Synthesis, tubulin assembly, and antiproliferative activity against MCF7 and NCI/ADR-RES cancer cells of 10-O-acetyl-5′-hydroxybutitaxel

Article abstract of DOI:10.1016/j.bmcl.2008.10.010

A highly efficient kinetic resolution of racemic cis-4-(2-tert-butyldimethylsilyloxy-1,1-dimethyl)ethyl-3-tert-butyldimethylsilyloxy-azetidin-2-one with 7-O-triethylsilylbaccatin III was carried out to furnish 10-O-acetyl-5′-hydroxybutitaxel after removal of the silyl protecting groups. The compound was 50% as active as paclitaxel in a tubulin assembly assay and showed significantly decreased activity against MCF7 cell proliferation compared to paclitaxel.

Full text of DOI:10.1016/j.bmcl.2008.10.010

Bioorganic & Medicinal Chemistry Letters 18 (2008) 6165–6167
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis, tubulin assembly, and antiproliferative activity against MCF7
and NCI/ADR-RES cancer cells of 10-O-acetyl-50-hydroxybutitaxel
Haibo Ge a, Jianmei Wang b, Margaret M. Kayser b, Richard H. Himes c, Gunda I. Georg a,
*
a Department of Medicinal Chemistry, University of Kansas, 1251 Wescoe Hall Drive, Lawrence, KS 66045, USA
b Department of Physical Sciences, University of New Brunswick, Saint John, Canada NB E2L 4L5
c Department of Molecular Biosciences, University of Kansas, 1200 Sunnyside Avenue, Lawrence, KS 66045, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A highly efficient kinetic resolution of racemic cis-4-(2-tert-butyldimethylsilyloxy-1,1-dimethyl)ethyl-3-
tert-butyldimethylsilyloxy-azetidin-2-one with 7-O-triethylsilylbaccatin III was carried out to furnish 10-
O-acetyl-50-hydroxybutitaxel after removal of the silyl protecting groups. The compound was 50% as
active as paclitaxel in a tubulin assembly assay and showed significantly decreased activity against
MCF7 cell proliferation compared to paclitaxel.
Received 25 July 2008
Revised 30 September 2008
Accepted 2 October 2008
Available online 7 October 2008
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
Paclitaxel analogues
Semisynthesis
Kinetic resolution
Tubulin assembly
Cytotoxicity
MCF7
NCI/ADR RES
The structurally and biologically unique diterpenoid paclitaxel
(1a, Fig. 1),1 and its semisynthetic derivative docetaxel (1b,
Fig. 1), are among the most effective anticancer agents known for
the treatment of metastatic breast cancer, refractory ovarian can-
cer and Kaposi’s sarcoma.2,3 Paclitaxel suffers from a number of
disadvantages, including low aqueous solubility, low oral bioavail-
ability, development of drug resistance, and inability to cross the
blood–brain barrier (BBB).4 Therefore, the continued synthesis
and evaluation of new analogues has been important and has led
to promising new results.5 For example, butitaxel analogue BMS-
275183 (1e, Fig. 1),6 is orally bioavailable, RPR-109881 A (2,
Fig. 1)7 is effective against MDR-positive and taxane resistant hu-
man tumor xenografts, and TX-67 (1d, Fig. 1)8 was shown to per-
meate the BBB in situ.
The efficient semisynthesis of paclitaxel analogues can be
achieved by reacting baccatin III derivatives with optically pure
b-lactams, the precursors for the C13 N-acyl-30-phenylisoserine
side chain. Enantiopure (3R,4S)-b-lactams can be obtained through
an asymmetric ester enolate-imine cyclocondensation or via en-
zyme-catalyzed resolutions.9–13 An alternative semisynthesis,
involving the kinetic resolution of racemic b-lactams with 7-O-tri-
ethylsilylbaccatin III derivatives has been studied for more than 10
years after being first described by Holton et al.14 This method has
O
R1
OR3
NH
O
O
10
OH
2'
R2
O
3'
13
7
2
OH
HO
H
BzO
R4O
O
1
2
3
4
1a Paclitaxel, R = R =Ph, R = R =Ac
Docetaxel, R = t-BuO, R = Ph, R = H, R = Ac
1c Butitaxel,R = t-BuO, R =t-Bu, R = H, R = Ac
1d TX-67, R = R = Ph, R = COCH CH CO H, R = Ac
BMS-275183, R = t-BuO, R = t-Bu, R = Ac, R = CO Me
1
2
3
4
1b
1
2
3
3
4
1
2
4
2
2
2
1
2
3
4
1e
2
O
OAc
O
NH
O
O
O
OH
HO
H
BzO
AcO
O
2 RPR-109881 A
* Corresponding author. Tel.: +1 612 626 6320; fax: +1 612 626 6318.
E-mail address: georg239@umn.edu (G.I. Georg).
Figure 1. Structures of paclitaxel (1a), docetaxel (1b) butitaxel (1c), Tx-67 (1d),
BMS-275183 (1e), and RPR-109881 A (2).
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.10.010

