Lobatamide C: Total synthesis, stereochemical assignment, preparation of simplified analogues, and V-ATPase inhibition studies

Article abstract of DOI:10.1021/ja0352350

The total synthesis and stereochemical assignment of the potent antitumor macrolide lobatamide C, as well as synthesis of simplified lobatamide analogues, is reported. Cu(I)-mediated enamide formation methodology has been developed to prepare the highly unsaturated enamide side chain of the natural product and analogues. A key fragment coupling employs base-mediated esterification of a β-hydroxy acid and a salicylate cyanomethyl ester. Three additional stereoisomers of lobatamide C have been prepared using related synthetic routes. The stereochemistry at C8, C11, and C15 of lobatamide C was assigned by comparison of stereoisomers and X-ray analysis of a crystalline derivative. Synthetic lobatamide C, stereoisomers, and simplified analogues have been evaluated for inhibition of bovine chromaffin granule membrane V-ATPase. The salicylate phenol, enamide NH, and ortho-substitution of the salicylate ester have been shown to be important for V-ATPase inhibitory activity.

Full text of DOI:10.1021/ja0352350

Published on Web 06/06/2003
Lobatamide C: Total Synthesis, Stereochemical Assignment,
Preparation of Simplified Analogues, and V-ATPase Inhibition
Studies
Ruichao Shen,† Cheng Ting Lin,† Emma Jean Bowman,‡ Barry J. Bowman,‡ and
John A. Porco, Jr.†*
Contribution from the Department of Chemistry and Center for Chemical
Methodology and Library DeVelopment, Boston UniVersity, 590 Commonwealth AVenue,
Boston, Massachusetts 02215, and Department of Molecular, Cell and DeVelopmental
Biology, Sinsheimer Labs, UniVersity of California, Santa Cruz, California 95064
Received March 19, 2003; E-mail: porco@chem.bu.edu
Abstract: The total synthesis and stereochemical assignment of the potent antitumor macrolide lobatamide
C, as well as synthesis of simplified lobatamide analogues, is reported. Cu(I)-mediated enamide formation
methodology has been developed to prepare the highly unsaturated enamide side chain of the natural
product and analogues. A key fragment coupling employs base-mediated esterification of a â-hydroxy
acid and a salicylate cyanomethyl ester. Three additional stereoisomers of lobatamide C have been prepared
using related synthetic routes. The stereochemistry at C8, C11, and C15 of lobatamide C was assigned by
comparison of stereoisomers and X-ray analysis of a crystalline derivative. Synthetic lobatamide C,
stereoisomers, and simplified analogues have been evaluated for inhibition of bovine chromaffin granule
membrane V-ATPase. The salicylate phenol, enamide NH, and ortho-substitution of the salicylate ester
have been shown to be important for V-ATPase inhibitory activity.
tunicate.2 Suzumura et al. also independently isolated YM-
Introduction
75518A-C, identical to lobatamides A-C, and the Z-oxime
Biological metabolites are the subject of ongoing investiga-
tions in the search for new medicinal leads including anti-
infective and anti-cancer agents. Recently, a number of unique
antitumor natural products containing a central benzolactone
core bearing an unusual enamide side chain have been reported.
Members include salicylihalamides A and B,1 lobatamides
A-F,2 apicularens A and B,3 CJ-12,950 and CJ-13,357,4 and
oximidines I and II5 (Figure 1). The lobatamides, containing a
15-membered ring macrodilactone and a divinylcarbinol moiety,
were isolated in 1998 by Boyd et al. from a southwestern Pacific
stereoisomer YM-75518D (Figure 1).6 The absolute stereochem-
istry of the C15 divinylcarbinol of YM-75518A was assigned
as (S) using modified Mosher ester analysis.6b However, the
configurations of the two remaining stereocenters were not deter-
mined and thus required chemical synthesis for full confirmation.
Extensive biological evaluation of lobatamides has been
performed against the National Cancer Institute’s (NCI) 60
human tumor cell line (mean panel GI50’s approximately 1.6
nM).2 Significantly, biological studies indicate that both sali-
cylihalamides and lobatamides represent antitumor natural
products with a novel mechanism of action.2 Recently, it has
been reported that the salicylate enamide macrolides selectively
inhibit vacuolar-type proton ATPases (V-ATPases),7 ubiquitous
proton-translocating pumps of eukaryotic cells.8 Moreover, it
has been found that proton-extruding V-ATPases are expressed
† Boston University.
‡ University of California.
(1) (a) Erickson, K. L.; Beutler, J. A.; Cardellina, J. H., II.; Boyd, M. R. J.
Org. Chem. 1997, 62, 8188. (b) Erickson, K. L.; Beutler, J. A.; Cardellina,
J. H., II.; Boyd, M. R. J. Org. Chem. 2001, 66, 1532. For total synthesis
of salicylihalamides, see: (c) Wu, Y.; Esser, L.; De Brabander, J. K. Angew.
Chem., Int. Ed. 2000, 39, 4308. (d) Labrecque, D.; Charron, S.; Rej, R.;
Blais, C.; Lamothe, S. Tetrahedron Lett. 2001, 42, 2645. (e) Snider, B. B.;
Song, F. Org. Lett. 2001, 3, 1817. (f) Smith, A. B., III; Zheng, J.
Tetrahedron 2002, 58, 6455. (g) Fu¨rstner, A.; Dierkes, T.; Thiel, O. R.;
Blanda, G. Chem. Eur. J. 2001, 7, 5286. (h) Holloway, G. A.; Hugel, H.
M.; Rizzacasa, M. A. J. Org. Chem. 2003, 68, 2200.
(5) (a) Kim, J. W.; Shin-ya, K.; Furihata, K.; Hayakawa, Y.; Seto, H. J. Org.
Chem. 1999, 64, 153. For synthetic studies on the oximidines, see: (b)
Kuramochi, K.; Watanabe, H.; Kitahara, T. Synlett 2000, 397. (c) Scheufler,
F.; Maier, M. E. Synlett 2001, 1221. (d) Coleman, R. S.; Garg, R. Org.
Lett. 2001, 3, 3487.
(6) (a) Suzumura, K.-i.; Takahashi, I.; Matsumoto, H.; Nagai, K.; Setiawan,
B.; Rantiatmodjo, R. M.; Suzuki, K.-i.; Nagano, N. Tetrahedron Lett. 1997,
38, 7573. (b) Suzumura, K.-i.; Takahashi, I.; Matsumoto, H.; Nagai, K.;
Suzuki, K.-i.; Nagano, N. 4th Symposium on the Chemistry of Natural
Products, Nagoya, 1999, Abstract No. 67.
(7) (a) Boyd, M. R.; Farina, C.; Belfiore, P.; Gagliardi, S.; Kim, J. W.;
Hayakawa, Y.; Beutler, J. A.; McKee, T. C.; Bowman, B. J.; Bowman, E.
J. J. Pharm. Exp. Therap. 2001, 297, 114. For a recent review, see: (b)
Beutler, J. A.; McKee, T. C. Curr. Med. Chem. 2003, 10, 787.
(8) For a recent review on V-(H+)-ATPases, see: Nishi, T.; Forgac, M. Nature
ReV. Mol. Cell Biol. 2002, 3, 94.
(2) (a) Galinis, D. L.; McKee, T. C.; Pannell, L. K.; Cardellina, J. H., II; Boyd,
M. R. J. Org. Chem. 1997, 62, 8968. (b) McKee, T. C.; Galinis, D. L.;
Pannell, L. K.; Cardellina, J. H., II; Laasko, J.; Ireland, C. M.; Murray, L.;
Capon, R. J.; Boyd, M. R. J. Org. Chem. 1998, 63, 7805.
(3) (a) Kunze, B.; Jansen, R.; Sasse, F.; Ho¨fle, G.; Reichenbach, H. J. Antibiot.
1998, 51, 1075. (b) Jansen, R.; Kunze, B.; Reichenbach, H.; Ho¨fle, G. Eur.
J. Org. Chem. 2000, 6, 913. For the total synthesis of apicularen A, see:
(c) Bhattacharjee, A.; Seguil, O. R.; De Brabander, J. K. Tetrahedron Lett.
2001, 42, 1217. (d) Nicolaou, K. C.; Kim, D. W.; Baati, R. Angew. Chem.,
Int. Ed. 2002, 41, 3701. (e) Lewis, A.; Stefanuti, I.; Swain, S. A.; Smith,
S. A.; Taylor, R. J. K. Tetrahedron Lett. 2001, 42, 5549.
(4) Dekker, K. A.; Aiello, R. J.; Hirai, H.; Inagaki, T.; Sakakibara, T.; Suzuki,
Y.; Thompson, J. F.; Yamaguchi, Y.; Kojima, N. J. Antibiot. 1998, 51, 14.
9
10.1021/ja0352350 CCC: $25.00 © 2003 American Chemical Society
J. AM. CHEM. SOC. 2003, 125, 7889-7901
7889

