A novel application of a Pd(0)-catalyzed nucleophilic substitution reaction to the regio- and stereoselective synthesis of lactam analogues of the epothilone natural products

Article abstract of DOI:10.1021/ja001899n

Several lactam analogues of the epothilones were prepared using a concise semisynthetic approach starting with the unprotected natural products. Highlighted in this strategy is a novel regio- and stereoselective Pd(0)-catalyzed azidation reaction of a macrocyclic lactone. Subsequent reduction and macrolactamization of the resulting azide acid intermediates provided the desired macrolactams in satisfactory overall yields. The entire three-step sequence was streamlined into a 'one-pot' process for the epothilone B-lactam, BMS-247550, which is currently undergoing phase I clinical trials. An initial total synthesis route to prepare the lactam analogue of epothilone C was completed and compared to the more direct semisynthesis approach. All of the lactam analogues were evaluated in vitro and the results are discussed.

Full text of DOI:10.1021/ja001899n

8890
J. Am. Chem. Soc. 2000, 122, 8890-8897
A Novel Application of a Pd(0)-Catalyzed Nucleophilic Substitution
Reaction to the Regio- and Stereoselective Synthesis of Lactam
Analogues of the Epothilone Natural Products
,
†
†
†
†
Robert M. Borzilleri,* Xiaoping Zheng, Robert J. Schmidt, James A. Johnson,
†
§
‡
§
Soong-Hoon Kim, John D. DiMarco, Craig R. Fairchild, Jack Z. Gougoutas,
Francis Y. F. Lee, Byron H. Long, and Gregory D. Vite
‡
‡
†
Contribution from the DiVisions of DiscoVery Chemistry, Oncology Drug DiscoVery, and Analytical
Research and DeVelopment, The Bristol-Myers Squibb Pharmaceutical Research Institute, P.O. Box 4000,
Princeton, New Jersey 08543-4000
ReceiVed May 30, 2000
Abstract: Several lactam analogues of the epothilones were prepared using a concise semisynthetic approach
starting with the unprotected natural products. Highlighted in this strategy is a novel regio- and stereoselective
Pd(0)-catalyzed azidation reaction of a macrocyclic lactone. Subsequent reduction and macrolactamization of
the resulting azide acid intermediates provided the desired macrolactams in satisfactory overall yields. The
entire three-step sequence was streamlined into a “one-pot” process for the epothilone B-lactam, BMS-247550,
which is currently undergoing phase I clinical trials. An initial total synthesis route to prepare the lactam
analogue of epothilone C was completed and compared to the more direct semisynthesis approach. All of the
lactam analogues were evaluated in vitro and the results are discussed.
Introduction
substantial resources toward developing synthetic routes to these
5
-10
11-13
natural products
and related analogues.
Elegant syn-
Epothilones A (1, Epo A) and B (2, Epo B) are the initial
members of a novel family of cytotoxic and antifungal mac-
rolides originally isolated at Gesellschaft f u¨ r Biotechnologische
Forschung (GBF) by H o¨ fle and Reichenbach via fermentation
thetic strategies incorporating the ring-closing olefin metathesis
reaction (RCM), macrolactonization, Suzuki coupling, or ester-
(5) For a general review of the “Chemical Biology of Epothilones,”
1
of the myxobacterium Sorangium cellulosum. Bollag and co-
see: Nicolaou, K. C.; Roschanger, F.; Vourloumis, D. Angew. Chem., Int.
Ed. Engl. 1998, 37, 2014 and references therein.
workers independently isolated these unique natural products
and discovered that their cytotoxic effects result from the
induction of tubulin polymerization and stabilization of micro-
(
6) (a) Meng, D.; Bertinato, P.; Balog, A.; Su, D.-S.; Kamenecka, T.;
Sorensen, E. J.; Danishefsky, S. J. J. Am. Chem. Soc. 1997, 119, 10073.
b) Balog, A.; Meng, D.; Kamenecka, T.; Bertinato, P.; Su, D.-S.; Sorensen,
(
2
tubules leading to G2-M arrest in the cell cycle and ultimately
E. J.; Danishefsky, S. J. Angew. Chem., Int. Ed. Engl. 1996, 35, 2801. (c)
Su, D.-S.; Meng, D.; Bertinato, P.; Balog, A.; Sorensen, E. J.; Danishefsky,
S. J.; Zheng, Y.-H.; Chou, T.-C.; He, L.; Horwitz, S. B. Angew. Chem.,
Int. Ed. Engl. 1997, 36, 757.
(7) (a) Nicolaou, K. C.; He, Y.; Vourloumis, D.; Vallberg, H.; Roschan-
gar, F.; Sarabia, F.; Ninkovic, S.; Yang, Z.; Trujillo, J. I. J. Am. Chem.
Soc. 1997, 119, 7960. (b) Nicolaou, K. C.; Ninkovic, S.; Sarabia, F.;
Vourloumis, D.; He, Y.; Vallberg, H.; Finlay, M. R. V.; Yang, Z. J. Am.
Chem. Soc. 1997, 119, 7974. (c) Nicolaou, K. C.; Winssinger, N.; Pastor,
J. A.; Ninkovic, S.; Sarabia F.; He, Y.; Vourloumis, D.; Yang, Z.; Li, T.;
Giannakakou, P.; Hamel, E. Nature 1997, 387, 268. (d) Nicolaou, K. C.;
Sarabia, F.; Ninkovic, S.; Yang, Z. Angew. Chem., Int. Ed. Engl. 1997, 36,
apoptosis.2a This mode of action for the epothilones, at least in
2
a
vitro and presumably in vivo, was shown by the Merck and
3
4
GBF groups to be identical to that first described by Horwitz
for the clinically successful chemotherapeutic agent Taxol
paclitaxel). More significantly, however, unlike paclitaxel the
(
epothilones are not affected by known mechanisms of resistance
such as P-glycoprotein overexpression (multidrug resistance)
or mutated tubulin.2
Following the initial report by H o¨ fle and co-workers, a
number of groups became intrigued by the epothilones’ unique
structure and impressive biological profile, and thus committed
5
25. (e) Yang, Z.; He, Y.; Vourloumis, D.; Vallberg, H.; Nicolaou, K. C.
Angew. Chem., Int. Ed. Engl. 1997, 36, 166.
8) (a) Schinzer, D.; Bauer, A.; Schieber, J. Chem. Eur. J. 1999, 5, 2492.
b) Schinzer, D.; Bauer, A.; B o¨ hm, O. M.; Limberg, A.; Cordes, M. Chem.
(
(
*
Corresponding author. E-mail: borzillr@bms.com.
Discovery Chemistry.
Oncology Drug Discovery.
Eur. J. 1999, 5, 2483. (c) Taylor, R. E.; Galvin, G. M.; Hilfiker, K. A.;
Chen, Y. J. Org. Chem. 1998, 63, 9580. (d) White, J. D.; Carter, R. G.;
Sundermann, K. F. J. Org. Chem. 1999, 64, 684. (e) May, S. A.; Grieco,
P. A. J. Chem. Soc., Chem. Commun. 1998, 1597. (f) Mulzer, J.;
Mantoulidis, A.; O¨ hler, E. Tetrahedron Lett. 1998, 39, 8633. (g) Martin,
H. J.; Drescher, M.; Mulzer, J. Angew. Chem., Int. Ed. Engl. 2000, 39,
581.
(9) (a) Harris, C. R.; Kuduk, S. D.; Balog, A.; Savin, K.; Glunz, P. W.;
Danishefsky, S. J. J. Am. Chem. Soc. 1999, 121, 7050. (b) Chou, T.-C.;
Zhang, X.-G.; Harris, C. R.; Kuduk, S. D.; Balog, A.; Savin, K. A.; Bertino,
J. R.; Danishefsky, S. J. Proc. Natl. Acad. Sci. U.S.A. 1998, 95, 15798. (c)
Chou, T.-C.; Zhang, X.-G.; Balog, A.; Su, D.-S.; Meng, D.; Savin, K.;
Bertino, J. R.; Danishefsky, S. J. Proc. Natl. Acad. Sci. U.S.A. 1998, 95,
9642. (d) Su, D.-S.; Meng, D.; Bertinato, P.; Balog, A.; Sorensen, E. J.;
Danishefsky, S. J.; Zheng, Y.-H.; Chou, T.-C.; He, L.; Horwitz, S. B. Angew.
Chem., Int. Ed. Engl. 1997, 36, 757.
†
‡
§
Analytical Research and Development.
(
1) H o¨ fle, G.; Bedorf, N.; Gerth, K.; Reichenbach, H. (GBF), German
Patent DE 91-4138042, 1993; Chem. Abstr. 1993, 120, 52841.
2) (a) Bollag, D. M.; McQueney, P. A.; Zhu, J.; Hensens, O.; Koupal,
L.; Liesch, J.; Goetz, M.; Lazarides, E.; Woods, C. M. Cancer Res. 1995,
5, 2325. (b) Bollag, D. M. Exp. Opin. InVest. Drugs 1997, 6, 867. (c)
Kowalski, R. J.; Giannakakou, P.; Hamel, E. J. Biol. Chem. 1997, 272,
(
5
2
1
534. (d) Wolff, A.; Technau, A.; Brandner, G. Int. J. Oncol. 1997, 11,
23.
(3) (a) Gerth, K.; Bedorf, N.; H o¨ fle, G.; Irschik, H.; Reichenbach, H. J.
Antibiot. 1996, 49, 560. (b) H o¨ fle, G.; Bedorf, N.; Steinmetz, H.; Schomburg,
D.; Gerth, K.; Reichenbach, H. Angew. Chem., Int. Ed. Engl. 1996, 35,
1
567.
4) (a) Schiff, P. B.; Fant, J.; Horwitz, S. B. Nature 1979, 277, 665. (b)
Hyams, J. S.; Loyd, C. W. Microtubules; Wiley-Liss: New York, 1994.
(
(10) Nicolaou, K. C.; He, Y.; Roschangar, F.; King, N. P.; Vourloumis,
D.; Li, T. Angew. Chem., Int. Ed. Engl. 1998, 37, 84.
