A unique approach to the concise synthesis of highly optically active spirooxazolines and the discovery of a more potent oxindole-type phytoalexin analogue

Article abstract of DOI:10.1021/ja106349m

Drug-lead synthesis through rapid construction of chiral molecular complexity around the biologically relevant framework using a highly efficient strategy is a key goal of organic synthesis. Molecules bearing a spirooxindole-type framework exhibit important bioactivities. Herein, we present a highly efficient and convenient strategy that allows rapid construction of unique optically active spiro[oxazoline-3,3′-oxindole]s through the organocatalyzed asymmetric synthesis of spirocyclic thiocarbamates via an aldol reaction. Preliminary biological evaluation of several of the spirooxazolines using a model of acute neuroinflammation revealed promising antipyretic activity and provided an opportunity to discover new antipyretic agents.

Full text of DOI:10.1021/ja106349m

Published on Web 10/12/2010
A Unique Approach to the Concise Synthesis of Highly
Optically Active Spirooxazolines and the Discovery of a More
Potent Oxindole-Type Phytoalexin Analogue
Xianxing Jiang,†,‡ Yiming Cao,† Yiqing Wang,† Luping Liu,† Fangfang Shen,† and
Rui Wang*,†,‡
Key Laboratory of Preclinical Study for New Drugs of Gansu ProVince, Institute of Biochemistry
and Molecular Biology, State Key Laboratory of Applied Organic Chemistry, Lanzhou
UniVersity, Lanzhou, China, and State Key Laboratory of Chiroscience, Department of Applied
Biology and Chemical Technology, The Hong Kong Polytechnic UniVersity, Kowloon, Hong Kong
Received July 28, 2010; E-mail: wangrui@lzu.edu.cn
Abstract: Drug-lead synthesis through rapid construction of chiral molecular complexity around the
biologically relevant framework using a highly efficient strategy is a key goal of organic synthesis. Molecules
bearing a spirooxindole-type framework exhibit important bioactivities. Herein, we present a highly efficient
and convenient strategy that allows rapid construction of unique optically active spiro[oxazoline-3,3′-
oxindole]s through the organocatalyzed asymmetric synthesis of spirocyclic thiocarbamates via an aldol
reaction. Preliminary biological evaluation of several of the spirooxazolines using a model of acute
neuroinflammation revealed promising antipyretic activity and provided an opportunity to discover new
antipyretic agents.
Scheme 1. Structure of Oxindole-Type Phytoalexins
Introduction
The oxindole framework bearing a spirocyclic quaternary
stereocenter at the C3 position represents a privileged hetero-
cyclic motif commonly found in clinical pharmaceuticals and
a number of natural spirooxindole alkaloids.1 The oxindole-
type phytoalexins2 isolated from the plant family Cruciferae have
shown potent antimicrobial, antitumor, and oviposition-stimulant
biological activities (Scheme 1).3 Since spiro[oxazoline-3,3′-
oxindole] can be considered as an important analogue of the
oxindole-type phytoalexins, the development of highly efficient
synthetic methods to access optically active spirooxindole
derivatives would be of great interest for drug-lead synthesis.
Furthermore, it remains a great challenge to develop an efficient
and convenient strategy to access these optically active natural
alkaloids and related analogues for further biological studies,
thereby contributing to the development of new therapeutic
agents. On the basis of our recent efforts to build upon chiral
precedents by using rosin-derived thiourea catalysts in our
group4,5 and to gain a better understanding of the scope of this
conceptually new catalytic system, we hoped to expand our
studies beyond model compounds to develop an efficient
protocol for accessing potentially bioactive chiral spiro[oxazo-
line-3,3′-oxindole]s and to provide a foundation for further
development of new types of therapeutic agents through pre-
liminary biological studies.
Results and Discussion
We envisioned that the intramolecular aldol6 cyclization
reaction of R-isothiocyanato imides7 to electron-deficient isatins
could analogously be initiated in the presence of a chiral tertiary
amine, leading to the generation of spiro[thiocarbamate-3,3′-
oxindole]s. Subsequent methylation would afford spirooxazo-
lines, which are isosteric analogues of natural spirobrassinin
(Scheme 2). To our knowledge, there is no precedent for the
catalytic asymmetric synthesis of highly optically active spiro[ox-
azoline-3,3′-oxindole]s, and the current protocol provides an
alternative asymmetric access to chiral spirooxazolines. Herein,
we first present our contribution to the successful development
of such a reaction and preliminary biological studies of the
activity toward the antipyretic role in the development of acute
neuroinflammation.
