Part 1: Stereoselective Synthesis of Quaternary Center BearingAzetines and β-Amino Acid Derivatives. Part 2: Natural andEnantiomeric Progesterone Analogues for the Treatment of TraumaticBrain Injury Open Access

Abstract

Part 1: A novel method for the synthesis of highly substituted -amino acid derivatives has been developed. The synthetic method described herein involves the use of a stable, four-membered azetine heterocycle that is generated from the reaction of a chlorotitanium enolate with an N-acyl imidazolidinone. The azetine has been shown to serve as a useful template for the introduction of an electrophile in order to form a quaternary center with complete diastereoselectivity. Treatment of the substituted azetine with benzoyl chloride and cleavage of its tethered chiral auxiliary afforded 2,2,3-amino acid derivatives in good yield.

Part 2: Pre-clinical and clinical research findings have revealed that the hormone progesterone, when acutely administered, can dramatically reduce cerebral edema, inflammation, tissue necrosis, and programmed cell death following traumatic brain injury (TBI). The use of progesterone as a therapeutic suffers however from a number of practical limitations, including its poor solubility and the instability of its formulation. Several chemically novel analogues of progesterone, its enantiomer, and its natural metabolite allopregnanolone have been synthesized and screened using both an in vitro assay and a whole animal model of TBI. All new derivatives demonstrated greatly improved solubility relative to progesterone and select compounds have shown equivalent effectiveness to progesterone in reducing cerebral edema after TBI.

Table of Contents

Table of Contents

Part 1: Stereoselective Synthesis of Quaternary Center Bearing Azetines and β-Amino Acid Derivatives...1

1. Introduction and Background...1

1.1 β-Amino Acids in Nature and Medicine...1 1.2 Asymmetric Synthesis of β-Amino Carbonyl Derivatives...3

1.2.1 General approaches...3 1.2.2 Stereoselective synthesis of β^2,2,3-amino acid derivatives...8

1.3 A New Route to β^2,2,3-Amino Acids from Azetines...14

1.3.1 Addition of chlorotitanium enolates to aldimines...14 1.3.2 The unexpected discovery of azetines...16

2. Results and Discussion...18

2.1 Azetine Alkylation Reactions with Traditional Auxiliaries...18

2.2 Imidazolidinone Based Auxiliaries...24

2.2.1 Free N-H auxiliaries...24 2.2.2 Substituted imidazolidinones...26 2.2.3 Ephedrine derived auxiliaries...32

2.3 Azetine Alkylation Reactions with Imidazolidinone Auxiliary...39

3. Conclusion...42

Part 2: Natural and Enantiomeric Progesterone Analogues for the Treatment of Traumatic Brain Injury...43

1. Introduction and Background...43

1.1 Traumatic Brain Injury: Incidence and Approaches to Treatment...43 1.2 Progesterone as Neuroprotectant...44

1.2.1 Pre-Clinical Evidence...44 1.2.2 Neuroprotective Mechanisms of Progesterone...46 1.2.3 Clinical Evidence...51

1.3 Progesterone as a Drug Candidate...53

1.3.1 Limitations of Natural Progesterone...53 1.3.2 Novel Progesterone Analogues...53

2. Results and Discussion...55

2.1 Synthesis of Progesterone Analogue Compounds...55

2.1.1 C-3 and C-20 Progesterone Derivatives...55 2.1.2 Allopregnanolone Series Derivatives...57 2.1.3 Progesterone Prodrug Compounds...59 2.1.4 Enantiomeric Progesterone Compounds...64

2.2 Solubility and Biological Testing Data...67

2.2.1 Solubility Data...67 2.2.2 In Vitro Assay Screening Data...68 2.2.3 In Vivo Cerebral Edema Assay Data...69 2.2.4 Pharmacokinetic Screening Data...71

3. Conclusion...74

Supporting Information...75

S1. Experimental Procedures and Characterization Data...75

S1.1 Part 1: Azetine Chemistry...75

General...75 Oximes...76 Traditional Auxiliaries, Azetines, and Alkylation Products...79 Free N-H Auxiliary Compounds...88 Isocyanate Derived Auxiliaries and Azetines...103 Ephedrine Derived Auxiliaries and Azetines...141 Azetine Addition Products...155 Optimization Study Raw Data and Analysis...178

S1.2 Part 2: Progesterone Chemistry...179

C-3 Progesterone Derivatives...179 C-20 Progesterone Derivatives...188 Allopregnanolone Derivatives...192 Progesterone Prodrug Series Compounds...203 Enantiomeric Progesterone Synthesis...228 Enantiomeric Progesterone Derivatives...236 Cerebral Edema Assay Methods...238