1
0.1021/ja001899n CCC: $19.00 © 2000 American Chemical Society
Published on Web 08/26/2000

Synthesis of Epothilone-Lactams
J. Am. Chem. Soc., Vol. 122, No. 37, 2000 8891
enolate-aldehyde condensation have been developed to complete
the total syntheses of these interesting compounds. In addition
In contrast, our preliminary studies in murine tumor models
indicated that at tolerated doses Epo B lacks in vivo activity,
resulting from rapid inactivation of the compound in the
systemic circulation (T1/2 ≈ 40 min in mouse plasma). It was
postulated that this lack of antitumor activity was due to esterase-
mediated hydrolysis of the lactone moiety to form an inactive
ring-opened species. This hypothesis was supported by the
finding that the cell growth inhibitory effects of Epo B could
be mitigated by preincubation of the drug substance in murine
plasma.
5
to these efforts, other members of the epothilone family, namely
epothilones C (3, Epo C), D (4, Epo D), E (5, Epo E), and F (6,
Epo F), were isolated as minor metabolites from the fermentation
19
14
6-8,10
13a
process or as synthetic intermediates
or via semisynthesis.
Currently, all three of these approaches are being explored by
multiple research groups to prepare various epothilone ana-
logues.
As part of our ongoing research program aimed at preparing
metabolically stable epothilone derivatives and establishing a
comprehensive structure-activity relationship (SAR) of these
15,16,19
analogues,
we have focused on replacing the metabolically
labile lactone moiety and altering this portion of the epothilone
core. In this article we describe the preparation of several
macrocyclic lactam derivatives of the epothilones based on a
semisynthetic approach from the corresponding natural mac-
rolides 1 and 2. In particular, we highlight the concise three-
step synthesis of the clinical agent BMS-247550 involving a
novel Pd(0)-catalyzed ring-opening reaction of unprotected Epo
B as the key step. In addition, we provide a description of our
initial attempts to prepare the Epo C-lactam via a total synthesis
route. Finally, a discussion of the in vitro biological profiles of
these analogues will be provided.
Moreover, major pharmaceutical companies have become
increasingly interested in the epothilones and/or related ana-
logues as potential chemotherapeutic agents capable of comple-
menting existing anticancer therapies.15-18 Several factors have
made this exciting class of natural products an attractive
oncology target, namely: (1) irreversible microtubule stabiliza-
tion is a clinically validated biological target; (2) epothilones
are active in vitro against a number of paclitaxel-resistant tumor
cell lines; (3) epothilones exhibit better water solubility than
taxanes; (4) epothilones are readily obtained by fermentation;
and (5) the epothilone structure is amenable to both total
synthesis and semisynthesis. Furthermore, Danishefsky and co-
workers demonstrated in vivo that Epo B and Epo D display
antitumor effects in both paclitaxel-sensitive and -resistant
murine tumor models.9
Results and Discussion
Soon after the discovery of the mechanism of action of the
epothilones a research collaboration was established between
GBF and our laboratories which provided us with access to
significant quantities of the epothilone natural products. How-
ever, from the outset, we were interested in comparing both
total synthesis and semisynthesis approaches, since either
strategy would clearly offer distinct advantages. Total synthesis
could provide access to a greater number of structurally diverse
analogues, and potentially lead to novel, biologically active
compounds. Conversely, semisynthesis, while not as flexible,
could allow for shorter synthetic routes and provide “unlimited”
scale-up potential.
Total Synthesis Approach.20 We set out to assess the
practicality of a total synthesis approach to epothilone analogues
from a pharmaceutical development perspective. Our initial
strategy to prepare the epothilone lactams was based on a RCM
route initially reported by Nicolaou for the syntheses of
epothilones A and C.
(11) (a) Nicolaou, K. C.; Hepworth, D.; Finlay, M. R. V.; King, N. P.;
Werschkun, B.; Bigot, A. Chem. Commun. 1999, 519. (b) Nicolaou, K. C.;
Sarabia, F.; Ninkovic, S.; Finlay, M. R. V.; Boddy, C. N. C. Angew. Chem.,
Int. Ed. Engl. 1998, 37, 81. (c) Nicolaou, K. C.; Ninkovic, S.; Finlay, M.
R. V.; Sarabia, F.; Li, T. Chem. Commun. 1997, 2343. (d) Nicolaou, K. C.;
Vourloumis, D.; Li, T.; Pastor, J.; Winssinger, N.; He, Y.; Ninkovic, S.;
Sarabia, F.; Vallberg, H.; Roschangar, F.; King, N. P.; Finlay, M. R. V.;
Giannakakou, P.; Verdier-Pinard, P.; Hamel, E. Angew. Chem., Int. Ed.
Engl. 1997, 36, 2097. (e) Nicolaou, K. C.; Vallberg, H.; King, N. P.;
Roschangar, F.; He, Y.; Vourloumis, D.; Nicolaou, C. Chem. Eur. J. 1997,
7
Thus, thiazole fragment 7, derived from commercially avail-
able L-allylglycine was coupled to the polypropionate acid 87a
(16) Johnson, J.; Kim, S.-H.; Bifano, M.; DiMarco, J.; Fairchild, C.;
Gougoutas, J.; Lee, F.; Long, B.; Tokarski, J.; Vite, G. Org. Lett. 2000, 2,
1537.
3
, 1957.
12) Su, D.-S.; Balog, A.; Meng, D.; Bertinato, P.; Danishefsky, S. J.;
(
(17) Novartis Pharma AG. TA Oncology Research, Basel, Switzerland
in collaboration with The Scripps Research Institute and Chemisches Institut
der Otto-von-Guericke-Universitat: Altmann, K.-H.; Caravatti, G.; Floer-
sheimer, A.; Wartmann, M.; Nicolaou, K. C.; Schinzer, D. Synthetic and
Semisynthetic Analogs of Epothilones: Chemistry and Biological Activity.
Book of Abstracts, 219th National Meeting of the American Chemical
Society, San Francisco, CA, March 26-30, 2000; American Chemical
Society: Washington, DC, 2000; Abstract 287.
Zheng, Y.-H.; Chou, T.-C.; He, L.; Horwitz, S. B. Angew. Chem., Int. Ed.
Engl. 1997, 36, 2093.
(13) (a) H o¨ fle, G.; Glaser, N.; Kiffe, M.; Hecht, H.-J.; Sasse, F.;
Reichenbach, H. Angew. Chem., Int. Ed. Engl. 1999, 38, 1971. (b) Sefkow,
M.; Kiffe, M.; Schummer, D.; H o¨ fle, G. Bioorg. Med. Chem. Lett. 1998, 8,
3
8
025. (c) Sefkow, M.; Kiffe, M.; H o¨ fle, G. Bioorg. Med. Chem. Lett. 1998,
, 3031. (d) Sefkow, M.; H o¨ fle, G. Heterocycles 1998, 48, 2485.
(
14) Reichenbach, H.; H o¨ fle, G.; Gerth, K.; Steinmetz, H. PCT Int. Appl.
WO9822461, 1998.
15) For a preliminary account of the work described herein and
(18) Schering AG. Preclinical Drug Research, Berlin, Germany: Klar,
U.; Skuballa, W.; Schwede, W.; Buchmann, B. Epothilones. Book of
Abstracts, 219th National Meeting of the American Chemical Society, San
Francisco, CA, March 26-30, 2000; American Chemical Society: Wash-
ington, DC, 2000; Abstract 288.
(
additional studies on the epothilones, see: (a) Vite, G.; Borzilleri, R.; Fooks,
C.; Johnson, J.; Kim, S.-H.; Leavitt, K.; Mitt, T.; Regueiro-Ren, A.; Schmidt,
R.; Zheng, P.; Lee, F. Epothilones A and B: Springboards for the Synthesis
of Promising Antimitotic Agents. Book of Abstracts, 219th National Meeting
of the American Chemical Society, San Francisco, CA, March 26-30, 2000;
American Chemical Society: Washington, DC, 2000; Abstract 286. (b) Vite,
G. D.; Borzilleri, R. M.; Kim, S.-H.; Johnson, J. A. PCT Int. Appl.
WO9902514, 1999.
(19) Bristol-Myers Squibb, PRI. Unpublished results.
(20) Following the publication of our patent application covering this
work (cf. ref 15b) and submission of this manuscript, an article describing
a different total synthesis approach to 21 appeared, see: Stachel, S. J.;
Chappell, M. D.; Lee, C. B.; Danishefsky, S. J.; Chou, T.-C.; He, L.;
Horwitz, S. B. Org. Lett. 2000, 2, 1637.

8892 J. Am. Chem. Soc., Vol. 122, No. 37, 2000
Borzilleri et al.
Scheme 1a
Scheme 2a
a
(
a) Pd(PPh
3
)
4
, 10 mol %, NaN
, THF, 45 °C for 14 h, then 28% NH
, EtOH, PtO , 50 wt %, 10 h, then an additional 25 wt
, 10 h, or PMe , THF-H O, 25 °C, 2 h, 53-89%; (c) DPPA,
3
, DMF (2.5 mM), 4 °C, 24 h or EDCl, HOBt, MeCN (0.03
3
, degassed THF-H
2
O, 45 °C, 1 h,
6
°
%
5-70%; (b) PPh
C for 4 h, or H
of PtO
NaHCO
3
4
OH, H O, 45
2
2
2
2
3
2
a
(
a) EDCl, HOBt, DMF, 15 h, 77%; (b) RuBnCl
2
(PCy
3
)
2
2
, C
Cl , 0 °C,
2
6 6
H (4
M), 25-65%.
mM), 22 h, 34% E-isomer and 7% Z-isomer; (c) TFA, CH
h, sat. aq NaHCO solution, 68%.
4
3
developments which had surfaced from our semisynthetic efforts
starting with natural product, led us to abandon the total
synthesis efforts toward these important lactam derivatives.
