOB” Ill-6 N-18 lV-23A lV-23B Mq'or Minor Diastereomer Diastereomer Methyl-Z-(benzamido)-3-(benzyloxy)-2-(hydroxymethyl)pentanoate (IV-23): Using the general procedure, a suspension of 4-carbmethoxy-2-phenyl-5(4H)- 290 oxazolone Ill-5 (100.0 mg, 0.46 mmol), 1-((prop-1-enyloxy)methyl)benzene lV-18 (102.3 mg, 0.69 mmol), and diphenyl phosphate lV-3 (11.5 mg, 0.05 mmol) in 20 mL of anhydrous benzene was stirred at room temperature for 21 hours. After washings, the crude reaction intermediate was diluted in 2 mL THF and cooled to 0 °C before NaBH4 (34.8 mg, 0.92 mmol) and water (1 mL) were added. Purification via silica gel chromatography (10% ether I 90% CH2CI2) afforded 0.08 g of the title compound (48% yield), separated as a 69 : 31 ratio of diastereomers. lV-23A (Major Diastereomer) (solid, mp. = 85 °C -— 87 °C): 1H NMR (500 MHz) (CDCI3): 6 1.09 (t, J = 7 Hz, 3H), 1.72-1.82 (m, 1H), 1.81-1.89 (m, 1H), 3.85 (s, 3H), 4.01 (dd, J1 = 5.5 Hz, J2 = 10 Hz, 1H), 4.23 (dd, J1 = 3 Hz, J2 = 9.5 Hz, 1H), 4.37-4.43 (m, 2H), 4.66 (d, J = 11.5 Hz, 1H), 4.71 (d, J = 11 Hz, 1H), 7.25-7.34 (m, 5H), 7.39 (bs, 1H), 7.40-7.44 (m, 2H), 7.50-7.54 (m, 1H), 7.73-7.75 (m, 2H); 13C NMR + DEPT (125 MHz) (CDCI3): 6 11.10 (-CH3), 24.53 (-CH2), 52.90 (- COZCH3), 64.48 (-CH2), 69.74 (quaternary C), 75.36 (~CH2), 81.59 (-CH), 127.04 (aromatic —CH), 127.61 (aromatic —CH), 127.77 (aromatic -CH), 128.40 (aromatic —CH), 128.62 (aromatic —CH), 131.90 (aromatic —CH), 133.84 (quaternary aromatic C), 137.85 (quaternary aromatic C), 167.70 (quaternary C), 171.35 (quaternary 6); IR (KBr): 3406 cm", 3390 cm", 2973 cm“, 1744 cm", 1651 cm", 1526 cm"; HRMS (FAB): m/z calcd for C21H26N05 [M + H], 372.1813; found, 372.1811. lV-23B (Minor Diastereomer) (oil): 1H NMR (500 MHz) (opera): 5 1.30 (t, J = 7.5 Hz, 3H), 1.52-1.64 (m, 2H), 3.77 (s, 3H), 3.88 (t, J = 11.5 Hz, 1H), 4044.09 291 (m, 2H), 4.51 (d, J = 11 Hz, 1H), 4.59 (d, J = 11 Hz, 1H), 5.60 (dd, J1 = 3 Hz, J2 = 11.5 Hz, 1H), 7.13 (bs, 1H), 7.27-7.36 (m, 5H), 7.40-7.43 (m, 2H), 7.49-7.52 (m, 1H), 7.74-7.76 (m, 2H); 13C NMR + DEPT (125 MHz) (CDCI3): 6 10.51 (-CH3), 23.26 (-CH2), 52.78 (-COZCH3), 64.01 (~CH2), 71.15 (quaternary C), 74.01 (- CH2), 82.72 (-CH), 127.08 (aromatic —CH), 127.89 (aromatic —CH), 128.08 (aromatic -CH), 128.49 (aromatic —CH), 128.66 (aromatic -CH), 131.89 (aromatic -CH), 134.34 (quaternary aromatic C), 137.36 (quaternary aromatic C), 169.18 (quaternary C), 171.51 (quaternary C); IR (neat): 3408 cm“, 3310 cm", 3030 cm“, 2951 cm", 1749 cm“, 1660 cm“; HRMS (FAB): m/z calcd for C21H26NO5 [M + H], 372.1813; found, 372.1811. e Nfo + — ’NYY COzMe MezOZNC +MBez02C Ill-5 lV-19 N-2C4A lV-ZB Ma'or Minor Diastereomer Diastereomer Methyl-Z-(benzamido)-3-(methoxy)-2-(hydroxymethyl)-4—methylpentanoate (IV-24): Using the general procedure, a suspension of 4-carbmethoxy-2-phenyl- 5(4H)-oxazolone Ill-5 (0.1 g, 0.46 mmol), 1-methoxy-2-methylprop-1-ene lV-19 (59.4 mg, 0.69 mmol), and diphenyl phosphate lV-3 (11.5 mg, 0.05 mmol) in 20 mL of anhydrous benzene was stirred at 50 °C for 15 hours. After washings, the crude reaction intermediate was diluted in 2 mL THF and cooled to 0 °C before NaBH4 (34.8 mg, 0.92 mmol) and water (1 mL) were added. Purification via silica 292 gel Chromatography (15% ether I 85% CH2CI2) afforded 0.05 g of the title compound (33% yield), separated as a 1.5:1 ratio of diastereomers. lV-24 (Major Diastereomer) (oil): 1H NMR (500 MHz) (CDCI3): 6 1.06 (dd, J1 = 7 Hz, J2 = 1.5 Hz, 6H), 1.86 (septet of doublets, J1 = 7 Hz, J2 = 3 Hz, 1H), 3.56 (s, 3H), 3.77 (s, 3H), 3.77-3.80 (m, 1H), 3.81 (d, J = 2.5 Hz, 1H), 4.21 (d, J = 7 Hz, 1 H), 4.40 (d, J = 11.5 Hz, 1H), 