Synthesis and SAR of a new series of COX-2-selective inhibitors: Pyrazolo[1,5-α]pyrimidines

Article abstract of DOI:10.1021/jm0009383

The synthesis and pharmacological activity of a series of bicyclic pyrazolo[1,5-α]pyrimidines as potent and selective cyclooxygenase-2 (COX-2) inhibitors are described. The new compounds were evaluated both in vitro (COX-1 and COX-2 inhibition in human whole blood) and in vivo (carrageenan-induced paw edema and air-pouch model). Modification of the pyrimidine substituents showed that 6,7-disubstitution provided the best activity and led to the identification of 3-(4-fluorophenyl)-6,7-dimethyl-2-(4-methylsulfonylphenyl) pyrazolo[1,5-α]pyrimidine (10f) as one of the most potent and selective COX-2 inhibitor in this series.

Full text of DOI:10.1021/jm0009383

350
J . Med. Chem. 2001, 44, 350-361
Syn th esis a n d SAR of a New Ser ies of COX-2-Selective In h ibitor s:
P yr a zolo[1,5-a ]p yr im id in es
Carmen Almansa,* Alberto F. de Arriba, Fernando L. Cavalcanti, Luis A. Go´mez, Agust´ı Miralles,
Manuel Merlos, J ulia´n Garc´ıa-Rafanell, and J avier Forn
Research Center, J . Uriach & Cı´a. S.A., Dega` Bahı´ 59-67, 08026 Barcelona, Spain
Received August 7, 2000
The synthesis and pharmacological activity of a series of bicyclic pyrazolo[1,5-a]pyrimidines
as potent and selective cyclooxygenase-2 (COX-2) inhibitors are described. The new compounds
were evaluated both in vitro (COX-1 and COX-2 inhibition in human whole blood) and in vivo
(carrageenan-induced paw edema and air-pouch model). Modification of the pyrimidine
substituents showed that 6,7-disubstitution provided the best activity and led to the identifica-
tion of 3-(4-fluorophenyl)-6,7-dimethyl-2-(4-methylsulfonylphenyl)pyrazolo[1,5-a]pyrimidine
(10f) as one of the most potent and selective COX-2 inhibitor in this series.
In tr od u ction
fonyl group can only be replaced by an SO2NH2,whereas
the second phenyl group accepts lower alkyl or halogen
substituents, the most common being 4-fluoro and
4-methyl. Two compounds in this class, celecoxib14 (2)
and rofecoxib15 (3), are already in the market for the
treatment of acute pain, osteoarthritis, and rheumatoid
arthritis and seem to be as effective as other established
NSAIDs and to have much better gastric tolerance.16
In addition they are undergoing clinical trials for the
treatment of certain forms of cancer and Alzheimer’s
disease.
Nonsteroidal antiinflammatory drugs (NSAIDs) exert
their antiinflammatory action mainly through inhibition
of cyclooxygenase (COX), one of the key enzymes in the
arachidonic acid (AA) cascade.1 This enzyme bis-oxygen-
ates AA to PGG2, which is subsequently degraded to
vasoactive and inflammatory mediators such as pros-
taglandins (PGs), prostacyclin (PGI2), and tromboxane
A2. The therapeutic use of NSAIDs has been associated
with well-known side effects at the gastrointestinal level
(mucosal damage, bleeding)2 and, less frequently, at the
renal level3, and thus their use in chronic diseases such
as rheumatoid arthritis has caused complications.
At the beginning of the 1990s two COX isoforms were
discovered:4 one (COX-1) constitutively present in many
tissues such as stomach, kidney, and platelets and the
other (COX-2) cytokine inducible and expressed mainly
in a wide range of inflammatory cells. This scenario led
to the recognition5 that selective COX-2 inhibitors could
provide antiinflammatory agents devoid of the undesir-
able effects associated with classical, nonselective
NSAIDs.
Several approaches based on modification of estab-
lished nonselective agents have been studied for the
design of COX-2-selective inhibitors,6 in a line of re-
search in which the crystal resolution of the structures
of both enzymes has also been somewhat helpful.7
Lengthening the carboxyl side chain of indomethacin8
(1) or substituting it with other groups9 and modifying
the preferential inhibitor nimesulide to flosulide,10
L-745337,11 and NS-39812 has led to selective com-
pounds, although none of them is being pursued for a
variety of reasons. Based on the structure of early
known antiinflammatory agents such as flumizole, a
wide range of diarylheterocycle COX-2-selective inhibi-
tors have been described.13 To obtain good activity and
selectivity such compounds require the presence of a
4-methylsulfonylphenyl group attached to an unsatur-
ated (generally five-membered) ring in which an ad-
ditional vicinal phenyl ring is present. The methylsul-
In a new approach to diarylheterocycle COX-2-selec-
tive inhibitors, we decided to investigate the attachment
of an additional six-membered ring to the usual five-
membered central ring. The pyrazolo[1,5-a]pyrimidine
nucleus I was selected for the SAR study in view of the
easier access to substituent variation in the extra ring
in comparison to other possible frameworks, which in
turn could be prepared once the key features of I were
identified. Two similar approaches taken with imidazo-
[1,2-a]pyridines17 and isoindoles18 have appeared in the
patent literature.
Ch em istr y
The basic SAR around the pyrazolopyrimidines I was
effected with the 3-(4-fluorophenyl)-2-(methylsulfo-
* To whom correspondence should be addressed. Tel: 933471511.
Fax: 934560639. E-mail: rd@uriach.com.
10.1021/jm0009383 CCC: $20.00 © 2001 American Chemical Society
Published on Web 01/04/2001