Semisynthesis Approach. With access to the natural prod-
ucts, we set out to develop a general strategy to directly convert
the lactone to a lactam. Careful analysis of the epothilone
structure led to the critical observation that the lactone is
actually an allylic ester and may be susceptible to palladium-
catalyzed ring-opening to afford an intermediate π-allylpalla-
using standard EDCI-HOBt [EDCI ) 1-(3-dimethylaminopro-
pyl)-3-ethylcarbodiimide, HOBt ) 1-hydroxybenzotriazole]
conditions in DMF at room temperature, to afford olefin
metathesis precursor 9 in good yield (Scheme 1). Surprisingly,
the RCM reaction21 of the bis-olefin 9 which was promoted
with the Grubbs ruthenium-based catalyst (∼0.7 mol %
RuBnCl2(PCy3)2) in benzene (0.004 M) at 25 °C (22 h) afforded
predominantly the E-isomeric macrolactam 10, along with only
a minor amount of the Z-olefin 11 in nearly a 5:1 ratio.
Subsequent deprotection of the silyl ether of 10 with trifluoro-
acetic acid in CH2Cl2 at 0 °C (4 h) followed by a sodium
bicarbonate workup gave the E-Epo C-lactam 12.
2
3
dium complex (cf. 16, Scheme 2). This latent species could
then be trapped in situ with a “soft” external nucleophile, such
as azide to form an allylic azide. If successful, this sequence of
events would provide the nitrogen atom necessary at C15 and
a readily available coupling partner (carboxylic acid) at C1.
Subsequent reduction of the azide followed by macrocyclization
would provide the epothilone-lactams. One concern of this
strategy, however, was the regio- and stereochemical outcome
of the Pd(0)-catalyzed nucleophilic substitution reaction involv-
ing a macrolactone substrate, since there was no precedent.
Certainly, the extensively studied palladium-catalyzed sub-
stitution of allylic acetates has become an important transforma-
tion in organic synthesis since it typically proceeds with a high
While this study demonstrated that the critical RCM reaction
was operative, the observed product ratio was unsatisfactory.
19
Based on the poor intrinsic biological activities of the unnatural
E-like geometric isomers of Epo C and Epo A, and the poor
activity observed for E-Epo C-lactam 12 (vide infra), it was
imperative that we discover a reliable method to access the more
desirable Z-stereochemistry.
From these preliminary results, it was unlikely that the lengthy
RCM approach would provide sufficient quantities of material
needed to sustain a significant medicinal chemistry effort, even
if conditions could be found that would reverse the stereochem-
ical outcome or allow ring closure of more highly substituted
olefins as required for Epo B/D analogues. Although the reaction
conditions and alternative catalysts such as the molybdenum-
2
3
degree of both regio- and stereochemical control. Depending
on the nature of the nucleophile, either overall retention or
inversion of configuration is observed for this process. Surpris-
ingly, there have been relatively few examples of palladium-
catalyzed substitution reactions of unsaturated lactones in the
2
4-29
22
literature.
In particular, only two types of systems have
based Schrock catalyst were not examined in detail, it was
6-8a,b,e
recognized even in the epothilone literature
that the
(23) For recent reviews on related π-allylpalladium chemistry, see: (a)
stereochemical outcome of the RCM process is highly substrate
dependent. These discouraging results combined with exciting
Tsuji, J. Palladium Reagents and Catalysts: InnoVations in Organic
Synthesis; Wiley and Sons: New York, 1995; p 291. (b) Heumann, A.;
R e´ glier, M. Tetrahedron 1995, 51, 975. (c) Frost, C. G.; Howarth, J.;
Williams, J. M. J. Tetrahedron: Asymmetry 1992, 3, 1089. (d) Godlesky,
S. A. In ComprehensiVe Organic Synthesis; Trost, B. M., Ed.; Pergamon
Press: Oxford, 1991; Vol. 4, p 585. (e) Consiglio, G.; Waymouth, R. M.
Chem. ReV. 1989, 89, 257.
(21) For recent reviews on RCM reactions, see: (a) Phillips, A. J.; Abell,
A. D. Aldrichimica Acta 1999, 32, 75. (b) Armstrong, S. K. J. Chem. Soc.,
Perkin Trans. 1 1998, 371. (c) Schmalz, H.-G. Angew. Chem., Int. Ed. Engl.
1
1
995, 34, 1833. (d) Grubbs, R. H.; Miller, S. J.; Fu, G. C. Acc. Chem. Res.
995, 28, 446.
(24) (a) Trost, B. M.; Verhoeven, T. R.; Fortunak, J. M. Tetrahedron
Lett. 1979, 20, 2301. (b) Trost, B. M.; Verhoeven, T. R. J. Am. Chem. Soc.
1980, 102, 4730.
(22) Schrock, R. R.; Murdzek, J. S.; Bazan, G. C.; Robbins, J.; DiMare,
M.; O’Regan, M. J. Am. Chem. Soc. 1990, 112, 3875.

Synthesis of Epothilone-Lactams
J. Am. Chem. Soc., Vol. 122, No. 37, 2000 8893
been explored. In the first case, bridged lactones were shown
to undergo ring-opening in the presence of a Pd(0) catalyst to
provide intermediate π-allylpalladium complexes which were
phine, in the absence of any base, cleanly reduced the azide
after a 1-2 h exposure at room temperature.
Several sets of reaction conditions were examined for the
2
4-28
30
then rapidly trapped in situ with a variety of nucleophiles.
final macrolactamization step in the synthesis of Epo B-lactam
Relief of ring strain was clearly the driving force behind these
transformations. In the second series, it was shown that unsatur-
ated, fused [3.3.0] bicyclic lactones also undergo Pd(0)-promoted
15b. Initially, diphenylphosphoryl azide (DPPA, 4 equiv) and
solid sodium bicarbonate (8 equiv) in degassed DMF (2.5 mM)
at 0 °C (24 h) were used to promote the macrocyclization of
amino acid 14b to provide the desired lactam 15b. An X-ray
crystal structure of the bis-triethylsilyl ether derivative of lactam
15b allowed for unambiguous structural assignment, including
confirmation of the regio- and C15-stereochemistry which was
obtained in the initial Pd(0)-catalyzed reactions (vide supra).31
In general, the solvents (DMF, CH2Cl2, or toluene), reaction
time (14-36 h), temperature (-10-25 °C), and concentration
2
4b,29
ring-opening to generate disubstituted cyclopentenes.
In
these examples, the requisite π-allylpalladium intermediates
possess an internal nucleophile (carboxylate) which effectively
competes with weak, external nucleophiles in an entropy driven
process.
We postulated that a Pd(0)-catalyzed azidation25a reaction
involving the epothilones should benefit from the entropy
associated with a ring-opened macrocycle, forcing the equilib-
rium toward the formation of an intermediate π-allylpalladium
species. We were therefore gratified to discover that when
unprotected Epo A (1) or Epo B (2) was treated with a catalytic
amount (10 mol %) of tetrakis(triphenylphosphine)palladium-
(0.05-0.5 mM) had varying effects on the overall yield of the
macrocyclization step which never exceeded 40%. At this point
we examined alternative coupling reagents which were known
3
0,32
to effect macrolactamization reactions,
such as PyBroP
[
bromo tripyrrolidinophosphonium hexafluorophosphate],
BOP-Cl [bis(2-oxo-3-oxazolidinyl)phosphinic chloride], and
HATU [O-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluro-
nium hexafluorophosphate]. In the attempts with PyBrop (PhMe,
(
0) and sodium azide in degassed THF-H2O at 45 °C (1 h) the
azide acids 13a/b were obtained as single diastereomers in good
yield (Scheme 2).
2
5-80 °C), BOP-Cl (CH2Cl2, 0-25 °C) or HATU (MeCN-
Mechanistically, this reaction can be considered to proceed
via anti-attack of palladium(0) on the allylic lactone to form
an initial π-allylpalladium complex 16. Subsequent attack by
azide at C15, from the opposite face of the palladium/ligands,
provides 13 with the desired regio- and stereochemistry (vide
infra). In effect, net retention of configuration at C15 was
observed. Surprisingly, all of the potentially labile functionality
found in the epothilone core, including the epoxide, remained
intact. Although not extensively investigated, use of other
nucleophiles such as benzylamine resulted in lower yields
DMF, 25 °C), significant decomposition occurred along with
the recovery of minor amounts of starting material or lactam
1
5b. However, when amino acid 14b was treated with EDCI
and HOBt or HOAt [1-hydroxy-7-azabenzotriazole] in aceto-
nitrile (0.03 M) at 25 °C (2 h), reproducible yields (∼65%) of
the desired macrolactam were obtained.
Epo F-lactam 17 was prepared in the same stepwise fashion
as outlined above beginning with readily available epothilone
13a
F. Analogously, unprotected Epo F underwent the Pd(0)-
catalyzed azidation to afford the intermediate azide acid, albeit
in much lower yields than in the cases of epothilones 1 and 2.
Macrolactamization of the subsequent amino acid furnished the
desired lactam 17.
(
∼18%), while methylamine only provided recovered starting
material.
Following our success in this critical step, we turned our
attention to completing the synthesis of the lactam analogues
of the epothilones. The initial reactions were carried out
exclusively on the Epo B series. Attempts to chemoselectively
reduce the azide acid 13b to amino acid 14b via a Staudinger
reaction were problematic. Treatment of azide 13b with tri-
phenylphosphine (2 equiv) in a THF-H2O (10:1) mixture at
4
5 °C (14 h) gave the intermediate iminophosphorane as the
major product along with a small amount of the desired amino
acid 14b. Addition of ammonium hydroxide to the reaction
mixture was required to complete the hydrolysis of this
unusually stable intermediate. Alternatively, reduction was
accomplished in the absence of base using polymer supported
triphenylphosphine or using hydrogenation conditions with
stoichiometric amounts of Adams’ catalyst (PtO2). In either case,
prolonged reaction times were required. Finally, after further
optimization it was found that the highly reactive trimethylphos-
We continued to optimize and scale up each of the three steps
in the semisynthesis of lactam analogues 15a/b in order to
provide sufficient quantities for biological testing. During this
time it became apparent that all of the reagents involved in the
sequence were completely compatible with each other. Thus,
in an unconventional approach, all three steps were carried out
sequentially in a single reaction vessel and without isolation of
intermediates (Scheme 3). Remarkably, Epo A and Epo B were
converted to the corresponding lactams 15a and 15b in
acceptable (20-25%) overall yields. In addition, the entire
process, including purification, could be accomplished in 1 day.
