Total synthesis of bryostatin 16 using a Pd-catalyzed diyne coupling as macrocyclization method and synthesis of C20-epi-bryostatin 7 as a potent anticancer agent

Article abstract of DOI:10.1021/ja105129p

Asymmetric total synthesis of bryostatin 16 was achieved in 26 steps in the longest linear sequence and in 39 total steps from aldehyde 10. A Pd-catalyzed alkyne-alkyne coupling was employed for the first time as a macrocyclization method in a natural pro

Full text of DOI:10.1021/ja105129p

Published on Web 11/02/2010
Total Synthesis of Bryostatin 16 Using a Pd-Catalyzed Diyne
Coupling as Macrocyclization Method and Synthesis of
C20-epi-Bryostatin 7 as a Potent Anticancer Agent
Barry M. Trost* and Guangbin Dong
Department of Chemistry, Stanford UniVersity, Stanford, California 94305-5080, United States
Received June 23, 2010; E-mail: bmtrost@stanford.edu
Abstract: Asymmetric total synthesis of bryostatin 16 was achieved in 26 steps in the longest linear
sequence and in 39 total steps from aldehyde 10. A Pd-catalyzed alkyne-alkyne coupling was employed
for the first time as a macrocyclization method in a natural product synthesis. A route to convert bryostatin
16 to a new family of bryostatin analogues was developed. Toward this end, 20-epi-bryostatin 7 was
synthesized from a bryostatin 16-like intermediate; the key step involves a Re-catalyzed epoxidation/ring-
opening reaction. Preliminary biological studies indicated that this new analogue exhibits nanomolar anti-
cancer activity against several cancer cell lines.
Introduction
containing functionalities and stereogenic centers. Previously,
only three of the 20 bryostatins have been accessed by total
Bryostatins 1-20 were first isolated in 1968 by Pettit and
co-workers from the marine bryozoan Bugula neritina (Figure
1).1,2 These structurally complex macrolides exhibit a remark-
able range of biological activities, including antineoplastic
activity,3 synergistic chemotheoreputic activity,4 cognition and
memory enhancement,5 recovery of brain damage,6 etc.
Stimulated by these appealing biological activities, total
syntheses of bryostatins and their analogues have been an
attractive goal. The structures of the bryostatins constitute
significant synthetic challenges, which include a 26-membered
lactone fused by three heavily substituted polyhydropyran (PHP)
rings, two acid/base-sensitive exo-cyclic unsaturated esters, one
congested C16-C17 trans-olefin, as well as numerous oxygen-
synthesis. In 1990, Masamune and co-workers accomplished
the synthesis of bryostatin 7.7 Their strategy involved use of a
highly chemoselective macrolactonization to construct the
macrocycle and use of Julia olefination8 to couple the northern
fragment and southern fragment. In 1998, another family
member, bryostatin 2, was synthesized by Evans et al.9 A key
feature of Evans’ synthesis is that they segregated the target
into three polyhydropyran-containing subunits with similar
complexity, and then assembled them via Julia olefination,
sulfone alkylation, and macrolactonization. Notably, Evans’
synthesis also constitutes a formal synthesis of bryostatin 1.10
In 2000, bryostatin 3, a structurally unique family member, was
synthesized by Ohmori, Nishiyama, Yamamura, et al.11 Julia
olefination was also employed to form the C16-C17 alkene,
and a high-yielding Yamaguchi esterification was used to furnish
the 26-membered lactone. Recently, Hale and co-workers
reported a concise synthesis of Masamune’s southern fragment,
which was recognized as a formal total synthesis of bryostatin
7.12 These elegant syntheses have illustrated the power of
organic synthesis for the creation of molecules of extreme
complexity; however, their lengths (>40 steps in the longest
linear sequence and >70 total steps) have so far restrained them
from serving as a practical supply source for this natural product.
(1) Pettit, G. R.; Day, J. F.; Hartwell, J. L.; Wood, H. B. Nature 1970,
227, 962.
(2) Pettit, G. R.; Herald, C. L.; Doubek, D. L.; Herald, D. L.; Arnold, E.;
Clardy, J. J. Am. Chem. Soc. 1982, 104, 6846.
(3) For some recent reviews, see: (a) Hale, K. J.; Hummersone, M. G.;
Manaviazar, S.; Frigerio, M. Nat. Prod. Rep. 2002, 19, 413. (b)
Wender, P. A.; Baryza, J. L.; Hilinski, M. K.; Horan, J. C.; Kan, C.;
Verma, V. A. Beyond Natural Products: Synthetic Analogues of
Bryostatin 1. In Drug DiscoVery Research: New Frontiers in the Post-
Genomic Era; Huang, Z., Ed.; Wiley-VCH: Hoboken, NJ, 2007; pp
127-162.
(4) For a review, see: (a) Newman, D. J.; Cragg, G. M. J. Nat. Prod.
2004, 67, 1216. (b) For ongoing studies, see http://clinicaltrials.gov/.
(c) Dowlati, A.; Lazarus, H. M.; Hartman, P.; Jacobberger, J. W.;
Whitacre, C.; Gerson, S. L.; Ksenich, P.; Cooper, B. W.; Frisa, P. S.;
Gottlieb, M.; Murgo, A. J.; Remick, S. C. Clin. Cancer Res. 2003, 9,
5929.
(7) Kageyama, M.; Tamura, T.; Nantz, M. H.; Roberts, J. C.; Somfai, P.;
Whritenour, D. C.; Masamune, S. J. Am. Chem. Soc. 1990, 112, 7407.
(8) (a) Julia, M.; Paris, J. M. Tetrahedron Lett. 1973, 4833. (b) Kocienski,
P. J.; Lythgoe, B.; Ruston, S. J. Chem. Soc., Perkin Trans. 1 1978,
829. Review: (c) Kocienski, P. Phosphorus Sulfur 1985, 24, 97.
(9) (a) Evans, D. A.; Carter, P. H.; Carreira, E. M.; Prunet, J. A.; Charette,
A. B.; Lautens, M. Angew. Chem., Int. Ed. 1998, 37, 2354. (b) Evans,
D. A.; Carter, P. H.; Carreira, E. M.; Charrette, A. B.; Prunet, J. A.;
Lautens, M. J. Am. Chem. Soc. 1999, 121, 7540.
(5) (a) Etcheberrigaray, R.; Tan, M.; Dewachter, I.; Kuiperi, C.; Van der
Auwera, I.; Wera, S.; Qiao, L.; Bank, B.; Nelson, T. J.; Kozikowski,
A. P.; Van Leuven, F.; Alkon, D. L. Proc. Natl. Acad. Sci. U.S.A.
2004, 101, 11141. (b) Alkon, D. L.; Sun, M.-K.; Nelson, T. J. Trends
Pharmacol. Sci. 2007, 28, 51. (c) Alkon, D. L.; Epstein, H.; Kuzirian,
A.; Bennett, M. C.; Nelson, T. J. Proc. Natl. Acad. Sci. U.S.A. 2005,
102, 16432. (d) Sun, M.-K.; Alkon, D. L. Eur. J. Pharmacol. 2005,
512, 43. (e) Hongpaisan, J.; Alkon, D. L. Proc. Natl. Acad. Sci. U.S.A.
2007, 104, 19571.
(10) Pettit, G. R.; Sengupta, D.; Herald, C. L.; Sharkey, N. A.; Blumberg,
P. M. Can. J. Chem. 1991, 69, 856.
(11) Ohmori, K.; Ogawa, Y.; Obitsu, T.; Ishikawa, Y.; Nishiyama, S.;
Yamamura, S. Angew. Chem., Int. Ed. 2000, 39, 2290.
(6) Sun, M.-K.; Hongpaisan, J.; Nelson, T. J.; Alkon, D. L. Proc. Natl.
Acad. Sci. U.S.A. 2008, 105, 13620.
(12) Manaviazar, S.; Frigerio, M.; Bhatia, G. S.; Hummersone, M. G.; Aliev,
A. E.; Hale, K. J. Org. Lett. 2006, 8, 4477.
9
10.1021/ja105129p  2010 American Chemical Society
J. AM. CHEM. SOC. 2010, 132, 16403–16416 16403

