Straightforward and diastereoselective synthesis of tetrafunctionalized thiol synthons for the design of metallopeptidase inhibitors

Article abstract of DOI:10.1016/S0040-4020(99)00922-9

Tetrafunctionalized thiol-containing synthons with different side-chains have been prepared with good yields by a straightforward diastereoselective and general methodology. The key step of the synthesis consisted of a tandem reduction and Wittig-Horner r

Full text of DOI:10.1016/S0040-4020(99)00922-9

TETRAHEDRON
Pergamon
Tetrahedron 56 (2000) 209–215
Straightforward and Diastereoselective Synthesis of
Tetrafunctionalized Thiol Synthons for the Design of
Metallopeptidase Inhibitors
*
´
Christelle David, Laurent Bischoff, Bernard P. Roques and Marie-Claude Fournie-Zaluski
´
´
´
Departement de Pharmacochimie Moleculaire & Structurale, INSERM U266-CNRS UMR 8600, Faculte de Pharmacie No. 4,
avenue de l’Observatoire, 75270 Paris Cedex 06, France
Received 28 June 1999; accepted 14 October 1999
Abstract—Tetrafunctionalized thiol-containing synthons with different side-chains have been prepared with good yields by a straightfor-
ward diastereoselective and general methodology. The key step of the synthesis consisted of a tandem reduction and Wittig–Horner reaction,
which conserved the stereochemistry of the starting material. The method was generalized to different side-chains, allowing synthons for
designing inhibitors of various classes of zinc metallopeptidases to be easily obtained. ᭧ 1999 Elsevier Science Ltd. All rights reserved.
Introduction
overall yields and racemization during the synthesis of the
synthon 1. To overcome these problems, we have developed
a rapid and diastereoselective strategy by modification and
improvement of a previously reported method in which
aspartic acid or glutamic acid diesters were used as chiral
precursors.6 This allowed any suitably protected ester side-
chain (sulfonate, carboxylate, phosphonate) to be intro-
duced. Moreover, synthon 1 contains two asymmetric
carbons whose configurations were controlled all along
the synthesis (Scheme 1).
Zinc metallopeptidases play a crucial role in the activation
or inactivation of peptides involved in the regulation of
biological functions and several inhibitors of these
enzymes1 are currently used in clinics. There is an increas-
ing interest in synthesizing potent inhibitors of metallo-
peptidases as new potential therapeutic agents. A large
part of the most active compounds developed to date contain
a thiol moiety as zinc chelating group. In continuation of our
studies2 about the physiological role of aminopeptidase A
(EC 3.4.11.7), a zinc metallopeptidase of the M1 family, we
have investigated the possibility of designing new potent
inhibitors bearing different types of hydrophilic side-chains.
Such compounds were derived from N,S-protected a-
mercapto-b-amino acids 1, the synthesis of which was
previously described3 by electrophilic sulfenylation of N-
protected b-amino esters, available from the corresponding
amino acids. However, in the case of a-amino acids bearing
a protected negatively charged side-chain (carboxylate,4
sulfonate), this synthesis led to the expected compound
with poor yields, whatever the protecting group used to
protect the charged moiety. Thus, one of the main by-
products isolated using a compound bearing a neopentyl
sulfonate5 side-chain resulted from an intramolecular
Claisen reaction during the sulfenylation step. Moreover,
when unnatural amino acids were used as starting materials,
this synthesis was often critical, the preliminary synthesis of
the suitably protected unnatural amino acids leading to poor
Finally, this synthetic approach was extended to the intro-
duction of other R1 side-chains in synthons 1 leading to a
generalization of the method.
Results
Commercially available Z-(l)-tert-butyl aspartate was used
as the starting material (Scheme 2).
The remaining carboxylic acid was esterified with diazo-
methane leading to compound 2 without racemization of
the amino acid observed when either cesium salts/CH3I7
or EDCI/DMAP/CH3OH8 were used. The next step
consisted of the introduction of the protected thiol group
in the b position of 2 by means of electrophilic sulfenylation
with the reagent described by Bischoff et al.4 The sulfenyl-
ation proceeded via an anti addition leading in this case
to formation of 3 with the (2R,3R) configuration with a
diastereoisomeric enrichment of 95:5 measured by HPLC.
Keywords: amino acids and derivatives; thiol enzyme inhibitors; Wittig–
Horner reaction.
* Corresponding author. Tel.: ϩ33-1-43-25-50-45; fax: ϩ33-1-43-26-69-
18; e-mail: roques@pharmacie.univ-paris5.fr
The a methyl ester was then used for the incorporation of
the side-chain aimed at recognizing the S1 subsite of the
0040–4020/00/$ - see front matter ᭧ 1999 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020(99)00922-9

210
C. David et al. / Tetrahedron 56 (2000) 209–215
Scheme 1. R1ˆSO3CH2t-Bu; COOBzl; P(O)(OBzl)2; CONHt-Bu; CN; COCH3; CON(CH3)–OCH3; C6H5.
Scheme 2. (i) CH2N2 2.5 equiv., Et2O, 0ЊC; (ii) (1) LiHMDS 2.2 equiv. (2) PMB-S-S-DNP 1.4 equiv., Ϫ78ЊC; (iii) 4a–g: Reagent Z 2 equiv., n-BuLi
2.1 equiv., Ϫ78ЊC, Dibal-H 1.95 equiv., Ϫ78ЊC !RT; 4h: Reagent Z 2 equiv., n-BuLi 2.1 equiv., Ϫ78ЊC, Dibal-H 1.05 equiv., Ϫ78ЊC !RT; (iv) 5a–c:
[(Ph3P)CuH]6 0.36 equiv., H2O 20 equiv.; 5h: KOOC–NyN–COOK 18 equiv., AcOH 36 equiv.; (v) TFA 15 equiv.; anisole 5% vol.