S2. References...240

Part 1: Stereoselective Synthesis of Quaternary Center Bearing Azetines and β-Amino Acid Derivatives List of Figures Figure 1. Pharmacologically active β-amino acid containing natural products...2 Figure 2. Designation of representative amino acids...3 Figure 3. ORTEP structures for cis and trans azetines derived from N-phenyl substituted imidazolidinone chiral auxiliary...28 Figure 4. Yield data for different imidazolidinone auxiliary aryl R groups...30 Figure 5. Randomized 3 factor/2 level optimization study results: response surface plot for total azetine yield...34 Figure 6. Crystal structure for alkylation addition product 146f...40 Figure 7. Substituent directed approach of electrophile to hypothesized azetine transition state...41

List of Schemes Scheme 1. General approaches to β-amino acid synthesis...4 Scheme 2. Yuan and Williams application of Arndt-Eistert reaction...4 Scheme 3. Asymmetric conjugate addition reactions of amines and acrylates...5 Scheme 4. Catalytic asymmetric reduction of enamine esters...6 Scheme 5. Chiral auxiliary mediated diastereoselective imine/enolate condensations...7 Scheme 6. Wyatt addition of imine to chlorotitanium enolate of oxazolidinone...8 Scheme 7. Ellman addition of ester enolate to tert-butanesulfinyl imine...9 Scheme 8. Kunz Mannich reaction with carbohydrate derived aldimine...9 Scheme 9. Waldmann's use of N,N-phthaloylamino acids as chiral auxiliaries...10 Scheme 10. Kobayashi catalytic enantioselective Mannich-type reaction...11 Scheme 11. Seebach and Podlech alkylation of α-methyl-β³-homophenylalanine...12 Scheme 12. Davies' tandem conjugate addition-alkylation sequence...12 Scheme 13. Cardillo ethylation of perhydropyrimidin-4-one...13 Scheme 14. Crimmins' asymmetric aldol additions with chlorotitanium enolates...14 Scheme 15. Effect of imine substituent on diastereoselectivity of enolate addition...15 Scheme 16. Imine/enamine tautomerization and polymerization...16 Scheme 17. Addition of oxime ether to chlorotitanium enolate gives an azetine...16 Scheme 18. Hydrolytic ring opening of azetine and cleavage of auxiliary...17 Scheme 19. Hypothesized route to β^2,2,3-amino acids and related compounds...17 Scheme 20. Synthesis of thiazolidinethione auxiliary 72 and oxime ether 77a...18

Scheme 21. Thiazolidinine-2-thione ring opening under strongly basic conditions...19 Scheme 22. Synthesis of oxazolidinethione auxiliary 90 and azetine 91...19 Scheme 23. Synthesis of thiazolidinone auxiliary 92...20 Scheme 24. First azetine alkylation reactions with thiazolidinone auxiliary...20 Scheme 25. Synthesis and alkylation of oxazolidinone azetine...21 Scheme 26. Competitive reaction pathways to either azetine or pyrimidinone...22 Scheme 27. Hypothesized advantages of imidazolidinone based chiral auxiliary...23 Scheme 28. Synthesis of imidazolidinone auxiliary 113...24 Scheme 29. Synthesis of imidazolidinethione auxiliary 120...25 Scheme 30. Retrosynthetic approach to imidazolidinone auxiliary from isocyanate...26 Scheme 31. Taguchi's Michael addition of substituted imidazolidinone...27 Scheme 32. Isocyanate route to imidazolidin-2-one auxiliaries...28 Scheme 33. Synthesis of ephedrine derived chiral auxiliary 132...32 Scheme 34. Azetine formation using ephedrine derived auxiliary 132...32 Scheme 35. Preparation of azetines from auxiliary 132 under optimized conditions...34 Scheme 36. Synthesis and azetine reactions of cyclohexyl auxiliary 136...36 Scheme 37. Titanium imine 142 as a common intermediate for azetine synthesis...37 Scheme 38. Hypothesized open transition states leading to azetines 133 and 134...38 Scheme 39. Hypothesized transition state leading to trans thiophenyl azetine 134f...39 Scheme 40. Alkylation/ring opening of 133a to give β^2,2,3-amino carbonyl derivatives...39 Scheme 41. Auxiliary cleavage to access β^2,2,3-amino acid and ester derivatives...41 Scheme 42. Imidazolidinone auxiliaries for the synthesis of azetines and β^2,2,3-amino acid derivatives...42

