Total synthesis and stereochemical reassignment of (+)-dolastatin 19

Article abstract of DOI:10.1021/ol060609q

A revised configurational assignment for the cytotoxic marine macrolide dolastatin 19 is proposed and validated by total synthesis. Key features of the route include an asymmetric vinylogous aldol reaction to install the isolated C13 stereocenter and (E)-

Full text of DOI:10.1021/ol060609q

ORGANIC
LETTERS
2006
Vol. 8, No. 10
2131-2134
Total Synthesis and Stereochemical
Reassignment of (
)-Dolastatin 19†
+
Ian Paterson,* Alison D. Findlay, and Gordon J. Florence
UniVersity Chemical Laboratory, Lensfield Road, Cambridge CB2 1EW, U.K.
ip100@cam.ac.uk
Received March 13, 2006
ABSTRACT
A revised configurational assignment for the cytotoxic marine macrolide dolastatin 19 is proposed and validated by total synthesis. Key
features of the route include an asymmetric vinylogous aldol reaction to install the isolated C13 stereocenter and (E)-trisubstituted alkene, two
sequential 1,4-syn boron-mediated aldol reactions, and a Mukaiyama glycosylation to append the L-rhamnose-derived pyranoside.
Dolastatin 19 is a polyketide of presumed cyanobacterial
origin, first isolated in 2004 by Pettit and co-workers from
the sea hare Dolabella auricularia collected in the Gulf of
California.1 Initial biological screening indicated significant
cancer cell growth inhibitory activity (GI50 values of 0.72
µg/mL and 0.76 µg/mL for breast MCF-7 and colon
KM20L2 cell lines, respectively). However, further biological
evaluation of dolastatin 19, including elucidation of the
mechanism of action, was precluded by the low isolation
yield (0.5 mg was obtained from 600 kg of D. auricularia),
inspiring our efforts toward the realization of a total
synthesis.2
14-membered glycosylated macrolide (1, Figure 1), contain-
ing a six-membered cyclic hemiacetal, appended with a
bromine-substituted (E,E)-diene and a 2,4-di-O-methyl-L-
rhamnopyranoside. It shares several structural features with
the 14-membered macrolides auriside A (2) and B (3)3 (also
isolated from D. auricularia) as well as with callipeltoside
A (4),4 which have previously been synthesized in our
laboratory.2a,5,6 However, the assignment of relative stereo-
chemistry for dolastatin 19, as shown in 1, appears unusual
on the basis of the anticipated common bacterial biogenesis
of these related polyketides.7 We now report the total
synthesis and revised configurational assignment of dolastatin
19.
Following extensive spectroscopic analysis of dolastatin
19 by the Pettit group,1 the structure was determined as a
At the outset, we considered the preferred conformation
of the related marine macrolides shown in Figure 1. Con-
† Dedicated to Professor K. C. Nicolaou on the occasion of his 60th
birthday.
(1) Pettit, G. R.; Xu, J.-P.; Doubek, D. L.; Chapuis, J.-C.; Schmidt, J.
M. J. Nat. Prod. 2004, 67, 1252.
(3) Sone, H.; Kigoshi, H.; Yamada, K. J. Org. Chem. 1996, 61, 8956.
(4) Zampella, A.; D’Auria, M. V.; Minale, L.; Debitus, C.; Roussakis,
C. J. Am. Chem. Soc. 1996, 118, 11085.
(2) For recent reviews on bioactive marine natural products, see: (a)
Yeung, K.-S.; Paterson, I. Chem. ReV. 2005, 105, 4237. (b) Blunt, J. W.;
Copp, B. R.; Munro, M. H. G.; Northcote, P. T.; Prinsep, M. R. Nat. Prod.
Rep. 2005, 22, 15. (c) Nicholas, G. M.; Phillips, A. J. Nat. Prod. Rep. 2005,
22, 144. (d) Newman, D. J.; Cragg, G. M. J. Nat. Prod. 2004, 67, 1216.
(5) (a) Paterson, I.; Davies, R. D. M.; Heimann, A.; Marquez, R.; Meyer,
A. Org. Lett. 2003, 5, 4477. (b) Paterson, I.; Davies, R. D. M.; Marquez,
R. Angew. Chem., Int. Ed. 2001, 40, 603.
(6) Paterson, I.; Florence, G. J.; Heimann, A. C.; Mackay, A. C. Angew.
Chem., Int. Ed. 2005, 44, 1130.
10.1021/ol060609q CCC: $33.50
© 2006 American Chemical Society
Published on Web 04/13/2006

