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Design and stereoselective synthesis of novel bicyclicbeta-turn dipeptide mimetics and cis-4-substitutedproline
analogues for peptides and peptidomimetics
Item Type text; Dissertation-Reproduction (electronic)
Authors Zhang, Junyi
Publisher The University of Arizona.
Rights Copyright © is held by the author. Digital access to this materialis made possible by the University Libraries, University of Arizona.Further transmission, reproduction or presentation (such aspublic display or performance) of protected items is prohibitedexcept with permission of the author.
Download date 20/03/2021 01:22:53
Link to Item http://hdl.handle.net/10150/289964
1
Design and Stereoselective Synthesis of Novel Bicyclic P-Tum
Dipeptide Mimetics and cw-4-Substituted Proline Analogues for
Peptides and Peptidomimetics
by
Junyi Zhang
A Dissertation Submitted to the Faculty of the DEPARTMENT OF CHEMISTRY
In Partial Fulfillment of the Requirements
For the Degree of
DOCTOR OF PHILOSOPHY
In the Graduate College
THE UNIVERSITY OF ARIZONA
2003
UMI Number: 3107058
UMI UMI Microform 3107058
Copyright 2004 by ProQuest Information and Learning Company.
All rights reserved. This microform edition is protected against
unauthorized copying under Title 17, United States Code.
ProQuest Information and Learning Company 300 North Zeeb Road
P.O. Box 1346 Ann Arbor, Ml 48106-1346
2
THE UNIVERSITY OF ARIZONA ®
GRADUATE COLLEGE
As members of the Final Examination Committee, we certify that we have
read the dissertation prepared by Junvi Zhang
entitled Design and Stereoselective Synthesis of Novel Bicyclic
beta—Turn Dipeptide Mimetics and cis—4—Substituted Proline
Analogues for Peptides and Peptidomimetics
and recommend that it be accepted as fulfilling the dissertation
requirement for the Degree of Doctor of Philosophy
Professor Victor J. Hruby
Professor Robert B. Bates
Professor Indraneel Ghosh o
Professor Zhiping Zheng
Date
Date
Date
Date
Final approval and acceptance of this dissertation is contingent upon
the candidate's submission of the final copy of the dissertation to the
Graduate College.
I hereby certify that I have read this dissertation prepared under my
direction and recommend that it be accepted as fulfilling the dissertation
requirement. / I /' i /?s i / /
Professor Victor J. Hruby'^^^^^w^/ Dissertation DirectorJ] Date
3
STATEMENT BY AUTHOR
This thesis has been submitted in partial fulfillment of requirements for an
advanced degree at The University of Arizona and is deposited in the University Library
to be made available to borrowers under rules of the Library.
Brief quotations from this thesis are allowable without special permission,
provided that accurate acknowledgment of source is made. Requests for permission for
extended quotation from or reproduction of this manuscript in whole or in part may be
granted by the head of the major department or the Dean of the Graduate College when in
his or her judgment the proposed use of the material is in the interests of scholarship. In
all other instances, however, permission must be obtained from the author.
SIGNED:
4
ACKNOWLEDGEMENTS
I would to express my sincerest thanks to the members of my committee members,
Professors Victor J. Hruby, Jacquelyn Gervay-Hague, Robert B. Bates, Zhiping Zheng.
Arnold R. Martin, Indraneel Ghosh, and Jon D. Rainier, for their advice and support. In
particular, I would like to thank Dr. Hruby for his assistance, excellent guidance, and
patience, and for his continued financial support throughout the course of my research.
This dissertation would not have been completed without them.
I would also like to thank the post-doctorates, staff and visiting professors in Dr.
Hruby's group: Drs. Wei Wang, Yeonsun Lee, Scott Cowell, Minying Cai, Trivedi Dev.
Josef Vagner, Ruben Vardanyan, Zhanna Zhilina, and Alexander Mayorov. I am also
very grateful to the former and current graduate students in this group: Dr. Xuejun Tang,
Dr. Chiyi Xiong, Jinfa Ying, Dr. Xuyuan Gu, and Dr. Richard Agnes.
Dedicated to
My wife Xiang Gao
My parents Xingkang Zhang and Quandi Gao
6
Table of Contents
List of Figures 9
List of Schemes 11
List of Tables 13
ABSTRACT ....14
CHAPTER 1. DESIGN OF PEPTIDOMIMETICS FROM PEPTIDE
LEADS CONFORMATIONAL AND TOPOGRAPHICAL
CONSIDERATIONS
1.1 Introduction 16
1.2 Conformational and Topographical Features of Peptides — Challenges in
the Studies of Biologically Active Conformation 19
1.3 Strategies in the Design of Pcptidomimetics 21
1.3.1. Modification of Amino Acid Side Chains 21
1.3.2 Modification of the Peptide Backbone 23
1.3.3 Global Restrictions of Conformation in Peptides and Pcptidomimetics...
25
CHAPTER 2. EFFICIENT AND STEREOSELECTIVE SYNTHESIS OF
NOVEL C/5-4-SUBSTITUTED PROLINE ANALOGUES
2.1 Introduction 28
2.2 Background and General Approach to 4-Substituted Proline
Derivatives 30
7
2.3 Stereoselective Synthesis of c/5-4-Substituled Proline Derivatives 31
2.4 Conclusion 35
2.5 Experimental Section 35
CHAPTERS. DESIGN AND SYNTHESIS OF CONFORMATIONALLY
CONSTRAINED REVERSE-TURN PEPTIDOMIMETICS OF
LEU-ENKEPHALIN
3.1 Introduction 45
3.2 Design of Leu-enkephalin Mimetics 47
3.3 Synthesis of Azabicyclo[X.Y.O]Alkane Amino Acids 52
3.4 Synthesis of Novel 4-Substituted Unsaturated and Saturated
Indolizidinone Amino Acids 54
3.5 Synthesis of Novel 4-Substituted Saturated Indolizidinone Amino
Acids 64
3.6 Synthesis of Leu-Enkephalin Mimetics 68
3.7 Conclusion 70
3.8 Experimental Section 71
CHAPTER 4. STEREOSELECTIVE SYNTHESIS OF 4, 8-DISUBSTITUTED
AZABICYCLO[4.3.0]NONANE AMINO ACIDS AS
PEPTIDOMIMETICS SCAFFOLDS OF MELANOCORTIN
RECEPTOR LIGANDS
4.1 Introduction ..78
4.2 Design of Conformationally Restricted Analogues. 79
8
4.3 Synthesis of Disubstituted Indolizidinone Amino Acids 82
4.3.1 B ackground and General Approach 82
4.3.2 Synthesis of P-Substituted Pyroglutamic Acid Ester. 83
4.3.3 Synthesis of 8-Substituted Azabicyclo[4.3.0]alkane Amino Acids 90
4.3.4 Synthesis of 4, 8-Disubstituted Indolizidinone Amino Acid Ester. 93
4.3.5 Synthesis of Pcplidomimetics of the Core Sequence in Melanotropin
Peptides...... 94
4.4 Future work 97
4.5 Conclusion 98
4.6 Experimental section 99
REFERENCES 118
APPENDIX; and NMR Spectra of Compounds 131
9
List of Figures
Figure 1-1. A de novo approach for peptidomimetic design ....18
Figure 1-2. Conformations of peptides 19
Figure 1-3 Torsional angles of an a-helix, (3-sheet and type I and I' P-turn
conformations 20
Figure 1-4 x-Constrained amino acids synthesized in the Hruby group 23
Figure 1-5 The most frequent modifications to the peptide backbone 24
Figure 1-6 Side-chain-to-side-chain cyclizations 25
Figure 1-7 Cyclic constrains 26
Figure 1-8 (3- and y-tuni mimetics of various types 27
Figure 2-1 Examples of proline-related ACE inhibitors 29
Figure 2-2 Transition states in alkylations 32
Figure 3-1 Structure of DPDPE 47
Figure 3-2 Schematic drawing of the three enkephalin conformations 48
Figure 3-3 Peptidomimetics of Leu-enkephalin 49
Figure 3-4 Reverse turn peptidomimetics of Leu-enkephalin 49
Figure 3-5 Azabicyclo[X. Y.Olalkane amino acids 50
Figure 3-6 General structure illustrating the dihedral angles constrained by an
azabicyclofX.Y.OJalkane amino acid in a peptide 51
Figure 3-7 Azabicycloalkane amino acids 53
Figure 3-8 Dipeptide mimetics designed for Leu-enkephalin peptidomimetics. 54
10
Figure 3-9 Transition state model for the generation of 12.... 59
Figure 3-10 The X-ray structure of compound 19a 63
Figure 3-11 Conformation of compound 18a suggested by modeling study 67
Figure 3-12 Classification of P-tums 67
Figure 4-1 Structure of MT-ll ...79
Figure 4-2 Core sequence of melanotropin peptides 80
Figure 4-3 The X-ray structure of compound 34... 90
11
List of Schemes
Schemc 2-1 Strategies for the asymmetric hydrogenation of pyrroline
intermediates 30
Scheme 2-2 Strategy for the preparation of cw-4-substituted proUne derivatives 1....
31
Scheme 2-3 Approach to ds-4-substituted proline derivatives 31
Scheme 2-4 Selective reduction of y-substituted glutamic acid ester 33
Scheme 2-5 Deprotection of c/.v-4-substituded proline derivatives 34
Scheme 2-6 Cyclization to pyroglutamic acid ester derivative 9 35
Scheme 3-1 Retrosynthetic analysis 56
Scheme 3-2 Synthesis of 4-subslituted unsaturated indolizidinone amino acid
esters 58
Scheme 3-3 Bromination of dehydroamino acids 60
Scheme 3-4 Tautomerization of a-bromoimine 61
Scheme 3-5 Mechanism of isomerization 62
Scheme 3-6 Hydrogenation of 18a and 19a ...65
Scheme 3-7 Synthesis of Leu-Enkephalin mimetic 24 69
Scheme 3-8 Decompose of 24 in TFA/HoO 70
Scheme 4-1 Analogues of melanotropin peptide core sequence 81
Scheme 4-2 Retrosynthetic analysis of mimetic 26 83
Scheme 4-3 Synthesis of Michael acceptor 30 84
12
Scheme 4-4 Metal-chelated intermediate in Michael addition ...86
Scheme 4-5 Preparation of the chiral Ni(ll) complex (S)-32 87
Schcme 4-6 Selective electrophilic attack on the complex enolate 88
Scheme 4-7 Asymmetric Michael addition between 30 and 32 88
Scheme 4-8 Hydrolysis of Ni(n) complex 33a 89
Scheme 4-9 Synthesis of dehydroamino acid derivative 36. 91
Scheme 4-10 Synthesis of 8-[2-(A'-phthalimido)-ethyl] indolizidinone amino acid
esters 92
Scheme 4-11 Synthesis of 4,8-disubstituted azabicyclo[4.3.0]alkane amino acid 94
Scheme 4-12 Synthesis of analogues 49-51 96
Scheme 4-13 Synthesis of analogue 53 97
Scheme 4-14 Future work: reduction of double bonds 98
13
List of Tables
Table 3-1 NOE Data for 17b 64
Table 3-2 NOE Data for 20a .66
Table 4-1 NOE Data for 47 96
14
ABSTRACT
A central goal of modem biology is to develop a detailed, predictive understanding
of the relationships of three-dimensional structure and biological function. However, to
establish the biologically active conformation is challenging because most small linear
peptides are inherently flexible, and at present, our knowledge of 3D structural
information of ligand-receptor complexes is very limited. Hence, some strategies have
been developed to prepare peptidomimetics with constrained conformations. Both local
conformational constraints and global conformational constraints can provide important
insights into the structural and topographical basis of biological activity.
A series of novel cw-4-substituted proline analogues were designed and synthesized.
Highly stereoselective alkylations at the y-position of glutamic ester were achieved,
followed by reduction, mesylation. and cyclization to afford the proline derivatives in
good yields and high diastereoselectivity. These c/.v-4-substituted proline analogues
could be used as conformationally restricted templates in local constrained
peptidomimetics.
We also have developed a general and efficient approach for the synthesis of
indolizidinone amino acids with stereospecific appendages of side chain functionality at
both the C-4 and C-8 positions, which can serve as restricted reverse turn mimetics in
global constrained peptidomimetics. Our synthetic reverse turn mimetic targets were
designed to serve as surrogates of the dipeptides Phe-Gly and Phe-Arg which contain two
important pharmacophore elements in Leu-Enkephalin and melanotropin peptides,
15
respectively. Introduction of side chain functionality at C-8 was achieved by using
P-substituted pyroglutamate as a synthetic precursor which was prepared via Michael
addition reaction between a Ni(II) complex of the chiral Schiff base of glycine with
(S)-o-[iV-(W-benzylproly])amino]benzophenone and 3-{trans-enoy\)- oxazolidin-2-one.
The side chains at C-4 were introduced by bromination of dehydroamino acid
intermediates followed by Suzuki cross-coupling.
16
CHAPTER 1. DESIGN OFPEPTIDOMIMETICS FROM PEPTIDE
LEADS-—CONFORMATIONAL AND TOPOGRAPHICAL CONSIDERATIONS
1.1 Introduction
In 1902, Emil Fischer and Frank Hofmeister revealed that proteins are composed of
amino acids, which are linked via '"peptide bonds".' The progress following this
finding was initially slow due to difficulties in synthesis and structure determination of
peptides. However, some great advances came in the 1950's. First of all, duVigneaud
isolated, determined the structure of, and accomplished the total synthesis of oxytoxin;^
then, Sanger elucidated the structure of bovine insulin;^'*' and the double helix of DNA
was discovered, suggesting that there is a connection between nucleotide and peptide
sequences.With discovery of more and more biologically active peptides, today it
has been well known that peptides exert essential influences on all vital physiological
processes via inter- and intracellular communication, and signal transduction mediated
through various classes of receptors.'" As neurotransmitters, neuromodulators,
hormones, antibiotics, growth factors, cytokines, antigens, etc., peptides play critical
roles in the maintenance of human health, in behavior and in many diseases. Therefore,
numerous native peptides and proteins have been isolated and applied as therapeutically
useful drugs.
Although numerous native peptides have great potential for medical applications and
the fact is that over 50% of all current drugs are peptide-based, the peptides have to be
modified to overcome certain problems, such as metabolic stability, selectivity, and
17
bioavailability, to be suitable therapeutic agents. Thus modified peptide and
peptidomimetic research has dramatically advanced during the last two decades.
One of the most challenging parts in the research is the rational design of
peptidomimetics. The first phase in this approach is to identify the key amino acid
residues which are necessary for receptor recognition. This is usually accomplished by
1 n single amino acid modification in the peptide, known as alanine & D-amino acid scans.
Once the SAR of the peptide ligand is elucidated, the conformation-activity relationship
has to be studied. The three-dimensional arrangement of critical side groups and
backbone functionalities can be analyzed via NMR spectroscopy,'^"''' X-ray
20 21 22 crystallography, circular dichroism measurements, and computational methods.
The next effort is to find a suitable organic moiety that can replace the peptide scaffold
and position the crucial recognition elements correctly. The general scheme of the de
novo design of peptidomimetics is outlined in Fig. 1-1. As we can see in the scheme,
understanding the conformation-activity relationships of biologically active peptides can
provide important guidance in the design of peptidomimetics and accelerate the process
from native peptides to biologically active peptidomimetics and small molecules.
18
Random Screening
Optimization
Non-Peptide Ligands
Peptidomimctic Design
Biologically Active Peptides
3D Receptor/Effector Modeling and Pharmacophore Docking
Bioactive Conformations and 3D Pharmacophore Models
Biophysical Studies
-NMR; -X-ray; -MM&MD; -QSAR
Non-Peptide Compounds
-natural products; -synthetic collections; -combinatorial libraries Structure-Activity Relationships
-truncations & deletions; -alanine & D-amino acid scans; -single & multiple substitutions
Linear Pharmacophore Model -AA/groups necessary for binding/single transduction; -targets for modifications
Conformationally Constrained Peptides
-global constraints (cyclization); -local constraints (specialized AA)
Figure 1-1. A de novo approach for peptidomimctic design."^
19
1.2 Conformational and Topographical Features of Peptides — Challenges in the
Studies of Biologically Active Conformation
As a key step in the de novo approach to peptidomimetics, to establish the
biologically active conformation is very challenging. This is partly due to our limited
knowledge of the conformations of ligand-receptor complexes. Another major problem
we have to overcome is the high degree of flexibility of small linear peptides.
gauche(+) trans gauche{-)
Figure 1-2. Conformations of peptides. (A) Definition of the (j), \j/, o), % torsional angles;
(B) Newman projection of the three staggered side-chain rotamers in L-amino acids.
20
As illustrated in Fig. 1-2, each side chain's dihedral angle, referred as chi (%) space,
can adopt three low energy staggered conformations (rotamers), which are referred to as
gauche{+), gauche{-) and trans. Therefore, even in a small peptide with four or five
residues, these different dispositions of side chain groups will give rise to a great
diversity in 3D conformations. Moreover, other torsional angles of the backbone, (j), \\i,
and to, make the topographical issue more complicated. These angles determine the
major secondary structures of peptides, such as a-helix, P-sheet, P-tum, and extended
conformations (Fig. 1-3), which have been shown to be the energetically preferred
backbone conformations for a peptide. 24,25
W I
R
A/ ^ o
H I
•N.
a-Helix 0 = 57° II O
P-Sheet (j) = -139° \!/ = 4-135°
Type I P-Tum V,-h-i = -30°
Type r P-Tum 002 =-90" ¥/+2=0°
/ - 1 i + 1
Figure 1-3. Torsional angles of an a-helix, P-sheet and type I and V P-tum
conformations.
Studies have shown that both the secondary structural features of peptide and the 3D
structure (topography) of the amino acid side chain moieties play critical roles in
ligand-receptor recognition events. Thus identification of the side chain rotomer and the
secondary structural feature that is present in the superpotent or bioselective ligand can
21
provide a valuable tool to aid in the development of peptidomimetics with better
potencies and selectivities. Some strategies to obtain such information will be discussed
next.
1.3 Strategies in the Design of Peptidomimetics
1.3.1. Modification of Amino Acid Side Chains
If conformational flexibility of the side chain groups can be restricted to a greater
degree, peptides can provide a more complete evaluation of their biologically active
three-dimensional topologies. Usually, the side chain conformation can be controlled in
several ways. One general approach is to introduce an alkyl group at the P-position or
on the aromatic ring of an amino acid residue. These kinds of modifications can
constrain x' and angles; on the other hand, they generally do not perturb the backbone
conformation drastically, and still allow the peptides to have some degree of flexibility.
In a similar way, substitution on the aromatic ring of an aromatic amino acid will limit
the conformational flexibility of a peptide to a moderate degree. Furthermore, the
introduction of alkyl groups will enhance the lipophilicity and thus help peptide binding
to receptors and crossing of membrane barriers.
The Hruby group has been engaged in the asymmetric synthesis of x-constrained
amino acids for over a decade. Fig. 1-4 gives structures of some novel amino acids
prepared in the Hruby group. Incorporation of these novel highly constrained amino
acids into peptides and studies of such peptidomimetics have provided a very valuable
approach to probe the stereochemical requirements of binding pharmacophores for
recognition of receptors."^'"'''
22
Among natural amino acids, proline is unique with a constrained cyclic system.
Proline can be used as a rigid template in design of conformationally constrained
peptidomimetics. Substituted proline derivatives are particularly attractive since the
substitution can influence not only the conformation of the pyrrolidine ring, but the rate
of cis-trans isomerization about the amide bond as well. In Chapter 2, a novel approach
to synthesis of 4-substituted prolines will be discussed.
23
COgH
HQC X1CO2H
Kazmierski. W. M.; Urbanczyk-Lipkowski, Z.; Hmby, V. J. J. Org. Chem. 1994,22, 231.
CO2H CH^^ CH
Boteju, L. W.; Wegner, K.; Qing, X.; Hruby, V. J. Tetrahedron 1994. 50,2391.
CO2H
Xiang. L.; \Vu. H.; Hruby. V. J. Tetrahedron 1995, 6, 83.
Qian, X.; Russel, K. C.; Boteju, L. W.; Hruby, V. J. Tetrahedron 1995,51, 1033.
CO2H CO2H
Liao, S.; Hruby, V. J. Tetrahedron Lett. 1996, 37, 1563.
H
NH
CH
Wang, S.; Tang, X.-J.; Hruby, V. J. Tetrahedron Lett. 2000, 41, 1307.
Han, Y.; Liao. S.; Qiu, W.; Cai, C.; Hruby, V. J. Tetrahedron Lett. 1997,58,5135.
R = H, CHg
Qiu, W.; Soloshonok, V. A.; Cai, €.; Hruby, V. J. Tetrahedron 2000, 56, 2511.
Figure 1-4. ^-Constrained amino acids synthesized in the Hruby group.
1.3.2 Modification of the Peptide Backbone
Another strategy in the design of peptidomimetics is peptide backbone modifications
which generally refer to the isosteric or isoelecironic exchange of NHCO units in the
24
peptide chain or introduction of additional groups?'' Some of the most frequent
modifications to the peptide backbone are listed in Fig. 1-5.
Exchange of individual units Extension of peptide chain
—N C —C— H H O
—O— —N— —C— S
—S C-alkyI -C—
H2
R
-NH-X-CH-CO-
X = O, S, CHa
Replacement of amide bond
-CO-NH-
-NH-CO- retro-inverso
-CH(0H)-CH2- hydroxyethylene
-CH=CH- E-alkene
-CH2-CH2- carba
Figure 1-5. The most frequent modifications to the peptide backbone.
The modification to the peptide backbone can also serve to introduce local backbone
constrains. For example, A'-alkylation greatly restricts the (j) torsional angle but
eliminates the hydrogen bonding capability of the amide bond. iV-Methyl amino acids
have been incorporated into bioactive mimetics of opioid peptides;^' bradykinin,^'
thyrotropin releasing hormone (THR), angiotensin II, and cholecystokinin (CCK). '
Other backbone modifications include retro-inverso,^^ reduced amide,thiomethylene,^^'
25
oxomethylene/' e thylene, th ioamide, '^ olef in ic '^^ ' '^ and kctomethylene"*^ analogues
and many others, each of which has its own unique stereoelectronic and stercostructural
features.
1.3.3 Global Restrictions of Conformation in Peptides and Peptidomimetics
Cyclization of a peptide is another general approach to constrain the conformation by
limiting the flexibility of the peptide. In this approach, the amino acid chain groups and
backbone moieties that are not important in biological activity are chosen as the sites to
construct a cyclic structure.'^*' The cyclization can be formed between side chains
through several different types of bonds, such as disulfide,'^"' lactam,"" and thioether (Fig.
1-6). Other kinds of cyclic constrain are also possible between side chain and C- or
A^-terminal or between side chain and backbone nitrogen (Fig. 1-7).
O H
HjC-S S-CHa HgC-C-——N-CH2
—N-C-C- - peptide- -C-C— —N-C-C- -peptide- -C-C— H H O HO H H O HO
Disulfide Lactam
HgC CH2 —N-C-C--peptide--C--C—
H H O HO
Ether/thioether
Figure 1-6. Side-chain-to-side-chain cyclizations.
26
H -C—-c- -N—C C-
O H H O
HpC-
-N-
-{CH2)n
Alkylation
H -C—C-
I O
H -C—c-
I ° C—(CH2)n
-N—-C-H H
-c-O
Acylation
R
-N C-H H
-C-O
-N-H
H -C C-
I o
HN
,(CH2)n HpC-
-N—( I (
-(CH2)n
-C-o
Trans-guanidation
R
Alkylation
-N-H
H -C C N-
I O I -c—-c-H O
-N-H
-C C N C C-I O I H O
H2C S—(CH2)n
Thioether
n(H2C)-
O
Acylation
Figure 1-7. Cyclic constrains.
As illustrated in Figs. 1-6 and 1-7, this approach allows considerable variability in
the design of ring size and ring type. With different sizes and types of cyclic structures,
cyclization not only can limit the flexibility of the conformation, but often can induce or
stabilize the secondary stmcture, such as P-tum. Secondary structures (a-helices,
P-sheets, and p-tums) play critical roles in biological activities.'^' Many novel types of
constraints have been employed to stabilize the secondary structures, especially reverse
turns, in peptides and peptidomimetics.^" This strategy is now universally accepted as a
method to design biologically active peptides and peptidomimetics with high potency and
improved selectivity. A selection of reverse turn mimicking scaffolds is shown in Fig.
1-8.
27
O Ra
Lactam constrains Bicyclic thiazolidincs
O
O
3-Amino- 10(R)-carboxy-1,6-dia2a-cyclodeca-2,7-dione
COO"
8-Aminomethyl-2-napthoic acids
O
4-Hydroxyproline dipeptide derivatives
N N-^O
H N ^
Spiro-bicyclic lactam analogues
Figure 1-8. P- and y-tum mimetics of various types.
As shown in Fig. 1-8, lactam constrains can cause a turn in the peptide backbone,
but it is difficult to place functional groups stereoselectively to match the side chains of
peptide. From this point of view, the bicyclic thiazolidincs are more promising P-turn
mimetics since they can be derived from a chiral pool of amino acids and side groups can
be introduced stereoselectively onto the scaffold during the synthesis. The synthesis of
this type of p-tum mimetic and a novel methodology to introduce functional groups at the
specific position on the backbone will be discussed in Chapters 3 and 4.
