Synthesis of 35-deoxy amphotericin B methyl ester: A strategy for molecular editing

Article abstract of DOI:10.1002/anie.200800589

(Chemical Presented) A modular strategy for the assembly of amphotericin B analogues with modifications in the macrolactone ring relies on the efficient gram-scale synthesis of all major and minor motifs of amphotericin B. Proof of concept has been achieved by the preparation of the 35-deoxy aglycone en route to the long-sought-after 35-deoxy analogue of amphotericin B.

Full text of DOI:10.1002/anie.200800589

Angewandte
Chemie
DOI: 10.1002/anie.200800589
Natural Products (1)
Synthesis of 35-Deoxy Amphotericin B Methyl Ester: A Strategy for
Molecular Editing**
Alex M. Szpilman, Damiano M. Cereghetti, Nicholas R. Wurtz, Jeffrey M. Manthorpe, and
Erick M. Carreira*
Amphotericin B (1) is the most prominent of the mycosamine
polyene macrolides and is of great importance in medicine.[1]
Various formulations of 1 have allowed its successful use as a
fungicide against aspergillosis, candidemia mucormycosis,
and multidrug resistant forms of leishmania.[2] Despite four
decades of research, there remain numerous questions con-
cerning its mechanism of action; the two most commonly
cited include its ability to induce oxidative damage and its
role in loss of electrochemical membrane potential.[3,4] The
latter is widely accepted and involves the formation of barrel-
stave ion channels that result in efflux of electrolytes through
the fungal cell membrane. Numerous experimental and
computational studies have led to various proposals for the
structure of these channels.[3,4] We have selected 35-deoxy
amphotericin B as a synthesis target because of the pivotal
role the hydroxy group at C35 has been suggested to play in
stabilizing channels. Herein we disclose the synthesis of 35-
deoxy amphotericin B aglycone, a necessary condition for the
subsequent biological studies described in the accompanying
paper.[5] More broadly, the approach we document should
enable access to designed analogues as powerful probes for
additional studies of the mechanism of action.
Modifications of the amphotericin B polyketide synthase
machinery through genetic engineering by Caffrey and co-
workers as well as semisynthetic alterations by other
researchers have provided access to derivatives that highlight
the importance of the polyene backbone, the carboxylic acid,
and the free amino group.[6–8] In an effort to access variants
not addressed by the reported approaches we chose to pursue
a modular strategy that would render the synthetic scheme
amenable to modification of various functional groups at
will.[9] It is important to note that numerous approaches to the
subunits of 1 have been developed,[4] in addition to the
synthesis of the related aglycones of candidin and rimoci-
din.[10,11] However, only one total synthesis of amphotericin B
(1) has been reported to date.[12]
The synthesis of the C1–C7 fragment 7 commenced with
dimethyl (S)-malate (2; Scheme 1).[13] Selective ester reduc-
tion[14] was followed by protection of the primary alcohol
Scheme 1. a) BH3·SMe2, cat. NaBH4, THF, 08C;b) TBSCl, imidazole,
CH2Cl2, 0–238C;c) LDA, tert-butyl acetate, THF, À78 to À108C;
54.5% over three steps;d) Bu 3B, NaBH4, MeOH, THF, À788C;
e) H2O2, water/THF;f) PPTS, 2-methoxypropene, À358C to RT;69%
over three steps, d.r.=15:1;g) LiAlH 4, THF, À108C;h) NaH, BnBr,
Bu4NI, THF/DMF (10:1);88% over two steps;i) HF/pyridine, THF,
08C;j) cat. TEMPO, cat. KBr, NaOCl, pH 8.6 buffer, CH Cl2, 0–78C;
k) EtO2CC(N2)P(O)(OEt)2, K2CO3, MeOH;51% over three steps. LDA
2
[*] Dr. A. M. Szpilman, Dr. D. M. Cereghetti, Dr. N. R. Wurtz,
Dr. J. M. Manthorpe, Prof. Dr. E. M. Carreira
ETH Zurich HCI H335
= lithium diisopropylamide;TBS = tert-butyldimethylsilyl;PPTS
pyridinium para-toluenesulfonate, Bn = benzyl.
=
Wolfgang-Pauli-Strasse 10, 8093 Zurich (Switzerland)
Fax: (+41)44-632-1328
E-mail: carreira@org.chem.ethz.ch
Homepage: http://www.carreira.ethz.ch
group. Chain elongation by Claisen condensation of 3 and
tert-butyl acetate gave b-ketoester 4 (54.5%, three steps).
Prasad reduction afforded the desired syn-3,5-diol with 15:1
selectivity.[15] Protection of the diol was effected by PPTS and
2-methoxypropene at low temperature (69%, three steps).
The ester 5 was converted into the benzyl ether 6. Subsequent
[**] This research was supported by ETH and agrant from the Swiss
National Science Foundation. Postdoctoral scholarships were
provided by the Carlsberg foundation (A.M.S.), Fonds NATEQ
(J.M.M.), and NSF (N.R.W.).
Supporting information for this article is available on the WWW
under http://www.angewandte.org or from the author.
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4335