A R T I C L E S
Trost and Dong
Figure 1. Bryostatins.
An attractive goal in synthesis is to access multiple targets with
a single route and multiple analogues via a common intermediate.
This concept particularly bodes well for the bryostatin synthesis
as this natural product contains 20 congeners and syntheses of their
analogues are equally valuable.13 Among all the 20 bryostatins,
we noticed that the structures of bryostatins 16 (1) and 1714 possess
a unique feature (Figure 1): their C-ring contains a relatively
reactive dihydropyran (DHP) moiety. By elaboration of this
electron-rich and relatively reactive C19-C20 olefin, we envisioned
that bryostatin 16 could act as a pivotal parent structure to allow
access to almost all the other naturally occurring bryostatins (except
bryostatin 3, 19 and 20).15 For example, oxidation of C19-C20
olefin could lead to bryostatins 1, 2, 4-9, 12, 14, and 15; a formal
hydration of this C-C double bond should provide potential access
to the C20-deoxy bryostatins (10, 11, 13, 18). Furthermore,
bryostatin 16 also offers an ideal forum for us to employ a Pd-
catalyzed tandem alkyne-alkyne coupling followed by 6-endo-
dig cyclization methodology to access the C-ring DHP motif in
an efficient and rapid fashion.16 In addition, simply by variations
in this natural product’s synthesis, we should be able to obtain
new analogues that might not be easily available from other
syntheses. In this full article, we provide a detailed account of our
completion of the synthesis of bryostatin 16 and demonstrate a
proof of principle in a concise synthesis of 20-epi-bryostatin 7 as
a potent anticancer agent from a bryostatin 16-like intermediate.17
accessed via either macrolactonization of a seco-acid or in-
tramolecular transesterification of a seco-methyl ester such as
2. The C-ring of bryostatin would be synthesized via the Pd-
catalyzed diyne coupling between donor alkyne 3 and acceptor
alkyne 4 followed by endo-cyclization of the secondary alcohol
to form the DHP entity. The A-ring could be formed through
acid-catalyzed tandem transesterification followed by methyl
ketal formation from lactone 5. The 4-methylene-2,6-cis-
tetrahydropyran (THP) moiety in 5 provides a perfect op-
portunity to examine another tandem transformation, the Ru-
catalyzed alkene-alkyne coupling/Michael addition methodology
(Scheme 1)19 between two rather complex fragments (6 and
7). Ideally, all of the three PHP rings of bryostatin 16 could be
accessible via three tandem, catalytic, and atom-economical
transformations.
Alkene 6, one of the key coupling partners, has been
previously synthesized in 16 overall steps from commercially
available (R)-pantolactone (8) (Scheme 2).13f Although this
synthesis is practical, to improve the efficiency our initial goal
was to shorten the synthesis of fragment 6.
Starting from aldehyde 10, the same aldehyde used in the
synthesis of fragment 7,20 aldehyde 9 was quickly afforded by
asymmetric Brown allylation with (-)-ꢀ-allyldiisopinocam-
pheylborane [Ipc2B(allyl)],21 followed by PMB protection with
PMBBr/NaH, and oxidative cleavage of the terminal olefin
(Scheme 3).22 As aldehyde 9 is a common intermediate with
our previous route, with this modification alkene 6 is now
available in 11 steps from aldehyde 10 (eq 1) which, in turn, is
commercially available or derived in two steps from 2,2-
dimethyl-1,3-propanediol.19
Results and Discussion
First-Generation Strategy toward the Synthesis of
Bryostatin 16. Given the difficulties of late-stage installation
of the C16-C17 olefin,18 we conceived of a strategy to
introduce this sterically hindered alkene at an earlier stage. From
a retrosynthetic viewpoint, the 26-membered lactone could be
(13) For recent examples of bryostatin analogues, see: (a) Keck, G. E.; Li,
W.; Kraft, M. B.; Kedei, N.; Lewin, E.; Blumberg, P. M. Org. Lett.
2009, 11, 2277. (b) Keck, G. E.; Poudel, Y. B.; Welch, D. S.; Kraft,
M. B.; Truong, A. P.; Stephens, J. C.; Kedei, N.; Lewin, N. E.;
Blumberg, P. M. Org. Lett. 2009, 11, 593. (c) Wender, P. A.; Horan,
J. C.; Verma, V. A. Org. Lett. 2008, 10, 3331. (d) Wender, P. A.;
DeChristopher, B. A.; Schrier, A. J. J. Am. Chem. Soc. 2008, 130,
6658. (e) Keck, G. E.; Kraft, M. B.; Truong, A. P.; Li, W.; Sanchez,
C. C.; Kedei, N.; Lewin, N. E.; Blumberg, P. M. J. Am. Chem. Soc.
2008, 130, 6660. (f) Trost, B. M.; Yang, H.; Thiel, O. R.; Frontier,
A. J.; Brindle, C. S. J. Am. Chem. Soc. 2007, 129, 2206.
With both alkene 6 and alkyne 7 in hand, the Ru-catalyzed
tandem alkene-alkyne coupling/Michael addition proceeded to
generate cis-tetrahydropyran 5 (eq 2). The chemoselectivity was
further demonstrated by the high compatibility of a ꢀ,γ-
unsaturated ketone, a six-membered lactone, an unprotected
allylic alcohol, a PMB ether, and two different silyl ethers in
(14) Pettit, G. R.; Gao, F.; Blumberg, P. M.; Herald, C. L.; Coll, J. C.;
Kamano, Y.; Lewin, N. E.; Shmidt, J. M.; Chapuis, J.-C. J. Nat. Prod.
1996, 59, 286.
(18) For an unsuccessful macrocyclic RCM or a RRCM strategy, see: (a)
Ball, M.; Bradshaw, B. J.; Dumeunier, R.; Gregson, T. J.; MacCormick,
S.; Omori, H.; Thomas, E. J. Tetrahedron. Lett. 2006, 47, 2223. (b)
Refs 7, 9, 11 and 13f.
(15) For biosynthetic approaches, see: (a) Sudek, S.; Lopanik, N. B.;
Waggoner, L. E.; Hildebrand, M.; Anderson, C.; Liu, H.; Patel, A.;
Sherman, D. H.; Haygood, M. G. J. Nat. Prod. 2007, 70, 67. (b) Kerr,
R. G.; Lawry, J.; Gush, K. A. Tetrahedron Lett. 1996, 37, 8305.
(16) Trost, B. M.; Frontier, A. J. J. Am. Chem. Soc. 2000, 122, 11727.
(17) For a preliminary report of a portion of this work, see: (a) Trost, B. M.;
Dong, G. Nature 2008, 456, 485.
(19) Trost, B. M.; Yang, H.; Wuitschik, G. Org. Lett. 2005, 7, 4761.
(20) For a synthesis of compound 7, see ref 17.
(21) Brown, H. C.; Jadhav, P. K. J. Am. Chem. Soc. 1983, 105, 2092.
(22) Yu, W.; Mei, Y.; Kang, Y.; Hua, Z.; Jin, Z. Org. Lett. 2004, 6, 3217.
9
16404 J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010

Syntheses of Bryostatin 16 and C20-epi-Bryostatin 7
A R T I C L E S
Scheme 1. First-Generation Strategy
Scheme 2. Previous Synthesis of Alkene 6
Scheme 3. Improved Synthesis of Alkene 6
this reaction. DCM was found to be the optimal solvent, while
acetone or a DCM-DMF mixed solvent gave either lower
conversion or more decomposition. Notably, only 1.2 equiv of
alkene 6 was required in this coupling reaction. Though the
yield is moderate, the yield based upon recovered starting
material is high (80%, both of the starting materials can be
recycled), presumably due to the fact that additional olefin
functionality in the alkyne fragment could limit turnover of the
Ru catalyst. This result has proved highly reproducible, and the
reaction is scalable to several grams.
MeOH, reflux), previously reported for a similar transesterifi-
cation/ketalization,13f gave a messy mixture. We envisaged that
the vinyl bromide functionality would serve as a convenient
handle in the future for the syntheses of bryostatin analogues
via the use of metal-catalyzed coupling reactions. As the natural
product contains a conjugated methyl ester at the C30 position,
a carbonylation reaction of 13 was next examined. On a smaller
scale (less than 100 mg), Pd(PPh3)4 acted as a good carbony-
lation catalyst, giving up to 78% yield (94% yield brsm);
however, on a larger scale, these conditions suffered from poor
conversion (less than 50% conversion). After surveying a
number of Pd catalysts, this problem was eventually solved by
using PdCl2(CH3CN)2-dppf as the catalyst, in which an 83%
yield (90% yield brsm) of ester 14 was obtained on a 0.76 g
scale.
The conditions for oxidizing primary alcohol 14 to
aldehyde 15 were next optimized (Table 1). It is known that
the methyl ketal of substrates like 14 is very sensitive to
acidic conditions.23,24 Thus, Ley oxidation (TPAP-NMO),25
known as a nonacidic oxidation method, was initially
employed. Using 0.1 equiv TPAP and 3 equiv NMO in
CH3CN, the reaction gave full conversion but only 65% yield
Advancement of lactone 5 is depicted in Scheme 4. Bromi-
nation of the exo-cyclic vinyl silane with NBS provided vinyl
bromide 12 with retention of the olefin geometry in 98% yield.
Subsequently, we attempted a one-pot acid-catalyzed transes-
terification-methyl ketalization-desilylation transformation. To
our delight, treatment of lactone 12 with a catalytic amount of
CSA in MeOH cleanly afforded the desired alcohol 13 contain-
ing both the A-ring and B-ring substructures in 93-96% yield
(on a gram scale). Use of the conditions (PPTS, HC(OMe)3,
(23) Yang, H. Ph.D. thesis, Stanford University, 2006.
(24) Similar observation has also been reported by Evans, see ref 9b.
(25) Ley, S. V.; Norman, J.; Griffith, W. P.; Marsden, S. P. Synthesis 1994,
639.
9
J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010 16405

