2,3-diarylcyclopentenones as orally active, highly selective cyclooxygenase-2 inhibitors

Article abstract of DOI:10.1021/jm980642l

Cyclopentenones containing a 4-(methylsulfonyl)phenyl group in the 3- position and a phenyl ring in the 2-position are selective inhibitors of cyclooxygenase-2 (COX-2). The selectivity for COX-2 over COX-1 is dramatically improved by substituting the 2-phenyl group with halogens in the meta position or by replacing the phenyl ring with a 2- or 3-pyridyl ring. Thus the 3,5-difluorophenyl derivative 7 (L-776,967) and the 3-pyridyl derivative 13 (L-784,506) are particularly interesting as potential antiinflammatory agents with reduced side-effect profiles. Both exhibit good oral bioavailability and are potent in standard models of pain, fever, and inflammation yet have a much reduced effect on the GI integrity of rats compared to standard nonsteroidal antiflammatory drugs.

Full text of DOI:10.1021/jm980642l

1274
J . Med. Chem. 1999, 42, 1274-1281
2,3-Dia r ylcyclop en ten on es a s Or a lly Active, High ly Selective Cyclooxygen a se-2
In h ibitor s
W. Cameron Black,* Christine Brideau, Chi-Chung Chan, Stella Charleson, Nathalie Chauret, David Claveau,
Diane Ethier, Robert Gordon, Gillian Greig, J ocelyne Guay, Gregory Hughes, Paule J olicoeur, Yves Leblanc,
Deborah Nicoll-Griffith, Nathalie Ouimet, Denis Riendeau, Denise Visco,† Zhaoyin Wang, Lijing Xu, and
Petpiboon Prasit
Merck Frosst Centre for Therapeutic Research, P.O. Box 1005, Pointe-Claire-Dorval, Que´bec, Canada H9R 4P8, and
Merck Research Laboratories, P.O. Box 2000, Rahway, New J ersey 07065
Received November 13, 1998
Cyclopentenones containing a 4-(methylsulfonyl)phenyl group in the 3-position and a phenyl
ring in the 2-position are selective inhibitors of cyclooxygenase-2 (COX-2). The selectivity for
COX-2 over COX-1 is dramatically improved by substituting the 2-phenyl group with halogens
in the meta position or by replacing the phenyl ring with a 2- or 3-pyridyl ring. Thus the 3,5-
difluorophenyl derivative 7 (L-776,967) and the 3-pyridyl derivative 13 (L-784,506) are
particularly interesting as potential antiinflammatory agents with reduced side-effect profiles.
Both exhibit good oral bioavailability and are potent in standard models of pain, fever, and
inflammation yet have a much reduced effect on the GI integrity of rats compared to standard
nonsteroidal antiflammatory drugs.
In tr od u ction
The conversion of arachadonic acid to prostaglandins
is mediated by the two-step action of prostaglandin H
synthase (PGHS). The first committed step in this
process is the oxidative cyclization of arachidonic acid
to PGG2, which is followed by peroxide reduction to
PGH2 at a second distinct binding site. PGHS, com-
monly known as cyclooxygenase or COX, is known to
be the principal target of nonsteroidal antiinflammatory
to be particularly interesting, and their pharmacological
drugs (NSAIDs). Recently, it has been found that there
profiles will be discussed.
are two isoforms of COX, each with a distinct physi-
ological role.1 One isoform, COX-1, is constitutively
Ch em istr y
produced in a variety of tissues and appears to be
The majority of the compounds listed in Table 1 were
prepared from a common intermediate, sulfone 28. This
intermediate was most efficiently prepared as outlined
in Scheme 1. 3-Ethoxycyclopentenone was treated with
bromine followed by triethylamine-mediated dehydro-
bromination to give 27 in 91% yield. 4-Bromothioanisole
was lithiated with nBuLi at -78 °C and then was
treated with 27 to give the 1,2 addition product, which
was converted to the unsaturated ketone on acidic
workup. Tungsten-catalyzed oxidation9 of the sulfide
then provided 28 in 71% yield for the two steps.
Substituted phenyl analogues 3-12 were prepared
from the appropriate boronic acids via palladium-
catalyzed Suzuki couplings with intermediate 28. The
base sensitivity of the cyclopentenone group prevented
the use of normal bases (e.g., Na2CO3) in these coupling
reactions. However, using 2 equiv of diethylamine in
refluxing 3:1:1 toluene/nPrOH/water generally provided
the product in good yield. In the case of the 3-pyridyl
analogue 13, the isolation of the required pyridine-3-
boronic acid was found to be problematic. As a result,
it was found that the Suzuki coupling could be carried
out very efficiently using the crude trialkylboronate.
Thus 3-bromopyridine was lithiated (nBuLi, ether, -78
°C) and then quenched with triisopropylborate (Scheme
important in the maintenance of normal physiological
functions including gastric cytoprotection. The second
isoform, COX-2, is induced by a wide variety of inflam-
matory stimuli and appears to be largely responsible
for the high-level production of prostaglandins that
results in inflammation.2 It has been proposed that a
selective COX-2 inhibitor could have useful antiinflam-
matory activity without the ulcerogenic side effects
associated with currently available NSAIDs, all of which
inhibit both COX-1 and COX-2.3 Indeed, evidence sup-
porting this hypothesis is now beginning to appear.4
Phenyl sulfone-containing tricyclic molecules have
proved to be a fertile area for the discovery of selective
inhibitors of COX-2. A number of central ring templates
have been successfully developed by ourselves5 and
others.6 Among these are the γ-lactone of MK-0966 (1,
rofecoxib, Vioxx)7 and the (trifluoromethyl)diazole of SC-
58635 (2, celecoxib, Celebrex).8
We wish to report here that 2,3-disubstituted cyclo-
pentenones also provide a useful template for the design
of COX-2-selective inhibitors. Two compounds in this
series, the 3,5-difluorophenyl derivative 7 (L-776,967)
and the 3-pyridyl derivative 13 (L-784,506), were found
† Merck Research Laboratories.
10.1021/jm980642l CCC: $18.00 © 1999 American Chemical Society
Published on Web 03/19/1999

2,3-Diarylcyclopentenones as COX-2 Inhibitors
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 7 1275
Sch em e 1a
Sch em e 4a
a
(a) Sia2BH, THF; (b) 28, Pd2(dba)3, PPh3, PhCH3/nPrOH/H2O.
Sch em e 5a
a
(a) Br2, Et3N, CHCl3; (b) 4-bromothioanisole, nBuLi, THF, then
27, 6 N HCl; (c) Na2WO4‚2H2O, H2O2, Aliquat 336, EtOAc/CH2Cl2;
(d) 3,5-difluorophenylboronic acid, Pd2(dba)3, PPh3, Et2NH, PhCH3/
nPrOH/H2O.
