Total synthesis of stevastelins B3 and C3: Structure confirmation of stevastelin B3 and revision of stevastelin C3

Article abstract of DOI:10.1016/j.tetlet.2004.11.121

The total synthesis of stevastelin B3, stevastelin C3 and 5-deoxy derivative of stevastelin C3, novel 13-membered cyclic depsipeptides, is described. This study unambiguously confirmed the proposed absolute structure of stevastelin B3, and revealed that t

Full text of DOI:10.1016/j.tetlet.2004.11.121

Tetrahedron
Letters
Tetrahedron Letters 46 (2005) 389–392
Total synthesis of stevastelins B3 and C3: structure confirmation
of stevastelin B3 and revision of stevastelin C3
Kazuo Kurosawa, Keigo Matsuura and Noritaka Chida*
Department of Applied Chemistry, Faculty of Science and Technology, Keio University, Hiyoshi,
Kohoku-ku, Yokohama 223-8522, Japan
Received 22 October 2004; revised 11 November 2004; accepted 24 November 2004
Abstract—The total synthesis of stevastelin B3, stevastelin C3 and 5-deoxy derivative of stevastelin C3, novel 13-membered cyclic
depsipeptides, is described. This study unambiguously confirmed the proposed absolute structure of stevastelin B3, and revealed that
the structure of stevastelin C3 is incorrect. The correct structure of stevastelin C3 was established by the total synthesis to be 5-deoxy
derivative of the proposed structure.
Ó 2004 Elsevier Ltd. All rights reserved.
Stevastelins B3 (1) and C3 (2), members of stevastelin
family, are novel cyclic depsipeptides isolated from a
culture broth of Penicillium by Nippon Kayaku group
and reported to show a potent immunosuppressive
activity.1,2 The structural study by spectral, degradation
and synthetic methods of stevastelin B (3), the most
abundant congener of stevastelin family, established
that stevastelin B consists of (2S,3S,4S,5R)-3,5-dihy-
droxy-2,4-dimethylstearic acid, L-serine, L-threonine
and L-valine, and possesses 15-membered ring struc-
ture.3 Based on spectroscopic analyses of other stevast-
elins, it has been proposed that stevastelin B3 (1) is an
isomer of stevastelin B with a 13-membered ring struc-
ture and stevastelin C3 (2) is a de-O-acetyl derivative
of stevastelin B3.1 Their interesting mode of action,
repression of both T cells and B cells,1 as well as their
unique structures have attracted the synthetic attention,
and total syntheses of stevastelins B4 and the proposed
structure of C3,5 synthetic approach6 and preparation
and biological assessment of simple analogues7 have
been reported to date. Although the proposed absolute
structure of stevastelin B has been determined by the
total synthesis,4b those of stevastelins B3 and C3 have
not been synthetically confirmed. Here we report a total
synthesis of stevastelins B3 (1), C3 (2) and a 5-deoxy
derivative of stevastelin C3 (4), which resulted in the
structure confirmation of the proposed absolute struc-
ture of stevastelin B3 (1) and the structure revision of
stevastelin C3 to 4.
1
3
n-C13H27
5
O
4
2
HO
O
O
HN
RO
R = Ac: stevastelin B3 (1)
NH HN
O
R = H: stevastelin C3 (2) (proposed)
H
O
OH
5
n-C13H27
O
1
O
3
n-C13H27
O
5
4
2
O
O
HN
OH
O
HN
O
val
HO
H
ser
H
NH HN
O
AcO
N
N
H
H
thr
O
O
OH
OH
stevastelin B (3)
stevastelin C3 (4) (revised)
Synthesis of stevastelin B3 commenced from acetonide
6, which was previously prepared from L-quebrachitol
(5) in our total synthesis of stevastelin B4b (Scheme 1).
Benzylation of the hydroxy group in 6, followed by acid
treatment gave diol 7.8 The primary hydroxy group in 7
was selectively protected as a TES ether to give 8 in 96%
yield from 6. Since direct acylation of 8 with Boc-Val-
Thr-Ser(Bzl)9 under various reaction conditions was
found to be fruitless due to the steric congestion of the
hydroxy group, stepwise introduction of the peptide
Keywords: Stevastelin B3; Stevastelin C3; Total synthesis; Structure
revision.