A R T I C L E S
Trost and Dong
Scheme 4. Synthesis of Aldehyde 15
Table 1. Oxidation of Alcohol 14
cleavage of the TBDPS group and provided alcohol 3 in 90%
entry
conditions
yield, %b
yield (96% brsm).
1
TPAP(0.1 equiv), NMO (3 equiv),
4A M.S., CH3CN
65
2
2
3
4
5
6
TPAP(0.1 equiv), NMO (2 equiv),
4A M.S., CH3CN
TPAP(0.1 equiv), NMO (2 equiv),
4A M.S., DCM
TPAP(0.08 equiv), NMO (2 equiv),
4A M.S., DCM
TPAP(0.1 equiv), NMO (1.5 equiv),
4A M.S., DCM
TPAP(0.05 equiv), NMO (1.5 equiv),
4A M.S., DCM
DMP(3 equiv), NaHCO3
(20 equiv), DCM
67
56-70
74a
77a
With alkyne 3 in hand, we continued to examine the plan of
uniting the two fragments together via diyne coupling and then
closing the macrocycle via lactonization. Pd-catalyzed alkyne-
alkyne coupling between fragments 3 and 4 proceeded very well,
giving enyne 16 in 80% yield (Scheme 5). A gold-catalyzed
6-endo-dig cyclization was subsequently employed to build the
C-ring subunit.26 A 4:1 mixture of DCM-CH3CN was found
to be the optimal choice of solvent, and dihydropyran 17 was
isolated as the major product in 65% yield.
77a
88 (0.83 g scale)
a The reaction did not give full conversion. b Isolated yield.
of aldehyde 15 (entry 1). By carefully tuning the reaction
conditions, we found that lowering the catalyst loading (entry
3) or the oxidant loading (entry 4) or both (entry 5), gave
cleaner reactions and higher yield (up to 77% yield). DCM
was also discovered to be a more suitable solvent than
CH3CN. Subsequently, a more effective oxidation was found;
treatment of alcohol 14 with Dess-Martin periodinane
(DMP) and excess NaHCO3, efficiently afforded aldehyde
15 in 88% yield on a 0.83 g scale.
The acetonide protecting group was efficiently removed with
a catalytic amount of CSA in MeOH. However, under those
conditions, the external C21-C34 olefin was isomerized,
yielding a ∼1:1 mixture of olefin geometric isomers (18).
Transesterification of seco-ester 18 to form the macrolactone
19 was briefly explored. Treatment of 18 with 10 mol % Otera’s
catalyst27 in refluxing toluene or hexanes, however, did not
provide any macrocycle, even when using 5 Å molecular sieves
to remove MeOH. Attempts to hydrolyze the C1 methyl ester
of 18 followed by macrolactonization was not fruitful either.
The major difficulties were due to the fragile nature of the
external C21-C34 olefin leading to the existence of both the
starting materials and the potential products as isomeric mixtures
which complicated product identification and NMR spectral
interpretation. Although this first-generation plan still represents
a valid route, given those late-stage difficulties, we resorted to
a new strategy.
Second-Generation Strategy toward the Synthesis of
Bryostatin 16. Given the severe acid sensitivity of the C-ring,
instead of carrying it through, we conceived of a strategy for
constructing the C-ring of bryostatin 16 at the very end of the
synthesis. The benefits of this strategy also include flexible late-
stage variations for access to other bryostatins or analogues, as
well as minimization of functional group transformations and
Advancement of aldehyde 15 to the key coupling partner 3
was achieved in two steps (eq 3). Treatment of 15 with the
Ohira-Bestmann reagent nicely provided the corresponding
terminal alkyne in quantitative yield. As removal of the TBDPS
protecting group at a late stage proved to be problematic,23 it
was removed before the subsequent coupling with the other
fragment. Due to its robust nature, the TBDPS deprotection
required some experimentation. When 2.5 equiv TBAF (1.0 M
in THF) was used, alcohol 3 was isolated in only 60% yield
along with a significant amount of retro-aldol, ꢀ-hydroxyl
elimination, and ester hydrolysis byproducts. Formation of those
byproducts was likely caused by the strong basic nature of
TBAF as well as the hydroxide present in commercial TBAF.
To minimize those undesired reaction pathways, we envisioned
that it would be helpful to add a buffer and to control the
reaction pH and concentration. Indeed, treatment of the TBDPS
ether with less TBAF (1.1 equiv) and buffered with 20 mol %
HOAc at a lower concentration, led to the chemoselective
(26) For a related model study, see: (a) Dong, G. Ph.D. thesis, Stanford
University, 2009.
(27) Otera, J.; Danoh, N.; Nozaki, H. J. Org. Chem. 1991, 56, 5307.
9
16406 J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010

Syntheses of Bryostatin 16 and C20-epi-Bryostatin 7
A R T I C L E S
Scheme 5. End Game of the First-Generation Strategy
Scheme 6. Synthetic Plan of the Second-Generation Strategy
protecting group usage. As early as in 1989, we have demon-
strated the principle of using Pd-catalyzed R,ω-diyne cycloi-
somerization to form macrocycles.28 Use of this method in
natural product synthesis, however, has not been previously
established. While all the previous bryostatin total syntheses
have relied on assembling the macrocycle by a demanding Julia
olefination followed by a lactonization, we foresaw that we could
employ the Pd-catalyzed alkyne-alkyne coupling as a novel
macrocyclization method followed by a metal-catalyzed 6-endo-
dig cyclization to construct both the macrolactone and the C-ring
of byostatin 16 simultaneously (Scheme 6). Esterification
between fragments 22 and 23 would provide the requisite diyne
precursor. Acid 22 and alcohol 23 would be synthesized
respectively from intermediates 3 and 4 from our first route.
Starting from the ꢀ-hydroxy methyl ester 3, the challenge
was to hydrolyze the C1 methyl ester chemoselectively in the
presence of the C31 methyl ester. We found that heating 3 with
trimethyltin hydroxide in DCE29 provided the desired C1 acid
(24) in 84% yield (Scheme 7). The desired chemoselectivity
was attributed to two factors: first, the conjugated esters are
generally less reactive toward hydrolysis than the nonconjugated
ones due to the more delocalized π-systems; second, the Lewis
acidity of trimethyltin hydroxide allows the adjacent ꢀ-hydroxy
group to act as a directing group in this saponification reaction.
Due to the acid-sensitivity of the methyl ketal functionality, acid
24 had to be handled with sufficient care.30 Subsequent TES
protection of the secondary alcohol completed the synthesis of
acid fragment 22.
(28) Trost, B. M.; Matsubara, S.; Caringi, J. J. J. Am. Chem. Soc. 1989,
111, 8745.
(30) For example, either extended exposure of 24 to silica gel or storage
in neat form led to the loss of the methyl ketal. It was also found that
the most desirable way to store acid 24 was to freeze it as a benzene
solution.
(29) Nicolaou, K. C.; Estrada, A. A.; Zak, M.; Lee, S. H.; Safina, B. S.
Angew. Chem., Int. Ed. 2005, 44, 1378.
9
J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010 16407