28
CHAPTER 2. EFFICIENT AND STEREOSELECTIVE SYNTHESIS OF
NOVEL CJS-4-SUBSTITUTED PROLINE ANALOGUES
2.1 Introduction
Among naturally occurring a-ami no acids, proline is the unique one that is
cyclic and a secondary amine, providing a conformational 1 y constrained system. The
importance of proline is reflected in its presence in many naturally occurring bioactive
peptides such as gramicidin""'''^ and a-melanotropin,^"^ both of which have important
biological activities. The studies of these bioactive peptides have shown that proline
exerts great influence on both the structure and function of peptides and protcins.^^
Peptides and proteins with a proline residue can influence important secondary structures
such as a j3-tum and an a-helix.^^ Due to its special structure, proline has been used as a
rigid template to design conformationally constrained amino acids for the study of the
interactions of peptide and protein ligands with their receptors/acceptors. When it is
incorporated into a peptide or protein, proline can induce a reverse turn which can
provide enhanced bioactivity.^*^'^^ Such effects on peptide conformations have created a
great deal of interest for the design of various substituted proline analogues.^'
Actually, proline and its derivatives have been extensively used in the pharmaceutical
industry, such as in angiotensin-converting enzyme (ACE) inhibitors, including
Captopril,®^ Enalapril,®"^ Fosinopril,''^ and Lisinopril (Fig. 2-1).^'^
29
COgH
Enalapril Captopril
HO. ^,0
O 0 COpH
O CO2H
Fosinopril Lisinoporil
Figure 2-1. Examples of proline-relaled ACE inhibitors.
Among these proline analogues, 4-substitued derivatives arc particularly attractive
since the C4 substituents can influence not only the conformation of the pyrrolidine ring,
but the rate of cis-trans isomerization about the amide bond as well.®*^'®^ The readily
available starting material 4-trans-hydroxyproline has been used as a versatile building
block for many biologically important compounds.In our ongoing melanocyte
stimulating hormone (MSH) project, the substitution of histidine with proline in MT-II
70 has generated a potent and selective analogue with agonist activity at the human MC5R.
To further explore the structure-activity relationship (SAR) of this ligand and its receptors,
we have designed novel 4-substituted prolines.
30
2.2 Background and Designed General Approach to cw-4-Siibstitiited Proline
Derivatives
Several methods for the synthesis of 4-substitued prolines have appeared in the
literature/'" ''^'"' Recently our and Goodman's group have reported the synthesis of cis-
and trans- 4-substituted proline analogues through hydrogenations of pyrroline
intermediates derived from 4-fran.v-hydroxyproline (Scheme 2-1
Scheme 2-1. Strategies for the asymmetric hydrogenation of pyrroline
Intermediates.
RiQ,
trans isomer
cis isomer
A: Hydroxy I directed hydrogenation B; Sterically directed hydrogenation
Meanwhile we have developed asymmetric hydrogenations/Suzuki-couplings for
the preparation of a number of novel x" constrained amino acids7^"^^ As an alternative
to these methods, herein we would like to disclose an efficient and stereoselective
approach to the synthesis of c/.v-4-subsititued prolines. The synthetic strategy for the
preparation of the title compounds 1 employs stereoselective alkylation at the y-position
31
of glutamic ester 2, followed by selective reduction, tosylation, and cyclization to obtain
c/.s-4-substituted prolines 1 (Scheme 2-2).
Scheme 2-2. Strategy for the preparation of cw-4-substituted proline derivatives 1.
MeO
O alkylation
OfBu ^
NHBoc
O O R
OfBu
R NHBoc N Boo
1
COOfBu
2.3 Stereoselective Synthesis of cii-4-Substituted Proline Derivatives
Scheme 2-3." Approach to czs-4-substituted proline derivatives.
O
HO OfBu NHBoc
MeO OfBu
NHBoc
O O
MeO
R
OfBu
NHBoc
3a: R = Allyl, 81% 3b: R = Benzyl, 81% 3c: R = Cinnamyl, 77% 3d: R = 4-Bromobenzyl, 72%
O
TsO
R
OfBu
NHBoc
5a; R = Allyl, 88% 5b: R = Benzyl, 85% 5c: R = Cinnamyl, 90% 5d; R - 4-Bromobenzyl, 92%
O
HO OfBu
NHBoc
4a: R = Allyl, 60% 4b: R = Benzyl, 66% 4c: R = Cinnamyl, 68% 4d: R = 4-Bromobenzyl, 74%
COOfBu N Boc
1a lb 1c Id
R = Allyl, 93% R = Benzyl, 88% R = Cinnamyl, 90% R = 4-Bromobenzyi, 85%
^Conditions; (a) MeOH, DCC, 5 hr, 76%; (b) LiHMDS, THF, -78 °C, 30 min, then RBr, 5-6 hr;
(c) NaBH4, MeOH, 16 hr; (d) TsCI, DMAP, Pyridine, 12 hr; (e) NaH, THF, r.t., 3-4 hr.
32
The synthesis of cw-4-substituted proHne analogues 1 started with commercially
available (45)-5-(ferf-butoxy)-4-[(?erf-butoxycarbonyl)amino]-5-oxopentanoic acid
(Scheme 2-3). The carboxylic acid was protected as a r-butyl ester, which can be
viewed as an orthogonal protection to the w-methyl ester in 2. Thus, the methyl ester
could be selectively reduced without affecting the r-butyl ester. The free O) carboxyl
functional group was converted into a methyl ester 2 using dicyclohexylcarbodiimide
(DCC) as an activating agent with methanol in the presence of a catalytic amount of
dimethylaminopyridine (DMAP) and triethylamine (TEA) in 76% yield.^^ The resulting
compound 2 was used for the alkylations. Alkylation at the C4 position of glutamates
has been reported in the literature.The studies indicated that the stereoselectivity
depended on the nature of the A^-substituents^'^ and the esters.Recently Hancssian
and co-workers achieved highly stereoselective alkylations with A''^-Boc or Cbz and the
methyl ester.^^'^® The stereoselectivity was attributed to a highly coordinated dianionic
chair-like transition state (Fig. 2-2).
Li /
Li O
Figure 2-2. Transition states in alkylations.
However, substrates with the bigger r-butyl ester, as in our case, had not been studied.
We rationalized that the size of the ester groups would play an important role in
33
controlling the stereoselectivity of the alkylations based on the proposed transition state.
The large r-butyl ester would further enhance the diastereoselectivity of the alkylations by
stabilization of the chair-like transition state. As expected, only one single diastereomer
{ami product) was obtained based upon 'H NMR analysis (Scheme 2-3).
Scheme 2-4." Selective reduction of y-substituted glutamic acid ester.
O O 0 0 0 0
R NHBoc R N(BOC)2 R N(BOC)2
3 6 ^
^Conditions; (a) (Boc)20, DMAP; (b) DIBAL, Ether.
With the optically pure y-substituted alkyl glutamic acid ester 3 in hand, initially
we planned to selectively reduce the methyl ester by diisobutylaluminumhydride (DIBAL)
at -78 "C to an aldehyde, which subsequently could undergo reductive amination to give
the final products 1. Based on our earlier study, the mono-A^^-Boc protected nitrogen
interfered with the reduction.^® Therefore, a second Boc protecting group was
introduced by reaction of 3 with di-r-butyl di carbon ate [(Boc)20] in the presence of a
catalytic amount of DMAP in acetonitrile after chromatography to give the bis-Boc
protected methyl ester 6 in over 80% yield (Scheme 2-4). However, the reduction of the
methyl ester in 6 using DIBAL at -78 "C did not give the desired aldehyde 7. Even
higher reaction temperatures (room temperature) did not work. In all cases, only the
starting materials were recovered. In contrast, our previous study with reduction of a
34
similar substrate without y-substituents under the same reaction conditions gave an
aldehyde in excellent yieldJ*^ The presumed reason was steric hindrance in the
y-substituted substrates 6.
Consequently, we modified our synthetic strategy (Scheme 2-3). We proposed the
conversion of the methyl ester 3 to an alcohol, which then was transformed into a good
leaving group for cyclization. The intramolecular nuclear substitution (cyclization)
would provide the target molecules. Reduction of the mono-Boc protected methyl
esters 3 with NaBRt to give alcohols 4 was achieved in good yields. Then the alcohols
were converted into tosylates 5 in high yields. The tosylated intermediates were treated
with NaH to give the cyclic proline derivati ves 1 in good yields. To test for
racemization during these conversions, we selectively deprotected the Boc group in la
(due to rotamers) to 8, which gave a "clean" NMR (Scheme 2-5). One isomer was
observed by NMR indicating that no isomerization occurred.
Scheme 2-5. Deprotection of c/.v-4-subslituded proline derivatives.
TFA, CH2CI2, r.t., 30 min COO®u COOffiu
H Boc
1a 8
This approach also provides important synthetic intermediates to other amino acid
derivatives. With compound 3a as an example, the pyroglutamate ester 9, which can be
35
used as starting material in our dipeptide |3-tum mimetic synthesis,was obtained in 3
steps (Scheme 2-6).
Scheme 2-6. Cyclization to pyroglutamic acid ester derivative 9.
MeO
O
1 NHBoc
O 1)TFA 2) TEA, toluene, reflux
•0©u 3) (Boc)20 [sj COOfBu
Boc
3a 9
2.4 Conclusion
In conclusion, a series of novel cw-4-substituted proline derivatives 1 were
efficiently synthesized from readily available starting materials. Highly stereoselective
alkylations at the y-position of the glutamic ester 2 were achieved. The resulting
alkylation compounds were transformed to the final products 1 through
reduction/tosy 1 ation/cyc 1 ization in high yields. Future work will aim at the
incorporation of these unnatural amino acids into a-MSH peptides and the study of
structure-activity relationships of the a-MSH peptides.
2.5 Experimental Section
General. ^H and NMR were performed on DRX-500 spectrometers using TMS
and CDCI3 as internal standards. High Resolution Mass Spectra (HRMS) were recorded
36
on a JOE HXllOA instrument in the University of Arizona Mass Spectrum Laboratory.
Optical rotations were measured on a JASCO-1020 polarimeter. Commercially
available starling materials and reagents were purchased from Aldrich and used as
received. THF was distilled from Na and benzophenone.
(S)-(-)-l-te/t-Butyl-5-methyl-[(2-^^rt-butoxycarbonyl)amlno]pentanedioate (2).
To a solution of (45)-5-(terf-butoxy)-4-[(terr-butoxycarbonyl)amino]-5-oxopentanoic
acid (10.0 g, 33 mmol) in 50 mL of CH2CI2 was added DCC (8.9 g, 43.1 mmol) at 0 "C
under argon. After 5 min, MeOH (2.7 mL, 26.6 mmol), TEA (6 mL, 43 mmol) and
DMAP (400 mg, 3.3 mmol) were added to the above mixture. After stirring 1.5 h at 0
"C and 5 h at rt, the solution was filtered and concentrated under reduced pressure. The
residue was redissolved in 500 mL of EtOAc, the organic solution was washed with IN
HCl (100 mL), saturated NallCOi (100 mL) and brine (100 mL), dried over MgS04 and
concentrated to give crude product as an oil. The crude product was purified by flash
column chromatography (hexanes/EtOAc : 6:1) to afford 2 (7.9 g, 76%) as a colorless oil.
'H NMR (500 MHz, CDCI3) § 1.44 (s, 9H), 1.47 (s, 9H), 1.85-1.98 (m, IH), 2.14-2.18 (m,
IH), 2.23-2.49 (m, 2H), 4.20 (m, IH), 5.08 (d, /= 8.1 Hz, IH).
Procedure A: To a solution of compound 2 in dry THF (6 mL/mmol) was added
LiHMDS (2.2 equiv.) at -78 "C under N2. After 30 min stirring, bromide (3 equiv.) was
added at -78 "C. The reaction solution was kept at this temperature for 6 h. The
reaction was quenched by H2O (2 mL) at -78 "C. The organic solution was washed with
NH4CI solution (10 mL/mmol) and brine (10 mL/mmol), dried over MgS04 and
evaporated. The crude product was purified by flash column chromatography
37
(hexanes/EtOAc ; 4:1) to give a colorless oil.
Procedure B: To a solution of compound 3 in methanol (10 mL/mmol) was added
sodium borohydride (10 equiv.) at rt. The mixture was stirred for 12 h at rt. The
solution was washed with NH4CI solution (10 mL/mmol) and brine (10 mL/mmol), dried
over MgS04 and evaporated. The crude product was purified by flash column
chromatography (hexanes/EtOAc : 2:1) to give a colorless oil.
Procedure C: To a solution of compound 4 in pyridine (10 mL/mmol) was added
toluenesulfonyl chloride (4 eq) and DMAP (0.1 eq). The reaction mixture was stirred at
room temperature for 12 h and diluted with EtOAc (30 mL/mmol). The organic
solution was washed with IN HCl (30 mL/mmol, 3x), NaHCOs (30 mL/mmol) and brine
(30 mL/mmol), dried over MgS04 and evaporated. The crude product was purified by
flash column chromatography (hcxanes/EtOAc : 4:1) to give a colorless oil.
Procedure D: To a solution of compound 5 in THF (10 mL/mmol) was added
sodium hydride (1.1 equiv.) at rt under N2. After 4 h stirring, the reaction was quenched
with NH4CI solution (5 mL/mmol). The organic solution was washed with NH4CI (10
mL/mmol) and brine (10 mL/mmol), dried over MgS04 and evaporated. The crude
product was purified by flash column chromatography (hexanes/EtOAc : 5:1) to give a
colorless oil.
(2S,45)-(+)-l-tert-Butyl-4-allyI-5-methyl-[(2-fe/f"butoxycarbonyl)aiiiino] pent-
anedioate (3a). Procedure A. 81% yield, [a]"^D +14.4 (c 3.66. CHCI3); 'H NMR (500
MHz, CDCI3) 5 5.70-5.61 (IH, m), 5.04-4.98 (2H, m), 4.88 (IH, d, /= 8.5 Hz), 4.16-4.15
(IH, m), 3.62-3.61 (3H, m), 2.56-2.50 (IH, m), 2.32-2.29 (2H, m), 1.90-1.84 (2H, m).
38
1.41 (9H, s), 1.39 (9H, s); '^C NMR (125 MHz, CDCl?) 8 175.7, 171.7, 155.6, 134.7,
117.7, 82.2, 79.9, 52.8, 51.8, 42.1, 36.5, 34.2, 28.5, 28.1; HRMS (FAB) calcd for
CjgHsaNOe 358.2230, found 358.2230.
(2S,4S)-(-)-l-tert-Butyl-4-alIyl-5-hydroxyI-[(2-terf-biitoxycarbonyI)amino] pent-
anoate (4a). Procedure B. 60% yield, [a]^^D -10.6 (c 1.74, CHCI3); 'H NMR (500
MHz, CDCI3) 5 5.78-5.70 (IE, m), 5.19-5.17 (IH, d, / = 6.2 Hz), 5.06-5.00 (2H, m),
4.19-4.18 (IE, m), 3.71-3.69 (IE, m), 3.53-3.49 (IH, m), 2.46 (IE, s), 2.13-2.10 (2H, m),
1.88-1.68 (2B, m), 1.65-1.59 (IE, m), 1.44 (9E, s), 1.43 (9H, s); NMR (125 MEz,
CDCI3) 5 172.3, 155.8, 136.6, 117.0, 82.2, 80.1, 65.2, 52.3, 37.6, 36.1, 35.2, 28.5, 28.2;
ERMS (FAB) calcd for C17H32NO5 330.2280, found 330.2287.
(2S,4S)-(+)-l-ter^-Butyi-4-allyl-5-(toIuene-4-sulfonyIoxy)-[(2-tert-butoxycarbonyl
)amino] pentanoate (5a). Procedure C. 88% yield, [a]-^) +5.3 (c 0.97, CECI3); 'E
NMR (500 MHz, CDCI3) 5 7.78 (2E, d, J = 8.3 Hz), 7.34 (2H, d, J = 8.3 Hz), 5.64-5.56
(IE, m), 5.01-4.96 (2E, m), 4.91 (IE, d, / = 8.4 Ez), 4.16-4.15 (IE, m), 3.97-3.90 (2E,
in), 2.45 (3E, s), 2.23-2.20 (IE, m), 2.12-2.07 (IE, m), 1.93-1.86 (IE, m), 1.74-1.68 (IE,
m), 1.54-1.47 (IE, m), 1.44 (9E, s), 1.43 (9E, s); "C NMR (125 MEz, CDCI3) 8 171.9,
155.6, 145.0, 134.6, 133.0, 130.1, 128.1, 118.2, 82.3, 80.0, 72.3, 52.1, 34.6, 34.3, 34.1,
28.5, 28.1, 21.8; ERMS (FAB) calcd for C24H38NO7S 484.2369, found 484.2372.
(S).(»)-4.Allyl-Boc-L-proline tert-\mtj\ ester (la). Procedure D. 93% yield,
[a]^^D -69.3 (c 1.56, CECI3); ^H NMR (rotamers) (500 MEz, CDCI3) 8 5.78-5.73 (IH,
m), 5.07-5.01 (2E, m), 4.10(0.7H, t, J= 8.0 Ez) (5 4.15, 0.3E), 3.78 (0.7E, dd, 7, = 10.5
Ez, J2 = 6.5 Ez) (8 3.65, 0.3H), 3.06-3.00 (IE, m), 2.45-2.38(lE, m), 2.23-2.13 (3E, m).
39
1.60-1.52 (IH, m), 1.49 (6H, s), 1.47 (6H, s), 1.45 (6H, s); '"C NMR (125 MHz, CDCI3)
5 172.5, 172.4, 154.3, 154.0, 136.3,136.2, 116.5,116.4, 81.0, 79.9, 79.7, 60.0, 59.9, 52.2,
51.9, 38.2, 37.4, 37.3, 37.2, 36.8, 35.8, 28.6, 28.5, 28.2, 28.1; HRMS (FAB) calcd for
C17H30NO4 312.2175, found 312.2164.
(2S,4S)-(-)-l-tert-Butyl-4-beiizyI-5-methyl-[(2-terf"butoxycarbonyl)amiiio] pent-
anedioate (3b). Procedure A. 81% yield, [a]^^D -3.5 (c 1.93, CHCI3); 'H NMR (500
MHz, CDCI3) 5 7.27-7.24 (2H, m), 7.20-7.14 (3H, m), 4.94 (IH, d, / = 9.0 Hz), 4.28-4.24
(IH, m), 3.57 (3H, s), 2.91-2.89 (2H, m), 2.81-2.75 (IH, m), 1.96-1.91 (2H, m), 1.44
(18H, s); NMR (125 MHz, CDCI3) 5 175.7, 171.6, 155.6, 138.7, 129.1, 128.5, 126.7,
82.2, 79.9, 52.7, 51.8, 44.6, 38.3, 34.8, 28.5, 28.1; HRMS (FAB) calcd for C22H34NO0
408.2386, found 408.2387
(2S,4S)-(+)-l-tert-ButyI-4-benzy!-5-hydroxyl-[(2-fert-butoxycarbonyl)amino]-
pentanoate (4b). Procedure B. 66% yield, la]\ +34.7 (c 1.28, CHCI3); 'H NMR
(500 MHz, CDCI3) 8 7.28-7.25 (2H, m), 7.19-7.17 (3H, m), 5.24 (IH, d, / = 6.8 Hz),
4.26-4.24 (IH, m), 3.76-3.74 (IH, m), 3.51-3.49 (IH, m), 2.72-2.64 (2H, m), 2.59 (IH, s),
2.01-1.96 (IH, m), 1.74-1.70 (2H, m), 1.45 (9H, s), 1.41 (9H, s); ''C NMR (125 MHz,
CDCI3) 5 172.2, 155.8, 140.4, 129.5, 128.5, 126.2, 82.3, 80.2, 64.3, 52.0, 39.9, 37.8, 35.4,
28.5, 28.2; HRMS (FAB) calcd for C21H34NO5 380.2437, found 380.2438.
(2S,45')-(+)-l-?e/t-Butyl-4-benzyl-5-(toluene-4-sulfonyloxy)-[(2-^ert-butoxy-carb-
onyl)-amino] pentanoate (5b). Procedure C. 85% yield, +1.3 (c 1.72, CHCI3);
'H NMR (500 MHz, CDCI3) 8 7.76-7.74 (2H, m), 7.32 (2H, d, / = 8.1 Hz), 7.19-7.14 (3H,
m), 7.03-7.02 (2H, m), 4.94-4.92 (IH. d, i = 8.5 Hz), 4.29-4.25 (IH, m), 3.88- 3.82 (2H,
40
m), (IH, dd, Jy = 13.5 Hz, Jj = 4.0 Hz), 2.57-2.52 (IH, m), 2.45 (3H, s), 2.07-2.03 (IH,
m), 1.78-1.73 (IH, m), 1.59-1.50 (IH, m), 1.45 (9H, s), 1.44 (9H, s); NMR (125 MHz,
CDCI3) 6 172.0, 155.7, 145.0, 138.8, 133.0, 130.1, 129.5, 128.6, 128.2, 126.5, 82.4, 80.1,
72.0, 52.0, 37.0, 35.8, 34.9, 28.5, 28.2, 21.9; HRMS (FAB) calcd for C28H40NO7S
534.2525, found 534.2540.
(S)-(-)-4-Benzyl-Boc-L-proIine tert-hutjl ester (lb). Procedure D. 88% yield,
-67.0 (c 1.65, CHCI3); 'H NMR (rolamers) (500 MHz, CDCI3) 5 7.31-7.25 (2H,
m), 7.23-7.13 (3H, m), 4.07 (0.7H, t, J= 8.0 Hz) (5 4.12, 0.3H), 3.71 (0.7H, dd, = 10.5
Hz, J2 = 7.5 Hz) (5 3.59, 0.3H), 3.13-3.08 (IH, m), 2.75-2.64 (2H, m), 2.45-2.29 (2H, m),
1.67-1.62 (IH, m), 1.47 (6 H, s) (5 1.46, 3 H), 1.42 (6H, s) (5 1.44, 3H); NMR (125
MHz, CDCI3) 8 172.5, 172.4, 154.3, 154.0, 140.3, 140.2, 128.8, 128.7, 128.6, 126.5, 81.1,
80.0, 79.8, 60.0, 52.3, 52.1, 40.5, 39.8. 39.3, 39.2, 37.0, 36.0, 28.6, 28.5, 28.3, 28.2;
HRMS (FAB) calcd for C21H32NO4 362.2331, found 362.2337.
(2SAS')-(-)-l-/ert-Butyl-4-(3-phenyIallyl)-5-methyl-[(2-f^rr-butoxycarbonyl)-
amino] pentanedioate (3c). Procedure A. 77% yield, [a]^^D -10.4 (c 2.35, CHCI3);
'H NMR (500 MHz, CDCI3) 8 7.34-7.26 (4H, m), 7.21-7.18 (IH, m), 6.43 (IH, d, J =
15.5 Hz), 6.13-6.07 (IH, m), 4.97 (IH, d, .7=8.8 Hz), 4.26 (IH, dd, J; = 14.5 Hz, .h = 8.7
Hz), 3.66 (3H, s), 2.69-2.63 (IH, m), 2.52 (2H, t, J = 6.9 Hz), 2.00-1.95 (2H, m), 1.44
(9H, s), 1.43 (9H, s); NMR (125 MHz, CDCI3) 8 175.7, 171.7, 155.6, 137.4, 132.8,
128.7, 128.6, 127.4, 126.4, 126.3, 82.2, 79.9, 52.7, 51.9, 42.4, 35.6, 34.4, 28.5, 28.1. 28.0,
27.9; HRMS (FAB) calcd for C24H36NO6 434.2543, found 434.2546.
(2S,4S)-(+)-l-fer?-Butyl-4-(3-ptienylallyI)-5-hydroxyI-[(2-terf-butoxycarboiiyl)-
41
aminojpentanoate (4c). Procedure B. 68% yield, [a]^^D +11-1 (c 1.53, CHCI3);
NMR (500 MHz, CDCI3) 6 7.34-7.24 (4H, m), 7.21-7.16 (IH, m), 6.43 (IH, d, i = 15.8
Hz), 6.21-6.15 (IH, m), 5.22 (IH, d, J = 7.5 Hz), 4.26-4.24 (IH, m), 3.78-3.76 (IH, m),
3.60-3.56 (IH, m), 2.52 (IH, br s), 2.30 (2H, t, J= 6.8 Hz), 1.91-1.84 (IH, m), 1.81-1.75
(IH, m), 1.72-1.67 (IH, m), 1.44 (9H, s), 1.43 (9H, s); NMR (125 MHz, CDCI3) 5
172.3, 155.8, 137.7, 132.3, 128.7, 128.2, 127.2, 126.2, 82.2, 80.1, 65.0, 52.3, 38.1, 35.3,
35.0, 28.5, 28.2; HRMS (FAB) calcd for C23H36NO5 406.2593, found 406.2582.
(2S,4S)-(-)-l-tert-Biityl-4-(3-phenylallyl)-5-(toliiene-4-sulfonyloxy)-[(2-terr-but-
oxycarbonyl)amino] pentanoate (5c). Procedure C. 90% yield, [aj"^) -7.0 (c 1.27,
CHCI3); 'H NMR (500 MHz, CDCI3) 6 7.77 (2H, d, / = 8.5 Hz), 7.30-7.19 (7H, m),
6.31 (IH, d, J= 16.0 Hz), 5.97-5.91 (IH, m), 4.92 (IH, d, i= 8.5 Hz), 4.24-4.20 (IH, m),
3.96 (2H, d, J = 5.3 Hz), 2.40 (3H, s), 2.27-2.21 (IH, m), 2.01-1.94 (IH, m), 1.80-1.74
(IH, m), 1.58-1.54 (2H, m), 1.44 (9H, s), 1.43(9H, s); NMR (125 MHz, CDCI3) 5
172.0, 155.7, 145.1, 137.4, 133.3, 133.0, 130.1, 128.7, 128.2, 127.4, 126.3, 126.2, 82.4,
80.1, 72.3, 52.1, 35.2, 34.4, 33.2, 28.5, 28.2, 21.8; HRMS (FAB) calcd for C30H42NO7S
560.2682, found 560.2686.