Sch em e 2a
a
(a) 4-(Methylthio)benzaldehyde, 3-benzyl-5-(2-hydroxyethyl)-
4-methylthiazolium chloride, Et3N, dioxane; (b) DBU, MeOH; (c)
Oxone, acetone/H2O.
tenone ring was synthesized by the intramolecular aldol
condensation of an appropriately substituted diketone
(Scheme 5). The lithium salt of phenoxyacetic acid was
treated with freshly prepared vinylmagnesium bromide
to give the vinyl ketone 32.10 A thiazolium salt-catalyzed
addition11 of (thiomethyl)benzaldehyde to 32 provided
diketone 33, which could be cyclized in the presence of
DBU to provide the desired 2-phenoxycyclopentenone
21.
a
(a) nBuLi, ether, -78 °C, (iPrO)3B; (b) 28, Pd2(dba)3, PPh3,
PhCH3/nPrOH/H2O.
Sch em e 3a
Resu lts a n d Discu ssion
Compounds were tested in initial screens for potency
against hCOX-2 in transfected CHO cells12 and against
hCOX-1 in U937 microsomes at low (100 nM) arachi-
donic acid concentration.13 Selected compounds were
subsequently tested in the human whole blood assay
against both COX-2 and COX-1.14 In vivo efficacy was
evaluated using a carageenen-induced rat paw edema
model.15
a
(a) Tf2O, iPr2NEt, CH2Cl2; (b) (Me3Sn)2, Pd2(dba)3, PPh3, LiCl,
PhCH3; (c) 28, Pd2(dba)3, AsPh3, NMP.
2). Removal of solvent provided the lithium trialkyl-
boronate 29 which was used directly in the Suzuki
coupling without the use of additional base. This method
obviated the problem of isolating the water-soluble
pyridineboronic acid. Similar couplings with the pyri-
dine trialkylboronates were used to provide analogues
15 and 17.
The first compound synthesized in this series, 3,
containing a 4-fluorophenyl group in the 2-postition, was
characterized by its high potency in COX-2 assays as
well as its efficacy in the rat paw edema model (ED50
)
For analogues 14, 16, and 18-20, where the required
boronic acids were deemed to be inaccessible, the
corresponding trimethylstannanes were prepared. These
were generally derived from the hydroxypyridine via the
triflate which was coupled with hexamethylditin as
illustrated for 4-chloro-2-pyridyl(trimethyl)stannane (30),
which was then coupled under Stille conditions (Pd2-
(dba)3, Ph3As, NMP, 100 °C) to provide analogue 20
(Scheme 3).
Styryl analogue 23 was prepared from (4-fluorophen-
yl)acetylene by treatment with disiamylborane and then
coupling the resulting crude vinylborane with 28 under
Suzuki conditions as described above (Scheme 4). Acety-
lenic derivatives 24 and 25 were prepared directly by
Heck coupling of 28 with phenylacetylene and tert-
butylacetylene, respectively (Pd(PPh3)2Cl2, Et3N, DMF,
60 °C).
3.0 mg/kg) (Table 1). However, its potency against
COX-1 (3.7 µM in the COX-1 whole blood assay) was
determined to be too great to provide an adequate safety
margin in our standard toxicology models at 100 mg/kg
(vide infra). Placing methyl substituents in either the
4- or 5-position of the cyclopentenone ring was not
productive: the former case led to a decrease in activ-
ity,16 while the latter substantially increased the COX-1
potency.17 We thus turned to modifications of the 2-aryl
moiety to achieve an acceptable level of potency and
selectivity.
Previous experience in other series has shown that
substitutions in the para position of this lower ring
increased potency against both COX-1 and COX-2, while
substitutions in the meta position increased the selec-
tivity of COX-2 inhibition.6d,7a As can be seen in Table
1, this trend generally held true in the cyclopentenone
series. Thus, in the highly sensitive U937 microsome
assay, the COX-1 potency decreases by 10-fold when the
Oxygen-linked compounds 21 and 22 could not be
prepared from intermediate 28. Instead, the cyclopen-

1276 J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 7
Black et al.
Ta ble 1. In Vitro and Rat Paw Edema Data for Diarylcyclopentenones
COX-2 (IC50, µM)
COX-1 (IC50, µM)
human
U937
human
rat paw edema
(ED50, mg/kg)d
entry
CHO cellsa
whole bloodb
microsomesc
whole bloodb
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
MK-0966
SC-58635
4-fluorophenyl
phenyl
0.02
0.5
1.0
1.7
0.05
0.5
3.1
6.6
1.7
11
9.2
0.3
0.7
2.5
0.1
34
19
1.7
3.2
3.0
10
0.002
0.019
0.011
0.014
0.014
0.015
0.014
0.009
0.015
0.006
0.014
0.17
0.087
0.30
0.44
0.15
1.53
6.3
3.7
4.1
0.08
0.19
0.34
0.21
0.62
0.28
0.09
0.15
0.93
0.08
0.64
1.5
3-fluorophenyl
6.4
3,4-difluorophenyl
3,5-difluorophenyl
3,5-dichlorophenyl
3-chloro-4-fluorophenyl
3-fluoro-4-chlorophenyl
3,4,5-trichlorophenyl
2-benzothiophenyl
3-pyridyl
5-chloro-3-pyridyl
5-bromo-3-pyridyl
4-methyl-3-pyridyl
4-methoxy-3-pyridyl
2-pyridyl
3.2
61
>100
2.6
>10
0.1
73
>100
1.7
1.3
>100
>100
5.5
0.06
>10
18
1.7
0.45
0.09
9.8
0.47
0.52
0.98
1.1
0.5
0.64
5.2
4.1
4-bromo-2-pyridyl
4-chloro-2-pyridyl
phenoxy
3,5-difluorophenoxy
4-fluorostyryl
0.63
0.19
27
11
2.7
34
2.5
0.011
0.017
0.175
0.006
0.15
94
27
6.6
>10
phenylacetylenyl
tert-butylacetylenyl
0.71
>100
a
b
Average of at least two independent determinations, each run in triplicate. Average of at least three determinations. c Average of
d
at least two independent determinations. ED50 values were determined using a minimum of four dose points, 5 animals/goup.
fluorine is moved from the para position (3) to the meta
position (5). When fluorine atoms are placed in both
meta positions (7), a further loss of COX-1 potency is
achieved. The intrinsic COX-2 potency as measured by
the CHO cell assay is not greatly affected by these
substituents, and while the COX-2 whole blood potency
is also decreased, it is still comparable to that of Vioxx
(1).
Several heterocycles were also examined. It was noted
that the 3-pyridyl moiety (13) provided a substantial
improvement in COX-2 selectivity. The potency against
COX-2 in the CHO cell assay was significantly reduced
relative to 7, but the potency in the human whole blood
assay was similar, and the compound was efficacious
in the rat paw edema model (ED50 ) 1.7 mg/kg).
