Semisynthesis and biological evaluation of a novel D-seco docetaxel analogue

Article abstract of DOI:10.1021/ol060531d

A 4-methyl-5-oxo docetaxel analogue has been prepared starting from 10-deacetylbaccatin III. This new D-seco docetaxel analogue is slightly less potent than docetaxel at microtubule stabilization in vitro and has about 1/1000th the cytotoxicity of docetax

Full text of DOI:10.1021/ol060531d

ORGANIC
LETTERS
2006
Vol. 8, No. 11
2301-2304
Semisynthesis and Biological Evaluation
of a Novel D-Seco Docetaxel Analogue#
Sylviane Thoret, Franc¸oise Gue´ritte, Daniel Gue´nard, and Joe1lle Dubois*
Institut de Chimie des Substances Naturelles-CNRS, aVenue de la Terrasse,
91198 Gif sur YVette Cedex, France
joelle.dubois@icsn.cnrs-gif.fr
Received March 3, 2006
ABSTRACT
A 4-methyl-5-oxo docetaxel analogue has been prepared starting from 10-deacetylbaccatin III. This new D-seco docetaxel analogue is slightly
less potent than docetaxel at microtubule stabilization in vitro and has about 1/1000th the cytotoxicity of docetaxel. The lack of improved
activity for this compound compared to other D-modified taxoids confirms that a C-5 oxygen atom is not required for biological activity.
Paclitaxel (Taxol, 1a)1 and its semisynthetic analogue
docetaxel (Taxotere, 1b) are important anticancer agents
useful for the treatment of breast, ovarian, and non-small
lung cancers2 and are also active against prostate cancer.3
They act by stabilizing microtubules, thereby blocking cell-
cycle progression during mitosis.4
the C-13 side chain, the ester groups at C-2 and C-4, and
the rigid core to which these moieties are attached are
essential for biological activity.5 The contribution of the
oxetane D-ring to the biological activity of taxoids has also
been the subject of extensive studies in the past decade. With
respect to microtubule binding, it may have two functions:
(i) it contributes to rigidify the taxoid core and thereby
impose a specific orientation of the C-2, C-4, and C-13 side
chains, or/and (ii) the oxygen atom at C-5 is involved in a
stabilizing dipolar or hydrogen-bonding interaction with an
amino acid of the tubulin binding site. Many derivatives have
been designed to elucidate the actual contribution of the
oxetane ring in microtubule binding.
Either D-modified or D-seco derivatives have been
synthesized. In the first series, the oxygen atom has been
substituted by nitrogen,6 sulfur,7 or selenium7a atoms or
Since their discovery, these compounds have been the
subject of intense chemical, biological, and clinical investiga-
tions. Especially, extensive SAR studies have shown that
(5) For recent reviews, see: (a) Gue´ritte, F. Curr. Pharm. Des. 2001, 7,
1229-1249. (b) Kingston, D. G. I.; Jagtap, P. J.; Yuan, H.; Samala, L. The
Chemistry of Taxol and Related Taxoids. In Progress in the Chemistry of
Organic Natural Products; Herz, W., Kirby, G. W., Moore, R. E., Steglich,
W., Tamm, C., Eds.; Springer-Verlag: Wien & New York, 2002; Vol. 84,
pp 53-225.
(6) Marder-Karsenti, R.; Dubois, J.; Bricard, L.; Gue´nard, D.; Gue´ritte-
Voegelein, F. J. Org. Chem. 1997, 62, 6631-6637.
(7) (a) Gunatilaka, A. A. L.; Ramdayal, F. D.; Sarragiotto, M.; Kingston,
D. G. I.; Sackett, D. L.; Hamel, E. J. Org. Chem. 1999, 64, 2694-2703.
(b) Merckle´, L.; Dubois, J.; Place, E.; Thoret, S.; Gue´ritte, F.; Gue´nard,
D.; Poupat, C.; Ahond, A.; Potier, P. J. Org. Chem. 2001, 66, 5058-5065.
# Dedicated to Pr Pierre Potier.
* To whom correspondence should be addressed. Phone: 33(0)-
169823058. Fax: 33(0)169077247.
(1) Wani, M. C.; Taylor, H. L.; Wall, M. E.; Coggon, P.; McPhail, A.
T. J. Am. Chem. Soc. 1971, 93, 2325-2327.
(2) Gue´nard, D.; Gue´ritte-Voegelein, F.; Lavelle, F. Curr. Pharm. Des.
1995, 1, 95-112.
(3) De Wit, R. Eur. J. Cancer 2005, 41, 502-507.
(4) Schiff, P. B.; Fant, J.; Horwitz, S. B. Nature 1979, 277, 665-667.
10.1021/ol060531d CCC: $33.50
© 2006 American Chemical Society
Published on Web 04/27/2006

