Transcript The Chemisty and Biology of Discodermolide

The Chemistry and Biology of
Discodermolide
HO
O
O
OH O
OH
O
NH2
OH
Yueheng Jiang
November 20, 2003
-1-
HO
Outline
O
O
OH O
OH
O
NH2
OH
 Discovery and Biological Activity
 Total Syntheses
 Structure-Activity Relationships (SAR)
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Discovery
 Isolated by Gunasekera and co-workers in 1990
from the Caribbean deep-sea sponge (Discodermia
dissoluta).
 0.002% w/w isolation yield (7 mg/434 g of
sponge).
 Found initially to have immunosuppressive and
antifungal activities.
 Further revealed to be a potent microtubule
stabilizer.
Gunasekera, S. P.; Gunasekera, M.; Longley, R. E.; Schulte, G. K. J. Org. Chem. 1991, 56, 1346.
Longley, R. E.; Caddigan, D.; Harmody, D.; Gunasekera, M.; Gunasekera, S. P.; Transplantation 1991, 52, 650.
ter Haar, E.; Kowalski, R. J.; Hamel, E.; Lin, C. M.; Longley, R. E.; Gunasekera, S. P.; Rosenkranz, H. S.; Day, B. W.
Biochemistry 1996, 35, 243.
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Microtubule-stabilizing agents
R
O
AcO
O
NH
OH
O
Ph
O
O
OH
HO
BzO
O
H
AcO
O
OH
OH
NH2
HO
HO
R = Ph
Taxol (BMS)
R = OtBu Taxotere (Aventis)
discodermolide
O
O
O
R
O
O
S
H
N
O
H
O
HO
O
NMe
N
N
O
H
OH O
R = H epothilone A
R = Me epothilone B
eleutherobin
H
OAc
O
OH
O
O
O
OMe
O
O
NMe
OH
OH
OH
CO2Me
sarcodictyin A
OH O
O
O
laulimalide
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Cytotoxicity
 Cytotoxic over a variety of cell lines (IC50 3-80 nM)
 More potent than Taxol
 Competitively inhibits the binding of Taxol to tubulin
 Active against multi-drug resistant (MDR) and Taxolresistant (Pgp mediated MDR) cell lines
ter Haar, E.; Kowalski, R. J.; Hamel, E.; Lin, C. M.; Longley, R. E.; Gunasekera, S. P.; Rosenkranz, H. S.; Day, B. W.
Biochemistry 1996, 35, 243.
Kowalski, R. J.; Giannakakou, P.; Gunasekera, S. P.; Longley, R. E.; Day, B. W.; Hamel, E.; Mol. Pharmacol. 1997,
52, 613.
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Mechanism of action




Promote tubulin polymerization in vitro
Stabilize microtubules against depolymerization
Interfere with Taxol-binding to microtubules
Induce microtubule bundles in cells
Consequences:
Interfere with proper formation of mitotic spindle
Cause arrest of cell cycle
Cell death by apoptosis
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Dynamic Equilibria of Tubulin-Microtubules
Heterodimer Formation
Initiation
Polymerzation/Elongation
(+) End (more grown)
Nucleation
center
5 nm
Microtubule
(-) End (less grown)
α-tubulin
~50 kD
KD = 10-5
(α.β)
β-tubulin
~50 kD
Taxol or
Discodermolide
Stabilized
Nucleation
centers
Stabilized
microtubules
Taxol or
Discodermolide
Nicolaou, K. C.; Roshangar, F.; Vourlounis, D. Angew. Chem. Int. Ed., 1998, 37, 2014.
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Microtubule Bundling
Influence of discodermolide on a
transformed mouse fibroblast.
Discodermolide induces
microtubule bundling (tubulin
appears green), which is clearly
seen in the pseudopodia and near
the nucleus (appears blue). As a
result of this microtubule
bundling, the cell is undergoing
apoptosis and fragmentation of
the nucleus can be seen.
