Total synthesis of apoptolidin

Article abstract of DOI:10.1002/anie.200460172

A ring-size-selective macrolactonization and the early introduction of the saccharide portions are the main features of a total synthesis of the 20-membered macrolide apoptolidin (see formula), which induces apoptosis in rat glia cells transformed with oncogenes.

Full text of DOI:10.1002/anie.200460172

Angewandte
Chemie
Natural Products Synthesis
Total Synthesis of Apoptolidin**
Hermut Wehlan, Mario Dauber,
M.-Teresa Mujica Fernaud, Julia Schuppan,
Rainer Mahrwald, Burkhard Ziemer,
M.-Elisa Juarez Garcia, and Ulrich Koert*
Apoptosis inducers specific for cancer cells are of interest for
tumor therapy.[1] Apoptolidin (1), isolated from Nocardiop-
[*] Dipl.-Chem. H. Wehlan, M. Dauber, Dr. M.-T. Mujica Fernaud,
Dr. J. Schuppan, Dipl.-Chem. M.-E. Juarez Garcia, Prof. Dr. U. Koert
Fachbereich Chemie
Philipps-Universität Marburg
Hans-Meerwein-Strasse, 35032 Marburg (Germany)
Fax: (+49)6421-282-5677
E-mail: koert@chemie.uni-marburg.de
Priv.-Doz. Dr. R. Mahrwald, Dr. B. Ziemer
Institut für Chemie
Humboldt-Universität zu Berlin
Brook-Taylor-Strasse 2, 12489 Berlin (Germany)
[**] This work was supported by Schering AGBerlin and the Fonds der
Chemischen Industrie. H.W. thanks the Schering Forschungsge-
sellschaft for a predoctoral fellowship. The authors thank Prof.
Dr. U. Eder for his helpful support.
Supporting information for this article is available on the WWW
under http://www.angewandte.org or from the author.
Angew. Chem. Int. Ed. 2004, 43, 4597 –4601
DOI: 10.1002/anie.200460172
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Communications
tion. The starting point for the synthesis of the O27
disaccharide portion was l-rhamnose (8) which was trans-
formed into the benzylidene-protected methyl acetal 9
(Scheme 2). Treatment of 9 with 6 equiv of methyllithium[10]
sis sp. by Hayakawa et al., selectively induces apoptosis in rat
glia cells transformed with the E1A oncogene (IC50
=
11 ngmLꢀ1) but not in untransformed cell lines.[2,3] The
apoptotic activity of 1 was correlated with its inhibition of
mitochondrial F0-F1-ATPase.[4] Apoptolidin (1) is a 20-
membered macrolactone with a side chain containing a
cyclic six-membered hemiketal. 6-Deoxy-4-o-methyl-l-glu-
cose (3) is attached to O9, and a disaccharide consisting of l-
olivomycose (4) and d-oleandrose (5) is linked to O27. The
sugar residues are a prerequisite for the potent bioactivity of
1. Treatment of 1 with MeOH/Amberlyst-15 produces 21-o-
methylapoptolidin (2).[5] Apoptolidin can rearrange into the
21-membered macrolactone isoapoptolidin by a O19!O20
acyl shift.[6] The noticeable biological activity of apoptolidin
(1) and its structural challenges make it a prominent synthetic
target.[7,8] In continuation of our synthesis of the aglycon
apoptolidinone[9] we communicate here the completion of the
total synthesis of apoptolidin.