together with the C-4 â-methyl group. The oxetane ring of
compound 6, obtained according to the reported procedure,10
was opened with BF3‚OEt2 to afford compound 7 bearing
an oxygen in the C-5 position (Scheme 1). Compound 7 was
Scheme 1
deleted.8 In the D-seco series, only compounds devoid of
an oxygen atom at the C-5 position and/or of the C-4 acetyl
group have been synthesized.9
deacetylated under mild conditions, and the C-20 OH was
reduced under the same conditions as those described by
Barboni et al.9f Formation of the epoxide 8 occurred in high
yield under the same conditions as those reported for the
synthesis of 5(20)deoxydocetaxel.9e Conditions for the ep-
oxide opening with an iodide ion had to be modified for
compound 8 because the reported conditions afforded the
C-20 iodo derivative 9 in very poor yield. The use of THF
as solvent instead of CH2Cl2 and 2 equiv of Lewis acid gave
compound 9 in good yield, and this was hydrogenolyzed to
afford compound 10 in 85% yield. The C-5 hydroxyl group
was then oxidized to ketone using trifluoroacetic anhydride-
activated DMSO in CH2Cl2, and finally, the C-4 OH was
acetylated under classical conditions (Scheme 1). Compound
11 was thus obtained in seven steps from modified 10-
deacetylbacatin III 6 with 21% overall yield. It should be
noted that only this sequence of reactions can afford
compound 11 from 6 in acceptable yield because oxidation
of the C-5 OH prior to deacetylation gives rise to unexpected
side reactions during acetyl removal.
Whereas the C-1,C-2 carbonate of 11 was readily opened
by phenyllithium in good yield (Scheme 2), further modi-
fications to complete the synthesis of the docetaxel analogue
5 were troublesome.
The main difficulty lies on the removal of the protecting
groups. Under the usual conditions to remove the silyl
protecting groups on taxoids (HF/pyridine complex), the
A-ring of compound 12 underwent a Wagner-Meerwein-
type rearrangement that has already been observed on taxoids
but under more acidic conditions (Scheme 2).11
The most active compounds in both series are 5(20)-
deoxydocetaxel 28 and the 4-methyl paclitaxel analogue 39f
which are almost as active as their parent compounds on
the tubulin assay but are much less cytotoxic. The closely
related compound 4 that lacks the 20-methyl group has
dramatically reduced microtubule-stabilizing activity as well
as low cytotoxicity. It has been suggested that this activity
loss could come from the modified C-ring conformation and
acetoxy orientation.9e Therefore, it has been proposed from
all these results that the D-ring is not necessary for tubulin
activity as long as the overall conformation of the taxane
skeleton, as well as the position of the C-4 acetoxy group
relative to the C-13 side chain, is maintained.9e,f However,
compounds 2 and 3 are less cytotoxic than their parent
compounds (100- and 400-fold respectively), and we won-
dered if the absence of the oxygen atom at C-5 could be
responsible for this reduced cytotoxicity. Therefore, to
evaluate the importance of the C-5 oxygen atom on biological
activity, we designed and synthesized the D-seco compound
5, bearing a methyl group at C-4 and a ketone at C-5.
Our starting material was 10-deacetylbaccatin III (DAB)
suitably modified to allow the construction of the C-5 ketone
(8) Dubois, J.; Thoret, S.; Gue´ritte, F.; Gue´nard, D. Tetrahedron Lett.
2000, 41, 3331-3334.
(9) (a) Barboni, L.; Dutta, A.; Dutta, D.; Georg, G. I.; Vander Velde, D.
G.; Himes, R. H.; Wang, M.; Snyder, J. P. J. Org. Chem. 2001, 66, 3321-
3329. (b) Samaranayake, G.; Magri, N. F.; Jitrangsri, C.; Kingston, D. G.
I. J. Org. Chem. 1991, 56, 5114-5119. (c) Beusker, P. H.; Veldhuis, H.;
Van den Bossche, B. A. C.; Scheeren, H. W. Eur. J. Org. Chem. 2001,
1761-1768. (d) Beusker, P. H.; Veldhuis, H.; Brinkhorst, J.; Hetterscheid,
D. G. H.; Feichter, N.; Bugaut, A.; Scheeren, H. W. Eur. J. Org. Chem.
2003, 689-705. (e) Deka, V.; Dubois, J.; Thoret, S.; Gue´ritte, F.; Gue´nard,
D. Org. Lett. 2003, 5, 5031-5034. (f) Barboni, L.; Giarlo, G.; Ricciutelli,
M.; Ballini, R.; Georg, G. I.; VanderVelde, D. G.; Himes, R. H.; Wang,
M.; Lakdawala, A.; Snyder, J. P. Org. Lett. 2004, 6, 461-464.
(10) Ojima, I.; Lin, S.; Inoue, T.; Miller, M. L.; Borella, C. P.; Geng,
X.; Walsh, J. J. J. Am. Chem. Soc. 2000, 122, 5343-5353.
2302
Org. Lett., Vol. 8, No. 11, 2006

