Organic Letters
Letter
face of the cis-fused CD ring, giving rise to pyrone-attached
epoxide 6 as a single diastereomer. The X-ray crystallographic
analysis of 6 uncovered the entire stereostructure, showing the
U shape of the steroidal skeleton and the α-configured 2-
pyrone at the hindered C17 position.
TMS group of 19 that was introduced at the prior step
shielded the β-face, NaBH4 reduction occurred from an
opposite side, stereoselectively leading to C16β-alcohol 24.
The two silyl protecting groups of 24 were then removed using
HF·pyridine in pyridine and THF to provide bufogenin B (2).
Alternatively, the free C16-alcohol of 24 was acetylated before
the HF·pyridine-mediated deprotection, giving rise to bufotalin
(3).
Reductive removal of the C16-alcohol of 24 in turn
produced bufalin (1). Application of the typical Barton−
McCombie-type deoxygenation to 24 was unsuccessful,
presumably due to the unwanted participation of 2-pyrone in
the reaction. Accordingly, alcohol 24 was subjected to a neutral
halogenation/dehalogenation process. CBr4, PPh3, and imida-
zole26 transformed 24 to the desired bromide 26 and the
undesired olefin 27 via an SN2 reaction and an E2 elimination,
respectively. Compounds 26 and 27 were together subjected to
radical deoxygenation conditions using Et3B and n-Bu3SnH
under an O2 atmosphere. In one pot, aqueous HCl and MeOH
were added for deprotection to afford 1 along with the
byproduct 28.27
Finally, esterification of the C3-hydroxy group of bufotalin
(3) delivered vulgarobufotoxin (4) and 3-(N-succinyl
argininyl) bufotalin (5). Argininyl suberic acid 29a and
argininyl succinic acid 29b were prepared as the protected
forms: the carboxylic acid and the guanidine were capped with
the t-Bu group and the 2,2,4,6,7-pentamethyldihydrobenzofur-
an-5-sulfonyl (Pbf) group,28 respectively. C3-alcohol 3 was
conjugated with the carboxylic acid of 29a/29b using N,N-
dimethyl-4-aminopyridine (DMAP) and N,N′-diisopropylcar-
bodiimide (DIC) to form 30a/30b. Removal of the t-Bu and
Pbf groups was realized by applying a mixture of CF3CO2H,
PhOMe, PhSMe, and HSCH2CH2SH in CH2Cl2,29 leading to
4/5.30
The epoxide rearrangement from 6 to 19 was not easily
accomplished because of the unusual reactivity of 2-pyrone.
The conditions greatly affected the reaction outcome. For
example, InCl3 activated the epoxide of 6, but the desired
product 19 was not detected. The major compound was fused-
hexacycle 16, which was isolated in 60% yield. Compound 16
was derivatized into p-bromobenzoate 18, whose structure was
determined by X-ray crystallographic analysis. After the
reagents were screened, the combination of trimethylsilyl
trifluoromethanesulfonate (TMSOTf) and 2,6-lutidine25 was
found to induce the requisite rearrangement. TMSOTf (2.4
equiv) and 2,6-lutidine (5.1 equiv) in CH2Cl2 promoted the
C14O-TMS ether formation and the 1,2-hydride shift from 6,
leading to 19. The β-orientation of all the substituents at C13,
C14, and C17 of 19 was confirmed by nuclear Overhauser
effect (NOE) correlations. Consequently, the three-step
transformations under mild conditions constructed the densely
substituted D ring structure without affecting the acid-sensitive
C14-tertiary alcohol.
A plausible mechanism of the two distinct pathways is
depicted in Scheme 3. The InCl3-induced opening of the
Scheme 3. Plausible Mechanism of Lewis Acid-Mediated
Reactions of Epoxide 6
The total synthesis of bufadienolides 1−5 permitted us to
study their structure−activity relationship (SAR).31 The cell
growth inhibitory activities of 1−5 and the byproduct C17-
olefin 28 were assessed against MCF-7 human breast cancer
cells using the sulforhodamine B assay (Table 1).4c,32 Among
the six tested compounds, 1 with no C16-oxygen functional
group exhibited the highest activity (50% growth inhibitory
concentration (GI50) = 13.3 nM). C16-OH 2 and C16-OAc 3
were 4.1-fold and 2.1-fold weaker than 1 while they retained
two-digit nanomolar activity. The three data suggested that the
C16-hydroxy group had an unfavorable effect on the potency.
The GI50 values of C3O-acylated 4 and 5 were at least 10-fold
larger than that of C3O-nonacylated 3, corroborating that their
C3O-acyl chains negatively influenced the activity. The least
potent compound was 28, which is the C16,17-didehydro
analogue of the most potent compound 1. The 180-fold
weaker activity (2390 nM) of 28 in comparison with 1
confirmed the biological significance of the β-orientation of the
2-pyrone group.
In summary, we accomplished a unified total synthesis of the
5 bufadienolides 1−5 in 13−16 steps from the commercially
available steroid 9. The two sequences of transformations
played crucial roles in the syntheses. C5-hydrogenation, C3-
reduction, and C14-hydroxylation constructed the AB-cis/CD-
cis ring system of 8, while Stille coupling, stereoselective
epoxidation, and a stereospecific TMSOTf-mediated 1,2-
hydride shift installed the β-oriented 2-pyrone of 19. The
following stereoselective hydride addition produced 24, which
served as the common intermediate for the total synthesis of
oxirane ring is assisted by electron donation from the oxygen
lone pair of the 2-pyrone ring, producing cation 21. The
nucleophilic indium alkoxide in 21 attacks the electrophilic
oxocarbenium ion to afford the chemically unstable acetal 22.
Further activation of 22 with InCl3 cleaves the acetal, and the
ejected carboxylate of 23 adds to the C17 position from the
opposite face of the C13-methyl group, providing the rigid
hexacycle 16. On the other hand, TMSOTf and 2,6-lutidine
activate the oxirane ring and cap both C14- and C16-alcohols
with TMS groups. Introduction of the bulky and electron-
donating C16O-TMS group of 20 decelerates the nucleophilic
attack on the oxocarbenium ion and accelerates the 1,2-hydride
shift. Hence, the α-oriented C16-hydride stereospecifically
adds to the C17 position to invert the C17 stereoconfiguration,
thereby rendering the β-configured 2-pyrone of 19.
The common intermediate 24 was prepared for the total
synthesis of bufadienolides 1−5 (Scheme 4). Since the C14O-
C
Org. Lett. XXXX, XXX, XXX−XXX