(S)-(-)-4-(3-Phenyl-allyl)-L-proline tert-hutyl ester (Ic). Procedure D. 90% yield,
[a]^^D -53.6 (c 1.80, CHCI3); ^H NMR (rotamers) (500 MHz, CDCI3) 8 7.35-7.26 (4H,
m), 7.23-7.19 (IH, m), 6.43-6.39 (IH, m), 6.16-6.10 (IH, m), 4.10 (0.7H, t, / = 8.0 Hz)
(5 4.16, 0.3H), 3.80 (0.7H, dd, 7, = 10.5 Hz, .h = 6.5 Hz) (5 3.67, 0.3H), 3.12-3.05 (IH,
m), 2.48-2.41 (IH, m), 2.35-2.24 (3H, m). 1.64-1.60(1H, m), 1.47 (6H, s) (5 1.46, 3H),
1.44 (6H, s) (5 1.45, 3H); '^C NMR (125 MHz, CDCI3) 5 172.5, 172.4, 154.3, 154.0,
42
137.5, 131.8,131.7, 128.7, 128.6, 128.0, 127.9, 127.4, 127.3, 126.2, 81.1, 80.0, 79.8, 60.0,
59.9. 52.2, 52.0, 38.7, 37.9, 36.8, 36.6, 36.5, 35.9, 28.6, 28.5, 28.2, 28.1; HRMS (FAB)
calcd for C23H34NO4 388.2488, found 388.2488.
(2S,4S)-(-)-l-tert-Butyl-4-(4-bromobeiizyl)-5-methyl-[(2-ferf"butoxycarbonyl)- ,
amino] pentanedioate (3d). Procedure A. 72% yield, [a]^^D -12.6 (c 1.79, CHCI3);
'H NMR (500 MHz, CDCI3) 6 7.38 (2H, d, / = 8.2 Hz), 7.04 (2H, d, / = 8.2 Hz), 4.95
(IE, d, J = 8.8 Hz), 4.28-4.27 (IH, m), 3.57 (3H, s), 2.89-2.83 (2H, m), 2.76-2.73 (IH,
m), 1.96-1.90 (2H, m), 1.45 (9H, s), 1.44 (9H, s); "C NMR (125 MHz, CDCI3) 5 175.4,
171.6, 155.7, 137.8, 131.7, 131.0, 120.6, 82.5, 80.1, 52.5, 51.9, 44.4, 37.4, 35.1, 28.5.
28.2; HRMS (FAB) calcd for C22H32BrN06Cs 618.0467, found 618.0445.
(25,4S)-(+)-l-^ert-ButyI-4-(4-bromo-benzyl)-5-hydroxyI-[(2-ter?-butoxycarbonyI)
amino] pentanoate (4d). Procedure B. 74% yield, [a]^^D +24.4 (c 3.51, CHCI3); 'H
NMR (500 MHz, CDCI3) 5 7.38 (2H, d,./= 8.3 Hz), 7.06 (2H, d, / = 8.3 Hz), 5.28 (IH, d,
/ = 7.4 Hz), 4.27-4.23 (IH, m), 3.40-3.68 (IH, m), 3.48-3.43 (IH, m), 2.84 (IH, br s),
2.70-2.59 (2H, m), 1.92 (IH, br s), 1.72-1.67 (2H, m), 1.44 (9H, s), 1.42 (9H, s); '^C
NMR (125 MHz, CDCis) 6 172.1, 155.8, 139.4, 131.5, 131.2, 120.0, 82.3, 80.2, 63.7,
51.9, 39.6, 36.8, 35.2, 28.5, 28.1; HRMS (FAB) calcd for C2iH33BrN05458.1542, found
458.1526.
(2S,4S')-(-)-l-tert-Butyi-4-(4-bromobenz:yl)-5-(toluene-4-siilfonyIoxy)-[(2-tert-
butoxycaronyl)amino] pentanoate (5d). Procedure C. 92% yield, [a]"\) -5.13 (c J .43,
CHCI3); 'H NMR (500 MHz, CDCI3) 5 7.73 (2H, d, J = 8.3 Hz), 7.32 (2H, d, J = 8.2
Hz), 7.28 (2H, d, J = 8.3 Hz), 6.91 (2H, d, 7 = 8.2 Hz), 4.99 (IH, d, / = 8.5 Hz),
43
4.30-4.26 (IH, m). 3.83-3.77 (2H, m), 2.86 (IH, dd, h = 13.5 Hz, J2 = 3.5 Hz), 2.47 (3H,
s), 2.01-1.98 (IH, m), 1.79-1.73 (IH, m), 1.57-1.50 (IH, m), 1.45 (18H, s); '^C NMR
(125 MHz, CDCI3) 5 171.8, 155.8, 145.1, 137.9, 132.8, 131.6, 131.1, 130.1, 128.1, 120.3,
82.5, 80.1, 71.7, 51.9, 36.9, 35.3, 35.0, 28.5, 28.1, 21.8; HRMS (FAB) calcd for
CigHsgBrNOvS 612.1631, found 612.1653.
(5)-(-)-4-(4-Bromobenzyl)-Boc-L-proIine fert-butyl ester (Id). Procedure D.
85% yield, [a]^^D -4.17 (c 1.76, CHCI3); ^H NMR (rotamers) (500 MHz, CDCI3) 5
7.44-7.40 (2H, m), 7.06-7.02 (2H, m), 4.09 (0.7H, t, / = 8.5 Hz) (5 4.14, 0.3H), 3.71
(0.7H, dd, ii = 10.5 Hz, J2 = 7.5 Hz) (8 3.59, 0.3H), 3.14-3.08 (IH, m), 2.71-2.63 (2H,
m), 2.42-2.32 (2H, m), 1.64-1.57 (IH, m), 1.49 (6H, s) (5 1.48, 3H), 1.44 (6H, s) (5 1.46,
3H) ; "C NMR (125 MHz, CDCI3) 5 172.4, 172.3, 154.2, 154.0, 139.2, 139.1, 131.8,
130.5, 120.3, 81.2, 80.1, 79.8, 59.9, 59.8, 52.2, 52.0, 40.2, 39.5, 38.7, 38.5, 36.8, 35.8,
28.6, 28.5, 28.2, 28.1; HRMS (FAB) calcd for C2iH3iBrN04440.1436, found 440.1436.
(5)-(-)-4-Allyl-L-prolme fe/f-butyl ester (8). A solution of la (20 mg, 0.064 mmol)
in 1.6 mL CH2CI2 and 0.4 mL TFA was stirred at rt under argon for 30 min. The
solution was diluted by CH2CI2 (2 mL) and the organic layer was washed with NaHCOs
(3 mL), dried over MgS04 and concentrated to give a lightly yellow oil with quantitative
yield. The crude product was pure enough for characterization, [a] d -29.0 (c 0.3,
CHCI3); ^H NMR (rotamers) (500 MHz, CDCI3) 5 5.78-5.69 (IH, m), 5.02-4.95 (2H,
m), 3.66 (IH, t, / = 7.8 Hz), 3.03 (IH, dd, 7, = 10.0 Hz, J2 = 6.0 Hz), 2.68 (IH, dd, Jy =
10.0 Hz, /2= 7.5 Hz), 2.45 (IH, br s), 2.29-2.26 (IH, m), 2.22-2.15 (IH, s), 2.14-2.02
(2H, m), 1.44 (9H, s), 1.43-1.39 (IH, m); NMR (125 MHz, CDCI3) 5 174.8, 137.2,
44
116.0, 81.4, 60.6, 52.6, 39.4, 38.1, 36.7, 28.3; HRMS (FAB) calcd for C12H22NO2
212.1651, found 212.1653.
(5)-4-AlIyI-5-oxo-Boc-L-proline fert-butyl ester (9). A solution of 3a (205 mg,
0.58 mmol) in 4 mL of CH2CI2 and 1 rtiL of TFA was stirred at room temperature for 30
min. The solution was diluted with CH2CI2 (5 mL). The organic layer was washed
with NaHCOs (2 x 10 mL), dried over MgS04, and concentrated to give a light yellow oil.
The crude product was redissolved in 5 mL of toluene with TEA (160 [ih, 1.14 mmol).
The solution was heated up to 90 "C for 12 h. After the oil bath was removed, the solvent
was removed under reduced pressure. To the solution of the above crude product in 5
mL of acetonitrile was added (Boc)20 (150 mg, 0.69 mmol) and DMA? (7.5 mg, 0.06
mmol). After stirring at rt for 3 h, the solvent was removed under reduced pressure. The
residue was redissolved in CH2CI2 (10 mL). The organic layer was washed with brine
(10 mL), dried over MgS04, and concentrated under reduced pressure. The crude
product was purified by flash column chromatography (hexanes/EtOAc : 3:1) to afford 9
(117 mg, 63%) as a colorless oil. NMR (500 MHz, CDCI3) 5 5.72-5.65 (IH, m),
5.04-5.00 (2H, m), 4.35 (IH, dd, h = 9.5 Hz, J2 =5.5 Hz), 2.61-2.54 (2H, m), 2.43-2.36
(IH, m), 2.21-2.14 (IH, m), 1.69-1.64 (IH. m), 1.46 (9H, s), 1.44 (9H, s); '-'C NMR (125
MHz, CDCI3) 6 174.9, 170.7, 149.6, 135.0, 117.7, 83.5, 82.3, 58.2, 42.3, 35.5, 28.1, 28.0,
26.5; HRMS (FAB) calcd for CiyHzgNOs 326.1967, found 326.1965.
45
CHAPTER 3. DESIGN AND SYNTHESIS OF CONFORMATIONALLY
CONSTRAINED REVERSE-TURN PEPTIDOMIMETICS OF
LEU-ENKEPHALIN
3.1 Introduction
In 1975, Hughes and Kosterlitz discovered the opioid pentapetides
Met(5)-enkcphalin (H-Tyr-Gly-Gly-Phe-Met-OH) and Leu(5)-enkephalin
(H-Tyr-GIy-Gly-Phe-Leu- Since then, the biology and chemistry of these
endogenous opioid peptides have been extensively studied. Some research results have
shown that these neurotransmitters regulate sensory functions including pain and control
of respiration in the central nervous system by binding to the G-protein coupled |i- and
6-opioid receptors, respectively.^^'^^ Among these studies, the structure-activity
relationships and the conformation of enkephalin in solution have been specially
interesting, because enkephalin is a noncyclic peptide binding to the same receptor site as
rigid non-peptide opiate agonists. The conformation of enkephalins has been
87 88 investigated by using a variety of methods, including NMR, X-ray diffraction,
conformational energy calculations,^^ CD,'**' and Raman.'"
However, studies that focus on elucidating the bioactive conformation of opioid
peptide ligands have been complicated by the fact that there are four main opioid
receptors (|i, 5, K, and a), some of which may also have distinct sub-types.*'^'^'^
Although some important pharmacophoric elements have been identified, such as the
relative location and orientation of the aromatic side chains and the relationship of the
46
N-terminal nitrogen to the phenolic oxygen,the large conformational freedom of
enkephalins leads to their nonspecific receptor affinity and thus makes the determination
of the bioactive conformation very challenging. On the other hand, the structure
determination of the membrane bound opioid receptors is extremely difficult. As
discussed in Chapter 1, a general and efficient strategy to study the bioactive
conformation of short linear peptides, like enkephalins, is to synthesize the
conformalionally restricted mimetics and test them in order to obtain some insight into
the SAR.
A successful example of this approach is c[DPen^,DPen"^Jenkephalin (DPDPE), a
cyclic conformalionally and topographically constrained analogue of enkephalin (Fig.
9 ^ c 3-1). In this analogue the Gly and Met (or Leu ) residues were replaced by
D-penicillamine (D-Pen, (3,P-dimethylcysteine) and formed a 14-membered disulfide
ring.^'' DPDPE was found to be highly potent and selective for the 5 opioid receptor and
to be completely stable to proteolytic breakdown in vitro and in vivo.'^^ ''^ Extensive
biophysical studies on DPDPE and some of its critical analogues using
molecular dynamics'^' and X-ray crystallography'^^'"'®" have clearly established the
preferred conformation of the 14-membered ring template of this cyclic analogue.
47
HpN
O NH
HOOC H
DPDPE
Figure 3-1. Structure of DPDPE.
Although considerable success has been obtained in designing non-peptide
peptidomimetics of enkephalins by incorporating some aspects of the overall structural
and topographical features into the peptides, like DPDPE, efforts to design in other
important features such as the proper topographical relationships of critical side chain
groups in ({), \|/ and % space, and of certain H-bond donating and accepting properties has
been difficult. As part of our continuing exploration in this field, we have designed and
synthesized novel Leu-enkephalin mimetics.
3.2 Design of Leu-enkeplialin Mimetics
Leu-enkephalin has been found to exist predominately in three different
conformations: extended,'"^ single-bend'^^"'"^ and double bend (Fig. 3-2).'""'' It has been
suggested that the single-bend form of enkephalin is representative of the bioactive
48
conformation at the 8-receptor. DPDPE was designed to mimic the single-bend
conformation and its enhanced 5 receptor selectivity supported this assumption. Some
other peptidomimetics have been also prepared to study this conformation (Fig.
3 _ 3 )_106,107
HgN
OH
H O
N'-r H 0
HO. HN
N-H
a) Extended b) Single-bend
OH yturn
OH
c) Double-bend
Figure 3-2. Schematic drawing of the three enkephalin conformations.
49
HOPC
HN O
-Fe-
CONH2
Figure 3-3. Peptidomimetics of Leu-enkephalin.
On the other hand, there was speculation that the double-bend conformation may
bind at fi-receptor sites.In order to further investigate the bioactive conformation of
Leu-enkephalin, we have designed novel double-bend form peptidomimetics of
enkephalin (Fig. 3-4).
tum
H /—?
,9 )=0---H-N ptum
H2N ° OH
Leu-enkephalin
0'-"^N b )=0
7/ 1 =0 H
y-Turn Peptidomimetics
H
HGN ^ OH
P-Tum Peptidomimetics
Figure 3-4. Reverse turn peptidomimetics of Leu-enkephalin.
50
One particular interesting approach in design of peptidomimetics is to replace a
dipeptide motif which adopts a biologically important confonnation in a given natural
peptide with a constrained or rigidified counterpart that simulates a so-called reverse turn.
It has been found that the double-bend conformation of Lcu-enkephalin is adopted in
biomimetic media where a y-tum is centered on Gly(2) and a p-tum is centered on
Gly(3)-Phe(4).^^'^°^ Thus we designed the reverse turn peptidomimetics of
Leu-enkephalin by incorporating a constrained dipeptide mimetic into the peptide to
replace dipeptide Tyr(l)-Gly(2) or Gly(3)-Phe(4) (Fig. 3-4)
Ideally, dipeptide mimetics should possess a scaffold with the right conformation and
appropriately positioned side chain functionalities in chiral space. The backbone of
such P-tum mimetics could then serve as a three-dimensional structural scaffold when the
full structures interact with their receptors/acceptors. Indeed, properly placed side chain
moieties involved directly in the interaction are critical for biological activities and
selectivities between peptide ligands and receptors/acceptors or sub-types
receptors.Several dipeptide mimetic systems have been proposed to mimic
different types of reverse turns.Among them, the azabicyclo[X.Y.O]alkane amino
acids potentially are a particularly important class due to their ability to provide
constrained backbone and side-chain conformations (Fig. 3-5).
COOH
R3 R|, Ri' R.v alkyl, aryl, etc n = ( ) , 1 , 2
Figure 3-5. Azabicyclo[X.Y.O]alkane amino acids.
51
Azabicyclo[X.Y.O]alkane amino acids are restricted dipeptide surrogates that
embody the peptide backbone within a bicyclic structure, in the bicyclic framework,
three contiguous (j)i-, and Wi-dihedral angles arc restricted by the structural
constraints of the heterocycle (Fig. 3-6). In addition, the outer two \\I2- and <])2-dihedral
angles are restricted by gauche interactions with the ring system. Thus this bicyclic
system provides the capacity of constraining five backbone bonds in a row within the
dipeptide. Moreover, it has been identified that the relative location and orientation of
the aromatic side chains is critical in the bioactivities and selectivities of Leu-enkephalin.
Hence, the constraints of % angles in azabicycloalkane amino acids can also provide
important information for the SAR studies (Fig. 3-6). Based on these advantages, we
have choscn azabicycloalkane amino acids as dipeptide mimetics incorporated into
Leu-enkephalin in order to study its bioactive conformation (Fig. 3-4). In our attempt to
implement this plan, we need to develop a flexible synthetic approach to such dipeptide
mimetics.
n=0, 1, 2 X=CH2, S, 0
Figure 3-6. General structure illustrating the dihedral angles constrained by an
azabicyclo[X.Y.O]alkane amino acid in a peptide.
52
3.3 Synthesis of Azabicyclo[X.Y.O]Alkane Amino Acids
Inherent in the synthesis of azabicycloalkane amino acids are three important
challenges: stereocontrol, side-chain attachment, and ring size (the three S's:
Stereochemistry. Side chains, and Size). The importance of stereochemistry is obvious,
since configuration can influence conformations. Thus chiral centers should be
introduced with control at the backbone carbons, ring fusion center, and attachment sites
of the side-chain appendages. The addition of various functional groups at appropriate
points along the azabicycloalkane heterocycle is also critical for mimicry of the nature
and the spatial orientation of amino acid side chains. Finally, the approach to a variety
of azabicycloalkane ring systems with different sizes is desired, because the size of the
heterocycle can bias the peptide conformation. Some synthetic methodologies have
been reported to prepare azabicycloalkane amino acids. Examples of the products
prepared by these methodologies are listed in Figure 3-7.
53
H
CbzHN N
O COOMe
J. E. Baldwin et a/. Tetrahedron 1989, 45, 4537
CbaHN
H
PhOCHgCHONH—K^^^
O OO2M©
J. E. Baldwin et al. Tetrahedron Lett. 1989, 30, 4019
H
CbaHN' Q CO2MG O COjBn
P. J. Belshaw et al. Syn/eff 1994, 381
J. E, Baldwin et al. J. Chem. Soc.; Chem. Commun. 1993, 935
H
PhthN
N-
Q CO2M©
J. A. RobI et al. Tetrahedron Lett. 1994, 35, 393.
Figure 3-7. Azabicycloalkane amino acids.
As listed in Figure 3-7, quite a few successes have been reported in obtaining
mimetics which can force or stabilize P-turas. However, little success has been reported
in incorporating mimetics for the active site of peptide hormone or neurotransmitter
receptors because of the lack of appropriately positioned side-chain groups. As
mentioned above, the side-chain functionalities are directly involved in the interaction
between the ligand and the receptor and thus are indispensable for the bioactivity,
especially for short peptides like Leu-enkephalins. Hence, there is quite a need for
methodology to introduce side-chain functionality on the backbone of the mimetics.
54
In order to prepare the Leu-enkephalin peptidomimetics shown in Figure 3-4, we
have developed an efficient approach to the stereoselective synthesis of azabicyclo[4.3.0]
(indolizidinone-type) alkane amino acids with appropriate side-chain appendages (Fig.
3-8). This approach can also be further explored to give rise to different size
azabicycloalkane ring systems through employment of starting materials of different
chain length. We chose 6,5-fuscd ring system as the first study target because modeling
studies showed that the dihedral angles in such systems mimic those angles in natural
reverse turn conformations better than other fused ring systems.
O H
HO OH
MeO.
COOH
y-Turn Peptidomimetics of Leu-enkephalin Dipeptide Mimetics
Figure 3-8. Dipeptide mimetics designed for Leu-enkephalin peptidomimetics.
3.4 Synthesis of Novel 4-Substituted Unsaturated Indolizidinone Amino Acids
We and other research groups have developed synthetic routes for the preparation of
enantiopurc indolizidinone type bicyclic lactam systems. " However, these
approaches suffer from some limitations. For example, the introduction of side chain
55
functionalities at the C4 position of azabicyclo[X.Y.O] alkane amino acids generally were
not accessible, or required a long synthetic sequence;^^"^ most methodologies have no way
to introduce a phenyl or a p-hydroxylphenyl group, which correspond to the side chains
in the amino acids Phe and Tyr, respectively. Moeller and co-workers chose a benzyl
group to substitute for a phenyl group. However, their studies showed that the extra
methylene group interfered badly in the binding to the TRH-R receptor.In this
section, we report a novel methodology which can allow for the synthesis of 4-phenyl- or
/7-hydroxylphenyl-substituted saturated and unsaturated indolizidinone amino acids.
Such reverse turn mimetics will be used to serve as surrogates of Tyr-Ala dipeptides and
be incorporated into peptides.
Some groups have reported on the preparation of indolizidinone type compounds
1 1 9 7 through dehydroamino acid intermediates. ' Further applications to these
methodologies have been successfully developed in our group to introduce side-chain
R9 functionalities at the C7 and C8 positions. Presently, we have extended these
methodologies to introduce aryl groups at position 4 through Suzuki cross-couplings.
The general retrosynthetic strategy is given in Scheme 3-1. The unsaturated bicyclic
lactam system could be approached from a dehydroamino acid intermediate, which can
be prepared by Horner-Emmons olefination of a proline aldehyde derivative. Two
challenges must be faced in our synthetic approach to these novel targets. The first is
the introduction of the side-chain functionality at the C4 position from the precursor
dehydroamino acid derivative. Secondly, the major Z dehydroamino acid products from
the Homer-Emmons reaction may restrict the cyclization in the following step. We
56
postulated that bromination of the dehydroamino acid could be a solution to these two
problems. Bromination can generate a reactive site for Suzuki cross-coupling and also
can provide the geometry required for cyclization. With this methodology, we also can
introduce side-chain functionalities at both C4 and C7 or C8 positions using the
corresponding chiral pyroglutamic ester derivatives prepared by chemistries previously
developed in our laboratory, which will be demonstrated in the next chapter. In this
chapter, we will demonstrate the synthetic method with side-chain functionalities at the
C4 position.
Scheme 3-1. Retrosynthetic analysis.
Bromination & Suzuki coupling
CbzHN COOMe
MeOOC CbzHN COOMe
Cyclization
Homer-Emmons olefination
MeOOCj_ r\
CbzHiPTN Allylation
O |\j COOMe Boc
Our approach to the synthesis of 4-substituted azabicyclo[4.3.0]alkane amino acid
derivatives 18a and 19a is illustrated in Scheme 3-2. The readily available
(5)-pyroglutamate 10 was reduced to the methoxy aminal 11 by treatment with
57
Super-Hydride (LiBEtsH) in THF at -78 °C, and then with methanol in the presence of a
catalytic amount of p-TsOH. The crude product 11 was directly subjected to
allyltrimethylsilane in the presence of boron trifluoride without further purification. The
allylsilane addition to the A'-acyUminium compound derived from 11 afforded a
3; 1 -c/.y/fra«.v-mixture of proline ester 12. The intermediate 12 underwent osmylation
and subsequent oxidation with NaI04 to give aldehyde 13. A cis/trans dehydroamino
acid mixture 14 was obtained in a 3:1 ratio via Homor-Emmons olefination of 13. The
mixture composition was based on 'H NMR spectra. The assignment of the major
product as a cis isomer was achieved according to the literature.
58
Scheme 3-2. Synthesis of 4-subslituted unsaturated indolizidinone amino acid esters.
O Iyj' 'COOMG Boc
10
COOMe
Boc
12
MeO ^QOOMe
Boc
11
O N Boc
13
COOMe
MeOOC.
CbzHN ^ COOMe
Boc 14
Br Br CbzHN^ / j \ CbzHN,^^ / / \
r^-^w-^COOMe * ^m-^COOMe MeOOC MeOOC
15a (53%)
R
15a CbzHN
MeOOC ^ COOMe
Boc
15b (crude)
. , f ^M''''^COOMe PhyHN (crude) MeOOC „ ObzHN
16a: R = phenyl (76%) 17a: R = 4-methoxyphenyl (79%)
CbzHN
CbzHN' Q COOMe
18a: R = phenyl (91%) 19a: R = 4-methoxyphenyl
(89%)
Boc
16b: R = 4-methoxyphenyl (crude)
COOMe
17b: R = 4-methoxyphenyl (two steps: 34%)
^Conditions: (a)(i) Super-Hydride, THF, -78 °C; (ii) p-TsOH (cat.), MeOH:
(b) BFg-EtaO, Me3SiCH2CH=CH2, three steps: 77%; (c) OSO4, Nal04,
THF/H2O, 4 h; (d) (MeO)2P(0)CH(NHCbz)COOCH3. DBU, DCM, rt, 8 h,
two steps: 63%; (e) (1) NBS, CHCI3, rt, 80 min; (ii) Dabco, CHCI3, rt,
24 h; (f) RB(0H)2, Pd(0Ac)2, P(o-tolyI)3, Na2C03, DME, 80 °C; (g) (i)
20% TFA, DCM, rt, 30 min; (ii) NaHC03; (iii) CHCI3, rt, 24 h.
59
The stereoselectivity favoring cw-isomer in the aliylation hinges on the facial
preference with which an allylsilane attacks the cyclic acyliminium intermediate
generated by the BF3 catalyzed OMe-elimination from aminal 11 (Fig. 3-9)/^^ It has
been suggested that the mechanistic pathway involved the formation of a BF3 complex
with the ester in which the fluoride acquired sufficient nucleophilicity to attack the
trimethylsilyl group and, hence, facilitate the concomitant allyl transfer to the iminium
function. Thus it can be suggested that the selective formation of the cis-product is the
result of a neighboring group participation of the methyl ester function.
OtBu
Figure 3-9. Transition state model for the generation of 12.