Scheme 1
Figure 1. Dolastatin 19 and related marine-derived macrolides.
sistent with detailed 1H NMR spectral analysis, callipeltoside
and the aurisides share a similar diamond lattice arrangement
for the macrolide with the six-membered hemiacetal ring
adopting a chair conformation, facilitating a stabilizing
hydrogen bond between the axial C3-OH and the lactone
carbonyl oxygen and minimizing steric interactions. In
contrast, the calculated lowest-energy conformation for 1,
the originally proposed structure for dolastatin 19, predicts
a boat conformation for the pyran ring8 and distortion of the
macrolactone relative to that in callipeltoside and the
aurisides. This conformational analysis prompted us to
propose the stereochemical inversion of the C5-C7 array
and the C13 carbinol in 1, giving the putative structure 5
(Scheme 1) for dolastatin 19, which fits better with a
common biogenesis for this family of cytotoxic polyketides.
As outlined in Scheme 1, our synthetic strategy for 5, the
proposed revised structure for dolastatin 19, envisaged an
R-selective glycosylation of the aglycon with L-rhamnose-
derived fluorosugar 6, following suitable elaboration steps
and macrolactonization of protected linear precursor 7. The
repeating 1,4-syn relationship found in 7 across the C2-C5
and C6-C9 arrays would, in turn, arise from two consecutive
boron-mediated aldol reactions with R-chiral methyl ketone
89 to chain extend aldehyde 9. Application of an asymmetric
*Denotes the stereocenter inverted in configuration relative to
the original assignment, i.e., C5, C6, C7, and C13.
vinylogous Mukaiyama10 aldol reaction would then allow
for the simultaneous introduction of the remote C13 stereo-
center and (E)-trisubstituted alkene in 9.
As shown in Scheme 2, preparation of the C9-C17
aldehyde 9 began with the vinylogous aldol reaction between
the silyl dienolate 1011 and the (E,E)-bromodienal 11.12
Following our standard conditions,5a,6 treatment of 11 with
the chiral Lewis acid promoter (R)-BINOL-Ti(OiPr)2 (gener-
ated in situ from (R)-BINOL and Ti(OiPr)4) in THF at -78
°C, followed by addition of 10, provided the adduct 12 in
93% yield and 94% ee. TBS ether formation, DIBAL
reduction, and subsequent reoxidation of the resulting
alcohol with MnO2 gave aldehyde 9 (75%). The stage was
now set for the first 1,4-syn aldol reaction with methyl ketone
8. Building on experience gained in our synthesis of
callipeltoside,5 we chose the 3,4-dimethoxybenzyl (DMB)
ether13 in 814 to alleviate later chemoselectivity complications
arising from competitive C13 oxidation by DDQ. Enolization
of 8 with (+)-Ipc2BCl/Et3N, followed by addition of aldehyde
9, generated the expected 1,4-syn aldol adduct 13 in 88%
yield and >95:5 dr.9
(7) Celmer’s configurational model, as developed for macrolide antibiot-
ics, may be usefully extended to predicting stereochemical homology in
marine-derived secondary metabolites produced by polyketide synthases
in genetically related bacteria. Celmer, W. D. J. Am. Chem. Soc. 1965, 87,
1801.
(8) The boat conformation of the pyran ring in dolastatin 19 was proposed
by the Pettit group from interpretation of NOE correlations in 2D-ROESY
experiments, leading to their stereochemical assignment in structure 1 (ref
1).
(10) Sato, M.; Sunami, S.; Sugita, Y.; Kaneko, C. Heterocycles 1995,
41, 1435.
(11) Savard, J.; Brassard, P. Tetrahedron 1984, 40, 3455.
(12) (a) Becher, J. Synthesis 1980, 589. (b) Soullez, D.; Ple´, G.; Duhamel,
L. J. Chem. Soc., Perkin Trans. 1 1997, 1639.
(13) Oikawa, Y.; Tanaka, T.; Horita, K.; Yoshioka, T.; Yonemitsu, O.
Tetrahedron Lett. 1984, 25, 5393.
(14) Methyl ketone 8 was prepared from methyl (R)-3-hydroxy-2-
methylpropionate: (i) DMBO(CCl3)CdNH, PPTS, CH2Cl2; (ii) (MeO)NHMe‚
(9) (a) Paterson, I.; Goodman, J. M.; Isaka, M. Tetrahedron Lett. 1989,
30, 7121. (b) Paterson, I.; Oballa, R. M. Tetrahedron Lett. 1997, 38, 8241.
i
HCl, PrMgCl, THF, -20 °C; (iii) MeMgCl, THF, 0 °C.
2132
Org. Lett., Vol. 8, No. 10, 2006