Our synthetic strategy for apoptolidin (Scheme 1) is based
on an early introduction of the sugar residues and a late cross-
coupling of the fully glycosylated southern half 6 with the
glycosylated northern half 7 followed by the macrolactoniza-
Scheme 2. a) 1. MeOH, DOWEX 50WX-8-200; 2. PhCH(OMe)2, pTsOH,
DMF 58%; b) 6 equiv MeLi, THF, 208C, 30 h, 43%; c) 1. Ac2O, Py,
DMAP, CH2Cl2; 2. TESOTf, lutidine; 3. DIBAH, CH2Cl2, ꢀ608C, 71%;
d) 1. tBu2Si(OTf)2, lutidine, DMF/CH2Cl2 1:1, ꢀ508C; 2. MeI, Ag2O,
87%; e) 1. TBAF, THF; 2. pTsCl, Py, CH2Cl2, 08C; 3. TBSOTf, lutidine,
CH2Cl2, 68%; f) 1. PhSCl, CH2Cl2; 2. Ag2CO3, CH3CN/H2O 9:1, 87%;
3. Cl3CCN, NaH, 91%; g) 11, TMSOTf, Et2O, ꢀ60!ꢀ408C, 1 h;
h) 1. NaI, DMF, 908C, 2 h; 2. Bu3SnH, AIBN, toluene, 1008C, 7 h, 70%
for three steps. AIBN=2,2’-azobis(isobutyronitrile), DIBAH=diisobu-
tylaluminum hydride, DMAP=4-dimethylaminopyridine, TES=triethyl-
silyl, TMS=trimethylsilyl, TBA=tetrabutylammonium.
gave via a cyclohexenone intermediate the glycal 10, which
was converted into the protected olivomycal building block
11. d-Glucal (12) was 4,6-silyl-protected and methylated at
O3 to give 13. Desilylation of the latter followed by tosylation
of the primary OH group and protection of the secondary
alcohol function with a tert-butyldimethylsilyl (TBS) group
gave 14. In preparation for a b-selective glycosylation, an
auxiliary SPh substituent was introduced at C2 of the d-
oleandrose building block.[11] To this end 14 was treated first
with PhSCl, then with Ag2CO3 in H2O/CH3CN, and the
Scheme 1. Retrosynthetic analysis of apoptolidin.
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resulting a-anomeric hemiacetal was converted into the
trichloroacetimidate 15. The TMSOTf-mediated glycosyla-
tion of 11 and 15 produced the disaccharide 16 with very high
b-selectivity (> 95:5). The conversion of the tosylate into an
iodide and the combined reductive removal of the iodo and
the thio function completed the preparation of the protected
2-deoxydisaccharide building block 17.
An inversion at C2 converts l-rhamnose into 6-deoxy-l-
glucose. Therefore, l-rhamnose is a suitable starting material
for the O9 sugar residue of apoptolidin (Scheme 3).[12] The
develop a new protecting group. Treatment of diol 21 with
1,4-dichloro-1,1,4,4-tetraphenyl-1,4-disilabutane (SIBACl2)[16]
gave the disilyl ether 22 in 92% yield. Oxidation of 22 with
mCPBA led to the desired SIBA-protected glycosyl sulfoxide
23.
The synthesis of the glycosylated northern half used the
unsaturated ester 26[9] from the aglycone synthesis as a
common building block (Scheme 4). Ester 26 was converted
into the glycosyl acceptor 27. Initial attempts to use the
Scheme 3. a) MeI, KOH, DMF, 08C, 99%; b) 1. pTsOH, MeOH, 89%;
2. TBSCl, imidazole, DMAP, CH2Cl2, 91%; 3. Dess–Martin periodinane,
80%; c) 1. NaBH4, MeOH, 08C, 15 min, 97%; 2. TBAF, THF, 94%;
d) Ph2Si(Cl)CH2CH2Si(Cl)Ph2, imidazole, DMF, 08C, 1 h, 92%;
e) mCPBA, CH2Cl2, ꢀ78!ꢀ208C, 2 h, 92%, 2:1 mixture of epimers.
mCPBA=meta-chloroperbenzoic acid.