One obvious issue that needed to be addressed was whether
global protection of the molecule would improve the yields in
(25) (a) Murahashi, S.-I.; Taniguchi, Y.; Imada, Y.; Tanigawa, Y. J. Org.
Chem. 1989, 54, 3292. (b) Murahashi, S.-I.; Tanigawa, Y.; Imada, Y.;
Taniguchi, Y. Tetrahedron Lett. 1986, 27, 227. (c) Murahashi, S.-I.; Imada,
Y.; Taniguchi, Y.; Kodera, Y. Tetrahedron Lett. 1988, 29, 2973.
(
26) Tsuda, T.; Okada, M.; Nishi, S.; Seagusa, T. J. Org. Chem. 1986,
1, 421.
27) (a) Negishi, E.; John, R. A. J. Org. Chem. 1983, 48, 4098. (b)
Matsushita, H.; Negishi, E. J. Chem. Soc., Chem. Commun. 1982, 160.
28) (a) Bystr o¨ m, S. E.; Aslanian, R.; B a¨ ckvall, J.-E. Tetrahedron Lett.
985, 26, 1749. (b) Genet, J. P.; Balabane, M.; B a¨ ckvall, J.-E.; Bystr o¨ m,
S. E. Tetrahedron Lett. 1983, 24, 2745.
29) (a) Aggarwal, V. K.; Monteiro, N.; Tarver, G. J.; McCague, R. J.
5
(
(30) For a comprehensive review on macrolactamization reactions, see:
Humphrey, J. M.; Chamberlin, A. R. Chem. ReV. 1997, 97, 2243.
(31) Crystallographic data (excluding structure factors) for the compound
have been deposited with the Cambridge Crystallographic Data Centre as
Supplementary Publication No. CCDC 147158. Copies of the data can be
obtained free of charge on application to CCDC, 12 Union Road, Cambridge
CB21EZ, UK (fax: (+44)1223-336-033; e-mail: deposit@ccdc.cam.ac.uk).
(32) Trost, B. M.; Ceschi, M. A.; K o¨ nig, B. Angew. Chem., Int. Ed. Engl.
1997, 36, 1486.
(
1
(
Org. Chem. 1997, 62, 4665. (b) Aggarwal, V. K.; Monteiro, N. J. Chem.
Soc., Perkin Trans. 1 1997, 2531. (c) Aggarwal, V. K.; Monteiro, N.; Tarver,
G. J.; Lindell, S. D. J. Org. Chem. 1996, 61, 1192.

8894 J. Am. Chem. Soc., Vol. 122, No. 37, 2000
Borzilleri et al.
Scheme 3a
Scheme 4a
a
(
a) Pd(PPh
3
)
4
, 10 mol %, NaN
3
2
, degassed THF-H O, 45 °C, 20
min, then PMe
C, 4-12 h, 20-25%.
3
, 1-2 h, then MeCN-DMF (20:1), EDCl-HOBt, 25
°
each reaction. Although this strategy would potentially add two
steps to the route, we set out to compare the efficiency of the
two sequences. Both hydroxyl groups of Epo B were protected
as either the trimethylsilyl or triethylsilyl ethers (10 equiv of
trimethylsilyl or triethylsilyl chloride, 15 equiv of i-Pr2EtN, 0.1
equiv of DMAP, CH2Cl2, 40 °C, 18 h), and these compounds
were subjected to the “one-pot” sequence. In each case only
minor improvement in the overall yield (25-35%) was ob-
served, and thus further work using fully protected epothilone
derivatives was suspended.
a
(
a) R ) H: TiCp
2
Cl
2
, Mg(s), THF, 49%, 20:1 ratio of 20 to 12;
(
6
b) R ) Me: WCl , n-BuLi, THF, 56%, 5:1 ratio of 21 to 22.
Table 1. In Vitro Data for Epothilones A-F and Lactam
Analogues
tubulin [a] EC0.01
HCT-116 [b] IC50
no.
compd
paclitaxel
Epo A
Epo B
Z-Epo C
E-Epo C
Z-Epo D
E-Epo D
Epo E
(µM)
(nM)
During the attempted semisynthesis of the lactam analogues
of Epo C (3) and Epo D (4), it became clear that there were
certain limitations associated with the Pd(0)-catalyzed nucleo-
philic substitution reaction. Unfortunately, lactones 3 and 4
failed to provide the corresponding azide acids when subjected
to our standard reaction conditions. In each case, a complicated
mixture of olefin isomers of trienes 18 and 19 was obtained as
-
1
5.0
2.3
1.4
2.3
3.2
0.42
2
3
5.0
21
64
160
-
4
0.80
11
17
6.5
97
6.0
0.77
NT
130
3.6
-
5
1
indicated by H NMR.
6
Epo F
1.8
1
1
1
1
2
2
2
2
E-Epo C-lactam
Epo A-lactam
Epo B-lactam
Epo F-lactam
Z-Epo C-lactam
Z-Epo D-lactam
E-Epo D-lactam
>1100
5a
5b
7
0
1
12
3.8
130
110
5.5
120
NT
65
2
>1200
>200
a
Tubulin polymerization assay performed using the method described
in refs 35a and 15b. b HCT-116, a human colon carcinoma cell line
cytotoxicity assay performed using the method described in ref 35b.
NT ) not tested.
To circumvent this problem, we utilized the stereoselective
deoxygenation chemistry recently disclosed by this laboratory
Biological Testing Results. The in vitro results for the lactam
to prepare Epo C and Epo D directly from Epo A and B,
1
9
respectively.16 Thus, unprotected Epo A-lactam 15a was treated
analogues and the natural epothilones are listed in Table 1. In
general, the tubulin-polymerizing and cytotoxic potencies of the
lactam analogues were clearly inferior to their natural epothilone
counterparts. One very important exception to this trend,
however, was lactam 15b (BMS-247550). This semisynthetic
analogue of Epo B was in fact comparable to paclitaxel in both
in vitro assays. After extensive in vitro testing in a number of
different cell lines, BMS-247550 was found to possess a superior
with the titanocene complex33 derived from TiCp2Cl2 and
magnesium to furnish Z-Epo C-lactam 20 along with a small
amount of the E-isomer 12 (Scheme 4). Interestingly, the
identical reaction conditions with unprotected Epo B-lactam 15b
led predominantly to decomposition products. Utilizing condi-
tions reported by Sharpless (WCl6/n-BuLi)34 for less complex
systems, Epo B-lactam 15b was converted to the lactams Z-21
and E-22 in good yields as a separable 5:1 mixture of geometric
isomers. The C12-C13 relative stereochemisty of the major
1
9
biological profile to that of paclitaxel. A second analogue
worth highlighting is lactam 21. This Epo D-lactam performed
well in the tubulin polymerization assay, but was 10- and 30-
fold less potent than Epo D and paclitaxel, respectively in cell
culture.
1
lactam isomers 20 and 21 was determined using 1D and 2D H
1
1
NMR techniques. In the case of lactam 20, the 2D H- H
NOESY NMR spectrum exhibited diagnostic H-12/H-13 and
H-11/H-14 NOE cross-peaks, thus indicating that these protons
When evaluated in vivo, BMS-247550 exhibited a broad
spectrum of antitumor activity against both paclitaxel-sensitive
and paclitaxel-resistant human tumor xenografts grown in
1
reside on the same side of the olefin (Z-relationship). 1D H-
1
H NOE NMR experiments on the Epo D-lactam 21 revealed
3
6
strong NOE enhancements between H-13 and the C-26 methyl
protons (irradiation on either peak provided enhancements). The
close proximity of these two groups again establishes the
Z-configurational assignment.
athymic mice. In addition, BMS-247550 was found to be
superior to the naturally occurring epothilones A-F in pre-
clinical antitumor models. It is postulated that this may be
partially attributed to the stabilization of the lactone functionality
toward metabolic cleavage. Due to its impressive in vivo profile
in several animal models, BMS-247550 was chosen for drug
development and clinical investigation.
(
33) Schobert, R.; Hoehlein, U. Synlett 1990, 8, 465.
(34) Sharpless, K. B.; Umbreit, M. A.; Nieh, M. T.; Flood, T. C. J. Am.
Chem. Soc. 1972, 94, 6538.

Synthesis of Epothilone-Lactams
J. Am. Chem. Soc., Vol. 122, No. 37, 2000 8895
Summary
(dd, 1 H, J ) 15.9, 3.3 Hz), 2.12 (dd, 1 H, J ) 15.9, 6.0 Hz), 2.06 (s,
3
1
H), 2.05-1.96 (m, 2 H), 1.76-1.70 (m, 1 H), 1.55-1.42 (m, 2 H),
.35-1.24 (m, 1 H), 1.21 (s, 3 H), 1.10 (s, 3 H), 1.02 (d, 3 H, J ) 6.6
After carefully evaluating a total synthesis approach, several
metabolically stable lactam analogues of the epothilones were
prepared via a semisynthesis route. Highlighted in this novel
strategy is a regio- and stereoselective π-allylpalladium nucleo-
philic substitution reaction which provided a pivotal ring-opened
intermediate with a nitrogen substituent at C15 of the epothilone
core. Chemoselective reduction of the intermediate azide fol-
lowed by macrolactamization of the resulting amino acid
afforded the desired lactams. In the case of the parent epo-
thilones 1 and 2, the entire sequence was telescoped into a three
step, “one-pot” process providing an efficient and practical
approach (20-25% yields) to the corresponding lactams. Further
optimization of this sequence is underway and will be reported
in due course. To prepare the macrolactams of Epo C and Epo
D, it was necessary to employ two highly effective deoxygen-
ation methods to convert the epoxides of 15a/b to the corre-
sponding olefins. The epothilone-lactams were evaluated in
vitro, and one analogue, 15b (BMS-247550) compared favorably
to paclitaxel and Epo B.