Halogen substituents on the pyridine ring reduced the
potency against both COX-1 and COX-2. It is interesting
to note the effect of the 4-methoxy substituent on COX-1
potency (17). The IC50 of 60 nM for compound 17 in
U937 microsomes is the most potent COX-1 inhibition
observed in this series and shows the dramatic effect of
a para substituent on COX selectivity.
The 2-pyridyl moiety (18) showed relatively poor
activity against both enzymes, but the addition of a
p-chloro substituent (20) gave a dramatic improvement
in COX-2 potency and provided a compound with good
rat paw edema efficacy (ED50 ) 2.5 mg/kg). Phenoxy
(21), styryl (23), and acetylenyl (24) substituents provide
an interesting measure of the flexibility possible in this
position of the molecule but did not provide the required
level of selectivity for COX-2.
As a result of their characteristics of COX-2 potency,
selectivity, and rat paw edema efficacy, compounds 7
and 13 were chosen for further examination. It was of
immediate interest that while the whole blood COX-2
potencies of 13, 7, and MK-0966 were comparable (0.64,
0.62, and 0.5 µM, respectively), the intrinsic potency of
13 was nearly 10-fold less as measured in the CHO cell
assay (170 nM vs 26 nM and 20 nM). We originally
speculated that 13 might be a dual inhibitor, with
activity against another proinflammatory enzyme such
as p38 kinase, but screening in relevant assays did not
support this hypothesis.18 Further, no effect was seen
on COX-2 expression, TNF-R or IL-1â production in
human mononuclear cells.19 A more likely cause for the
discrepancy was a differential shift in potency by protein
present in the human whole blood assay. Indeed, when
MK-0966 was tested in a human blood mononuclear cell
assay in the presence of 10% human serum, it showed
an IC50 of 0.04 µM, 16-fold more potent than observed
in the protein-rich whole blood assay and comparable
to the value obtained in the CHO cell assay. In contrast,
compound 13 showed an IC50 of 0.62 µM in the human
blood mononuclear cell assay, not significantly different
from the 0.64 µM value obtained in the human whole
blood assay. In a separate protein-binding experiment,
13 was found to be 55% protein-bound in human
plasma, while MK-0966 was found to be 85% protein-
bound. Thus the favorable, low protein-binding charac-
teristics of 13 appear to compensate for its lower
intrinsic potency, thus giving an effective potency in
blood comparable to that of MK-0966.
Detailed kinetic studies were carried out on compound
7 to determine the mechanism of inhibition of COX-2.
These studies indicated that 7 is a time-dependent,
reversible inhibitor of COX-2 that forms a noncovalent
enzyme-inhibitor complex similar to other COX-2-
selective inhibitors which have been studied.20
Compounds 7 and 13 had substantially different
pharmacokinetics in the rat (Figure 1). Compound 7

2,3-Diarylcyclopentenones as COX-2 Inhibitors
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 7 1277
Ta ble 2. In Vivo Profiles
7
13
efficacy:
rat paw edema (ED50, mg/kg)a
2.7
0.8
3.0
4.0
1.7
0.7
0.7
0.6
rat pyresis (ED50, mg/kg)a
rat hyperalgesia (ID50, mg/kg)a
adjuvant arthritis, 21 day
(ID50, mg/kg)a
F igu r e 1. Rat plasma levels, 3 mg/kg po.
safety:
% change in rat liver weight,
chronic dosing (M/F, 4 days
at 100 mg/kg/day)c
+7/+9
+12/+15
rat GI tolerance, 51Cr leakage
(10 days at 100 mg/kg/day)
12/12 clean
11/12 clean
(1 death)
a
ED50 values were determined using a minimum of four dose
b
points, 5 animals/goup. ID50 values were determined using a
minimum of four dose points, 10 animals/goup. c Mean change
relative to control using 4 animals/group.
observed (7-15%). Finally, to test the hypothesis that
a selective COX-2 inhibitor would not be associated with
increased levels of gastrotoxicity, rats were treated with
either 7 or 13 at 100 mg/kg for 10 days and then given
an intravenous dose of 51Cr to test for any increased
gastric permeability.15 The amount of 51Cr in the feces
was determined to be indistinguishable from control for
all 12 rats treated with 7, indicating a high level of GI
tolerance. In contrast, a single dose of a nonselective
inhibitor such as diclofenac or indomethacin at 10 mg/
kg caused a significant increase in 51Cr excretion.
Eleven out of 12 of the rats treated with 13 were also
found to have an insignificant level of 51Cr excretion;
however one rat died during the course of the study due
to acute peritonitis. This appears to indicate that
compound 13 at high dosages is still associated with
some gastric toxicity. As the mean plasma concentration
of 13 observed in the surviving rats 1 h postdose was
185 µM, it is possible that COX-1 inhibition is a factor
at these high dosages.
F igu r e 2. Dog plasma levels, 3 mg/kg po.
showed 55% bioavailability in male rats and gave a 1.6
µM Cmax when dosed orally at 3 mg/kg. In contrast,
compound 13 gave 100% bioavailability with a 10 µM
Cmax when dosed under the same conditions. Addition-
ally, the observed half-life in rats based on intravenous
dosing was considerably longer for 13: 3 h vs 1 h for
compound 7. When dosed in male beagle dogs, the
pharmacokinetic differences between the two com-
pounds were much less apparent (Figure 2). Both 7 and
13 showed excellent oral bioavailability (90% and 84%,
respectively). In this species, compound 7 was found to
have the longer intravenous half-life: 4 h compared to
2 h for compound 13.
These studies have shown that a COX-2-selective
inhibitor can be a potent antiinflammatory, analgesic,
and antipyretic drug with a much reduced risk of GI
toxicity compared to conventional NSAIDs. Further, the
2,3-diarylcyclopentenone moiety has been shown to be
a useful template for the construction of such COX-2-
selective inhibitors. As a result of these studies, it was
decided to take compound 7 forward for evaluation in
preclinical safety assessment.
Both compounds were pursued in other in vivo models
of efficacy and tolerability that were at our disposal
(Table 2). Both 7 and 13 were efficacious in standard
rat models of inflammation, pyresis, and hyperalgesia,15
with 13 being slightly superior in all three models. In
the case of the rat adjuvant arthritis model,21 a chronic
inflammation model with b.i.d. dosing over 21 days,
compound 13 showed marked superiority, but these
data must be interpreted in light of the differing
pharmacokinetics between the two compounds. With
only a 1-h half-life in rats, compound 7 would not be
expected to provide protection against chronic inflam-
mation at doses that are efficacious in acute efficacy
models.
Exp er im en ta l Section
1
Ch em istr y. H NMR spectra were determined on Bruker
AM300, AMX300, or AMX400 spectrometers, and proton
chemical shifts are relative to tetramethylsilane as internal
standard. Elemental analyses were performed by Oneida
Research Services, Whitesboro, NY, or by the Universite´ de
Montreal, Department of Chemistry.