*
Corresponding author. Tel./fax: +81 45 566 1573; e-mail: chida@
applc.keio.ac.jp
0040-4039/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2004.11.121

390
K. Kurosawa et al. / Tetrahedron Letters 46 (2005) 389–392
OMe
OH
n-C13H27
BzlO
n-C13H27
CH2OTES
n-C13H27
e
HO
HO
OH
OH
a
b, c
O
1'
OBzl OH OR
OH
O
O
O
7
8
R = H
R = TES
BzlO
d
2'
NHCbz
6
L
-quebrachitol (5)
9
n-C13H27
O
H
n-C13H27
BzlO
5
R1
R2HN
NH HN
f
l
j
RO
O
O
HN
HN
stevastelin B3 (1)
O
O
1'
BocHN
BzlO
RO
NH
O
2'
H
N
H
HO2C
O
H
O
O
OH
O
OH
OH
11 R1 = CH2OTES, R2 = Boc
12 R1 = CH2OH, R2 = Boc
10
g
14 R = Bzl
R = H (proposed structure of stevastelin C3)
k
2
h, i
13 R1 = CO2H, R2 = H•CF3CO2H
Scheme 1. TES = –SiEt3, Bzl = –CH2Ph, Cbz = –C(O)OCH2Ph. Reagents and conditions: (a) see Ref. 4b; (b) (Me3Si)2NK, BzlBr, THF; (c) AcOH–
H2O (4/1); (d) TESCl, Et3N, CH2Cl2; (e) Cbz–Ser(Bzl), 2,4,6-trichlorobenzoyl chloride, Et3N, DMAP, THF; (f) H2, 10% Pd–C ethylenediamine
complex, MeOH, rt, then 10, WSCÆHCl, HOBt, DMF, rt; (g) AcOH–THF–H2O (3/3/1); (h) TEMPO, KBr, NaOCl, NaHCO3, aq CH2Cl2, 0 °C, then
NaClO2, HOSO2NH2, NaH2PO4, t-BuOH–H2O, 0 °C; (i) TFA, CH2Cl2, ꢀ18 °C; (j) DEPC, Et3N, DMF; (k) H2, 10% Pd–C, MeOH; (l) Ac2O,
pyridine, rt.
moiety was carried out. Condensation of 8 with Cbz–
Ser(Bzl) under the modified Yamaguchiꢀs conditions10
afforded 9. Unfortunately racemization of the serine
moiety during the coupling process was observed and
9 and its diastereoisomer were obtained as an insepar-
able mixture in a ratio of ca. 1:1 in 80% yield. Hydrogen-
olysis of a mixture of 9 and its diastereomer in the
presence of 10% Pd–C ethylenediamine complex11
deprotected N-Cbz group selectively to give an amine,
which, without isolation, was coupled with Boc-Val-
Thr 109 by the action of ethyl-3-(3-dimethylaminoprop-
yl)-carbodiimide hydrochloride (WSCÆHCl) and 1-
hydroxybenzotriazole (HOBt) to provide 11 and its dia-
stereomer in 94% yield. Acid hydrolysis of 11 gave diol
12. At this stage, diastereomers were cleanly separated
by silica gel chromatography and 12 was obtained in
pure form in 45% yield from 11. The primary hydroxy
group in 12 was selectively oxidized to give a carboxylic
acid, whose N-Boc group was deprotected to generate
13. Macrolactamization of 13 was successfully accom-
plished under Shioiriꢀs protocol12 [(diethyl phosphoro-
cyanidate (DEPC) in DMF (0.01 mol dmꢀ3)] to give
macrocycle 14 in 29% yield from 12. Removal of the
O-benzyl group in 14 by hydrogenolysis afforded the
compound possessing the proposed structure of stevast-
elin C3 213,14 in 74% yield. Treatment of 2 with acetic
anhydride in pyridine at room temperature furnished
stevastelin B3 (1)14 (38% yield).
To elucidate the correct structure of stevastelin C3, the
natural product was subjected to degradation (Scheme
2). Treatment of natural stevastelin C3 with LiBH4
afforded diol 1514 in 24% yield. The structure of 15
was determined based on spectral analysis, and finally
confirmed by comparison with the synthetic specimen
(vide infra). On the other hand, acid hydrolysis of the
natural product, followed by HPLC analysis (MIC
GEL CRS 10W) of the hydrolysates revealed that the
amino acids constituting stevastelin C3 are the same as
those of stevastelins B and B3 (L-serine, L-threonine
and L-valine). From these results, it was presumed that
natural stevastelin C3 should be a 5-deoxy derivative
of the proposed structure.
