Total synthesis of a potent hybrid of the anticancer natural products dictyostatin and discodermolide

Article abstract of DOI:10.1039/b811575c

A potent dictyostatin-discodermolide hybrid was designed and synthesised; it showed enhanced cell growth inhibitory activity relative to discodermolide in four human cancer cell lines including the Taxol-resistant NCI/ADR-Res cell line. The Royal Society of Chemistry.

Full text of DOI:10.1039/b811575c

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Total synthesis of a potent hybrid of the anticancer natural products
dictyostatin and discodermolidewz
Ian Paterson,*a Guy J. Naylora and Amy E. Wrightb
Received (in Cambridge, UK) 10th July 2008, Accepted 30th July 2008
First published as an Advance Article on the web 28th August 2008
DOI: 10.1039/b811575c
A potent dictyostatin–discodermolide hybrid was designed and
synthesised; it showed enhanced cell growth inhibitory activity
relative to discodermolide in four human cancer cell lines
including the Taxol-resistant NCI/ADR-Res cell line.
there appears to be minimal spatial correlation between the
d-lactone of discodermolide and the dienoate of dictyostatin.
In addition, the AutoDock-derived model for tubulin binding
indicates they both occupy the taxoid site and share similar
interactions with the protein residues of the receptor.
Discodermolide1 (1, Fig. 1) and dictyostatin2,3 (2) are marine
sponge-derived antimitotic polyketides which exhibit potent
growth inhibition against a wide range of human cancer cell
lines, including multidrug-resistant cancer cells.4 Functioning
by the same microtubule-stabilising mechanism as Taxol, they
cause an accumulation of cells in the G2/M phase and sub-
sequent cell death via apoptosis. In a comparison of the
tubulin polymerising ability of natural products that bind at
the taxoid site on b-tubulin, discodermolide and dictyostatin
were found to be the most potent, with dictyostatin displaying
the strongest assembly inducing abilities.3a Notably, Novartis
undertook the large-scale total synthesis of discodermolide4a
and advanced it into clinical trials as a novel anticancer agent.
Recently, the bioactive conformations of dictyostatin and
discodermolide were elucidated using a combination of NMR
analysis, molecular modelling and docking studies.5 The over-
lay of these tubulin-bound structures (Fig. 1) revealed some
striking conformational similarities. The overlap is most pro-
nounced from the common terminal diene moiety through
to C9 on dictyostatin and C7 on discodermolide. Whereas,
With this information in hand and building on our previous
synthetic work,6,7 we sought to rationally design an active
hybrid of these two anticancer natural products. Herein, we
report an efficient total synthesis of the novel dictyostatin–
discodermolide hybrid 3 (Fig. 1) and disclose that it has
enhanced (low nanomolar) antiproliferative activity in vitro
relative to discodermolide, which is retained against the Taxol-
resistant NCI/ADR-Res cell line. As part of efforts to develop
a practical dictyostatin synthesis, we also describe an im-
proved route to a common C4–C10 intermediate exploiting
our boron aldol methodology.
The structural similarities between dictyostatin and disco-
dermolide coincide with the regions of best overlap on the
tubulin-bound conformers. However, the superior binding
ability of dictyostatin could potentially be aided by the
dienoate, occupying a region where the polyketide-derived
structures differ. Hence, in our designed hybrid 3, the stereo-
chemistry and substitution from C8 to C26 are identical to
those of discodermolide (lacking the carbamate), while the C1
to C7 region, incorporating the dienoate moiety, and
22-membered macrolactone are dictyostatin8 derived. Our
a University Chemical Laboratory, Lensfield Road, Cambridge, UK
CB2 1EW. E-mail: ip100@cam.ac.uk; Fax: +44 (0) 1223 336362;
Tel: +44 (0)1223 336407
retrosynthetic analysis (Scheme 1) for
3 envisaged a
cross-coupling–macrolactonisation endgame, preceded by
installation of the (10Z)-alkene by a complex Still–Gennari
olefination between aldehyde 47 and b-ketophosphonate 5.6
Synthesis of phosphonate 5 was initiated by enolisation of
the lactate-derived ketone 79 with c-Hex2BCl–Me2NEt
(Scheme 2) and addition of aldehyde 8, affording the anti
b Harbor Branch Oceanographic Institution at Florida Atlantic
University, 5600 US 1 North, Ft. Pierce, FL 34946, USA
w Dedicated to Professor Andrew B. Holmes on the occasion of his
65th birthday.
z Electronic supplementary information (ESI) available: Characterisa-
tion data for new compounds. See DOI: 10.1039/b811575c
Fig. 1 Structures of discodermolide (1), dictyostatin (2) and designed hybrid analogue 3. Overlay of the NMR-derived bioactive conformations of
discodermolide (green) and dictyostatin (blue) at the taxoid binding site on b-tubulin.