6.96 (bs, 1H), 7.41-7.45 (m, 2H), 7.49-7.52 (m, 1H), 7.74-7.76 (m, 2H); 13C NMR + DEPT (125 MHz) (CDCI3): 5 16.48 (-CH3), 22.19 (-CH3), 31.12 (-CH), 52.55 (~COZCH3), 62.52 (-CH3), 65.02 (-CH2), 68.29 (quaternary -C), 84.41 (-CH), 127.05 (aromatic -CH), 128.67 (aromatic —CH), 131.87 (aromatic -CH), 134.02 (quaternary aromatic C), 167.49, 171.88; IR (neat): 3416 cm", 2959 cm", 1736 cm", 1662 cm", 1514 cm", 1481 cm"; HRMS (FAB): m/z calcd for CrsH24N05 [M + H], 310.1652; found, 310.1654. lV-24B (Minor Diastereomer) (oil): 1H NMR (500 MHz) (CDCI3): 6 0.96 (d, J = 6.5 Hz, 3H), 1.05 (d, J = 7 Hz, 3H), 1.87 (septet of doublets, J1 = 6.5 Hz, J2 = 2 Hz, 1H), 3.41 (s, 3H), 3.80 (s, 3H), 3.87 (t, J = 11.5 Hz, 1H), 4.02 (dd, J1 = 12 Hz, J2 = 2.5 Hz, 1H), 5.84 (dd, J1 = 11.5 Hz, J2 = 2.5 Hz, 1H), 7.13 (bs, 1H), 7.43- 7.47 (m, 2H), 7.51-7.53 (m, 1H), 7.77-7.80 (m, 2H); 13C NMR + DEPT (125 MHz) (CDCI3): 5 16.35 (-CH3), 21.86 (-CH3), 29.33 (-CH3), 52.96 (-002CH3), 61.14 (- CH3), 63.78 (-CH2), 70.57 (quaternary C), 86.40 (-CH), 127.02 (aromatic -CH), 128.74 (aromatic —CH), 131.93 (aromatic -CH), 134.37 (quaternary aromatic - C), 168.54, 172.00; IR (neat): 3408 cm", 3306 cm", 2959 cm", 1749 cm“, 1658 cm", 1522 cm“; HRMS (FAB): m/z calcd for C15H24N05 [M + H], 310.1652; found, 310.1654. 293 Y0 O H 030 N + =_"—OBu ——-> BZ’N c 002,“ Meozc‘ OH Ill-5 Ill-29 lV-25 Methyl-2-(benzamido)-3-butoxy-2-(hydroxymethyl)but-3-enoate (IV-25): Using the general procedure, a suspension of 4£arbmethoxy-2-phenyl-5(4H)- oxazolone Ill-5 (0.11 g, 0.49 mmol) and butoxyethyne Ill-29 (72.7 mg, 0.74 mmol) in 20 mL of anhydrous dichloromethane was stirred at room temperature overnight. After concentration, the crude reaction intermediate was diluted in 3 mL THF and cooled to -41°C before anhydrous EtOH (1.5 mL) and NaBH4 (92.7 mg, 2.45 mmol) was added. Purification via silica gel Chromatography (15% ether / 85% CH2CI2) afforded 0.11 g of the title compound (67% yield) as an oil. 1H NMR (500 MHz) (CDCI3)I 6 0.90 (t, J = 7.4 Hz, 3 H), 1.33 - 1.41 (m, 2 H), 1.60 - 1.68 (m, 2 H), 3.40 (dd, J1 = 5.1 Hz, J2 = 8.7 Hz, 1 H), 3.70 - 3.76 (m, 2 H), 3.80 (s, 3 H), 4.15 (dd, J1 = 8.7 Hz, J2 = 11.3 Hz, 1 H), 4.26 (d, J = 3.7 Hz, 1 H), 4.32 (d, J = 3.7 Hz, 1 H), 4.47 (dd, J1 = 4.9 Hz, J2 = 11.4 Hz, 1 H), 7.33 (bs, 1 H), 7.41 - 7.46 (m, 2 H), 7.49 — 7.54 (m, 1 H), 7.77 — 7.81 (m, 2 H). 13C NMR + DEPT (125 MHz) (CDCl3): 6 13.7 (-CH3), 19.2 (-CH2), 30.6 (~CH2), 53.3 (-CH3), 64.1 (-CH2), 67.9 (-CH2), 84.6 (-CH2), 127.1 (aromatic CH), 128.7 (aromatic CH), 131.9 (aromatic CH), 134.0 (aromatic quaternary C), 156.6 (quaternary C), 167.2 (quaternary C), 170.8 (quaternary C); IR (neat): 3575-3125 cm", 3416 cm", 1742 cm", 1651 cm", 1287 cm", 1225 cm"; HRMS (FAB): m/z calcd for C17H24N05 [M+H], 322.1656; found, 322.1654. 294 methyl-Z-benzamido-3-benzyloxy-2- (hydroxymethyl)butanoatle (IV-20), methyl-2-benzamido—3-ethoxy-2-(hydroxymethyl)butanoatle (IV-21), Methyl- 2-(benzamido)-2-(hydroxymethyl)- 3-methoxy-3-methylbutanoate (IV-26), med-lyl-Z-(4-methoxybenzamido)-3-tert-butoxy-2-(hydroxymethyl)butanoalle (IV-30), methyl-Z-(4-(1rifluoromethyl)benzamido) -3-tert-butoxy -2- (hydroxymethyl) bumnoalle (IV-31), mefliyla-tert-butoxy-Z-(hydroxymefliyl)- 2-(propionamido)butanoate (IV-32) and mefllyI-Z-(Z-phenylacetamido)-3- ten-butoxy-Z-(hydroxymethyl)butanoatle (IV-33): Tart-alkyl amino hydroxy carboxylic esters lV-20, lV-21, lV-26, and lV-30 to lV-33 were all prepared using the general procedure written above. These compounds were prepared and fully Characterized by Robert A. Mosey. For further details regarding either the synthesis or characterization of these compounds, please see the Ph.D. thesis of Robert A. Mosey or the supporting information of the following publication: Mosey, R. A.; Fisk, J. 5.; Friebe, T. L.; Tepe, J. J. Org. Lett. 2008, 10, 825-828. 