18O assisted analysis of a γ,δ-epoxyketone cyclization: Synthesis of the C16-C28 fragment of ammocidin D

Article abstract of DOI:10.1021/ol103003f

The C16-C28 fragment common to the cytotoxic macrolide ammocidin D has been prepared by a stereospecific 5-exo closure of a γ,δ-epoxyketone followed by a rearrangement to a pyran acetal. The reaction pathway was traced by ¹⁸O labeling of the ke

Full text of DOI:10.1021/ol103003f

ORGANIC
LETTERS
18O Assisted Analysis of a
2011
Vol. 13, No. 4
756–759
γ,δ-Epoxyketone Cyclization: Synthesis
of the C16-C28 Fragment of Ammocidin D
Stephen T. Chau,† Yoichi Hayakawa,‡ and Gary A. Sulikowski*,†
Departments of Chemistry and Biochemistry, Vanderbilt University,
Vanderbilt Institute of Chemical Biology, Nashville, Tennessee 37235, United States,
and Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki,
Noda, Chiba 278-8510, Japan
gary.a.sulikowski@vanderbilt.edu
Received December 10, 2010
ABSTRACT
The C16-C28 fragment common to the cytotoxic macrolide ammocidin D has been prepared by a stereospecific 5-exo closure of a
γ,δ-epoxyketone followed by a rearrangement to a pyran acetal. The reaction pathway was traced by 18O labeling of the keto carbonyl and
observation of 18O induced 13C shifts in the pyran acetal product. NMR data of the synthetic C16-C28 fragment compared favorably to the natural
product providing support of the assigned stereochemistry.
During the course of screening extracts for apoptosis
inducers in Ras-dependent Ba/F3-V12 cells, Hayakawa
and co-workers identified the macrolide ammocidin A
(Figure 1) from the culture broth of Sacchaarothrix sp.
AJ9571.1 In 2009, the same research group reported on the
isolation and antiproliferative properties of ammocidins
B-D.2 Structurally, ammocidins A-D share a common
20-membered macrolactone and differ primarily in glyco-
sylation at C24. For example, ammocidin A incorporates a
β-D-olivomycose-β-D-digitoxose disaccharide at C24 while
ammocidin D isdevoidofsugarsatthisposition(Figure1).
Only the two-dimensional structure of the common agly-
cone (ammocidinone) was assigned by NMR analysis with
degradation by acidic methanolysis providing full assign-
ment of the deoxy sugars by isolation of the derived methyl
acetals. Hayakawa has noted that the structure of ammo-
cidin A bears a resemblance to apoptolidin A,3 a macrolide
of fully assigned stereochemistry.1b Among the common
structural features shared by these polyketides are a central
20-membered macrolactone, a 28-carbon seco-acid incor-
porating a C21-C25 cyclic acetal (Figure 1) and 6-deoxy
glucose at C9. Based on comparison to apoptolidin A and
analysis of NOE data, the stereochemistry of ammocidin
A/D was tentatively assigned as shown in Figure 1.4 Given
the significant antiproliferative properties of ammocidins
† Vanderbilt University.
‡ Tokyo University of Science.
(1) (a) Murakami, R.; Tomikawa, T.; Shin-Ya, K.; Shinozaki, J.;
Kajiura, T.; Kinoshita, T.; Miyajima, K.; Seto, H.; Hayakawa, Y. J.
Antibiot. 2001, 54, 710–713. (b) Murakami, R.; Tomikawa, T.; Shin-Ya,
K.; Shinozaki, J.; Kajiura, T.; Kinoshita, T.; Miyajima, A.; Seto, H.;
Hayakawa, Y. J. Antibiot. 2001, 54, 714–717.
(3) Hayakawa, Y.; Kim, J. W.; Adachi, H.; Shin-Ya, K.; Fujita, K.;
Seto, H. J. Am. Chem. Soc. 1998, 120, 3524–3525.
(2) Murakami, R.; Shinozaki, J.; Kajiura, T.; Kozone, I.; Takagi, M.;
Shin-Ya, K.; Seto, H.; Hayakawa, Y. J. Antibiot. 2009, 62, 123–127.
(4) The relative stereochemistry between C8-C9, C16-C20, and
C21-C25 remains ambiguous.
r
10.1021/ol103003f
Published on Web 01/19/2011
2011 American Chemical Society