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Scheme 1 Retrosynthesis of dictyostatin–discodermolide hybrid 3.
adduct 9 cleanly (89%, 497 : 3 dr) via the bicyclic aldol
transition state shown. Following formation of the PMB ether,
a one-pot reduction and hydrolysis sequence gave the corre-
sponding 1,2-diol (84%). Periodate cleavage revealed the
aldehyde 10, which was then converted into the b-ketophos-
phonate 5 (52%) via 11 and 12 by a similar procedure to that
developed previously.6,8a In our 2004 dictyostatin total syn-
thesis,6,10 the requisite anti C6/C7 stereocentres were configured
through a Brown crotylation reaction. In comparison, this new
aldol-based route to 5 was found to be more readily scaleable
and achieved enhanced stereoselectivity.
With both 4 (a key intermediate in our discodermolide
synthesis)4a,7 and 5 in hand (Scheme 3), the pivotal Still–
Gennari olefination11 was performed on a gram scale to afford
the desired (Z)-enone 13 in 67% isolated yield with good
selectivity (6.9 : 1 Z/E). Oxidative cleavage of the PMB ether
at C7 afforded the corresponding b-hydroxy ketone in readi-
ness for reduction to install the C9 stereocentre. Following
related studies,12a use of (R)-CBS13 and BH3ꢁTHF generated
the required 1,3-anti diol cleanly (495 : 5 dr), which was
transformed into acetonide 14 (70% from 13).
Scheme 3 Completion of the synthesis of 3 and 15.
The endgame commenced with a copper-mediated Stille
cross-coupling14 between vinyl iodide 14 and stannane 6 to
install the (2Z,4E)-dienoate. Macrolactonisation under
Yamaguchi conditions then smoothly afforded the protected
macrolactone (74% from 14). While HFꢁpyridine gave incon-
sistent results, global deprotection with HCl–MeOH (1 : 3)
proved more reliable, generating the hybrid 3 (72%) with
negligible translactonisation onto the C19 hydroxyl. In order
Scheme 2 Synthesis of b-ketophosphonate 5.
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Table 1 Human cancer cell growth inhibitory properties of hybrid 3
and acetonide derivative 15 relative to discodermolide (1),8a dictyos-
tatin (2) and Taxol, as determined by MTT metabolism after 72 h
exposure to the test agent
In conclusion, we have completed an efficient total synthesis
of the most potent cytotoxic hybrid of dictyostatin and
discodermolide reported to date.12 We attribute the enhanced
cell growth inhibitory activity of 3 relative to discodermolide
to the more constrained macrocyclic structure and the dictyos-
tatin-like C1–C7 region playing a significant role in binding to
tubulin. Efforts are ongoing to further probe the pharmaco-
phore and anticancer profiles of these fascinating marine
natural products and their hybrids.
Cytotoxicity IC50/nM
PANC-1
AsPC-1
DLD-1
NCI/ADR-Res
1
2
59 ꢂ 34
98 ꢂ 34
29 ꢂ 8
160 ꢂ 34
4.2 ꢂ 0.5
9.9 ꢂ 1.3
12.9 ꢂ 2.0
6.2 ꢂ 0.6
150 ꢂ 32
33.9 ꢂ 6.4
2.2 ꢂ 0.5
22.4 ꢂ 1.4
5.9 ꢂ 1.1
6.6 ꢂ 0.4
Taxol
3
15
1260 ꢂ 140
66.4 ꢂ 15.2
Financial support was provided by the EPSRC, Merck
Research Laboratories and NIH Grant no. CA-93455. We
thank Dr J. Fernando Dıaz (CSIC, Madrid) for providing 3D
´
4860 ꢂ 150 4850 ꢂ 450
2350 ꢂ 180 2930 ꢂ 300
structures, Dr Stuart Mickel (Novartis) for chemicals, Nicola
Gardner (Cambridge) for helpful discussions, Ms P. Linley
for cytotoxicity assays and Ms T. Pitts (HBOI) for immuno-
fluorescence and flow cytometry assays.