Although the P-bromination of dehydroamino acid esters has been well
d o c u m e n t e d , t h e l i t e r a t u r e r e p o r t e d i n c o n s i s t e n t s t e r e o s e l e c t i v i t y , a n d s u g g e s t e d
1^1 various intermediates in this reaction, including dibromides, N-hmmo
compounds, ' and a-bromoiminium species. In most cases, Z-selectivity was
observed in the bromination. Das and co-workers reported a 10:90 E/Z ratio in the
bromination of N-acyIdchydroalanine derivatives with Bra and EtsN; Danion and
co-workers described Z-selectivity in bromination of ethyl
2~(methoxycarbonylamino)cinnamate using NBS and EtsN;'^'^ Olsen and co-workers^^^
60
and Shin and co-workers'^^ also reported Z-selective halogenation under similar
conditions. On the other hand, some groups have reported /^-selective bromination
using the substrates with a particular substituent attached to the double bond.^^^''^^
Coleman and Carpenter have extensively examined the mechanism of bromination of
dehydroamino acid esters.They isolated a-bromoimine as the intermediate produced
by reaction of dehydroamino acids with NBS (Scheme 3-3). The two possible
ground-state conformers of a-bromoimine, A and B, undergo base-promoted
tautomerization to give E- and Z-vinyl bromide, respectively (Scheme 3-4).
Scheme 3-3. Bromination of dehydroamino acids.
R-iOCHN. XOpMe RiOCN^COgMe RiOCN^COgMe
Y T . T Rz H ^2 "'Br Rg-^Br
Base
RiOCHN^COaMe MeOaC^NHCORi
y + Ji Rg^Br Rg-^Br
Z
61
Scheme 3-4. Tautomerizalion of a-bromoimine.
H
R1OC—
RiOCHN^COsMe
COaMe Br
Base
Ro Br
Br
R1OC—N
Ra
COaMe Base
MeOaC^NHCORi
Ro Br H
B
Based on the results of molecular mechanics calculations, conformer A is energetic
favored and thus £-vinyl bromide is the kinetically favored product. However, this
difference in stability of conformer A and B does not account for the observed
Z-selectivity in the reaction. Meanwhile, Coleman and Carpenter observed that the
mixture of E and Z isomers underwent isomerization in the presence of DABCO and
predominately afforded Z-vinyl bromide. They also noticed that other strong but
sterically hindered amine bases are inefficient at promoting this isomerization. Based
on these observations, they proposed a mechanism for interconversion of the kinetically
formed £-vinyl bromide to the thermodynamically more stable Z-isomer, which contains
a Michael addition-elimination reaction sequence (Scheme 3-5). This assumption is
supported by the fact that DABO, the least hindered and most nucleophilic amine, is the
most effective agent promoting the isomerization.
62
Scheme 3-5. Mechanism of isomerization.
RjOCHN^COOCHg DABCO 1,4-addn R2OCHN.. ^
u ,x Ri Br R,—T"iBr
^ NRg
HsCOOC-^NHCORa retro OCH3 1,4-addn .Q/L/NHCOR2
Ri Br ' R3N
2
In our case, we treated the dehydroamino acid ester 14 with A^-bromosuccinimide
(NBS) to produce a-bromoimines, which underwent tautomerization to afford
(Z)-|3-bromo-a,p-dehydroamino acids 15ab upon treatment with an amine base. We
examined various amine bases which could be used in the tautomerization step. The
results showed that the use of DABCO instead of EtsN and DBU as a base improved
Z-selectivity and gave the Z-isomer exclusively. This result was consistent with the
observations of Coleman and Carpenter. The cisltrans isomers 15ab could be separated
at this point by column chromatography eluting with hexanes/ethyl acetate (4/1).
However, attempts to purify compound 15b were not successful due to a contamination
of the a-bromoimine intermediate in this reaction.
We then investigated the Suzuki cross-coupling of 15a with arylboronic acids. We
chose phenylboronic acid and 4-methoxyphenylboronic acid to use in the couplings
63
because Phe and Tyr arc two important amino acid moieties in our cx-MSH and 5-opioid
studies. Suzuki coupling went smoothly to afford 16a and 17a, in 76 and 79% yields,
respectively. Deprotection of A^'^-Boc group was performed in 20% TFA in
dichloromethane (DCM) at room temperature. The TFA salt was neutralized with
NaHCOs, and the reaction was stirred in chloroform at room temperature for 48 h.
Surprisingly the deprotected 16a, 17a underwent cyclization smoothly under this mild
condition without base and heat and afforded 18a, 19a in good yields. Crystallization of
19a from EtOAc and X-ray crystallographic analysis confirmed the configuration of the
bridgehead proton (Figure 3-10).
Figure 3-10. The X-ray structure of compound 19a.
On the other hand, we used the crude compound 15b in Suzuki cross-coupling and
the resulting crude product 16b was cyclized to afford 17b in 34% overall yield from 15b.
64
The stereochemistry of 17b was assigned with the use of ID-NOE experiments (Table
3-1).
Table 3-1. NOE Data for 17b
MeO. I7a
CbzHN Hg COOMe
17b
protons NOE (%)
He Hsa 1.93
Hg Hap 0.39
He Haa n.o.^
He Hap 0.68
Hsp H7„ 0.47
CO.
CO I
Hyp 1.30
Hsa Hra 1.46
Hsa HTP n.o.^
He H/A 0.26
He Hyp 2.01
a NOE not observed.
3.5 Synthesis of Novel 4-Substituted Saturated Indolizidinone Amino Acids
Once the unsaturated bicyclic lactams 18a, 19a were obtained, we investigated the
possibility of converting the unsaturated bicyclic structures to the saturated structures by
hydrogenation. The literature reported that the hydrogenation of dehydroamino acids in
open chain substrates preceded with poor stereoselectivity.''^ However, the restricted
conformation in the bicyclic compound provided an asymmetric environment in
65
hydrogenation. As we expected, compound 18a and 19a were hydro gen ated (Pd-C, Ha,
75 psi) to give 20a and 21a exclusively (Scheme 3-6).
Scheme 3-6. Hydrogenation of 18a and 19a.
H;, 75 psi
Pd-C, MeOH CbzHN
COOMe
18a: R = phenyl 20a: R = phenyl 19a: R = 4-methoxyphenyl 21a: R = 4-methoxyphenyl
The stereochemistry of 20a was determined based on the NOE data (Table 3-2). In
order to further confirm the relative configuration between the C-3 and C-4 protons, we
performed a modeling study which showed that the dihedral angles between H? and H4
arc 40" and 57° (two conformations) in the syn isomer, and -169" in the anti isomer. The
coupling constants were calculated on the basis of this finding as Jsyn = 4.1 Hz and Janti =
12.5 Hz. The observed coupling constant was 6.9 Hz. Considering the additional fact
that syn addition has been observed in most metal catalyzed hydrogenations, we drew the
conclusion that the relative configuration of H3 and H4 in compound 20a is syn.
66
Table 3-2. NOE Data for 20a
'So
Ph
COOMe
20a
protons NOE (%)
Hsp He 0.35
Hsa He 1.15
Hsp H4 0.50
Hsa H4 1.25
Hsp Ha n.o.^
^5a Ha 0.05
" NOE not observed.
We also performed some modeling studies in order to explain the stereoselectivity of
hydrogenation. The conformations of 18a and 19a suggested by these studies (Fig. 3-11)
were later confirmed by the X-ray structure (Fig. 3-10). The stereoselectivity of this
hydrogenation is probably due to the steric hindrance of the axial y proton adjacent to the
olefin group. Although the bottom face approach is sterically hindered by the
bridgehead proton, the axial y proton placed a greater steric effect in this case due to its
proximity to the reaction site.
67
Figure 3-11. Conformation of compound 18a suggested by modeling study.
As discussed above, wc chose the indolizidinone type system because it is expected
to be a good mimetic of a reverse turn conformation. Our modeling studies showed that
superposition of the two lowest energy conformations of the compound 20a onto a
peptide with various types of P-tum structures (Fig. 3-12) using backbone heavy atoms
gave an RMSD of 0.42 A for the type 11' structure for one conformation, and an RMSD
of 0.56 A for the type V' structure for the other conformation. In other words, the
backbone conformations of the compound best fit the criteria for type 11' and V P-tum
structures.
'vvrcn.
Turn Type 02 \|/2 <1)3 ii/3 I -60 -30 -90 0 r 60 30 90 0 U -60 120 80 0 ir 60 -120 -80 0 III -60 -30 -60 -30 Iir 60 30 60 30 V -80 80 80 -80 V 80 -80 -80 80
Figure 3-12. Classification of P-turns.
68
3.6 Synthesis of Leu-Enkephalin Minietics
Once the methodologies for preparing bicyclic P-tum dipeptide mimetics were
established, we investigated the incorporation of these mimetics into the bioactive peptide,
Leu-enkephalin. The Leu-enkephalin mimetic 24 was synthesized by the usual coupling
protocols (Scheme 3-7). Deprotection of 19a using the LiOH/MeOH system and
coupling with tripeptide H-Gly-Phe-Leu-OMe in DMF using PyBOP and HOBt as the
coupling reagents gave the protected Leu-enkephalin mimetic 22. Treatment of 22 with
IN aqueous Li OH in MeOH released the C-terminal of the peptide and treatment of
resulting product 23 with HBr (30% in AcOH) cleavcd the A^-Cbz protecting group.
However, a problem occurred in the step of purification by HPLC. Free peptidomimetic
24 was unstable in TFA/H20(1%) and decomposed to yield the corresponding hydroxyl
I -V-J -I IJQ j-y
product 25 (Scheme 3-8). ' Therefore, an A protecting group was required for the
stability of the peptidomimetic. Although it has been suggested that the cationic amine
is necessary to the biological activity of Leu-enkephalin, literature has reported the
1 -30 identification of a |i-selective opioid receptor agonist without a cationic amine group.
Scheme 3-7. Synthesis of Leu-Enkcphalin mimetic 24.
MeO
COOMe CbzHN
+ H-Gly-Phe-Leu-OMe
1)LiOH, MeOH 2) DMF, PyBop, HOBt, DIPEA
CbzHN CbzHN
MeO MeO OMe
23 22
HBr(30% in AcOH)
70
Scheme 3-8. Decompose of 24 in TFA/H2O.
MeO OH
24
HO,
MeO OH
25
3.7 Conclusion
In conclusion, we have successfully developed an approach to the synthesis of
unsaturated and saturated azabicyclo[4.3.0]alkane amino acids with an aryl side chain at
the C4 position. This methodology potentially can be extended to the preparation of
more complex molecules, such as 7/5 and 5/5 azabicyclo[X.Y.O]aIkane amino acids.
The unsaturated azabicyclo[4.3.0]alkane amino acid has been incorporated into the
biologically active peptide, Leu-Enkephalin. The study of structure-activity
relationships is still under investigation.
71
3.8 Experimental Section
(2S,5jR/S)-l-(iert-Butyloxycarbonyl-5-methoxy)-proline methyl ester (11). To a
solution of 10 (2.1 g, 8.63 mmol) in THDF (60 mL) was added Super-Hydride (1.0 M in
THF, 13 mmol) at -78 °C under Ar. After stirring at -78 "C for 40 min, the reaction
mixture was quenched with saturated NaHCOs (10 mL) and warmed to 0 °C. Then 30
drops of 30% aqueous H2O2 solution was added at this temperature. After the mixture
warmed to rt, THF was removed under reduced pressure. The reaction residue was
extracted with diethyl ether (3 x 25 mL). The organic extracts were combined, dried
over Na2S04, filtered and concentrated under vacuum to give a colorless oil. To the oil
in McOH (50 mL) was added p-TsOH-H^O (165 mg, 0.86 mmol) and the mixture was
stirred at rt overnight. After quenching with saturated NaHCOs (12 mL) and removal of
the solvent, the reaction mixture was extracted with diethyl ether (3 x 25 mL). The
combined organic extracts were dried over Na2S04, filtered and concentrated under
vacuum to give crude 11 as a colorless oil (2.03 g, 91%).
Methyl (2S,5/f/5)-l-(<ert-butyloxycarbonyl)-5-allylprolinate (12). To a solution
of crude 11 (2.03 g, 7.83 mmol) in diethyl ether (40 mL) was added allyltrimethylsilane
(5.5 mL, 34.6 mmol) and BF3«Et20 (1.10 mL, 9.1 mmol) at -40 "C under Ar. The cold
bath was removed after stirring at -40 "C for 15 min. The reaction mixture was stirred
for and additional 40 min, then quenched with NaHCO:, (20 mL) and extracted with
diethyl ether (3 x 30 mL). The organic extracts were combined, dried over Na2S04,
filtered and concentrated under vacuum to afford the crude product as a yellow oil.
Purification of the crude product by flash column chromatography (hexanes:EtOAc = 6:1)
72
gave a cis/trans mixture (3:1) of 12 as a colorless oil (1.78 g, 84%, and 77% overall yield
from 10). HRMS (FAB) calcd for C14H24NO4 (M+H) 270.1705, found 270:1703.
Methyl (25,5/?/S)-l-(fert-butyloxycarbonyl)-5-(formylmethyl)prolinate (13). To
a solution of 12 (1.76 g, 6.53 mmol) in THF (30 mL) and H2O (15 mL) was added OSO4
(80 mg, 0.32 mmol) in the dark. After 5 min, NaI04 (3.5 g, 16.4 mmol) was added in
small portions in 15 min. The reaction mixture was stirred at rt for 4 hr, then filtered,
and washed with MeOH (3 x 10 mL). After removal of the solvent, the residue was
redissolved in dichloromethane (30 mL). The organic solution was washed with brine
(3 X 30 mL), dried over Na2S04, filtered and concentrated under vacuum to give the
crude product 13 as a brown oil (1.55 g. 87%).
Methyl (2S)-cisltrans-(Z)-1 - (tert-butyloxycarbonyl)-5 - [(3' -amino- (A'-benzyloxy-
carbonyl)-3'-methoxycarbonyl)-2'-propenyI]prolinate (14). To a solution of
(Me0)2P(0)CH(NHCbz)C02Me (2.2 g, 6.64 mmol) in dichloromethane (50 mL) was
added DBU (990 fiL, 6.62 mmol) slowly at rt. After 10 min, to the above solution was
added a solution of 13 (1.55 g, 5.71 mmol) in dichloromethane (10 mL) and the reaction
mixture was stirred overnight at rt. After removal of the solvent, the residue was
redissolved in EtOAc (50 mL), washed with IN HCl (30 mL) and brine (50 mL), dried
over Na2S04, filtered and concentrated under vacuum to afford a brown oil. The crude
product was purified by flash column chromatography (hexanes:EtOAc = 3:1) to give a
cishrans (3:1) mixture of Z isomers 14 as a colorless oil (1.95 g, 72%, 63% overall yield
from 12). HRMS (FAB) calcd for C24H33N2O8 (M+H) 477.2237, found 477.2234.
Methyl {2S)-cis- and fr£ins-(Z)-l-(fert-Biityloxycarbonyl)-5-[2'-bromo-(3'-aniino-
73
(A'-benzyloxycarbon\i)-3'-niethoxycarbonyI)-2'-propenyl]prolinate (15ab). To a
solution of 14 (240 mg, 0.5 mmol) in chloroform (5 mL) was added NBS (98 mg, 0.55
mmol) at rt. After 80 min, Dabco (85 mg, 0.76 mmol) was added. The reaction
mixture was stirred at rt for 24 h. Then the reaction mixture was washed with saturated
NH4CI (3 X 5 mL) and dried over Na2S04. After removal of the solvent, the reaction
residue was subjected to flash column chromatography (Hexanes:EtOAc = 4;1) to afford
the cis isomer 15a as a light yellow oil (148 mg, 53% yield from cisltrans mixture 14)
and the trans isomer 15b (mixture). 15a. [a]'^D -10.3 (c 4.39, CHCI3); 'll NMR (500
MHz, CDCI3) (two rotamers) 5 7.36 (brs, 5H), 5.14-5.11 (m, 2H), 4.33-4.29 (m, IH),
4.23-4.19 (m, IH), 3.81 (brs, 3H), 3.73 (s, 3H), 3.17-3.12 (m, 1.5H), 2.93 (dd, 0.5H, 7=
14.4, 3.4 Hz), 2.24-2.21 (m, IH), 2.04-1.86 (m, 3H), 1.48 (s, 4H), 1.40 (s, 5H); NMR
(125 MHz, CDCI3) ( two rotamers) 5 174.0, 173.8, 163.0, 162.9, 154.0, 153.5, 153.3,
153.2, 135.7. 135.6, 128.8, 128.7, 128.6, 128.5, 128.4, 128.3, 80.7, 80.5, 68.2, 68.0, 60.3,
59.8, 57.6, 53.0, 52.4, 52.2, 40.0, 39.3, 29.1, 28.7, 28.6, 28.5, 28.4, 28.0; HRMS (FAB)
calcd for C24H32BrN208 (M+H) 555.1342, found 555.1337.
Methyl (25)-CM-(Z)-l-(fert-ButyIoxycarbonyI)-5-[2'-phenyI- and 4-
methoxyphenyl- (3' -amino- (/V-beiiz;yloxycarbonyl)-3' -methoxycarbonyl)-2' -propenyl]
prolinate (16a, 17a). To a solution of 15a (175 mg, 0.32 mmol) in DME (2 mL) and
degassed water (400 jiL) was added phenylboronic acid (80 mg, 0.65 mmol), NaiCOs (70
mg, 0.66 mmol), Pd(0Ac)2 (8 mg, 0.035 mmol) and P(o-tolyl)3 (10 mg, 0.033 mmol).
Then the reaction mixture was stirred at 80 "C overnight. The reaction mixture was
passed through a short column containing a bottom 1" layer of silica gel and a top 1"
74
layer of NaHCOs using ethyl acetate as eluent. After removal of the solvent, the crude
product was subjected to flash column chromatography (hexanes:EtOAc = 3:1) to afford
16a as a colorless oil (134 mg, 76%). The identical procedure afforded 17a as a
colorless oil with a yield of 79%. 16a. [a]^^D -142.3 (c 8.72, CHCI3); 'H NMR (500
MHz, CDCI3) (rotamers) 5 7.34-7.18 (m, lOH), 5.98 (brs, IH), 5.08 (s, 2H), 4.24 (I, 0.6H,
J = 8.0 Hz), 4.14 (t, 0.4H, J = 8.0 Hz), 3.84 (brs, 3H), 3.74 (s, 3H), 3.70-3.65 (m, 0.4H),
3.57-3.52 (m, 0.6H), 3.18-3.15 (m, 0.6H), 3.14-3.05 (m, 0.6H), 3.04-2.99 (m, 0.4H),
2.91-2.88 (m, 0.4H), 2.19-2.02 (m, 3H), 1.74-1.63 (m, IH), 1.39 (s, 4H), 1.38 (s, 5H); '''C
NMR (125 MHz, CDCis) (rotamers) 5 174.2, 173.9, 165.6, 165.4, 154.1, 154.0, 153.9,
153.4, 137.1, 136.8, 136.1, 136.0, 134.5, 134.2, 129.4, 129.3, 128.8, 128.7, 128.6, 128.5,
128.3, 128.2, 125.9, 125.7, 80.2, 80.0, 67.6, 60.5, 60.0, 56.6, 56.5, 52.5, 52.3, 52.1, 36.7,
35.6, 28.9, 28.7, 28.5, 28.3, 28.2, 27.8; HRMS (FAB) calcd for C30H37N2O8 (M+H)
553.2550, found 553.2554. 17a. -214.1 {c 2.30, CHCI3); 'H NMR (600 MHz,
CDCI3) (rotamers) 6 7.36-7.31 (m, 5H), 7.21-7.13 (m, 2H), 6.91-6.88 (m, 2H), 6.00 (brs,
IH), 5.09 (s, 2H), 4.25 (t, 0.5H, J = 8.0 Hz), 4.14 (t, 0.5H, J = 8.0 Hz), 3.83-3.81 (m, 6H),
3.75 (s, 3H), 3.74-3.70 (m, 0.5H), 3.61-3.57 (m, 0.5H), 3.15-3.11 (m, 0.5H), 3.10-3.04 (m,
0.5H), 3.00-2.98 (m, 0.5H), 2.90-2.88 (m, 0.5H), 2.18-2.06 (m, 3H), 1.71-1.63 (m, IH),
1.39 (s, 4H), 1.37 (s, 5H); '^C NMR (150 MHz, CDCI3) (rotamers) 8 174.2, 173.9, 165.8,
165.6, 159.8, 154.1, 153.5, 153.3, 136.1, 134.9, 129.7, 129.6, 129.0, 128.7, 128.6, 128.5,
128.4, 125.6. 125.2, 114.8, 114.7, 80.2, 80.0, 67.6, 60.5, 60.0, 56.7, 56.6, 55.5, 55.4, 52.5,
52.3, 52.1, 36.6, 35.5, 29.0, 28.7, 28.5, 28.3, 28.2, 27.7; HRMS (FAB) calcd for
C31H39N2O9 (M+H) 583.2656, found 583.2634.
75
(6/?,9S)-l-Aza-3-iV-benzoxycarbonylamino-9-methoxycarbonyl-4-phenyl- and 4-
methoxyphenyi-2-oxobicyclo[4.3.0]non-3-enes (18a and 19a). To a solution of 16a
(65 mg, 0.12 mmol) in dichloromelhane (2.4 mL) was added TFA (0.6 mL) at rt. The
mixture was stirred for 30 min, then saturated NaHCOs (10 mL) was added. After
stirring for 20 min, the organic solution was dried over Na2S04 and concentrated under
vacuum. The residue was redissolved in chloroform (5 mL) and stirred at rt for 48 h.
After removal of the solvent, the crude product was chromatographed (hexanes:EtOAc =
1:2) to afford 18a as a white solid (45 mg, 91%). The identical procedure afforded 19a
as a white solid with a yield of 89%. 18a. mp 169-171 "C; [a]'"^D -24.8 (c 3.53. CHCI3);
NMR (600 MHz, CDCI3) 5 7.37-7.20 (m, lOH), 6.51 (brs, IH), 4.99 (d, IH, J = 12.0
Hz), 4.91 (d, IH, /= 12.0 Hz), 4.61 (d, IH, /= 8.5 Hz), 4.07-4.02 (m, IH), 3.77 (s, 3H),
2.89-2.81 (m, 2H), 2.25-2.19 (m, 2H), 2.17-2.13 (m, IH), 1.90-1.83 (m, IH); "C NMR
(150 MHz, CDCI3) 5 172.6, 161.7, 153.7, 138.5, 138.2, 136.4, 128.7, 128.6, 128.4, 128.2,
127.4, 124.5, 67.2, 57.9, 56.2, 52.7, 36.6, 31.7, 29.0; HRMS (FAB) calcd for
C24H25N2O5 (M+H) 421.1763, found 421.1752. 19a. mp. 170-172 "C; [af'o-19.6 (c 0.94,
CHCI3); 'H NMR (600 MHz, CDC13) 8 7.34-7.22 (m, 7H), 6.85 (d, 2H, J = 8.6 Hz), 6.52
(brs, IH), 5.01 (d, IH, J = 12.3 Hz), 4.94 (d, IH, J = 12.3 Hz), 4.60 (d, IH, J = 8.6 Hz),
4.04-4.00 (m, IH), 3.81 (s, 3H), 3.76 (s, 3H), 2.88-2.77 (m, 2H), 2.24-2.12 (m, 3H),
1.89-1.84 (m, IH); NMR (125 MHz, CDCI3) 5 172.6, 161.9, 159.6, 153.8, 138.2,
136.5, 130.6, 128.9, 128.5, 128.2, 128.1, 123.7, 114.1, 67.1, 57.8, 56.1, 55.4, 52.6, 36.5,
31.7, 29.0; HRMS (FAB) calcd for C25H27N2O6 (M+H) 451.1869, found 451.1862.
(6S',95)-l-Aza-3-A^-benzoxycarbonylamino-9-methoxycarbonyl-4-phenyl-2-oxo-
76
bicycle[4.3.0]non-3-ene (17b). Prepared according to the method used for 16a and 18a,
23 starting from mixture 15b, in 34% overall yield from 15b, as a colorless oil. [a] o
+43.9 (c 3.29 , CHCI3); 'li NMR (500 MHz, CDCI3) 5 7.34-7.19 (m, 7H), 6.84 (d, 211, J
= 9.0 Hz), 6.69 (brs, IH), 4.97 (d, IH, /= 12.5 Hz), 4.88 (d, IH, /= 12.5 Hz), 4.54 (t, IH,
/= 8.0 Hz), 4.25-4.19 (m, IE), 3.81 (s, 3H), 3.77 (s, 3H), 2.92 (dd, IH, /= 17.0, 4.9 Hz),
2.60 (dd, IH, J = 17.0, 14.1 Hz), 2.47-2.40 (m, IH), 2.36-2.30 (m, IH), 2.01-1.93 (m,
IH), 1.77-1.69 (m, IH); NMR (125 MHz, CDCI3) 5 172.7, 162.0, 159.5, 153.3, 136.5,
136.0, 131.1, 128.7, 128.6, 128.2, 128.1, 123.1, 114.1, 67.1, 58.5, 55.9, 55.4, 52.6, 36.5,
32.9, 28.8; HRMS (FAB) calcd for C25H27N2O6 (M+H) 451.1869, found 451.1851.
(3S',4i?,6if,95)-l-Aza-3-amino-9-methoxycarbonyI-4-phenyl-2-oxobicyclo [4.3.0]-
nonane (20a). A solution of 18a (60 mg, 0.14 mmol) in degassed MeOH (5 mL) with
Pd/C (10 wt.%, catalytic amount) was reacted at an initial H2 pressure of 75 psi at rt.
After 24 h, the mixture was filtered and the solvent was removed under reduced pressure.