was obtained by an Evans asymmetric glycolate aldol starting
from (E)-6-methoxy-2-methylhexenal, prepared in four steps
from 4-methoxy methyl butanoate.7 Conversion of 1 to a
Weinreb amide followed by ethylation and removal of
the TES protecting group provided allylic alcohol 2,
which was poised for a hydroxyl directed epoxidation.
Our synthetic plan as shown in Figure 2 required an anti-
2,3-epoxy alcohol; therefore we conducted a vanadium
catalyzed epoxidation of 2.8
Scheme 1. Unexpected Isolation of Furan Acetal 4
Figure 1. Tentative stereochemical assignment of ammocidin A/D.
A-D, we initiated a program directed toward the total
synthesis of ammocidin D in order to assign the full struc-
ture and subsequently advance biological studies. Herein
we describe synthetic and mechanistic studies leading to a
stereocontrolled assembly of the C16-C28 fragment of
ammocidin D.
When compared to the apoptolidins, a distinguishing
structural feature of ammocidins A-D is incorporation of
a tertiary alcohol at C24. The C24 hydroxyl group and
substitution pattern of the pyran-acetal spanning the
C21-C25 region suggested the synthetic strategy shown
in Figure 2 beginning with a 5-exo opening of the neighbor-
ing methyl substituted epoxide by the C21 keto group (I).5
When accompanied by hydration, the 5-exo cyclization of I
would afford furan acetal II that we anticipated would
isomerize to the ammocidin pyran-acetal III.6
Epoxidation of allylic alcohol 2 did not provide epoxide 3
but instead furan acetal 4 (Scheme 1). The structure and
stereochemistry of 4 was assigned based on the combined
analysis of 4 and the derived TES ether 5. The acetal carbon
(C21) of both 4 and 5 resonated at 112 ppm, approximately
12 ppm further downfield than expected for the ammocidin
pyran acetal (III, Figure 2).1b Second, the observed cou-
pling constant between H22 and H23 for both 4 and 5 was 2
Hz while the corresponding coupling constant for ammo-
cidin D was 9.9 Hz. Further evidence for furan acetal 4 was
provided by two-dimensional NMR data whereby an NOE
cross-peak between H22 and Me24 was observed in the
NOSY spectrum and an HMBC cross-peak between C21
and H25 was absent. Thus, acetal 4 not only possessed the
incorrect ring tautomer but also inverted stereochemistry at
C24 and C25 relative to that required for the ammocidins
(cf. II, Figure 2).
Two possible reaction pathways, outlined in Scheme 2,
could account for the unexpected production of furan
acetal 4 starting from vanadium catalyzed epoxidation of
allylic alcohol 2. In path A, epoxidation leads to a syn-2,3-
epoxy alcohol (syn-3) followed by a 5-exo closure of the
γ,δ-epoxyketone leading to furan acetal 4. Path B presents
a second scenario starting from the anti-2,3-epoxy alcohol
(anti-3) followed by a 6-endo closure to intermediate pyran
(5) (a) Morten, C. J.; Jamison, T. F. J. Am. Chem. Soc. 2009, 131,
6678–6679. (b) Morten, C. J.; Byers, J. A.; Van Dyke, A. R.; Vilotijevic,
I.; Jamison, T. F. Chem. Soc. Rev. 2009, 38, 3175–3192.
(6) (a) Holland, J. M.; Lewis, M.; Nelson, A. Angew. Chem., Int. Ed.
2001, 40, 4082–4084. (b) Holland, J. M.; Lewis, M.; Nelson, A. J. Org.
Chem. 2003, 68, 747–753.
Figure 2. Synthetic strategy for acetal pyran formation.
(7) Martin, S. F.; Dodge, J. A.; Burgess, L. E.; Limberakis, C.;
Hartmann, M. Tetrahedron 1996, 52, 3229–3246.
(8) (a) Adam, W.; Wirth, T. Acc. Chem. Res. 1999, 32, 703–710. (b)
Sharpless, K. B.; Verhoweven, T. R. Aldrichimica Acta 1979, 12, 63–74.
Our first evaluation of this proposal examined cyclization
of the epoxy alcohol derived from aldol adduct 2, the C20-
C28 fragment of ammocidin D (Scheme 1). Oxazolidinone 1
Org. Lett., Vol. 13, No. 4, 2011
757