A R T I C L E S
Porco et al.
Figure 1. Salicylate enamide natural products.
on the plasma membranes of human tumor cells.9 Accordingly,
these natural products are exciting new targets for chemical
synthesis, lead optimization studies, and preparation of designed
analogues and molecular probes to further define interactions
with the molecular target. The salicylihalamides have been
synthesized in several laboratories.1c-h De Brabander and co-
workers have reported the preparation and structure-function
analysis of a number of promising salicylihalamide derivatives.10
Herein, we report a full account of the total synthesis and
stereochemical assignment of lobatamide C,11 as well as
structure-activity studies in the lobatamide series.
R-substituted amides,16 isomerization of N-allylamides,17 pal-
ladium(II)-catalyzed amidation of alkenes,18 direct addition of
amides to alkynes,19 acid-catalyzed condensation of aldehydes
and amides,20 amide Peterson olefination,21 Horner-Wittig and
Wadsworth-Emmons reactions,20b,22 and N-acylation of pro-
tected enamines.1f,23 Other recent enamide formation methods
have also been developed, including organometal addition to
vinyl isocyanates,5b,24 oxidative decarboxylation-elimination,25
Ru-catalyzed chain extension,26 and rearrangement of N-(R-
silyl)allyl amides.27 Some of these methods have been employed
in the synthesis of enamide-containing natural products such
as lycorine,28 mycalolide A,29 chondriamides,25 crocacin D,30
TMC-95 A and B,27 and the salicylihalamides.1c-f
Development of Methodology for Synthesis of the
Enamide Side Chain
In considering potential new methods for the formation of
enamides, we have focused our efforts on transition metal-
catalyzed vinylic substitution reactions of vinyl halides and
amides, due to the ready availability and stability of the reaction
partners and the possibility that such C-N bond constructions
may occur in a stereocontrolled manner (Figure 2). It was
envisioned that a coupling process could be developed wherein
E and Z vinyl halides could be converted to the corresponding
To enable the synthesis of the salicylate enamides as an entire
class, we required a general method to synthesize highly
unsaturated enamides. In general, enamides have been shown
to have ambident reactivity, both electrophilic at the R-carbon
and nucleophilic at the â-carbon. Enamides may be regarded
as deactivated enamines and will react with powerful electro-
philes such as bromine, peracids, and lead(IV) acetate.12 They
are stable compounds under neutral or basic conditions and with
Brønsted acids give rate-determining protonation on carbon,
leading to a reactive N-acyliminium ion intermediate that may
either undergo hydrolysis of the double bond to form carbonyl
compounds and amides13 or react with a range of nucleophiles,
including oxygen, sulfur, or π-based nucleophiles.14
(16) Ben-Ishai, D.; Giger, R. Tetrahedron Lett. 1965, 6, 4523.
(17) (a) Tischler, A. N.; Tischler, M. H. Tetrahedron Lett. 1978, 19, 3407. (b)
Stille, J. K.; Becker, Y. J. Org. Chem. 1980, 45, 2139.
(18) Hosokawa, T.; Takano, M.; Kuroki, Y.; Murahashi, S.-I. Tetrahedron Lett.
1992, 33, 6643.
(19) Kondo, T.; Tanaka, A.; Kotachi, S.; Watanabe, Y. J. Chem. Soc., Chem.
Commun. 1995, 413.
(20) (a) Zezza, C. A.; Smith, M. B. Synth. Commun. 1987, 17, 729. (b) Paterson,
I.; Cowden, C.; Watson, C. Synlett 1996, 209.
(21) (a) Palomo, C.; Aizpurua, J. M.; Legido, M.; Picard, J. P.; Dunogues, J.;
Constantieux, T. Tetrahedron Lett. 1992, 33, 3903. (b) Fu¨rstner, A.; Brehm,
C.; Cancho-Grande, Y. Org. Lett. 2001, 3, 3955.
(22) Couture, A.; Deniau, E.; Grandclaudon, P. Tetrahedron Lett. 1993, 34,
1479.
(23) (a) Brettle, R.; Mosedale, A. J. J. Chem. Soc., Perkin Trans. 1 1988, 2185.
(b) Shimasaki, H.; Kagechika, H.; Fukasawa, H.; Kawachi, E.; Shudo, K.
Chem. Pharm. Bull. 1995, 43, 100.
(24) (a) Ramamurthy, B.; Sugumaran, M. Synthesis 1987, 523. (b) Snider, B.
B.; Song, F. Org. Lett. 2000, 2, 407. (c) Stefanuti, I.; Smith, S. A.; Taylor,
R. J. K. Tetrahedron Lett. 2000, 41, 3735.
Enamides have been previously synthesized using a number
of methods, including N-acylation of imines,15 elimination of
(9) (a) Martinez-Zaguilan, R.; Lynch, R. M.; Martinez, G. M.; Gillies, R. J.
Am. J. Physiol. 1993, 265, C1015. (b) Martinez-Zaguilan, R.; Raghunand,
N.; Lynch, R. M.; Bellamy, W.; Martinez, G. M.; Rojas, B.; Smith, D.;
Dalton, W. S.; Gillies, R. J. Biochem. Pharmacol. 1999, 57, 1037.
(10) Wu, Y.; Liao, X.; Wang, R.; Xie, X-s.; De Brabander, J. K. J. Am. Chem.
Soc. 2002, 124, 3245.
(11) For a preliminary Communication, see: Shen, R.; Lin, C. T.; Porco, J. A.,
Jr. J. Am. Chem. Soc. 2002, 124, 5650.
(12) (a) Morzycki, J. W.; Wilczewska, A. Z. Tetrahedron, 1996, 52, 14057. (b)
Back, T. G.; Ibrahim, N.; McPhee, D. J. J. Org. Chem. 1982, 47, 3283. (c)
Lessor, R. A.; Rafalko, P. W.; Lenz, G. R. J. Chem. Soc., Perkin Trans. 1
1989, 11, 1931.
(13) Csizmadia, V. M.; Koshy, K. M.; Lau, K. C. M.; McClelland, R. A.;
Nowlan, V. J.; Tidwell, T. T. J. Am. Chem. Soc. 1979, 101, 974.
(14) Speckamp, W. N.; Hiemstra, H. Tetrahedron 1985, 41, 4367.
(15) Lenz, G. R. Synthesis, 1978, 489.
(25) Wang, X.; Porco, J. A., Jr. J. Org. Chem. 2001, 66, 8215.
(26) Trost, B. M.; Surivet, J.-P. Angew. Chem., Int. Ed. 2001, 40, 1468.
(27) Lin, S.; Danishefsky, S. J. Angew. Chem., Int. Ed. 2002, 41, 512.
(28) Boeckmann, R. K., Jr.; Goldstein, S. W.; Walters, M. A. J. Am. Chem.
Soc. 1988, 110, 8250.
(29) Liu, P.; Panek, J. S. J. Am. Chem. Soc. 2000, 122, 1235.
(30) (a) Feutrill, J. T.; Lilly, M. J.; Rizzacasa, M. A. Org. lett. 2002, 4, 525. (b)
Chakraborty, T. K.; Laxman, P. Tetrahedron Lett. 2002, 43, 2645.
9
7890 J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003