The crude product was purified by flash column chromatography (5% MeOH in
dichloromethane) to give 20a as colorless oil (34 mg, 84%). [aj^^o -67.8 (c 0.73 ,
CHCI3); 'H NMR (600 MHz, CDCI3) 5 7.35-7.23 (m, 5H), 4.54 (d, IH, J = 8.9 Hz),
3.81-3.76 (m, 4H), 3.65 (d, IH. J = 6.9 Hz), 3.51-3.47 (m, IH), 2.48-2.44 (m, IH),
2.25-2.17 (m, 2H), 2.13-2.07 (m, 2H), 1.85-1.78 (m, IH), 1.35 (brs, 2H); NMR (150
MHz, CDCI3) 5 172.5, 172.2, 141.6, 128.9, 128.6, 127.2, 58.4, 57.7, 54.4, 52.6, 44.5,
34.8, 31.9, 29.1; HRMS (FAB) calcd for C16H21N2O3 (M+H) 289.1552. found
289.1558.
(35,4/?,6/?,95)-l-Aza-3-amino-9-methoxycarbonyI-4-methoxyphenyl-2-oxo-
77
bicyclo[43.§]Bonaiie (21a). In a manner similar to the preparation of 2§a, using 19a as
starting material gave 21a in 67% yield. 21a. 'H NMR (600 MHz, CDCI3) 6 7.22 (d,
2H, J = 8.6 Hz), 6.87 (d, 2H, J = 8.6 Hz), 4.53 (d, IH, / = 8.5 Hz), 3.84-3.74 (m, 7H),
3.61 (d, IH, 7.1 Hz), 3.48-3.43 (m, IH), 2.48-2.43 (IH), 2.25-2.17 (m, 2H), 2.16-2.04 (m,
2H), 1.85-1.77 (m, IH); HRMS (FAB) calcd for C17H23N2O4 (M+H) 319.1658, found
319.1648.
78
CHAPTER 4. STEREOSELECTIVE SYNTHESIS OF 4,8-DlSLIBSTITUTED
AZABICYCLO[4.3.0]NONANE AMINO ACIDS AS PEPTIDOMIMETICS
SCAFFOLDS OF MELANOCORTIN RECEPTOR LIGANDS
4.1 Introduction
a-Melanotropin (a-melanocyte stimulating hormone, a-MSH) is a linear
tridecapeptide consisting of the amino acid sequence Ac-Scr-Tyr-Ser-Met-Glu-His-
Phe-Arg-Trp-Gly-Lys-Pro-Val-NHa. It is synthesized in the vertebrate pars intermedia
and in the brain.Biologically, a-MSH is known primarily for its ability to
stimulate the integumental melanocyte, in addition to interacting with several other
proposed peripheral and central biological systems.'^
To determine the biologically active conformation of a-MSH, numerous analogues
have been designed and synthesized. Substitution of Nle (sidechain = CH2CH2CH2CH3)
for Met'^ resulted in peptides with more potent biological activity. Inversion of
configuration of L-Phe^ to D-Phe' led to the analogue [Nle'^,DPhe^]a-MSH, which is
referred as "MT-I" and "NDP-a-MSH". This analogue showed a substantial increase in
potency as well as prolonged activity.*'^^ ''"' The conformationally restricted analogue
MT-II possesses potency up to 90 times greater than a-MSH (Figure 4-1).^°'^'^'' Because
cyclization leads to considerable restriction of conformational flexibility of the peptide
backbone and, to a lesser degree, the side-chain groups, MT-Il lends itself to a more
complete evaluation of the biologically active 3D topologies.
79
Ac-Nle-Asp-His-D-Phe-Arg-Trp-Lys-NH2
MT-li
Figure 4-1. Structure of MT-II.
To understand further the effects of conformation on biological potency, we would
like to design and synthesize novel analogues with conformationally constrained
dipeptide mimetics.
4.2 Design of Conformationally Restricted Analogues
Recently the major effort in peptidomimetic research has been to find the organic
moieties that can replace the peptide scaffold and position the crucial recognition
elements in 3D space correctly. Of particular interest are the constrained scaffolds that
not only possess the right conformation but also allow for a wide-ranging display of
substitutes. Azabicyclo[4.3.0]alkane amino acids are unique dipeptide P-tum mimetics.
As we introduced in previous chapters, these scaffolds have been applied as secondary
structure replacement due to their capacity of restricting five backbone bonds in a row
within the peptide. Although the studies have shown that indolizidinone systems can
mimic |3-tum confonnation, little success in the stereoselective incorporation of diverse
substituents on their backbones has limited their application as peptidomimetic scaffolds.
Herein we would like to develop an efficient approach to the synthesis of
4,8-disubstituted azabicyclo[4.3.0]nonane amino acids as peptidomimetic scaffolds of
melanocortin receptor ligands.
80
As part of our ongoing program for the design of novel melanotropin peptide
mimetics, we have identified the core bioactive sequence of melanotropin peptides as
His-(D/L)Phe-Arg-Trp (Figure and found a P-turn structural feature centered in
these core residues.The studies of structure-activity relationships of melanotropin
peptides have resulted in the development of various more potent, prolonged acting and
enzymatically stable analogues, such MT-I and MT-II, which have been widely used in
biological studies. However, these analogues did not show significant selectivity for the
different melanocortin receptors. Recently, we reported the first example of a selective
and potent antagonist at the hMC3 receptor, MK-9.'^'* Based on our conformational
studies of melanotropin peptides and their analogues, we postulated that indolizidinone
systems can serve as peptidomimetic scaffolds of the core sequence in melanotropin
peptides which not only mimic P-tum conformation but also place amino acid side-chain
functionalities in space correctly (Scheme 4-1).
a-MSH Ac-Ser-Tvr-Ser-Met-Glu-Hls- Phe-Arg-Trp-Glv-Lvs-Pro-Val-NHo
MT-I Ac-Ser-Tvr-Ser-Nle-Glu-His-D-Phe-Ara-Trp-Glv-Lvs-Pro-Val-NHo
MT-II Ac-Nle-Asp-Hls-D-Phe-Arg-trp-Lvs-NHo
Figure 4-2. Core sequence of melanotropin peptides.
81
Scheme 4-1. Analogues of melanotropin peptide core sequence.
His-D-Phe-Arg-Trp
For designing the synthetic targets 26. we chose the benzyloxycarbonyl group as a
bioisosteric replacement of His because it can be readily introduced from commercially
available reagents. The phenyl group as R2 corresponds to the side-chain of Phe. The
2-naphthyl group is also an interesting element to be incorporated here because we found
that introducing a Nal residue in place of Phe in MT-II converts this potent agonist to a
potent antagonist (SHU9119).^^^ For R3, both guanidine and amine groups are placed at
this site since some literature suggested that the guanidine group might not be essential in
MC4R agonists when a heteroatomic substituent is present in a proper position. Lastly,
tryptamine and phenethylamine are chosen to serve as bioisosteric replacements of
tryptophane amide at the Rj position.
82
4.3 Synthesis of Disubstituted Indolizidinone Amino Acids
4.3.1 Background and General Approach
Our group has developed a strategy to introduce a sidc-chain group at the C-8
position of indoHzidinone amino acids.The strategy involves employing ^-substituted
pyroglutamates as chiral synthetic precursors which could be prepared stereoselectively
by methods recently developed in our laboratory.Stercoselectively introducing
allyl groups at the C-5 position of pyroglutamates and their appropriate elaboration could
afford dehydroamino acid intermediates, which could undergo asymmetric
hydrogenations and cyclizations to afford C8-substituted indolizidinone amino acids. In
addition, as discussed in the previous chapter, we recently have developed a novel
methodology which could readily prepare C4-substituted unsaturated and saturated
indolizidinone amino acid esters.The functional groups at C-4 were introduced by
bromination of dehydroamino acid intermediates followed by Suzuki couphng.
Hydrogenation of the unsaturated bicyclic dehydroamino acid esters provided saturated
bicyclic lactams. These successful synthetic results directed our attention to developing
a convergent synthetic approach using a combination of these two methods to synthesize
peptidomimetics 26. The retrosynthetic analysis of the target analogues 26 is described
in Scheme 4-2.
83
Scheme 4-2. Retrosynthctic analysis of mimctic 26.
8>""H3
cyclization coupling
O
bromination & Suzuki coupling
RIHN
MeOOC jvj' 'COOM0 Boc
Michael addition
J I cyclization
COOMe
Boc
O
R3 OH J
Wittig olefination
O R3
allylation r„
"••">-00. r ^^^^Ki^COOMe
Horner-Emnnons olefination
^OH
4.3.2 Synthesis of p-Substituted Pyroglutamic Acid Ester.
As the first goal in our studies, a method for introducing an appropriate side-chain
group at C-8 was investigated. As discussed above, the side-chain functionality could
be introduced at the C8 position by using the pyroglutamic acid ester with a
corresponding substituent at the (3-position as the starting material. Therefore, a novel
P-substituted pyroglutamic acid was the first synthetic target in the approach.
84
Scheme 4-3." Synthesis of Michael acceptor 30.
29
c, d, e
Of-Bu
N O
30
"Conditions; (a) PCC, silica gel, DCM; (b) ;BuOCOCH2PPh3Br, NaOH, TEA,
CH2CI2, H2O; (c) TFA (50% in CH^Cl.); (d) rBuCOCl, TEA, THE, -78 °C;
(e) (S)-4-phenyl-2-oxazolidinone, nBuLi, THE, -78 °C;
As illustrated in Scheme 4-2, the proposed approach would include diverse reactions.
Therefore, it was important to identify the proper protecting group for the amino group in
the starting material which would later be functionalized in the final step. This
protecting group should be sufficiently robust to survive the projected reactions, and also
should be orthogonal to other functionalities and be labile enough to be removed in the
final step. Furthermore, based on our earlier studies, it seemed likely that a
mono-protected nitrogen would interfere with the aldehyde intermediate.^^ All of these
requirements prompted us to choose the phthalimide group to doubly protect the amine
group. Aldehyde 28 was prepared in good yield by PCC oxidation of alcohol 27
85
following the literature procedure (Scheme 4-3)."'"^ However, when the reaction was
performed on a large scale, the reduced chromium byproduct made workup difficult.
Thus we employed the Swem oxidation as a good alternative synthetic method to prepare
28 on a large scale. Wittig olefination of aldehyde 28 with (?-butoxycarbonylmethyl)
triphenylphosphonium bromide in the presence of NaOH and triethylamine in a
two-phase system of dichloromethane/HaO gave the £'-5-A'-phlhalimido-a,P-unsaturatcd
/-butyl ester 29 in excellent yield. The /-butyl protecting group was removed by
treatment with 50% TFA in dichloromethane, and 5-4-phenyl-2-oxazolidinone was
coupled to the deprotected 29 as the chiral auxiliary for asymmetric functionalization of
the P-position in the next step.
Optically pure 4-substituted oxazolidin-2-ones were introduced as chiral auxiliaries
by Evans"^'*' and later were widely used due to their excellent stereoconlrolling power in
acylation,'^^ aldol condensation,"'^"^''' and Diels-Alder reactions.'™ According to the
literature, successful application of chiral 4-substitutcd oxazolidin-2-ones in conjugate
addition reactions always required the use of a chelating agent (e.g. LiCl, MgC104, or
CuBr) to form the corresponding metal-chelated intermediates, in which the
stereocontrolling 4-substituted group could be located in close proximity to the CC
double bond so as to efficiently control its facial selectivity (Scheme 4-4).'^'''^"
Furthermore, the eiectrophilicity of the CC double bond is substantially enhanced, as
compared with the corresponding esters, due to the fact that an oxazolidin-2-one is a
substantially stronger electron-withdrawing substituent than alkoxy groups. Thus
4-substituted oxazolidin-2-ones can not only provide excellent diastereoselectivity, but
86
also enhanced reactivity in asymmetric Michael addition.
Scheme 4-4. Metal-chelated intermediate in Michael addition.
Ph. \—V
V\ R ^ R. CuBr-SMe2 Y
T n ^ P—CU Q O o o
R—cu q p ' \ / R Mg
^Br
Meanwhile, chiral Ni(II) complex (S)-32 was prepared according to a literature
procedure (Scheme 4-5).'"^ First the amination of (5)-proline with benzyl chloride was
conducted at 40 °C in /-PrOH in the presence of KOH with 78% yield to give
/V-benzylproline in hydrochloride salt form. Then //-benzylproline was coupled with
2-aminobenzophenone to give the chiral ligand,
A'-(A^-benylprolyl)-2-aminobenzophenonc (BPB, 31). After simple work-up by washing
with deionized water, the ligand was used for the next step without further purification.
The Ni(Il) complex 32 was formed between the chiral ligand and glycine in the presence
of NiCl-6H20 under strong base in MeOH at 55 "C for 6 hours.
87
Scheme 4-5. Preparation of the chiral Ni(TI) complex (S)-32.
NH2 O
H
(S)-Pro
Ph^ CI
(S)-BP
Ph
BOP
NH O Glycine, NiCl2 KOH/MeOH
Ph O'
N
O ^
Ni^N ^Ph
(S)-BPB 31 32
The presence of the proline stereogenic center in the complex imparts an asymmetric
distortion to the rigid polycyclic system of the chelate rings, which brings about a steric
shielding of the corresponding enolate on the re face by the ketimine phenyl group.
Accordingly, in the Michael addition, the electrophile approaches preferentially from the
si face to form a new complex with pseudoaxial orientation of the introduced substituent
which usually leads to an a-(5) absolute configuration (Scheme 4-6).
88
Scheme 4-6. Selecti ve electrophilic attack on the complex enolate.
32
a-(S}
Ph-electrophile Base
'N--NI—N -N Ph
si favorable
In agreement with the above explanations, Michael acceptor 30 underwent
asymmetric Michael addition with the NiCII) complex 32 to give a mixture of (2S,35)-33a
and (2R,3S)-33h in a ratio of 9/1 (Scheme 4-7). The stereochemistry of products was
assigned based on our detailed knowledge of similar Ni(II) complexes.
Scheme 4-7. Asymmetric Michael addition between 30 and 32.
O
N +
30
Ni-^N / ^Ph
32
DBU(15 mole%) DMF,RT
Ph , / ° / VX -Ph
(25, 35)-33a {2R, 35)-33b
89
Hydrolysis of the Ni(ll) complex 33a followed by cyclization under the basic
condition afforded the corresponding {5-substituted pyroglutamic acid (Scheme 4-8).
Attempts to purify the amino acid using a Dowex ion-exchange resin column were not
successful probably due to a contamination of the uncyclized product. Instead the crude
amino acid was protected directly as the N^-Boc pyroglutamate 34. Compound 34 was
purified by flash column chromatography and the configuration was confirmed by X-ray
crystallographic analysis (Figure 4-3).
Scheme 4-8." Hydrolysis of Ni(II) complex 33a.
Ph A / ? i «N-^Nh—N
COOMe
" (a) 3N HCl, MeOH; (b) NH4OH; (c) SOjCl, MeOlI; (d) (Boc)20, DMAP.
Acetonitrile
90
Figure 4-3. The X-ray structure of compound 34.
4.3.3 Synthesis of 8-Substituted Azabicyclo[4.3.()]Alkane Amino Acids
The lactam moiety of pyroglutamate usually could be reduced selectively by
Super-Hydride (LiBEtsH).'^'* However, in agreement with a previous report,''^ the
treatment of 34 with Super-Hydride (LiBEtsH) also resulted in the reduction of the
phthalimide protecting group. Therefore, we chose to use DIBAL-H which the
phthalimide group is stable to. The reduction by DIBAL-H followed by treatment with
methanol in the presence of a catalytic amount of p-TsOH afforded the methoxyaminal
which was directly subjected to allyltrimethylsilane and boron trifluoride in ether without
further purification (Scheme 4-9). Consistent with our previous observations, the
allylsilane addition to the vV-acyliminium intermediate gave exclusively the trans product
related to the p-substituent as a result of a neighboring group participation of the methyl
91
ester and the steric effect of the P-substituent. The optically pure intermediate 35
underwent osmylation and subsequent oxidation with NaI04 to afford the aldehyde
intermediate. The dchydroamino acid ester 36 was obtained via the Homer-Emmons
1 nft olefination of the aldehyde intermediate.
Scheme 4-9/' Synthesis of dehydroamino acid derivative 36.
a, b. c
d, e
MeOOC.
CbzHN
COOM©
Boo
35
|yj COOM©
Boo
36
" (a) DIBAL-H, THF, -78 °C; (b) Ts-OH, MeOH; (c) BFj-EtjO,
MesSiCHjCH^CHj, EtjO, -40 °C; (d) OSO4, NaI04, THF/HjO;
(e) DBU, (Me0)2P(0)CH(NHCbz)C00CH3, CHjClj, rt.
92
With the important intermediate 36 in hand, we employed asymmetric
hydrogenations on this substrate to prepare a-amino acid derivatives 37a and 37b
(Scheme 4-10). The Bark's catalysts [Rh(I) (COD) (R.R)- or (S,S)-Et-DuPHOS] OTf
were used as the catalyst in hydrogenation with high yields (>90%) and high
diastereoselectivity (>96% ee)/^^'^^^ Deprotection of N^-Boc group was performed in
20% TFA in dichloromethane at rt. The TFA salt was neutralized with NaHCOs and the
reaction was stirred in chloroform at rt. The cyclization proceeded smoothly under
these mild conditions to give bicyclic lactams 38 and 39 in good yield.
Scheme 4-10." Synthesis of 8-[2-(iV-phthalimido)-ethyl] indolizidinone amino acid esters.
MeOOC,
COOMe CbzHN CbzHN
COOMe
MeOOC J S)
COOMe CbzHN CbzHN
COOMe
37b 39
® (a) Rh(l)(COD)-(afl)-Et-DuPHOS, Hg (75 psi), MeOH; (b) Rh(l)(COD)-(S,S)-Et-DuPHOS, Hg (75 psi),
MeOH; (c) TFA (20% in CH2CI2); (d) CHCI3, rt, 24 h.
93
4.3.4 Synthesis of 4,8-Disubstituted Indolizidinone Amino Acid Ester
Having developed the methods to introduce side-chain groups at the C-4 and C-8
positions, respectively, we were able to carry out experiments aimed at preparing
peptidomimetics 26 by combining the two methodologies. With the dehydroamino acid
ester 36 in hand, we treated it with iV-bromosuccinimide (NBS) in chloroform to producc
the a-bromoimines (Scheme 4-11). Upon treatment with an amine base, the
a-bromoimine intermediates underwent tautomerization to afford the
(Z)-P-bromo-a,(3-dehydroamino acid 40. Consistent with our previous observation,
using DABCO in stead of TEA and DBU in the tautomerization step resulted in the Z
isomers exclusively. Suzuki coupling of 40 with phenylboronic acid and
2-naphthalcneboronic acid introduced the phenyl and 2-naphthyl groups at the P-position
of the dehydroamino acid. Deprotection of the V-Boc group in 41 and 42 was
performed with 20% TFA in dichloromethane at it. The TFA salt was neutralized with
NaHCOs solution and the reaction was stirred in chloroform at rt. The cyclization
proceeded smoothly to give bicyclic lactam 43 and 44 in good yields after 24 h.
94
Scheme 4-11/ Synthesis of 4,8-disubstituted azabicyclo[4.3.0]alkane amino acids.
MeOOC.
CbzHN COOMe
Boc
36
a,b
CbzHN
MeOOC COOMe
Boc
40
CbzHN
MeOOC COOMe
Boc
41; Ri = phenyl 42; Ri = 2-naphthyl
d, e f o
CbzHN
O - COOMe
43; Ri = phenyl 44: R, = 2-naphthyl
^(a)NBS, CHCI3; (b) Dabco, CHCI3; (c) PhB(0H)2, Pd(OAc)2, P(o-tolyl)3, NagCOs, DME, 80 °C;
(d) 20% TFA,CH2Cl2, rt; (e) CHCI3, rt, 24 h.
4.3.5. Synthesis of Peptidomimetics of the Core Sequence in Melanotropin
Peptides
Deprotection of the phthalimide protecting group in 43 and 44 was accomplished by
treatment with hydrazine at rt for 24 h (Scheme 4-12). Work-up and purification of free
amine gave poor yields. Thus the crude free amines 45 and 46 were directly
guanidinated to give 47 and 48, using the commercial available guanidinating reagent.
95
A^,iV'-bis(te?t-butoxycarbonyl)-//"-triflylguanidine. Although an initial attempt using
DMAP as base failed, the guanidinating reaction with triethylamine afforded 47 and 48 in
good yields. The configuration of 47 was confirmed by NOE (Table 4-1). Hydrolysis
of the methyl ester group using IN Li OH and MeOH (1:2) went slowly due to the steric
hindrance of the neighboring group. Subsequently, deprotected 47 and 48 were treated
with tryptamine or phenethylamine in DMF using BOP and HOBt as the coupling reagent
to yield 49- 51. On the other hand, hydrazinolysis of 44 followed by protection of
amine with Boc afforded 52 (Scheme 4-13). In a similar way, tryptamine was coupled
to 52 to afford 53. Compounds 49-51 and 53 were treated with 20% TFA in
dichloromethane to remove the Boc group. The deprotected products were purified by
HPLC and submitted for biotests.
96
COOMe
Scheme 4-12/' Synthesis of analogues 49-51.
Q
HN^
43: Ri = phenyl
44; Ri = 2-naphthyl
H NH,
COOMe
45: R-i = phenyl
46: R-i = 2-naphthyl
BocN '^NHBoc
-NH
HN' O COOMe
47: R-I = phenyl
48: R-I = 2-naphthyl
c, d
BocN ^-NHBcc
NH
49 50 51
R-I = phenyl; Ra = tryptamine
Ri = phenyl; R2 = phenethylamine
Ri = 2-naphthyl; R2 = tryptamine
®(a) NH2NH2, EtOH, CHCI3; (b) A/,W-Bis(fert-butoxycarbonyl)-A/"-triflylguanldine, TEA, CH2CI2:
(c) LiOH(IN), MeOH; (d) PyBOP. HOBt, DIPEA, R-NH2, CH2CI2.
Table 4-1. NOE Data for 47
BocN
CbzHN
NHBoc ^H7a NH protons NOE
Hg H7„ 0.80
Hs Htp 1.46
bOOMe He Hva 1.05
He H7P 0.66
He Hg n.o.®
He H<, 0.20
"NOE not observed.
97
Scheme 4-13." Synthesis of analogue 53.
NHBoc
b HN
COOMe
52
HN COOMe
44
NHBoc
HN
HN
NH 53
^(a) NH2NH2, Eton, CHCI3; (b) (BocjgO, TEA, CH2CI2: (c) IN LiOH, MeOH; d) PyBOP, HOBt, DIPEA, DCM, tryptamine.
4.4 Future Work
In our ongoing opioid and a-MSH program, the studies have indicated that the
conformational requirements for optimal interaction with different receptors differ in a
subtle manner.On the other hand, the configurations of the peptidomimetics
influence their conformations. Thus it is very important for us to have the approach to
every individual stereoisomer of the peptidomimetics. As discussed in Chapter 3,
98
hydrogenalion of unsaturated indolizidinone amino acid only afforded one stereoisomer
due to the steric effect. Therefore, other methods to reduce the double bonds will be
investigated (Scheme 4-14). Once other stereoisomers are available, they will be
incorporated into the peptidomimetics and provide us with more valuable information.
Scheme 4-14. Future work: reduction of double bonds.
CbzHN CbzHN COOMe COOMe
HN
HN AA -AA
4.5 Conclusion
4,8-Disubstituted azabicyclo[4.3.0]alkane amino acid analogues (26) were designed
to serve as peptidomimetic scaffolds of the core sequence in melanotropin peptides due to
their ability to provide constrained backbone and side-chain conformations. We have
developed efficient synthetic methodologies to introduce side-chain functionalities at the
C-4 and C-8 positions and also demonstrated the synthesis of analogues 49-51 and 53.
99
In our approach, the pyroglutamates with appropriate P-substituents were prepared
stereoselectively via asymmetric Michael additions between a Ni(II) complex and a chiral
oxazolidin-2-one derivative. The aliylation of the p-substituted pyroglutamates
followed by osmylation and Horner-Emmons olefination gave the dehydroamino acid
ester intermediates in good yield. The aryl side-chain at the C-4 position was
introduced via bromination of the dehydroamino acid ester and Suzuki cross-coupling.
The cyclization afforded bicyclic lactam structures which were subsequently
functionalized and coupled with tryptamine or phenethylamine to provide the target
analogues. The biological activities of these analogues are under investigation.
4.6 Experimental Section
3-(A'-Phthaiimido)propionaidehyde (28). Commercial grade pyridinium
chlorochromate (PCC, 21 g, 97.4 mmol) was ground with silica gel (1 wt eq) in a mortar.
The resulting free-running light orange solid was suspended in CH2CI2 (200 mL) at rt and
iV-(3-hydroxypropyl)phthalimide (10 g, 48.7 mmol) was added in one portion. The
resulting brown suspension was stirred for 2 h and filtered through a Buchner funnel
packed with Celite, and the granular brown residue was washed with ether (80 mL).
The filtrate was concentrated, purified by flash chromatography (hexanes:EtOAc = 1:1),
and recrystallized (Hexanes/EtOAc) to give 28 as white crystals (B.Ig, 82%). 28. 'H
NMR (500 MHz, CDCI3) S 9.83 (t, IH, /= 1.3 Hz), 7.86-7.84 (m, 2H), 7.74-7.72 (m, 2H),
4.04 (t, 2H, J = 7.0), 2.90-2.87 (m, 2H); '^C NMR (125 MHz, CDCI3) 8 199.4, 168.0,
100
134.1,131.9, 123.3,42.3,31.7.