P r ep a r a tion of In ter m ed ia te 28. 2-Br om o-3-(4-(m eth -
ylsu lfon yl)p h en yl)-2-cyclop en ten -1-on e. Step 1. 2-Br om o-
3-eth oxy-2-cyclop en ten -1-on e (27). To a -15 °C solution of
3-ethoxy-2-cyclopenten-1-one (26) (165 g, 1.30 mol) in 1.8 L of
To evaluate the initial toxicology of these two com-
pounds, male and female rats were treated with either
7 or 13 at 100 mg/kg for 4 days and then examined for
changes in liver weight. Both compounds were well-
tolerated, with only minor liver weight increases being

1278 J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 7
Black et al.
CHCl3 was added a solution of bromine (219 g, 1.37 mol) in
CHCl3 (200 mL) via dropping funnel over 1.5 h to give a thick
yellow slurry. After 30 min of additional stirring, a solution
of Et3N (210 mL, 1.5 mol) in CHCl3 (300 mL) was added via
dropping funnel over 30 min. The resulting mixture was
concentrated, suspended in EtOAc, and filtered through a pad
of 2-cm Celite over 1-cm silica gel. The filtrate was concen-
trated and the resulting material recrystallized from 4:1 ether/
hexanes to provide 245 g of the title compound: 1H NMR
(CD3COCD3) δ 4.43 (2H, q), 2.95 (2H, m), 2.47 (2H, m), 1.39
(1H, t).
Step 2. 2-Br om o-3-(4-(m eth ylth io)p h en yl)-2-cyclop en -
ten -1-on e. To a -78 °C solution of 4-bromothioanisole (253
g, 1.25 mol) in THF (5 L) was added nBuLi (2.5 M in hexanes,
500 mL, 1.25 mol) over 20 min to give a thick slurry which
was stirred 2 h at -78 °C. A solution of 2-bromo-3-ethoxy-2-
cyclopenten-1-one (27) (242 g, 1.18 mol) in THF (1 L) was then
added via cannula, and the mixture was allowed to warm to
-10 °C; 6 N HCl (430 mL) was then added, and the resulting
mixture stirred 20 min. EtOAc (3 L) was added, and the
aqueous phase was separated. The organic layer was concen-
trated. The residue was suspended in ether and filtered to give
287 g of the title compound: 1H NMR (CD3COCD3) δ 8.12 (4H,
m), 3.22 (2H, m), 3.20 (3H, s), 2.69 (2H, m).
Step 3. 2-Br om o-3-(4-(m eth ylsu lfon yl)p h en yl)-2-cyclo-
p en ten -1-on e (28). To a solution of Na2WO4‚2H2O (6.7 g, 20.3
mmol) in water (70 mL) was added a few drops of concentrated
H2SO4 to give a solution of pH 6. This solution was added to
a room temperature suspension of 2-bromo-3-(4-(methylthio)-
phenyl)-2-cyclopenten-1-one (304.9 g, 1.07 mol) in EtOAc (3
L) and dichloromethane (300 mL). Aliquat 336 (22 g, 54.4
mmol) was added, and the mixture was warmed to 40 °C.
Hydrogen peroxide (30%, 350 mL, 3.1 mol) was added dropwise
to maintain an internal reaction temperature of approximately
50 °C (external cooling with a water bath was used). After the
addition was complete, stirring was continued for 3 h at 50
°C; then the aqueous phase was removed by cannula. Two
portions of warm water were added, stirred, and removed by
cannula. The organic phase was then concentrated to ap-
proximately 1 L, and the crystalline product was filtered to
provide 290.6 g (86%) of the title compound: 1H NMR (CD3-
COCD3) δ 8.12 (4H, m), 3.22 (2H, m), 3.20 (3H, s), 2.69 (2H,
m).
3,5-Diflu or op h en ylbor on ic Acid . To a -78 °C solution
of 1-bromo-3,5-difluorobenzene (50 g, 0.26 mol) in ether (860
mL) was added nBuLi (2.4 M in hexanes, 108 mL, 0.26 mol)
dropwise over 20 min. The resulting solution was stirred for
10 min and then treated with triisopropylborate (61 mL, 0.27
mol). The reaction mixture was warmed to 0 °C for 30 min
and then quenched with 1 M HCl. After stirring 30 min, the
mixture was partitioned between ethyl acetate and water. The
organic layers were washed with brine, dried over MgSO4,
filtered, and evaporated. The resulting product was dried
under high vacuum overnight to give 38 g of the title
compound: 1H NMR (CD3COCD3) δ 7.50 (2H, br s), 7.40 (2H,
m), 7.04 (1H, m).
δ 7.95 (2H, m), 7.66 (2H, m), 6.98 (1H, m), 6.80 (2H, m), 3.17
(2H, m), 3.12 (3H, s), 2.68 (2H, m). Anal. (C18H14F2O3S) C,H,N.
2-(4-F lu or op h en yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-2-cy-
clop en ten -1-on e (3): H NMR (CD3COCD3) δ 7.93 (2H, m),
7.63 (2H, m), 7.23 (2H, m), 7.07 (2H, m), 3.16 (2H, m), 3.14
(3H, s), 2.67 (2H, m). Anal. (C18H15FO3S) C,H,N.
2-P h en yl-3-(4-(m eth ylsu lfon yl)p h en yl)-2-cyclop en ten -
1-on e (4): 1H NMR (CD3COCD3) δ 7.90 (2H, m), 7.62 (2H, m),
7.31 (3H, m), 7.15 (2H, m), 3.14 (2H, m), 3.12 (3H, s), 2.65
(2H, m). Anal. (C18H16O3S) C,H,N.
1
2-(3-F lu or op h en yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-2-cy-
1
clop en ten -1-on e (5): H NMR (CD3COCD3) δ 7.93 (2H, m),
7.65 (2H, m), 7.34 (1H, m), 7.10 (1H, m), 6.95 (2H, m), 3.16
(2H, m), 3.13 (3H, s), 2.68 (2H, m); HRMS calcd for C18H16
-
FO3S 331.08042, found 331.08034. Anal. (C18H15FO3S‚1/2H2O)
C,H,N.
2-(3,4-Diflu or op h en yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-2-
cyclop en ten -1-on e (6): 1H NMR (CD3COCD3) δ 7.95 (2H, m),
7.65 (2H, m), 7.25 (2H, m), 6.95 (1H, m), 3.16 (2H, m), 3.12
(3H, s), 2.67 (2H, m); HRMS calcd for C18H14F2O3S 349.07099,
found 349.07115.
2-(3,5-Dich lor op h en yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-2-
cyclop en ten -1-on e (8): 1H NMR (CD3COCD3) δ 7.95 (2H, m),
7.68 (2H, m), 7.44 (1H, t), 7.18 (2H, d), 3.18 (2H, m), 3.12 (3H,
s), 2.68 (2H, m); HRMS calcd for C18H15Cl2O3S 381.01190,
found 381.01187. Anal. (C18H14Cl2O3S‚1/4H2O) C,H,N.