Lobatamide C
A R T I C L E S
(5),39 related transformations provided the desired enamides 6
and 7 (entries 3 and 4). The coupling reaction was also
successful for secondary amides. Amidations of 1 and 5 with
N-methylformamide provided enamides 8 and 9 as a mixture
of rotamers in good yields (entries 5 and 6). With 2-pyrrolidi-
none, enamide 1020a was obtained in 99% yield (entry 7).
Amidation of a Z-vinyl iodide 1140 provided Z-enamide 12 in
only 23% yield (entry 8) likely due to competitive elimination
of the Z-vinyl iodide to the corresponding terminal alkyne under
the reaction conditions.41
Figure 2. Stereoselective C-N bond construction for enamide synthesis.
Scheme 1 a
To investigate the nature of the halogen substituent, we
evaluated (E)-1-bromo-1-heptene (13)42 and (3Z)-(4-bromo-3-
butenyl)benzene43 in coupling reactions with benzamide. How-
ever, only trace amounts of enamide products were obtained in
these experiments. To compare the reactivities of vinyl iodides
and vinyl bromides in the C-N bond formation, we conducted
the competition experiment depicted in Scheme 2. Cu-mediated
amidation was performed with vinyl bromide 13, (E)-1-iodo-
1-pentene (14),34 and benzamide in NMP (90 °C). In this case,
a 7:1 ratio of enamides 15 and 2 was detected by HPLC44
analysis of the crude reaction mixture, which further supports
that vinyl bromides are significantly less reactive than vinyl
iodides as amidation substrates.45
a Reagents and conditions: (a) CuI (0.10 equiv), Cs2CO3 (1.2 equiv),
PPh3 (0.20 equiv), NMP, 90 °C, 12 h, 31% (1H NMR); (b) CuTC (0.10
equiv), Cs2CO3 (1.2 equiv), NMP, 90 °C, 12 h, 59% (1H NMR).
E and Z enamides. This is significant since many of the existing
approaches to prepare enamides are not stereoselective. As an
entry to our studies, we were attracted to a study by Ogawa et
al. who reported the copper iodide-promoted substitution of vinyl
bromides and potassium amides (1 equiv of CuI, HMPA, 130
°C) to afford enamides in low to moderate (38-45%) yields.31
On the basis of this precedent and related Cu(I)-catalyzed C-N
cross coupling reactions,32 we focused on developing a Cu(I)-
catalyzed amidation method that would occur at milder tem-
peratures and would be suitable for the installation of potentially
labile enamides on complex substrates.33
Using benzamide and (E)-1-iodo-1-heptene (1)34 as model
substrates, we initially compared Cu(I) phosphine35 and Cu(I)
carboxylate catalysts with Cs2CO3 as base (Scheme 1). We
obtained a higher conversion (59%) for enamide 2 using
Liebeskind’s Cu(I) thiophenecarboxylate (CuTC),36 which led
us to undertake further optimization with this catalyst.
To prepare enamides related to the lobatamides and related
salicylate natural products, we next prepared 4-(methoxyimino)-
2-butenamides 16 and 17 (Scheme 3). Treatment of 5-hydoxy-
2(5H)-furanone46 with aqueous methoxyamine hydrochloride led
to the formation of 4-(methoxyimino)-(2Z)-butenoic acid 18
(92%).47 The corresponding 4-(methoxyimino)-(2Z)-butenamide
16 was prepared in 88% yield by formation of the mixed
anhydride of 18 and subsequent reaction with aqueous ammonia.
(2Z)-Butenamide 16 could be fully isomerized to (2E)-buten-
amide 17 in 81% yield under acidic conditions.
Next, we conducted model amidation reactions with buten-
amides 16 and 17 and representative vinyl iodides (Table 2).
Treatment of (2E)-butenamide 17 with vinyl iodide 1 or 5 using
N,N-dimethylacetamide (DMA) as solvent afforded unsaturated
enamides 19 and 20 in 57% and 52% yield, respectively (entries
1 and 2). However, cross-coupling of (2Z)-butenamide 16 with
vinyl iodide 1 under similar conditions (90 °C, 12 h) did not
afford the desired enamide 21. In an effort to improve this
reaction, 2,2,6,6-tetramethyl-3,5-heptanedione48 was used as
After reaction optimization, optimal conditions for amidations
were discovered by employing CuTC (30 mol %), Cs2CO3 as
base, and rigorous vacuum purge degassing of the reaction
mixture in 1-methyl-2-pyrrolidinone (NMP) prior to heating (90
°C, 12 h). Using these conditions, a number of enamides were
prepared as shown in Table 1. Both benzamide and (E,E)-2,4-
hexadienamide (3) 37 participated in vinylic substitution of 1 to
afford enamides 2 and 4 in good yields (entries 1 and 2). It
should be mentioned that, under the same conditions using an
excess of vinyl iodide 1 (2.2 equiv) and Cs2CO3 (2.0 equiv),
the coupling reaction with benzamide afforded only enamide 2
(77%) and recovered vinyl iodide 1 (70%) without any evidence
of an N,N-divinyl amide product.38 Using (E)-â-iodostyrene
(38) N,N-Divinylureas and N,N-divinylamides have been previously prepared
by isomerization of N,N-diallylic ureas and amides, see: (a) Murai, T.;
Kasai, Y.; Ishihara, H.; Kato, S. J. Org. Chem. 1992, 57, 5542. (b)
Krompiec, S.; Suwinski, J.; Grobelny, J.; Wagner, P. Pol. J. Chem. 1997,
71, 747.
(39) Takai, K.; Ichiguchi, T.; Hikasa, S. Synlett 1999, 1268.
(40) Prepared according to: Stork, G.; Zhao, K. Tetrahedron Lett. 1989, 30,
2173.
(31) Ogawa, T.; Kiji, T.; Hayami, K.; Suzuki, H. Chem. Lett. 1991, 1443.
(32) (a) Kiyomori, A.; Marcoux, J.-F.; Doye, S.; Buchwald, S. L. Tetrahedron
Lett. 1999, 40, 2657. For recent studies on Cu(I)-catalyzed C-N bond
formation, see: (b) Wolter, M.; Klapars, A.; Buchwald, S. L. Org. Lett.
2001, 3, 3803. (c) Kwong, F. Y.; Klapars, A.; Buchwald, S. L. Org. Lett.
2002, 4, 581. (d) Klapars, A.; Huang, X.; Buchwald, S. L. J. Am. Chem.
Soc. 2002, 124, 7421. (e) Antilla, J. C.; Klapars, A.; Buchwald, S. L. J.
Am. Chem. Soc. 2002, 124, 11684. (f) Job, G. E.; Buchwald, S. L. Org.
Lett. 2002, 4, 3703. (g) Kwong, F. Y.; Buchwald, S. L. Org. Lett. 2003, 5,
793.
(41) In a control experiment, 4-phenyl-1-butyne was formed when 11 (1.0 equiv)-
was heated with Cs2CO3 (1.0 equiv) in NMP (60 °C, 14 h).
(42) Brown, H. C.; Larock, R. C.; Gupta, S. K.; Rajagopalan, S.; Bhat, N. G. J.
Org. Chem. 1989, 54, 6079.
(43) Uenishi, J.; Kawahama, R.; Shiga, Y.; Yonemitsu, O.; Tsuji, J. Tetrahedron
Lett. 1996, 37, 6759.
(44) HPLC analysis was performed using a Waters 2700/600 HPLC system:
Symmetry C18 column (19 × 50 mm), λ ) 269 nm; 40-100% CH3CN/
H2O, 10 min; tR(15) ) 5.06 min, tR(2) ) 6.82 min.
(45) For recent reports concerning the relative reactivity of bromo and iodo
substrates in Cu(I)-catalyzed couplings, see: (a) Klapars, A.; Buchwald,
S. L. J. Am. Chem. Soc. 2002, 124, 14844. (b) Zanon, J.; Klapars, A.;
Buchwald, S. L. J. Am. Chem. Soc. 2003, 125, 2890.
(46) Doerr, I. L.; Willette, R. E. J. Org. Chem. 1973, 38, 3878.
(47) Schroeter, S. H.; Appel, R.; Brammer, R.; Schenck, G. O. Justus Liebigs
Ann. Chem. 1966, 697, 42.
(48) Buck, E.; Song, Z. J.; Tschaen, D.; Dormer, P. G.; Volante, R. P.; Reider,
P. J. Org. Lett. 2002, 4, 1623.
(33) For a preliminary account of Cu(I)-catalyzed enamide formation, see: Shen,
R.; Porco, J. A., Jr. Org. Lett. 2000, 2, 1333.
(34) Zweifel, G.; Whitney, C. C. J. Am. Chem. Soc. 1967, 89, 2753.
(35) Okuro, K.; Furuune, M.; Enna, M.; Miura, M.; Nomura, M. J. Org. Chem.
1993, 58, 4716.
(36) (a) Allred, G. D.; Liebeskind, L. S. J. Am. Chem. Soc. 1996, 118, 2748.
(b) Zhang, S.; Zhang, D.; Liebeskind, L. S. J. Org. Chem. 1997, 62, 2312.
(37) Pellegata, R.; Italia, A.; Villa, M. Synthesis 1985, 517.
9
J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003 7891