† Lanzhou University.
‡ The Hong Kong Polytechnic University.
(1) For recent reviews, see: (a) Trost, B. M.; Jiang, C. Synthesis 2006,
369. (b) Williams, R. M.; Cox, R. J. Acc. Chem. Res. 2003, 36, 127.
(c) Galliford, C. V.; Scheidt, K. A. Angew. Chem., Int. Ed. 2007, 46,
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(e) Lin, H.; Danishefsky, S. J. Angew. Chem., Int. Ed. 2003, 42, 36.
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M.; Dzurilla, M.; Balentova´, E. J. Org. Chem. 2001, 66, 3940. (b)
Takasugi, M.; Monde, K.; Katsui, N.; Shirata, A. Chem. Lett. 1987,
1631.
(3) For selected examples, see: (a) Pedras, M. S. C.; Okanga, F. I.; Zaharia,
I. L.; Khan, A. Q. Phytochemistry 2000, 53, 161. (b) Baur, E.; Sta˜dler,
K.; Monde, K.; Takasugi, M. Chemoecology 1998, 8, 163. (c) Mehta,
R. G.; Liu, J.; Constantinou, A.; Hawthorne, M.; Pezzuto, J. M.; Moon,
R. C.; Moriarty, R. M. Anticancer Res. 1994, 14, 1209.
To explore the possibility of the proposed aldol cyclization
process, our investigation began with screening of several
9
15328 J. AM. CHEM. SOC. 2010, 132, 15328–15333
10.1021/ja106349m  2010 American Chemical Society

Concise Synthesis of Highly Optically Active Spirooxazolines
A R T I C L E S
Scheme 2. Strategy for Synthesis of Chiral Spirooxazolines Using
Chiral Tertiary Amines
Table 2. Spiro[thiocarbamate-3,3′-oxindole]s Formed by
Rosin-Derived Tertiary Amine-Thiourea-Catalyzed Asymmetric
Aldol Reactiona
Table 1. Synthesis of Spiro[thiocarbamate-3,3′-oxindole] 3a Using
Various Organocatalysts and Optimization of the Reaction
Conditionsa
Entry
Cat.
Yield (%)b
drc
ee (%)d
1
2
3
4
5
6
7
8
TEA
DIEA
L1
L2
L3
L4
L5
L6
L7
L8
L3
L3
L3
71
76
92
93
98
10
65
90
79
76
98
96
36
>99:1
>99:1
>99:1
>99:1
>99:1
n.d
>99:1
>99:1
>99:1
>99:1
>99:1
>99:1
>99:1
0
0
80
91
96
n.d.
62
86
76
75
95
96
66
a For experimental details, see the Supporting Information. b The
configuration was assigned by comparison with the HPLC and X-ray
crystal data for 3b. c Isolated yield. d Determined by 1H NMR spec-
troscopy and chiral HPLC. e The ee values were determined by HPLC.
f Using 10 mol % ligand loading.
9
10
11e
12f
13g
no ee value was inevitably observed in the reactions (entries 1
and 2). Gratifyingly, except for L4, this catalytic asymmetric
process generally exhibited efficacy in the presence of chiral
organocatalysts. The results showed that tertiary amine-thiourea
catalysts L1-L3, L5, and L6 gave the products with the same
excellent diastereoselectivities (>99:1 dr; entries 3-5, 7, and
8). Thiourea catalysts L2 and L3 provided excellent results in
terms of yield and stereochemical outcome (entries 4 and 5),
but L3 furnished the products with slightly higher yield and
enantioselectivity (98% yield and 96% ee; entry 5). In contrast,
thiourea catalyst L4 proved to be essentially inactive for this
transformation (entry 6). Under the same reaction conditions,
quinidine derivatives L7 and L8 showed relatively low catalytic
activities in comparison with L1-L3 and L6, affording the
product 3a in yields ranging from 79 to 76% and moderate
enantioselectivities (76 and 75% ee, respectively; entries 9 and
10). The tertiary amine-thiourea catalyst L3 was found to be
the most promising catalyst for further investigation. In recent
studies,5b,c we also tried to explain the role of structural features
of thiourea catalysts in obtaining high enantioselectivity by
a Unless otherwise noted, the reaction was conducted with 1a (0.24
mmol) and 2a (0.20 mmol) in CH2Cl2 (1.0 mL) for 12 h at room
temperature. b Isolated yield. c Determined by 1H NMR spectroscopy.