Notes and references
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Schmidt, J. Chem. Soc., Chem. Commun., 1994, 1111; (b) R. A.
Isbrucker, J. Cummins, S. A. Pomponi, R. E. Longley and A. E.
Wright, Biochem. Pharmacol., 2003, 66, 75; (c) I. Paterson, R.
Britton, O. Delgado and A. E. Wright, Chem. Commun., 2004, 632.
3 (a) R. M. Buey, I. Barasoain, E. Jackson, A. Meyer, P.
Giannakakou, I. Paterson, S. Mooberry, J. M. Andreu and J. F.
Fig. 2 Immunofluorescence images of PANC-1 cells stained with
anti-a-tubulin (green) and propidium iodide (red) and observed by
confocal microscopy. Cells were exposed to 100 nM concentrations of
dictyostatin (left image) and analogue 3 (right image). Typical dense
intracellular bundling of microtubules (green) can be seen around the
nuclei (red) in both images.
´
Dıaz, Chem. Biol., 2005, 12, 1269; (b) C. Madiraju, M. C. Edler,
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A. Vogt, Y. Shin, J. H. Fournier, Y. Fukui, A. M. Bruckner,
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D. P. Curran and B. W. Day, Biochemistry, 2005, 44, 15053.
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J. Am. Chem. Soc., 2001, 123, 9535.
8 For representative dictyostatin analogues, see: (a) I. Paterson, N. M.
Gardner, K. P. Poullennec and A. E. Wright, Bioorg. Med. Chem.
Lett., 2007, 17, 2443; (b) I. Paterson, N. M. Gardner, K. P. Poullennec
and A. E. Wright, J. Nat. Prod., 2008, 71, 364; (c) W. H. Jung,
C. Harrison, Y. Shin, J. H. Fournier, R. Balachandran, B. S. Raccor,
R. P. Sikorski, A. Vogt, D. P. Curran and B. W. Day, J. Med. Chem.,
2007, 50, 2951; (d) B. S. Raccors, A. Vogt, R. P. Sikorski, C. Madiraju,
R. Balachandran, K. Montgomery, Y. Shin, Y. Fukui, W. H. Jung,
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10 A Brown crotylation was also used by the Curran group, see: Y.
to probe the contribution of the C7,C9-diol to the pharmaco-
phore, 3 was treated with 2,2-dimethoxypropane–PPTS to
reinstate the acetonide in 15.
Following HPLC purification, the antiproliferative activities
of 3 and 15 were evaluated in vitro against four human cancer
cell lines (Table 1): PANC-1 (pancreatic), AsPC-1 (pancrea-
tic), DLD-1 (colon), and NCI/ADR-Res (Taxol-resistant
ovarian). Importantly, hybrid 3 demonstrated low nanomolar
cell growth inhibitory activity that was intermediate between
that measured for discodermolide (1)8a and dictyostatin (2)
and similarly maintained this potent activity against the NCI/
ADR-Res cell line (IC50 = 66.4 ꢂ 15.2 nM), where the
overexpression of a P-glycoprotein drug efflux pump in the
cell membrane gives rise to Taxol resistance. As with dictyos-
tatin and discodermolide, hybrid 3 led to an accumulation of
cells at the G2/M phase. In contrast, acetonide 15 was found
to have greatly reduced cytotoxicity (low micromolar), sug-
gesting that one or both of the C7,C9 hydroxyls plays a key
role in interacting with tubulin or in maintaining the bioactive
conformation. Anti-a-tubulin staining of PANC-1 pancreatic
carcinoma cells treated with 100 nM of hybrid 3 (Fig. 2) shows
the characteristic patterns of microtubule bundling observed
for other tubulin polymerising agents such as Taxol, disco-
dermolide and dictyostatin.2b Similar to what is observed for
dictyostatin, treatment with 10 nM of 3 shows a large number
of cells undergoing apoptosis as evidenced by high levels of
nuclear fragmentation observed in the confocal images and a
large sub-G0 population in the cell cycle analysis.
Shin, J. H. Fournier, Y. Fukui, A. M. Bruckner and D. P. Curran,
¨
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Madiraju, B. W. Day and D. P. Curran, Org. Lett., 2002, 4, 4443.
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14 G. D. Allred and L. S. Liebeskind, J. Am. Chem. Soc., 1996, 118,
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