A R T I C L E S
Porco et al.
Table 1. CuTC-Catalyzed Enamide Coupling of Amides and Vinyl Iodidesa
a
b
Performed using 30 mol % of CuTC, 1.0 equiv of vinyl halide, 1.5 equiv of amide, and 1.5 equiv of Cs2CO3, NMP, 90 °C, 12 h. Isolated yields after
c
d
e
silica gel chromatography. 3:1 mixture of rotamers by NMR (400 MHz). 1.3:1 mixture of rotamers by NMR (400 MHz). Reaction performed at 60
°C.
Scheme 2. Vinyl Bromide vs Iodide Competition Experiment
Scheme 3
ligand to increase the reactivity and turnover of CuTC (Scheme
4). However, the reaction afforded only 20% of enamide 21
along with the unexpected N,N-divinyl amide 23 (17%).38
Finally, enamide 21 was obtained in 52% yield using Rb2CO3
as base49 and N,N′-dimethylethylenediamine (24) as ligand32d,e
(entry 3). Entry 4 illustrates the stereoselective coupling of
Z-amide 16 and Z-vinyl iodide 11. The reaction provided a
modest yield (30%) of the desired Z-enamide 22 when only
CuTC was used as catalyst. However, when diamine ligand 24
was added, the yield of enamide 22 was enhanced to 55%. This
initial study provided encouragement for the stereospecific
construction of the Z-enamide side chain of oximidines (cf.
Figure 1).
Since diamine ligand 24 remarkably facilitated the production
of enamides 21 and 22, we employed 24 as ligand in couplings
to prepare enamides 19 and 2. However, the yield of 19 (54%)
was slightly lower than that solely using CuTC as catalyst
(57%), while the yield of 2 was only 40%.50 These results
(49) For use of Rb2CO3 as a base in Pd-catalyzed C-N bond formation, see:
Watanabe, M.; Nishiyama, M.; Yamamoto, T.; Koie, Y. Tetrahedron Lett.
2000, 41, 481.
9
7892 J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003

Lobatamide C
A R T I C L E S
Table 2. Cu(I)-Catalyzed Vinylic Substitution of Butenamides 16
and 17
Scheme 5
amidation to construct enamides may be difficult on fragile and
potentially elimination prone macrodilactone substrates. As we
were later to discover, this convergent approach creates synthetic
challenges with regard to manipulation and reactivity of
enamide-containing intermediates but also adds an element of
flexibility for construction of analogues that vary at the salicylate
portion.
a
Isolated yields after silica gel chromatography. b Reaction performed
with 0.3 equiv of CuTC, 1.5 equiv of 17, 1.5 equiv of Cs2CO3, DMA, 90
°C, 12 h. 0.5 equiv of CuTC, 0.5 equiv of 24, 1.5 equiv of 16, 1.1 equiv
of Rb2CO3, DMA, 60 °C, 12 h. 0.3 equiv of CuTC, 0.6 equiv of 24, 1.5
equiv of 16, 1.1 equiv of Rb2CO3, DMA, 60 °C, 14.5 h.
c
d
Scheme 4
Retrosynthetic analysis of the lobatamide C skeleton reveals
two principal fragments: the C1-C10 enamide sector 28 and
the C11-C26 salicylate subunit 29 containing a divinylcarbinol
moiety and an acetonide protecting group. We planned to
prepare the precursor 30 from esterification of subunits 28 and
29, which could be desilylated and macrocyclized by removal
of the 1,3-dioxin-4-one to furnish the salicylate ester under
basic51 or potentially photochemical conditions.52 Although the
configuration of lobatamide C at C8 and C11 was not deter-
mined,2 we first focused on preparation of the 8S enantiomer
of 30 in light of the stereochemistry of salicylihalamides1c and
indicate that ligand 24 is most effective when used with (2Z)-
butenamide 16. It is conceivable that enamide 21 may chelate
with CuTC to form complex 25 (Scheme 5), which may
substantially deactivate the catalyst. Addition of diamine 24 may
compete with enamide 21 to transform a complex such as 25
to 26, which may be available for further catalysis.
First Retrosynthetic Analysis of Lobatamide C
After the establishment of a workable methodology for the
synthesis of the enamide side chain, we initiated studies toward
the total synthesis of the lobatamides. Our plan for the synthesis
of our first target, lobatamide C (27), is outlined in Figure 3. In
contrast to previous syntheses of the salicylate enamide natural
products1c-g,3c,d in which the enamide side chain was typically
installed at a late stage in the synthesis, our general plan for
lobatamide C was to pursue a convergent coupling of fragments
with the enamide side chain preinstalled. This synthetic plan
was primarily based on our general concern that base-catalyzed
(50) Conditions for preparation of 19: 30 mol % of CuTC, 60 mol % of 24,
1.0 equiv of vinyl iodide 1, 1.5 equiv of amide 17, 1.1 equiv of Cs2CO3,
DMA, 60 °C, 12 h. For preparation of 2: 30 mol % of CuTC, 60 mol %
of 24, 1.0 equiv of vinyl iodide 1, 1.5 equiv of benzamide, 1.5 equiv of
Cs2CO3, NMP, 90 °C, 12 h.
Figure 3. First retrosynthetic analysis for lobatamide C.
9
J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003 7893