Scheme 3. Aluminoxyacetal intermediate for the one-pot reduction and Wittig–Horner olefination.
1
metallopeptidase. The synthon 3 underwent a smooth and
regioselective reduction of the methyl ester with Dibal-H,
leading to a postulated aluminoxyacetal intermediate,6
which was not isolated and was in situ submitted to a
Wittig–Horner reaction (Scheme 3).
without loss of optical purity. Moreover, H NMR data (i.e.
coupling constants) showed that only the E-isomer of the
double bond was formed. It can be seen that a complete
racemization at both C2 and C3 (Scheme 2) was observed
when the same reaction was performed with the correspond-
ing free a-aminoaldehyde.
Some examples illustrating this reaction are given in
Table 1. The reaction of 3 with Dibal-H in the presence of
the lithium salt of the dialkylphosphorylmethyl derivatives
bearing a neopentyl sulfonate,9 a benzyl carboxylate10 or a
dibenzyl phosphonate11 group, respectively, resulted exclu-
sively in the formation of the unsaturated compounds 4a–c
The a,b-unsaturated esters 4a–c needed to be subsequently
reduced in conditions keeping the stereoisomeric enrich-
ment unchanged. Indeed, sodium borohydride in ethanol
resulted in a rapid reduction but a complete epimerization
at the C2 carbon. The same result was expected with Mg/
MeOH as reagent.12 Moreover, the presence of the thioether
and benzyloxycarbonyl protecting groups was incompatible
with catalytic hydrogenation methods. Therefore, another
reagent, [(Ph3P)CuH]6, specific for the reduction of
a,b-unsaturated esters in neutral media,13,14 was tested.
The reduction was obtained at 25ЊC in 3 h with only a
small change in the diastereomeric excess at the C2 carbon
(80:20 for compound 5a, 70:30 for compounds 5b and 5c).
Table 1. Synthesis of compounds 4
Entry Reagent Z
R1
Yield (%)
4a
4b
4c
4d
4e
4f
(EtO)2P(O)CH2SO3CH2–tBu SO3CH2–tBu
68
58
44
17
38
51
(EtO)2P(O)CH2CO2Bzl
[(BzlO)2P(O)]2CH2
(EtO)2P(O)CH2CONH-tBu
(EtO)2P(O)CH2CN
(EtO)2P(O)CH2COCH3
(EtO)2P(O)CH2CON(CH3)–
OCH3
CO2Bzl
P(O)(OBzl)2
CONH-tBu
CN
COCH3
CON(CH3)–OCH3 72
4g
The last step consisted of a deprotection of the tert-butyl
ester with trifluoroacetic acid/anisole at 25ЊC, giving the
synthons 1a–h with high yields.
4h
Ph3PϩCH2C6H5, BrϪ
C6H5 60

C. David et al. / Tetrahedron 56 (2000) 209–215
211
As far as the stereochemistry is concerned, the three
synthons described for APA, containing the side-chains of
a glutamate residue 1b or those of the unnatural sulfonic 1a
or phosphinic 1c acids, had essentially the 2R,3R absolute
configuration when the l-aspartate was the starting material
and the 2S,3S configuration from the d-aspartate. The other
stereoisomers, if needed, (2R,3S or 2S,3R), can be obtained
in small quantities with the latter procedure but the epimer-
izing reduction of the unsaturated ester 4 by NaBH4 can
allow their production in 50% yield after separation. Indeed,
this reagent induced a complete epimerization at the C2
carbon and the two diastereoisomers obtained could be
separated by chromatography on silica gel.
The synthons 1a–c could therefore be coupled to various
well-chosen amino acids by classical methods of peptide
synthesis followed by deprotections, allowing the design
of inhibitors of the different families of zinc exo- or endo-
metallopeptidases depending on the addition0 of different
moieties expected to interact with the Sn–Sn subsites of
the enzyme. For instance, this synthesis has led to the
most potent inhibitors of aminopeptidase A reported to
date.20
Experimental
General
To generalize this methodology, the sulfenylated derivative
of aspartic acid 3 could also be treated by Dibal-H in the
presence of phosphonate anions or triphenyl phosphonium
ylides bearing other side-chains. In most cases, the unsatu-
rated compounds 4d–g were obtained with good yields and
good diastereoisomeric enrichments whatever the side-
chain. In the case of the synthesis of 4h, the E/Z ratio was
2/1. Furthermore, unactivated double bonds (e.g. CHyCH–
Ph) could not be reduced by [(Ph3P)CuH]6. Consequently,
we used diimide, previously reported to afford reduction in
relatively mild conditions.15–17 This reagent has been
prepared in situ as described by acidic decomposition of
dipotassium azodicarboxylate.18 The reduction was
performed with a slight modification of the procedure,19
since 18 equiv. of dipotassium azodicarboxylate and
36 equiv. of acetic acid were required. The reaction was
monitored by HPLC and was complete after 80 h. This
reagent was also used instead of [(Ph3P)CuH]6. Thus, only
the double bond in the cyano derivative 4e was reduced in
the same conditions as reported above, allowing the produc-
tion of the saturated cyano compound in good yields with no
change in the diastereoisomeric excess.
Materials were used without further purification unless
otherwise stated. Z-l-Asp(OtBu)–OH was purchased from
Bachem (Budendorf, Switzerland), reagents as n-butyl-
lithium, 1,1,1,3,3,3-hexamethyldisilazane, diisobutylalumi-
num hydride, sodium borohydride, triphenylphosphine
copper(I) hydride hexamer were purchased from Aldrich
(Saint Quentin Fallavier, France) and Diazogen᭨ was
purchased from Acros Organics (Noisy-Le-Grand, France).