Hz), 0.89 (s, 9 H), 0.83 (d, 3 H, J ) 6.6 Hz), 0.11 (s, 3 H), 0.071 (s,
3 H); 13C NMR (125 MHz, CDCl
) δ 222.6, 169.8, 164.4, 152.7, 139.1,
138.5, 134.0, 119.4, 118.1, 115.7, 114.2, 74.6, 73.8, 55.4, 54.2, 42.3,
3
4
1
1.5, 37.7, 35.5, 34.2, 32.4, 26.1 (3C), 26.0, 22.6, 19.7, 19.2, 18.1,
+
+
6.1, 15.3, 9.6, -4.2, -4.8; HRMS (ESI ) m/z (M + H) calcd for
SSi: 619.3965, found: 619.3969.
4S-[4R*,7S*,8R*,9R*,15R*(E)]]-4-tert-Butyldimethylsilyloxy-8-
34 58 2 4
C H N O
[
hydroxy-5,5,7,9-tetramethyl-16-[1-methyl-2-(2-methyl-4-thiazolyl)-
ethenyl]-1-aza-13(E)-cyclohexadecene-2,6-dione (10). A solution of
9
(17 mg, 27 µmol) in degassed benzene (8.0 mL) was treated with
Grubbs’ catalyst ([bis(tricyclohexyl-phosphine)benzylidine ruthenium
dichloride, Strem Chemicals, 11 mg, 14 µmol) under Ar. The reaction
mixture was stirred at 25 °C for 15 h and treated again with an
additional portion of catalyst (5.0 mg, 4.5 µmol). After 7 h, the benzene
was removed in vacuo, and the black viscous residue was passed
through a pad of silica gel (1.0 × 3 cm) eluting with Et O (25 mL).
2
The eluent was concentrated in vacuo to afford a separable 5:1 (E/Z)
2
mixture of geometric isomers. PTLC (SiO , 1 mm plate, 2 elutions
with a 1:1:1 solution of hexane-toluene-ethyl acetate) afforded the
E-isomer 10 (5.1 mg, 34%) along with the corresponding Z-isomer
Taxanes (e.g., paclitaxel and docetaxel) are among the most
effective chemotherapeutic agents used for the treatment of
ovarian, breast, and lung cancers. However, many patients
develop resistance to taxane therapy, and several additional
cancers (e.g., colon and prostate) are refractory to taxanes. BMS-
+
+
(
1.0 mg, 6.7%). For 10: LRMS (ESI ) 591.4 (M + H). For the
+
+
-
-
Z-isomer 11: LRMS (ESI ) 591.2 (M + H); (ESI ) 589.3 (M - H) .
4S-[4R*,7S*,8R*,9R*,15R*(E)]]-4,8-Dihydroxy-5,5,7,9-tetramethyl-
6-[1-methyl-2-(2-methyl-4-thiazolyl)ethenyl]-1-aza-13(E)-cyclo-
hexadecene-2,6-dione (12). To a 1 dram vial charged with 10 (2.3
mg, 3.9 µmol) in CH Cl (0.4 mL) at 0 °C was added trifluoroacetic
[
1
247550 represents a novel “non-taxane” that has the potential
2
2
to extend the clinical activity of the tubulin-polymerizing class
of anticancer agents beyond that demonstrated with taxanes.
acid (0.1 mL). The reaction mixture was sealed under a blanket of Ar
and stirred at 0 °C. After 4 h, the volatiles were removed under a
3
constant stream of Ar at 0 °C. Saturated aqueous NaHCO (1 mL) and
Experimental Section
EtOAc (1 mL) were added to the residue, and the two layers were
separated. The aqueous phase was extracted with EtOAc (4 × 1 mL),
(
3S,6R,7S,8S)-N-(S)-[1-(2-Methyl-4-thiazolyl)-2-methyl-1(E)-5-
2 4
and the combined EtOAc layers were dried (Na SO ) and concentrated
hexadien-3-yl]-3-tert-butyldimethylsilyloxy-4,4,6,8-tetramethyl-7-hy-
droxy-5-oxo-12-trideceneamide (9). A solution of amine 7 (88 mg,
in vacuo. PTLC (SiO
CHCl
.2, CHCl
H), 6.07 (d, 1 H, NH, J ) 7.1 Hz), 5.54-5.47 (m, 1 H), 5.38-5.31
2
, 20 × 10 × 0.025 cm, eluting with 5% MeOH-
22
3
) afforded 12 (1.3 mg, 68%) as a white film: [R]
D
-93.5 (c
0
.42 mmol, cf. Supporting Information) in DMF (1.3 mL) at 0 °C was
1
0
3 3
); H NMR (400 MHz, CDCl ) δ 6.96 (s, 1 H), 6.44 (s, 1
7
a
treated sequentially with acid 8 (0.15 g, 0.35 mmol), 1-hydroxyben-
zotriazole (49 mg, 0.36 mmol), and 1-(3-dimethylaminopropyl)-3-
ethylcarbodiimide hydrochloride (0.10 g, 0.52 mmol). The reaction
mixture was gradually warmed to 25 °C, stirred for 15 h, and diluted
(m, 1 H), 4.48-4.43 (m, 1 H), 4.22 (d, 1 H, J ) 11.5 Hz), 3.67-3.58
(m, 2 H), 3.29-3.22 (m, 1 H), 3.21 (br d, 1 H, OH, J ) 3.8 Hz), 2.71
(s, 3 H), 2.44-2.29 (m, 3 H), 2.28-2.19 (m, 1 H), 2.17 (dd, 1 H, J )
with H
mL). The combined organic phases were washed with aqueous 5% HCl
10 mL), saturated aqueous NaHCO (10 mL), and saturated aqueous
NaCl (10 mL) and were dried (Na SO ) and concentrated in vacuo.
Flash chromatography (SiO
, 1.5 × 20 cm, 2.5% MeOH-CHCl
afforded 9 (0.17 g, 77%) as a white foam: [R] -54.1 (c 9.9, CHCl
) δ 6.90 (s, 1 H), 6.41 (s, 1 H), 5.83-5.68
m, 3 H), 5.13-5.07 (m, 2 H), 4.99-4.90 (m, 2 H), 4.52 (dd, 1 H,
J ) 14.3, 7.1 Hz), 4.41 (dd, 1 H, J ) 6.0, 3.8 Hz), 3.39 (s, 1 H),
.33-3.29 (m, 2 H), 2.69 (s, 3 H), 2.43-2.38 (m, 2 H), 2.33
2
O (3 mL). The mixture was extracted with EtOAc (3 × 10
1
1
4.5, 1.7 Hz), 2.08 (s, 3 H), 1.95-1.83 (m, 1 H), 1.71-1.62 (m, 2 H),
.51-1.38 (m, 1 H), 1.32 (s, 3 H), 1.27-1.18 (m, 2 H), 1.17 (d, 3 H,
(
3
+
J ) 6.8 Hz), 1.06 (s, 3 H), 1.01 (d, 3 H, J ) 6.8 Hz); HRMS (ESI )
m/z (M + H) calcd for C26
2
4
+
40 2 4
H N O S: 477.2709, found: 477.2798.
2
3
)
3
);
22
(3S,6R,7S,8S,12R,13S,15S)-15-Azido-12,13-epoxy-4,4,6,8,16-pen-
tamethyl-3,7-dihydroxy-17-(2-methyl-4-thiazolyl)-5-oxo-16(E)-hep-
tadecenoic acid (13a). A solution of epothilone A (1, 2.48 g, 5.02
mmol) and sodium azide (0.489 g, 7.53 mmol) in a degassed THF-
H O mixture (10:1, 55 mL) was treated with a catalytic amount (0.580
2
g, 0.502 mmol) of tetrakis(triphenylphosphine) palladium(0) under Ar.
D
1
H NMR (400 MHz, CDCl
3
(
3
The suspension was warmed to 45 °C for 1 h, and the resulting bright
(
35) (a) Tubulin polymerization assays were performed as described in
Swindell, C. S.; Krauss, N. E.; Horwitz, S. B.; Ringel, I. J. Med. Chem.
991, 34, 1176. Effective concentration (EC0.01) is defined as the interpolated
yellow, homogeneous solution was diluted with H
extracted with EtOAc (3 × 100 mL). The organic extracts were washed
with saturated aqueous NaCl (150 mL), dried (Na SO ), and concen-
trated in vacuo. The residue was purified by flash chromatography
SiO -MeOH to 95:5.0:0.5 CHCl -MeOH-
, 4.5 × 30 cm, 95:5 CHCl
AcOH gradient elution) to afford 13a (1.84 g, 69%) as a colorless oil:
2
O (50 mL) and
1
concentration capable of inducing an initial slope of 0.01 OD/min rate and
is calculated using the formula: EC0.01 ) concentration/slope. EC0.01 values
are expressed as the mean with standard deviation obtained from 3 different
concentrations. (b) Cytotoxicity was assessed in HCT-116 human colon
carcinoma cells by MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethox-
yphenyl)-2-(4-sulphenyl)-2H-tetrazolium, inner salt) assay as reported in
Riss, T. L.; Moravec, R. A. Mol. Biol. Cell 1992, 3 (Suppl.), 184a (Abstract
No. 1067).
2
4
(
2
3
3
[R]22
1
D
3 3
-60.7 (c 1.0, CHCl ); H NMR (400 MHz, CDCl ) δ 7.05 (s, 1
H), 6.60 (s, 1 H), 4.28-4.24 (m, 1 H), 4.18 (dd, 1 H, J ) 8.9, 5.1 Hz),
3
3
.40 (d, 1 H, J ) 8.8 Hz), 3.28-3.24 (m, 1 H), 2.88 (dd, 1H, J ) 7.8,
.9 Hz), 2.73 (s, 3 H), 2.48-2.36 (m, 2 H), 2.06 (s, 3 H), 1.92-1.88
(
36) Complete details of the preclinical efficacy data for BMS-247550
will be published elsewhere. Preliminary preclinical efficacy data was
recently presented: Lee, F. Y. F.; Vite, G. D.; Borzilleri, R. M.; Arico, M.