2-(3-Ch lor o-4-flu or oph en yl)-3-(4-(m eth ylsu lfon yl)ph en -
yl)-2-cyclop en ten -1-on e (9): 1H NMR (CD3COCD3) δ 7.95
(2H, m), 7.67 (2H, m), 7.38 (1H, dd), 7.25 (1H,t), 7.12 (1H, m),
3.17 (2H, m), 3.13 (3H, s), 2.18 (2H, m). Anal. (C18H14ClFO3S)
C,H,N.
2-(4-Ch lor o-3-flu or oph en yl)-3-(4-(m eth ylsu lfon yl)ph en -
1
yl)-2-cyclop en ten -1-on e (10): H NMR (CD3COCD3) δ 7.95
(2H, m), 7.65 (2H, m), 7.45 (1H, t), 7.18 (1H, dd), 6.98 (1H,
dd), 3.15 (2H, m), 3.12 (3H, s), 2.69 (2H, m). Anal. (C18H14
ClFO3S) C,H,N.
-
2-(3,4,5-Tr ich lor oph en yl)-3-(4-(m eth ylsu lfon yl)ph en yl)-
2-cyclop en ten -1-on e (11): 1H NMR (CD3COCD3) δ 7.97 (2H,
m), 7.80 (4H, m), 7.68 (1H, s), 7.32 (2H, m), 3.19 (3H, s), 3.15
(2H, m), 2.72 (2H, m). Anal. (C18H13Cl3O3S) C,H,N.
2-(2-Ben zoth iop h en yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-2-
1
cyclop en ten -1-on e (12): H NMR (CD3COCD3) δ 8.05 (2H,
m), 7.72 (2H, m), 7.35 (2H, s), 3.19 (2H, m), 3.14 (3H, s), 2.69
(2H, m); MS (CI, CH4) m/z 369 (M + 1, 100). Anal. (C20H16O3S2‚
1/2H2O) C,H,N.
Rep r esen ta tive P r oced u r e for Su zu k i Cou p lin g w ith
P yr id yltr ia lk ylbor on a tes: 3-(4-(Meth ylsu lfon yl)p h en yl)-
2-(3-p yr id in yl)-2-cyclop en ten -1-on e (13). Step 1. Lith iu m
3-P yr id in yltr iisop r op ylbor on a te (29). To a -73 °C solution
of 3-bromopyridine (960 g, 6.08 mol) in diethyl ether (20 L)
was added n-butyllithium (2.5 M in hexanes, 2.43 L, 6.08 mol)
over 4.5 h, maintaining an internal temperature below -70
°C. To the resulting thick yellow slurry was added triisopro-
pylborate (1165 g, 6.19 mol) over 2 h, again maintaining the
internal temperature below -70 °C. The resulting solution was
warmed to room temperature and concentrated until the
product began to separate (∼10 L). Hexane (6 L) was added
with stirring to give a gum. The remaining solvent was
removed and the residue placed under vacuum for 3 days to
give 1.7 kg of the crude salt as a light-brown solid. This
material was used in the next step without further purifica-
tion: 1H NMR (CD3OD, 400 MHz) δ 8.50 (1H, m), 8.15 (1H,
m), 7.85 (1H, m), 7.15 (1H, m), 1.15 (21H, s). (Alternatively,
the salt could be purified by evaporating twice from EtOH and
swishing in ether/hexanes to give lithium 3-pyridinyltrieth-
ylboronate as a white, hygroscopic crystalline solid.)
Rep r esen ta tive P r oced u r e for Su zu k i Cou p lin g w ith
P h en ylbor on ic Acid s: 2-(3,5-Diflu or op h en yl)-3-(4-(m eth -
ylsu lfon yl)p h en yl)-2-cyclop en ten -1-on e (7). A mixture of
2-bromo-3-(4-(methylsulfonyl)phenyl)-2-cyclopenten-1-one (28)-
(52.2 g, 166 mmol), 3,5-difluorophenylboronic acid (31.9 g, 202
mmol), tris(dibenzylideneacetone)dipalladium(0) (3.3 g, 3.6
mmol), and triphenylphosphine (1.89 g, 7.2 mmol) were
dissolved in toluene (800 mL) and nPrOH (250 mL) and
degassed. After the mixture stirred for 10 min, diethylamine
(21 mL, 203 mmol) and water (250 mL) were added. The
mixture was degassed again, heated to reflux for 1 h, then
cooled, and poured into EtOAc (2 L). The aqueous layer was
separated, and the organic layer was washed with 0.2 N NaOH
(500 mL), 0.5 N HCl (500 mL), and brine. The organic phase
was then dried over MgSO4, filtered through Celite, and
evaporated. The resulting light-brown solid was slurried with
hot EtOAc (250 mL), cooled, and filtered to give 45.8 g (75%)
of the title compound: mp 174-175 °C; 1H NMR (CD3COCD3)
Step 2. 3-(4-(Meth ylsu lfon yl)p h en yl)-2-(3-p yr id in yl)-2-
cyclop en ten -1-on e (13). A mixture of 2-bromo-3-(4-(methyl-
sulfonyl)phenyl)-2-cyclopenten-1-one (28) (120 g, 381 mmol),
lithium 3-pyridinyltriisopropylboronate (29) (122 g, 447 mmol),
and PPh3 (3.99 g, 15.2 mmol) was suspended in 3:1:1 toluene:
n-propanol:water (2.5 L). The mixture was degassed, and Pd2-
(dba)3 (6.95 g, 7.59 mmol) was added. The mixture was heated
to reflux for 3 h, then cooled, and diluted with EtOAc (1.5 L).

2,3-Diarylcyclopentenones as COX-2 Inhibitors
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 7 1279
The layers were separated, and the organic phase was washed
with water (1 L). The aqueous layers were back-extracted with
EtOAc, and the combined organic layers were washed with
water, dilute aqueous NaHCO3, and brine. The organic phases
were then filtered through Celite and concentrated to give 148
g of a black oil. Purification by flash chromatography (100%
EtOAc) followed by swishing in EtOAc gave 87.6 g (73%) of
the title compound: 1H NMR (CDCl3) δ 8.55 (1H, m), 8.34 (1H,
m), 7.40 (2H, m), 7.65 (1H, m), 7.49 (2H, m), 7.33 (1H, m),
3.12 (2H, m), 3.05 (3H, s), 2.80 (2H, m); MS (CI, CH4) m/z 314
(M + 1, 100).