Communications
removal of the TBS group gave the free alcohol which was
Studies in our laboratory had shown that coupling of
lithiated 7 to 8 would produce the (S)-C8 epimer of 9 as the
major and undesired product.[18] This step thus presented an
opportunity to examine the asymmetric zinc acetylide addi-
tion to aldehydes in a highly complex setting.[19] Accordingly,
alkyne 7 was treated with Zn(OTf)2, (À)-N-methyl-ephe-
drine, Et3N, and aldehyde 8 to afford propargylic alcohol 9 in
98% yield and d.r. = 16:1 (Scheme 2). Hydrogenation of the
alkyne was followed by protection of the secondary alcohol
function. Ester 10 was converted to oxime 11 in 84% overall
yield in three steps without purification of intermediates.
Alcohol 12 was accessed through an Evans aldol addition
reaction and converted to 13 following deprotection, oxida-
tion, and esterification.[20] Coupling of 11 and 13 was then
performed by a nitrile oxide cycloaddition (Scheme 3), in
analogy to that reported by McGarvey et al. for a different
homoallylic alcohol.[21] Under these conditions, the desired
isoxazoline 14 was formed as a single regioisomer in 95%
yield and d.r. = 88:12. Treatment of 14 with LiOOH afforded
the carboxylic acid, which was converted to bis-methyl ester
15. Reductive opening of the isoxazoline ring with [Mo(CO)6]
led to unmasking of a hydroxy ketone,[22] which underwent
cyclization during purification to hemiketal 16. Protection of
16 as the methyl ketal proved difficult because of low
reactivity combined with the acid sensitivity of the acetonides.
This problem was solved by employing 2-chloropyridinium
camphorsulfonate (pKa = 0.8) in a mixture of 2,2-dimethox-
ypropane and MeOH. Protection of the secondary alcohol
using TBSOTf afforded the fully protected C1–C19 polyol 17.
The treatment of 17 with 1.8 equiv of (MeO)2P(O)CH2Li at
08C gave the desired keto-phosphonate in 71% yield.
Removal of the benzyl group and two-step oxidation of the
resulting C1 alcohol afforded carboxylic acid 18 (99%), which
oxidized to the corresponding aldehyde. Finally, treatment
with Ohiraꢀs reagent[16] in the presence of potassium carbon-
ate afforded the desired alkyne 7 in 51% overall yield. Only
four chromatographic purification steps were necessary for
the entire sequence, a fact that expedited the synthesis of
more than one hundred grams of this material.
Aldehyde 8 (Scheme 2), corresponding to the C8–C13
part of amphotericin B, was synthesized in an analogous
manner in eight steps starting from readily available diethyl
(R)-malate.[17]
Scheme 2. a) 7, Zn(OTf)2, (À)-NME, Et3N, toluene, then 8;98%,
d.r.=16:1;b) H 2, cat. Pd/C, NaHCO3, MeOH;c) TBSCl, imidazole,
DMF, 408C;d) LiAlH 4, THF, 08C;e) cat. TEMPO, cat. KBr, NaOCl,
pH 8.6 buffer, CH2Cl2, 08C;f) HONH 2·HCl, pyridine;84% over five
steps. NME = N-methyl ephedrine;TEMPO = 2,2,6,6-tetramethyl-
piperidine-1-oxyl.
Scheme 3. a) HF/pyridine, THF, 08C;b) cat. TEMPO, cat. KBr, NaOCl, pH 8.6 buffer/CH 2Cl2, 08C: c) NaClO2, tBuOH/2-methyl-2-butene/2m
NaH2PO4, 08C;d) TMSCHN 2, MeOH/EtOAc;63% over three steps;e) (Bu 3Sn)2O, 11, then 13, tBuOCl, CH2Cl2, À35 to 238C;95%, d.r. =88:12;
f) LiO2H, water/dioxane;g) TMSCHN 2, MeOH/EtOAc, 67% over two steps;h) [Mo(CO) 6], MeCN/water, 808C, 86%;i) 2-chloropyridine·CSA,
(MeO)2CMe2, MeOH;j) TBSOTf, 2,6-lutidine, CH Cl2, 75% over two steps;k) (MeO) 2PCH2Li, THF, À35–08C;71% at 73% conversion;l) cat.
2
Pd(OH)2, H2, EtOAc;m) DMP, pyridine, CH 2Cl2;n) NaClO 2, tBuOH, 2-methyl-2-butene, 2m NaH2PO4;99% over three steps;o) 24, 2,4,6-
trichlorobenzoylchloride, pyridine, CH2Cl2, 08C;48%;p) K 2CO3, [18]crown-6, toluene, 608C;52%;q) NaBH 4, MeOH, 08C;73%. TMS
trimethylsilyl;CSA = camphor sulfonic acid;DMP = Dess–Martin periodonane;X N = Evans’ nor-ephedrine derived auxiliary.
=
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Chemie
intercepts the Nicolaou synthesis of Amphotericin B (1).[12]