A.; Clark, J. L.; Fager, K. L.; Kan, D.; Kennedy, K. A.; Kim, A. S.-H.;
Smykla, R. A.; Wen, M.-L.; Kramer, R. A. BMS-247550: An Epothilone
Analog Possessing Potent Activity Against Paclitaxel-Sensitive and -Re-
sistant Human Tumors. Book of Abstracts, 91st Annual Meeting of the
American Association for Cancer Research, San Francisco, CA, April 1-5,
(m, 1 H), 1.73-1.61 (m, 1 H), 1.61-1.33 (a series of multiplets, 7 H),
1.31 (s, 3 H), 1.21 (s, 3 H), 1.12 (s, 3 H), 1.06 (d, 3 H, J ) 6.8 Hz),
13
0
1
4
9
.86 (d, 3 H, J ) 6.7 Hz); C NMR (100 MHz, CDCl
3
) δ 221.9, 175.8,
65.4, 151.2, 136.9, 121.8, 116.5, 74.6, 72.1, 68.3, 57.3, 54.1, 51.9,
0.8, 36.2, 35.2, 32.3, 31.5, 28.1, 23.5, 21.1, 18.6 (2C), 15.3, 14.3,
+
+
.9; HRMS (ESI ) m/z (M + H) calcd for C26
40 4 6
H N O S: 537.2669,
2
000; American Association for Cancer Research: Philadephia, PA, 2000;
LB-34.
found: 537.2745.

8
896 J. Am. Chem. Soc., Vol. 122, No. 37, 2000
Borzilleri et al.
Alternatively, a solution of 13b (29 mg, 53 µmol) in THF-H
(
3S,6R,7S,8S,12R,13S,15S)-15-Azido-12,13-epoxy-4,4,6,8,12,16-
2
O
hexamethyl-3,7-dihydroxy-17-(2-methyl-4-thiazolyl)-5-oxo-16(E)-
heptadecenoic acid (13b). By using epothilone B(2) as the starting
material and following the same procedure outlined above for epothilone
(0.5 mL) was treated with trimethylphosphine (1.0 M in toluene, 0.11
mL, 0.11 mmol) under Ar. The reaction mixture was stirred at 25 °C
(2 h) and concentrated in vacuo, and the residue was purified as
described above to afford 14b (20 mg, 71%).
[1S-[1R*,3R*(E),7R*,10S*,11R*,12R*,16S*]]-7,11-Dihydroxy-
8,8,10,12,16-pentamethyl-3-[1-methyl-2-(2-methyl-4-thiazolyl)ethe-
nyl]-4-aza-17-oxabicyclo[14.1.0]heptadecane-5,9-dione (15b). A so-
lution of 14b (0.95 g, 1.8 mmol) in degassed DMF (700 mL) was treated
2
2
A, 13b (1.9 g, 70%) was obtained as a colorless oil: [R]
D
-66.8 (c
) δ 7.05 (s, 1 H), 6.61 (s, 1
H), 4.25 (dd, 1 H, J ) 7.0, 5.5 Hz), 4.19 (dd, 1 H, J ) 9.4, 5.0 Hz),
.43 (dd, 1 H, J ) 8.4, 2.0 Hz), 3.28-3.24 (m, 1 H), 2.86 (dd, 1 H, J
7.6, 4.4 Hz), 2.73 (s, 3 H), 2.45-2.40 (m, 2 H), 2.06 (s, 3 H), 1.96-
1
1
.4, CH
3
OH); H NMR (400 MHz, CDCl
3
3
)
1
.88 (m, 1 H), 1.70-1.61 (m, 1 H), 1.61-1.33 (a series of multiplets,
H), 1.31 (s, 3 H), 1.25 (s, 3 H), 1.13 (s, 3 H), 1.08 (d, 3 H, J ) 6.9
3
with solid NaHCO (1.2 g, 14 mmol) and diphenylphosphoryl azide
7
(1.6 mL, 7.2 mmol) at 0 °C under Ar. The resulting suspension was
stirred at 4 °C for 24 h, diluted with cold phosphate buffer (750 mL,
pH ) 7) at 0 °C, and extracted with EtOAc (5 × 500 mL). The organic
extracts were washed with cold 10% aqueous LiCl (750 mL), dried
1
3
Hz), 0.88 (d, 3 H, J ) 6.8 Hz); C NMR (100 MHz, CDCl
3
) δ 222.2,
76.2, 165.5, 151.5, 137.1, 121.9, 116.7, 74.8, 72.3, 68.5, 61.6, 52.1,
1.0, 36.4, 35.5, 33.2, 32.8, 32.4, 22.5, 22.1, 21.2, 18.9, 18.8, 18.7,
1
4
1
5
+
+
5.5, 14.5, 10.2; HRMS (ESI ) m/z (M + H) calcd for C27
H
42
N
4
6
O S:
(Na
by flash chromatography (SiO
gradient elution) and then re-purified using a Chromatotron (2 mm SiO
GF rotor, 2-5% MeOH-CHCl gradient elution) to afford 15b (0.39
g, 43%) as a colorless oil: [R]22
MHz, CDCl ) δ 6.98 (s, 1 H), 6.71 (d, 1 H, NH, J ) 8.1 Hz), 6.56 (s,
2
SO
4
), and concentrated in vacuo. The residue was first purified
, 2.0 × 10 cm, 2-5% MeOH-CHCl
2
51.2903, found: 551.2916.
2
3
(3S,6R,7S,8S,12R,13S,15S)-15-Amino-12,13-epoxy-4,4,6,8,16-pen-
tamethyl-3,7-dihydroxy-17-(2-methyl-4-thiazolyl)-5-oxo-16(E)-hep-
tadecenoic acid (14a). To a 100 mL round-bottom flask charged with
3
1
D
3
-40.7 (c 1.0, CHCl ); H NMR (400
1
3a (0.11 g, 0.20 mmol) and PtO
EtOH (30 mL) under Ar. The resulting black mixture was stirred under
one atmosphere of H for 10 h, after which time the system was purged
with N , and an additional portion of PtO (55 mg, 50 wt %) was added.
Once again the reaction mixture was stirred under a blanket of H for
0 h. The system was then purged with N , and the reaction mixture
was filtered through a Celite pad eluting with CH Cl
(3 × 75 mL).
The solvents were removed in vacuo, and the residue was purified by
2
(55 mg, 50 wt %) was added absolute
3
1 H), 4.69-4.62 (m, 1 H), 4.28 (d, 1 H, J ) 6.6 Hz), 4.06-3.99 (m,
1 H), 3.86-3.81 (m, 1 H), 3.38-3.34 (m, 1 H), 2.82 (dd, 1 H, J ) 7.6,
5.7 Hz), 2.71 (s, 3 H), 2.58 (br s, 1 H, OH), 2.43 (dd, 1 H, J ) 8.7,
14.5 Hz), 2.34 (dd, 1 H, J ) 3.0, 14.5 Hz), 2.14 (s, 3 H), 2.05-1.92
(m, 2 H), 1.82-1.74 (m, 2 H), 1.69-1.40 (m, 5 H), 1.35 (s, 3 H), 1.28
(s, 3 H), 1.18 (d, 3 H, J ) 6.8 Hz), 1.14 (s, 3 H), 1.00 (d, 3 H, J ) 6.8
2
2
2
2
1
2
2
2
Hz); 13C NMR (100 MHz, CDCl
) δ 220.8, 170.9, 165.2, 152.7, 138.1,
3
flash chromatography (SiO
CHCl
as a foam: [R]
2
, 1.5 × 10 cm, 95:5.0:0.5 to 90:10:1.0
119.5, 116.5, 75.4, 73.9, 61.8, 61.4, 54.7, 52.7, 44.2, 40.4, 38.0, 32.1,
3
-MeOH-AcOH gradient elution) to afford 14a (92 mg, 89%)
31.8, 31.2, 24.0, 23.3, 21.9, 21.5, 19.5, 17.4, 17.3, 15.0; HRMS (FAB)
2
2
1
+
D
-19.0 (c 7.7, CH
3
OH); H NMR (400 MHz, CD
3
-
m/z 507.2892 (M + H) calcd for C27
H
42
N
2
O
5
S, found: 507.2875.
OD) δ 7.34 (s, 1 H), 6.65 (s, 1 H), 4.17 (dd, 1 H, J ) 9.9, 3.0 Hz),
“One-Pot” Procedure: A suspension of epothilone B (2, 5.06 g,
4
3
1
.02 (dd, 1 H, J ) 9.7, 5.1 Hz), 3.58 (dd, 1 H, J ) 7.2, 4.0 Hz), 3.33-
.30 (m, 1 H), 2.95-2.89 (m, 2 H), 2.70 (s, 3 H), 2.29 (dd, 1 H, J )
5.2, 3.0 Hz), 2.21-2.10 (m, 2 H), 1.96 (s, 3 H), 1.61-1.05 (a series
9.97 mmol) and sodium azide (0.777 g, 12.0 mmol) in a THF-H
2
O
mixture (5:1, 96 mL) was degassed for 20 min with nitrogen and treated
with a catalytic amount of tetrakis(triphenylphosphine) palladium(0)
(Lancaster, 1.2 g, 0.997 mmol) under Ar. The reaction mixture was
warmed to 45 °C for 20 min and cooled to 25 °C. The resulting bright
yellow, homogeneous solution was directly treated with a 1.0 M solution
of trimethylphosphine in THF (24.9 mL, 24.9 mmol) at 25 °C, and the
of multiplets, 8 H), 1.25 (s, 3 H), 1.12 (d, 3 H, J ) 6.8 Hz), 1.03 (s,
1
3
3
2
5
1
5
H), 0.95 (d, 3 H, J ) 6.8 Hz); C NMR (125 MHz, DMSO-d ) δ
6
18.8, 175.3, 164.1, 152.0, 137.6, 121.0, 117.5, 75.5, 72.0, 56.8, 55.5,
3.1, 51.7, 43.6, 37.8, 35.1, 30.8, 29.2, 28.0, 23.4, 21.2, 20.9, 18.7,
7.0, 14.6, 13.6; HRMS (ESI ) m/z (M + H) calcd for C26
11.2842, found: 511.2852.