295 ._ uczanoo so 86on £22 .72 2:9". 3. o 2 cm 8 9. S 8 2 8 8 o: 8. co; at 2: 2m _ p - u b P — P — P P F - . — p p p p L P u b P FLl- - n P b . P llll' l I? Dr I. it: i _ _ E :l b p 0.0 o... o.~ od 9* 9m 0.0 0.» od o6 Q? of. 0.3 44 41:4 1 .lflalJ- 8.2622me 632 ( v.2 Q3... 36 : 296 67>. 2868 so good... «:22 .o.>_ use: u..- c or cm on 04 on oo as co co 0:. O9. 8.. at. cm? ova m6. m6 m... m.~ md mi m6 m6 m.» md md mdw m._._. m.N_. u . . 4 . d. a 6:60.235 352 9.2 :ol. 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Duos... >\Az \ a I cmm 301 .<.VN..>. uczanoo .0 6.60% E22 .0 ....>. 0.59.... o..- 0 or on on ow on on on em cm 0.... cm... 05 2... 8’ EN rbrrr5—pbP—p—pP-PFbP555L Lb hrh’p—lrh-n-nLl—u—nF 5121111.}...llll md. m6 m4 md 0.» m6 9m m6 m... mew md mdp m: ode 36.1.. 4 i 6Eoo66a5 5.5: (3.2 :0 L/VMDNOos ' ‘3 . z 2:“. I 302 .m¢~.>. 9:62:60 5 9.06% 1.22 ._....>. 9:5...— o 3 cm on av om cm on om ca 5.... 00.. 02 2... CE. o..~ m6- md m .. md m m m v mam m o m h nd md m.o.. 9:. a..- - r .3 6:60.235 .055 303 .32 2:828 .6 968m «22 5.2 2.5.“. 3. o 0.. ON on ow om cm on on ea o: car on? at our 9N m o- m6 m _. mN 06 m6 ad ad mK m6 m6 mdw m... md # . p L . .4 d . ‘lr L r L . p L ht »_‘+ p p . p . . . . b on.)— 10 @822 .. \Nm 3 :mO 304 N. REFERENCES (1) (2) (3) (4) (5) (6) (7) (8) (9) Venkatraman, J.; Shankaramma, S. C.; Balaram, P.; Design of folded peptides. Chem. Rev. 2001, 101, 3131-3152. (a) Cativiela, C.; Diaz-de-Vrllegas, M. D.; Stereoselective synthesis of quaternary alpha-amino acids. Part 1: Acyclic compounds. Tetrahedron- Asymmetry. 1998, 9, 3517-3599; (b) Cativiela, C.; Diaz-de-Villegas, M. D.; Stereoselective synthesis of quaternary alpha-amino acids. Part 2. Cyclic compounds. Tetrahedron-Asymmetry. 2000, 11, 645-732. Ohfune, Y.; Shinada, T.; Enantio- and diastereoselective construction of alpha,alpha-disubstituted alpha-amino acids for the synthesis of biologically active compounds. Eur. J. Org. Chem. 2005, 5127-5143. Terada, M.; Tanaka, H.; Sorimachi, K.; Enantioselective Direct Aldol-Type Reaction of Azlactone via Protonation of \finyl Ethers by a Chiral Bronsted Acid Catalyst. J. Am. Chem. Soc. 2009, 131, 3430. Vogt, H.; Brase, 8.; Recent approaches towards the asymmetric synthesis of alpha,alpha—disubstituted alpha-amino acids. Org. Bio. Chem. 2007, 5, 406-430. (a) O'Connor, S. J.; Liu, 2.; A concise synthesis of sterically hindered 3- amino-2-oxindoles. Synlett. 2003, 2135-2138; (b) Polinelli, 8.; Broxterman, Q. 8.; Schoemaker, H. E.; Boesten, W. H. J.; Crisma, M.; Valle, G.; Toniolo, C.; Kamphuis, J.; New Aspartame-Like Sweeteners Containing L- (Alpha-Me)Phe. BioMed. Chem. Lett. 1992, 2, 453-456. (a) Almond, H. R.; Manning, D. T.; Niemann, C.; Interaction of Alpha- Chymotrypsin with Several Alpha-Methyl—Alpha—Acylamino Acid Methyl Esters. Biochemistry. 1962, 1, 243; (b) Khosla, M. C.; Stachowiak, K.; Smeby, R. R.; Bumpus, F. M.; Piriou, F.; Lintner, K.; Ferrnandjian, 8.; Synthesis of [Alpha-Methyltyrosine-41Angiotensin-li - Studies of Its Conformation, Pressor Activity, and Mode of Enzymatic Degradation. Proc. Natl. Acad. Sci. U. SA. 1981, 78, 757-760. Kang, S. H.; Kang, S. Y.; Lee, H. S.; Buglass, A. J.; Total synthesis of natural tert-alkylamino hydroxy carboxylic acids. Chem. Rev. 2005, 105, 4537-4558. Abellan, T.; Chinchilla, R.; Galindo, N.; Guillena, G.; Najera, C.; Sansano, J. M.; Glycine and alanine imines as templates for asymmetric synthesis of alpha-amino acids. Eur. J. Org. Chem. 2000, 2689-2697. 