d The ee values were determined by HPLC, and the configuration was
assigned by comparison with HPLC and X-ray crystal data for 3b.
e Using 5.0 mol % ligand loading. f Using 3.0 mol % ligand loading.
g Using 1.0 mol % ligand loading.
organocatalysts to evaluate their catalytic activities. The model
reaction of N-methylisatin (1a) with isothiocyanate 2a was
performed in CH2Cl2 at room temperature in the presence of a
10 mol % loading of ligand (Table 1). Besides the chiral tertiary
amine-thiourea catalysts L1-L6 with diversely structured
scaffolds, the quinidine-derived bifunctional catalysts L7 and
L88 as well as the two organic bases triethylamine (TEA) and
diisopropylethylamine (DIEA) were also tested. While the
desired products could be obtained in moderate yields with
excellent diastereoselectivities in the presence of TEA or DIEA,
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A R T I C L E S
Jiang et al.
Figure 1. X-ray crystal structure of 3b.
Figure 2. Proposed model of the reaction transition state.
carrying out reactions with catalysts bearing different configura-
tions of the chiral scaffold moiety. The effects of the rosin-
derived thiourea catalysts were investigated in comparison with
other thiourea catalysts. We disclosed that the two chiral
moieties of the thiourea are mutually reinforcing for the high
efficacy of the catalyst. The stereochemical control of the
reaction is mainly provided by the 1,2-diaminocyclohexane
moiety of the thiourea. The catalytic activity depends mainly
on the remaining chiral scaffold moiety of the thiourea (the
inherent property of the stereochemical structure of the dehy-
droabietic amine moiety of the thiourea) and also on the suitable
matching of the configuration of the 1,2-diaminocyclohexane
moiety with the remaining chiral scaffold moiety. In addition,
the excellent structural backbone and well-defined stereocenters
of the dehydroabietic amine moiety of L3 also have an important
effect on the high enantioselectivity for the formation of adduct.
These results also indicated that the excellent diastereoselectivity
results from substrate stereocontrol in the reaction. To our
delight, we further lowered the loading of catalyst L3 to 3.0
mol % and still obtained 96% ee without a significant decrease
in yield (96% yield; entry 12).
are summarized in Table 2. The results showed that variation
of the electronic properties of the substituent at either C4, C5,
or C7 of the N-protected spiro[thiocarbamate-3,3′-oxindole] with
different steric parameters was tolerated, affording the products
with excellent enantioselectivities (91%-99% ee) and diaste-
reoselectivities in good to excellent yields (70%-99%; entries
1-18). To our delight, spirocyclic thiocarbamate 3s was also
favorably formed with 99% yield, >99:1 dr and >99% ee (entry
19). It is worth noting that the spiro[thiocarbamate-3,3′-
oxindole]s bearing contiguous tetrasubstituted chiral carbon
stereocenters9 (3n-r) were still obtained in good yields and
excellent stereoselectivities (entries 14-18). Additionally, as
expected, the catalytic system also proved to be efficient for
construction of contiguous dispiro[thiocarbamate-3,3′-oxin-
dole]s, again leading to excellent yields (92-99%), albeit with
a decrease in diastereoselectivity (entries 20-23). The relative
and absolute configurations of the products were determined
by X-ray crystal structure analysis of 3b (see Figure 1).
On the basis of the experimental results described above and
recent studies,10 a possible model to account for the high
enantioselectivity of the present reaction is shown in Figure 2.