The hydrochloride salt could be prepared: 40 g of the title
compound was dissolved in 2:1 CH2Cl2:hexanes (700 mL), and
a stream of HCl gas was bubbled through the solution, giving
an oil which separated. Excess HCl was flushed with a stream
of nitrogen. The oil solidified on standing, and the solvent was
decanted. The salt was crystallized from hot EtOH/iPrOH to
give 37.1 g of the hydrochloride salt: 1H NMR (DMSO-d6) δ
8.79 (1H, m), 8.61 (1H, m), 8.06 (1H, m), 7.93 (2H, m), 7.88
(1H, m), 7.65 (2H, m), 3.24 (3H, s), 3.17 (2H, m), 2.74 (2H, m).
Anal. (C17H15NO3S‚HCl‚H2O) C,H,N.
MS (CI, CH4) m/z 350 (30), 348 (M + 1, 100), 237 (20). Anal.
(C17H14ClNO3S) C,H,N.
2-(5-Ch lor o-3-p yr id in yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-
2-cyclop en ten -1-on e (14): 1H NMR (CD3COCD3) δ 8.50 (1H,
d), 8.20 (1H, d), 7.95 (2H, m), 7.22, (1H, dd), 7.18 (2H, m), 3.22
(2H, m), 3.16 (3H, s), 2.71 (2H, m); MS (CI, CH4) m/z 350 (38),
348 (M + 1, 100), 312 (5). Anal. (C17H14ClNO3S‚1/2H2O) C,H,N.
2-(4-Meth yl-3-p yr id in yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-
2-cyclop en ten -1-on e (16): 1H NMR (CD3COCD3) δ 8.20 (1H,
d), 7.94 (2H, m), 7.65 (2H, m), 7.47 (1H, dd), 7.20 (1H, d), 3.18
(2H, m), 3.14 (3H, s), 2.69 (2H, m); HRMS calcd for C18H18
NO3S 328.10074, found 328.10062.
-
2-(2-P yr id in yl)-3-(4-(m et h ylsu lfon yl)p h en yl)-2-cyclo-
p en ten -1-on e (18): 1H NMR (CD3COCD3) δ 8.46 (1H, m), 7.88
(2H, m), 7.80 (1H, m), 7.62 (2H, m), 7.45 (1H, m), 7.30 (1H,
m), 3.21 (2H, m), 3.12 (2H, m), 2.71 (2H, m); MS (CI, CH4)
m/z 314 (M + 1, 100), 100 (10).
2-(4-Br om o-2-p yr id in yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-
2-cyclop en ten -1-on e (19): 1H NMR (CDCl3) δ 8.58 (1H, d),
7.90 (3H, m), 7.49 (2H, m), 7.39 (1H, d), 3.10 (2H, m), 3.04
(3H, s), 2.79 (2H, m). Anal. (C17H14BrNO3S‚1/2H2O) C,H,N.
2-((E)-2-(4-Flu or oph en yl)eth en yl)-3-(4-(m eth ylsu lfon yl)-
p h en yl)-2-cyclop en ten -1-on e (23). To a 0 °C solution of
freshly prepared disiamylborane (from 2.5 mmol of BH3‚DMS)
in THF (4 mL) was added (4-fluorophenyl)acetylene (0.30 mL,
2.6 mmol). The solution was stirred for 1 h, then warmed to
room temperature, stirred for 1 h, and concentrated in vacuo.
The residue was then placed under high vacuum to romove
traces of DMS. 2-Bromo-3-(4-(methylsulfonyl)phenyl)-2-cyclo-
penten-1-one (28) (467 mg, 1.48 mmol), tris(dibenzylidene-
acetone)dipalladium(0) (86 mg, 0.094 mmol), and triphenylphos-
phine (98 mg, 0.37 mmol) were dissolved in toluene (15 mL),
nPrOH (10 mL), and water (8 mL). The freshly prepared
vinylborane was added as a solution in toluene (10 mL), and
the mixture was degassed. After the mixture stirred for 15
min, diethylamine (0.53 mL, 5.1 mmol) was added, and the
mixture was heated to reflux for 1.5 h. The mixture was then
cooled and partitioned between EtOAc and water. The organic
phase was washed with 1 M HCl and brine and dried over
MgSO4. Purification by flash chromatography (60% EtOAc/
hexanes), followed by crystallization from CH2Cl2/ether/hex-
anes, provided 253 mg of the title compound: 1H NMR
(CD3COCD3) δ 8.09 (2H, m), 7.92 (1H, d, J ) 15 Hz), 7.87 (2H,
m), 7.54 (2H, m), 7.10 (2H, m), 6.88 (1H, d, J ) 15 Hz), 3.18
(3H, s), 3.05 (2H, m), 2.62 (2H, m). Anal. (C20H17FO3S) C,H,N.
Rep r esen ta tive P r oced u r e for P r ep a r a tion of 2-P h e-
n oxy Der iva tives: 3-(4-(Meth ylsu lfon yl)p h en yl)-2-p h e-
n oxy-2-cyclop en ten -1-on e (21). Step 1. 1-(4-(Meth ylth io)-
p h en yl)-5-p h en oxy-1,4-p en t a d ion e (33). A mixture of
phenoxymethyl vinyl ketone (32)10 (1.00 g, 6.17 mmol), 4-
(methylthio)benzaldehyde (0.62 g, 4.1 mmol), 3-benzyl-5-(2-
hydroxyethyl)-4-methylthiazolium chloride (0.11 g, 0.41 mmol),
and triethylamine (0.34 mL, 2.44 mmol) in dioxane (20 mL)
was heated to 70 °C for 5 h and then to 100 °C for 18 h. The
reaction mixture was poured into water and extracted with
EtOAc. The organic phase was dried over Na2SO4 and con-
centrated. Purification by flash chromatography provided 140
mg of the desired diketone.
2-(5-Br om o-3-p yr id in yl)-3-(4-(m eth ylsu lfon yl)p h en yl)-
2-cyclop en ten -1-on e (15): 1H NMR (CD3COCD3) δ 8.59 (1H,
m), 8.24 (1H, m), 7.98 (2H, m), 7.86 (1H, m), 7.68 (2H, m),
3.23 (2H, m), 3.14 (3H, m), 2.70 (2H, m); MS (CI, CH4) m/z
394 (90), 392 (M + 1, 100), 314 (8), 237 (5). Anal. (C17H14
-
BrNO3S‚1/2H2O) C,H,N.
2-(4-Meth oxy-3-pyr idin yl)-3-(4-(m eth ylsu lfon yl)ph en yl)-
2-cyclop en ten -1-on e (17): 1H NMR (CD3COCD3) δ 7.98 (3H,
m), 7.67 (2H, m), 7.45 (1H, dd), 6.72 (1H, d), 3.86 (3H, s), 3.14
(5H, m), 2.65 (2H, m). Anal. (C18H17NO4S) C,H,N.