A R T I C L E S
Porco et al.
Scheme 6
Scheme 7
oximidines5 (cf. Figure 1). The stereochemistry at C8 could
easily be altered in the event that this prediction was found
to be incorrect. Further disconnection of fragment 29 at the
C18-C19 and C14-C15 bonds led to benzyl bromide 31,
Z-vinyl stannane 32,53 and nonracemic alkyne 33. The divinyl-
carbinol moiety may be obtained by addition of a vinylzinc
species (derived from 33 by hydrozirconation and transmeta-
lation),54 to an enal which will be prepared by Stille coupling55
of 31 and 32, desilylation, and alcohol oxidation. It was
anticipated that stereocontrol at C15 could be established using
chiral ligands in the vinylzinc addition step.56 Enamide subunit
28 may be prepared by Cu(I)-catalyzed amidation of vinyl iodide
34 with amide 17 according to the C-N bond formation
protocols previously described.
establish the C8 stereocenter and efficiently prepare multigram
amounts of (R)-35 (>99% ee). (R)-35 was next employed in
epoxide ring-opening reactions in order to prepare a vinyl iodide
substrate for enamide synthesis. Addition of the acetylenic alane
reagent59 derived from trimethylsilylacetylene led to facile
epoxide opening and afforded 36 in 80% yield. Silyl protection
of 36 using chlorodiethylisopropylsilane (DEIPSCl), followed
by treatment of the resulting silyl ether 37 with AgNO3/NBS/
H2O,60 afforded the bromoalkyne intermediate 38 that was
converted to the (E)-stannylalkene 39 using Pattenden’s method61
(78%, three steps). Iodine exchange of 39 furnished vinyl iodide
34 in 73% yield with full retention of olefin stereochemistry.
In contrast, iodination of the desilylated analogue of 39 afforded
a 5:1 E:Z mixture of vinyl iodides, indicating the importance
of protecting the secondary alcohol. After considerable experi-
mentation, we found that Cu(I)-mediated vinylic substitution
of 34 with the butenamide 17 (CuTC, 1,10-phenanthroline,32a
dba, Cs2CO3, DMA) led to a 52% yield of the C1-C10 enamide
fragment 40. Critical to the success and reproducibility of this
reaction were the use of a moderate reaction temperature (65
Synthesis of C1-C10 Subunit (28)
Preparation of a protected form of the C1-C10 enamide
fragment 28 is shown in Scheme 6. Ethyl vinyl acetate was
transformed in quantitative yield into (()-ethyl-3,4-epoxybu-
tanoate 35 using m-CPBA.57 Hydrolytic kinetic resolution
(HKR) developed by Jacobsen and co-workers58 was used to
(58) Schaus, S. E.; Brandes, B. D.; Larrow, J. F.; Tokunaga, M.; Hansen, K.
B.; Gould, A. E.; Furrow, M. E.; Jacobsen, E. N. J. Am. Chem. Soc. 2002,
124, 1307.
(59) (a) Fried, J.; Sih, J. C.; Lin, C. H.; Dalven, P. J. Am. Chem. Soc. 1972, 94,
4343. (b) Larcheveque, M.; Petit, Y. Bull. Chim. Soc. Fr. 1989, 130.
(60) Tobe, Y.; Nakanishi, H.; Sonoda, M.; Wakabayashi, T. Achiba, Y. J. Chem.
Soc., Chem. Commun. 1999, 17, 1625. Optimization studies indicate that
inclusion of water (2.0 equiv) is important for high yield and reproducibility
for the bromination reaction.
(51) Bhattacharjee, A.; De Brabander, J. K. Tetrahedron Lett. 2000, 41, 8069.
(52) (a) Chapman, O. L.; McIntosh, C. L. J. Am. Chem. Soc. 1970, 92, 7001.
(b) Liu, R. C.-Y.; Lusztyk, J.; McAllister, M. A.; Tidwell, T. T.; Wagner,
B. D. J. Am. Chem. Soc. 1998, 120, 6247.
(53) Han, Q.; Wiemer, D. F. J. Am. Chem. Soc. 1992, 114, 7697.
(54) Wipf, P.; Ribe, S. J. Org. Chem. 1998, 63, 6454.
(55) Farina, V.; Krishnamurthy, V.; Scott, W. J. Org. React. 1997, 50, 1. For a
similar Stille coupling in synthesis of the salicylihalamides, see ref 1h.
(56) For a review on catalytic asymmetric organozinc addition to carbonyls,
see: Pu, L.; Yu, H.-B. Chem. ReV. 2001, 101, 757.
(61) (a) Boden, C. D. J.; Pattenden, G.; Ye, T. J. Chem. Soc., Perkin Trans. 1
1996, 2417. (b) Claus, E.; Kalesse, M. Tetrahedron Lett. 1999, 40,
4157.
(57) Mohr, P.; Roesslein, L.; Tamm, C. Tetrahedron Lett. 1989, 30, 2513.
9
7894 J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003

Lobatamide C
A R T I C L E S
Scheme 8
Scheme 9
Scheme 11
Scheme 12
Synthesis of the C11-26 Fragment (29)
Synthesis of the C11-C26 salicylate fragment 29 commenced
with preparation of salicylate benzyl bromide 31, prepared in
two steps from the known aryl triflate 4163 (Scheme 7).
Treatment of 41 with lithium dimethylcuprate64 and MeI65 led
to the production of 6-methylsalicylate derivative 42 that was
brominated66 to afford benzylic bromide 31 (78%). Coupling
of 31 with vinylstannane 3253 using the conditions reported by
Kamlage67 (Pd2dba3-CHCl3, AsPh3, THF) afforded Z-allylic
silyl ether 43 (79%). Silyl deprotection using HF/pyridine
provided allylic alcohol 44, which was oxidized to Z-enal 45
using Dess-Martin periodinane.68 However, 45 was found to
be readily isomerized to E-enal 46 during purification on silica
gel (46:45 ) 6:1). Hence, crude 45 was used in subsequent
coupling reactions. Hydrozirconation of DEIPS-protected (R)-
3-butyn-2-ol (33), followed by transmetalation with Et2Zn and
addition to 45 according to the general procedure of Wipf and
Scheme 10
°C) to suppress elimination of the sensitive â-silyloxy ester
coupling substrate, the use of 1,10-phenanthroline as sup-
porting ligand for CuTC, and use of high purity cesium
carbonate.62 Synthesis of the target C1-C10 fragment was
completed by hydrolysis of 40 to afford the labile enamide acid
28 (73%).
(63) Fu¨rstner, A.; Konetzki, I. Tetrahedron 1996, 52, 15071.
(64) McMurry, J. E.; Mohanraj, S. Tetrahedron Lett. 1983, 24, 2723.
(65) Quenching the reaction with MeI was found to significantly improve the
reaction yield, likely due to methylation of an aryl copper side product.
Cf. Cohen, T.; Wood, J.; Dietz, A. G., Jr. Tetrahedron Lett. 1974, 15, 3555.
(66) Eicher, T.; Tiefensee, K.; Pick, R. Synthesis 1988, 525.
(67) (a) Kamlage, S.; Sefkow, M.; Peter, M. G. J. Org. Chem. 1999, 64, 2938.
(b) Farina, V.; Krishnan, B. J. Am. Chem. Soc. 1991, 113, 9585.
(68) Dess, D. B.; Martin, J. C. J. Org. Chem. 1983, 48, 4155.
(62) Yin, J.; Buchwald, S. L. Org. Lett. 2000, 2, 1101.
9
J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003 7895