Scheme 2. Possible Routes Leading to Furan Acetal 4
Scheme 3. 18O-Assisted Analysis of Epoxidation-Cyclization
Pathwaya
acetal 6 and isomerization to the more stable furan acetal
4.9 In support of path A (cyclization by way of syn-3),
epoxidation of allylic alcohol 2 with m-CPBA did lead to
an isolable epoxide that on treatment with borontrifluor-
ide etherate provided acetal furan 4. Based on literature
precedent, we assumed theepoxide produced in theperacid
epoxidation was syn-3.10 However, direct assignment of
the syn/anti relative stereochemistry of 2,3-epoxy alcohols
by NMR analysis is historically difficult.11 In order to
provide evidence for syn-3 we labeledthe C21 carbonyl of 2
with an 18O isotope. Epoxidation of 18O-2 followed by
cyclization of the intermediate γ,δ-epoxyketone to furan
18O-4 would allow path A (syn-3) and path B (anti-3) to be
distinguished by assignment of the 18O position in furan 18O-4
based on the expected 18O-induced shift of the adjacent carbon
signal in the 13C NMR spectrum (Scheme 2).12
a NMR analysis assisted by premixing of 16O- and 18O-labeled 4.
2 to afford γ,δ-ketoepoxide syn-3 followed by a rapid
cyclization starts with the preorganization of aldol adduct
2 by an intramolecular hydrogen bond (Figure 3).14 We
speculated incorporation of a second hydrogen bond by
way of the C19 hydroxyl group may alter stereoselectivity
to favor the anti epoxy alcohol or allow isolation of the syn-
2,3-epoxy alcohol by slowing the rate of keto-epoxide
cyclization (Scheme 4). In the latter case we would examine
a 6-endo closure of the syn epoxy alcohol promoted by
aqueous solvent to afford the ammocidin acetal-pyran
based on the work of Jamison.5
The 18O-labeled ketone (18O-2) was readily prepared by
stirring ketone 2 in THF using 18O-labeled water and trace
HCl. After 1 h, 18O-2 was isolated and determined to be
>99% 18O labeled.13 Exposure of 18O-2 to VO(acac)2/
TBHP led to the isolation of 18O-4. Examination of the 13
C
NMR spectrum of 18O-4 indicated resonances correspond-
ing to C21 and C24 carbons were accompanied by 18O
induced shifts (Scheme 3). The combination of assigned
stereochemistry and isotope position in 18O-4 (Scheme 3)
indicatessyn-2,3-epoxyalcohol3underwenta5-exoclosure
to 4 (path A, Scheme 2). In contrast, 18O incorporation was
not observed at the C25 carbon as would be expected for a
6-endo closure (path B, Scheme 2).
A tentative model used to rationalize the observed
proclivity for stereoselective epoxidation of aldol adduct
Figure 3. Assumed hydrogen bonding of C20-C28 and
C16-C28 fragments.
(9) Merck Molecular Force Field calculations (MMFF94) estimate
furan acetal 4 to be 1.3 kcal more stable then pyran acetal 6.
(10) (a) Loh, T. P.; Feng, L. C. Tetrahedron Lett. 2001, 42, 3223–
3226. (b) Smith, A. B., III; Adams, C. M.; Barbosa, S. A. L.; Degnan,
A. P. J. Am. Chem. Soc. 2003, 125, 350–351. (c) Fleming, K. N.; Taylor,
R. E. Angew. Chem., Int. Ed. 2004, 43, 1728–1730.
(11) (a) Mihelich, E. D. Tetrahedron Lett. 1979, 4729–4732. (b)
Rossiter, B. E.; Verhoeven, T. R.; Sharpless, K. B. Tetrahedron Lett.
1979, 4733–4736.
Preparation of the C16-C28 fragment of ammocidin
(Figure 3) started with a double diastereoselective Mu-
kaiyama aldol reaction between aldehyde 8 and silyl enol
ether 9. Aldehyde 8 was prepared in five steps from (-)-malic
(14) In the case of the vanadium-mediated epoxidation coordination
of either the C23 hydroxyl group and/or C21 carbonyl may be con-
sidered.
(15) (a) Saito, S.; Hasegawa, T.; Inaba, M.; Nishida, R.; Fujii, T.;
Nomizu, S.; Moriwake, T. Chem. Lett. 1984, 1389–1392. (b) Saito, S.;
Ishikawa, T.; Kuroda, A.; Koga, K.; Moriwake, T. Tetrahedron 1992,
48, 4067–4086.
(12) (a) Gree, D.; Gree, R.; Lowinger, T. B.; Martelli, J.; Negri, J. T.;
Paquette, L. A. J. Am. Chem. Soc. 1992, 114, 8841–8846. (b) Risley,
J. M.; Vanetten, R. L.; Uncuta, C.; Balaban, A. T. J. Am. Chem. Soc.
1984, 106, 7836–7840. (c) Wilgis, F. P.; Neumann, T. E.; Shiner, V. J.
J. Am. Chem. Soc. 1990, 112, 4435–4446.
(13) The extent of 18O incorporation was determined by LC-MS.
758
Org. Lett., Vol. 13, No. 4, 2011