Trifluoroacetic acid and solvents were purchased from SDS
(Peypin, France). All reactions involving strong bases and
air-sensitive reagents were carried out under an argon atmo-
sphere, and starting materials were thoroughly dried under
vacuum prior to use. THF was distilled from sodium/benzo-
phenone. HPLC analyses were run on a Shimadzu LC-10AT
with a reverse-phase column (Touzart & Matignon, Vitry-
sur-Seine, France) with CH3CN/H2O containing 0.05% TFA
as a mobile phase. Flash chromatography was carried out
with Merck silica gel Geduran 60 (40–63 mm). TLC was
performed on precoated silica gel plates (60F-254, 0.2 mm
thick, Merck) with the solvents indicated. Plates were devel-
1
oped with UV light or iodine vapor. H NMR spectra were
recorded using a Bruker AC spectrometer (270 MHz) at
25ЊC. Spectra were internally referenced to HMDS and
peaks are reported in ppm downfield of HMDS. Multiplici-
ties are reported as singlet (s), doublet (d), triplet (t), some
combinations of these, broad (br) or multiplet (m). 13C
NMR spectra were recorded at 68 MHz on the same spectro-
meter as 1H NMR spectra, at 25ЊC. Spectra were referenced
to HMDS and are reported in ppm downfield of HMDS.
Melting points were measured on a Bu¨chi B-540 and are
given uncorrected. Mass spectra were performed by Quad
Service (Poissy, France) using electrospray. Only molecular
ions are assigned. Satisfactory elemental analyses,
performed at the University of Paris VI, were obtained (C,
H, N) for all compounds. Optical rotations were measured
on a Perkin–Elmer 241 polarimeter for CHCl3 solutions at
25ЊC.
Moreover, by further reactions involving the conjugated
double bond (Michael addition, dihydroxylation) or modifi-
cations of the side-chain moiety R1, compounds 4a–g, could
lead to a variety of functionalized a-mercapto-b-amino acid
derivatives. These completely protected compounds could
be introduced in different kinds of sequences by appropriate
deprotection, to yield potential inhibitors of many different
metallopeptidases. For instance, the amino group of the
synthon can be kept free, leading to P1 moieties required
for aminopeptidase inhibition. Conversely, the a-mercap-
to-b-amino acid derivatives could be introduced in different
positions in a pseudopeptide or non-peptide backbone
through coupling with various amino acids or derivatives
to furnish endopeptidase inhibitors.
The following abbreviations are used: cHex, cyclohexane;
EtOAc, ethyl acetate; THF, tetrahydrofuran; TFA, tri-
fluoroacetic acid; Dibal-H, diisobutylaluminum hydride;
HMDS, 1,1,1,3,3,3-hexamethyldisilazane.
Conclusions
We have developed a new and rapid methodology to synthe-
size in five steps N,S-protected a-mercapto-b-amino acids 1
bearing various side-chains without racemization of the two
asymmetric centers present on the common aspartic acid-
derived precursor. Moreover, we have demonstrated that
the tandem reduction and Wittig–Horner reaction allowed
the introduction of different side-chains, which could be the
precursors of novel unnatural amino acid derivatives.
General procedure for the one-pot reduction Wittig–
Horner olefination: procedure A
To a solution of the phosphonate reagent Z (2.0 equiv.) in
dry THF (4.5 mmol/ml) was added dropwise, at Ϫ78ЊC, a

212
C. David et al. / Tetrahedron 56 (2000) 209–215
solution of n-butyllithium (2.1 equiv. of a 2.5M hexane
solution). After stirring for 30 min at this temperature, a
solution of 3 (1 equiv.) in dry THF (1 ml/mmol) was
added, immediately followed by dropwise addition of diiso-
butylaluminum hydride (1.95 equiv. of a 1.5M toluene
solution). The resulting mixture was stirred for 3 h at
Ϫ78ЊC and warmed to room temperature for 1 h. 2N HCl
and water were then added, the organic layer was separated
and the aqueous phase was further extracted with ethyl
acetate. The combined organic fractions were washed with
brine, dried (Na2SO4), filtered and concentrated in vacuo.
The product was purified by flash chromatography on silica
gel.
(2R,3R,4E)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-5-[2,2-dimethylpropanoxy sulfonyl]-
pent-4-enoic acid tert-butyl ester (4a). Starting from 3
(12 g, 24.5 mmol) and following the general procedure A,
the crude product was purified by flash chromatography on
silica gel using cHex:CH2Cl2:EtOAc 7:1.5:1.5 as an eluent
(Rfˆ0.4) giving 4a as a yellow solid, 11.6 g (68%,
d.e.ˆ90%).
Mpˆ56.3–59.0ЊC.
Anal.
calc.
for
C30H41NO8S C, 59.29; H, 6.80; N, 2.30. Found C, 59.35;
H, 6.74; N, 2.66. [a]Dˆϩ74.9 (cˆ1.25; CHCl3). NMR
(CDCl3) dH 0.8 (s, 9H, SO3–CH2–tBu), 1.4 (s, 9H, tBu),
3.1 (d, 1H, J=4.4 Hz, CH–S), 3.65 (s, 2H, SO3–CH2), 3.75
(s, 5H, S–CH2, OCH3), 4.7 (m, 1H, CH–CHyCH), 5.1 (s,
2H, C6H5–CH2), 5.7 (d, 1H, Jˆ9.3 Hz, Z–NH), 6.2–6.3 (dd,
1H, Jˆ2.0, 13.1 Hz, CH–CHyCH), 6.7–6.9 (m, 3H, CH–
CHyCH, CH arom. ortho OCH3), 7.2 (m, 2H, CH arom.