A R T I C L E S
Porco et al.
Figure 4. Proposed transesterification using intramolecular general base
catalysis.
Scheme 13
Figure 5. Revised retrosynthesis of lobatamide C
Scheme 14
co-workers,54 led to the target divinylcarbinol fragment 47 as a
1:1 mixture of diastereomers (30% yield). Although the Z-olefin
stereochemistry of 47 was verified using NOE experiments, the
presence of significant amounts of isomerized E-enal 46 led to
rapid alteration of the synthetic sequence and use of configu-
rationally stable Z-enal 4869(Scheme 8). In the event, hydrozir-
conation, zirconocene-zinc transmetalation and addition to enal
48 afforded divinylcarbinol 49 as a 1:1 nonseparable mixture
of diastereomers (87%). Compound 49 was further advanced
by silylation of the secondary alcohol, followed by lithiation-
trimethylstannylation, to afford vinyl stannane 51 which was
coupled with benzyl bromide 31 to furnish C11-C26 fragment
52 (56%). Selective deprotection of 52 using HF/pyridine at 0
°C afforded the target alcohol 29 (69%).
Scheme 15
Fragment Coupling and Attempted Macrocyclization
With fragments 28 and 29 in hand, we attempted their
coupling using standard acylation methods. However, both
Yamaguchi70 and Keck71 esterification conditions did not furnish
the desired ester C11-epi-30, presumably due to decomposition
of the sensitive enamide side chain. However, the esterification
was successfully accomplished using Mitsunobu conditions to
afford 30 (70%, Scheme 9). Selective deprotection of the C8
DEIPS ether of 30 using HF/pyridine afforded acyclic alcohol
53 (41%). Extensive studies were performed to prepare mac-
rolactone 54 by intramolecular transesterification of 53. Treat-
ment of 53 under basic conditions (NaH51 or NaHMDS, THF)
or distannoxane transesterification catalysts72 did not furnish
the desired cyclized product 54. Under these conditions,
compound 53 was substantially decomposed. Attempted pho-
tolysis of 53 (Ace-Hanovia UV lamp 450 W, Pyrex filter, 20
min) in an effort to form the macrolactone by trapping of a
keto-ketene intermediate52 led to complete decomposition of
the substrate. Due to a concern that the C15 TBS ether may
block macrocyclization, we next prepared compound 55 by
removing the TBS ether with HF/pyridine. However, treatment
of 55 under basic conditions (NaH, THF) led to the production
of the eight-membered lactone enamide 56 (dr 1:1) instead of
the desired natural product. Compound 56 is formally an isomer
of lobatamide C in which the divinylcarbinol forms a salicylate
ester, leaving a pendant C8 hydroxyl. Attempts to rearrange 56
to 15-membered ring lobatamide C (27) by translactonization
(69) Larock, R. C.; Doty, M. J.; Han, X. J. Org. Chem. 1999, 64, 8770.
(70) Inanaga, J.; Hirata, K.; Saeki, H.; Katsuki, T.; Yamaguchi, M. Bull. Chem.
Soc. Jpn. 1979, 52, 1989.
(71) Boden, E. P.; Keck, G. E. J. Org. Chem. 1985, 50, 2394.
(72) (a) Otera, J. Chem. ReV. 1993, 93, 1449. (b) Shimizu, I.; Nakagawa, H.
Tetrahedron Lett. 1992, 33, 4957.
9
7896 J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003

Lobatamide C
A R T I C L E S
Scheme 16
Scheme 17
dimethylcyanamide, ethyl vinyl ether, ethoxyacetylene, and
N-benzylmaleimide failed to provide the corresponding Diels-
Alder adducts.77 Interestingly, treatment of 60 and methoxy
cyanomethyl ester 64 under the same conditions afforded the
ester product 65, albeit in lower chemical yield (28%) and
limited conversion (Scheme 11). These results, in conjunction
with conformational analysis of the sodium salt of 61,78
suggested an alternative mechanistic pathway involving in-
tramolecular general base catalysis. In the conformer shown in
Figure 4, the ester carbonyl is out of planarity, which is expected
to increase the reactivity of the carbonyl toward transesterifi-
cation. In addition, the o-hydroxyl may be oriented to act as a
general base catalyst and direct attack of the alcohol to the π*
of the salicylate carbonyl. This proposed transesterification
mechanism is substantiated by literature precedent.79
protocols73 (e.g., catalytic PPTS/CH2Cl2 or Et2Zn/ CH2Cl2) were
not successful.
Development of Methodology for the Construction of
Salicylate Esters
To ultimately apply this transesterification methodology to
the construction of the salicylate portion of lobatamide C, we
next prepared analogues 66-68. In particular, compounds 66
and 68 contain o-salicylate alkyl substituents. Hydrolysis of
benzodioxinone 42 with KOH quantitively provided 6-methyl-
salicylic acid80 (Scheme 12), which was converted to cyano-
methyl ester 69 (80%).81 Acylation of enamide alcohol 60 with
69 in the presence of K2CO3 in DMA afforded analogue 66
(70%). Analogue 67 was prepared from secondary alcohol 70
(prepared by TBAF desilylation of 40), which exhibited
significantly lower reactivity in acylation with cyanomethyl ester
61 (25% yield, Scheme 13). In addition, 15% of the â-elimina-
tion product, as well as considerable amounts of recovered 70,
were obtained. Under similar conditions, acylation of 70 with
cyanomethyl ester 69 did not afford the desired salicylate
product. However, the derived acid 71, after neutralization with
Bu4NOH and heating with 69 and Na2CO3 (1.0 equiv) in DMA/
2-butanone (80 °C, 2 h), furnished the desired salicylate ester.
The tetrabutylammonium salt of 71 both increases the solubility
of the enamide alcohol fragment82 and likely blocks R-depro-
tonation/elimination of the â-salicyloxyester product. A number
In parallel with our total synthesis efforts, we initiated studies
toward the synthesis and biological evaluation of simplified
analogues of the lobatamides in order to clarify the minimal
core structure (pharmacophore) required for V-ATPase inhibi-
tion. In initial studies, a series of acyclic analogues were
prepared using the C1-C10 enamide fragment. Our first target
was simplified salicylate enamide 57 that eliminates the C8
stereogenic center (Scheme 10). TIPS-protected vinyl iodide 5874
underwent CuTC-mediated cross-coupling with (2E)-butenamide
17 to furnish enamide 59, which was desilylated using TBAF
to afford enamide alcohol 60. Encouraged by patent literature
for esterification of salicylate cyanomethyl esters75 and general
stability of enamides to basic conditions, we first treated enamide
alcohol 60 with cyanomethyl ester 6175 in the presence of a
catalytic amount of K2CO3 (DMA, 90 °C). Salicylate enamide
57 was rapidly and cleanly produced under these conditions
(91%).
As a control experiment, we found that the acylation using
benzoyloxyacetonitrile 62 with alcohol 60 under similar reaction
conditions did not afford the desired ester product (Scheme 11).
We initially suspected that keto-ketene 63 was the active
acylating agent.76 However, trapping experiments with N,N-
(77) (a) Kaneko, C.; Sato, M.; Sakaki, J-i.; Abe, Y. J. Heterocycl. Chem. 1990,
27, 25. (b) Stadler, A.; Zangger, K.; Belaj, F.; Kollenz, G. Tetrahedron
2001, 57, 6757.
(78) A conformational search was performed using PC Spartan Pro version 1.0.7
(Wavefunction, Irvine, CA).
(73) (a) For acid-catalyzed translactonization to prepare macrocyclic lactones,
see: Corey, E. J.; Brunelle, D. J.; Nicolaou, K. C. J. Am. Chem. Soc. 1977,
99, 7359. (b) For an example of base-catalyzed translactonization, see:
Duret, P.; Figadere, B.; Hocquemiller, R.; Cave, A. Tetrahedron Lett. 1997,
38, 8849.
(79) (a) Bender, M. L.; Kezdy, F. J.; Zerner, B. J. Am. Chem. Soc. 1963, 85,
3017. (b) Khan, M. N. Int. J. Chem. Kinet. 1988, 20, 443.
(80) Bray, L. G.; Dippy, J. F. J.; Hughes, S. R. C.; Laxton, L. W. J. Chem. Soc.
1957, 2405.
(81) For preparation of cyanomethyl esters, see: (a) Byers, J. H.; Baran, R. C.;
Craig, M. E.; Jackman, J. T. Org. Prep. Proced. Int. 1991, 23, 373. (b)
Hugel, H. M.; Bhaskar, V.; Longmore, R. W. Synth. Commun. 1992, 22,
693.
(74) Posner, G. H.; Weitzberg, M.; Hamill, T. G.; Asirvatham, E. Tetrahedron
1986, 42, 2919.
(75) Sherlock, M. H. S. African Pat. ZA 6802187, 1968; Chem. Abstr. 1969,
70, 106224.
(82) For use of tetrabutylammonium salts to improve solubility of protected
amino acids in solid-phase peptide synthesis, see: Nagase, T.; Kumagai,
U.; Niiyama, K.; Mase, T.; Ishikawa, K. Tetrahedron Lett. 1993, 34, 1173.
(76) For a recent review on keto-ketenes, see: Simion, C.; Costea, I.; Badea,
F.; Iordache, F. Roum. Chem. Q. ReV. 2001, 8, 131.
9
J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003 7897