The rosin-derived chiral tertiary amine-thiourea is proposed
as a bifunctional catalyst. The electron-deficient N-protected
Results of experiments in which a variety of spirocyclic
thiocarbamates were synthesized under the optimized conditions
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Concise Synthesis of Highly Optically Active Spirooxazolines
A R T I C L E S
Scheme 3. Synthesis of Biologically Active Chiral Spirooxazolinesa
a Synthesis details are given in the Experimental Section.
isatin is activated by the two thiourea hydrogen atoms through
weak hydrogen bonds, while the acidic R-carbon atom of the
isothiocyanate could be activated by an interaction between the
neighboring tertiary amine moiety of the catalyst and the
isothiocyanate. The si-face of the N-protected isatin is predomi-
nantly approached by the incoming nucleophile generated from
the isothiocyanate and the tertiary amine group of the bifunc-
tional catalyst (attack on the re-face of the N-protected isatin is
restricted by the cyclohexyl scaffold of the catalyst). Subsequent
intramolecular cyclization reactions of intermediates afford the
chiral spirocyclic products, which is consistent with the exper-
imental results.
With the successful construction of spirocyclic thiocarbamates
as described above, various optically active spirooxazolines were
synthesized under the asymmetric protocol established next
(Scheme 3; for details, see the Supporting Information). The
substituted spiro[oxazoline-3,3′-oxindole]s SPX-A through
SPX-N having various steric and electronic parameters, includ-
ing in some cases N-methyl or N-allyl protection, were smoothly
formed with good to excellent enantioselectivities (83% to 99%
ee) and diastereoselectivities (up to >99:1 dr) in yields ranging
from 60 to 82%.
The oxindole-type phytoalexin is widely known as an
important component of the multifaceted defense systems of
plants against microbial attack. Especially, in recent years, the
interesting antimicrobial activity has been reported.11 In view
of the ease with which we were able to access diverse analogues
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A. G.; Jacobsen, E. N. Chem. ReV. 2007, 107, 5713. (b) Connon, S. J.
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A R T I C L E S
Jiang et al.
spirooxazolines on the fever by icv injection of LPS. The current
results demonstrate a role of the analogues in an animal model
of acute neuroinflammation, pointing out their critical role in
the fever induced by central administration of LPS. Excitingly,
LPS-induced fever was significantly reduced by coinjection of
several of the analogues, including SPX-F (20 nmol, p < 0.05)
and SPX-I (20 nmol, p < 0.05); SPX-F (20 nmol) or SPX-I
(20 nmol) given alone into the third ventricle did not signifi-
cantly alter the body temperature. The two analogues admin-
istered at 2 nmol along with LPS slightly but not significantly
reduced the LPS-induced fever. The realization that this kind
of analogue plays a key role in the control of the process of
neuroinflammation is a new concept and may well lead to a
fruitful approach for identifying novel therapies for neuroin-
flammatory conditions. For example, it may be possible to use
this kind of analogue as a therapeutic strategy to prevent and
treat brain diseases associated with neuroinflammation (e.g.,
multiple sclerosis, Alzheimer’s disease).
Conclusion
In summary, we have disclosed the synthesis of highly
optically active spirooxazolines through organocatalyzed asym-
metric synthesis of spirocyclic thiocarbamates with high levels
of enantio- and diastereoselectivity (up to >99% ee and >99:1
dr) via an aldol reaction. Several of the new spirooxindoles were
found to significantly reduce LPS-induced fever using a model
of acute neuroinflammation. The preliminary biological studies
on the activity toward the antipyretic role provide a foundation
for further development of new spirooxindole-type antipyretic
agents.
Figure 3. Biological activity of SPX-F and SPX-I in acute neuroinflam-
mation. Time courses of the change in body temperature induced by LPS
in the absence or presence of (top) SPX-F or (bottom) SPX-I injected into
the third ventricle in mice are shown. The rectal temperature was recorded
after injection of control, LPS, or the coapplication of LPS and SPX-F or
SPX-I. Each data point represents the mean ( standard error of the mean
from experiments conducted on 6-8 mice per group. Points labeled with
# were significantly different (p < 0.05) from the corresponding points for
LPS alone. (For experimental details, see the Supporting Information).