stevastelin C3
(natural)
6
a
b, c, d
n-C13H27
BzlO
CH2OTBS
e, f
O
O
1
3
n-C13H27
5
1'
4
2
CH2OH
NHCbz
2'
OH
15
16
g, h
n-C13H27
O
H
5
n-C13H27
BzlO
R1
O
O
O
O
O
HN
R2HN
k
RO
NH HN
O
NH HN
O
OH
H
O
OH
The direct comparison of synthetic 1 and 2 with natural
stevastelins B3 and C3, kindly provided by Nippon
Kayaku group revealed that the synthetic 1 is unambig-
uously identical with natural stevastelin B3. Thus, the
proposed structure of stevastelin B3 was fully confirmed
by this first total synthesis. However, spectral data of
synthetic 2 were not identical with those of natural ste-
vastelin C3. Further analysis of natural stevastelin C3
by NMR suggested the possibility that the natural prod-
uct might be a deoxy derivative of the proposed struc-
ture. The molecular ion peak observed at m/z 598
(C32H59N3O7 + H) by FABMS also supported the deoxy-
genated structure.
19 R = Bzl
R = H
17 R1 = CH2OH, R2 = Boc
18 R1 = CO2H, R2 = H•CF3CO2H
l
i, j
4
revised structure
of stevastelin C3
Scheme 2. TBS = –SiMe2 (t-Bu). Reagents and conditions: (a) LiBH4,
THF, reflux; (b) NaH, CS2, THF, then MeI; (c) Bu3SnH, AIBN,
toluene, reflux; (d) CSA, MeOH, rt; (e) TBSCl, Et3N, CH2Cl2; (f) Cbz–
Ser(Bzl), 2,4,6-trichlorobenzoyl chloride, Et3N, DMAP, THF, 0 °C;
(g) H2, 10% Pd–C ethylenediamine complex, MeOH, rt, then 10,
WSCÆHCl, HOBt, DMF; (h) AcOH–THF–H2O (14/3/6); (i) TEMPO,
KBr, NaOCl, NaHCO3, aq CH2Cl2, 0 °C, then NaClO2, HOSO2NH2,
NaH2PO4, t-BuOH–H2O, 0 °C; (j) TFA, CH2Cl2, ꢀ18 °C; (k) DEPC,
Et3N, DMF; (l) H2, 10% Pd–C, MeOH.

K. Kurosawa et al. / Tetrahedron Letters 46 (2005) 389–392
391
6. (a) Chakraborty, T. K.; Ghosh, S.; Dutta, S. Tetrahedron
Lett. 2001, 42, 5085–5088; (b) Sarabia, F.; Chammaa, S.;
With these structural information, we turned to the syn-
thesis of the expected structure of stevastelin C3. The
hydroxy group in 6 was removed via xanthate ester by
Bartonꢀs method,15 and the product was treated with
acid to give diol 15 in 73% yield from 6. The spectral
data of 15 were fully identical with those of the diol ob-
tained by reductive degradation of natural stevastelin
C3. The primary hydroxy group in 15 was selectively
protected as a TBS ether to give secondary alcohol,
which was then condensed with Cbz–Ser(Bzl) under
the modified Yamaguchiꢀs conditions to give 16 and its
C-20 epimer as an inseparable mixture in a ratio of
1.5:1 in 90% yield. Deprotection of N-Cbz group in 16
and subsequent condensation with 10, followed by
deprotection of O-TBS group gave 17 and its diastereo-
mer, which were cleanly separated by silica gel chroma-
tography to afford pure 1713 in 37% from 15. Compound
17 was transformed into macrocycle 19 in 28% overall
yield by the same procedure as described for preparation
of 14 from 12. Removal of the O-benzyl group in 19 fur-
nished 4,14 whose spectral data as well as [a]D value
showed good accordance with those of natural stevast-
elin C3. Based on this synthesis, it was concluded that
the structure of natural stevastelin C3 should be revised
to 4.
´
Ruiz, A. S.; Lopez-Herrera, F. J. Tetrahedon Lett. 2003,
44, 7671–7675.
7. Hamaguchi, T.; Masuda, A.; Morino, T.; Osada, H.
Chem. Biol. 1997, 4, 279–286.
8. All new compounds described in this paper were charac-
terized by 300 MHz 1H NMR, 75 MHz 13C NMR, IR and
mass spectrometric and/or elemental analyses.
9. These peptides were prepared by condensation (WSCÆHCl,
HOBt, DMF) of appropriate protected amino acids which
were purchased from Peptide Institute, Inc. (Osaka,
Japan).
10. (a) Hikota, M.; Tone, H.; Horita, K.; Yonemitsu, O. J.