Scheme 4. a) 1. DIBAH, toluene, ꢀ788C, 98%; 2. Ac2O, Et3N, DMAP,
CH2Cl2, 08C; 3. TBAF, THF, 0!208C, 92%; b) 1 equiv 23, 1.5 equiv
Tf2O, DTBMP, ꢀ808C, 10 min; addition of 27, ꢀ80!ꢀ358C, 2 h;
65%, a/b=85:15; c) 1. chromatographic separation of the anomers;
2. LiEt3BH, THF, ꢀ508C, 1 h, 75%; 3. MnO2, CH2Cl2;
acetonide-protected l-rhamnose thioglycoside 18[13] was O-
methylated to obtain 19. Acetonide cleavage, TBS protection
of O3, and subsequent oxidation of the remaining 2-OH
group provided the ketone 20. A highly stereoselective
reduction of 20 with NaBH4 gave the corresponding alcohol,
which was desilylated to provide the diol 21. The relative
configuration of 21 was confirmed by X-ray structure analysis
(Scheme 3).[14] The following choice of the right O2/O3 l-
glucose protecting groups was crucial for the successful
glycosylation of the northern half. An a-selective glycosyla-
tion required a passive O2 protecting group that can be
removed at the end without affecting the highly unsaturated
and acid-sensitive target molecule. Silyl ethers should be the
best choice. After several unsuccessful glycosylation attempts
(trichloroacetimidate, glycosyl fluoride, thioglycoside activa-
tion by PhSOTf) we focused on the Kahne glycosylation by
means of sulfoxide activation.[15] Initially we prepared and
examined the bis(TES)- and bis(TBS)-protected sulfoxides 24
and 25. However, these glycosyl donors gave unsatisfying
glycosylation results (vide infra), which prompted us to
=
4. Ph3P C(CH3)CO2Me, toluene, 908C, 44 h, 91%; d) 1. TBAF, THF,
208C, 99%; 2. TESCl, imidazole, 208C, 99%; 3. LiOH, THF/MeOH/
H2O 2:1:1, 08C, 1 h, 83%; 4. MnO2, CH2Cl2, chromatographic separa-
tion of the anomers, 78% a-anomer, 13% b-anomer; 5. (EtO)2-
P(O)CH(CH3)CO2H, NaH, THF, 128C, 14 h, 87%; 6. ClCH2CN, Et3N,
MeCN, 208C, 14 h, 92%. DTBMP=2,6-di-tert-butyl-4-methylpyridine.
bis(TES)-protected glycosylsulfoxide 24 as a glycosyl donor
failed due to loss of the TES groups under the reaction
conditions (Tf2O, DTBMP, ꢀ80!ꢀ358C). The more stable
bis(TBS)-protected glycosylsulfoxide 25 gave the desired
glycoside in 50% yield but with an unacceptably low stereo-
selectivity (a/b = 66:33). In contrast, the reaction of the
SIBA-protected glycosylsulfoxide 23 with 27 gave the
expected product 28 in 65% yield with an acceptable
stereoselectivity (a/b = 85:15). The final task for the comple-
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Scheme 5. a) TBAF, THF, 08C, 90%; b) 17, NIS, 4-Š molecular sieves, CH2Cl2, 0!208C, 72 h, 53%; anomeric ratio >95:5; c) Bu3SnH, AIBN, tolu-
ene, 1008C, 15 min, 96%; d) 1. H2, Pd/C, EtOH, 90%; 2. Dess–Martin periodinane, pyridine, CH2Cl2, 208C, 1 h, 84%; e) 1. Mg, BrCH2CH2Br, Et2O,
ꢀ788C, 74%, ds>95:5 ; 2. KCN, MeOH, 408C, 16 h, 86%. NIS=N-iodosuccinimide.
Scheme 6. a) 3 equiv CuI thiophenecarboxylate, NMP, 1 h, 08C, 89%; b) 3 equiv LiOH, THF/MeOH 3:1, 208C, 2 h, 88%; c) 20 equiv 2,4,6-tri-
chlorobenzoyl chloride, 40 equiv Et3N, THF, 6 h; toluene, 80 equiv DMAP, c=3”10ꢀ4 m, 75%; d) HF, pyridine, THF, 208C, 5 d, 50%; e) H2SiF6
(25% in H2O), CH3CN, ꢀ40!ꢀ208C, 2 d, ꢀ108C, 1 d, 71% 1 and 27% 41. NMP=N-methylpyrrolidone.
tion of the northern half was the introduction of the C1–C3
unsaturated ester fragment. This was accomplished using
standard Wittig and Horner—Wadsworth–Emmons proce-
dures to give the methyl ester 29 and the cyanomethyl ester
30, respectively.