Sasse, F. Current Biology, 2000, 10, R469.
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Unique Activities
 Discodermolide could not substitute for Taxol in a
Taxol-resistant cell line (A549-T12) that requires low
concentrations of Taxol for normal growth.
 Exhibits synergistic effects with Taxol in several
cultured cell lines (not observed with Taxol/epothilones
or Taxol/eleutherobin).
Martello, L. A.; McDaid, H. M.; Regl, D. L.; Yang, C. H.; Ment, D. T.; Pettus, R. R.; Kaufman, M. D.; Arimoto, H.;
Danishefsky, S. J.; Smith, A. B. III; Horwitz, S. B. Clin. Cancer Res. 2000, 6, 1978.
Giannakakou, P.; Fojo, T. Clin. Cancer Res. 2000, 6, 1613.
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Potential Candidate for Cancer Chemotherapy





Novel structure
Greater or comparable efficacy to Taxol
Poor substrate for P-glycoprotein (Pgp).
Synergistic effect in combination with Taxol
Greater water solubility (100-fold > Taxol)
Entered Phase I clinical trials in 2002 as a
chemotherapeutic agent for use against solid tumors.
Myles, D. C. Annual Reports in Med. Chem., 2002, 37, 125.
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 Taxol (semi-synthesis)
 Epothilone (fermentation)
 Discodermolide ( ???)
Total Synthesis of Discodermolide !
(-)-Discodermolide
(+)-Discodermolide
S. L. Schreiber (1993)
A. B. Smith (1995)
D. C. Myles (1997)
S. L. Schreiber (1996)
J. A. Marshall (1998)
A. B. Smith (1999, 2003)
I. Paterson (2000, 2002)
Novartis (2003)
D. C. Myles (2003)
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Selected Total Synthesis of (+)-Discodermolide
Schreiber
1st total synthesis of (+/-)-discodermolide
established absolute configuration
Smith
Delivered ~1 g of (+)-discodermolide (2nd generation)
Paterson
Novel approaches
Novartis
Meet supply needs for clinical studies by total synthesis
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Structure and conformation of (+) discodermolide
24
HO
17
19
7
O
O
21
OH
5
1
O
O
13
OH
NH2
OH
X-ray structure of discodermolide;
hydrogen atoms omitted for clarity
Paterson, I.; Florence, G. J. Eur. J. Org. Chem., 2003, 2193.
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General Features
24
HO 7
O
*
O
1
*
*
17
*
**
* * *
OH
O
O
13
OH
NH2
OH
* * *
OH
repeating anti, syn-stereotriad
divided into three fragments
of roughly equal complexity
HO
Introduction of two Z-alkenes
and terminal Z-diene
CO2Me
Roche ester
Fragment Coupling
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Strategy by Scheiber
enolate alkylation
Nozaki-Kishi coupling
HO
24
8
15
OH
O
O
1
5
OH
OH
13
(+)-discodermolide 1
O
O
NH2
I
8
1
O
A
24
15 OPiv
7 H
O
PhS
+
+
16
OTBS
OTBS
Still-Gennari
HWE olefination
B
OPMB
O
C
Roush crotylation
Roush crotylation
TBSO
Negishi coupling
OH
2
2
3
CO2Me
OH
TBSO
OH
TBSO
Roche ester
OH
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Wittig olefination
Strategy by Smith
2nd Generation
Negishi coupling
24
HO
O
19
15
9
O
OH
1
OH
O
Yamamoto
olefination
O
13
NH2
(+)-discodermolide 1
OH
Evans aldol
O
O
I
O
8 H
1
OTBS
+
14
PMBO
+
9
TBSO
OTBS
OTBS
A
CO2Me
OH
Roche ester
B
Zhao-Wittig
olefination
PMBO
N
OH
19
I 15
O
O
O
C
O
PMP
Common Precursor 2
Evans aldol
reaction
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Strategy by Paterson
1st generation
lithium aldol
boron aldol
24
HO 7
O
16
9
O
OH
1
O
(+)-discodermolide 1
O
13
OH
NH2
OH
boron aldol
boron aldol
6
MeO2C
A
TBSO
O
+
PMBO
ArO
O
16
9
Nozaki-Hiyama/
Peterson elimination
24
+
H 17
B
TBSO
Claisen [3,3]
BnO
C
O
O
PMBO
2
O
OBz
3
HO
CO2Me
Roche ester
boron aldol
PMB
O
O
EtO2C
4
OH
ethyl-(S)-lactate
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Synthesis of Stereotriad
by Schreiber
CO2iPr
O
B O
CO2iPr
Roche-ester
CO2iPr
H
(R,R)-(E)-crotylboronate
TBSO
OH
2
TBSO
O
B O
CO2iPr
(S,S)-(Z)-crotylboronate
TBSO
O
Roush crotylation
Subunit A & C
OH
3
Subunit B
Hung, D. T.; Nerenberg, J. B.; Schreiber, S. L. J. Am. Chem. Soc. 1996, 118, 11054.