A R T I C L E S
Trost and Dong
Scheme 7. Synthesis of Acid Fragment 22
Scheme 8. Esterification between Acid 22 and Alcohol 23
Alcohol fragment 23 was synthesized in three straightforward
steps from acetonide intermediate 4.31 Cu(OTf)2-catalyzed PMB
protection of the secondary alcohol,32 acid-mediated hydrolysis
of the acetonide moiety to provide the vicinal diol, and selective
TBS-ether protection of the less hindered alcohol gave the
alcohol fragment 23 (eq 4). A somewhat diminished yield was
observed on a 0.4 g scale in contrast to a 66% yield oVer three
steps on a 3.7 mg scale, which is caused by incomplete
conversion of the acetonide hydrolysis on a large scale. As a
higher yield can be obtained, this step will be validated by future
optimization studies.
traditional Yamaguchi method,34 led to satisfactory and con-
sistent results, and ester 26 was isolated in 70-92% yield.
Removal of both PMB protecting groups in one step from
26 proved to be nontrivial (Scheme 9). Treatment of ester 26
with excess DDQ under buffered conditions gave a mixture
containing mono-deprotection product 28, diol 21 and lactol
29. One interesting observation was that cleavage of the C7
PMB ether was much faster than the one at the C23 position.
However, this reaction did not afford complete conversion to
diol 21 without generating byproduct 29. Addition of 2,2-
dimethoxypropane has been attempted to prevent the undesirable
ketal hydrolysis, but this did not help. After fine-tuning of
reaction concentration and the DDQ stoichiometry, we settled
on quenching the reaction at a point, which provided 52% yield
of mono-deprotection product 28 and 46% yield of double-
deprotection product 21 with minimum formation of byproduct
29. The mono-PMB ether 28 can be recycled and resubjected
to the DDQ deprotection conditions to provide more diol 21 in
an overall yield of around 72% from ester 26.
With diyne 21 in hand, the stage was now set for the
macrocyclization (eq a). To our delight, the desired macro-
cycle 30 was provided in 22% yield (44% brsm) when a
solution of diyne 21 in benzene was slowly added via syringe
pump to the catalyst solution that contains 12 mol %
Pd(OAc)2 and 15 mol % tri(2,6-dimethoxyphenyl)-phosphine
(TDMPP) (Table 2, entry 1). Note that a slightly higher
ligand/Pd ratio (1.25:1) was used here, since a 1:1 ratio
proved less efficient. Solvent effects were next examined.
Use of toluene as the solvent proved to be most effective,
providing up to 56% yield of macrocycle 30 (entry 3). In
contrast, use of THF as the solvent gave a sluggish reaction
and only 23% yield was obtained after 5 days (entry 2). Like
other macrocyclizations, low concentration (0.002M) proved
to be critical; otherwise, formation of the dimeric byproducts
Esterification between acid 22 and alcohol 23 was studied
under several conditions (Scheme 8). Use of anhydride 27 as
the coupling reagent has recently been developed by Shiina,33
which has been demonstrated as a mild and efficient esterifi-
cation method. Indeed, use of Shiina’s method provided the
desired ester (26), albeit only in 15-19% yield. Under the
reaction conditions, acid 22 was partially decomposed to its
desilylated precursor 24. This reaction also suffered from
reproducibility issues on larger scale. Switching to the more
(31) For the synthesis of compound 4, see ref 13f.
(32) Rai, A. N.; Basu, A. Tetrahedron Lett. 2003, 44, 2267.
(33) (a) Shiina, I.; Kubota, M.; Oshiumi, H.; Hashizume, M. J. Org. Chem.
2004, 69, 1822. (b) Shiina, I.; Kubota, M.; Ibuka, R. Tetrahedron Lett.
2002, 43, 7535. (c) Shiina, I.; Ibuka, R.; Kubota, M. Chem. Lett. 2002,
286.
(34) Inanaga, J.; Hirata, K.; Saeki, H.; Katsuki, T.; Yamaguchi, M. Bull.
Chem. Soc. Jpn. 1979, 52, 1989.
9
16408 J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010

Syntheses of Bryostatin 16 and C20-epi-Bryostatin 7
A R T I C L E S
Scheme 9. PMB Deprotection
Table 2. Macrocyclization via Alkyne-Alkyne Coupling
entry
solvent
yield, %
1
benzene
THF
toluene
toluene
22 (44% brsm)
23 (5 d)
56
2
3a
4b
36 (57% brsm)
a On 3.2 mg scale. b On 45 mg scale.
could be observed. This macrocyclization was also performed
on a 45-mg scale, but the conversion was lower than the
smaller scale reaction (entry 4). To the best of our knowledge,
this represents the first example of using a Pd-catalyzed
alkyne-alkyne coupling as a macrocyclization method for
complex natural product synthesis, which illustrates a new
avenue of using C-C bond formation to construct a
macrocycle.
bias of the macrocycle may interfere with the desired process.
To our delight, use of the cationic gold catalyst gave satisfactory
results (Scheme 11). When 20 mol % catalyst was employed,
the THP product (34) was isolated in 56-73% yield. Notably,
likely due to the Lewis acidity of the catalyst, the methyl ketal
moiety was hydrolyzed under the cyclization conditions.
Subsequent pivalation of the hindered secondary alcohol under
rather forcing conditions (Piv2O 50 equiv, DMAP 80 equiv, 50
°C)36 did afford the pivalate ester (35) in 62% yield. Other
acylation methods, such as use of pivaloyl chloride/Py/DMAP,
proved to be less efficient.
Mechanistically,35 the Pd catalyst chemoselectively inserts
into the C-H bond of the terminal alkyne and then eliminates
one molecule of acetic acid. Subsequent coordination with
the disubstituted alkyne then sets the stage for the chemo-
and regioselective intramolecular migratory insertion. The
resulting vinylpalladium motif is next protonated by the acetic
acid to regenerate the Pd(OAc)2-TDMPP catalyst (Scheme
10).
The following global deprotection turned out to be nontrivial.
Treatment of pivalate ester 35 with HF-pyridine or aqueous HF
provided a four-component mixture, which could be further
separated into two mixtures: one containing bryostatin 16 (36)
and one containing bryostatin 17 (37) (eq 6). The ratio between
The remaining challenge was to conduct a 6-endo-dig
cyclization to form the C-ring of bryostatin. This cyclization
has to proceed within the macrocycle, and the conformational
1
mixtures 36 and 37 was roughly 1.5 to 1, determined via H
NMR spectroscopy. Bryostatin 17 was likely formed via the
acid-catalyzed isomerization of the C21-C34 olefin, but what
(35) For a mechanistic discussion and a full paper, see: (a) Trost, B. M.;
Sorum, M. T.; Chan, C.; Ruehter, G. J. Am. Chem. Soc. 1997, 119,
698.
(36) For a similar pivalation, see ref 10.
9
J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010 16409

A R T I C L E S
Trost and Dong
Scheme 10. Plausible Mechanism for the Pd-Catalyzed Macrocyclization
Scheme 11. Synthesis of THP 35
were the other two compounds isolated with bryostatin 16 and
17?
that this may not be true when the THP was fused in a macrocycle
and there is a possibility to form both hemiketal isomers. In
addition, when the A-ring conformation of the 36/37 mixtures was
locked by forming the methyl ketals, the formed products contained
only two compounds instead of four (eq 7), which serves as another
evidence for our hypothesis, that the two unknown compounds
are the C9 epimers of bryostatins 16 and 17.
We hypothesized that these two mysterious compounds could
be the C9 hemiketal diastereomers of the natural products. Although
the hemiketal isomer of the THP A-ring drawn in the natural
product could be the thermodynamically most stable one if secluded
by itself, our previous bryostatin analogue synthesis13f indicated
9
16410 J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010