305 (10) (11) (12) (13) (14) (15) (16) Groger, H.; Catalytic enantioselective Strecker reactions and analogous syntheses. Chem. Rev. 2003, 103, 2795-2827. Carter, H. E.; Azlactones. Org. React. 1946, 3, 198-239; (D) Mukerjee, A. K.; Azlactones - Retrospect and Prospect. Heterocycles. 1987, 26, 1077- 1097. Fisk, J. S.; Mosey, R. A.; Tepe, J. J.; The diverse chemistry of oxazol-5- (4H)-ones. Chem. Soc. Rev. 2007, 36, 1432-1440. Dejersey, J.; Willadse.P; Zemer, B.; Oxazolinone lnterrnediates in Hydrolysis of Activated N-Acylamino Acid Esters . Relevance of Oxazolinones to Mechanism of Action of Serine Proteinases. Biochemistry 1969, 8, 1959. Goodman, M.; Levine, L.; Peptide Synthesis Via Active Esters . 4 . Racemization + Ring-Opening Reactions of Optically Active Oxazolones. J. Am. Chem. Soc. 1964, 86, 2918-2922. (a) Obrecht, D.; Altorfer, M.; Lehmann, C.; Schonholzer, P.; Muller, K.; An efficient strategy to orthogonally protected (R)- and (S)-alpha- methyl(alkyl)serine-containing peptides via a novel azlactone/oxazoline interconversion reaction. J. Org. Chem. 1996, 61, 4080-4086; (b) Obrecht, D.; Bohdal, U.; Broger, C.; Bur, D.; Lehmann, C.; Ruffieux, R.; Schonholzer, P.; Spiegler, C.; Muller, K.; L-Phenylalanine Cyclohexylamide - a Simple and Convenient Auxiliary for the Synthesis of Optically Pure Alpha,AIpha-Disubstituted (R)-Amino and (S)-Amino Acids. Helv. Chim. Acta. 1995, 78, 563-580; (c) Obrecht, D.; Bohdal, U.; Ruffieux, R.; Muller, K.; A Reinvestigation of the Alpha-Alkylation of 4- Monosubstituted 2-Phenyloxazo|-5(4h)-Ones (Azlactones) - a General Entry into Highly Functionalized Alpha,AIpha-Disubstituted Alpha-Amino- Acids. Helv. Chim. Acta. 1994, 77, 1423-1429; (d) Obrecht, D.; Lehmann, C.; Ruffieux, R.; Schonholzer, P.; Muller, K.; Novel Open-Chain and Cyclic Conformationally Constrained (R)-Alpha,Alpha-Disubstituted and (S)- Alpha,Alpha-Disubstituted Tyrosine Analogs. Helv. Chim. Acta. 1995, 78, 1567-1587; (e) Obrecht, D.; Spiegler, C.; Schonholzer, P.; Muller, K.; Heimgartner, H.; Stierli, F .; A New General-Approach to Enantiomerically Pure Cyclic and Open-Chain (R)-Alpha,Alpha-Disubstituted and (S)- Alpha,AIpha-Disubstituted Alpha-Amino-Acids. Helv. Chim. Acta. 1992, 75, 1666-1696. (a) Trost, B. M.; Ariza, X.; Catalytic asymmetric alkylation of nucleophiles: Asymmetric synthesis of alpha-alkylated amino acids. Angew. Chem, Int. Ed. 1997, 36, 2635—2637; (b) Trost, B. M.; Ariza, X.; Enantioselective allylations of azlactones with unsymmetrical acyclic allyl esters. J. Am. Chem. Soc. 1999, 121, 10727-10737; (c) Trost, B. M.; Dogra, K.; 306 (17) (18) (19) (20) Synthesis of novel quaternary amino acids using molybdenum-catalyzed asymmetric allylic alkylation. J. Am. Chem. Soc. 2002, 124, 7256-7257. (a) Morgan, J.; Pinhey, J. T.; Reaction of Organolead Triaoetates with 4- Ethoxycarbonyl-2-Methyloxazol-5-One - the Synthesis of Alpha-Aryl and Alpha-Vinyl N-Acetylglycine Ethyl-Esters and Their Enzymatic Resolution. Tetrahedron Lett. 1994, 35, 9625-9628; (b) Morgan, J.; Pinhey, J. T.; Sherry, C. J.; Reaction of organolead triacetates with 4-ethoxycarbonyl-2- methyl-4,5-dihydro-1,3-oxazol-5—one. The synthesis of alpha-aryl— and alpha-vinyl-N-acetylglycines and their ethyl esters and their enzymic resolution. J. Chem. Soc, Perkin Trans. 1 1997,613-619. Liu, X. X.; Hartwig, J. F.; Palladium-catalyzed alpha-arylation of azlactones to form quaternary amino acid derivatives. Org. Lett. 2003, 5, 1915-1918. (a) Byun, H. 8.; Lu, X. 0.; Bittman, R.; Stereoselective total synthesis of serine palmitoyl—CoA transferase inhibitors. Synthesis. 