The synthesis of the glycosylated southern half made use
of the bis(TES) ether 31 from the aglycone synthesis
(Scheme 5).[9] Selective deprotection of the 27-O-TES group
in 31 gave 32, which was allowed to react with the
disaccharide glycal 17 to produce the glycoconjugate 33
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with a high a-selectivity of > 95:5.[17] Reductive removal of
the auxiliary iodo function led to the benzyl ether 34. In order
to prevent side reactions with the subsequent hydrogenolysis,
it was necessary to remove all tin impurities from 34 by
treatment with fluoride at this stage. The corresponding
primary alcohol could be oxidized to the aldehyde 35. The
final step for the completion of the southern half was the
chelation-controlled addition of the Grignard reagent 36[9] to
35 to produce the corresponding alcohol with a diastereose-
lectivity of 96:4. Cleavage of the two acetates led to the triol
37.
Keywords: cross-coupling · glycosylation · macrolides ·
natural products · total synthesis
.
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The cross-coupling of either the northern half, 29 or 30,
with the southern half 37 was possible with CuI thiophene-2-
carboxylate in NMP (Scheme 6).[18] The methyl ester coupling
product caused trouble in the subsequent methyl ester
hydrolysis. We therefore focused on the cyanomethyl ester
coupling product 38. The cyanomethyl ester function in 38
could be hydrolyzed under very mild conditions (LiOH, 208C,
2 h) to the acid 39 without affecting the triene system or the
TES protecting groups. A remarkable ring-size-selective
macrolactonization of 39 produced the 20-membered lactone
40 in 75% yield. The final deprotection step required a
careful examination of reagents and optimization of reaction
conditions. The use of HF/pyridine in THF/pyridine at room
temperature for 5 d cleaved all silyl ethers but left the methyl
ketal intact and gave synthetic 21-o-methyl apoptolidin (2)
which proved to be identical with 2 derived from natural
sources ([a]22 = ꢀ76 (c = 0.55 in CHCl3),[5] [a]2D2 = ꢀ67 (c =
D
1.28 in CHCl3); for 1H and 13C NMR data see the Supporting
Information).
We then turned our attention to the complete deprotec-
tion of 40 leading to apoptolidin (1). Treatment with 25%
aqueous H2SiF6 in CH3CN at ꢀ40!ꢀ108C proved to be
effective for cleavage of all silyl ethers[19] and noteworthy the
methyl ketal. Apoptolidin (1) could be isolated in 71% yield
by chromatographic separation on deactivated silica gel[20]
with CH2Cl2/MeOH. In addition, the cleavage of the 27-O-
disaccharide was observed and compound 41 was isolated in
27% yield. The physical and spectroscopic data of the
synthetic apoptolidin (1) matched those published by Haya-
kawa et al. (m.p. 129–1318C (MeOH), ref. [2]: 128–1308C;
([a]2D2 = ꢀ4.4 (c = 0.70 in MeOH), ref. [2] [a]2D2 = ꢀ5.2 (c = 1.0
1
in MeOH); for H and 13C NMR data see the Supporting
Information).
In conclusion an efficient, convergent, and stereoselective
total synthesis of apoptolidin has been achieved. The distinct
features of this synthesis are the early introduction of the
sugar residues using a new sugar protecting group (SIBA), a
CuI-mediated cross-coupling followed by a ring-size-selective
macrolactonization, and mild deprotection conditions. This
strategy should be applicable to the synthesis of apoptolidin
derivatives with potential applications in apoptosis studies
and tumor therapy.
[20] Chromatotrex silicagel MB 100-40/75 from Fuji Silysia Chemical
LTD.
Received: March 31, 2004
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