A R T I C L E S
Porco et al.
Scheme 18
Scheme 19
of carbonate bases (Li, K, Rb, and Cs) were also evaluated, but
interestingly only stoichiometric levels of Na2CO3 were effec-
tive. To facilitate purification, the unstable enamide acid product
was methylated with trimethylsilyl diazomethane to afford
methyl ester 68 (34%, three steps).
Revised Retrosynthesis of Lobatamide C
Due to the difficulties in forming the C8 salicylate ester using
macrocyclization (cf. Scheme 9), we altered our synthetic route
by constructing this bond at an earlier stage using the cyano-
methyl ester transesterification methodology successfully em-
ployed in our simplified analogue synthesis. Figure 5 illustrates
our revised retrosynthetic analysis of lobatamide C (27) by
macrolactonization of hydroxy acid 72 to form the C10-C11
ester bond and deprotection of the silyl group at the C15
divinylcarbinol. Acid 72 may be prepared by base-catalyzed
transesterification of cyanomethyl ester fragment 73 and hydroxy
enamide acid 71. Further, disconnection of fragment 73 at
C18-C19 bond reveals benzylic bromide and vinylstannane
fragments which may be merged by sp2-sp3 coupling (cf.
Scheme 8).
Completion of the Total Synthesis of Lobatamide C
Synthesis of fragment 73 requires diastereomerically pure
divinylcarbinol 49. Compound 49 (dr 1:1) was prepared from
alkyne 33 (Scheme 8). To establish the proposed S configuration
of the divinylcarbinol at C15, extensive studies were performed,
including evaluation of numerous amino alcohol controllers.83
However, the best result (2:1, 15S:15R) was obtained using
Wipf’s amino thiol ligand 7454 (Scheme 14). Compound 49a
(2:1 dr) was next advanced to vinyl stannane 51a. Silyl
protection of cyanomethyl ester 69, followed by benzylic
bromination, afforded benzylic bromide 76 (Scheme 15). Stille
coupling of 76 and vinyl stannane 51a afforded C11-C26
fragment 77. Selective desilylation of 77 (TBAF, 0 °C) afforded
the target salicylate cyanomethyl ester fragment 73.
After preparation of fragments 71 and 73, we found that the
tetrabutylammonium salt of enamide acid 71 participated in
smooth esterification reactions with cyanomethyl ester 73
(Na2CO3, DMF/2-butanone, 80 °C, 2 h) to provide the desired
salicylate 78 (Scheme 16). Treatment of 78 with HF/pyridine
(rt, 1 h) afforded seco acid 72 (43%, two steps). Both 78 and
72 are highly labile compounds and could only be purified using
reverse phase (C18) silica. This instability is likely due to the
pendant carboxylic acid that may protonate and decompose the
enamide functionality.13 Interestingly, when acid 78 was ex-
posed to HF/pyridine for extended reaction times (4 h), the
eight-membered ring lactone 79 was obtained along with de-
sired compound 72 in modest yield (Scheme 17). To further
confirm the structure, compound 79 was coupled with enamide
acid 28 under Mitsunobu conditions and desilylated to 56 (50%,
2 steps), which was obtained in our earlier synthetic route (cf.
Scheme 9).
Gratifyingly, seco acid 72 was smoothly macrolactonized
using intramolecular Mitsunobu conditions84 to afford separable
macrolactones 54a (26%) and 54b (26%) (Scheme 18). How-
ever, the formation of 54a and 54b in a 1:1 ratio indicates
influence of the protected divinylcarbinol stereocenter on the
macrocyclization and thus necessitated independent confir-
mation of the C15 stereochemistry. Desilylation of 54a and 54b
with HF-pyridine/pyridine led to efficient production of
lobatamide C (27a) (52%) and its C15 epimer 27b (78%).
Synthetic 27a was confirmed to be identical to data reported
for natural lobatamide C by 1H and 13C NMR, [R]D (-18.8°, c
(83) In addition to chiral ligand-controlled vinyl zinc addition to aldehydes, other
methods were also evaluated to establish the C15 (S) configuration,
including lipase resolution, asymmetric reduction of the corresponding
divinyl ketone, and zinc triflate-mediated asymmetric alkynylation of enal
48 (cf. Frantz, D. E.; Fa¨ssler, R.; Carreira, E. M. J. Am. Chem. Soc. 2000,
122, 1806). However, none of these methods afforded useful levels of
selectivity.
(84) Li, K. W.; Wu, J.; Xing, W.; Simon, J. A. J. Am. Chem. Soc. 1996, 118,
7237.
9
7898 J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003

Lobatamide C
A R T I C L E S
Figure 6. X-ray Crystal Structure of 84.
0.17, MeOH), TLC Rf values in three solvent systems, and
COMPARE profiles in NCI’s 60 tumor cell line.85
Stereochemical Assignment of Lobatamide C (27a)
The absolute configuration of 27a at C15 was determined to
be S using a modified Mosher’s ester analysis according to
Suzumura’s procedure.86 In addition, 11R diastereomers of
lobatamide C (27c and 27d) were prepared independently from
(S)-3-butyn-2-ol (Scheme 19). TIPS-protected (S)-3-butyn-2-
ol 80 was converted to divinylcarbinol 81 (dr ) 1:1) using
hydrozirconation-transmetalation-enal addition. Fortunately,
diol 82 (dr ) 1:1, obtained by TBAF desilylation of 81) was
resolved by recrystallization in CHCl3. Using this method,
diastereomers 82a and 82b were obtained in high diastereomeric
purity (>95% de).87 Selective protection of diols 82a and 82b
with DEIPSCl furnished divinylcarbinols 83a and 83b. Lobata-
mide C diastereomers 27c and 27d were synthesized from 83a
and 83b, respectively, employing the same route. Both HPLC
and NMR studies indicated that 27c and 27d did not match
natural lobatamide C.
Although numerous allylic alcohols have been reported to
undergo inversion in Mitsunobu reactions,88 recent work has
documented examples of retention of stereochemistry in Mit-
sunobu esterifications.89 To fully confirm the stereochemical
assignment of lobatamide C, compound 54a was derivatized to
afford bromophenyl acetate 84 (Figure 6). Single X-ray crystal
structure of 84 indicates that the stereochemistry at C11 is S,
clearly showing that the C11 stereogenic center was inverted
in the Mitsunobu macrocyclization. The crystal structure of 84
thus fully confirms the stereochemistry of lobatamide C as 8S,
11S and 15S without ambiguity.
Figure 7.
amide and a macrocyclic analogue 86, which replaces the fragile
divinylcarbinol moiety of lobatamide C with a Z-olefin and a
simplified three-carbon segment (Figure 7).90 The preparation
of 86 is shown in Scheme 20. Stille coupling of vinyl stannane
8791 and benzylic bromide 76 afforded salicylate 88. Selective
desilylation (TBAF) provided cyanomethyl ester 89, which was
coupled with enamide acid 71 to furnish the salicylate 90.
Desilylation (HF-pyridine/pyridine) provided hydroxy acid 91
which underwent Mitsunobu macrolactonization (0.005 M) to
afford 14-membered macrolactone analogue 86 (65%).
Synthetic lobatamide C (27a), stereochemical isomers
(27b-d), and lobatamide analogues were evaluated for activity
against bovine V-ATPase from chromaffin granule membranes.
As shown in Table 3, both lobatamide C and its C15 epimer
27b showed potent inhibition (2.1 and 3.6 nM), while the 11R
isomers of lobatamide C (27c,d) showed lower activity (20 and
21 nM). The eight-membered ring lobatamide C isomer 56 had
significantly reduced activity. For simplified lobatamide ana-
logues, compounds 57 were found to inhibit bovine V-ATPase
with modest activity. Methylated analogues 65 and 85 had
significantly reduced activities against V-ATPase, which indi-
cates the importance of a free phenol and enamide NH.
Macrodilactone 86 showed good V-ATPase inhibition (1.4 µm),
but not at the nanomolar potency of the lobatamides, which
indicates that the ring size and substitution of the macrolactone
are critical for potent V-ATPase inhibition. Interestingly, per-
mutation of the ortho hydrogen of analogue 57 to a methyl group
substantially increased the activity (66, 1.6 µM). 6-Methyl
salicylate compound 68 also showed potent inhibition of
V-ATPase (0.1 µM), approximately 180 times more active than
compound 67 without a methyl group ortho to the carbonyl.
Conformational analysis78 of 67 and 68 (Figure 8) indicates that
conformers of ortho-H substituted salicylate 67 maintain near
planarity of the carbonyl with the aromatic ring. In contrast, in
analogue 68 the o-methyl substituent forces the carbonyl out-
of-plane by approximately 60°, presumably because of steric
Synthesis and Biological Evaluation of Simplified
Lobatamide Analogues
Although several simplified analogues (57, 65-68) were
prepared during the fragment coupling studies, we synthesized
an additional lobatamide derivative 85 bearing an N-methyl en-
(85) Synthetic 27a and 27b were evaluated against NCI’s 60 tumor cell line.
Mean panel GI50’s: 3.7 nM (natural lobatamide C), 5.1 nM (27a), and
227 nM (27b); GI50 (COMPARE correlation): 1.00 (natural lobatamide
C), 0.83 (27a), 0.80 (27b).
(86) See Supporting Information. We thank Dr. K. Suzumura (Yamanouchi
Pharmaceutical Co.) for providing experimental details on the Mosher ester
analysis of YM75518A.
(87) The absolute configurations of diols 82a and 82b were assigned by HPLC
analysis and comparison of optical rotations of diols obtained from
compound 49. See Supporting Information.
(88) Hughes, D. L. Org. React. 1992, 42, 335.
(89) (a) Ahn, C.; Correia, R.; DeShong, P. J. Org. Chem. 2002, 67, 1751. (b)
Smith, A. B., III.; Safonov, I. G.; Corbett, R. M. J. Am. Chem. Soc., 2002,
124, 11102. (c) Liao, X.; Wu, Y.; De Brabander, J. K. Angew. Chem., Int.
Ed. 2003, 42, 1648.
(90) For a preliminary account, see: Shen, R.; Lin, C. T.; Bowman, E. J.;
Bowman, B. J.; Porco, J. A., Jr. Org. Lett. 2002, 4, 3103.
9
J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003 7899