6166
H. Ge et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6165–6167
Table 1
not been used widely for the semisynthesis of paclitaxel analogues
because of unpredictable diastereoselectivities obtained during the
kinetic resolution.6,15,16
ED50 values for tubulin assembly and cytotoxicity assays
Compound
Assemblya
(l
M)
MCF7b (nM)
NCI/ADR-RES (lM)
Our recent study of the kinetic resolution of racemic b-lac-
tams with 7-O-triethylsilylbaccatin III17 showed that the struc-
ture of the b-lactam played an important role in controlling
the diastereoselectivity of the kinetic resolution. High diastere-
oselectivities were observed when sterically demanding C3-hy-
droxy protecting groups or sterically demanding C4
substituents at the b-lactam rings were present. In this commu-
nication we are reporting the semisynthesis of 10-O-acetyl-50-
hydroxybutitaxel (9) via kinetic resolution with racemic b-lactam
6 that carries two sterically demanding groups: the tert-butylsi-
lyoxy group at C3 and the 1-hydroxy-2-methylpropan-2-yl group
at C4 of the b-lactam ring (Scheme 2). The new hydroxylated
butitaxel analogue 9 was expected to possess better aqueous sol-
ubility (cLogP = 3.7) than paclitaxel (cLogP = 4.2). In addition,
analogue 9 is a potential template for structure-activity studies
as it allows the facile attachment of a variety of groups at the
50-hydroxyl group of the C13 side chain.18
Paclitaxel (1a)
9
1.2
2.2
2.1
560
2.0
12.5
a
The assembly assay was conducted as described in Ref. 23.
The cytotoxicity assay was conducted as described in Ref. 24.
b
(PMP) of 5 was removed oxidatively with ammonium cerium(IV)
nitrate (CAN),20 and the resulting N-unsubstituted b-lactam was
then treated with tert-butyl dicarbonate to afford racemic N-Boc-
b-lactam 6.
We then carried out the kinetic resolution between racemic
b-lactam 6 and 7-O-triethylsilylbaccatin III (8)21 using the previ-
ously reported reaction conditions (Scheme 2).17 Thus, four
equivalents of 6 were used to react with the baccatin derivative
8 in the presence of 1.5 equivalents of LiHMDS in THF at ꢀ40 °C.
The crude reaction product was subsequently dissolved in pyri-
dine and treated with hydrogen fluoride to furnish derivative
9. The isomeric ratio of 9 was determined by HPLC analysis as
described before.22 Excellent diastereoselectivity (216:1) was ob-
served. In our previous study we had observed that diastereose-
lectivities increased with the size of the C4-substutients of N-
Boc-3-OTBS-b-lactams. The 4-phenyl-b-lactam showed a 41:1
diastereomeric ratio and the 4-tert-butyl-b-lactam an 82:1 dia-
stereomeric ratio.17 In the present study, the even more steri-
The synthesis of the racemic cis-b-lactam 6 is shown in Scheme
1, starting with the hydrolysis (KOH) of the acetate in b-lactam 3,19
followed by deprotection of the benzyl group to provide diol 4. Pro-
tection of the two hydroxyl groups of 4 with tert-butyldimethylsi-
lyl chloride provided intermediate 5. The 4-methoxyphenyl moiety
OBn
OH
cally
demanding
4-(2-tert-butyldimethylsilyloxy-1,1-
AcO
O
HO
O
dimethyl)ethyl group in b-lactam 6 provided an enhanced dia-