ether at C13. Employing the optimized conditions from our
callipeltoside synthesis,5 treatment of DMB ether 16 with
DDQ in CH2Cl2 and pH 7 buffer (60 °C, 10 min) gave
alcohol 17 in 77% yield (99% based on recovered 16).
Conversion into the C5-C17 aldehyde 14 was then achieved
by Dess-Martin oxidation of 17, in preparation for the
second 1,4-syn aldol reaction with 8. Enolization of methyl
ketone 8 with c-Hex2BCl/Et3N and reaction with 14 pro-
ceeded under efficient substrate control, imparted from both
aldol partners, to give the expected Felkin-Anh adduct 7 in
excellent yield and selectivity (89%, >95:5 dr).
Scheme 2
With the full C1-C17 carbon backbone in place, attention
was now directed to assembling the putative aglycon 18 of
dolastatin 19. Treatment of 7 with PPTS and trimethyl
orthoformate in MeOH led to removal of the TES ether at
C7, with concomitant cyclization and methyl acetal forma-
tion, giving 19 (70%) after TBS protection of the C5
hydroxyl (TBSOTf/2,6-lutidine).16 Cleavage of the DMB
ether in 19 was now required. However, employing the
conditions used successfully for 16 led only to poor conver-
sion, and prolonged exposure of 19 to DDQ led to hydrolysis
of the methyl acetal and C13 oxidation. After extensive
optimization, treatment of 19 with DDQ (5 equiv) in pH 9
buffer and CH2Cl2 (0 °C, 10 min) provided the desired
primary alcohol 20 in 53% yield. Next, oxidation of 20 with
Dess-Martin periodinane gave the intermediate aldehyde,
which was readily oxidized with NaClO2 to carboxylic acid
21 (96%). Selective cleavage of the allylic TBS ether at C13
with TBAF then provided the required seco-acid 22 (98%),
in preparation for the key macrolactonization step. Under
Yamaguchi conditions,17 22 was converted into the 14-
membered macrolide 23 in 64% yield. Cleavage of the
remaining silyl group at C5 with TBAF, followed by acid
hydrolysis of the methyl acetal 24, provided the putative
With aldol adduct 13 in hand, elaboration to C5-C17
aldehyde 14 was required (Scheme 3). Evans-Tischenko 1,3-
anti reduction15 of â-hydroxy ketone 13 using SmI2 and
propionaldehyde provided the intermediate hydroxy ester,
which was readily converted into TES ether 15 (72% over
two steps, >95:5 dr). Reductive cleavage of the propionate
ester in 15 by DIBAL and O-methylation of the resulting
C9 alcohol with Meerwein’s salt provided 16 (84%).
Oxidative cleavage of the primary DMB ether was now
required in the presence of the potentially labile allylic TBS
Scheme 3
Org. Lett., Vol. 8, No. 10, 2006
2133