Roush, W. R.; Palkowitz, A. D.; Ando, K. J. Am. Chem. Soc. 1990, 112, 6348.
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Synthesis of stereotriad
by Smith
Ph
HO
1 PMBO
CO2Me
O
N
NH
CCl3, PPTS
PMBO
2 LAH
3 Swern Oxidation
H
O
O
O
n-Bu2BOTf, Et3N
52-55%, 4 steps
3
Ph
PMBO
O
N
OH
4
O
O
MeNH(OMe).HCl
AlMe3, THF, 98%
PMBO
N
OH
O
O
Common Precursor 2
Smith, A. B. III; Beauchamp, T. J.; LaMarche, M. J.; Kaufman, M. D.; Qiu, Y.; Arimoto, H.; Jones, D. R.; Kobayashi,
K. J. Am. Chem. Soc. 2000, 122, 8654.
Smith, A. B. III; Kaufman, M. D.; Beauchamp, T. J.; LaMarche, M. J.; Arimoto, H. Org. Lett. 1999, 1, 1823.
Iversen, T.; Bundle, D. R. J. Chem. Soc., Chem. Commun. 1981, 1240.
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Synthesis of stereotriad
by Paterson
cHex2BCl
Et3N, Et2O BnO
BnO
LiBH4
O
86%
> 30:1 dr
+O - O
O
B
2
H
5
L
O
3
6
OH
O
6 steps
94%, > 30:1 dr PMBO
CHO
cHex2BCl, Me2NEt
> 97% ds
Subunit B
OH
OH
7
8
TBSO
OBz
OBz
4
OH
Me4NBH(OAc)3
95%, >30:1 dr PMBO
TBSO
O
Subunit A
OH
L
cHex2BCl, Et3N
methacrolein
PMBO
6 steps
BnO
PMBO
6 steps
Subunit C
O
10
Paterson, I.; Florence, G. J.; Gerlach, K.; Scott, J. P. Angew. Chem. Int. Ed. 2000, 39, 377.
Paterson, I.; Arnott, E. A. Tetrohedron Lett. 1998, 39, 7185.
Paterson, I.; Wallace, D. J.; Cowden, C. J. Synthesis 1998, 639.
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Synthesis of trisubstituted (Z)-alkene
Schreiber
KHMDS,
(CF3CH2O)2POCHMeCO2Me
O
CO2Me
14
TBSO
OTBS
93%, Z:E >20:1
(Still-Gennari HWE olefination)
Sch-5
TBSO
OTBS
Sch-8
Smith
PMBO
13 H
1 Ph3PEtI, n-BuLi, 0oC
2 0.1 M I2 in THF, -78oC
3 NaHMDS, -23oC
I
PMBO 9
14
o
TBSO
Sm-5
O
4 Add 6, -33 C
46%, 3 steps, Z:E = 8-17:1
(Zhao-Wittig olefination)
TBSO
Subunit B
Still, W. C.; Gennari, C. Tetrahedron Lett. 1983, 24, 4405.