reported procedure15 and then to protect the C-7 alcohol with
a TES group to introduce the docetaxel side chain selectively
at the C-13 position. Surprisingly, the formation of the
7-triethylsilyl-10-acetylbaccatin III derivative 16 proved to
be difficult. Silylation conditions had to be modified, and
the addition of a catalytic amount of DMAP together with
heating were the only conditions that were able to afford
16. After acylation of the C-13 OH, full deprotection of
compound 17 was carried out in two steps: removal of the
TES group and then opening of the oxazolidine ring of the
docetaxel side chain (Scheme 4).
Scheme 2
Scheme 4
Many other deprotection conditions (HCOOH, PTSA, HCl,
TFA, and CAN) were tried, leading either to rearranged
compound 13 or to starting material 12. We suspected
compound 12 to be more sensitive to an acidic medium than
other DAB derivatives, and so we added pyridine to the HF/
pyridine complex to buffer the reaction medium. As ex-
pected, the fully deprotected compound 14 was obtained in
85% yield under these conditions (Scheme 2).
The next step in our synthesis was to introduce the
docetaxel side chain selectively at C-13. It has been reported
that a sterically bulky group at C-7, such as TES, prevents
enzymatic acylation at C-10.12 Furthermore, the phenyliso-
serine side chain has been introduced selectively at C-13 on
2-debenzoyl 4-deacetyl 7-TES DAB13a and 2-debenzoyl
7-TES DAB.13b We thought the chemical acylation by the
protected docetaxel side chain after selective silylation at
the C-7 OH of compound 14 would be worth trying.14
However, under our usual conditions, both the C-10 and C-13
hydroxyl groups were acylated affording compound 15
(Scheme 3).
Compound 15 was also deprotected under the same
conditions, affording compound 18 for biological evaluation
(Scheme 4).
Scheme 3
Then, biological activities of compounds 5 and 18 were
evaluated on a cold-induced microtubule disassembly assay
and in an antiproliferative assay on KB cell lines.
Both compounds are less active than docetaxel 1b and
5(20)deoxydocetaxel 2 (Table 1). Addition of an oxygen
atom to the same position as in the oxetane ring does not
improve the biological activity of D-seco taxoids. On the
basis of NMR data, it can be stated that the overall
conformation of compound 5 is identical to that of docetaxel.
As described for docetaxel and the two biologically active
D-modified taxoids, 5(20)deoxydocetaxel 28 and compound
3,9f the coupling constant between H-2 and H-3 is 7.0 Hz;
H-13 has nearly equal coupling to both protons at C-14, and
the signal is a broad triplet (J13-14 ) 8 Hz). Therefore, the
slight loss of activity may not be due to conformational
changes in compound 5.
Because 10-acetyldocetaxel is as active as docetaxel, we
decided to acetylate the C-10 OH first according to the
(11) (a) Samaranaryake, G.; Magri, N. F.; Jitrangari, C.; Kingston, D.
G. I. J. Org. Chem. 1991, 56, 5114-5119. (b) Wahl, A.; Gue´ritte-Voegelein,
F.; Gue´nard, D.; Le Goff, M.-T.; Potier, P. Tetrahedron 1992, 48, 6965-
6974.
(12) Lee, D.; Kim, K.-C.; Kim, M.-J. Tetrahedron Lett. 1998, 39, 9039-
9042.
(13) (a) Gue´ritte-Voegelein, F.; Gue´nard, D.; Dubois, J.; Wahl, A.;
Marder, R.; Mu¨ller, R.; Lund, M.; Bricard, L.; Potier, P. In Taxane
Anticancer Agents: Basic Science and Applications; Georg, I. G., Chen,
T. T., Ojima, I., Vyas, D. M., Eds.; ACS Symposium Series Nï. 583;
American Chemical Society: Washington, DC, 1995; pp 189-202. (b)
Kingston, D. G. I. Ibid. pp 203-216.
(14) Denis, J.-N.; Greene, A. E.; Gue´nard, D.; Gue´ritte-Voegelein, F.;
Mangatal, L.; Potier, P. J. Am. Chem. Soc. 1988, 110, 5917-5919.
(15) Holton, R. A.; Zhang, Z.; Clarke, P. A.; Nadizadeh, H.; Procter, D.
J. Tetrahedron Lett. 1998, 39, 2883-2886.
Org. Lett., Vol. 8, No. 11, 2006
2303