meta), 7.3 (s, 5H, C6H5–CH2). dC 24.1 (CH2–C(CH3)3),
26.1 (COO–C(CH3)3), 34.1 (S–CH2), 46.3 (CH–S), 50.5
(CH–CHyCH), 53.3 (OCH3), 65.4 (C6H5–CH2), 77.9
(COO–C(CH3)3), 81.4 (CH2–C(CH3)3), 112.3 (CH arom.
ortho OCH3), 124.6, 126.1, 126.4, 126.6, 128.3 (CH
arom., CH–CHyCH, C arom.), 142.8 (CH–CHyCH),
153.6, 157.2 (CONH, C–OCH3), 167.9 (COOtBu). HPLC
General procedure for the diastereoselective reduction
using [(Ph3P)CuH]6: procedure B. The reaction was
performed as described.13,14 To a solution of a,b-unsatu-
rated ester 4 under argon in degassed benzene (15 ml/
mmol) was added degassed water (20 equiv.) quickly
followed by the bright red [(PPh3)CuH]6 (0.36 equiv.) and
the mixture was stirred at room temperature. The reaction
was monitored by means of HPLC and quenched when
complete (3 h). After addition of ammonium chloride, the
mixture was extracted with ethyl acetate. The organic layer
was washed with brine, dried over Na2SO4 and the solvent
removed under reduced pressure.
˚
C18 Kromasil (5 m, 100 A) CH3CN/H2O (TFA) 75/25,
tRˆ17.9 min. SM (ESI), m/zˆ629.8 (MNaϩ).
(2R,3R,4E)-3-Benzyloxycarbonylamino-2-(4-methoxy-
benzylsulfanyl)-hex-4-ene-1,6-dioic acid 6-benzyl ester
1-tert-butyl ester (4b). Starting from 3 (2.9 g, 5.92 mmol)
and following the procedure A, the product was purified
by flash chromatography on silica gel eluting with
cHex:CH2Cl2:EtOAc 7:1.5:1.5 (Rfˆ0.36) affording 4b as
white crystals, 2.03 g (58%, d.e.ˆ90%). Mpˆ89.1–91.3ЊC.
Anal. calc. for C33H37NO7S C, 66.98; H, 6.30; N, 2.37.
Found C, 66.70; H, 6.45; N, 2.52. [a]Dˆϩ54.6 (cˆ1.15;
CHCl3). NMR (CDCl3) dH 1.4 (s, 9H, tBu), 3.1 (d, 1H,
Jˆ4.4 Hz, CH–S), 3.7 (2s, 5H, OCH3, S–CH2), 4.6 (m, 1H,
CH–CHyCH),5.0(s,2H, COOCH2–C6H5),5.1(s,2H, C6H5–
CH2–NH), 5.75 (d, 1H, Jˆ9.4 Hz, Z–NH), 5.85–6.0 (dd, 1H,
Jˆ1.2, 15.9 Hz, CH–CHyCH), 6.8 (m, 3H, 2CH arom. ortho
OCH3, CH–CHyCH), 7.2 (d, 2H, Jˆ8.7 Hz, CH arom. meta),
7.3 (s, 5H, C6H5–CH2–NH). dC 26.0 (COO–C(CH3)3), 33.9
(S–CH2), 46.9 (CH–S), 48.6 (CH–CHyCH), 53.3 (OCH3),
64.5 (C6H5–CH2OCO–NH), 65.1 (C6H5–CH2–OCO), 81.0
(COO–C(CH3)3), 112.2 (CH arom. ortho OCH3), 120.6,
126.1, 126.2, 126.3, 126.6, 128.3, 134.0 (CH arom., CH–
CHyCH, C arom.), 142.9 (CH–CHyCH), 153.8, 157.1
(CONH, C–OCH3), 163.6 (CH–CHyCH–COO), 167.5
General procedure for the deprotection of tert-butyl
esters: procedure C. The deprotection was achieved
by treatment with trifluoroacetic acid (15 equiv.) in
CH2Cl2 (50/50, v/v) using anisole (5 vol%) as a scavenger,
for 4 h.
(2S)-2-Benzyloxycarbonylamino-succinic acid 1-methyl
ester 4-tert-butyl ester (2). Starting from 20 g of
Z-Asp(OtBu)–OH (61.5 mmol), the methyl ester was
obtained as a yellow oil (20.7 g, 99%) via diazomethane
esterification21 using Diazogen᭨ (26.3 g, 123 mmol). Rf
(cHex:EtOAc:CH2Cl2 7:1.5:1.5)ˆ0.31. NMR (CDCl3) dH
1.3 (s, 9H, COOtBu), 2.7–2.9 (m, 2H, CH–CH2), 3.7 (s,
3H, COOCH3), 4.5 (m, 1H, CH–CH2), 5.0 (s, 2H, C6H5–
CH2), 5.7 (d, 1H, Jˆ9.5 Hz, Z–NH), 7.3 (m, 5H, C6H5–
˚
CH2). HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O
(TFA) 70/30, tRˆ6.1 min.
(2R,3R)-2-Benzyloxycarbonylamino-3-(4-methoxy-
benzylsulfanyl)-succinic acid 1-methyl ester 4-tert-butyl
ester (3). The sulfenylation of 2 (20.7 g, 61 mmol) was
performed following the strategy described in the litera-
ture.4 The crude product was then diluted with cold ether
in order to precipitate the remaining sulfenylating reagent
which was filtered off. The residue was purified by flash
chromatography on silica gel using cHex:CH2Cl2:EtOAc
8: 1:1 (Rfˆ0.27) as an eluent, giving 3 as yellow crystals
(20 g, 68%). Mpˆ71.6–73.3ЊC. [a]Dˆϩ72.1 (cˆ0.79;
CHCl3). NMR (CDCl3) dH 1.4 (s, 9H, tBu), 3.6 (s, 5H,
S–CH2, OCH3), 3.7 (s, 3H, COOCH3), 3.8 (d, 1H,
Jˆ5.4 Hz, CH–S), 4.6 (dd, 1H, Jˆ9.7, 5.4 Hz, CH–
COOCH3), 5.1 (s, 2H, C6H5–CH2), 5.9 (d, 1H, Jˆ9.7 Hz,
Z–NH), 6.8 (d, 2H, Jˆ8.7 Hz, CH arom. ortho OCH3), 7.2
(d, 2H, Jˆ8.7 Hz, CH arom. meta), 7.3 (m, 5H, C6H5–CH2).