{E)-5-(A'-PhthaIimido)-pent-2-enoic acid tert-butyl ester (29). To a solution of 28
(500 mg, 2.46 mmol) in CH2CI2 (20 mL) and H2O (10 mL) was added
(terf-butoxycarbonylmethyl)triphenylphosphonium bromide (1.13 g, 2.47 mmol), NaOH
(200 mg, 4.92 mmol) and EtsN (1 mL, 7.38 mmol) at rt. After stirring at rt for 1 h, the
organic phase was separated and washed with IN HCl (30 mL) and saturated aqueous
NaHCOs (30 mL). The organic phase was dried over Na2S04, filtered, and concentrated
to give a crude product, which was purified by flash column chromatography
(hexanes:EtOAc = 3;1) to afford pure product 29 as a white solid (704 mg, 95%). 29.
mp 91-93 °C; 'H NMR (500 MHz, CDCI3) 5 7.85-7.84 (m, 2H), 7.74-7.71 (m, 2H), 6.81
(dt, IH, J = 15.6, 7.1 Hz), 5.82 (dt, IE, / = 15.6, 1.5 Hz), 3.81 (t, 2H, J = 7.3 Hz),
2.60-2.55 (m, 2H), 1.46 (s, 9H); NMR (125 MHz, CDCI3) 5 168.0, 165.3, 142.6,
134.0, 132.0, 125.5, 123.3, 80.3, 36.4, 31.0, 28.1; HRMS (FAB) calcd for C17H20NO4
(M+H) 302.1392, found 302.1405.
(4S,2E)-3-[5-(iV-Phthalimido)-l-oxo-2-penteiiyI]-4-phenyl-2-oxazolidinone (3§).
A solution of 29 (2 g, 6.64 mmol) in 50% TEA in dichloromcthane (15 mL) was stirred at
rt for 2 h. The solution was concentrated, neutralized by saturated aqueous NaHCOs
and extracted with EtOAc (3 x 30 mL). The organic layers were combined, dried over
Na2S04 and evaporated in vacuo to give a crude product. To a solution of the crude
product in dry THE (20 mL) were added triethylamine (970 fiL, 6.96 mmol) and
trimethylacetyl chloride (900 ^iL, 7.31 mmol) at -78 "C. The mixture was stirred at 0 "C
for 1 h and recooled to -78 °C. Meanwhile, the chiral auxiliary.
101
(S)-4-phenyl-2-oxazolidinone (1.08 g, 6.62 mmol), in THF (40 niL) at -78 "C was treated
with the dropwise addition of nBuLi (1.6 M in hexanes, 4.15 mL, 6.64 mmol). This
reaction was stirred at -78 °C for 25 min. The preformed mixed anhydride was
cannulated into the lithiated chiral auxiliary solution at -78 °C and stirred in an ice bath
afterward, allowing the reaction mixture to achieve rt overnight. Water (100 mL) and
diethyl ether (50 mL) were added and the organic phase was separated, washed with
water (50 mL) and saturated aqueous NaCl (80 mL), dried over Na2S04 and concentrated
to give a crude product, which was purified by flash column chromatography
(hexanes:EtOAc = 1:1) to afford pure product 30 as a white solid (2.31 g, 89%). 30.
mp 139-141 °C; +65.4 (c 1.14, CHCI3); 'H NMR (500 MHz, CDCI3) 8 7.84-7.82
(m, 2H), 7.73-7.70 (m, 2H), 7.39-7.26 (m, 6H), 7.01 (dt, IH, J = 15.4, 7.1 Hz), 5.46 (dd,
IH, J = 8.7, 3.9 Hz), 4.68 (t, IH, J = 8.8 Hz), 4.26 (dd, IH, J = 8.9, 3.9 Hz), 3.84-3.82 (m,
2H), 2.68-2.64 (m, 2H); NMR (125 MHz, CDCI3) S 168.0, 164.0, 153.5, 146.4, 138.9,
134.0, 132.0, 129.2, 128.7, 125.9, 123.3, 122.5, 69.9, 57.7, 36.3, 31.6; HRMS (FAB)
calcd for C22H19N2O5 (M+H) 391.1294, found 391.1289.
A^-(A^-Benzylprolyl)-2-aminobenzophenone (31). To a clear solution of (S)-proline
(173 g, 1.5 mol) and potassium hydroxide (252.5 g, 4.5 mol) in 1 L of ?-PrOH at 40 "C
was added benzyl chloride (207.4 mL. 1.8 mol) dropwise in 3 h. The mixture was
stirred at 40 "C overnight, acidified to pH 5-6 with concentrated hydrochloric acid, mixed
with 200 mL of chloroform, and allowed to stand overnight. The white precipitate (KCl)
was filtered and washed with chloroform thoroughly. The filtrate was evaporated in
vacuo to give crude product that was dispersed in acetone and allowed to stand overnight.
102
Pure product was obtained after liltration and washing with a small amount of cold
acetone and dried in vacuum. To a solution of the above product (50 g, 0.207 mol),
2-aminobenzophone (40.8 g, 0.207 mol) and BOP (91.5 g, 0.207 mol) in 300 mL of
freshly distilled dichloromethane was added triethylamine (87 mL, 0.621 mol). The
mixture was stirred at it until the 2-aminobenzophone completely disappeared. The
reaction was quenched by adding IN hydrochloric acid with strong agitation. The
aqueous layer was separated from the organic layer and washed with methylene chloride
(2 X 100 mL). The combined methylene chloride layer was dried over anhydrous
MgS04 and evaporated in vacuo to afford the crude product which was used in the next
step.
(S)-NiGlyBPB (32). The solution of (S)-BPB generated above, NiCl2»6H20 (2 eq),
potassium hydroxide (7 eq) and glycine (5 eq) in methanol was stirred at 40 °C until the
starting ligand (5)-BPB was completely consumed as monitored by TLC (1:1
acetone/hexanes). The mixture was poured into ice water (four times the volume of
methanol) with 1% acetic acid. The precipitate was filtered, washed with water, and
dried in vacuum to afford the crude product. The crude product was dissolved in a large
quantity of chloroform and the precipitate was filtered and washed with chloroform.
The filtrate was evaporated in vacuo and dried under vacuum to afford the product.
Further recrystallization with acetone/hexanes gave the pure product 32 as a red powder
(yield: 75%). 32. mp 219-221 °C; [af\ +2117 (c 0.0145, CHCI3); 'll NMR (500 MHz,
CDCI3) 6 8.29 (d, IH, /= 8.5 Hz). 8.07 (d, 2H, J = 8.0 Hz), 7.49-7.53 (m, 3H), 7.41-7.44
(m, 2H). 7.30-7.33 (m, IH), 7.19-7.23 (m, IH), 7.10 (d, IE, J = 7.0 Hz), 6.97-6.99 (m,
103
IH), 6.80 (d, m,J= 8.5 Hz). 6.69-6.72 (m, IH), 4.48 (d, IH, J = 12.5 Hz), 3.66-3.80 (m,
4H), 3.47 (dd, IH, /= 11.0, 5.5 Hz), 3.32-3.38 (m, IH), 2.56-2.59 (m, IH), 2.40-2.47 (m,
IH), 2.13-2.18 (m, IH), 2.06-2.10 (m, IH); "C NMR (125 MHz, CDCI3) 181.3, 177.3,
171.6, 142.5, 134.6, 133.3, 133.2, 132.1, 129.8, 129.7, 129.6, 129.3, 129.1, 128.9, 126.2,
125.7, 125.1, 124.2, 120.8, 69.9, 63.1, 61.3, 57.5, 30.7, 23.7. HRMS (FAB) calcd for
C27H25N3Ni03 (M-i-H) 498.1328, found 498.1328.
Ni(II)-Coiiiplex of the ScMff base of (S)-BPB with (28- and i?,3S)-3-[2-(Af-
phthalimido)-ethyl]-5-[(4S)-3-(4-phenyl-2-oxazolidinonyl)] glutamic acids (33ab).
To a suspension of complex (S)-32 (1.2 g, 2.4 mmol) in DMF (8 mL), complex 30 (1 g,
2.56 mmol) was added with stirring. The mixture was stirred at rt for 10 min to get a
homogeneous solution and DBU (35 jxL, 0.24 mmol) was added dropwise. After 6 min,
the reaction was quenched with 5% aqueous acetic acid (10 mL) and the product was
extracted with dichloromethane (2 x 20 mL). The combined organic phase was dried
over Na2S04 and concentrated to give a crude product. Diastereomerical 1 y pure 33a
was recrystallized from ethyl acetate/hcxanes as a red solid (1.82 g, 85%).
Diastereomerically pure 33b was obtained by flash column chromatography
(DCM:acetone = 2:1) as a red oil (200 mg, 9%). 33a. mp 186-188 °C; [a]~^D +1522 (c
0.05, CHCI3); NMR (500 MHz, CDCI3) S 8.30 (dd, IH, / = 8.7, 0.8 Hz), 8.02 (d, 2H,
J= 7.1 Hz), 7.69-7.65 (m, 2H), 7.59-7.55 (m, 2H), 7.46-7.43 (m, IH), 7.40-7.37 (m, 2H),
7.35-7.22 (m, 6H), 7.16-7.04 (m, 4H), 6.66 (d, IH, J = 7.7 Hz), 6.56-6.47 (m, 2H), 5.19
(dd, IH, .7=8.5, 3.2 Hz), 4.44 (t, IH, /= 8.6 Hz), 4.37 (d, IH, J= 12.7 Hz). 4.12 (dd, IH,
J = 8.7, 3.2 Hz), 4.07 (d, IH, J = 6.2 Hz), 4.00-3.95 (m, IH), 3.77-3.67 (m, 2H),
104
3.50-3.42 (m, 4H), 3.17 (dd, IH, J = 17.9, 8.3 Hz), 2.96-2.85 (m, 2H), 2.47-2.38 (m 2H),
2.07-2.03 (m, 3H); '"'C NMR (125 MHz, CDCI3) 5 180.3, 177.4, 172.3, 170.2, 167.9,
153.5, 142.8, 139.3, 133.8, 133.7, 133.6, 133.5, 132.3, 131.8, 131.4, 129.5, 129.1, 129.0,
128.9, 128.7, 128.6, 128.3, 128.0, 127.0, 125.9, 125.6, 123.1, 123.0, 120.2, 72.0, 70.5,
70.0, 63.3, 57.5, 57.1, 36.7, 36.0, 35.9, 30.3, 29.7, 23.3; HRMS (FAB) calcd for
C49H44N5Ni08 (M+H) 888.2543, found 888.2553. 33b. [a]-\j +1068.6 (c 0.07, CHCI3);
^H NMR (500 MHz, CDCls) S 8.55 (d, IH, / = 9.0 Hz), 7.65-7.62 (m, 2H), 7.55-7.49 (m,
4H), 7.44-7.38 (m, 6H), 7.37-7.30 (m, 2H), 7.27-7.25 (m, 2H), 7.18-7.14 (m, 2H), 7.11 (d,
IE, J = lA Hz), 6.75 (d, IH, J = lA Hz), 6.56 (d, 2H, J = 4.0 Hz), 5.28-5.25 (m, 2H),
4.61 (t, IH, /= 8.7 Hz), 4.36-4.29 (m, IH), 4.19 (dd, IH, 7=8.7, 3.4 Hz), 4.15-4.10 (m,
2H), 3.92-3.76 (m, 4H), 3.21 (dd, IH, J = 17.4, 8.0 Hz), 2.94 (dd, IH, J = 17.4, 5.9 Hz),
2.66-2.60 (m, IH), 2.37-2.29 (m, 2H), 2.06-2.03 (m, IH), 1.92-1.86 (m, IH), 1.67-1.57
(m, 2H); NMR (125 MHz, CDCI3) 5 182.5, 177.7, 172.7, 170.2, 168.3, 153.4, 143.1,
139.3, 133.9, 133.8, 133.7, 132.4, 132.3, 131.9, 131.6, 129.5, 129.1, 128.9, 128.8, 128.7,
128.4, 128.2, 126.8, 126.0, 125.5, 123.5, 123.2, 120.4, 72.0, 69.9, 68.4, 60.6, 57.6, 55.8,
37.3, 36.7, 36.6, 31.2, 29.6, 23.6; HRMS (FAB) calcd for C49H44N5Ni08 (M+H)
888.2543, found 888.2543.
Methyl (25,35)wV®'-fifrt-butoxycarbonyl-3-[2-(A'-phthaIimido)-ethyl]pyroglu-
tamate (34). Diastereomerically pure complex 33a (7 g, 7.88 mmol) was dissolved in
melhanol (30 mL) and dichloromethane (10 mL) and added to a 1/1 mixture (50 mL) of 3
N HCI and water dropwise at 70 °C. After decomposition of the complex was
completed (disappearance of the orange color), the mixture was evaporated in vacuo.
105
treated with conc. ammonia and washed with CHCI3. The aqueous phase was stirred for
2 h before the solvent was evaporated in vacuo. The resultant crude product was
dissolved in methanol (60 mL), and thionyl chloride (4.5 mL, 61.8 mmol) was added
drop wise at 0 °C. The mixture was allowed to come to rt and stirred overnight. After
evaporation, the residue was dissolved in CH2CI2 (50 mL), washed with saturated
aqueous NaHCOs solution (50 mL) and brine (50 mL), and dried over Na2S04. After
filtration and rotary evaporation, the crude ester was dissolved in CH3CN (15 mL) and
di-ferr-butyl dicarbonate (1.7 g, 7.79 mmol) and DMAP (50 mg, 0.41 mmol) were added.
The mixture was stirred for 5 h. After removal of the solvent, the residue was purified
by column chromatography on silica gel (hexanes:EtOAc = 3:2) to give the pure product
34 as a colorless oil(1.77 g, 54%). 34. [a]^^D +26.4 (c 0.99, CHCI3); 'H NMR (500
MHz, CDCI3) 5 7.87-7.84 (m, 2H), 7.76-7.74 (m, 2H), 4.32 (d, IH, J = 3.8 Hz), 3.80-3.75
(m, 5H), 2.86 (dd, IH, J = 17.6, 8.8 Hz), 2.36 (dd, IH, / = 17.6, 4.6 Hz), 2.31-2.27 (m,
IH), 2.09-2.05 (m, IH), 1.88-1.84 (m, IH), 1.49 (s, 9H); NMR (125 MHz, CDCI3) 5
172.0. 171.1. 168.1, 149.1, 134.1, 131.9, 123.3, 83.8, 64.2, 52.6, 37.3, 35.4, 33.5, 32.5,
27.8; HRMS (FAB) calcd for C21H25N2O7 (M+H) 417.1662, found 417.1675.
Methyl (2.S',3S,55)-1 -(tert-biityloxycarbonyl)-3-[2-(iV-phthalimido)-ethyl]-5-aIlyI-
prolinate (35). To a solution of 34 (220 mg, 0.53 mmol) in dry THF (5 mL) was added
DIBAL-H (IM solution in hexanes, 2.1 mL) at -78 °C under Ar. After stirring at -78 °C
for 15 min, the reaction was quenched with saturated aqueous NH4CI (3 mL) and allowed
to come to rt. The solution was passed through a short column of Celite and dried over
Na2S04. After removal of the solvent, the residue was dissolved in methanol (4 mL)
106
and /7-Ts0H«H20 (10 mg, 0.05 mmol) was added. After stirring for 3 h, the solution
was quenched with saturated NaHCOs (4 mL), the solvent was removed, the mixture was
extracted with diethyl ether (3 x 8 mL), and the combined organic extracts were dried
over Na2S04, filtered and concentrated under vacuum. The residue was dissolved in
diethyl ether (4 mL) and allyltrimethylsilane (335 p-L, 2.1 mmol) and BFs'EtiO (75 |iL,
0.59 mmol) were added at -40 °C under Ar. The cold bath was removed after stirring at
-40 "C for 15 min. The reaction mixture was stirred for additional 40 min, then
quenched with NaHCOs (2 mL) and extracted with diethyl ether (3x5 mL). The
organic extracts were combined, dried over Na2S04, filtered and concentrated under
vacuum to afford the crude product. Purification by flash column chromatography
(hexanes;EtOAc = 3:1) afforded pure product 35 as a colorless oil (150 mg, 64%). 35.
[a]-^, +24.8 (c 1.84, CHCI3); 'H NMR (500 MHz, CDCI3) (two rotamers) 5 7.86-7.83 (m,
2H), 7 .74-7.71 (m, 2H), 5 .84-5.76 (m, IH), 5 .10 (dd, IH, J= 17.1, 1 .6 Hz) , 5 .03 (d, IH, J
= 10.1 Hz), 4.00 (brs, 0.6H), 3.94-3.90 (m, 0.9H), 3.84 (d, 0.5H, J = 8.6 Hz), 3.76-3.65
(m, 5H), 2.68-2.66 (m, 0.6H), 2.57-2.54 (m, 0.4H), 2.34-2.30 (m, IH), 2.23-2.16 (m, IH),
2.11-2.05 (m, 2H), 1.74-1.66 (m, 2H), 1.46 (s, 4H), 1.40 (s, 5H); '"'C NMR (125 MHz,
CDCI3) (two rotamers) 8 173.3, 173.1, 168.1, 154.0, 153.3, 135.2, 134.0, 132.0, 123.2,
117.1, 80.1, 65.6, 65.1, 57.8, 57.6, 52.2, 52.0, 40.2, 39.3, 39.1, 38.3, 36.2, 35.4, 34.5, 32.2,
32.1, 28.4, 28.2; HRMS (FAB) calcd for C24H31N2O6 (M+H) 443.2182, found 443.2177.
Methyl (2S,3S,5R)-1 -(tert-butyloxycarbonyl)-3- [2-(7V-phtlialimido)-ethy 1]-5-[(3-
amino-(iV-benzyIoxycarbonyl)-3-methoxycarbonyI)-2-propenyl]-proiinate (36). To a
solution of 35 (100 mg, 0.23 mmol) in THF (6 mL) and H2O (3 mL) was added OSO4
107
(cat.) in the dark. After 5 min, NaI04 (120 mg, 0.56 mmol) was added. The mixture
was stirred at rt for 4 h, filtered, and washed with MeOH (3x5 mL). After removal of
the solvent, the residue was redissolved in dichloromethane (10 mL). The organic
solution was washed with brine (3 x 10 mL), dried over Na2S04, filtered and
concentrated under vacuum to give crude aldehyde. Meanwhile, to a solution of
(Me0)2P(0)CH(NHCbz)C02Me (75 mg, 0.23 mmol) in dichloromethane (3 mL) was
added DBU (34 jiL, 0.23 mmol) at rt. After 10 min, to the above solution was added a
solution of the crude aldehyde in dichloromethane (2 mL) and the reaction mixture was
stirred overnight at rt. After removal of the solvent, the residue was redissolved in
EtOAc (5 mL), washed with IN HCl (5 mL) and brine (5 mL), dried over NajSOa,
filtered and concentrated under vacuum. The crude product was purified by flash
column chromatography (hexanes.EtOAc = 1:1) to give product 36 as a colorless oil (117
mg, 79%). 36. [a]'\, +49.7 (c 1.84, CHCI3); 'H NMR (500 MHz, CDCI3) (two
rotamers) 5 7.83 (brs, 2H), 7.67 (brs, 2H), 7.58-7.29 (m, 5H), 6.65 (brs, IH), 5.19-5.12
(m, 2H), 4.17-4.10 (m, IH), 3.91 (d, 0.5H, 7=9.5 Hz), 3.80-3.65 (m, 8.5H), 2.70-2.61 (m,
IH), 2.45-2.42 (m, 0.5H), 2.36-2.33 (m, 1.5H), 2.18-2.05 (m, 2H), 1.83-1.65 (m, 2H),
1.39 (s, 4H), 1.35 (s, 5H); "C NMR (125 MHz, CDCI3) (two rotamers) 8 173.8, 173.6,
168.3, 165.1, 154.9, 154.8, 153.9, 153.4, 136.3, 134.0, 133.9, 133.4, 131.8, 128.3, 128.2,
127.9, 123.3, 80.6, 80.5, 66.9, 65.5, 65.3, 57.2, 57.0, 52.6, 52.3, 52.0, 40.4, 39.5, 37.0,
36.1, 36.0, 35.8, 34.2, 33.5, 32.2, 32.0, 28.1, 28.0; HRMS (FAB) calcd for C34H40N3O10
(M+H) 650.2714, found 650.2703.
Methyl (25,35,55)-l-(/e/t-butyIoxycarbonyl)-3-[2-(iV-phthaliniido)-ethyI]-5-
108
[(3R)-(3-atnino-(A'-benzyloxycarbonyl)-3-methoxycarbonyl)-propyI]-prolinate (37a).
A hydrogenation bottle was charged with 36 (90 mg, 0.14 mmol) in degassed
methanol (10 mL, HPLC grade) and then purged with argon for 15 min, followed by
adding [(R,R)-(COD)-Et-DuPHOS Rh(I)]OTf (10 mole%). After five vacuum/hydrogen
cycles, the reaction bottle was pressurized to an initial pressure of 75 psi. The reaction
proceeded for 24 h. After evaporation of solvent, the crude product was purified by
flash column chromatography (hexanes:EtOAc = 1:1) to afford pure product as a
colorless oil (84 mg, 92%). 37a. [af'o +24.1 (c 1.71, CHCI3); 'H NMR (500 MHz,
CDCI3) (two rotamers) 5 7.95-7.63 (m, 4H), 7.36-7.28 (m, 5H), 5.89 (d, 0.6H, J= 7.8 Hz),
5.59 (d, 0.4H, J = 7.8 Hz), 5.15-5.10 (m, 2H), 4.35 (brs, IH), 4.01-3.81 (m, 2H),
3.77-3.65 (m, 8H), 2.29 (brs, IH), 2,11-2.02 (m, 2H), 1.96-1.63 (m, 5H), 1.49-1.47 (m,
IH), 1.43 (s, 4H), 1.39 (s, 5H); "C NMR (125 MHz, CDCI3) (two rotamers) § 173.2,
173.1, 172.9, 172.7, 168.3, 156.2, 156.0, 153.9, 153.6, 136.4, 136.3, 134.0, 131.8, 128.4,
128.0, 127.9, 123.5, 123.3, 80.2, 66.8, 66.7, 65.3, 64.8, 57.6, 57.3, 54.2, 53.8, 52.2, 52.0,
40.2, 39.4, 36.1, 35.5, 32.3, 32.1, 30.6, 30.3, 29.1, 28.7, 28.3, 28.1; HRMS (FAB) calcd
for C34H42N3O10 (M+H) 652.2870, found 652.2884.
Methyl (2S,3S,55)-l-(/e/t-butyloxycarbonyl)-3-[2-C\'-phthalimido)-ethyI]-5-
[(3S)-(3-aniino-(A^-benzyloxycarbonyl)-3-methoxycarbonyI)-propyl]-prolinate (37b).
In a manner similar to the preparation of 37a, using [(5,5)-(COD)Et-DuPHOS
Rh(I)]OTf as a catalyst gave 37b in 94% yield. 37b. [a]^^D +20.56 (c 1.49, CHCI3);
NMR (500 MHz, CDCI3) (two rotamers) 6 7.84-7.82 (m, 2H), 7.70 (brs, 2H), 7.37-7.28
(m, 5H), 5.56-5.50 (m, IH), 5.12 (s, 2H), 4.38 (brs, IH), 4.01-3.84 (m, 2H), 3.75-3.65 (m.
109
8H), 2.31 (brs, IH), 2.12-J.90 (m, 3H), 1.79-1.71 (m, 4H), 1.53-1.47 (m, IH), 1.44 (s,
4H), 1.39 (s, 5H); '^C NMR (125 MHz, CDCI3) (two rotamers) 5 173.3, 173.2, 172.9,
172.8, 168.1, 156.0, 154.0, 153.5, 136.4, 136.3, 134.0, 131.9, 128.4, 128.0, 127.9, 123.3,
80.3, 80.2, 66.7, 65.3, 64.8, 57.5, 57.2, 53.8, 53.7, 52.2, 52.0, 40.3, 39.5, 36.8, 36.2, 36.0,
32.6, 32.3, 30.6, 29.3, 29.1, 28.3, 28.2; HRMS (FAB) calcd for C34H42N3O10 (M-^H)
652.2870, found 652.2877.
i3R, 6S, 8S, 9S)-Methyl 2-oxo-3-A'-(benzoxycarbonyl)amino-8-[2-(iV-phthalimido)
-ethyl]-l-azabicyco[4.3.0]nonane-9-carboxylate (38). To a solution of 37a (80 nig,
0.12 mmol) in dichloromethane (2.4 mL) was added TFA (0.6 mL) at rt. The mixture
was stirred for 30 min, then saturated NaHCOa (10 mL) was added. After stirring for 20
min, the organic solution was dried over Na2S04 and concentrated under vacuum. The
residue was redissolved in chloroform (5 mL) and stirred at rt for 48 h. After removal
of the solvent, the crude product was chromatographed (hexanes:EtOAc = 1:3) to afford
38 as a colorless oil (50 mg, 80%). 38. [a]^^D +20.2 (c 1.16, CHCI3); 'H NMR (500 MHz,
CDCI3) 5 7.87-7.84 (m, 2H), 7.75-7.71 (m, 2H), 7.36-7.26 (m, 5H), 5.42 (brs, IH),
5.12-5.06 (m, 2H), 4.17 (s, IH), 4.08-4.05 (m, IE), 3.82-3.69 (m, 6H), 2.55 (brs, IH),
2.29-2.25 (m, IH), 2.14 (d, IH, / = 11.5 Hz), 2.05-1.87 (m, 3H), 1.86-1.75 (m, 2H),
1.74-1.64 (m, IH); "C NMR (125 MHz, CDCI3) 5 171.4, 168.2, 168.1, 156.4, 136.3,
134.1, 132.0, 128.5,128.1, 123.3, 66.8, 64.1, 58.2, 52.5, 52.4, 39.2, 36.4, 36.0. 32.9, 28.7,
27.7; HRMS (FAB) calcd for C28H30N3O7 (M+H) 520.2084, found 520.2077.