stereomeric ratio (216:1), further supporting the hypothesis that
C4-substituents play an important role in controlling the diaste-
reoselectivity of this kinetic resolution.
To verify the efficiency of the kinetic resolution, we coupled 8
with enantiopure b-lactam 7 prepared according to the reported
procedure19 under the standard reaction conditions to generate
analogue 9 in 74% yield after deprotection of the silyl groups
(Scheme 2). Spectral data and HPLC retention time of this analogue
are identical to the product obtained in the kinetic resolution. This
result demonstrated the highly efficient feature of the kinetic res-
olution method.
Analogue 9 was tested in a tubulin assembly assay and as an
inhibitor of proliferation of the MCF7 and NCI/ADR-RES breast can-
cer lines (Table 1).23,24 In the tubulin assembly assay analogue 9
was just slightly less active than paclitaxel. However, it showed
significantly reduced activity against MCF7 proliferation (267-
fold).
b
a
N
N
50%
74%
PMP
PMP
(+)
(+)
—
—
3
4
OTBS
OTBS
TBSO
O
TBSO
c
NBoc
68%
N
PMP
O
(+)
—
(+)
—
6
5
Scheme 1. Synthesis of racemic b-lactam 6. Reagents and conditions: (a) KOH, THF,
0 °C; then 10% Pd/C, HCO2NH4, MeOH, reflux; (b) TBSCl, imidazole, DMF, 35 °C; (c)
CAN, CH3CN, ꢀ10 °C; then Boc2O, DMAP, Et3N, CH2Cl2, rt.
In conclusion, a new paclitaxel analogue was synthesized
through a highly efficient kinetic resolution of racemic b-lactam
6 with 7-O-triethylsilylbaccatin III. The observed high diastereose-
lectivity confirms earlier observations that sterically demanding
C4-b-lactams substituents are a key factor in the efficiency of this
kinetic resolution. Although the new paclitaxel analogue showed
excellent activity in the tubulin assembly assay, it had significantly
decreased activity against MCF7 cell proliferation. The results sug-
gest that the compound may not enter the cells as well as paclit-
axel because it is more hydrophilic or possibly because it is a
much better substrate for the multidrug transporter than paclit-
axel. A similar result was observed for the phenolic paclitaxel
metabolite that carries a 4-hydroxy group at the C3 phenyl
group.25
OAc
O
OTBS
OTES
O
TBSO
O
+
HO
a
NBoc
HO
H
BzO
AcO
6: Racemic
7: (3R, 4S)
8
O
OAc
O
NH
O
O
OH
O
2
4
1
3
O
HO
5
OH
HO
H
BzO
AcO
Acknowledgments
9 (72% from 6; 74% from 7)
Scheme 2. Synthesis of 10-O-acetyl-50-hydroxybutitaxel. Reagents and conditions:
(a) LiHMDS, THF, ꢀ40 °C; then HF-pyridine, pyridine, 0 °C-rt.
The authors wish to thank the National Cancer Institute for
financial support of this research (NIH CA82801 and NIH

H. Ge et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6165–6167
6167
11. a Kayser, M. M.; Yang, Y.; Mihovilovic, M. D.; Feicht, A.; Rochon, F. D. Can. J.
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CA105305), NBIF Research Assistantship Initiative, and Tapestry
Pharmaceuticals (Boulder, CO) for a generous gift of 10-deacetyl-
baccatin III. We wish to thank Christopher Schneider for his help
with the HPLC analysis. We also thank Jacquelyn K. Huff for her
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