2006, 2447-2474; (b) Li, M.; Wu, A. M.; Total synthesis of sphingofungin F. Synlett. 2006, 2985-2988. (a) Balskus, E. P.; Jacobsen, E. N.; alpha,beta-unsaturated beta-silyl imide substrates for catalytic, enantioselective conjugate additions: A total synthesis of (+)-lactacystin and the discovery of a new proteasome inhibitor. J. Am. Chem. Soc. 2006, 128, 6810-6812; (b) Brennan, C. J.; Pattenden, G.; Rescourio, G.; Formal synthesis of (+)-lactacystin based on a novel radical cyclisation of an alpha-ethynyl substituted serine. Tetrahedron Lett. 2003, 44, 8757-8760; (c) Corey, E. J.; Li, W. D. 2.; Total synthesis and biological activity of lactacystin, omuralide and analogs. Chem. Phan'n. Bull.1999, 47, 1-10; (d) Donohoe, T. J.; Sintim, H.; Sisangia, L.; Harling, J. D.; An efficient synthesis of lactacystin beta- lactone. Angew. Chem, Int. Ed. Engl. 2004, 43, 2293-2296; (e) Fukuda, N.; Sasaki, K.; Sastry, T. V. R. 8.; Kanai, M.; Shibasaki, M.; Catalytic asymmetric total synthesis of (+)-lactacystin. J. Org. Chem. 2006, 71, 1220-1225; (7) Gilley, C. B.; Buller, M. J.; Kobayashi, Y.; New entry to convertible isocyanides for the ugi reaction and its application to the stereocontrolled formal total synthesis of the proteasome inhibitor Omuralide. Org. Lett. 2007, 9, 3631-3634; (9) Green, M. P.; Prodger, J. C.; Hayes, C. J.; An enantioselective formal synthesis of the proteasome inhibitor (+)-lactacystin. Tetrahedron Lett. 2002, 43, 6609-6611;(h) Hayes, C. J.; Sherlock, A. E.; Selby, M. D.; Enantioselective total syntheses of (-)- clasto-lactacystin beta-lactone and 7-epi-(-)-clasto-lactacystin beta- Iactone. Org. Bio. Chem. 2006, 4, 193-195; (i) Masse, C. E.; Morgan, A. J.; Adams, J.; Panek, J. 8.; Syntheses and biological evaluation of (+)- lactacystin and analogs. Eur. J. Org. Chem. 2000, 2513-2528; (i) Ooi, H.; lshibashi, N.; lwabuchi, Y.; lshihara, J.; Hatakeyama, 8.; A concise route 307 (21) (22) (23) (24) (25) (26) to (+)-lactacystin. J. Org. Chem. 2004, 69, 7765-7768; (k) Yoon, C. H.; Flanigan, D. L.; Yoo, K. 8.; Jung, K. W.; Stereogenic evolution of clasto- lactacystin beta-lactone from L-serine. Eur. J. Org. Chem. 2007, 37—39. Kende, A. S.; Liu, K.; Brands, K. M. J.; Total Synthesis of (-)-Altemicidin - a Novel Exploitation of the Potier-Polonovski Rearrangement. J. Am. Chem. Soc. 1995, 117, 10597-10598. (a) Moloney, M. G.; Trippier, P. C.; Yaqoob, M.; Wang, 2.; The oxazolomycins: A structurally novel class of bioactive compounds. Curr. Dmg Disc. Tech. 2004, 1, 181-199; (b) Onyango, E. O.; Tsurumoto, J.; lmai, N.; Takahashi, K.; lshihara, J.; Hatakeyama, 8.; Total synthesis of neooxazolomycin. Angew. Chem, Int. Ed. 2007, 46, 6703-6705. (a) Bon'ssenko, L.; Groll, M.; 208 proteasome and its inhibitors: Crystallographic knowledge for drug development. Chem. Rev. 2007, 107, 687-717; (b) Fenteany, G.; Standaert, R. F.; Lane, W. S.; Choi, 8.; Corey, E. J.; Schreiber, S. L.; Inhibition of Proteasome Activities and Subunit- Specific Amino-Terminal Threonine Modification by Lactacystin. Science. 1995, 268, 726-731; (c) Powers, J. C.; Asgian, J. L.; Ekici, O. D.; James, K. E.; Irreversible inhibitors of serine, cysteine, and threonine proteases. Chem. Rev. 2002, 102, 4639-4750. (a) Banwell, M. G.; Crasto, C. F.; Easton, C. J.; Forrest, A. K.; Karoli, T.; March, D. R.; Mensah, L.; Naim, M. R.; O'Hanlon, P. J.; Oldham, M. D.; Yue, W. M.; Analogues of SB-203207 as inhibitors of tRNA synthetases. Bioorg. Med. Chem. Lett. 2000, 10, 2263-2266; (b) Takahashi, A.; Kurasawa, 8.; Ikeda, D.; Okami, Y.; Takeuchi, T.; Altemicidin, a New Acaricidal and Antitumor Substance .1. Taxonomy, Fermentation, Isolation and Physicochemical and Biological Properties. J. Antibiot. 