Org. Chem. 1990, 55, 7–9; (b) Inanaga, J.; Hirata, K.;
Saeki, H.; Katsuki, T.; Yamaguchi, M. Bull. Chem. Soc.
Jpn. 1979, 52, 1989–1993.
11. Sajiki, H.; Hattori, K.; Hirota, K. J. Org. Chem. 1998, 63,
7990–7992.
12. (a) Shioiri, T.; Yokoyama, Y.; Kasai, Y.; Yamada, S.
Tetrahedron 1976, 32, 2211–2217; (b) Yamada, S.; Kasai,
Y.; Shioiri, T. Tetrahedron Lett. 1973, 14, 1595–1598; (c)
Takuma, S.; Hamada, Y.; Shioiri, T. Chem. Pharm. Bull.
1982, 30, 3147–3153.
13. The absolute structures of amino acids in this compound
were confirmed by acid hydrolysis, followed by HPLC
analyses (MIC GEL CRS 10W, Mitsubishi Chemical
Industries, Ltd) of the hydrolysates.
25:5
14. Spectral data of compound 2: ½aꢁD ꢀ 39 (c 0.1, MeOH); 1H
In summary, total synthesis of stevastelins B3 and C3,
which fully established the absolute structures of these
natural products has been achieved. The methodology
developed in this work would be applicable to the syn-
thesis of other stevastelins as well as other cyclic depsi-
peptides. It is also interesting that the fatty acid
moiety of stevastelin C3 is different from that of stevast-
elins B and B3 although the stevastelins are produced by
the same microorganism.
NMR (300 MHz, DMSO-d6) d 0.52 (3H, d, J = 6.9 Hz),
0.86 (3H, t, J = 6.9 Hz), 0.88 (3H, d, J = 6.0 Hz), 0.93 (3H,
d, J = 6.6 Hz), 1.07 (3H, d, J = 6.3 Hz), 1.09 (3H, d,
J = 7.2 Hz), 1.14–1.41 (24H, m), 1.66 (1H, m), 2.04 (1H,
m), 2.86 (1H, m), 3.54 (1H, m), 3.85 (1H, m), 3.92 (1H, m),
4.03 (2H, m), 4.13 (1H, dd, J = 9.9 and 2.4 Hz), 4.31
(1H, d, J = 5.7 Hz), 4.60 (1H, m), 4.91 (1H, m), 5.03 (1H,
dd, J = 4.8 and 5.1 Hz), 5.14 (1H, d, J = 4.8 Hz), 7.54
(1H, d, J = 9.9 Hz), 7.84 (1H, d, J = 9.9 Hz) and
7.88 (1H, d, J = 9.9 Hz); HRMS (FAB) m/z 614.4407,
calcd for C32H60N3O8 (M+H) 614.4380. Spectral data
26
of compound 1: ½aꢁD ꢀ 53 (c 0.1, CHCl3), {natural stevast-
25:5
elin B3, ½aꢁD ꢀ 51 (c 0.255, CHCl3) (measured in
Acknowledgements
1
our laboratory)}; H NMR (300 MHz, DMSO-d6) d 0.54
We thank Dr. T. Nishikiori (Pharmaceutical Group,
Nippon Kayaku Co., Ltd, Tokyo, Japan) for providing
us with natural stevastelins B3 and C3. This work was
supported by Grants-in-Aid for the 21st Century COE
program ꢁKEIO LCCꢀ from the Ministry of Education,
Culture, Sports, Science and Technology, Japan.
(3H, d, J = 6.8 Hz), 0.85 (3H, t, J = 6.6 Hz), 0.90 (3H, d,
J = 6.8 Hz), 0.93 (3H, d, J = 6.6 Hz), 1.08 (3H, d, J = 6.1
Hz), 1.12 (3H, d, J = 6.8 Hz), 1.17–1.35 (24H, m), 1.72
(1H, m), 2.01 (3H, s), 2.05 (1H, m), 2.90 (1H, m), 3.84 (1H,
m), 4.01 (2H, m), 4.13 (1H, m), 4.18 (1H, m), 4.35 (1H, m),
4.40 (1H, d, J = 5.4 Hz), 4.92 (2H, m), 5.24 (1H, d,
J = 4.4 Hz), 7.50 (1H, m), 7.67 (1H, br) and 7.72 (1H, d,
J = 9.0 Hz) ; 13C NMR (75 MHz, DMSO-d6) d 9.2, 13.8,
13.9, 19.0, 19.2, 20.7, 20.9, 22.1, 25.7, 28.7, 29.0, 29.1 and
29.1, 31.3, 34.9, 39.1, 41.1, 50.2, 59.4, 61.6, 63.2, 65.2, 69.0,
80.2, 168.7, 170.1, 170.5, 170.7 and 170.8; HRMS (FAB)
m/z 656.4494, calcd for C34H62N3O9 (M+H) 656.4486.