New Series of COX-2-Selective Inhibitors
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3 351
Sch em e 1a
or in toluene (methods B, D). In the remaining cases,
the presence of masked carbonyl groups in the reactants
required acidic conditions and one of methods A (HCl/
ZnCl2/EtOH), C (HCl/EtOH), E (AcOH), or G (p-TsOH/
toluene) was used. The 6-bromo derivative 10a b (Table
2) was obtained from 3-bromo-4-diethylamino-3-buten-
2-one (20, R2 ) Br, R3 ) Me) using HBr (method F),
since the standard HCl gave its 6-chloro analogue,
product of a halogen exchange.
The parent compound 10a (Table 1) was obtained
using the diprotected malonaldehyde 12 in the presence
of ZnCl2 (method A), while the 6-methyl 10d was
obtained from 14 under the conditions of method C. The
isomeric 5-methyl 10b and 7-methyl 10c were obtained
as a 1:4 mixture by reaction of 9a with 13 (R3 ) Me)
under the conditions of method B. The structural
a
(i) Thioanisole, AlCl3, CH2Cl2, -10 °C, 1 h; (ii) 1. POCl3/DMF,
1
assignment was made on the basis of their H and 13C
CHCl3, 80 °C, 18 h, 2. NH2OH‚HCl, DMF, 20 °C, 4 h; (iii)
NH2NH2‚H2O, EtOH, 78 °C, 18 h; (iv) Oxone, MeOH/THF, 20 °C,
1 h.
NMR spectra. The methyl groups of 10b and 10c
showed proton shifts of 2.63 and 2.88 ppm, respectively,
and carbon shifts of 26.5 and 18.5 ppm, respectively, in
accordance with results previously reported for related
compounds.20a,b The structural identity of 10b,c was
further confirmed by additional chemical trans-
formations.20c,d Hence, 10s was converted to 10b upon
hydrogenolysis with H2 over Pd/C/NaOAc,20c whereas
10x was converted to 10c upon decarboxylation with
H2SO4, using the same conditions described below for
the preparation of 10j.
nylphenyl) derivatives 10, which could be prepared from
amine 9a . Amines 8 and 9 were obtained from ketones
4 and 519 in a process involving reaction with POCl3/
DMF followed by treatment with hydroxylamine in
DMF to give a 3-chloropropenenitrile intermediate (6,
7), which was cyclized with hydrazine in ethanol
(Scheme 1).
The cyclization of 3-aminopyrazoles with a 1,3-dicar-
bonyl compound is the usual method for preparing
pyrazolopyrimidines.20 When a nonsymmetrical reagent
is used, usually isomeric mixtures are obtained. The
proportion of isomers is quite erratic and depends on
the substrate, the nature of the reagent, and the type
of groups used to mask the carbonyl compounds.20a,b
As outlined in Scheme 2, compounds 10 were obtained
by cyclization of 9a with an appropriately substituted
free or masked 1,3-dicarbonyl derivative (11-17, 19-
21), under conditions A-G indicated for each compound
in Tables 1-3. When a free dicarbonyl derivative (11
or 15) was used, or in the case of the ethoxymethylene
derivatives 13, the reactions were run either in ethanol
The 5,7-dimethyl derivative 10e was prepared from
9a and acetylacetone (11, R2 ) H, R1 ) R3 ) Me),
whereas the isomeric 10f,g were obtained in different
proportions depending on the reactant employed. Thus,
the R-hydroxymethylene derivative 21 (R2 ) R3 ) Me)
led to 10f,g in a 1:9 ratio, while the R-methoxymethyl-
ene derivative 19 (R2 ) R3 ) Me)21 led to 10f,g in a 9:1
ratio. In both cases the most abundant isomer was
isolated by silica gel chromatography in 62% (10g) and
70% (10f) yield. The assignment of the structure to 10g
was based on its unambiguous synthesis by hydro-
genolysis of the 7-chloropyrazolopyrimidine 10t over Pd/
Sch em e 2a
a
(i) Method A: HCl, ZnCl2, EtOH, 78 °C, 1 h; Method B: piperidine, EtOH, 78 °C, 18 h; Method C: HCl, EtOH, 78 °C, 1-18 h; Method
D: toluene, 111 °C, 8 h; Method E: AcOH, 117 °C, 1 h; Method F: HBr, EtOH, 78 °C, 1 h; Method G: p-TsOH, toluene, 111 °C, 8 h.
Specific methods and reagents used are indicated in Tables 1-3.