Syntheses of Bryostatin 16 and C20-epi-Bryostatin 7
A R T I C L E S
methanol was used as the additive. With this variation, the yield
for this macrocyclization was increased to 65% (72% brsm),
and the reaction rate was also enhanced (40 h). The exact reason
why an alcohol additive improved both the reaction yield and
rate is unclear. We conjecture that a proton source might
facilitate protonation of the vinyl palladium intermediate gener-
ated in the catalytic cycle (see Scheme 10), which eventually
increases the turnover efficiency of the catalyst. The conditions
for the 6-endo-dig cyclization were next optimized. When
macrocycle 40 was subjected to the conditions developed earlier
[AuCl(PPh3) (20 mol %), AgSbF6 (20 mol %), NaHCO3, DCM/
CH3CN, 0 °C to rt], in contrast to the reaction of 30, a complex
mixture, containing the desired product (41), the product with
methyl-ketal hydrolyzed, the starting material (40) and the
starting material with methyl-ketal hydrolyzed was observed.
Switching to a platinum catalyst38 [(PtCl2(CH2dCH2))2 (20 mol
%), NaHCO3, DCM or ether, rt] did save the methyl ketal,
however, the C21-C34 olefin was isomerized under those
reaction conditions. Finally, the problem was solved by adding
2,2-dimethoxypropane to the Au-catalyzed cyclization reaction.
To our delight, under the modified conditions, dihydropyran 41
was obtained in 80-83% yield with the methyl ketal remaining
intact. During this reaction, 2,2-dimethoxypropane was believed
to act as both an acid scavenger and a methyl ketal repairer.
With a scalable route to access the bryostatin 16 intermediate
(41), efforts were next taken to explore the conditions for the
chemoselective and diastereoselective oxidation of the C19-C20
olefin in the presence of three other olefins. An asymmetric
dihydroxylation reaction seemed to be the most straightforward
method to install two oxygen functionalities stereoselectively
on both of the C19 and C20 carbons. Under the Sharpless’
asymmetric dihydroxylation (SAD) conditions,39 to our surprise,
the oxidation occurred almost exclusively at the C13-C30
olefin, the olefin that was supposed to be least reactive toward
oxidation due to its electron-deficient nature (eq 9). As a result,
diol 43 was isolated as the major product in 70% yield; while
the more electron-rich C19-20 alkene remains intact. The
structure of diol 43 has been carefully confirmed by 1H NMR,
IR, gCOSY, gHSQC, gHMBC, ROSEY, and HRMS. The
stereochemistry of the formed diols was tentatively assigned
according to the Sharpless model.39 This unusual chemoselec-
tivity could be explained plausibly by the fact that SAD reaction
is more sensitive to steric factors than electronic ones due to
the bulkiness of the chiral Os catalyst.
Figure 2
Given the extreme acid-sensitivity of this natural product,
we switched to basic desilylation conditions. Fortunately,
treatment of 35 with 5 equiv of TBAF and direct purification
by reverse phase HPLC37 successfully provided bryostatin 16
(eq 8), which was spectroscopically identical to the literature
(reported optical rotation [R]D + 84, c 0.43, MeOH; found [R]D
+ 81, c 0.04, MeOH).14
Total Syntheses of 20-epi-Bryostatin 7. With completion of
the total synthesis of bryostatin 16, we decided to address the
question of how to convert brystatin 16 or a bryostatin 16-like
compound to other structurally related bryostatins or their
analogues (Figure 2).
As illustrated in Scheme 12, during our bryostatin 16
synthesis, the yields for the last several steps were only
moderate; and for certain steps, such as DDQ deprotection and
macrocyclization, conversion was an issue. To permit enough
materials to advance those intermediates, the initial goal was
to improve or modify these challenging steps.
As we observed earlier, the PMB group on the C7 alcohol
was cleaved faster than the one on the C23 alcohol. By carefully
controlling the DDQ stoichiometry (2 equiv) and the reaction
temperature (0 °C), the C7 PMB group was chemoselectively
removed, providing alcohol 28 in 77% yield (Scheme 13).
Subsequent acylation of the secondary C7 alcohol with acetic
anhydride and pyridine afforded the corresponding ester in 91%
yield. Due to the electron-withdrawing nature of the OAc group,
the C9 ketal of the resultant ester became somewhat stabilized
and thus less sensitive to acid than its precursor 28. Conse-
quently, treatment with excess DDQ (10 equiv) at room
temperature cleanly led to the second PMB-cleaved product (39)
in 90% yield.
Macrocyclization with diyne precursor 39 under the condi-
tions developed earlier was effective; however, the yield for
macrocycle 40 was only moderate: 41% (71% brsm), and the
reaction was slow (3 days). Having surveyed several factors
for this Pd-catalyzed alkyne-alkyne coupling, we eventually
found that addition of a mild proton source, like an alcohol,
was beneficial. As precursor 39 has a methyl acetal moiety,
Epoxidation is known to be more sensitive to electronic
effects compared to steric effects, thereby a strategy of epoxi-
(38) Liu, B.; De Brabander, J. K. Org. Lett. 2006, 8, 4907.
(39) For a review, see: Kolb, H. C.; Van Nieuwenhze, M. S.; Sharpless,
K. B. Chem. ReV. 1994, 94, 2483.
(37) Isomerization occurred even when trying to purify bryostatin 16 via
prep-TLC.
9
J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010 16411

A R T I C L E S
Trost and Dong
Scheme 12. Some Challenging Steps in the Bryostatin 16 Synthesis
Scheme 13. Synthesis of Key Intermediate 41
dation followed by ring-opening with an alcohol was next
pursued (eq 14). Although trifluoroperacetic acid (TFPAA) was
used to oxidize a less complicated substrate containing a similar
C-ring motif,13f reaction of 41 under the same conditions only
resulted in decomposition of the starting material (Table 3, entry
1). Payne oxidation,40 with use of either CH3CN or CCl3CN as
the oxidant presursor, failed to provide any identifiable products
(entries 2 and 3). On the other hand, a Re-catalyzed epoxidation,
using methyl rhenium trioxide (MTO) as the catalyst and
urea-hydrogen peroxide (UHP) as the oxidant was promising.41
Under these conditions, the epoxidation/ring-opening product
(44) was isolated in around 10% yield, along with some TES
cleavage products (entry 4). Although the yield was low, we
were encouraged about the chemoselectivity.
We assumed that TES cleavage was caused by the acid
generated during the reaction, thus buffering of the reaction with
a base should minimize the undesirable desilylation. Moreover,
it is also known that Lewis base can act as a ligand to accelerate
the Re-catalyzed epoxidation reaction.42 Indeed, when N-
methylimidazole (50 mol %) was added, the reaction proceed
with full conversion in two hours at 0 °C. Surprisingly, epoxide
45 turned out to be very stable under those reaction conditions,
and it could be isolated via aqueous workup (Scheme 14).43
Epoxide 45 was subsequently ring-opened with MeOH, either
by addition of ZnCl2 solution to the reaction mixture in a one-
pot fashion or treatment of the crude epoxide with dilute HOAc
in MeOH, providing C20 alcohol 44 in 48% or 64% yield
respectively. It is not totally unexpected that the stereochemistry
(40) (a) Payne, G. B.; Deming, P. H.; Williams, P. H. J. Org. Chem. 1961,
26, 659. (b) Arias, L. A.; Adkins, S.; Nagel, C. J.; Bach, R. D. J.
Org. Chem. 1983, 48, 888.
(41) For a review, see: Kuhn, F. E.; Santos, A. M.; Herrmann, W. A. Dalton
Trans. 2005, 2483. Also, see: (a) Herrmann, W. A.; Fischer, R. W.;
Marz, D. W. Angew. Chem., Int. Ed. 1991, 30, 1638. (b) Boehlow,
T. R.; Spilling, C. D. Tetrahedron Lett. 1996, 37, 2717. (c) Soldaini,
G.; Cardona, F.; Goti, A. Tetrahedron Lett. 2003, 44, 5589.
(42) Rudolph, J.; Reddy, K. L.; Chiang, J. P.; Sharpless, K. B. J. Am. Chem.
Soc. 1997, 119, 6189.
(43) Epoxide 45 decomposed on silica gel.
9
16412 J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010

Syntheses of Bryostatin 16 and C20-epi-Bryostatin 7
A R T I C L E S
Table 3. An Epoxidation/Ring-Opening Strategy
entry
conditions
results
1
2
3
4
TFPAA, CH3CN, Na2HPO4, DCM/MeOH 0 °C to rt
CCl3CN, UHP, Na2HPO4, DCM/MeOH 0 °C to rt
CH3CN, UHP, KHCO3, MeOH rt
decomposition of 41
complete decomposition
complex mixture
MTO (10 mol %), UHP (2 equiv), MeOH, 0 °C
44 (ca. 10%) + TES cleavage products
Scheme 14. Synthesis of Alcohol 44
Scheme 15. End Game of the Synthesis of 20-epi-Bryostatin 7 (47)
of the C20 alcohol in 44 was opposite to the one in the natural
bryostastin,44 as the Re catalyst likely attacks at the less hindered
face of the C-ring at the epoxidation stage. However, it
represented the feasibility to function the C19-C20 olefin
chemoselectiVely in a bryostatin-16 like compound!
Synthesis of 20-epi-Bryostatin 7. We envisaged that the
unnatural C20-epimer of bryostatins, could serve as an attractive
new family of bryostatin analogues for two reasons: first, they
might retain the biological activities that are comparable or
complementary to those of the natural products due to their
closely related structures; second, study of these analogues could
contribute to our understanding of the structure-activity
relationships (SAR) of bryostatins, especially about the role of
the C-ring unit generally and the C20 stereocenter specifically.
Consequently, we explored the synthesis of the analogue 20-
epi-bryostatin 7. Acylation of the C20 alcohol with acetic
anhydride, followed by global deprotection with aqueous HF,
afforded 20-epi-bryostatin 7 (47) in 63% yield over two steps
(Scheme 15). The structure of 47 was confirmed by 1H, gCOSY,
gHSQC, gHMBC, IR and HRMS.45 Note that we are able to
install the C7 and C20 ester groups in 47 at different stages,
thus this route also represents a valid access to other bryostatin
analogues.
20-epi-Bryostatin 7 was next tested against several cancer
cell lines in a biological assay. Initial biological studies showed
that this bryostatin analogue possessed nanomolar potency
against DOHH2 (a lymphoma cancer cell line), Granta 519 (a
lymphoma cancer cell line), and Jurkat (a T-lymphocyte cancer
cell line) (Table 4). These biological studies indicate that this
new bryostatin analogue could be a potential drug lead for anti-
cancer chemotherapy.
Conclusion
In summary, we have developed a unique and highly concise
strategy (26 steps in the longest linear sequence, 39 total steps
(44) A very small ratio of the desired diastereomer was observed, and the
dr was about 7:1 favoring the undesired one.
(45) The stereochemistry of the C19 hemi-ketal was tentatively assigned
as the same one in the natural product.
9
J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010 16413