˚
(COOtBu). HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O
(TFA) 75/25, tRˆ18.2 min. SM (ESI), m/zˆ613.9 (MNaϩ),
629.9 (MKϩ).
(2R,3R,4E)-2-Benzyloxycarbonylamino-3-[4-methoxy-
benzylsulfanyl]-5-[dibenzyloxyphosphoryl]-pent-4-enoic
acid tert-butyl ester (4c). Following the procedure A and
starting from 3 g of compound 3 (6.13 mmol), the compound
4c was obtained after purification by flash chromatography on
silica gel (eluent: cHex:CH2Cl2:EtOAc 5:2.5:2.5), (Rfˆ0.32)
giving a yellow solid, 1.9 g (44%, d.e.ˆ90%). Mpˆ74.5–
76.1ЊC. Anal. calc. for C39H43NO8SP C, 65.35; H, 6.05; N,
1.95. Found C, 65.52; H, 6.14; N, 2.07. [a]Dˆϩ72.8
(cˆ1.00; CHCl3). NMR (CDCl3) dH 1.35 (s, 9H, tBu), 3.15
(d, 1H, Jˆ4.4 Hz, CH–S), 3.7(2s,5H, OCH3, S–CH2), 4.6(m,
˚
HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O (TFA)
70/30, tRˆ14.1 min.

C. David et al. / Tetrahedron 56 (2000) 209–215
213
1H, CH–CHyCH), 4.85–5.1 (m, 6H, 3×C6H5–CH2), 5.6–5.8
(m, 2H, Jˆ1.5, 8.4, 17.3 Hz, CH–CHyCH, Z–NH), 6.5–6.7
(m, 1H, CH–CHyCH), 6.7 (d, 2H, Jˆ8.5 Hz, CH arom. ortho
OCH3), 7.1 (d, 2H, CH arom. meta), 7.1–7.4 (m, 15H,
3×C6H5). dC 26.0 (COO–C(CH3)3), 34.0 (S–CH2), 46.5
(CH–S), 52.0 (CH–CHyCH), 53.3 (OCH3), 65.1, 65.3, 65.5
(C6H5–CH2), 81.0 (COO–C(CH3)3), 112.2 (CH arom. ortho
OCH3), 126.0, 126.2, 126.4, 126.6, 128.3, 134.2 (CH arom.,
CH–CHyCH, C arom.), 147.2 (CH–CHyCH), 153.8, 157.1
(CONH, C–OCH3), 168.1 (COOtBu). HPLC C18 Kromasil
N, 2.80. Found C, 65.28; H, 6.58; N, 2.60. [a]Dˆϩ94.1
(cˆ1.07; CHCl3). NMR (CDCl3) dH 1.4 (s, 9H, tBu), 2.15
(s, 3H, COCH3), 3.15 (d, 1H, Jˆ4.5 Hz, CH–S), 3.7 (s, 5H,
S–CH2, OCH3), 4.65 (m, 1H, CH–CHyCH), 5.0 (s, 2H,
C6H5–CH2), 5.7 (d, 1H, Jˆ9.6 Hz, Z–NH), 6.0 (dd, 1H,
Jˆ1.2, 16.1 Hz, CH–CHyCH), 6.5–6.7 (dd, 1H, Jˆ4.9,
16.1 Hz, CH–CHyCH), 6.8 (d, 2H, Jˆ8.6 Hz, CH arom.
ortho OCH3), 7.1 (m, 2H, CH arom. meta OCH3), 7.2 (s,
5H, C6H5–CH2). dC 25.6 (COCH3), 26.1 (COO–C(CH3)3),
34.0 (S–CH2), 47.0 (CH–S), 51.0 (CH–CHyCH), 53.3
(OCH3), 65.2 (C6H5–CH2), 81.0 (COO–C(CH3)3), 112.2
(CH arom. ortho OCH3), 126.1, 126.2, 126.6, 128.3,
129.3, 134.2 (CH arom., CH–CHyCH, C arom.), 141.2
(CH–CHyCH), 153.8, 157.1 (CONH, C–OCH3), 168.3
(COOtBu), 195.8 (COCH3). HPLC C18 Kromasil (5 m,
˚
(5 m, 100 A) CH3CN/H2O (TFA) 75/25, tRˆ17.3 min. SM
(ESI), m/zˆ740.1 (MNaϩ).
(2R,3R,4E)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-5-[tert-butylcarbamoyl]-pent-4-enoic
acid tert-butyl ester (4d). Starting from 410 mg of 3
(0.84 mmol) and following the procedure A, the crude
product was purified by flash chromatography on silica gel
using cHex:EtOAc 7:3 as an eluent (Rfˆ0.44) giving 4d as a
yellow oil (80 mg, 17%, d.e.ˆ90%). Anal. calc. for
C30H40N2O6S C, 64.73; H, 7.24; N, 5.03. Found C, 64.89;
H, 6.90; N, 5.10. NMR (CDCl3) dH 1.3 and 1.45 (2s, 18H,
2×tBu), 3.15 (d, 1H, Jˆ4.1 Hz, CH–S), 3.75 (s, 5H, S–CH2,
OCH3), 4.6 (m, 1H, CH–CHyCH), 5.1 (s, 2H, C6H5–CH2),
5.7 (m, 1H, CONH), 5.6–5.8 (m, 1H, CH–CHyCH), 5.9 (d,
1H, Jˆ9.5 Hz, Z–NH), 6.4–6.6 (dd, 1H, Jˆ6.5, 16.4 Hz,
CH–CHyCH), 6.75 (d, 2H, Jˆ6.7 Hz, CH arom. ortho
OCH3), 7.15 (m, 2H, CH arom. meta OCH3), 7.3 (s, 5H,
˚
100 A) CH3CN/H2O (TFA) 75/25, tRˆ8.99 min. SM
(ESI), m/zˆ499.5 (MHϩ).