Using this route, more than five grams of 18 were produced.
The work outlined in Scheme 1–3 constitutes the shortest
(27 steps from dimethyl malate) and most efficient (4.1%
yield) synthesis of 18 to date. Importantly, we had fulfilled the
first objective of our strategy, namely to have access to all
three major fragments of amphotericin B by efficient syn-
thetic routes on a gram-scale.[9]
zation afforded protected 19-keto 35-deoxy amphotericin B
aglycone.
In a final step, substrate-controlled reduction[10,12a] of the
resulting macrocyclic ketone afforded protected 35-deoxy
amphoteronolide as a single diastereomer.[12a]
In summary, we have reported a modular strategy that is
adaptable to the efficient assembly of amphotericin B ana-
logues bearing modifications in the macrolactone ring. This
strategy relies on the gram-scale efficient synthesis of all the
subunits.[9] For example, we have prepared more than
100 grams of each of the C1–7 and C8–C13 units and more
than five grams of the complex C1–C20 polyol 18. Addition-
ally, the reagent-controlled coupling of alkyne and aldehyde
provides full configurational control over the relevant frag-
ment assembly. The ready availability of these subunits should
expedite the preparation of a large variety of amphotericin B
analogues that may not be accessed by semi-synthesis or
genetic engineering.[6–8] The implementation of the approach
is showcased with the preparation of the aglycone en route to
the 35-deoxy analogue of amphotericin B.[25]
The assembly of the 35-deoxy analogue of the C21–C38
fragment 24 commenced with a Frµter–Seebach alkylation of
(S)-3-hydroxy-butyric acid ethyl ester (20, Scheme 4). After
Received: February 5, 2008
Revised: March 18, 2008
Published online: May 2, 2008
Keywords: amphotericin B · antifungal agents ·
.
asymmetric synthesis · natural products · total synthesis
Scheme 4. a) LDA, MeI, HMPA/THF (1:10), À788C;92%, d.r. =95:5;
b) TIPSOTf, 2,6-lutidine, CH2Cl2, 08C;c) DIBAL, Et O, À78 to 08C;
[1] T. H. Sternberg, E. T. Wright, M. Oura, Antibiot. Annu. 1955–
2
47% over two steps;d) Ph 3P, I2, THF;86%;e) 25, LDA, LiCl, THF,
1956, 566 – 573.
À78 to 08C;80%, d.r. =95:5;f) LDA, BH ·NH3, THF;72%;g) cat.
[2] a) H. A. Gallis, R. H. Drew, W. W. Pickard, Rev. Infect. Dis.
1990, 12, 308 – 329; b) I. M. Hann, H. G. Prentice, Int. J.
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Kiderlen, O. Kayser, Appl. Microbiol. Biotechnol. 2005, 68, 151 –
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[3] For a leading review, see: S. C. Hartsel, C. Hatch, W. Ayenew, J.
Liposome Res. 1993, 3, 377 – 408.
[4] For a recent review see: D. M. Cereghetti, E. M. Carreira,
Synthesis 2006, 914 – 942.
3
TEMPO, cat. KBr, NaOCl, pH 8.6 buffer/CH2Cl2, 08C;h) 26, LDA, THF,
À78 to 08C;94%;i) HF, aq. MeCN;67%;j) TESOTf, 2,6-lutidine,
CH2Cl2, 08C;81%;k) DIBAL, CH 2Cl2, À788C;l) MnO 2, CH2Cl2;m) 26,
LDA, THF, À78 to 08C;55% over three steps;n) HF/pyridine, THF,
08C;o) DIBAL, CH 2Cl2, À788C;p) MnO , CH2Cl2;62% over three
2
steps. TIPS = triisopropylsilyl;Tf = trifluoromethanesulfonyl;TES
triethylsilyl;DIBAL = diisobutylaluminum hydride;X a = Myers’
pseudo-ephedrine derived auxiliary.
=
[5] A. M. Szpilman, J. M. Manthorpe, E. M. Carreira, Angew. Chem.
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alcohol protection, ester reduction, and conversion of the
resulting primary alcohol to iodide 21, Myers diastereoselec-
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alcohol that was converted into hexaene-aldehyde 24.
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the 3,5-acetonide. In addition, the low solubility of 24 in any
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[9] a) An efficient synthesis of the C33–C38 fragment has already
been disclosed: J. Tholander, E. M. Carreira, Helv. Chim. Acta
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