A R T I C L E S
Trost and Dong
Table 4. Anticancer Activity of 20-epi-Bryostatin 7 (47)a
-21.1 (c 1.27 g cm-3 in DCM); H NMR (CDCl3, 500 MHz) δ
7.63-7.59 (m, 4H), 7.47-7.44 (m, 2H), 7.43-7.37 (m, 4H), 7.13
(d, J ) 8.5 Hz, 2H), 6.76 (d, J ) 8.5 Hz, 2H), 5.63 (dd, J ) 0.5,
16 Hz, 1H), 5.40 (dd, J ) 7.0, 16 Hz, 1H), 5.26 (s, 1H), 4.55 (d,
J ) 11 Hz, 1H), 4.34 (d, J ) 11 Hz, 1H), 4.03-3.98 (m, 2H),
3.88-3.80 (m, 2H), 3.73 (m, 1H), 3.73 (s, 3H), 3.25 (s, 2H), 3.00
(dd, J ) 5.5, 17 Hz, 1H), 2.53 (dd, J ) 7.0, 18 Hz, 1H), 2.47-2.38
(m, 3H), 2.23 (br d, J ) 13 Hz, 1H), 1.97 (br dd, J ) 12, 24 Hz,
2H), 1.90 (ddd, J ) 2.5, 5.5, 14 Hz, 1H), 1.58-1.55 (m, 2H), 1.48
(dd, J ) 1.5, 10.5, Hz, 1H), 1.18 (s, 3H), 1.08 (s, 3H), 1.03 (s,
9H), 0.93 (s, 3H), 0.92 (s, 3H), 0.88 (s, 9H), 0.09 (s, 9H), 0.00 (s,
6H); 13C NMR (CDCl3, 125 MHz) δ 212.3, 170.4, 159.2, 152.9,
139.2, 135.73, 135.69, 133.4, 133.2, 130.7, 130.12, 130.06, 129.6,
128.0, 127.9, 127.7, 123.7, 113.8, 79.4, 79.3, 75.1, 74.6, 73.2, 71.8,
65.2, 55.3, 52.6, 45.3, 45.2, 40.6, 39.7, 38.9, 38.1, 37.9, 30.4, 26.9,
26.0, 24.0, 23.7, 20.9, 20.8, 19.1, 18.3, 0.4, -5.4; IR (film): 2956,
2858, 1744, 1702, 1612, 1514, 1249, 1094, 838 cm-1; HRMS
(C57H86O8Si3): calcd 1005.5528 (M + Na+), found 1005.5520.
Compound 13. CSA (18 mg, 0.080 mmol) was added to a
solution of compound 12 (0.88 g, 0.89 mmol) in MeOH (dry, 18
mL) at 0 °C. The resulting solution was stirred at rt for 12 h, before
it was poured into saturated aqueous NaHCO3. The mixture was
extracted with ethyl acetate three times and the combined organic
fractions were dried over Na2SO4. Compound 13 was purified via
silica gel flash column chromatography (15%, then 30% ethyl
acetate/petroleum ether) to give a colorless oil (0.76 g, 93%). RF:
1
cell lines
DOHH2
Granta 519
Jurkat
IC50 (nM)
22.5
17.6
44.7
a DOHH2, a lymphoma cancer cell line; Granta 519, a lymphoma
cancer cell line; Jurkat, a T-lymphocyte cancer cell line.
from aldehyde 10, eq 11) for the asymmetric total synthesis of
bryostatin 16. A Pd-catalyzed alkyne-alkyne coupling was
employed for the first-time as a macrocyclization method in
natural product synthesis. The efficiency of our synthesis can
also be attributed to a tandem Ru-catalyzed alkene-alkyne
coupling/Michael addition to form the B-ring, an acid-catalyzed
one-pot cascade to form the A-ring, a directed chemoselective
ester hydrolysis, and a palladium/gold-catalyzed cascade to form
the C-ring of bryostatin 16. These atom-economical and/or
chemoselective approaches not only are useful in bryostatin
syntheses, but should also be indicative for the synthesis of
numerous other polyacetate-polypropionate-derived natural
products.
0.2 (ethyl acetate:petroleum ether, 1:4 v/v); [R]D (deg cm3 g-1
20
dm-1): +27.5 (c 0.85 g cm-3 in DCM); 1H NMR (C6D6, 400 MHz)
δ 7.84-7.80 (m, 4H), 7.26-7.21 (m, 6H), 6.80 (d, J ) 8.8 Hz,
2H), 5.98 (s, 1H), 5.71 (dd, J ) 1.2, 16 Hz, 1H), 5.52 (dd, J ) 5.2,
16 Hz, 1H), 4.58 (m, 1H), 4.38 (d, J ) 11.2, 1H), 4.18 (d, J )
11.2 Hz, 1H), 3.71-3.64 (m, 2H), 3.57 (m, 1H), 3.45 (m, 1H),
3.38 (s, 3H), 3.31 (s, 3H), 3.13 (s, 2H), 2.97 (m, 1H), 2.90 (s, 3H),
2.75-2.65 (m, 2H), 2.26 (brd, J ) 12 Hz, 1H), 2.18 (dd, J ) 4.8,
16 Hz, 1H), 1.89 (m, 1H), 1.81-1.65 (m, 4H), 1.51 (m, 1H), 1.20
(s, 3H), 1.17 (s, 9H), 1.14 (s, 3H), 0.91 (s, 3H), 0.90 (s, 3H); 13C
NMR (C6D6, 100 MHz) δ 171.6, 159.6, 140.8, 138.2, 136.30,
136.26, 134.4, 134.2, 131.8, 130.1, 129.3, 129.2, 128.0, 127.9,
114.0, 104.3, 100.5, 78.1, 77.2, 74.6, 71.4, 71.2, 69.7, 66.5, 54.7,
51.2, 48.0, 44.7, 43.6, 43.5, 42.4, 39.0, 38.2, 37.6, 33.2, 27.1, 23.9,
23.7, 21.1, 19.5, 16.8; IR (film): 3444, 2933, 1738, 1614, 1514,
1248 cm-1; HRMS (C50H69O9BrSi): calcd 943.3792 (M + Na+),
found 943.3801.
Compound 26. To a solution of hydroxyacid 24 (74 mg, 0.12
mmol) in DCM (2.5 mL) was added freshly distilled 2,6-lutidine
(62 mg, 0.58 mmol) at -10 °C, followed by dropwise addition of
freshly distilled TESOTf (67 mg, 0.25 mmol). The resulting solution
was stirred at the same temperature for 20 min, before poured into
pH 7.0 buffer. The mixture was extracted with ethyl acetate five
times and the combined organic fractions were dried over Na2SO4.
The TES ether-acid 22 was purified via quick silica gel flash
column chromatography (10%, 20% then 30% ethyl acetate/
petroleum ether) to give a colorless foam (73 mg, 79%). (Significant
decomposition has been observed when slower chromatography was
applied.)
In addition, we demonstrated the feasibility to functionalize
the C-ring of bryostatin 16 by a highly chemo- and stereose-
lective Re-catalyzed epoxidation/ring-opening reaction. By
accomplishing a concise synthesis of a potent anti-cancer agent
20-epi-bryostatin 7, we also proved the principle for the first
time that novel bryostatin analogues can be derived from a
bryostatin 16-like intermediate (eq 12). Extension of this strategy
in the synthesis of various other natural bryostatins and their
analogues, and performance of more systematic biological
studies are currently being undertaken.
To a solution of TES ether-acid 22 (17.0 mg, 0.0225 mmol) in
dry toluene (0.5 mL) was added Et3N (4.8 mg, 0.047 mmol) at rt
under N2, followed by dropwise addition of freshly distilled 2,4,6-
trichlorobenzoyl chloride (5.6 mg, 0.024 mmol) at rt. The resulting
solution was stirred at rt for 1 h, before a solution of alcohol 23
(10.1 mg, 0.0225 mmol) and DMAP (6.9 mg, 0.056 mmol) in
toluene (0.75 mL) was added. The resulting mixture was stirred at
rt for another 1 h, before poured into pH 7.0 buffer. The mixture
was extracted with ethyl acetate four times and the combined
organic fractions were dried over Na2SO4. The ester 26 was purified
via silica gel flash column chromatography (10%, then 20% ethyl
acetate/petroleum ether) to give a colorless foam (25 mg, 92%).
Experimental Section
Compound 5. CpRu(CH3CN)3PF6 (4.0 mg, 0.0092 mmol) was
added to a solution of compound 6 (50 mg, 0.080 mmol) and
compound 7 (24.5 mg, 0.069 mmol) in DCM (0.4 mL) at 0 °C.
The resulting yellow solution was stirred at rt for 12 h. Compound
5 was purified directly via silica gel flash column chromatography
(10%, then 15% ethyl acetate/petroleum ether) to give a colorless
foam (23.1 mg, 34%; 80% brsm, 45.6 mg 6 + 7 can be recovered).
At a larger scale, compound 6 (969 mg, 1.59 mmol) and
compound 7 (471 mg, 1.33 mmol) with CpRu(CH3CN)3PF6 (86
mg, 0.199 mmol) in DCM (3 mL), according to the same procedure,
gave compound 5 (0.45 g, 35%, 0.19 g, 7 was recovered). RF: 0.3
(ethyl acetate:petroleum ether, 1:9 v/v); [R]D20 (deg cm3 g-1 dm-1):
RF: 0.35 (ethyl acetate:petroleum ether, 1:4 v/v); [R]D (deg cm3
20
1
g-1 dm-1): +62.6 (c 0.11 g cm-3 in DCM); H NMR (C6D6, 500
9
16414 J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010