(2R,3R,4E)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-5-[N-methyl-N-methoxy
carbamoyl]-
pent-4-enoic acid tert-butyl ester (4g). Starting from 3
(359 mg, 0.73 mmol) and following the procedure A, the
crude product was purified by flash chromatography on silica
gel (eluent: cHex:EtOAc 7:3, Rfˆ0.05) affording 4g as a
yellow oil, 280 mg (72%, d.e.Ͼ98%). Anal. calc. for
C28H36N2O7S C, 61.75; H, 6.66; N, 5.14. Found C, 61.58; H,
6.98; N, 5.23. [a]Dˆϩ58.2 (cˆ1.49; CHCl3). NMR (CDCl3)
dH 1.4 (s, 9H, tBu), 3.15 (m, 4H, CH–S, N–CH3), 3.5 (s, 3H,
OCH3), 3.7 (s, 5H, S–CH2, OCH3), 4.65 (m, 1H, CH–
CHyCH), 5.0 (s, 2H, C6H5–CH2), 5.9 (d, 1H, Jˆ9.5 Hz,
Z–NH), 6.0 (dd, 1H, Jˆ1.4, 15.6 Hz, CH–CHyCH),
6.7–6.8 (m, 3H, CH–CHyCH, CH arom. ortho OCH3),
7.1–7.3 (m, 7H, CH arom. meta OCH3, C6H5–CH2). dC 26.0
(COO–C(CH3)3), 30.5 (N–CH3), 34.0 (S–CH2), 46.9
(CH–S), 51.7 (CH–CHyCH), 53.3 (OCH3), 59.9 (OCH3),
65.1 (C6H5–CH2), 81.0 (COO–C(CH3)3), 112.2 (CH arom.
ortho OCH3), 118.4 (CH–CHyCH), 126.1, 126.2, 126.6,
˚
C6H5–CH2). HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O
(TFA) 75/25, tRˆ10.6 min.
(2R,3R,4E)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-5-(cyano)-pent-4-enoic acid tert-butyl
ester (4e). Starting from 3 (383 mg, 0.78 mmol) and follow-
ing the procedure A, the crude product was purified by flash
chromatography on silica gel (eluent: cHex:EtOAc 4:1)
affording 4e as a yellow oil, 143 mg (38%, d.e.ˆ40%). Rf
(cHex:EtOAc 7:3)ˆ0.45. Anal. calc. for C26H30N2O5S C,
64.71; H, 6.27; N, 5.80. Found C, 65.01; H, 6.35; N, 5.52.
[a]Dˆϩ73.2 (cˆ1.26; CHCl3). NMR (CDCl3) dH 1.4 (s,
9H, tBu), 3.15 (2d, 1H, Jˆ4.6, 6.2 Hz, CH–S), 3.7 (s, 5H,
S–CH2, OCH3), 4.6 (m, 1H, CH–CHyCH), 5.0 (s, 2H,
C6H5–CH2), 5.1–5.5 (m, 1H, CH–CHyCH), 5.6 (d, 1H,
Jˆ9.2 Hz, Z–NH), 6.5 (dd, 1H, Jˆ4.9, 11.3 Hz, CH–
CHyCH), 6.75 (m, 2H, CH arom. ortho OCH3), 6.8 (m,
2H, CH arom. meta OCH3), 7.2 (s, 5H, C6H5–CH2). dC
26.1 (COO–C(CH3)3), 33.9 (S–CH2), 46.4 (CH–S), 51.4
(CH–CHyCH), 53.4 (OCH3), 65.4 (C6H5–CH2), 81.4
(COO–C(CH3)3), 100.0 (CH–CHyCH), 112.3 (CH arom.
ortho OCH3), 114.5 (CN), 126.2, 126.3, 126.4, 126.5,
126.7, 128.3 (CH arom.), 134.0 (C arom.), 148.8 (CH–
CHyCH), 153.7, 157.2 (CONH, C–OCH3), 167.0
126.7, 128.3, 134.4 (CH arom.,
C arom.), 141.2
(CH–CHyCH), 154.0, 157.1 (CONH, C–OCH3), 164.0
(CO–NCH3), 168.4 (COOtBu), 195.8 (COCH3). HPLC C18
˚
Kromasil (5 m, 100 A) CH3CN/H2O (TFA) 75/25,
tRˆ7.83 min. SM (ESI), m/zˆ545.3 (MHϩ).
(2R,3R,4E,Z)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-5-phenyl-pent-4-enoic acid tert-butyl
ester (4h). To a solution of benzyl triphenylphosphonium
bromide (442 mg, 1.02 mmol) in dry THF (5 ml) was added
dropwise, at Ϫ78ЊC, n-butyllithium (0.44 ml of a 2.5M hex-
ane solution, 1.1 mmol). After stirring for 30 min at this
temperature, a solution of 3 (250 mg, 0.51 mmol) in dry
THF (1 ml) was added, immediately followed by a dropwise
addition of Dibal-H (0.37 ml of a toluene 1.5M solution,
0.56 mmol). The resulting mixture was stirred 3 h at
Ϫ78ЊC, then warmed at room temperature and stirred for
1 h. 2N HCl (2 ml) and water (1 ml) were added, the organic
layer was separated and the aqueous phase was extracted
with ether (50 ml). The combined organic fractions were
washed with brine (10 ml), dried (Na2SO4), filtered and
the solvent was removed in vacuo. The product was purified
by flash chromatography on silica gel eluting with
cHex:CH2Cl2:EtOAc 7:1.5:1.5 (Rfˆ0.48) affording 4h as a
yellow oil, 164 mg (60%, d.e.ˆ90%, E/Zˆ2/1). Anal. calc.
for C31H35NO5S C, 69.77; H, 6.61; N, 2.62. Found C, 70.14;
˚
(COOtBu). HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O
(TFA) 75/25, tRˆ9.6 and 9.9 min. SM (ESI), m/zˆ483.1
(MHϩ).