1989, 42, 1556-1561. Steglich, W.; Hofle, 6.; Simple Synthesis of Acyl-Oxazolin-5-Ones from 5- Acyloxy-Oxazoles .2. Information on Hypemucleophilic Acylation Catalysts. Tetrahedron Lett.1970, 4727. (a) Shaw, 8. A.; Aleman, P.; Christy, J.; Kampf, J. W.; Va, P.; Vedejs, E.; Enantioselective TADMAP-catalyzed carboxyl migration reactions for the synthesis of stereogenic quaternary carbon. J. Am. Chem. Soc. 2006, 128, 925-934; (b) Shaw, 8. A.; Aleman, P.; Vedejs, E.; Development of chiral nucleophilic pyridine catalysts: Applications in asymmetric quaternary carbon synthesis. J. Am. Chem. Soc. 2003, 125, 13368-13369; (c) Thomson, J. E.; Rix, K.; Smith, A. 0.; Efficient N-heterocyclic carbene- catalyzed O- to C-acyl transfer. Org. Lett. 2006, 8, 3785-3788. 308 (27) (28) (29) (30) (31) (32) (33) (34) (35) Ruble, J. C.; Fu, G. C.; Enantioselective construction of quaternary stereocenters: Rearrangements of O-acylated azlactones catalyzed by a planar-chiral derivative of 4-(pyrrolidino)pyridine. J. Am. Chem. Soc. 1998, 120, 11532-11533. Trost, B. M.; Jakel, C.; Plietker, B.; Palladium-catalyzed asymmetric addition of pronucleophiles to allenes. J. Am. Chem. Soc. 2003, 125, 4438-4439. (a) Trost, B. M.; Lee, C. B.; A new strategy for the synthesis of sphingosine analogues. Sphingofungin F. J. Am. Chem. Soc. 1998, 120, 6818-6819; (b) Trost, B. M.; Lee, C. B.; gem-diacetates as carbonyl surrogates for asymmetric synthesis. Total syntheses of sphingofungins E and F. J. Am. Chem. Soc. 2001, 123, 12191-12201. Horn, W. 8.; Smith, J. L.; Bills, G. F.; Raghoobar, S. L.; Helms, G. L.; Kurtz, M. 8.; Marrinan, J. A.; Frommer, B. R.; Thornton, R. A.; Mandala, S. M.; Sphingofungin-E and Sphingofungin-F - Novel Serinepalmitoyl Transferase Inhibitors from Paecilomyces-Variotii. J. Antibiot. 1992, 45, 1692-1696. Zweerink, M. M.; Edison, A. M.; Wells, G. B.; Pinto, W.; Lester, R. L.; Characterization of a Novel, Potent, and Specific Inhibitor of Serine Palmitoyltransferase. J. Biol. Chem. 1992, 267, 25032-25038. Fisk, J. S.; Tepe, J. J.; lnterrnolecular Ene Reactions Utilizing Oxazolones and Enol Ethers. J. Am. Chem. Soc. 2007, 129, 3058-3059. Mosey, R. A.; Fisk, J. 8.; Friebe, T. L.; Tepe, J. J.; Synthesis of tert-Alkyl amino hydroxy carboxylic esters via an intermolecular ene-type reaction of oxazolones and enol ethers. Org. Lett. 2008, 10, 825-828. (a) Garcia, J.; Mata, E. G.; Tice, C. M.; Hormann, R. E.; Nicolas, E.; Albericio, F.; Michelotti, E. L.; Evaluation of solution and solid-phase approaches to the synthesis of libraries of alpha,alpha-disubst'rtuted-alpha- acylaminoketones. J. Comb. Chem. 2005, 7, 843-863; (b) Tice, C. M.; Hormann, R. E.; Thompson, C. 8.; Friz, J. L.; Cavanaugh, C. K.; Michelotti, E. L.; Garcia, J.; Nicolas, E.; Albericio, F.; Synthesis and SAR of alpha-acylaminoketone ligands for control of gene expression. Bioorg. Med. Chem. Lett. 2003, 13, 475-478. (a) Berkessel, A.; Cleemann, F.; Mukherjee, 8.; Muller, T. N.; Lex, J.; Highly efficient dynamic kinetic resolution of azlactones by urea-based bifunctional organocatalysts. Angew. Chem, Int. Ed. Engl. 2005, 44, 807- 811; (b) Berkessel, A.; Mukherjee, S.; Cleemann, F.; Muller, T. N.; Lex, J.; 309 (36) (37) (38) (39) (40) Second-generation organocatalysts for the highly enantioselective dynamic kinetic resolution of azlactones. Chem. Commun. 2005, 1898- 1900; (c) Gottwald, K.; Seebach, D.; Ring opening with kinetic resolution of azlactones by Ti-TADDOLates. Tetrahedron. 