+
+
37
H
42
N
2
6
O S:
reaction mixture was stirred for 2 h at ambient temperature.
The amino acid mixture was then diluted38 with MeCN-DMF (20:
1, 450 mL), cooled to 0 °C and treated with HOBt (1.35 g, 9.97 mmol)
followed by EDCI (4.78 g, 24.9 mmol). The reaction mixture was
warmed to 25 °C, stirred for 12 h, and extracted with EtOAc (4 × 200
(3S,6R,7S,8S,12R,13S,15S)-15-Amino-12,13-epoxy-4,4,6,8,12,16-
hexamethyl-3,7-dihydroxy-17-(2-methyl-4-thiazolyl)-5-oxo-16(E)-
heptadecenoic acid (14b). A solution of 13b (0.23 g, 0.42 mmol) in
THF (4.0 mL) was treated with H
supported triphenylphosphine (Aldrich, polystyrene cross-linked with
% DVB, 0.28 g, 0.84 mmol) at 25 °C. The resulting suspension was
2
O (23 µL, 1.25 mmol) and polymer
2
mL). The organic extracts were washed with H O (400 mL), saturated
aqueous NaHCO
and were dried (Na
initially purified by flash chromatography (SiO
MeOH-CHCl
) and then further purified by HPLC39 to afford 15b
3
(400 mL), and saturated aqueous NaCl (400 mL)
SO ), and concentrated in vacuo. The residue was
, 5.0 × 25 cm, 2%
2
2
4
stirred at 25 °C under Ar (32 h), filtered through a Celite pad, and
concentrated in vacuo. The residue was purified by flash chromatog-
raphy (SiO
AcOH gradient elution) to afford 14b (96 mg, 44%) as a colorless oil,
which was used directly in the macrolactamization reaction: HRMS
2
3
40
2
, 1.5 × 10 cm, 95:5.0:0.5 to 90:10:1.0 CHCl
3
-MeOH-
(1.2 g, 23% overall yield, >95% purity as judged by HPLC: retention
time ) 2.01 min) as a white lyopholizate which possessed the same
characterization data as listed above.
+
+
(
ESI ) m/z (M + H) calcd for C27
25.3000.
Alternatively, to a 100 mL round-bottom flask charged with 13b
1.9 g, 3.5 mmol) and PtO (0.95 g, 50 wt %) was added absolute
EtOH (30 mL) under Ar. The resulting black mixture was stirred under
one atmosphere of H for 10 h, after which time the system was purged
with N , and an additional portion of PtO (0.48 g, 25 wt %) was added.
Once again the reaction mixture was stirred under a blanket of H for
0 h. The system was then purged with N , and the reaction mixture
was filtered through a Celite pad eluting with CH Cl
H N
44 2
6
O S: 525.2998, found:
[1S-[1R*,3R*(E),7R*,10S*,11R*,12R*,16S*]]-7,11-Dihydroxy-
8,8,10,12-tetramethyl-3-[1-methyl-2-(2-methyl-4-thiazolyl)ethenyl]-
4-aza-17-oxabicyclo[14.1.0]heptadecane-5,9-dione (15a). By using
epothilone A as the starting material and following the “one-pot”
5
(
2
(
37) The reaction was monitored by HPLC: YMC S5 ODS (4.6 × 50
2
mm) column, 0 to 100% B, 4 min gradient with 1 min hold time, 4 mL/
min flow rate; A Solvent: 95% H2O-5% MeCN-0.1% AcOH, B
Solvent: 95% MeCN-5% H2O-0.1% AcOH. The retention times were
as follows: For azide acid intermediate 13b, Rt ) 3.35 min; For amino
acid intermediate 14b, Rt ) 2.18 min.
2
2
2
1
2
2
2
(3 × 75 mL).
(38) Alternatively, at this point the reaction mixture could be concentrated
The solvents were removed in vacuo, and the residue was purified as
described above to afford 14b (0.95 g, 53%).
Alternatively, a solution of 13b (20 mg, 36 µmol) in THF (0.4 mL)
was treated with triphenylphosphine (19 mg, 73 µmol) under Ar. The
reaction mixture was warmed to 45 °C, stirred for 14 h, and cooled to
in vacuo and the residual H2O removed by azeotroping with toluene (the
yield remained the same; however, the reaction time was reduced from 12
to 4 h).
(39) HPLC conditions: YMC S-15 ODS (50 × 500 mm) column, 40 to
100% B gradient (40 min), 50 mL/min flow rate; A Solvent: 95% H2O-
5
% MeCN, B Solvent: 95% MeCN-5% H2O. The appropriate fractions
2
5 °C. The resulting iminophosphorane was treated with ammonium
were concentrated in vacuo, and the residue was lyophilized from aqueous
acetonitrile.
hydroxide solution (28%, 0.1 mL), and once again the reaction mixture
was warmed to 45 °C. After 4 h, the volatiles were removed in vacuo,
and the residue was purified as described above to afford 14b (13 mg,
(
40) HPLC conditions: YMC S5 ODS (4.6 × 50 mm) column, 20 to
100% B, 4 min gradient with 1 min hold time, 4 mL/min flow rate; A
Solvent: 95% H2O-5% MeCN, B Solvent: 95% MeCN-5% H2O.
7
0%).

Synthesis of Epothilone-Lactams
J. Am. Chem. Soc., Vol. 122, No. 37, 2000 8897
procedure outlined above, 15a was obtained as a white lyopholizate
colorless oil and 12 (1.0 mg, 2.6%). For 20: [R]22
D
-42.4 (c 1.6,
2
2
1
1
(
18-25% yield): [R]
D
-41.4 (c 10, CHCl
3
); H NMR (400 MHz,
) δ 6.91 (d, 1 H, NH, J ) 8.1 Hz), 6.88 (s, 1 H), 6.45 (s, 1 H),
CHCl
3
); H NMR (400 MHz, CDCl
3
) δ 6.95 (s, 1 H), 6.49 (s, 1 H),
CDCl
4
3
6.00 (d, 1 H, NH, J ) 7.0 Hz), 5.54-5.51 (m, 1 H), 5.42-5.37 (m, 1
H), 4.42-4.36 (m, 1 H), 4.12-4.06 (m, 1 H), 3.76 (br s, 1 H), 3.59 (d,
1 H, J ) 5.5 Hz), 3.14 (q, 1 H, J ) 6.9 Hz), 2.92 (d, 1 H, J ) 2.6 Hz),
2.70 (s, 3 H), 2.49-2.40 (m, 3 H), 2.29 (dd, 1 H, J ) 15.0, 2.7 Hz),
2.05 (s, 3 H), 2.82-2.66 (m, 5 H), 1.31-1.28 (m, 3 H), 1.31 (s, 3 H),
.53 (dd, 1 H, J ) 12.5, 5.1 Hz), 4.40 (d, 1 H, J ) 6.9 Hz), 3.99-3.94
(
(
9
m, 1 H), 3.77 (d, 1 H, J ) 3.1 Hz), 3.26-3.19 (m, 1 H), 2.98-2.93
m, 1 H), 2.85-2.71 (m, 2 H), 2.62 (s, 3 H), 2.39 (dd, 1 H, J ) 15.2,
.5 Hz), 2.27 (dd, 1 H, J ) 15.2, 2.8 Hz), 2.03 (s, 3 H), 2.02-1.95 (m,
H), 1.87-1.78 (m, 1 H), 1.63-1.35 (a series of multiplets, 7 H),
.27 (s, 3 H), 1.09 (d, 3 H, J ) 7.0 Hz), 1.05 (s, 3 H), 0.91 (d, 3 H, J
1
3
1
1
)
1
3
1.19 (d, 3 H, J ) 6.9 Hz), 1.11 (s, 3 H), 1.00 (d, 3 H, J ) 7.0 Hz);
NMR (125 MHz, CDCl ) δ 220.7, 170.4, 164.7, 152.5, 139.3, 134.2,
125.2, 118.4, 115.5, 74.4, 73.2, 55.8, 53.0, 42.4, 40.0, 38.5, 33.0, 30.8,
C
3
1
3
3
7.0 Hz); C NMR (100 MHz, CDCl ) δ 221.7, 171.4, 165.6, 153.0,
+
+
38.8, 120.1, 116.8, 75.4, 74.8, 58.1, 55.3, 54.2, 52.9, 44.7, 40.7, 37.5,
27.6 (2C), 22.3, 20.2, 19.1, 16.9, 15.5, 13.7; HRMS (ESI ) m/z (M
+ H) calcd for C26 S: 477.2709, found: 477.2794.
+
1.9, 31.4, 27.9, 25.1, 22.6, 22.3, 19.9, 17.9, 17.7, 15.0; HRMS (ESI )
41 2 4
H N O
+
m/z (M + H) calcd for C26
H N O
40 2 5
S: 493.2736, found: 493.2746.
[4S-[4R*,7S*,8R*,9R*,15R*(E)]]-4,8-Dihydroxy-5,5,7,9,13-pen-
tamethyl-16-[1-methyl-2-(2-methyl-4-thiazolyl)ethenyl]-1-aza-13(Z)-
cyclohexadecene-2,6-dione (21). Tungsten hexachloride (0.76 g, 2.0
mmol) was dissolved in THF (20 mL) and the solution was cooled to
-78 °C. n-Butyllithium (1.6 M in hexane, 2.5 mL, 4.0 mmol) was
added in one portion and the reaction mixture was allowed to warm to
25 °C over 20 min. A 0.1 M solution of the prepared dark green tungsten
reagent (13.8 mL, 1.38 mmol) was added to 15b (0.35 g, 0.69 mmol)
in THF (2 mL) at 25 °C. The reaction mixture was stirred at ambient
temperature for 30 min, cooled to 0 °C, and then quenched with
[
1S-[1R*,3R*(E),7R*,10S*,11R*,12R*,16S*]]-7,11-Dihydroxy-
8
,8,10,12,16-pentamethyl-3-[1-methyl-2-(2-hydroxymethyl-4-thia-
zolyl)ethenyl]-4-aza-17-oxabicyclo[14.1.0]heptadecane-5,9-dione (17).