Scheme 4. Preparation of Ketone 18O-11, Cyclization to Pyran
Acetal 12 and Analysis of 13C NMR Spectrum of 18O-13
Figure 4. Revised pathway leading to acetal pyran formation.
tentatively assigned to ammocidin D (Figure 2). Further-
more, the position of 18O labels in 18O-13 and the product
stereochemistry supported epoxidation of 18O-10 afford
anti-2,3-epoxy alcohol (11) that upon heating in water
yielded 12 via a 5-exo epoxide opening. However, upon
heating a solution (MeOH-CH2Cl2) of pyran acetal 13 in
the presence of catalytic PPTS complete isomerization to
the corresponding furan acetal (C21: 13C NMR 112 ppm)
was observed indicating the latter to be thermodynamically
favored.17 This finding does not support the pathway
proposed in Figure 2 but that shown in Figure 4. In this
case, intermediate oxonium ion IV is intercepted by the C25
hydroxyl group leading to intermediate V and upon hydra-
tion provides pyran acetal III. A closely related cyclization of
a γ,δ-epoxyketone has been proposed to account for the
conversion of myriaporone 1 to myriaporone 2.18
In conclusion we have developed a synthetic route to
access the C16-C28 fragment of ammocidins A-D. Com-
parison of the spectral data of ammocidin A-D and pyran
acid.15 The aldol reaction between 8 and 9 proceeded with
>20:1 stereoselectivity to afford syn aldol 10 following
removal of the C23 TES ether.16 Incorporation of 18O at
theketo carbonyl proceededwith50% incorporationusing
conditions described earlier to provide 18O-10.13 Epoxida-
tion of 18O-10 with VO(acac)2-TBHP afforded a single 2,3-
epoxy alcohol (18O-11) isolable by flash chromatography,
implying hydrogen bonding of the C19 hydroxyl group
retarded the rate of cyclization as anticipated (Figure 3).
To our delight, heating a solution of 18O-11 in 18O-water at
60 °C for 3 days afforded pyran acetal 18O-12. Following
acetylation of 18O-12, analysis of the 13C NMR spectrum
of diacetate 18O-13 indicated 18O-incorporation at C21
and C24. Upon examination of the HMBC and NOESY
spectrum of the 18O-13, the chemical shift of the C21
carbon (99 ppm) and H22-H 23 coupling constant (10 Hz)
indicated pyran acetal 13 possessed the stereochemistry
1
acetal 13 (NOESY and H-1H coupling constants) sup-
port the stereochemical assignment of the C16-C28 frag-
ment shown in Figure 1.
Acknowledgment. We thank the National Institutes of
Health (CA059515). We also acknowledge Professor Ri-
chard E. Taylor (University of Notre Dame) for helpful
discussions and Donald F. Stec(Vanderbilt University) for
assistance with NMR data collection and analysis.
Supporting Information Available. Experimental proce-
dures and full spectroscopic data for all new compounds.
Comparison of NMR data for pyran acetal 14 to ammo-
dicidin D. This material is available free of charge via the
Internet at http://pubs.acs.org.
(17) In contrast heating
a solution of furan acetal 4 in
MeOH-CH2Cl2 and PPTS resulted in no change.
(18) Cheng, J.-F.; Lee, J.-S.; Sakai, R.; Jares-Erijman, E. A.; Silva,
M. V.; Rinehart, K. L. J. Nat. Prod. 2007, 70, 332–336.
(16) (a) Evans, D. A.; Yang, M. G.; Dart, M. J.; Duffy, J. L.; Kim,
A. S. J. Am. Chem. Soc. 1995, 117, 9598–9599. (b) Evans, D. A.; Dart,
M. J.; Duffy, J. L.; Yang, M. G. J. Am. Chem. Soc. 1996, 118, 4322–4343.
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