1999, 55, 723-738. (a) Kolka, A. J.; Napolitano, J. P.; Ecke, G. G.; The Ortho-Alkylation of Phenols. J. Org. Chem. 1956, 21, 712-713; (b) Kolka, A. J.; Napolitano, J. P.; Filbey, A. H.; Ecke, G. G.; The Ortho-Alkylation of Phenols. J. Org. Chem. 1957, 22, 642-646; (c) Ma, Q. 8.; Chakraborty, D.; Faglioni, F.; Muller, R. P.; Goddard, W. A.; Harris, T.; Campbell, C.; Tang, Y. C.; Alkylation of phenol: A mechanistic view. J. Phys. Chem. 2006, 110, 2246-2252; (d) Yadav, G. D.; Kumar, R; Alkylation of phenol with cyclohexene over solid acids: Insight in selectivity of 0- versus C- alkylation. Appl. Catal., A. 2005, 286, 61-70. Akiyama, T.; ltoh, J.; Fuchibe, K.; Recent progress in chiral Bronsted acid catalysis. Adv. Synth. Catal. 2006, 348, 999-1010. (a) Bihovsky, R.; Kumar, M. U.; Ding, 8.; Goyal, A.; Oxonium Ions in Organic-Synthesis - Condensation of 2,3—Dihydrofuran and 3,4-Dihydro— 2H-Pyran with 1,3-Dicarbonyl Compounds. J. Org. Chem. 1989, 54, 4291- 4293; (b) Nguyen, R. V.; Yao, X. Q.; Bohle, D. 8.; Li, C. J.; Gold- and silver-catalyzed highly regioselective addition of active methylenes to dienes, triene, and cyclic enol ethers. Org. Lett. 2005, 7, 673-675. Feling, R. H.; Buchanan, G. O.; Mincer, T. J.; Kauffman, C. A.; Jensen, P. R.; Fenical, W.; Salinosporamide A: A highly cytotoxic proteasome inhibitor from a novel microbial source, a marine bacterium of the new genus Salinospora. Angew. Chem, Int. Ed. Engl. 2003, 42, 355. (a) Cusack, J. C.; Liu, R.; Xia, L. J.; Chao, T. H.; Pien, C.; Niu, W.; Palombella, V. J.; Neuteboom, S. T. C.; Palladino, M. A.; NPl-0052 enhances tumoricidal response to conventional cancer therapy in a colon cancer model. Clin. Cancer Res. 2006, 12, 6758-6764; (b) Roccaro, A. M.; Leleu, X.; Sacco, A.; Jia, X. Y.; Melhem, M.; Moreau, A. 8.; N90, H. T.; Runnels, J.; Azab, A.; Azab, F.; Burwick, N.; Farag, M.; Treon, S. P.; Palladino, M. A.; Hideshima, T.; Chauhan, D.; Anderson, K. C.; Ghobrial, l. M.; Dual targeting of the proteasome regulates survival and homing in Waldenstrom macroglobulinemia. Blood. 2008, 111, 4752-4763; (c) Ruiz, 8.; Krupnik, Y.; Keating, M.; Chandra, J.; Palladino, M.; McConkey, D.; The proteasome inhibitor NPI-0052 is a more effective inducer of apoptosis than bortezomib in lymphocytes from patients with chronic Iymphocytic leukemia. Mol. Cancer Ther. 2006, 5, 1836-1843. 310 (41) (42) (43) (44) Williams, P. G.; Buchanan, G. O.; Feling, R. H.; Kauffman, C. A.; Jensen, P. R.; Fenical, W.; New cytotoxic salinosporamides from the marine actinomycete Salinispora tropica. J. Org. Chem. 2005, 70, 6196-6203. Groll, M.; Huber, R.; Potts, B. C. M.; Crystal structures of salinosporamide A (NPI-0052) and B (NPl-0047) in complex with the 208 proteasome reveal important consequences of beta-lactone ring opening and a mechanism for irreversible binding. J. Am. Chem. Soc. 2006, 128, 5136- 5141. Macherla, V. R.; Mitchell, 8. 8.; Manam, R. R.; Reed, K. A.; Chao, T. H.; Nicholson, 3.; Deyanat-Yazdi, 6.; Mai, B.; Jensen, P. R.; Fenical, W. F.; Neuteboom, S. T. C.; Lam, K. S.; Palladino, M. A.; Potts, B. C. M.; Structure-activity relationship studies of salinosporamide a (NPl-0052), a novel marine derived proteasome inhibitor. J. Med. Chem. 2005, 48, 3684- 3687. (a) Caubert, V.; Langlois, N.; Studies toward the synthesis of salinosporamide A, a potent proteasome inhibitor. Tetrahedron Lett.2006, 47, 4473-4475; (b) Caubert, V.; Masse, J.; Retailleau, P.; Langlois, N.; Stereoselective formal synthesis of the potent proteasome inhibiton salinosporamide A. Tetrahedron Lett. 2007, 48, 381-384; (c) Endo, A.; Danishefsky, S. J.; Total synthesis of salinosporamide A. J. Am. Chem. Soc. 2005, 127, 8298-8299; (d) Eustaquio, A. 8.; Moore, B. S.; Mutasynthesis of fluorosalinosporamide, a