(2R,3R,4E)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-6-oxo-hept-4-enoic acid tert-butyl ester
(4f). Starting from 3 (338 mg, 0.69 mmol) and following
the procedure A, the crude product was purified by flash
chromatography on silica gel (eluent: cHex:EtOAc 7:3,
Rfˆ0.32) affording 4f as a yellow oil, 175 mg (51%,
d.e.ˆ98%). Anal. calc. for C27H33NO6S C, 64.91; H, 6.66;

214
C. David et al. / Tetrahedron 56 (2000) 209–215
H, 6.81; N, 2.54. NMR (CDCl3) dH 1.4 (s, 9H, tBu), 3.15 (d,
1H, Jˆ4.2 Hz, CH–S), 3.7 (m, 5H, S–CH2, OCH3), 4.65 (m,
1H, CH–CHyCH), 5.1 (s, 2H, C6H5–CH2), 5.7 (d, 1H,
Jˆ8.2 Hz, Z–NH), 6.0 (dd, 1H, Jˆ6.1, 14.5 Hz, CH–
CHyCH), 6.45 (dd, 1H, Jˆ1.2, 14.5 Hz, CH–CHyCH),
6.7 (m, 2H, CH arom. ortho OCH3), 7.0–7.4 (m, 12H, CH
arom. meta, 2×C6H5–CH2). HPLC C18 Kromasil (5 m,
(80 h). Finally, 1.96 g of dipotassium azodicarboxylate
(3.36 mmol) and 384 ml of acetic acid (6.72 mmol) were
required. After addition of water, the product was extracted
with diethyl oxide. The organic layer was washed with
NaHCO3, brine, dried over Na2SO4 and evaporated to
dryness, giving a yellow oil, 100 mg (100%), which was
not further purified. NMR (CDCl3) dH 1.4 (s, 9H, tBu),
1.6–1.9 (m, 2H, CH–CH2–CH2), 2.4–2.7 (m, 2H, CH–
CH2–CH2), 3.1 (d, 1H, CH–S), 3.7 (m, 5H, S–CH2,
OCH3), 4.0 (m, 1H, CH–CH2–CH2), 5.1 (s, 2H, C6H5–
CH2), 5.5 (d, 1H, Jˆ9.6 Hz, Z–NH), 6.7 (m, 2H, CH
arom. ortho OCH3), 7.0–7.4 (m, 12H, CH arom. meta,
˚
100 A) CH3CN/H2O (TFA) 75/25, tRˆ21.7 (E) and 23.0
(Z) min.
(2R,3R)-3-Benzyloxycarbonylamino-2-[4-methoxybenzyl-
sulfanyl]-5-[2,2-dimethylpropanoxy sulfonyl] pentanoic
acid tert-butyl ester (5a). Starting from 4a (12 g,
19.7 mmol) and following the general procedure B, the
crude product was purified on silica gel (eluent: cHex:
EtOAc:CH2Cl2 7:1.5:1.5, Rfˆ0.1) affording predominantly
the stereoisomer (2R,3R)-5a, 8.99 g (78%, d.e.ˆ60%). NMR
(CDCl3) dH 0.9 (s, 9H, CH2-tBu), 1.8–2.2 (m, 2H, CH–CH2–
CH2), 3.1 (m, 3H, CH–S, CH–CH2–CH2), 3.7 (s, 7H, S–CH2,
OCH3, SO3–CH2), 3.9 (m, 1H, CH–CH2–CH2), 5.1 (m, 2H,
C6H5–CH2), 5.6 (d, 1H, Jˆ9.5 Hz, Z–NH), 6.8 (d, 2H,
Jˆ8.7 Hz, CH arom. ortho OCH3), 7.2 (d, 2H, Jˆ8.7 Hz,
CH arom. meta), 7.3 (s, 5H, C6H5–CH2). HPLC C18 Kromasil
˚
2×C6H5–CH2). HPLC C18 Kromasil (5 m, 100 A) CH3CN/
H2O (TFA) 75/25, tRˆ23.6 min.
(2R,3R)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-5-[2,2-dimethylpropanoxy sulfonyl]-
pentanoic acid (1a). Starting from 5a (10 g, 16.4 mmol),
following the procedure C, and after purification by flash
chromatography on silica gel (cHex:Et2O:HCOOH 4:6:0.1;
Rfˆ0.4) the pure product was obtained as a red foam, 6.1 g
(69%). NMR (CDCl3) dH 0.8 (s, 9H, SO3–CH2–tBu), 1.8–
2.2 (m, 2H, CH–CH2–CH2), 3.1 (m, 1H, CH–S), 3.1 (t, 2H,
Jˆ6.9 Hz, CH–CH2–CH2), 3.65 (s, 2H, SO3–CH2), 3.75 (s,
5H, S–CH2, OCH3), 5.1 (s, 2H, C6H5–CH2), 5.7 (d, 1H,
Jˆ9.5 Hz, Z–NH), 6.7–6.9 (m, 2H, CH arom. ortho
OCH3), 7.2 (d, 2H, Jˆ8.6 Hz, CH arom. meta), 7.3 (s, 5H,
˚
(5 m, 100 A) CH3CN/H2O (TFA) 75/25, tRˆ17.5 and
18.4 min.