Rep r esen ta tive P r oced u r e for (Tr im eth ylsta n n yl)-
p yr id in e F or m a tion : 2-(Tr im eth ylsta n n yl)-5-ch lor op yr -
idin e (30). Step 1. Tr iflu or om eth a n esu lfon ic Acid 5-Ch lo-
r o-2-p yr id in yl Ester . To a -78 °C solution of 5-chloro-2-
hydroxypyridine (1.0 g, 7.72 mmol) and diisopropylethylamine
(1.88 mL, 10.81 mmol) in CH2Cl2 (35 mL) was added trifluo-
romethanesulfonic anhydride (1.56 mL, 9.26 mmol), and the
reaction mixture was allowed to warm to room temperature.
After 30 min, the mixture was washed with water and brine,
filtered through cotton, and concentrated to dryness. The
residue was purified by flash chromatography (8% EtOAc/
hexanes) to give 1.0 g of the title compound: 1H NMR (CD3-
COCD3) δ 8.50 (1H, d), 8.23 (1H, dd), 7.58 (1H, d).
Step 2. 2-(Tr im eth ylsta n n yl)-5-ch lor op yr id in e (30). To
a degassed room temperature mixture of trifluoromethane-
sulfonic acid 5-chloro-3-pyridinyl ester (0.510 g, 2.06 mmol),
hexamethylditin (0.443 mL, 2.16 mmol), lithium chloride
(0.262 g, 6.18 mmol), and a few crystals of butylated hydroxy-
toluene in dioxane (20 mL) was added a freshly prepared
solution of 0.1 M solution of Pd(PPh3)4 in toluene (0.807 mL,
0.081 mmol). The resulting mixture was heated to reflux for
1.5 h before it was cooled to room temperature, diluted with
EtOAc, washed with 10% NH4OH and brine, dried over
MgSO4, filtered, and concentrated to dryness. The residue was
used in step 3 without further purification.
Rep r esen ta tive P r oced u r e for Stille Cou p lin g w ith
(Tr im eth ylsta n n yl)p yr id in es: 2-(4-Ch lor o-2-p yr id in yl)-
3-(4-(m eth ylsu lfon yl)p h en yl)-2-cyclop en ten -1-on e (20).
To a degassed room temperature solution of 2-bromo-3-(4-
(methylsulfonyl)phenyl)-2-cyclopenten-1-one (28) (0.630 g, 2.0
mmol), Pd2(dba)3 (0.036 g, 0.04 mmol), and AsPh3 (0.098 g,
0.32 mmol) in NMP (5 mL) was added a degassed NMP
solution (8 mL) of 2-(trimethylstannyl)-5-chloropyridine (30)
(∼4.0 mmol). The resulting mixture was heated to 60 °C for
16 h and then to 100 °C for a further 2 h. The mixture was
then cooled to room temperature, diluted with EtOAc, washed
two times with 10% NH4OH and brine, dried over MgSO4, and
concentrated to dryness. The residue was purified by flash
chromatography (70% EtOAc/hexanes) followed by a CH2Cl2/
Et2O swish to provide 130 mg of the title compound: 1H NMR
(CD3SOCD3) δ 8.54 (1H, d), 8.00 (1H, dd), 7.89 (2H, m), 7.55
(2H, m), 7.45 (1H, d), 3.23 (3H, s), 3.13 (2H, m), 2.69 (2H, m);
Step 2. 3-(4-(Meth ylth io)p h en yl)-2-p h en oxy-2-cyclo-
p en ten -1-on e. To a solution of diketone 33 (70 mg, 0.22 mmol)
in MeOH (40 mL) was added 14 drops of DBU, and the solution
was stirred at 60 °C for 18 h. The reaction mixture was poured
into aqueous ammonium chloride and extracted with EtOAc.
The organic phase was dried over Na2SO4 and concentrated.
Purification by flash chromatography (20% EtOAc/hexanes)
provided 60 mg of the desired cyclopentenone.
Step 3. 3-(4-(Meth ylsu lfon yl)p h en yl)-2-p h en oxy-2-cy-
clop en ten -1-on e (21). To a solution of phenoxy-3-(4-(thio-
methyl)phenyl)-2-cyclopenten-1-one (60 mg, 0.20 mmol) in
acetone was added a solution of Oxone (0.45 g, 0.73 mmol) in
1.6 mL water. After stirring 1 h at room temperature, the
reaction mixture was diluted with water and extracted with
CH2Cl2. The organic phase was dried over Na2SO4 and
concentrated. Purification by flash chromatography provided

1280 J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 7
Black et al.
50 mg of the title compound: 1H NMR (CD3COCD3) δ 8.10
(4H, m), 7.20 (5H, m), 3.20 (2H, m), 3.15 (3H, s), 2.60 (2H, m);
HRMS calcd for C18H17O4S 329.0847, found 329.0847.
to compression of the carrageenan-injected paw was measured
1 h later by an algesiometer (Ugo-Basile, Italy).
Rat Adju van t-In du ced Ar th r itis Assay.21 Seventy 7-week-
old female Lewis rats (144-172 g) were used. Adjuvant-
induced arthritis (AIA) was induced in 6 groups of 10 rats each
by an intradermal injection of 0.5 mg of Mycobacterium
butyricum in light mineral oil in the left hind foot pad. Ten
rats were not injected and served as nonadjuvant controls.
Body weights, radiographs, and foot volumes of the noninjected
(secondary) paws were determined on various days (0, 14, and
21). Test compound (0.1, 0.3, 1.0, and 3.0 mg/kg/day po b.i.d.),
indomethacin (1 mg/kg/day po b.i.d.), and appropriate vehicles
were started on day 0 and continued thoughout the experi-
ment. Rats were euthanized by carbon dioxide inhalation on
day 21. Two-factor (‘treatment’ and ‘time’) analysis of variance
with repeated measures on ‘time’ was applied to the percent
changes for body weight and foot volumes and to the rank-
transformed radiographic total scores. A post hoc Dunnett’s
test was conducted to compare the effect of treatments to
vehicle.
P r elim in a r y Hep a totoxicity Stu d y in Ra ts. Four male
and four female rats (100-200 g) were used for each of three
groups: a vehicle control group (0.5% methylcellulose at 5 mL/
kg, used as the vehicle for all other dosage groups), a positive
control group (phenobarbitol/bezafibrate at 50/50 mg/kg/day),
and the test compound (100 mg/kg/day). All animals were
dosed by gavage once a day for 4 days, then were weighed,
and euthanized by carbon dioxide inhalation on the fifth day,
approximately 24 h after the final dose. The livers were
harvested and weighed. The weights as a fraction of total body
weight were then compared to those of the vehicle-control
group.
2-(3,5-Diflu or op h en oxy)-3-(4-(m eth ylsu lfon yl)p h en yl)-
2-cyclop en ten -1-on e (22): 1H NMR (CD3COCD3) δ 8.10 (4H,
m), 6.70 (3H, m), 3.20 (2H, m), 3.15 (3H, s), 2.80 (2H, m);
HRMS calcd for C18H15F2O4S 365.0659, found 329.0657. Anal.
(C18H14F2O4S) Calcd: C, 59.33; H, 3.87. Found: C, 58.86; H,
3.79.