List of Tables Table 1. Azetine/pyrimidinone product distribution and yield relative to auxiliary type...22 Table 2. Azetine yield and selectivity data for variably substituted imidazolidinones...29 Table 3. Effect of Lewis acid on yield and diastereoselectivity of azetine reaction with auxiliary 126a...31 Table 4. Experimental design and raw data for optimization study...33 Table 5. Gram scale additions of oximes to ephedrine derived auxiliary 132...35 Table 6. Alkylation of α-substituted azetines and hydrolysis to β-amino acid derivatives...40 Table 7. Crystal data and structure refinement for compound 127a...118 Table 8. Atomic coordinates ( x 10⁴) and equivalent isotropic displacement parameters (Ųx 10³) for 127a...119 Table 9. Bond lengths [Å] and angles [°] for 127a...120 Table 10. Anisotropic displacement parameters (Ųx 10³) for 127a...124 Table 11. Hydrogen coordinates ( x 10⁴) and isotropic displacement parameters (Ųx 10³) for 127a...125 Table 12. Torsion angles [°] for 127a...126 Table 13. Crystal data and structure refinement for 128a...129 Table 14. Atomic coordinates ( x 10⁴) and equivalent isotropic displacement parameters (Ųx 10³) for 128a...130 Table 15. Bond lengths [Å] and angles [°] for 128a...131 Table 16. Anisotropic displacement parameters (Ųx 10³) for 128a...135 Table 17. Hydrogen coordinates ( x 10⁴) and isotropic displacement parameters (Ųx 10³) for 128a...136 Table 18. Torsion angles [°] for 128a...137 Table 19. Crystal data and structure refinement for 146f...160 Table 20. Atomic coordinates ( x 104) and equivalent isotropic displacement parameters (Ųx 10³) for 146f...162 Table 21. Bond lengths [Å] and angles [°] for 146f...164 Table 22. Anisotropic displacement parameters (Ųx 10³) for 146f...170 Table 23. Hydrogen coordinates ( x 10⁴) and isotropic displacement parameters (Ųx 10³) for 146f...173 Table 24. Hydrogen bonds for 146f [Å and °]...175

Part 2: Natural and Enantiomeric Progesterone Analogues for the Treatment of Traumatic Brain Injury List of Figures Figure 1. Clincally studied compounds with negligible effects for the treatment of TBI...44 Figure 2. Initial findings correlate progesterone levels to reduction in cerebral edema...45 Figure 3. Overview of approach to the development of progesterone analogues...54 Figure 4. Imidazolidin-4-one tethered PROG derivative 36...63 Figure 5. PROG analogue solubilities in phosphate buffered saline...68 Figure 6. Cerebral edema level assay data for PROG and test compounds...70 Figure 7. Pharmacokinetic data for P1-31 and P1-33 IV dosed at 10 mg/kg...72 Figure 8. Pharmacokinetic data for P1-131 IV dosed at 10 mg/kg...72 Figure 9. Pharmacokinetic data for P1-186 IV dosed at 10 mg/kg...73 Figure 10. Water soluble lead compounds for the treatment of TBI...74

List of Schemes Scheme 1. Enzymatic conversion of progesterone (1) to allopregnanolone (2)...49 Scheme 2. Preparation of C-3 amino acid tethered PROG analogues...55 Scheme 3. Synthesis of analogue P1-163 by Mitsunobu inversion...56 Scheme 4. Preparation of C-20 valine tethered PROG analogue P1-57...56 Scheme 5. Preparation of 5α ALLO isomer derivatives P1-123 and P1-131...58 Scheme 6. Preparation of 5β ALLO isomer derivatives P1-133 and P1-135...58 Scheme 7. Hypothetical pathway leading from PROG prodrug to PROG in vivo...59 Scheme 8. Selective PROG ketalization and oxime formation...60 Scheme 9. Preparation of oxime based PROG prodrug compounds P1-185 and P1-186...61 Scheme 10. Hydrazide formation and C-20 ketal removal with thiourea...62 Scheme 11. Fmoc deprotection and completion of hydrazide prodrug P2-29 synthesis...63 Scheme 12. Synthesis of enone 45, CD ring fragment of ent-PROG...65 Scheme 13. Preparation of β-keto ester annulating agent 50...65 Scheme 14. Completion of ent-PROG (57) synthesis...66 Scheme 15. Synthesis of ent-PROG derivative P2-13...67

List of Tables Table 1. Major findings from Phase II clinical trial of progesterone treatment for TBI...52 Table 2. Reduction in cortical neuron cell death caused by glutamate toxicity...69 Table 3. Crystal data and structure refinement for compound 36...214 Table 4. Atomic coordinates ( x 10⁴) and equivalent isotropic displacement parameters (Ųx 10³) for compound 36...215 Table 5. Bond lengths [Å] and angles [°] for compound 36...216 Table 6. Anisotropic displacement parameters (Ųx 10³) for compound 36...223 Table 7. Hydrogen coordinates ( x 10⁴) and isotropic displacement parameters (Ųx 10³) for compound 36...224

Table 8. Torsion angles [°] for compound 36...225

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School	Laney Graduate School
Department	Chemistry
Degree	PhD
Submission	Dissertation
Language	English
Research Field	Chemistry, Organic
Keyword	allopregnanolone azetine imidazolidinone progesterone traumatic brain injury TBI
Committee Chair / Thesis Advisor	Liotta, Dennis C, Emory University
Committee Members	McDonald, Frank, Emory University Liebeskind, Lanny S, Emory University

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