1
aglycon 18 (67%). At this stage, comparison of the H and
conditions (SnCl2/AgClO4),19 followed by cleavage of the
residual TBS ether with HF‚pyridine, afforded 5 in 39% yield
with complete R-selectivity. Gratifyingly, the spectroscopic
data18 obtained for 5 (1H and 13C NMR, MS) and the
measured specific rotation, [R]D20 +2.2 (c 0.18, MeOH) cf.
+7.5 (c 0.04, MeOH), correlated fully with that of natural
dolastatin 19.1 Together with the biological data obtained,20
this provided convincing evidence that the relative and
absolute configuration of (+)-dolastatin 19 is that indicated
in structure 5 (i.e., 2S, 3S, 5S, 6R, 7S, 9S, 13R in the aglycon).
In conclusion, we have resolved the stereochemical
ambiguities surrounding the structure of the cytotoxic mac-
rolide (+)-dolastatin 19, isolated from the sea hare Dolabella
auricularia, by completing the first total synthesis (23 steps,
1.7% overall yield), and have enabled further biological
studies.20 The present work further demonstrates the versatil-
ity of our aldol methodology for providing synthetic access
to rare bioactive marine polyketides, as well as for establish-
ing their full stereochemistry.21
13C NMR spectra of aglycon 1818 with the reported data for
the macrolide region of dolastatin 19 showed close agree-
ment, providing an early indication of the likely validity of
our proposed stereochemical reassignment.
Completion of the synthesis of dolastatin 19 (Scheme 4)
required the stereocontrolled glycosylation of the aglycon
Scheme 4
Acknowledgment. We thank the EPSRC (EP/C541677/
1), Emmanuel College, Cambridge (Research Fellowship to
G.J.F.), and Merck Research Laboratories for support;
Professor G. R. Pettit (Arizona State University) for copies
of NMR spectra for natural dolastatin 19; Dr. Carmen Cuevas
(PharmaMar) for providing cytotoxicity assays; and Dr.
Jeremy Scott (Merck Sharp & Dohme Research Laboratories,
Hoddesdon) and Robert Paton (Cambridge) for helpful
discussions.
Supporting Information Available: Representative ex-
perimental details and spectroscopic data for all new
1
compounds and copies of H and 13C NMR spectra for
synthetic and natural dolastatin 19 with full assignments. This
material is available free of charge via the Internet at
http://pubs.acs.org.
OL060609Q
18 with rhamnose-derived fluorosugar 6, previously utilized
for auriside A.6 Coupling of 18 and 6 under Mukaiyama
(18) See the Supporting Information.
(19) Mukaiyama, T.; Murai, Y.; Shoda, S. Chem. Lett. 1981, 431.
(20) GI50 values measured for 5 against representative cancer cell lines:
0.88 µM for HT29 (colon), 1.04 µM for NSCLC (lung), 1.20 µM for MDA-
MB-231 (breast). These biological results are consistent with that reported
for dolastatin 19 (ref 1).
(21) For a review on the role of total synthesis in the reassignment of
natural product structures, see: Nicolaou, K. C.; Snyder, S. A. Angew.
Chem., Int. Ed. 2005, 44, 1012.
(15) Evans, D. A.; Hoveyda, A. H. J. Am. Chem. Soc. 1990, 112, 6447.
(16) The stereochemistry of 19 was confirmed by 1H NMR analysis,
with irradiation of H5 providing diagnostic NOE enhancements with the
C3-OMe and C6-Me, consistent with their 1,3-diaxial and cis orientations,
respectively.
(17) Inanaga, J.; Hirata, K.; Saeki, H.; Katsuki, T.; Yamaguchi, M. Bull.
Chem. Soc. Jpn. 1979, 52, 1989.
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Org. Lett., Vol. 8, No. 10, 2006