potent and reversible inhibitor of the proteasome. Angew. Chem, Int. Ed. Engl. 2008, 47, 3936-3938; (e) Hogan, P. C.; Corey, E. J.; Proteasome inhibition by a totally synthetic beta-lactam related to salinosporamide A and omuralide. J. Am. Chem. Soc. 2005, 127, 15386-15387; (f) Ling, T. T.; Macherla, V. R.; Manam, R. R.; McArthur, K. A.; Potts, B. C. M.; Enantioselective total synthesis of (-)- salinosporamide A (NPI-0052). Org. Lett. 2007, 9, 2289-2292; (9) Ma, G.; Nguyen, H.; Romo, 0.; Concise total synthesis of (+l-)-salinosporamide A, (+l-)—cinnabaramide A, and derivatives via a bis-cyclization process: Implications for a biosynthetic pathway? Org. Lett. 2007, 9, 2143-2146; (h) Margalef, I. V.; Rupnicki, L.; Lam, H. W.; Formal synthesis of salinosporamide A using a nickel-catalyzed reductive aldol cyclization- lactonization as a key step. Tetrahedron. 2008, 64, 7896-7901; (i) Mulholland, N. P.; Pattenden, G.; Walters, L. A. 8.; A concise total synthesis of salinosporamide A. Org. Bio. Chem. 2006, 4, 2845-2846; (j) Reddy, L. R.; Foumier, J. F.; Reddy, B. V. 8.; Corey, E. J.; An efficient, stereocontrolled synthesis of a potent omuralide-salinosporin hybrid for selective proteasome inhibition. J. Am. Chem. Soc. 2005, 127, 8974-8976; (k) Reddy, L. R.; Foumier, J. F.; Reddy, B. V. 8.; Corey, E. J.; New synthetic route for the enantioselective total synthesis of salinosporamide A and biologically active analogues. Org. Lett. 2005, 7, 2699-2701; (I) Shibasaki, M.; Kanai, M.; Fukuda, N.; Total synthesis of lactacystin and 311 (45) (46) (47) (48) (49) (50) (51) (52) (53) salinosporamide A. Chemistry-an Asian Joumal 2007, 2, 20—38; (m) Takahashi, K.; Midori, M.; Kawano, K.; lshihara, J.; Hatakeyama, 8.; Entry to heterocycles based on indium-catalyzed Conia-ene reactions: Asymmetric synthesis of (-)-salinosporamide A. Angew. Chem, Int. Ed. 2008, 47, 6244-6246. Akiyama, T.; Stronger bronsted acids. Chem. Rev. 2007, 107, 5744-5758. Reddy, L. R.; Saravanan, P.; Corey, E. J.; A simple stereocontrolled synthesis of salinosporamide A. J. Am. Chem. Soc. 2004, 126, 6230- 6231. Mosey, R. A.; Tepe, J. J.; New synthetic route to access (+l-) salinosporamide A via an oxazolone mediated ene-type reaction. Tetrahedron Lett. 2009, 50, 295-297. Li, B.; Beriiner, M.; Buzon, R.; Chiu, C. K. F.; Colgan, S. T.; Kaneko, T.; Keene, N.; Kissel, W.; Le, T.; Leeman, K. R.; Marquez, 3.; Morris, R.; Newell, L.; WundenNald, S.; Witt, M.; Weaver, J.; Zhang, Z. J.; Zhang, Z. L.; Aqueous phosphoric acid as a mild reagent for deprotection of tert- butyl carbamates, esters, and ethers. J. Org. Chem. 2006, 71, 9045-9050. Connon, S. J.; Chiral phosphoric acids: Powerful organocatalysts for asymmetric addition reactions to imines. Angew. Chem. Int. Ed. 2006, 45, 3909-3912. (a) Guillerrn, G.; Muzard, M.; Glapski, C.; Inactivation of S- adenosylhomocysteine hydrolase with haloethyl and dihalocyclopropyl esters derived from homoadenosine-6 '-carboxylic acid. Bioorg. Med. Chem. Lett. 2004, 14, 5799-5802; (b) Nielsen, T. E.; Le Quement, S.; Juhl, M.; Tanner, D.; Cu-mediated StiIIe reactions of sterically congested fragments: towards the total synthesis of zoanthamine. Tetrahedron. 2005, 61, 8013-8024. Barbot, F.; Miginiac, P.; New Way to \frnylic Ethers from Acetals. Helv. Chim. Acta. 1979, 62, 1451-1457. Dickinson, J. M.; Murphy, J. A.; Patterson, C. W.; Wooster, N. F.; A Novel Probe for Free-Radicals Featuring Epoxide Cleavage. J. Chem. Soc, Perkin Trans. 1. 1990, 1179-1184. Hamon, D. P. G.; Trenerry, V. C.; Cyclopropylidene Insertion Reactions - 2-Methoxy-1,2-Dimethylbicyclo[1,1,0]Butane. Aust. J. Chem. 1980, 33, 809-821. 312