Chen, J.; Wang, T.; Zhao, K. Tetrahedron Lett. 1994, 35, 2827.
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Synthesis of trisubstituted (Z)-alkene
Paterson
+
+
1 NaIO4, NaHCO3
MeOH, H2O, rt
9
PMBO
O
O
[3, 3]
O
O
2 DBU, xylene, reflux
82%
9
PhSe
Me
PMBO
Me
14
AcO
11
PMBO
Me
PMBO
O
Me
O
O
15
O
13
PMBO
3 steps, 81%
9
O
16
OTBS
Subunit B
Burton, J. W.; Clark, J. S.; Derrer, S.; Stork, T. C.; Bendall, J. G.; Holmes, A. B. J. Am. Chem. Soc. 1997, 119, 7483.
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Synthesis of terminal (Z)-diene
Schreiber
O
75%
TBSO
OH
I
NaHMDS
Ph3P+CH2I ITBSO
(Stork-Zhao
Wittig olefination)
Sch-4
Pd(PPh3)4
H2C CHZnBr
OPMB
80%
TBSO
Sch-6
OPMB
Sch-7
Smith
21 O
O
TrO
OPMB
TBS
Ph2P
76%, 2 steps Z:E = 12:1
O
TrO
9
OTBS
Sm-7
24
Ti(OiPr)4Li
then MeI
Yamamoto Olefination
OTBS
OPMB
TBS
Sm-8
1. Stock, G.; Zhao, K. Tetrahedron Lett. 1989, 30, 2173.
Ikeda, Y.; Ukai, J.; Ikeda, N.; Yamamoto, H. Tetrahedron 1987, 43, 723.
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Synthesis of terminal (Z)-diene
Paterson
SiMe3
TBSO
H
PMBO
O
11
LnCr
SiMe3
TBSO
PMBO
Nozaki-Hiyama
allylation
OH
12
KH, 98%
24
Z:E = > 20:1
Peterson
elimination
TBSO
OPMB
13
Takai, K.; Kuroda, T.; Nakatsukasa, S.; Oshima, K.; Nozaki, H. Tetrahedron Lett. 1985, 26, 5585.
Aicher, T. D.; Kishi, Y. Tetrahedron Lett. 1987, 28, 3463.
Ager, D. Org. React., 1990, 38, 1.
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Fragment coupling
Schreiber
I
PhS
8
7
O
15 OPiv
15
Br
O
OTBS
10
5 steps
1
OTBS
OTBS
OPMB
C
9
PhS
LDA (1.1 eq), THF, 0oC
65%
24
16
TBSO
24
16
O
1
OTBS
A
O
(Nozaki-Kishi coupling)
TBSO
15 OPiv
7
PhS
0.011% NiCl2/CrCl2, 65%
2:1 dr at C7
Subunit B
PhS
HO
O
OTBS
OTBS
H
1
15
O
O
OTBS
OPMB
11
OTBS
24
19
TBSO
16
LiN(SiMe2Ph)2 (5 eq)
MeI, 47%
PhS
O
O
OTBS
3:1 dr at C16
OTBS
12
Takai, K.; Kuroda, T.; Nakatsukasa, S.; Oshima, K.; Nozaki, H. Tetrahedron Lett. 1985, 26, 5585.
Aicher, T. D.; Kishi, Y. Tetrahedron Lett. 1987, 28, 3463.