Taking the results on D-modified taxoids together, it can
be concluded that the oxetane ring is not essential for tubulin
binding. The contribution of the oxygen atom to microtubule
interaction seems to be weak, in disagreement with the
hypothesis of a hydrogen bond between the oxetane and Thr
274 in the binding site.17 With regard to the oxetane binding
site, it is to be noted that it may be relatively restricted
because C-20 acetoxy derivatives9a and N-20 Me or N-20
Ac azetidine analogues18 are completely inactive. The most
important element for microtubule interaction is definitely
the conformational properties of the taxane diterpene that
can be preserved by the presence of either a small D-cycle
(four- or three-membered ring) or a C-4 â-Me as in
compounds 3 and 5.
Table 1. Results of the Biological Evaluation of Compounds 5
and 18
microtubule-disassembly
inhibitory activitya
cytotoxicity against
KB cell lineb
compound
IC50/IC50(paclitaxel)
IC50 (nM)
1b
2
5
0.5
1.2
3.5
10
0.6
60
600
18
3000
a IC50 is the concentration that inhibits 50% of the rate of microtubule
disassembly. The ratio IC50/IC50(paclitaxel) gives the activity with respect
to paclitaxel. IC50(paclitaxel) ) 1 µM. b IC50 measures the drug concentra-
tion required for the inhibition of 50% cell proliferation after 72 h of
incubation.
The absence of the oxetane ring is more crucial for
cytotoxicity and may reflect other factors besides microtubule
interaction. Addition of an oxygen atom was thought to
increase the bioavailability of this D-seco derivative. Because
no cytotoxicity improvement has been observed, other
elements have to be further investigated to explain this
reduced cytotoxicity of D-modified taxoids.
Compound 18 is less active than 5 showing that the
addition of a second docetaxel side chain at C-10 is
detrimental to the activity. This result is not surprising
because it has already been reported that the addition of a
cinnamyl side chain at C-10 reduces the tubulin activity.16
In summary, a novel D-seco taxoid 5 bearing an oxygen
atom at the same position as in the oxetane ring has been
synthesized. The 10-deacetylbaccatin III derivative 12 dis-
plays unusal chemical behavior, in particular, an increased
sensitivity to an acidic medium compared to the other
D-modified DAB derivatives already described.6,7b,8,9e,10 This
reactivity has made the complete transformation of 12 to
docetaxel analogue 5 very complicated.
Acknowledgment. The authors thank Dr. A. Commerc¸on
(Sanofi-Aventis) for a gift of the docetaxel side chain. J. F.
Gallard and M. T. Martin are acknowledged for performing
NMR spectra, and C. Gaspard is acknowledged for cyto-
toxicity evaluations. The authors are very grateful to Prof.
G. Appendino (University of Torino, Italy) for helpful
discussions on silyl group removal.
Contrary to our expectation, addition of an oxygen atom
at C-5 on D-seco taxoids does not improve the antiprolif-
erative nor the microtubule-stabilizing activities. These
activities are even slightly reduced in the tubuline assay and
are lower by an order of magnitude for cytotoxicity.
However, they are comparable to that of thia derivatives of
docetaxel.7b
Supporting Information Available: Experimental pro-
cedures and spectral data for compounds 5 and 7-18. This
material is available free of charge via the Internet at
http://pubs.acs.org.
OL060531D
(17) Wang, M.; Cornett, B.; Nettles, J.; Liotta, D. C.; Snyder, J. P. J.
Org. Chem. 2000, 65, 1059-1068.
(18) Unpublished results. 5(20)-Azadocetaxel is 16-fold less active than
docetaxel.
(16) Gue´nard, D.; Thoret, S.; Dubois, J.; Adeline, M.-T.; Wang, Q.;
Gue´ritte, F. Bioorg. Med. Chem. 2000, 8, 14-156.
2304
Org. Lett., Vol. 8, No. 11, 2006