(3S,6S,8S,9S)-Methyl 2-oxo-3-yV-(benzoxycarbonyl)amino-8-[2-(A'-phthalinii(io)-
ethyI]-l-azabicyco[4.3.0]nonane-9-carboxylate (39). In a manner similar to the
110
23 preparation of 38, using 37b as starting material gave 39 in 92% yield. 39. [a] o
+24.1 (c 2.25, CHCI3); NMR (500 MHz. CDCI3) 5 7.87-7.84 (ra, 2H), 7.75-7.72 (m,
2H), 7.35-7.26 (m, 5H), 5.81 (d, IH, J= 5.0 Hz), 5.11 (s, 2H), 4.27 (s, IH), 4.24-4.20 (m,
IH), 3.86-3.82 (m, IH), 3.80-3.73 (m, 2H), 3.71 (s, 3H), 2.55-2.51 (m, IH), 2.34 (dd, IH,
J= 14.6, 7.3 Hz), 2.18 (dd, IH, /= 13.0, 6.3 Hz), 2.12-2.09 (m, IH), 1.93-1.87 (m, 2H),
1.78-1.65 (m, 3H); NMR (125 MHz, CDCI3) § 171.4, 169.1, 168.1, 156.0, 136.5,
134.1, 132.0, 128.4, 128.0, 127.9, 123.3, 66.7, 63.8, 54.6, 52.4, 50.3, 39.9, 36.5, 35.9,
32.4, 26.9; HRMS (FAB) calcd for C28H30N3O7 (M+H) 520.2077, found 520.2077.
Methyl (2S,35',5i?)-l-(te?f-butyloxycarbonyl)-3-[2-(iV-phthalimido)-ethyl]-5-[2-
bromo-(3-amino-(A'-benzyloxycarbonyl)-3-methoxycarbonyl)-2-propenyl]prolinate
(40). To a solution of 36 (405 mg, 0.62 mmol) in chloroform (10 mL) was added NBS
(122 mg, 0.69 mmol) at rt. After 80 min, Dabco (140 mg, 1.25 mmol) was added. The
mixture was stirred at rt for 24 h, washed with saturated NH4CI (3 x 10 mL), and dried
over Na2S04. After removal of the solvent, the residue was subjected to flash column
chromatography (hexanes:EtOAc = 4:1) to afford 40 in 78% yield as a colorless oil. 40.
+10.4 (c 1.05 , CHCI3); 'H NMR (500 MHz, CDCI3) (Rotamer) S 7.85-7.82 (m,
2H), 7.72-7.69 (m, 2H), 7.36-7.27 (m, 5H), 6.61 (brs, IH), 5.14-5.11 (m, 2H), 4.38-4.35
(m, 0.6H), 4.34-4.28 (m, 0.4H), 3.95-3.84 (m, 1.5H), 3.83-3.62 (m, 7.5H), 3.12-3.10 (m,
1.5H), 2.91-2.87 (m, 0.5H), 2.44-2.41 (m, IH), 2.24 (brs, IH), 2.10-2.03 (m, IH),
1.72-1.66 (m, 2H), 1.47-1.26 (m, 9H); "C NMR (125 MHz, CDCI3) (rotamers) 5 173.3.
173.1, 168.1, 162.7, 162.5, 153.5, 153.0, 152.9, 152.8, 135.4, 134.1, 134.0, 133.9, 132.1,
132.0, 128.6, 128.5, 128.4, 128.3, 128.2, 123.4, 123.2, 123.1, 80.5, 80.4, 67.9, 67.7, 65.5,
I l l
65.0, 57.0, 56.9, 52.7, 52.3, 52.1, 40.3, 39.8, 39.5, 39.2, 36.2, 36.1, 34.4, 33.9, 32.1, 32.0,
28.4, 28.3; HRMS (FAB) calcd for C34H39BrN30,o (M+H) 728.1819, found 728.1804.
Methyl (2S,3S,5/?)-l-(^t;/t-butyloxycarbonyl)-3-[2-C\'-phthalimido)-ethyl]-5-[2-
phenyl- and 2-naphthyl-(3-ainino-(A'-beiizyioxycarbonyl)-3-methoxycarbonyl)-2-
propenyl]-prolinate (41 and 42). To a solution of 40 (375 rag, 0.52 mmol) in DME
(3 mL) and degassed water (520 |xL) were added phenylboronic acid (185 mg, 1.52
mmol), Na2C03 (165 mg, 1.56 mmol), Pd(0Ac)2 (25 mg, 0.11 mmol) and P(o-tolyl)3 (30
mg, 0.10 mmol). The mixture was stirred at 80 "C overnight and passed through a short
column containing a bottom 1" layer of silica gel and a top 1" layer of NaHC03 using
ethyl acetate as eluent. After removal of the solvent, the crude 41 was subjected to the
next step without further purification. The identical procedure starting with 40 afforded
42 as crude product for the next step.
(6/?,8S,9S)-l-Aza-3-A^-benzoxycarbonylamino-9-methoxycarbonyl-4-phenyl-
and 2-naphthyl-8-[2-(iV-phthallmido)-ethyl]-2-oxobicyclo[4.3.0]non-3-ene (43 and
44). To a solution of above crude product 41 in dichloromethane (2.4 mL) was added
TFA (0.6 mL) at rt. The mixture was stirred for 30 min, then saturated NaHC03 (10 mL)
was added. After stirring for 20 min, the organic solution was dried over Na2S04 and
concentrated under vacuum. The residue was redissolvcd in chloroform (5 mL) and
stirred at rt for 48 h. After removal of the solvent, the crude product was
chromatographed (hexanesiEtOAc = 1:1) to afford 43 as a colorless oil in 61% overall
yield from 40. The identical procedure starting with 42 afforded 44 in 63% overall yield
from 40.43. [af\^ -5.7 (c 1.04, CHCI3); 'H NMR (500 MHz, CDC13) 5 7.88-7.85 (m,
112
2H), 7.75-7.72 (m, 2H), 7.37-7.33 (m, 4H), 7.31-7.23 (m, 4H), 7.20-7.19 (m, 2H), 6.51
(brs, IH), 4.97 (d, IH, J = 12.3 Hz), 4.89 (d, IH, i= 12.3 Hz), 4.36 (s, IH), 4.22-4.15 (m,
IH), 3.84-3.72 (m, 5H), 2.90-2.80 (m, 2H), 2.40 (q, IH, J = 7.0 Hz), 2.19 (dd, IH, / =
12.8, 5.7 Hz), 2.07-2.02 (m, IH), 2.01-1.92 (m, IH), 1.83-1.76 (m, IH); 'T. NMR (125
MHz, CDCI3) 5 171.7, 168.2, 161.8, 153.4, 138.2, 137.9, 136.2, 134.1, 132.0, 128.5,
128.4, 128.3, 127.9, 127.2, 124.2, 123.4, 67.0, 63.4, 54.0, 52.6, 39.7, 36.4, 36.0, 35.8,
32.7; HRMS (FAB) calcd for C34H32N3O7 (M-t-H) 594.2240. found 594.2222. 44.
[a]^^D -8.8 (c 2.12, CHCI3); NMR (500 MHz, CDCO) 5 7.90-7.72 (m, 8H), 7.52-7.46
(m, 3H), 7.22-7.16 (m, 3H), 7.09-7.07 (m, 2H), 6.65 (s, IH), 4.94-4.81 (m, 2H), 4.39 (s,
IH), 4.27-4.22 (m, IH), 3.85-3.73 (m, 5H), 3.04-2.99 (m, IH), 2.93-2.87 (m, IH), 2.43
(dd, IH, J = 14.6, 7.3 Hz), 2.24-2.20 (m, IH), 2.10-2.04 (m, IH), 2.00-1.93 (m, IH),
1.85-1.80 (m, IH); NMR (125 MHz, CDCI3) 8 171.6, 168.2, 161.8, 153.3, 137.5,
136.1, 135.7, 134.1, 133.1, 132.9, 132.0, 128.5, 128.4, 128.3, 128.1, 127.9, 127.6, 126.6,
126.5, 126.3, 124.9, 124.4, 123.3, 66.9, 63.4, 54.1, 52.6, 39.8, 36.4, 35.9, 35.8, 32.7;
HRMS (FAB) calcd for C38H34N3O7 (M+H) 644.2397, found 644.2402.
(6/?,85,95)-l-Aza-3-A^-benzoxycarbonylamino-9-methoxycarbonyI-4-phenyI-
and 2-naphthyl-8-[2-(A^^'-di-Boc-guanidino)-ethyI]-2-oxobicyclo[43.0]non-3-ene
(47 and 48). To a solution of 43 (31 mg, 0.052 mmol) in absolute ethanol (2 niL) and
chloroform (400 |nL) was added anhydrous hydrazine (11 fxL, 0.35 mmol) at rt. After
stirring at rt for 24 h, the solution was filtered to remove the white precipitate and the
filtrate was concentrated in vacuum to afford crude 45. The crude 45 was dissolved in
113
dichloromethane (2 niL) and TEA (45 fiL, 0.33 mmol) and N.N'-
bis{rt'?t-butoxycarbonyl)-A^"-lriflylguanidine (31 mg, 0.079 mmol) were added at rt.
The mixture was stirred at rt for 24 h before the solvent was removed in vacuo. The
crude product was chromatographed (hexanes:EtOAc = 1:1) to afford 47 as a colorless oil
(30.5 mg, 83%). 47. -7.8 (c 2.07, CHCI3); NMR (500 MHz, CDC13) 511.48
(brs, IH), 8.43 (brs, IH), 7.37-7.25 (m, 8H), 7.21-7.19 (m, 2H), 6.48 (brs, IH), 4.99 (d,
IH, J= 12.3 Hz), 4.91 (d, IH, /= 12.3 Hz), 4.36 (s, IH), 4.20-4.13 (m, IE), 3.78 (s, 3H(,
3.66-3.59 (m. IH), 3.52-3.47 (m, IH), 2.84 (d, 2H, J= 8.9 Hz), 2.43 (q, IH, /= 14.7, 7.4
Hz), 2.11-2.07 (m, IH), 2.05-1.99 (m, IH), 1.80-1.75 (m, 2H), 1.50 (s, 18H); NMR
(125 MHz, CDCI3) 8171.7, 163.5, 161.8, 156.2, 153.5, 153.3, 138.3, 138.2, 136.2, 128.5,
128.4, 128.3, 128.0, 127.9, 127.2, 124.3, 83.3, 79.4, 67.0, 62.8, 54.0, 52.6, 39.5, 38.6,
36.4, 36.3, 33.3, 28.3, 28.1, 27.8; HRMS (FAB) calcd for C37H48N5O9 (M-i-H) 706.3452,
found 706.3445. The identical procedure starting with 46 afforded 48 in 80% overall
yield. 48. [aj^'o -16.6 (c 1.14, CHCI3); 'H NMR (500 MHz, CDC13) 5 11.51 (s, IH),
8.44-8.43 (m, IH), 7.87-7.73 (m, 6H), 7.52-7.47 (m, 3H), 7.24-7.15 (m, 3H), 6.65 (brs,
IH), 4.95-4.85 (m, 2H), 4.39 (s, IH), 4.22 (brs, IH), 3.79 (s, 3H), 3.66-3.62 (m, IH),
3.52-3.48 (m, IH), 3.01-2.97 (m, IH), 2.92-2.87 (m, HI), 2.47-2.43 (m, IH), 2.13-2.11
(m, IH), 2.08-2.03 (m, IH). 1.80-1.75 (m, 2H), 1.51 (s, 9H), 1.50 (s, 9H); NMR (125
MHz, CDCI3) 8 171.7, 166.6, 163.5, 161.8, 156.2, 153.3, 137.7, 136.1, 135.7, 134.2,
133.1, 132.9, 130.2, 128.3, 128.1, 127.9, 127.6, 126.6, 126.5, 126.3, 124.9, 123.4, 83.2,
114
79.3, 66.9, 62.8, 54.1, 52.6, 39.6, 38.6, 36.4, 36.3, 33.4, 28.3, 28.0; HRMS (FAB) calcd
for C41H50N5O9 (M+H) 756.3609, found 756.3611.
(6/e,85,9S)-l-Aza-3 -A^-benzoxycarbonylamino-9-niethoxycarbonyl-4-2'-naphthyl
-8-[2-/e/t-butoxycarbonylaniino-ethyI]-2-oxobicyclo[4.3.0]non-3-ene (52). To a
solution of 44 (22 mg, 0.034 mmol) in absolute ethanol (2 mL) and chloroform (1 mL)
was added anhydrous hydrazine (8 jiL, 0.25 mmol). After stirring at rt for 24 h, the
solution was filtered to remove the white precipitate and the filtrate was concentrated in
vacuum and redissolvcd in dichloromethane (2 mL). To the solution were added TEA
(29 p,L, 0.21 mmol) and di-?e/t-butyl dicarbonate (15 mg, 0.069 mmol). The mixture
was stirred at rt for 6 h before the solvent was removed in vacuo. The crude product
was chromatographed (hexanes:EtOAc = 1:2) to afford 52 as a colorless oil (18 mg, 86%).
52. [aJ--^D-10.7 (c 1.11, CHCI3); 'H NMR (500 MHz, CDC13) 5 7.82-7.76 (m, 4H),
7.51-7.47 (m, 3H), 7.24-7.10 (m, 5H), 6.61 (brs, IH), 4.95-4.83 (m, 2H), 4.63 (brs, IH),
4.35 (s, IH), 4.24-4.19 (m, IH), 3.78 (s, 3H), 3.30-3.24 (m, 2H), 3.01-2.96 (m, IH),
2.92-2.86 (m, IH), 2.45-2.41 (m, IH), 2.12-2.02 (m, 2H), 1.70-1.66 (m, 2H), 1.46 (s, 9H);
NMR (125 MHz, CDCI3) 5 171.8, 161.9, 155.9, 153.4, 137.8, 136.1, 135.7, 133.1,
133.0, 128.4, 128.3, 128.2, 127.9, 127.7, 126.6, 126.5, 126.3, 124.9, 124.4, 79.5, 67.0,
63.1, 54.2, 52.6, 39.6, 38.6, 36.5, 36.2, 34.3, 28.4; HRMS (FAB) calcd for C35H40N3O7
(M-fH) 614.2866, found 614.2888.
General procedure of coupling reactions: The solution of starting material in Li OH
115
(IN) and methanol (1:2) was stirred at rt for 3 h. After the solution was neutralized by
HCl (IN), it was washed with dichloromethane. The organic phase was dried over
Na2S04 and concentrated in vacuo. To the solution of the residue in dichloromethane
was added PyBOP (1.1 eq), HOBt (1.1 eq), DIPEA (3 eq) and amine (2 eq). The
mixture was stirred at rt for 2 h before washing with NH4CI. The organic phase was
collected and dried over Na2S04. After the solvent was removed, the crude product was
purified by column chromatography (hexanes:EtOAc = 1:2).
(6/f,85,9iS')-l-Aza-3-iV-benzoxycarbonylamino-9-[2-(indol-3-yl)-ethylcarbanioyl]-
4-phenyl-8-[2-(iV,A'''-di-Boc-guanidino)-ethyl]-2-oxo-bicyclo[4.3.0Jnon-3-ene (49).
Yield: 68%. 'H NMR (600 MHz, CDC13) 5 11.45 (s, IH), 8.72 (brs, IH), 8.33 (brs, IH),
7.60 (d, IH, J = 7.9 Hz), 7.37-7.27 (m, 9H), 7.24-7.08 (m, 4H), 7.00-6.99 (m, IH),
6.50-6.46 (m, 2H), 5.00-4.93 (m, 2H), 4.14 (s, IH), 4.01-3.98 (m, IH), 3.73-3.70 (m, IH),
3.50-3.43 (m, 2H), 3.35-3.30 (m, IH), 3.04-2.95 (m, 2H), 2.68-2.65 (m, IH), 2.52-2.47
(m, IH), 2.40-2.37 (m, IH), 1.94-1.88 (m, 2H), 1.65-1.55 (m, 2H), 1.49 (s, 9H), 1.48 (s,
9H); '^C NMR (125 MHz, CDCI3) 5 170.0, 163.4, 162.5, 156.3. 153.6, 153.2, 139.3,
138.4, 136.4, 136.1, 128.5, 128.4, 128.3, 128.1, 128.0, 127.1, 127.0, 124.1, 122.9, 121.9,
119.2, 118.7, 112.1, 111.3, 83.4, 79.6, 67.1, 64.7, 54.7, 38.9, 38.8, 36.3, 36.1, 33.1, 29.7,
28.3. 28.1, 24.9; HRMS (FAB) calcd for C46H56N7O8 (M-l-H) 834.4190, found
834.4205.
(6/?,8,S,95)-l-Aza-3-A'-benzoxycarbonylaniino-9-phenethylcarbamoyl-4-phenyl-
116
8-[2-(A^,A^'-di-Boc-guanidino)-ethyl]-2-oxo-bicyclo[4.3.0]non-3-ene (50). Yield: 73%.
'H NMR (600 MHz, CDC13) S 11.47 (s, IH), 8.38 (brs, IH), 7.38-7.15 (m, 15H), 6.99
(brs, IH), 6.50 (brs, IH), 5.02-4.89 (m, 2H), 4.30 (s, IH), 4.09-4.05 (m, IH). 3.68-3.63
(m, IH), 3.61 (m, IH), 3.48-3.38 (m, 2H), 2.82-2.77 (m, 3H), 2.71-2.66 (m, IH),
2.55-2.51 (m, IH), 2.04-1.99 (m, IH), 1.97-1.94 (m, IH), 1.66-1.60 (m, 2H), 1.50 (s, 9H),
1.49 (s, 9H); NMR (125 MHz, CDClg) S 170.3, 163.4, 162.6, 156.5, 153.5, 153.3,
139.1, 139.0. 138.1, 136.2, 128.8, 128.5, 128.4, 128.3, 128.2, 128.0, 127.9, 127.2, 126.3,
124.3, 83.4, 80.0. 67.0, 64.2, 54.7, 40.7, 38.7, 38.3, 36.3, 36.2, 35.6, 33.0, 28.3, 28.1;
HRMS (FAB) calcd for C44H55N6O8 (M+H) 795.4081, found 795.4069.
(6/?,8S,9S)-l-Aza-3-A'-benzoxycarbonylamino-9-[2-(indol-3-yl)-ethylcarbanioyl]-
4-(2-naphthyI)-8-[2-(A^'-di-Boc-guanidino)-ethyI]-2-oxo-bicyclo[4.3.0]noii-3-ene
(51). Yield: 63%. 'H NMR (600 MHz, CDC13) 5 11.45 (s, IH), 8.75 (brs, IH),
8.34-.32 (m, IH), 7.83-7.77 (m, 4H), 7.61-7.33 (m, 5H), 7.27-6.99 (m, 8H), 6.61-6.54 (m,
2H), 4.95-4.86 (m, 2H), 4.17 (s, IH), 4.07-4.02 (m, IH), 3.76-3.71 (m, IH), 3.51-3.45 (m,
2H), 3.36-3.31 (m, IH), 3.06-2.94 (m, 2H), 2.81-2.77 (m, IH), 2.57-2.50 (m, IH),
2.43-2.40 (m, IH). 1.94-1.91 (m. 2H), 1.66-1.54 (m, 2H), 1.49 (s, 9H), 1.48 (s, 9H); "C
NMR (125 MHz, CDCI3) 5 169.9, 163.4, 162.5, 156.2, 153.6, 153.2, 138.8, 136.4, 136.0,
135.9, 133.1, 132.9, 128.3, 128.1, 128.0, 127.9, 127.7, 127.1, 126.6, 126.4, 126.3, 124.6,
124.3, 122.9. 121.9, 119.2, 118.7, 112.1. 111.3, 83.4, 79.5. 67.1, 64.8, 54.7, 38.9, 38.8,
36.2, 36.1, 33.0, 28.3, 28.0, 24.9; HRMS (FAB) calcd for C50H58N7O8 (M-L-H) 884.4347,
117
found 884.4362.
(6JR,8S,9S)-l-Aza-3-A?-benzoxycarbonylamino-9-[2-(indol-3-yl)-ethylcarbamoyl]-
4-(2-naphthyl)-8-[2-re/t-butoxycarbonylamino-ethyl]-2-oxo-bicycIo[4.3.0]non-3-ene
(53). Yield; 69%. 'H NMR (500 MHz, CDC13) 5 8.40 (brs, IH), 7.84-7.78 (m, 4H).
7.61-7.46 (m, 4H), 7.34-7.04 (m, 8H), 6.99 (brs, IH), 6.88 (brs, IH), 6.62 (s, IH),
4.95-4.88 (m, 2H), 4.77 (brs, IH), 4.32 (s, IH), 4.08-4.03 (m, IH), 3.60-3.58 (m, 2H),
3.32-3.28 (m, IH), 3.20-3.13 (m, IH), 3.08-3.03 (m, IH), 2.97-2.91 (m, IH), 2.80-2.76
(m, IH), 2.56-2.50 (m, 2H), 1.90-1.84 (m, 2H), 1.54-1.51 (m, 2H), 1.42 (m, 9H);
NMR (125 MHz, CDCI3) 6 170.2, 162.7, 156.3, 153.7, 139.0, 136.4, 136.0, 135.9, 133.1,
133.0, 128.4, 128.1, 128.0, 127.9, 127.7, 127.2, 126.7, 126.4, 126.3, 124.6, 124.4, 122.8,
121.9, 119.2, 118.7, 112.3, 111.3, 79.6, 67.1, 64.2, 54.8, 39.1, 38.7, 38.2, 36.3, 36.2, 34.0,
28.4, 25.0; HRMS (FAB) calcd for C44H48N5O6 (M+H) 742.3605, found 742.3614.