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Fragment coupling
Smith
1 ZnCl2 (1.15 eq.)
2 t-BuLi (3.45 eq.), -78oC to rt
I 15
TBSO
O
19
3 Pd(PPh3)4 (0.05 eq.), 66%
O
O
14
I
PMP
14
PMBO
PMBO
O
TBS
OTBS
O
8 steps
PMP
subunit C
6
TBSO
subunit B
NaHMDS (0.95 eq.), THF, -20oC
O
-IPh +P
3
O
9
OPMB
TBS
O
O
OTBS
H
OTBS
10
OTBS
TBSO
O
24
8
8
69%
Z:E = 24:1
A
(1.05 eq.)
9
O
O
OPMB
TBS
OTBS
11
OTBS
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Fragment coupling
Paterson
O
PMBO
1 LiTMP, LiBr
2 LiAlH4, 62%
> 30:1 dr
O
HO
PMBO
17
9 steps
16
16
24
OH
OPMB
H 17
OTBS
OTBS
O
Subunit B
16
OPMB
Subunit C
24
HO
(+)-Ipc2BCl
O
7
7
87%, 5:1 at C7
H
O
OTBS
TBS
17
O
O
6
1
NH2
MeO2C
TBSO
MeO2C
O
O
O
OTBS
TBS
2
O
O
NH2
OTBS
18
Subunit A
Paterson, I.; Goodman, J. M.; Lister, M. A.; Schumann, R. C.; McClure, C. K.; Norcross, R. D.
Tetrahedron, 1990, 46, 4663.
Mulzer, J.; Berger, M. J. Am Chem. Soc. 1999, 121, 8393.
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Improvement
Smith
2nd Generation: ultrahigh pressure
Undesired intramolecular cyclization
1 PPh3, I2
2 PPh3, i-Pr2NEt
12.8 kbar, 6d
82%, 2 steps
9
OH
O
11
OPMB
TBS
OTBS
H
I
11
H
Me
R
I
-
+
OPMB
TBS
OTBS
R
R
H
OTBS
Me
H
O
9
H
H
R
-IPh +P
3
12.8 kbar =
12600 atm = 186,000 psi !!
Me
+
OTBS
Me
H
TBSO
TBSO
3rd Generation: improvement
Reduction of the steric bulk at C11
9
11
OH
OMOM
O
OPMB
TBS
1 PPh3, I2
2 PPh3, i-Pr2NEt, 100oC
70%, 2 steps
ambient pressure
-IPh +P
3
11
9
O
OPMB
TBS
OMOM
Smith, A.B. III; Kaufman, M. D.; Beauchamp, T. J.; LaMarche, M. J.; Arimoto, H. Org. Lett. 2003, 5, 4405.
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Paterson
1st generation
Problem
O
R2
R1
(+)-Ipc2BCl
7
87%, 5:1 at C7
H
O
O
TBS
OTBS
O
6
1
NH2
MeO2C
OTBS
O
Nu
H
H
Me
10
O
7
H H
10
O
TBS
2
TBSO
+
+
7
OTBS
MeO2C
17
O
24
reagent
c-Hex2BCl
(+)-Ipc2BCl
18a : 18b
7 :
1 :
1
5
yield
O
O
NH2
18
18a: R1 = OH, R2 = H
18b: R1 = H, R2 = OH (desired)
67%
87%
RL
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Paterson
2nd Generation
boron aldol
24
HO
O
O
100% substrate control
OH
6
O
O
5
1
OH
NH2
1
lithium aldol
OH
5 H
MeO2C
TBSO
A
+
24
16
O
11
6
O
OH
OH
alkylation
PMBO
ArO
O
NH2
O
16
9
+
24
H 15
14
O
OPMB
C
TBSO
B
O
Still-Gennari
HWE olefination
boron aldol
PMBO
PMBO
O
OH
reduced overall steps
from 42 to 35
OH
common precursor 19
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Paterson
2nd Generation
Improvement
cHex2BCl, Et3N
O
16
6
O
10
TBS
OTBS
O
O
NH2
1
5 H
MeO2C
O
TBSO
19
Subunit A
+
+
RL
10
H
L
H
B
O
O
64%, 20:1 dr at C5
L
HO
MeO2C
+
O
5
TBS
OTBS
O
R
24
8
OH
O
O
NH2
20
Paterson, I.; Delgado, O.; Florence, G. J.; Lyothier, I.; Scott, J. P.; Sereinig, N. S. Org. Lett. 2003, 5, 35.