The 1H and 13C NMR data were fully identical with those
References and notes
1. Morino, T.; Masuda, A.; Yamada, M.; Nishimoto, M.;
Nishikiori, T.; Saito, S.; Shimada, N. J. Antibiot. 1994, 47,
1341–1343.
2. Morino, T.; Shimada, K.-i.; Masuda, A.; Nishimoto, M.;
Saito, S. J. Antibiot. 1996, 49, 1049–1051.
3. (a) Morino, T.; Shimada, K.-i.; Masuda, A.; Yamashita,
N.; Nishimoto, M.; Nishikiori, T.; Saito, S. J. Antibiot.
1996, 49, 564–568; (b) Shimada, K.-i.; Morino, T.;
Masuda, A.; Sato, M.; Kitagawa, M.; Saito, S. J. Antibiot.
1996, 49, 569–574.
4. (a) Kohyama, N.; Yamamoto, Y. Synlett 2001, 694–696;
(b) Kurosawa, K.; Nagase, T.; Chida, N. Chem. Commun.
2002, 1280–1281.
of natural stevastelin B3.2 Spectral data of compound 15:
21
½aꢁD þ 10 (c 1.74, CHCl3); mmax (neat) 3320 cmꢀ1
;
1H NMR (CDCl3, 300 MHz) d 0.79 (3H, d, J = 7.1 Hz),
0.86 (3H, d, J = 7.1 Hz), 0.87 (3H, t, J = 7.0 Hz), 1.15–
1.42 (24 H, m), 1.60 (3 H, m), 1.85 (1H, dddq, J = 9.0, 7.8,
3.4 and 7.1 Hz), 2.12–3.18 (2H, m), 3.46 (1H, dd, J = 9.0
and 2.4 Hz), 3.63 (1H, dd,J = 10.7 and 7.8 Hz) and
3.71 (1H, dd, J = 10.7 and 3.4 Hz); 13C NMR (CDCl3,
75 MHz) d 12.2, 13.5, 14.1, 22.7, 27.4, 29.4, 29.7, 29.9,
31.9, 34.0, 35.1, 37.3, 68.8 and 80.3; HRMS (FAB) m/z
315.3283, calcd for C20H43O2 (M+H) 315.3263. Spectral
25
´
5. Sarabia, F.; Chammaa, S.; Lopez-Herrera, F. J. Tetrahe-
dron Lett. 2002, 43, 2961–2965.
data of compound 4: ½aꢁ ꢀ 67 (c 0.13, MeOH), {natural
stevastelin C3,½aꢁD ꢀD66 (c 0.305, MeOH) (measured in
27:5

392
K. Kurosawa et al. / Tetrahedron Letters 46 (2005) 389–392
1
our laboratory)}; H NMR (300 MHz, DMSO-d6) d 0.59
(3H, d, J = 6.8 Hz), 0.85 (3H, t, J = 6.8 Hz), 0.87 (3H, d,
J = 6.3 Hz), 0.92 (3H, d, J = 6.6 Hz), 1.07 (3H, d, J = 6.3
Hz), 1.10 (3H, d, J = 7.6 Hz), 1.13–1.30 (25H, m), 1.49
(1H, m), 1.74 (1H, m), 2.05 (1H, m), 2.74 (1H, m), 3.54
(1H, m), 3.92 (1H, m), 4.03 (2H, m), 4.13 (1H, dd, J = 10.3
and 2.4 Hz), 4.61 (2H, m), 5.04 (1H, dd, J = 5.1 and 4.9
Hz), 5.16 (1H, d, J = 4.6 Hz), 7.52 (1H, m) and 7.85 (2 H,
m); 13C NMR (75 MHz, DMSO-d6) d 13.9, 15.4, 18.9,
19.3, 21.0, 22.1, 25.9, 28.7, 28.9, 29.0, 29.2, 31.3, 32.3, 34.4,
42.1, 53.7, 59.5, 61.0, 65.2, 80.5, 170.0, 170.4 and 170.8;
HRMS (FAB) m/z 598.4422, calcd for C32H60N3O7
1
(M+H) 598.4431. The H and 13C NMR data were fully
identical with those of natural stevastelin C3.
15. Barton, D. H. R.; McCombie, S. W. J. Chem. Soc., Perkin
Trans. 1 1975, 1574–1585.