Syntheses of Bryostatin 16 and C20-epi-Bryostatin 7
A R T I C L E S
MHz) δ 7.41 (d, J ) 8.5 Hz, 2H), 7.24 (d, J ) 8.5 Hz, 2H), 6.85
(d, J ) 8.5 Hz, 2H), 6.79 (d, J ) 8.5 Hz, 2H), 6.07 (dd, J ) 5.0,
15.5 Hz, 1H), 5.93 (s, 1H), 5.79 (dd, J ) 1.5, 15.5 Hz, 1H), 5.48
(ddd, J ) 2.0, 4.0, 10.5 Hz, 1H), 4.60 (m, 1H), 4.53 (d, J ) 10.5,
1 H), 4.47 (d, J ) 11.5 Hz, 1H), 4.42 (d, J ) 10.5 Hz, 1H), 4.32
(brd, J ) 15.5 Hz, 1H), 4.27 (d, J ) 11.5 Hz, 1H), 3.97 (m, 1H),
3.88-3.83 (2H), 3.79 (m, 1H), 3.67 (m, 1H), 3.45 (s, 3H), 3.33 (s,
3H), 3.30 (s, 3H), 3.28 (s, 3H), 3.27 (s, 3H), 2.70 (br s, 1H), 2.69
(d, J ) 2.0 Hz, 1H), 2.38-2.25 (4H), 2.07-1.89 (6H), 2.01 (s,
1H), 1.84 (dd, J ) 5.0, 16 Hz, 1H), 1.80 (m, 1H), 1.65 (m, 1H),
1.57 (dd, J ) 9.5, 19.5 Hz, 2H), 1.29 (s, 3H), 1.22 (s, 3H), 1.21 (s,
6H), 1.18 (d, J ) 6.0 Hz, 3H), 1.07 (t, J ) 6.5 Hz, 9H), 1.00 (3,
9H), 0.72 (m, 6H), 0.17 (s, 3H), 0.12 (s, 3H); 13C NMR (C6D6,
125 MHz) δ 170.9, 166.7, 159.8, 159.6, 157.7, 153.9, 136.8, 131.7,
130.5, 130.1, 129.3, 128.7, 128.3, 128.1, 127.9, 114.9, 114.1,
114.03, 113.96, 104.5, 89.4, 85.8, 78.5, 77.5, 75.6, 75.1, 73.6, 73.4,
72.1, 71.5, 70.5, 68.7, 67.8, 66.2, 54.74, 54.70, 52.0, 50.6, 48.3,
44.9, 44.3, 43.7, 43.6, 39.5, 36.4, 34.4, 33.8, 33.5, 29.8, 26.01,
25.96, 24.5, 21.2, 18.6, 18.2, 17.0, 7.3, 5.7, -4.6, -4.7; IR (film):
1605, 1614, 1433, 1375, 1259, 1154, 1099 cm-1; HRMS
(C42H62O14): calcd 913.4037 (M + Na+), found 913.4038.
Compound 41. To a mixture of Au(PPh3)3Cl (10.2 mg, 0.020
mmol) and AgSbF6 (7.0 mg, 0.020 mmol) was added dry DCM
(0.5 mL) at rt under N2. The resulting mixture was stirred in the
dark for 15 min, and a purple precipitate was formed. The
supernatant solution (0.015 mL, ca. 0.0006 mmol) was transferred
to a mixture of compound 40 (2.9 mg, 0.0030 mmol) and NaHCO3
(2.4 mg, 0.03 mmol) in DCM/CH3CN/2,2-dimethoxypropane, (10:
1:2, 0.4 mL) at 0 °C under N2. The resulting reaction mixture was
stirred vigorously overnight, before it was poured into a mixture
of saturated aqueous NaHCO3 and saturated aqueous Na2S2O3 (ca.
1:1), and then the mixture was extracted with ethyl acetate four
times and the combined organic fractions were dried over Na2SO4.
Dihydropyran 41 was purified via quick silica gel flash column
chromatography (10%, then 20% ethyl acetate/petroleum ether) to
give a colorless foam (2.4 mg, 83%): Rf: 0.3 (10% ethyl acetace in
1
petroleum ether); [R]D: 42.5 (c 0.17, DCM); H NMR (C6D6, 500
MHz): δ 6.13 (d, J ) 15.5 Hz, 1H), 5.74 (s, 1H), 5.73 (dd, J )
4.5, 15.5 Hz, 1H), 5.67 (dd, J ) 5.0, 12 Hz, 1H), 5.61 (s, 1H),
5.46 (dd, J ) 4.5, 11 Hz, 1H), 5.37 (s, 1H), 4.63 (m, 1H), 4.37 (d,
J ) 13 Hz, 1H), 4.02-3.91 (m, 4H), 3.77 (t, J ) 12 Hz, 1H), 3.43
(s, 3H), 3.41 (s, 3H), 3.11 (s, 3H), 2.68 (dd, J ) 5.0, 15 Hz, 1H),
2.42-2.34 (m, 2H), 2.18 (dd, J ) 8.5, 16 Hz, 1H), 2.05-1.84 (m,
5H), 1.78 (m, 1H), 1.75 (s, 3H), 1.65-1.45 (5H) 1.30 (s, 3H), 1.24
(s, 3H), 1.12 (s, 3H), 1.09 (s, 3H), 1.05 (d, J ) 7.0 Hz, 3H), 1.03
(s, 9H), 0.97 (t, J ) 8.0 Hz, 9H), 0.57 (m, 6H), 0.23 (s, 3H), 0.13
(s, 3H); 13C NMR(C6D6, 125 MHz) δ 170.0, 169.8, 169.3, 167.3,
158.2, 150.5, 136.2, 129.1, 128.5, 127.8, 125.0, 114.7, 108.9, 103.2,
100.9, 77.0, 73.8, 73.7, 73.4, 72.4, 68.1, 66.8, 64.7, 50.5, 50.4, 48.2,
44.8, 44.0, 42.7, 42.2, 41.0, 39.7, 37.0, 33.8, 33.7, 32.4, 30.2, 26.0,
25.0, 24.9, 20.72, 20.70, 18.3, 18.0, 17.9, 7.2, 6.0, -4.7; IR (film)
2927, 2240, 1717, 1651, 1614, 1514, 1463, 1377, 1250, 1075 cm-1
;
HRMS (C66H100O15Si2): calcd 1211.6499 (M + Na+), found
1211.6484.
Compound 30. To a mixture of Pd(OAc)2 (4.4 mg, 0.02 mmol)
and TDMPP [tris(2,6-dimethoxyphenyl)phosphine] (11.2 mg, 0.025
mmol) was added freshly distilled toluene (1 mL). The mixture
was stirred at rt for 30 min, and the resulting red solution (0.02
mL, ca. 0.0004 mmol) was slowly added a solution of diyne 21
(3.2 mg, 0.0034 mmol) in freshly distilled toluene (1.6 mL) under
N2. The reaction was stirred at rt for 3 days, before it was filtered
through a short plug of silica gel. The solvent was removed under
vacuum and the macrocycle 30 was purified via silica gel flash
column chromatography (20%, 30% then 40% ethyl acetate/
petroleum ether) to give a white paste (1.8 mg, 56%).
The same reaction was also carried out with Pd(OAc)2 (1.1 mg,
0.0048 mmol), TDMPP (1.6 mg, 0.0075 mmol), diyne 21 (45 mg,
0.048 mmol) in toluene (20 mL) to give macrocycle 30 (16.0 mg,
36% yield; 16.7 mg 21 was recover, 57% yield brsm). RF: 0.35
(ethyl acetate:petroleum ether, 3:7 v/v); [R]D20 (deg cm3 g-1 dm-1):
-43.8 (c 0.21 g cm-3 in DCM); 1H NMR (C6D6, 500 MHz) δ 6.32
(s, 1H), 6.12 (dd, J ) 3.5, 15.5 Hz, 1H), 5.89 (d, J ) 15 Hz, 1H),
5.73 (s, 1H), 5.34 (d, J ) 10.5 Hz, 1H), 4.67 (m, 1H), 4.36 (d, J
) 13 Hz, 1H), 4.31 (m, 1H), 4.12 (d, J ) 11 Hz, 1H), 4.03-4.00
(2H), 3.91 (m, 1H), 3.86 (m, 1H), 3.40 (s, 3H), 3.38 (t, J ) 7.5
Hz, 1H), 3.26 (s, 3H), 3.21 (s, 3H), 3.16 (dd, J ) 5.0, 14.5 Hz,
1H), 2.68 (dd, J ) 3.0, 16 Hz, 1H), 2.62 (dd, J ) 9.0, 16 Hz, 1H),
2.21 (dd, J ) 8.0, 16 Hz, 1H), 2.00 (m 1H), 1.92-1.84 (4H),
1.65-1.53 (3H), 1.46 (m, 1H), 1.36-1.29 (2H), 1.24 (s, 3H), 1.22
(s, 3H), 1.19 (d, J ) 6.5 Hz, 3H), 1.10 (s, 3H), 1.09 (s, 3H), 1.07
(t, J ) 8.0 Hz, 9H), 0.98 (s, 9H), 0.66 (m, 6H), 0.16 (s, 3H), 0.11
(s, 3H); 13C NMR (C6D6, 125 MHz) δ 172.3, 166.5, 157.7, 140.2,
134.9, 129.1, 128.2, 127.9, 125.1, 114.8, 103.5, 102.0, 83.9, 76.4,
75.1, 74.3, 70.0, 68.7, 67.4, 66.9, 65.9, 50.9, 50.6, 49.1, 46.5, 44.6,
43.2, 42.8, 40.6, 40.2, 37.2, 36.8, 36.4, 34.6, 30.2, 29.8, 29.6, 26.0,
20.6, 18.5, 18.3, 16.6, 7.35, 6.0, -4.68, -4.72; IR (film): 3442
(br), 2924, 2853, 1717, 1650, 1614, 1435, 1376, 1256, 1151, 1107
cm-1; HRMS (C50H84O13Si2): calcd 971.5348 (M + Na+), found
971.5341.
2953, 2929, 1734, 1608, 1614, 1435, 1375, 1245, 1150, 1102 cm-1
;
HRMS (C52H86O14Si2): calcd 1013.5454 ([M + Na]+), found
1013.5453.
Compound 44. UHP (37.6 mg, 0.4 mmol) was added to a
solution of MTO (5.0 mg, 0.02 mmol), N-methylimidazole (8.2
mg, 0.1 mmol) in freshly distilled MeOH (2 mL) at rt. The resulting
solution was stirred at rt for 5 min, during which the color of the
reaction turned to yellow. A portion of the above solution (0.026
mL) was added to a solution of 41 (2.5 mg, 0.0026 mmol) in MeOH
(0.2 mL) at 0 °C. The resulting reaction mixture was stirred at °C
for 6 h (monitored by TLC) before it was quenched with saturate
aqueous NaHCO3 and Na2S2O3. The mixture was extracted with
ethyl acetate four times and the combined organic fractions were
dried over Na2SO4. The crude epoxide product was obtained after
the solvent was removed under vacuum. To the above crude epoxide
was added a solution of HOAc in MeOH (0.2 mL, obtained from
1 drop of HOAc in 1 mL MeOH) at 0 °C. The resulting solution
was stirred at 0 °C for 3 h (monitored by TLC), before it was
quenched with saturate aqueous NaHCO3. The mixture was
extracted with ethyl acetate four times and the combined organic
fractions were dried over Na2SO4. Compound 44 was purified via
silica gel flash column chromatography (10%, then 20% ethyl
acetate/petroleum ether) to give a colorless foam (1.7 mg, 64%).
A One-Pot Protocol. UHP (19 mg, 0.2 mmol) was added to a
solution of MTO (5.0 mg, 0.02 mmol), N-methylimidazole (8.2
mg, 0.1 mmol) in freshly distilled MeOH (1 mL) at rt. The resulting
solution was stirred at rt for 5 min, during which the color of the
reaction turned to yellow. A portion of the above solution (0.021
mL, 10 mol % MTO) was added to a solution of 41 (4.3 mg, 0.0044
mmol) in MeOH (0.3 mL) and DCM (0.2 mL) at 0 °C for 30 min,
before another portion of the oxidant (0.022 mL) was added. The
resulting reaction mixture was stirred at °C for 3 h (monitored by
TLC) before it was cooled to -78 °C and ZnCl2 (0.020 mL, 1.0 M
in ether) was added. The resulting solution was stirred at 4 °C for
4 h (monitored by TLC), before it was quenched with saturate
aqueous NaHCO3 and Na2S2O3. The mixture was extracted with
ethyl acetate four times and the combined organic fractions were
Bryostatin 16. To a solution of pivalate ester 35 (1.0 mg, 0.001
mmol) in THF (0.05 mL) was added TBAF (0.005 mL, 0.005 mmol,
1M) at 0 °C. The resulting solution was allowed to slowly warm
to rt and stirred for 4 h. The reaction mixture was diluted with
ethyl acetate and pH 7.0 buffer was added. The mixture was
extracted with ethyl acetate five times and the combined organic
fractions were dried over Na2SO4. The residue was purified by
reverse phase HPLC (RP C-18 column, CH3CN in H2O from 65%
to 95%) to give 1 as a white paste (0.4 mg, ca. 52%). RF: 0.35
(ethyl acetate:petroleum ether, 4:1 v/v); [R]D20 (deg cm3 g-1 dm-1):
+81 (c 0.04 g cm-3 in MeOH); For NMR data, see the Supporting
Information; IR (film): 3359 (br), 2958, 2917, 2849, 1722, 1702,
9
J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010 16415