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Strategy by Novartis
boron aldol (Paterson)
24
HO
O
O
18
9
OH
6
O
O
14
1
OH
Mg2+-chelated aldol (novel)
NH2
OH
O
6
N
MeO2C
+
O
O
18
9
OH
O
TBS
OH
MeO2C
9
18
O
PMBO
NH2
OH
OH
+
H 19
O
13
Negishi coupling
(Smith)
Nozaki-Hiyama/
Peterson elimination
(Paterson)
PMBO
H
HO
O
O
Zhao-Wittig Olefination
(Smith)
Roush Crotylation
(Schreiber)
PMBO
O
14
CO2Me
HO
CO2Me
H
O
commercially available
Francavilla, C.; Chen, W.; Kinder, F. R. Jr. Org. Lett., 2003, 5, 1233.
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Next Generation ???
Longest Linear Sequences
Steps
Yield
Schreiber
24
4.3%
Smith
24
6%
Myles
25
1.4%
Marshall
29
2.2%
Paterson
23
10.3%
Novartis
21
N.A.
Despite considerable synthetic efforts, there continues to be a pressing
demand for a more practical and scaleable total synthesis …
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SAR Summary
Antiproliferative Potencies (IC50, nM) Against A549 or MG63
Cells of Discodermolide (1), and Analogues 2-3e.
Discodermolide 1
(R = Me)
2 (R = H; 16-demethyl)
R
HO
17
16
7
O
O
11
3
OH
OH
OH O
A549
3.6
MG63
6
n.d.
10
O
NH2
Acetylated analogues:
3a (3-OAc)
3b (7-OAc)
3c (3,7-OAc)
3d (3,11-OAc)
3e (3,17-OAc)
3.8
0.8
0.8
164
524
N. Choy, Y. Shin, P. Q. Nguyen, D. P. Curran, R. Balanchandran, C. Madiraju, B. W. Day, J. Med. Chem., 2003,
46, 2846-2864.
Nerenberg, J. B.; Hung, D. T.; Schreiber, S. L. J. Am. Chem. Soc. 1996, 118, 11054.
Gunasekera, S. P.; Longley, R. E.; Isbrucker, R. A. J. Nat. Prod. 2001, 64, 171.
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SAR Summary
Antiproliferative Potencies (IC50, nM) Against A549 Cells of
Discodermolide (1), and Analogues 4a-4d.
HO
O
R
O
14
OH
OH O
O
7
O
O
NH2
OH
A549
4a (14-cis, demethyl)
7.8
4b (14-trans, demethyl) 485
OH
OH
O
O
NH2
3
A549
4c (3-dehydro, R = OH) 1.8
4d (3-dehydro, R = H)
11.4
Martello, L. A.; LaMarche, M. J.; He, L.; Beauchamp, T. J.; Smith, A. B. III; Horwitz, S. B. Chem. Biol. 2001, 8,
843.
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Summary
 Discodermolide, a marine natural product, shares the same
microtubule-stabilizing mechanism as Taxol and has a
promising anticancer profile.
 However, the supply problem is still hampering further
biological and SAR studies. To date, total synthesis is the
only economical means of providing useful quantities of
Discodermolide. Despite considerable synthetic efforts,
there continues to be a demand for a more practical and
scaleable total synthesis.
-36-
Acknowledgements
Prof. Stephen F. Nelsen, Prof. Steven D. Burke
Brian Lucas, Andy Hawk
Dr. Jian Hong, Dr. Lei Jiang
Dr. Stuart Rosenblum, Dr. Michael Wong, Dr. Ron Kuang
Lei Chen, Dr. Gopinadhan Anilkumar
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