To a 10 mL round-bottom flask charged with (3S,6R,7S,8S,12R,13S,-
1
(
7
5S)-15-azido-12,13-epoxy-4,4,6,8,12,16-hexamethyl-3,7-dihydroxy-17-
2-hydroxymethyl-4-thiazolyl)-5-oxo-16(E)-heptadecenoic acid (40 mg,
1 µmol, cf. Supporting Information) and PtO (20 mg, 50 wt %) was
added absolute EtOH (3 mL) under Ar. The resulting black mixture
was stirred under one atmosphere of H for 10 h. The system was then
purged with N , and the reaction mixture was filtered through a nylon
2
2
saturated aqueous NaHCO
with H O (50 mL) and extracted with CH
combined organic extracts were dried (Na SO
trated in vacuo. The inorganics were removed by passing the residue
through a silica gel plug (eluting with 19:1 CHCl -MeOH). The eluent
3
(10 mL). The resulting mixture was diluted
Cl
(4 × 30 mL). The
), filtered, and concen-
2
membrane (washing with 25 mL of MeOH). The solvents were removed
in vacuo to afford the intermediate amino acid (29 mg, 76%) as a foam,
2
2
2
2
4
+
which was sufficiently pure to use in the next step. LCMS (ESI ):
+
5
41.3 (M + H).
3
3
9
was concentrated in vacuo and the residue was purified by HPLC to
A solution of the amino acid (29 mg, 54 µmol) in degassed DMF
21 mL) was treated with solid NaHCO (36 mg, 0.43 mmol) and
3
afford 21 (0.16 g, 47%) as a white foam along with minor E-isomer
(
2 (0.031 g, 9.1%). For 21: [R]22
-44.4 (c 5.5, CHCl
); H NMR
1
2
D
3
diphenylphosphoryl azide (46 µL, 0.21 mmol) at 0 °C under Ar. The
resulting suspension was stirred at 4 °C for 19 h, cooled to -40 °C,
diluted with 25 mL of pH 7 phosphate buffer (carefully adding such
that the internal temperature remained below -30 °C), and extracted
with EtOAc (4 × 10 mL). The organic extracts were washed with cold
(400 MHz, CDCl
3
) δ 6.95 (s, 1 H), 6.48 (s, 1 H), 5.99 (d, 1 H, NH, J
)
6.4 Hz), 5.13 (dd, 1 H, J ) 7.7, 7.5 Hz), 4.36-4.29 (m, 1 H), 4.07-
4
1
.01 (m, 1 H), 3.78-3.75 (m, 2 H), 3.14 (q, 1 H, J ) 6.9 Hz), 2.98 (d,
H, J ) 2.5 Hz), 2.70 (s, 3 H), 2.48-2.35 (m, 3 H), 2.23 (dd, 1 H, J
)
14.6, 2.6 Hz), 2.07 (s, 3 H), 2.07-2.00 (m, 1 H), 1.82-1.69 (m, 4
H), 1.69 (s, 3 H), 1.32 (s, 3 H), 1.32-1.21 (m, 2 H), 1.19 (d, 3 H, J )
.9 Hz), 1.12 (s, 3 H), 1.00 (d, 3 H, J ) 7.0 Hz); 13C NMR (125 MHz,
CDCl ) δ 221.1, 170.5, 164.7, 152.6, 139.6 (2C), 120.6, 118.5, 115.5,
1
0% aqueous LiCl (25 mL), dried (Na
2
SO
4
), and concentrated in vacuo.
GF rotor,
3
gradient elution) to afford 17 (9.1 mg, 34%) as
The residue was purified using a Chromatotron (1 mm SiO
2
a colorless oil: [R]
CDCl ) δ 7.13 (s, 1 H), 6.94 (d, 1 H, NH, J ) 7.6 Hz), 6.58 (s, 1 H),
2
6
-5% MeOH-CHCl
2
2
1
-23 (c 0.06, CHCl
3
); H NMR (400 MHz,
3
D
7
2
4.2, 73.9, 56.5, 52.9, 42.4, 40.3, 38.3, 32.7, 31.5, 31.3, 25.7, 23.2,
2.8, 20.0, 19.1, 16.6, 15.7, 13.5; HRMS (ESI ) m/z (M + H) calcd
S: 491.2944, found: 491.2926.
3
+
+
4
1
1
.94 (s, 2 H), 4.69-4.65 (m, 1 H), 4.08 (d, 1 H, J ) 7.3 Hz), 3.83 (s,
42 2 4
for C27H N O
H), 3.39-3.34 (m, 1 H), 2.83 (dd, 1 H, J ) 7.1, 6.4 Hz), 2.68 (br s,
H), 2.45 (dd, 1 H, J ) 14.8, 9.4 Hz), 2.34 (dd, 1 H, J ) 14.8, 2.5
For [4S-[4R*,7S*,8R*,9R*,15R*(E)]]-4,8-Dihydroxy-5,5,7,9,13-
pentamethyl-16-[1-methyl-2-(2-methyl-4-thiazolyl)ethenyl]-1-aza-13-
Hz), 2.14 (s, 3 H), 2.09-1.96 (m, 2 H), 1.77-1.40 (a series of
multiplets, 8 H), 1.37 (s, 3 H), 1.31 (s, 3 H), 1.20 (d, 3 H, J ) 6.9 Hz),
22
(
E)-cyclohexadecene-2,6-dione (22): white foam, [R]
D
-91.2 (c 2.8,
1
1
d
6
1
.14 (s, 3 H), 1.02 (d, 3 H, J ) 6.9 Hz); 1 C NMR (125 MHz, acetone-
6
3
CHCl ); H NMR (400 MHz, CDCl ) δ 6.94 (s, 1 H), 6.44 (s, 1 H),
3
3
6
.15 (d, 1 H, NH, J ) 8.0 Hz), 5.15-5.06 (m, 1 H), 4.49-4.46 (m, 1
) δ 217.9, 172.9, 170.8, 154.3, 140.3, 119.3, 117.2, 77.5, 71.7, 63.1,
2.6, 61.5, 54.6, 54.2, 45.8, 40.2, 37.4, 34.3, 34.0, 24.5, 22.9, 22.5,
8.9 (2C), 18.6, 17.1, 15.8; HRMS (ESI ) m/z (M + H) calcd for
S: 523.2842, found: 523.2850.
4S-[4R*,7S*,8R*,9R*,15R*(E)]]-4,8-Dihydroxy-5,5,7,9-tetramethyl-
6-[1-methyl-2-(2-methyl-4-thiazolyl)ethenyl]-1-aza-13(Z)-cyclo-
H), 4.24 (dd, 1 H, J ) 10.7, 2.0 Hz), 3.80 (d, 1 H, J ) 3.5 Hz), 3.58-
3.52 (m, 1 H), 3.43-3.39 (m, 1 H), 3.31-3.24 (m, 1 H), 2.70 (s, 3 H),
+
+
2
.57-2.49 (m, 1 H), 2.39-2.30 (m, 3 H), 2.22-2.17 (m, 1 H), 2.07
27 42 2 6
C H N O
(
s, 3 H), 1.97-1.88 (m, 1 H), 1.75-1.56 (m, 4 H), 1.63 (s, 3 H), 1.44-
[
1
0
1
4
1
4
.33 (m, 1 H), 1.33 (s, 3 H), 1.15 (d, 3 H, J ) 7.0 Hz), 1.04 (s, 3 H),
1
.99 (d, 3 H, J ) 6.8 Hz); 13C NMR (125 MHz, CDCl
) δ 219.6, 171.3,
3
hexadecene-2,6-dione (20). To a round-bottom flask was added
chopped pieces of magnesium turnings (0.12 g, 5.0 mmol). The reaction
vessel was flame-dried under vacuum and cooled under Ar. Bis-
64.6, 152.5, 139.4, 138.8, 120.3, 117.9, 114.9, 72.0 (2C), 55.1, 53.6,
3.4, 40.2, 39.9, 37.3, 32.8, 30.5, 24.6, 22.3, 19.0, 17.8, 17.2, 16.8,
+
+
5.9, 14.0; HRMS (ESI ) m/z (M + H) calcd for C27
42 2 4
H N O S:
(cyclopentadienyl)titanium dichloride (1.25 g, 5.0 mmol) was added
91.2944, found: 491.2943.
followed by anhydrous THF (25 mL). The stirring suspension was
evacuated with low vacuum, and the reaction flask was back-filled with
Ar. The red suspension became dark and eventually turned a homo-
geneous deep green after 1.5 h with nearly all of the Mg metal being
consumed. An aliquot (6.0 mL, 3.0 equiv) of the 0.2 M titanocene
reagent in THF was removed from the solution and cooled to -78 °C
under Ar. To this solution was added 15a (40 mg, 81 µmol). The
reaction mixture was stirred at -78 °C for 15 min and diluted with
Acknowledgment. We thank Sarah Traeger and Yolanda
Pan for their assistance in obtaining NMR data for select
compounds.
Supporting Information Available: Experimental proce-
dures for the preparation of compounds 7, 8, bis-TES 15b, and
1
13
EtOAc (10 mL) and saturated aqueous NaHCO
was extracted with EtOAc (3 × 10 mL), and the organic layers were
washed with brine (25 mL) and were dried (Na SO ) and concentrated
in vacuo. The residue was first purified by flash chromatography (SiO
.5 × 20 cm, 0-5% MeOH-CHCl gradient elution) to give a
separable 20:1 mixture of Z-20 and E-12. Further purification of the
geometric isomers using a Chromatotron (1 mm SiO GF rotor, 0-5%
MeOH-CHCl gradient elution) afforded 20 (18.4 mg, 47%) as a
3
(10 mL). The mixture
the precursor to compound 17, copies of the H and C NMR
1
1
spectra for select compounds, the 2D H- H NOESY NMR
1
1
2
4
spectrum of compound 20, the 1D H- H NOE NMR spectra
of compound 21, and X-ray crystallographic parameters for the
bis-TES ether derivative of compound 15b (PDF). This material
is available free of charge via the Internet at http://pubs.acs.org.
2
,
1
3
2
3
JA001899N