Experimental Section
General Procedure for the Synthesis of Highly Optically
Active Spiro[oxazoline-3,3′-oxindole] (SPX-A). To a stirred
solution of L3 (0.012 mmol, 3.0 mol %) and N-methylisatin (1a)
(0.48 mmol, 77.3 mg) in dry CH2Cl2 (2.5 mL) was added
isothiocyanate 2a (0.40 mmol, 85.6 mg) under Ar. The solution
was stirred at room temperature for 12 h. After the reaction was
complete (as determined by TLC), the resulting mixture was
concentrated under reduced pressure, and the residue was purified
through column chromatography on silica gel to give the optically
pure spiro[thiocarbamate-3,3′-oxindole] 3a (144.0 mg). To a mixture
and the unique architecture of the optically active spiro[oxazo-
line-3,3′-oxindole]s formed, we decided to evaluate the biologi-
cal activities of several spirooxazolines on fever by intracere-
broventricular (icv) injection of lipopolysaccharide (LPS, a
component of the outer membrane of Gram-negative bacteria)
using a model of acute neuroinflammation in mice (Figure 3).
The injection of LPS directly into the brain has been recognized
as an animal model for the study of neuroinflammation.12 Fever
is part of the acute-phase reaction to infection, which is char-
acterized by a raised thermoregulatory set point, leading to an
elevation in body temperature.13 Although fever is an important
indicator for the severity of the inflammation, no one has
investigated and/or linked this parameter with spirooxazolines
during neuroinflammation. Using a model of acute neuroin-
flammation, we sought to determine the effects of the several
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A R T I C L E S
of 3a and anhydrous K2CO3 (60.7 mg, 1.10 equiv) in 5.0 mL of
acetone was added MeI (62.5 mg, 1.10 equiv) dropwise at 0 °C,
after which the reaction was left overnight and the concentrated in
vacuo. The mixture was subjected to chromatography to afford the
purified product SPX-A (126.0 mg, 81% yield). The enantiomeric
purity of the product was determined using HPLC, and the dr was
determined using 300 Hz 1H NMR spectroscopy. [R]2D0 ) +115 (c
) 1.0, CHCl3). Mp: 209 °C. Major diastereomer: ee ) 96%, as
determined by HPLC analysis (Chiralcel AD-H, i-PrOH/hexane )
30/70, 1.0 mL/min, 254 nm). Retention times: tmajor ) 11.76 min,
tminor ) 35.99 min.
Materials and General Procedure for the Biological Studies.
Male Kunming mice weighing 27-30 g were used. Each animal
was used only once. The experiments were performed between 10:
00 and 17:00. All mice were housed one per cage in a room
maintained at 22 ( 0.5 °C and a relative humidity of 52 ( 2%
with free access to food and water. The mice were placed in the
specially designed restraining device as described by Rosow et al.14
with their tails taped lightly to horizontal posts. Rectal temperatures
were measured with a thermistor probe (Machine Equipment
Corporation, GaoBeiDian, China) inserted to a depth of 2.5 cm
into the rectum and linked to a recorder system (model BL-420E+,
Taimeng Technology Corporation, Chengdu, China). The icv
administration in the third ventricle was performed following the
method described by France´s et al.15 The drugs were coinjected to
investigate whether the fever of LPS could be antagonized by the
spiro[oxazoline-3,3′-oxindole]. Body temperature was recorded
before and then at 120, 180, 240, and 300 min after icv injection
in the third ventricle of control or various treatments. Changes in
body temperature after injection and before drug administration were
calculated for each animal. The time courses of the change in body
temperature of mice subjected to different treatments are shown in
Figure 3. Data were given as mean ( standard error of the mean.
One-way ANOVA followed by Bonferroni’s posthoc test was used
to establish statistical significance; a probability level of p < 0.05
was considered to be significant.
Acknowledgment. This work is dedicated to Professor Albert
S. C. Chan on the occasion of his 60th birthday. We are grateful
for the grants from the NNSF of China (20932003 and 90813012)
and the National S & T Major Project of China (2009ZX09503-
017).
Supporting Information Available: Experimental details,
compound characterization, and X-ray crystallographic data for
3b (CIF). This material is available free of charge via the
Internet at http://pubs.acs.org.
JA106349M
(14) Rosow, C. E.; Miller, J. M.; Pelikan, E. W.; Cochin, J. J. Pharmacol.
Exp. Ther. 1980, 213, 273.
(15) Frances, B.; Lahlou, H.; Zajac, J. M. Regul. Pept. 2001, 98, 13.
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