(2R,3R)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-hexan-1,6-dioic acid, 6-benzyl ester
1-tert-butyl ester (5b). Compound 5b was obtained from
1.4 g of 4b (2.3 mmol) following the procedure B. After
purification by flash chromatography on silica gel (eluent:
cHex:EtOAc:CH2Cl2 7:1.5:1.5, Rfˆ0.3) a yellow oil was
obtained, 1.2 g (87%). NMR (CDCl3) dH 1.4 (s, 9H, tBu),
1.6–2.0 (m, 2H, CH–CH2–CH2), 2.3 (t, 2H, Jˆ7.2 Hz,
CH–CH2–CH2), 3.1 (m, 1H, CH–S), 3.7 (s, 5H, S–CH2,
OCH3), 3.9 (m, 1H, CH–CH2–CH2), 5.1 (2s, 4H,
2×C6H5–CH2), 5.5 (d, 1H, Jˆ9.5 Hz, Z–NH), 6.8 (d, 2H,
Jˆ8.7 Hz, CH arom. ortho OCH3), 7.2 (d, 2H, Jˆ8.7 Hz,
CH arom. meta), 7.3 (s, 10H, 2×C6H5–CH2). HPLC C18
˚
C6H5–CH2). HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O
(TFA) 75/25, tRˆ5.6 min.
(2R,3R)-3-Benzyloxycarbonylamino-2-[4-methoxybenzyl-
sulfanyl]-6-benzyl ester hexanoic acid (1b). Compound 1b
was obtained from 1.0 g of 5b (1.68 mmol) according to the
procedure C. After purification by flash chromatography on
silica gel (eluent: cHex:Et2O:HCOOH 4:6:0.1, Rfˆ0.31),
540 mg (60%) of 1b were obtained. NMR (CDCl3) dH 1.6–
2.0 (m, 2H, CH–CH2–CH2), 2.4 (t, 2H, Jˆ7.1 Hz, CH–CH2–
CH2), 3.25 (m, 1H, CH–S), 3.75 (s, 5H, S–CH2, OCH3), 4.05
(m, 1H, CH–CH2–CH2), 5.0 (2s, 4H, 2×C6H5–CH2), 5.4 (d,
1H, Jˆ9.5 Hz, Z–NH), 6.75 (2, 2H, Jˆ8.6 Hz, CH arom.
ortho OCH3), 7.2 (d, 2H, Jˆ8.6 Hz, CH arom. meta), 7.3
˚
Kromasil (5 m, 100 A) CH3CN/H2O (TFA) 75/25, tRˆ17.2
and 17.3 min.
˚
(m, 10H, 2×C6H5–CH2). HPLC C18 Kromasil (5 m, 100 A)
(2R,3R)-3-Benzyloxycarbonylamino-2-[4-methoxybenzyl-
sulfanyl]-5-[dibenzylphosphoryl]pentanoic acid tert-butyl
ester (5c). Following the procedure B and starting from 2.1 g
(2.9 mmol) of 4c, compound 5c was obtained after puri-
fication by flash chromatography on silica gel (eluent:
cHex:EtOAc:CH2Cl2 5:2.5:2.5, Rfˆ0.13) giving a brown
oil, 1.7 g (81%). NMR (CDCl3) dH 1.4 (s, 9H, tBu), 1.5–
2.0 (m, 4H, CH–CH2–CH2, CH–CH2–CH2), 3.1 (m, 1H,
CH–S), 3.7 (s, 5H, S–CH2, OCH3), 3.9 (m, 1H, CH–CH2–
CH2), 4.8–5.1 (m, 6H, 3×C6H5–CH2), 5.5 (d, 1H, Jˆ9.4 Hz,
Z–NH), 6.7 (d, 2H, Jˆ8.4 Hz, CH arom. ortho OCH3), 7.1
(d, 2H, Jˆ8.4 Hz, CH arom. meta), 7.3 (s, 15H, 3×C6H5–
CH3CN/H2O (TFA) 75/25, tRˆ4.2 min.
(2R,3R)-3-Benzyloxycarbonylamino-2-[4-methoxybenzyl-
sulfanyl]-5-[dibenzylphosphoryl]pentanoic acid (1c).
Compound 1c was obtained from 1.6 g of 5c (2.2 mmol)
according to the procedure C. After purification by flash
chromatography on silica gel (eluent: Et2O:HCOOH 10:0.1,
Rfˆ0.2), 974 mg (70 %) of 1c were obtained. NMR (CDCl3)
dH 1.6–2.0 (m, 4H, CH–CH2–CH2, CH–CH2–CH2), 3.75 (m,
7H, CH–S, S–CH2, OCH3, CH–CH2–CH2), 4.8–5.1 (m, 6H,
3×C6H5–CH2), 5.8 (d, 1H, Jˆ9.5 Hz, Z–NH), 6.7 (CH arom.
ortho OCH3), 7.2–7.4 (m, 17H, CH arom. meta, 3×C6H5–
˚
˚
CH2). HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O (TFA)
CH2). HPLC C18 Kromasil (5 m, 100 A) CH3CN/H2O (TFA)
75/25, tRˆ15.3 and 16.1 min.
75/25, tRˆ4.4 min.
(2R,3R)-3-Benzyloxycarbonylamino-2-[4-methoxy-
benzylsulfanyl]-5-phenylpentanoic acid tert-butyl ester
(5h). To a solution of 4h (100 mg, 0.19 mmol) in ethanol
(2.5 ml), CH2Cl2 (1 ml) and methanol (0.5 ml) was added at
0ЊC, dipotassium azodicarboxylate salt followed by acetic
acid until the reaction, monitored by HPLC, was complete
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