A R T I C L E S
Porco et al.
Scheme 20
Table 3. Effect of Simplified Analogues of the Lobatamides
Scheme 21
against Bovine V-ATPasea
compd
IC50 (µM)
compd
IC50 (µM)
27a
27b
27c
27d
56
57
65
0.0021
0.0036
0.020
0.021
b
66
67
68
85
86
92
93
1.6
18
0.10
>200
1.4
0.21
0.06
c
no effect
a ATPase activity determined as described in the Supporting Information
using 20 µg of membrane protein and the indicated amount of inhibitor.
c
b 27% inhibition at 30 µm, higher concentrations not soluble. 25%
inhibition at 20 µm, higher concentrations not soluble.
Scheme 22
Figure 8. Representative minimum energy conformations of 67 (A) and
68 (B) (Chem 3D, enamide side chain omitted for clarity).
hindrance effects.92 This steric effect is also seen in the X-ray
crystal structure of apicularen A3b and compound 84 (salicylate
ester carbonyl out of planarity by 80° and 52°, respectively)
and may be an important conformation of the salicylate moiety
in potent V-ATPase inhibitors.
On the basis of this observation, compound 92 and 93 with
sterically demanding substituents (iodine and prenyl) ortho to
the carbonyl were prepared in order to potentially increase
V-ATPase inhibition potency. Na2CO3-mediated coupling of
enamide acid 71 and iodo cyanomethyl ester 94 (prepared from
6-iodosalicylic acid93) afforded analogue 92 after methylation
(Scheme 21). Preparation of prenylated analogue 93 commenced
with triflate 41 (Scheme 22). Stille coupling of 41 and prenyl
stannane 95 afforded acetonide 96, which was hydrolyzed and
reprotected as cyanomethyl ester 98. Analogue 93 was obtained
using similar transesterification conditions. Evaluation of 92 and
93 against bovine V-ATPase showed that 92 has lower inhibition
activity (210 nM) than analogue 68, while 93 showed slightly
higher activity (60 nM). These results indicate that it is pos-
sible to produce relatively potent, acyclic salicylate enamide
V-ATPase inhibitors. However, in addition to steric hindrance
effects, specific orientation of the functional groups by the
macrolactone scaffold may be necessary to achieve low nano-
molar inhibition against mammalian V-ATPases.
Conclusion
We have achieved a highly convergent synthesis of the potent
antitumor and V-ATPase inhibitory natural product lobatamide
C. For construction of the enamide side chain of lobatamides
and related natural products, we have developed a mild and
stereoselective Cu(I)-catalyzed vinylic substitution protocol
which complements related C-N bond formation methodolo-
(91) Lipshutz, B. H.; Keil, R.; Barton, J. C. Tetrahedron Lett. 1992, 33, 5861.
(92) Cf. (a) Pinkus, A. G.; Lin, Ellen Y. J. Mol. Struct. 1975, 24, 9. (b) Decouzon,
M.; Ertl, P.; Exner, O.; Gal, J. F.; Maria, P. C. J. Am. Chem. Soc. 1993,
115, 12071.
(93) Kobayashi, S.; Tagawa, S.; Nakajima, S. Chem. Pharm. Bull. 1963, 11,
123.
9
7900 J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003

Lobatamide C
A R T I C L E S
gies.32 A key step in the total synthesis involves base-mediated
esterification of hydroxy enamide and salicylate cyanomethyl
fragments. Macrocyclization was achieved using an intramo-
lecular Mitsunobu reaction. The stereochemistry of lobatamide
C was assigned as 8S,11S,15S by comparison to that of
synthesized diastereoisomers. Preparation and X-ray crystal-
lographic analysis of a lobatamide C derivative fully confirmed
the stereochemical assignment. A number of simplified lobata-
mide analogues have been prepared in an effort to probe
structure-activity relationships. Lobatamide C isomers and
simplified analogues were evaluated against bovine V-ATPase
from chromaffin granule membranes, which showed that the
salicylate phenol, enamide NH, and ortho-substitution of the
salicylate ester are important for V-ATPase inhibition. In
addition, a number of nanomolar acyclic salicylate enamides
inhibitors of bovine V-ATPase were uncovered in this study.
Further studies on the application of Cu(I)-catalyzed vinylic
substitution and preparation of other lobatamide derivatives are
in progress and will be reported in due course.
Acknowledgment. We thank Mr. Scott Magee for initial
experiments on the lobatamides, Dr. Michael R. Boyd for
providing authentic lobatamide C, Prof. J. Snyder (Boston
University) for helpful comments, Dr. Emil Lobkovsky (Cornell
University) for X-ray crystallographic analysis, Waters, Inc. for
providing HPLC equipment, Rhodia Chirex for providing salen
catalysts, and Chemetall GmBH (Boston, MA) for providing
Cs2CO3. Financial support for this study was provided by
NIH (GM62842, J.A.P, Jr., and GM28703, GM58903, B.J.B.).
C.-T.L. was supported in part by the Boston University Un-
dergraduate Research Opportunities Program and Pfizer Summer
Undergraduate Research Fellowship.
Supporting Information Available: Experimental procedures
and characterization data for all new compounds, including
X-ray crystal structure coordinates for 84. X-ray crystallographic
file in CIF format. This material is available free of charge via
the Internet at http://pubs.acs.org
JA0352350
9
J. AM. CHEM. SOC. VOL. 125, NO. 26, 2003 7901