A R T I C L E S
Trost and Dong
dried over Na2SO4. Compound 44 was purified via silica gel flash
column chromatography (10%, then 20% ethyl acetate/petroleum
ether) to give a colorless foam (2.2 mg, 48%): Rf: 0.33 (15% ethyl
acetace in petroleum ether); [R]D: 57.7 (c 0.17, DCM); For NMR
data, see the Supporting Information; IR (film): 3400 (br), 2954,
2928, 2856, 1722, 1651, 1378, 1247, 1098 cm-; HRMS (C53H90-
O16Si2): calcd 1061.5665 ([M + Na]+), found 1061.5640.
ether); [R]D: 27.6 (c 0.07, DCM); For NMR data, see the Supporting
Information; IR (film): 3455, 3300(br), 2924, 2853, 1722, 1652,
1374, 1243, 1153, 1077 cm-1; HRMS (C41H60O17): calcd 847.3728
([M + Na]+), found 847.3741.
Acknowledgment. We acknowledge the National Institutes of
Health (GM 13598) for their support of our programs. G.D. was a
Stanford Graduate Fellow. We thank Dr. H. Yang for discussions
and for help characterizing compound 35, and we additionally thank
him and Dr. C. S. Brindle for providing synthetic intermediates.
Palladium and ruthenium salts were supplied by Johnson Matthey.
We are grateful to Dr. J. Flygare and Genentech for performing
the preliminary biological test of 20-epi-bryostatin 7. We also
acknowledge Dr. S. R. Lynch for his help with two-dimensional
nuclear magnetic resonance analysis. We also thank the Wender
group’s assistance with reverse-phase high-performance liquid
chromatography.
Compound 47. To a solution of alcohol 44 (1.0 mg, 0.001 mmol)
in pyridine (0.15 mL) was added acetic anhydride (0.1 mL) at 0
°C, followed by DMAP (1.0 mg, 0.008 mmol). The resulting
solution was stirred at rt for 4 h, before it was quenched with MeOH
(0.1 mL), followed by pH 7.0 Buffer at 0 °C. The mixture was
extracted with ethyl acetate four times and the combined organic
fractions were dried over Na2SO4. The residue was purified by silica
gel column chromatography (10%, then 20% ethyl acetate/petroleum
ether) to give compound 46 (ca. 1.0 mg) as a colorless thick oil.
To a solution of the above acetate (46, ca. 1.0 mg) in CH3CN
(0.2 mL) was added aqueous HF (2 drops, conc. 48-53%) at 0
°C. The resulting solution was stirred at a warming ice-bath for
2.5 h, before solid K2HPO4 and saturated aqueous NaHCO3 were
added. The mixture was extracted with ethyl acetate four times and
the combined organic fractions were dried over Na2SO4. The residue
was purified by preparative TLC (60% ethyl acetace in petroleum
ether) to give 20-epi-bryostatin 7 (47) as a white paste (0.5 mg,
ca. 63% over two steps): Rf: 0.30 (60% ethyl acetace in petroleum
Supporting Information Available: Experimental procedures
and characterization data for all new compounds. This material
is available free of charge via the Internet at http://pubs.acs.org.
JA105129P
9
16416 J. AM. CHEM. SOC. VOL. 132, NO. 46, 2010