Rep r esen ta tive P r oced u r e for Heck Cou p lin g w ith
Acetylen es: 2-(2-P h en yleth yn yl)-3-(4-(m eth ylsu lfon yl)-
p h en yl)-2-cyclop en ten -1-on e (24). To a suspension of 2-bro-
mo-3-(4-(methylsulfonyl)phenyl)-2-cyclopenten-1-one (28) (500
mg, 1.58 mmol) and dichloropalladium(II) bistriphenylphos-
phine (56 mg, 0.08 mmol) in triethylamine (13 mL) and DMF
(5 mL) was added phenylacetylene (0.26 mL, 2.4 mmol), and
the mixture was degassed. The reaction was heated to 60 °C
for 90 min, giving an orange solution. This solution was cooled
and partitioned between 1 M HCl and dichloromethane. The
aqueous layer was washed with 1 M HCl, water, and brine,
filtered through cotton, and concentrated. Purification by flash
chromatography (60% EtOAc/hexanes) provided 36 mg of the
title compound: 1H NMR (CD3COCD3) δ 8.48 (2H, m), 8.13
(2H, m), 7.60 (2H, m), 7.42 (3H, m), 3.28 (2H, m), 3.20 (3H, s),
2.68 (2H, m); HRMS calcd for C20H17O3S 337.08984, found
337.08973. Anal. (C20H16O3S‚1/2H2O) C,H,N.
2-(3,3-Dim eth yl-1-bu tyn yl)-3-(4-(m eth ylsu lfon yl)ph en yl)-
2-cyclop en ten -1-on e (25): 1H NMR (CD3COCD3) δ 8.43 (2H,
m), 8.06 (2H, m), 3.17 (5H, m), 2.57 (2H, m), 1.34 (9H, s);
HRMS calcd for C18H21O3S 317.12114, found 317.12124. Anal.
(C18H20O3S) C,H,N.
51Cr F eca l Excr etion Assa y in Ra ts.15 Male Sprague-
Dawley rats (150-200 g) were administered orally a test
compound either once (acute dosing) or b.i.d. for 5 days (chronic
dosing). Immediately after the administration of the last dose,
the rats were injected via a tail vein with 0.5 mL of 51Cr-labeled
red blood cells from a donor rat. The animals were placed
individually in metabolism cages with food and water ad lib.
Feces were collected for a 48-h period, and 51Cr fecal excretion
was calculated as a percent of total injected dose. 51Cr-labeled
red blood cells were prepared using the following procedures:
10 mL of blood was collected in heparinized tubes via the vena
cava from a donor rat. Plasma was removed by centrifugation
and replenished with equal volume of HBSS. The red blood
cells were incubated with 400 Ci of sodium chromate-51 for
30 min at 37 °C. At the end of the incubation, the red blood
cells were washed twice with 20 mL of HBSS to remove free
sodium chromate-51. The red blood cells were finally recon-
stituted in 10 mL of HBSS, and 0.5 mL of the solution (about
20 Ci) was injected per rat.
Mea su r em en t of P la sm a Levels in Ra ts. For po studies,
the drug was suspended in 1% methocel by mixing with a
plantary micromill (Fritsch Pulverizette 7) for 10 min. Rats
were dosed by gavage at 3 mg/kg with a dose volume of 10
mL/kg. Blood samples (1 mL) were taken at 0, 0.25, 0.5, 1, 2,
4, and 6 h after dosing. For iv studies, the drug was dissolved
in 60% PEG-200 and injected intravenously into the jugular
vein at a dose of 3 mg/kg with a dose volume of 1 mL/kg. Blood
samples (1 mL) were taken at 0, 0.08, 0.5, 1, 2, 4, and 6 h
after dosing. The blood was centrifuged and the plasma
collected. To 150 µL of each plasma sample was added an equal
volume of acetonitrile to precipitate the protein. The sample
was centrifuged, and a 100-µL aliquot of the supernatant was
analyzed by reverse-phase HPLC on a NovaPak C18 column
(3.9 × 150 mm). The solvent system used for compound 7 was
33% CH3CN in aqueous NH4OAc (1 g/L) at a flow rate of 1
mL/min with UV detection at 270 nM. The solvent system used
for compound 13 was 20% CH3CN in aqueous NH4OAc (1 g/L)
at a flow rate of 1 mL/min with UV detection at 275 nM. The
area of the resulting peak was quantified by comparison to a
standard (plasma sample at time 0 spiked with the parent
compound corresponding to a plasma concentration of 6.7 µg/
mL).
Biologica l Met h od s. The hCOX-2 transfected CHO cell
assay,12 the U937 microsome assay,13 and the COX-1 and
COX-2 human whole blood assays14 have been described
previously.
Ra t P a w Ed em a Assa y.15 Male Sprague-Dawley rats
(150-200 g) were fasted overnight and were given, po, either
vehicle (1% methocel or 5% Tween 80) or a test compound.
After 1 h, a line was drawn using a permanent marker at the
level above the ankle in one hind paw to define the area of
the paw to be monitored. The paw volume (V0) was measured
using a plethysmometer (Ugo-Basile, Italy) based on the
principle of water displacement. The animals were then
injected subplantarly with 50 µL of 1% carrageenan solution
in saline (FMC Corp., Maine) into the paw using an insulin
syringe with a 25-gauge needle (i.e., 500 µg of carrageenan/
paw); 3 h later, the paw volume (V3) was measured, and the
increases in paw volume (V3 - V0) were calculated. The
animals were sacrificed by CO2 asphyxiation, and the absence
or presence of stomach lesions was scored. Data were compared
with the vehicle-control values, and percent inhibition was
calculated. All treatment groups were coded to eliminate
observer bias.
Ra t P yr esis Assa y.15 Male Sprague-Dawley rats (150-
200 g) were fasted overnight, and the baseline rectal temper-
ature was recorded using a flexible temperature probe (YSI
series 400) connected to a digital thermometer (model 08502,
Cole Parmer). At time 0, the rats were injected interperito-
neally with either saline or LPS (0.36 mg/kg), and the rectal
temperature was measured 5, 6, and 7 h following LPS
injection. After the measurement at 5 h, when the increase in
rectal temperature had reached a plateau, the LPS-injected
rats were given orally either the vehicle or a test compound
to determine whether the rise in temperature could be
reversed. Percent reversal (antipyretic activity) was calculated
using the rectal temperature obtained at 7 h, taking this value
in the vehicle-control group as zero reversal.
Ra t Hyp er a lgesia Assa y.15 Hyperalgesia to mechanical
compression of the hind paw of male Sprague-Dawley rats
(90-110 g) was induced by intraplantar injection of carra-
geenan (4.5 mg/paw) 3 h previously. A test compound was
given orally 2 h after carrageenan. The vocalization response

2,3-Diarylcyclopentenones as COX-2 Inhibitors
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 7 1281
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