118
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131
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I ! I I I * ' A h I I
-A A_jA—^—_—iL/>_A»fcJ
T-I ro TJ TE SJ J.C «J
y y y \i y yw
Dwd in-lKfmte CSMTRAB iKfvete
NAME 5»-T« EXFNQ 1 niOOfO I P2 - AEQ^NIIOB HNN DM 20911210 TUDC >5.35 CiSTKUM MTOBHD ; rumoc 1IM30 TD US» SOLV&RR AX33 US vm OS * SWH KI ra»es o.47?mHj AQ l.(M20TM» SC *09S iiJQOm oe •re tflOSa ft-DE
PL12 PL(3 smi
TS.OODS UJDA
'*9.»3I}00» M F1 - PMCJTEG IIMIMIIINIIIII SI tS33i 5F 123 im«792MH«
w \ l V
HO NHBoc
Compound 4d
P3 I'O IJ5 1«> i;S !5fl U5
143
IE 5=£s ie=«-sE£;8l5i«5ssj; = l=s-:55 = HE=!iiis5;iES:lii5jpS^
TsO'
NHBoc
Compound 5a
SJ 3.0
Siamtert^C Spetns
F2 - ADwiuM ptfsaon f-'r InsDutl)
",'S.SS JO
k'r.r
PI I FI.OO 4B
PC?D1 7I6#»tc
sis 'ES
i 1 iiii i
TsO"^ ^ Y "°®" "\ NHBoc
II Compound Ss
S £ C n
Wl
5SI55 S
yv I
Ji r— SF ia70l«7fc!MHi
IR :(; ,5
144
F1 - ACQVUIRIDD ( DK zoeiuM TM li.li wsrew ipsn fRWKD ^ itsa Dual I) PUIPROG ig)3 TD 143M SOLVENT CDQJ HS 16 OS I SWH fIDKES 9.»;U4»H> AQ LJ9N5*9ME
OfBu V. NMBoC
Ph •
Compound 5b
R - PROCNXM NVEMRIDI SI }27«« if WOW EM
IL JL 11 I i I
101 wo »J 9il 1.5 S.0 IJ 6S
\i |i|
I illlHi iiiis? B ill II i I i W7 \Wl I \l/ V I
CSROADATEFBI NAME EXPHO vRocm
MSTItUM PRQBMD PULTROO JMOOAIU Ctfg'* iSiX T C&CU I5IS »**& Ml HI ftonsstb I.M21IT2»BK 4091 L5.NO MR $.OE« <LDK
LA.TIISONMHI
crontfu Mun POTO PL? Pl-tl n.» SFOl >
IH Tl.£OaMC LLOOOFFL T».9AO 25.!8dS
MHI R - {¥>CU><U PSWIUIUI SI USlfc Sf 125 Tft7»w MHt
Of8u ^ NH0OC
Ph
Compound 5b
)
145
NAME EXPNQ KTAO«D
F2-Aeqai« PM. jmtiiii rat !»« (NSTSUM ttKO PAOBKD SMNDBK PinjiROG u TO 16»« SOLVIT csa) t6
TsO ^ y offlu NMBoc
Ph Compound 5c
ai&(9SLrk
1 H I jiL I J I* 9 Aim 'J '-0 tJ ».0 13 S.9 4i *0 5-1 »,® iJ UO IJ
1 Y I I l |i|14l
Suctesl l»; Sp«Rn.n AOBT M TYIKMLS FA> I3C
Sttcrai }~m }-uii (ndwH ptsbe
NAME }S»~M EKPNO 3 n>oc«o I
7i«c iNSTRUM MOBKO PULPROO
ltU6 W( Hi S 4T98)6 Ht ).!M2{M4 RC IAZ1.5
=.— CKA^MEL r» ~ IW
•JOODS tlS.TJ|«71MHi
"CHWWELO = CPDPBCJ Nuc; iM p-crc? a)iCuiu PL} LIOOODFI
I2I RCI'J* M>H
I S=SiliaE3 - < m M g l i t j
! \W Wi \ I /V
TsO" V 'Y SiHBoc
Ph Compounci Sc
146
SU-42 SMtafI to IN
t-Ro. ?'au> Coiio
c- Esas ;2 V \y V
EKPua WfOCW
T»i»w INSTSUM PKOBHD rvmno
lUB* PtfjumKn 10QIIU1 i541
62«l.7]0Hz ammut 1.}II93» >et
TsO' V Y NHSoc
Br Compound S6
F2 - Prvmoii eMiwrUn 51 377H SF 499.n<SiaSMHf
11 I I ± 'I « «
6^ 6J) )J SJB 4J i.5 i.e »J
Y i li\i\i
Csans Dttk Fmaeitn NAME EXPND 3 ntocNo i n - Pu liiHJin Doe. UOIUie Tw 17.it !N9>TRUM MCI ni(»HD 5s«D>J«IO puuRoa mio TD 655J6 SOLVQJT C»03 MS jm D£ 4 SWM PlDfiBS 0.47WHHt AQ l.»4Xr724>s RC U5I OW DE SISwE TS O.DC D> 1i.tm9SKlK dtl ojjMnogoHe
Fl - W'eeee^ piraeeuet a 63ns Sf I25.717747B MHt
I £ S i^SSlS a c S3 KSKsisis a
1\V//
TsO Y 0(Bu
NHBoc
Br Compound 5d
Mm! W l
c IB
I W V
147
mSTRUM
coomu
H48.736 auMt. iJiionzi Compound 6
'COOSu
Compour>d 9
±
148
n - Acawwitran Peeween OKC mitizs Tine llSi ff4ST»UM tpert PRDfiKD insNtlcnc I'ULnMX: n TO 'B3t4 SCHVEKT dJCn NS IS DS 2 SWH »»S.7}0HB HZMtes ajBO?] Ke AO i.iiinziMc
•COOfBi
Compound 9
SwiiiuiJ >3C5ij«iiWiMB rntaBMSpted'O' ISC
NfthBK J-m
NAME EKPNO WOCNO
m-n I i S S c SstBc 5 3 5 sis
I I j W I i l w
2I»I«I3S T« IS-t} WCTBUM ^ reOBHD iawNaiaw n'=^Kj'^COOfflu PULPS OG BS& N TD m* Boc stn.vBNT CDoa NS I7J
svn Compounds nDRS a«73»3il& AQ i iManu •« nc imj
IZJTIIWJJMH*
CTDTttC? NVC2 fCPDJ
l«b
149
537-11 .ftCsCU IH Spenrna
D>aim-nc \l/ Cvncnl D«il Ftrvtftn NAME «n-it EKPMQ I PROCNO I
fj ~ Ac^ovsft hmnenn Dik ^0121 Timt I4<7 {NSTRUM HMU FftOeXt} SmDMtO fl/LPROO tfJO T13 163M SOLVENT CDQ! KS OS
6Z64.4I I Hx OJRlMHi
C b z H N ^ J j \
Boc Compound 15^ Standard ^HNMR
fl - Pracuoas SI n7« SF 4n»3<»l«lMHl WO« EM
iU A ! . i M i l
Ur7'tBiaCDCQ SuBdMil 1K SpomB Dtt pnta
CwnM Daa PwMum MAME un-ll OfPKK) J PnOCNO I
P2 - Afi^eiwK ^inawicn Datt UDJ0}2t T»Be >4 Se tNSniJM ipcU PROBHD S an Dvu) n PULPROO ttpt3« TD tSSM SOLVENT 0303
«ns
V V W 'fw ' Ssfas 2S sSSsSB ssccs s» sacssi: §5 ?555»£
SSSSiSS
S W V V I V V w
DS
fAJCI PI
STOl
3I446S4I H« e.mmHt
I iHim* >«
IS.9QDSMK
m/ci pcpDj n»iH PL2 iaQ.ai AB
•voMt pmn»tn
)?5 MHJ
CbzHN,
MeOOC ,r N
Compound {5c^ standard "c NMR
«4««
(14.010 til.m in.90) in.«H uriiM I MM mm iM.m t2»^ 121 T5i I2I.M] I1I.ST7 mm 1M.M7 lU.tTS mT13
151
o«, :aix»«J
IF toD.imuxiMi
\l ^ I ^// \.y>^ I OMe
6 MeOOC L:
Boc
Compound
SUndard NMR
JL J[ u Vu_iV
w • li/ kffl V W i
S3c;:ccc{:& SSSSSSCSS sscsssssc
CbzHN,
•COOMB MEOOC Boc
Compound
Standard NMR
l'> !• 1(1} lo; If
152
stn-u-soM' tHS^ECBWB C>1»( iK'DrPmbc — CMni Out Puancnn NAME EKPND I HIOCMO 1
FZ - Ar^waWA PwwwKn Dali 2n>}0«t9 Tiiw IDS iNrreuw sfo nOSHD JaunOualia riAMOC J^w TO 163»* SOLVEfT CBCU NS 1$
2 i2<«.4ll Hz 8.}UM»Hc l.]ff7754aMc
SF «»JXBt52 MHx wnw Sti
CbzHN
Compound
Stand^d H ^MR
i i «
• 0 3,3 J.O
gl 8 seiSSiBI | SS = ^5£^5SrlS 2
CbzHN
Lampeuitd
Standard '^CNMR
153
Ou*/ IH-1JC Freer
NAJKE fcOrH EXPNO I PRorKO I F1 • ^c^^^9a^ Ptf •mien nw !oa}oi2S Tubt n <$ INSTOUM Iprct PAOBHD 5fenO»>l>) puuftoc teJo TD SCM-VEKT a>a>
«76«»lt V O.J833«
1 .« rOi.« rs.tU»<r Ct»mpound
Standard 'h NMR
i i ) b _L
JiJ 4.J J# *0 f.S i.B LS 10
y y y \i y
JIS5=s£s£5-5Si stfrfs-aaSaSsass 15 = 11
tivnuvM 1^ »BMKO *i-.TI nn.»«on ir<
Cb?HN 0 COOMB
Compound
Standard '^C NMR
154
M// V \|/ U — •
T« iKSntlAl rftOBHO 1 ruLmoo
iW TJ9TJB* ncM£3 »«nu]t« AQ IMltlMof
CtJzHN Q COOM9
Compound Standard NMR
LJL
y ¥• « 111 i ¥ y
=1 ^gfSSS 8 snssn;: s
WW! Ic£ B s^fx CR?? S S'SRSi
CiMCM DM PifMtBm NAME m-iA-rnitat EXPNO 2 PROCNO >
Dae. Tocnun Ti« W-J4 tKSntUM sped PaOBKD Sm)«D»c>0 p\?Lni(X> taiii TD 45516 SOLVENT CDCII N5 I5Xlfi OS « 3W>t Hi FUMES 9 479»3AKl AO i.wimiiitt RC 6Mn DW llSOOswc DE &DO«K TE 0.0 R D» O.H9»»»K* dii nojnxioniu:!
Compound Standard NMR
«»0*ANNSi n »
lii niWT3 MHi
«9J 9JI5COI MHi
H'i 707«H Mill
155
IKTTRUM PffOVlW i fVLrtoc
SFO) MBUWKIH
f2 - ^rvccUiH
N^N'V"
Compound
Standard NMR
.lii
COOMe
Compound 30ci Standard ^'c NMR
in
~ >a»sjc?0»B m -o — -l-Mrjrj— 93BBW03 iw. r^" ^ r-' \a <£> \C >d
w w 607-50-li
SfSftdaTri lO IW ^pccirum Nnlorac 3-Rej. l-aaa Ovadient Probe
CurrenJ Data P?i»<Mtf(eri
NAME 607-50-11
EXP/VO f PROCNO I
P2 - AcquisKton Pnfnmctcrj Dale. 200206 )^ Time {2.15
INSTRUM tpevj PROBHD 5 mm NnlorPv J'ULPROG If TD 16AM solvent CDCt3 NS 64 OS 2 SWH 624S.750H?
FIDRES 0,3Rt59> Hi
AQ I..TJJD32: RC m DW 80.0U usee DE 6.00 usee TE 31X1.0 K Dl 2 00000Q00iec
Re=a«:s==*==»a CHANNEL fl «»**=*saa«*»s
Nuc» m P! T.OOiisci:
PL J Q.OOdB SFOJ 499.9.^8752 Mkt
F2 - Procejiiiif pnraiMeieu
SI 32-76S
SF 499,9J0013» MH2
WDW EM
SS8 0
LB 0.20 Hz
CB 0 PC 1.00
MeO.
COOMe
Compound
Standard 'H NMR
157
Stante4 iO IH ^ctintn •eJ-Rej )-^u OftAaw^nXK
ssj»2s;r
HAMC Wrf~M exrim i WIOCWD T F1 - AiquiMAW PwUMcn Ds* 20SZ07I8 Tint I2.i8 ENSntVM Iftsi noam > m Ntt<» rVlTROC 4 TO IU»« SOLV0A CDCIJ MS IS DS 2 SWH 62M.7}0H> FSHies CM>nt3Hi AQ tJliUZlM*
izisll
Compound SlS
Standard V NMR
MMt
wow EM 5S9 B CB » ?C 1 w
.
i 3.J J-U J $
CoiiM Date PirweNn NAME EXWO^ *1 R - Ac^ritna PBsmert Om 7lB3tr/fl Tine I1J< EMSTHUM tfia mOBHD SusNriatK PUtPROO »#4t
^1/
TT» JD4.V6WT CD03
}iW6 3«) Hi 0.47M16 H> IMUmtM Z89ej
All o.(i»»ocianK< «=«=rw«*i CHArfl^L » Ntfci nc n n.i -JtOiB SPOl UJ.731W71 MH>
Compound
Standard "c NMR
(7CPRC2 NUCI rcsTO
n - Praclln«t pirmsi Si 6U)« sr m7tI730]«M WOW EM
158
V
CmvM &ftu P. NAME 697-69 EKVfrtl t ntocNo I fl - Acttt«uuM>P(*«m>c» DMT. mivnx T\m f«7 iKrreuiu tpxi PROBHO SoffiNaimc
FVim<K tt TO t6)H SDLVEHT CCCIl DS 1 SWM 62«B.7)!' Hi FBTRES OJtl}})Kt AQ iHtfaUnc
Compound 29 Siandard NMR
- fmeuMa^MONten 499.9)900*4 M(b
i SJ w •J *.9 ZJ 2,8
S(n-60iBcici3
NAME ExrrK) PBOCNO
nJLfaoG yp TO 6S5« 5«a.VtN7 CDOS
SCiCS
Of'Bu M9 NS DS SWH lIM&Mt Hi FTDEIE5 0.mS3S Ht AQ IMiOTZ* uc BC zmi l5-9W«iee
Compound
standard "c NMR
OS tE O.OK
t I
CTDMIG2 KUCI «TDJ
Fl - f>KUi\M psmwRn 51 S5i36 Sr (25'<r7'Qi7 MMi WDW EM
159
-ssg jsac
W/ HV
CoRcM DkU 9tmt>tvn NAME 601-M EXPMQ > nocno 1
tlBK fusnxM PROBHD niLraoG
MOB PaniKsKn ame?i« iO.<«
MWW H» SHtJSlHi I SI1UZ2 e«c
P2 - PtocauTU SHI S! JVU Sf «V9.»ia>40«4Hi WQW EM
CompountJ ,"10
Stanilard NMR
IJ 1.9 «« 1) ).0 13
i 111 i i
wr?-M SoadDtf iKSvnms fMta Mc G9t(^ t3C
NAME EXIW PROCND F7 -Dae 21H3V7lt TioT lO.M WSTSUW "pea PitOBKD S K"® pULmoc *«4«-Tt> aSJA SDLVHWT N.<1 OS * >1446.541 H> 3.419BM HI
>2WJ *S.»0 «>i<« SSDuai OOK
I § 9 i S s clSil^S « • S::;SS!3SR
=85 S
Compound JO Standard NMR
iPOl lM7JI»0T>MHi «=«»«=.!?=« CMAWWEV n CPOPHCJ •riJBlfe m7C2 IM pcroz »o »«=
160
_
li 12 5™k:
rt c^KNEL —
PI Tl»»»t
a T-pr (jxt
9 0 SJ BS «• 1> t.V
iMfflaM IB
fS. 'SSS PtlJ I3.M4B SFO? m.»»JW( V
'Jo. '"""«' ^t"
ii ii iiiliiiiiilliiiiliiliii
I M
Hsff^SS 9 I? CC';g?:ge 5 saaaE =
M / i i / 1 V \ y \ / I
i:/ !>(. •»; .»< •)' i:i 'i' i^'.
161
F} - Ae^tMSiicn finn"^
Compound UR. Standard NMR
9.» 8,3 ».» 1i *5 8J »J) J> >8 «4 "-O iJ >B 1! ;0
5 - S S S f55sSg53S2SSs5S= B :S355s = z ES5 BicBsB^cESSSB^SSsssaaRa
P O -nA
/-^ I T ft /__., ,1 »N-^Nh--N pw'
r'V T ;u°
Compound (2R. 3S) 33^
Standard "c NMR
ll'iid ffflfiii lSiilili'Tilti1i|^iliiflliii!iil'1
EiSSis isl v n i 1 1 1 \ n /
162
S>1-M-p»i IH Syu-Watt Dual1H-l)CPri>»
CuflCM DlU PvMKEn NAME wn-i^-r" EHTNO I PROCMD t
IJIS Fl-Ac«iui Oau. ZODIMII TiOH iWTRUM PflOBW i 1MB uw I.' niLPROO uSS TO lOM SOtVEX? CDOJ N5 U OS 2 JWH FIDftES »)aU«*Ht AQ • WTHM Kt
Compound 34 Standard NMR
MVCI tH PI IS«eat< PLt O.QOCB SPDI «** «3UTU MHi
1 JLl
i i \i yi III \i i
SummijC ipcEtaa DnltK.i)Crn(«
CancMOati Pvtacttn NAME fi(n-7«-pe EMWO t PROCNO I n - Aartiaiim hneHsa Due, U)62aiI7 ruM IJ.U JHS7WM Ipm PHOBHD SiM(iD«alt) PULPROC ccpgW
SWH Fnȣ5 a.4nauHi AQ lOOmMBe
lii
9a K aojooaooote
JD^ O^^-^COCMe
Compound 31^
Standard "c NMR
cpsreci mtliit KVfCJ IH pcn>2 TZ-tB MCS PL2 imoeds Ptl2 ItOStt n.11 U.OBdB 5F07 WSMJOBIMHi F2 - Fnxeuut; panavim SI 6SS3fi SF nS.TinSMT MH» WDW EM
M.Q93 m
}7.Wi n.y4»
" i?2p|| i? = ij
to.5n ro.jM TT,13«
6S.U7 3T.t« S1M9 Si.5Ti lUTQ SLWi <W.J« M9<4 M.OH 3La]« )3.I)S XKM llMl )IU) » coe U.03t 37,978
165
fNST*\JM MtDBHD PUUWOC
i s:.r
~~T—-S-"""'" h, JH-h.
FI-Pr»cw«»>8Sww««»
!?!= !!! !!l!ls! £3n?5;5!si5Ss5ili5lii5: = i;!5s5|;s;55 V V w
MeOOCjH)
Compound
standard 'h NMR
JLJ Jv 1 III
I, I
"isrV W lif Y Y/W i!ri?rt=ii
Si>wiann)C%MV«n Dv«l IK-llCprate
Fl - Ac4<iinii«e Pv»a>e
r t 411 OLinoaooroKt
,jri PLJ nonoiB n,u i^nntjB Cll) JMMifB
iiiil iiii mm NK \ \ \v^
Compound !^7£i Slandard "c NMR
i l l ; imiimmimmii
ji_«L iJ.
166
fULPROC If soLVEHT corn KS lb
rt
jni-t i i-i-
\\V'l I
r .S...
0^ "H. ( °
Compound
Standart) 'H NMR
¥ l^f' 'ii/
'S~T
NS
s. "ksx. 'S?-
s tF" «' ' OOKOMSI^ m^T" PI
PCP02 P12 n.u SP03 n - Pwc
r".pr®' t) 90 .MT IIS 00 da nradB •*S»>I5I»1 MHJ
94ift| p»/«mcteT> »
SSM 0» iX
MHi
iyii III "Wl \ \ /
mm "iWi
Sill! iiiliilsliiliiiiiiiii
N -
Compound Standard '^C NMR
U-
I
J,.
167
£:si = i55=j i5g5j ; iJ5=SssMs5£ss£RBpi5= ; =i i l £5|I- I=M
2,'; i;, laJiiSiii?'"" - •" j \y7
Suniard ID IH SMITMR Naiffrw: ?—Mif Qfvdicw Pra^
UI
Compound 33
Standard V NMR
lil'^ ¥ W W\i4^
Cmkm Das Pi N>M4E W7'9I IXTND 3 PROCNO t Fl - Ac4aiiitiBfiP«Min<n DMT 2M2W)F Tiae IIJI OCnmUM wen PHOWFD JMCBDUFLLO ruu«oc tm^O TO hSUt CtJZHN'
0 47WH HI I (HjaTUtcc M9<:t
0.0 K 0 «»V»»n (a RBToanoaia
SFOI kU 72)907.1 lufHi VT-TJWT^CT^ CMANWEL R ir CPOPftCI mjcj iH PCFtn IISOmm pu ud-ooa PL>} 19.0}» PLO IJOOdS SFD2 «»797iStlOI MHT n - Pn>r»».
OR::'
Compound
Slandar i ) "c NMR
3iS I? 5 J l ?5~i sj
JL
168
i::H = l;5?6S5?fSa5;;EsS5l; -""" i,;^ y"" - "i ii,iiTi" •""
= i V
l53=5j;|l:|!?FjSIJEr!:;iJ«; = riE .ililSiSSiiiSE
;;:s—
:r
Compounif 3? Standard NMR
± u ilidiL
y i 1^ IS iiiittyw
i i i i mm m u p i p s j =
——- \\i I !ii\n V ii III iVi I "s:s"
Y o IKmUM mOBHD SmsNaim ^rooc Cb?HN soLVEWT ccoj o uuufvre SI FTOftES "o'iTOJsm Compound 3 ]
Standard "c NWR
169
:; : 7: V! ? ; 3 r f" 5 r - - - -''" - - - ^ ^ ^ ^ ^ ^ 5 ' y' ^\i/' ]j
{
MeOOC ^ Boc
Compound li^
SUndard V NMR
- IP " ' '-Uj
¥ ¥ ¥ iti w vi^ w
r-uri" 1 - Acqiiiiil>«<i FmoecBt }«ii zoQiiita IK 1121 •ROMS) SMDittli;
SOtVEKT CDOJ T
\ fr
il' ,s,." ==^3=——«(^.>«r«eL n
,Ill i „li.
i 'V PC i-W
i i 111 i i i i mmmm VI V T
Cbzm,
MeOOC
Compound
Standard ''c NMR
j U
mn mrnmimmmmm
170
MAME tCTNO M«OCXO
WJTRUM raoBHO : PULPBOC
62M4M Hf g 1B2)44 H> I wsas Ut 456 I
2 nnnoocwm
F7 - Praceuis^ MiuniWJt SI )27U SF M9 93aOtJEMHt WOW EM
CbzHN COOMe
Compound
Standard NMR
t-B 7J 1.0 U
¥ 3.} >0
"is" y y
S:€issiall§S
mW \¥i NAME S61-113 EXPHO 1 PHOWO 1 n - Ac«tJ*^M farwwtm OaM icntii? TM I1.I» rNTHlUM «pe*i PAOBHD SmmDwiO nitrRDC igpB'O Tt> 65J0S SOL-VEKT COO? MS as )ia«6.>al H<
i g47H3&H> t MUr72« )tt USI IJ MO utcc LOS ««s SDK 0.SSWW99 KTT O.tDOOWOO tms o-ixncino Mc
wuci n.1 iFDI UJ.7J»W7JMH>
CPDPBC? MUCJ »C«>3 n.2 PLll fL»J SR)1
CbzHN
Compound
Slarjdard "C NMR
171
iiiiniiihiiiimii;;:^,::;' 'J
PBO^O 'l fl - Atow...t.fr^^l«««cl T\^' 11-5)
p£ ;::fs-g.v„, ";;»a,
?k, 4h^x B 'EC 7,
Ei, .3£».,
« I 00
Q COOMe
Compound 44
JUlUIUjL
eo <0 7.9 »-J <uO
v \i w y
*.J
III 111 w 5.1 >.e 1.5 io
k W isj [31 mi
•sr™
fS^:
si 'Eir E s.'sss
la..
?U l2fl SIJ sss 5K): »!»<l>ISOW)M»i
ii 1 I iiiiiiliiiiiiiiiiiil
Compound 44
m 11 ii M/ i l I V /
172
f 5sS®5a-'is»5 = ss~-i2ss--''S5S£sss?=35E£3a'--®-'
«n-in Suetlfi IP tWi^ul-WMB I
Nairn >-*n )'4u>0ii&aiPTe^ |
BocN ^NHBoc •NH
COOMe
Compound 47
I
J
tm-m StaaAaiA t3C Spoanra QkdiH-lX:pRte
Sr 5sB §essssg|cs •35 fiss SISSSSSIhRRS
I I I V V / W
BocN ^ y-NH0oc NH
HN „ o
Compound 47
|5=|3iiS IS Is ^sssSSsS
I 1 11 W? V
173
Sf5=Sa!Siijl!5;55;S5S5 = nES!;S;|S5||I ==5H;;nnHiH:?£-HiS;^
MN \ At o COOMe
Compound 48
w ^ « w i i y
OE"
Cr°
i" 7"
ti
i i i f i l i l i a m u i i l \ V I I I I \ \ \ / l V V
174
i.=5"=»ss"ss"5E553H=5S3s=uinn5=55=8i55issis:fH!!fl;;;;l!Hi!5!=55!H£=?;===5H!li!l;!
BocN p H }~msQc
NH
jPp0-^0° \J \ r
: Jlll_i i ii_iALU^_JjtiL.i
11 ill lillillliilillllli \i IV WM^/Wy
CPCS s s
'•ssrH-s:"
PE="
S es A.O
SS 'S,„.
'.s.™
•£'1' I?OftdB
0-^0°
Compound 49
ll 1 i
1
\\F
BocN
L
s£»is sSIs
Wi wi
loM
175
BacN ^NHBoc -NH
Compound SO
aoMCwCcoi
NHBoc -NH
Compound SO
176
1 ' • 'iiiiiiii
M1-U7 Siindtrt to IH SfesBW )-^a. V-Ufti Pnrfrc
riiiTni fiTif Pinimirrn («W<E SOT-H7 EXFMO I HtOCNO I
Out lOSMit} Tint IJ.d INfTKl/M tost mttKD SsnNtlm PUIAOC tt TP \S1W SOtVEKT CDQJ
t~w-? T
BocN ^NH0OC
0^0°
Compound 51
I I J *_ I 7 iN M ' *1
•1.0 u> 9J 9.B tS S.# 7J 70 JJ J.« IJ 2B M
»s&r 1 ii iiiiiiiiliiiiiiliiiiiliili I y
,3
BocN ^NHBoc
NH
WN y \
Compound 51
im ii i mum m I I I N w v i
2,T 155 IW iri l^n nj 70 t? y !:
177
(W lN-|]Cr'ttx
Csiun Dwt PanawKs NAW 638-n EKFMO I raocNO I
One. I»»il6 Tuat il.a INSmUM (pott P80B(« SiasD<i4lt] niLTSOC zg]0 719 I6)H SOtVENT CI>CB HS 16 DS 1 SWH ftUJlSSHt FotEs Oman Ml AQ iJonmya HC 3Z2.S Dw mnuiK
> CMANNB- (I 2
JPOl F7 - fTBccuMi MraaetH » lll£» 5f 4««.M(»l)3MH KTDW S>4
HN ^ L O COOMe O 0
Compound S2
T-5 IS
w « i a
iS 9fl •J ».# jj ,J> JJ 1.0 13
Hi y iy i ]i i \i w ii( y
S3MI li»dw< IJCSfmsnua tW l»f-nc pabe
NAME 67«-il Exmo 1 mocNo I
CWt. fftSTKUM fVQBHD nnnoo ID SOLVBHT
CFDPRC3 okteti Kua m PCPtt: 7Z.UWBC pu lat.ceda Pttl lf.OOiB rtn n.QQ4B SPD3 mHiMlDCMHs n - Pmcunnjps SI MSH SF tUMU5n
n iisssaii i i fs i is 11
Sail El is 8 2 llsSS I
W
L o COOMe "O'^O
Compound 52
\1V/
11
178
!5!EE!EES!!5=!!E=!!EE5!!5!5l53!!i!|£!E£!!ii^
MHSoc
SZniHHz yoBQ\a Zompound 53
»o.ox
4»9inrs>wii
Cmat Dca ParasKSn NAME S3»-n EXPMO i psocm I
10DM3U Tun li.U 0C7BUH tftes PSC»ifi> jOSBthMitl PUmtOG UKM TO S5534 iSOtVEJ^T CDOJ NS WMS £35 « SWH 31440.052 Hz nDltfS 0.«77T40 HI I^Q l.W22t091« RO lUSJ
cpcmcn •nibit wun iH rcPO! 71.6<IM« fl3 IIOQOdB PLI2 l9Q0rfB PLil noma 5(^2 «HMt5Cn>Mlb - Pncnilitrpi
iSlit (25.M33y5M>t
"O 0 HN
?3|§ ? 5
W
Compound 53
iJ
\\l(//