352 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3
Ta ble 1. Methylpyrazolopyrimidines of Formula 10
Almansa et al.
a
Numbers indicate the type of reagent used as indicated in Scheme 2 and letters the cyclization method used for the preparation of
b
compounds, as described in the Experimental Section. Isomeric ratio of 7- vs 5-monosubstituted, 6,7- vs 5,6-disubstituted, or 5,7- vs
7,5-disubstituted compounds. c Percentage of inhibition of COX-1 (10 µM) or COX-2 (10 and 1 µM) activity in human whole blood. Assays
d
were performed in duplicate or repeated, so that standard errors were inferior to 10%. IC50 for whole cell inhibition in U-937 cells
(COX-1) and 143982 cells (COX-2). Assays were performed in duplicate. e 82.3% inhibition at 1 µM.
Sch em e 3a
1
C/NaOAc.20c It is noteworthy that the H NMR spectra
of the two isomers were characterized by the chemical
shift of the 7-methyl (10f) at 2.85 ppm and the 5-methyl
(10g) at 2.60 ppm. The downfield shift generally ob-
served for the 7- over the 5-substituted derivatives
allowed the identification of the remaining compounds
of Tables 2 and 3.
As can be seen from the results outlined in Tables 2
and 3, where the observed isomeric ratios are indicated
for every compound, the 7-substituted and 6,7-disub-
a
(i) 19 (R2 ) R3 ) Me), HCl, EtOH, 78 °C, 4 h; (ii) 1. MCPBA,
stituted isomers are favored over their 5-substituted and
5,6-disubstituted counterparts. This result is consistent
with the proposed mechanism for this reaction, in which
the most reactive carbonyl group (in this case the
masked aldehyde function) first interacts with the
amino group of the aminopyrazole, after which cycliza-
tion takes place.
Some compounds were obtained by further transfor-
mations of the pyrimidine substituents. Hence, the
7-hydroxy derivative 10q was converted to the methoxy
10r with methyl sulfate and to the chloro 10s on
treatment with POCl3. Compound 10t was prepared
from its hydroxyl counterpart in an analogous way.
Standard ester hydrolysis provided acids 10n ,x,a f.
Decarboxylation at position 6 by refluxing in H2SO4 for
18 h provided access to 7-monosubstituted derivatives
such as 10j. The preparation of the monohydroxymethyl
derivatives 10y,a d was more difficult. Attempts to
reduce 10w with LiAlH4 or NaBH4 were unsuccessful,
CH2Cl2, 20 °C, 2 h, 2. Ac2O, AcONa, 140 °C, 8 h, 3. MMPP, CH2Cl2/
MeOH, 20 °C, 18 h, 4. 1 N NaOH, THF/MeOH, 20 °C, 1 h, 5.
NH2OSO3H, AcONa, H2O, 20 C, 18 h.
since partial saturation of the pyrimidine ring was
observed while the ethoxycarbonyl group was unaf-
fected. However, using diisobutylaluminum hydride
(DIBAL) in THF, 10y could be prepared in good yield.
Various approaches were studied for the preparation of
10a d , but only the oxidation of 10f with KMnO4/NEt3/
22
H2SO4 allowed its isolation in moderate yields. The
fluoromethyl derivative 10a a was obtained upon treat-
ment of 10y with diethylaminosulfur trifluoride (DAST).
Other 6,7-dimethyl sulfones (23) were prepared from
the corresponding amines 9 as described for 10f. The
sulfonamides 26 (Table 4, Scheme 3) were prepared
from the methylthio derivatives 22 using a procedure23
which involves monooxidation to the sulfoxide, Pum-
merer rearrangement to the acetoxymethylthio com-
pound 24, and oxidation with magnesium monoperoxy-

New Series of COX-2-Selective Inhibitors
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3 353
Ta ble 2. Pyrazolopyrimidines of Formula 10
d
a,b,e See footnotes a-c of Table 1. c Melting points correspond generally to chromatographed products. Elemental analyses for C, H,
g
N, and S were within 0.4% of the theoretical values indicated. f Mixture of regioisomers. See text and Experimental Section for method
h
i
j
k
l
of synthesis. 6.8% inhibition at 1 µM. 0% inhibition at 1 µM. 53.9% inhibition at 1 µM. 18.9% inhibition at 1 µM. 44.6% inhibition
at 1 µM.
phthalate (MMPP), followed by treatment with NaOH
to give the sodium sulfinate 25. The reported process
involved subsequent treatment with sulfuryl chloride
and ammonia, but we found that on treatment of 25
with hydroxylamine O-sulfonic acid and NaOAc in H2O,
the sulfonamides 26 were produced in one step and in
somewhat better yields.
Triazines 29 and 33 (Table 5) were prepared as
described in Scheme 4. Formation of the diazonium salt
of amine 9a with NaNO2 followed by treatment24 with
the in situ generated â-keto acids from 28 afforded the
[1,2,4]triazines 29. The [1,3,5]triazines 33 were obtained
from 9a upon treatment25 with a suitable ethyl imidate
30, followed by boiling in a triethyl orthoacylate 32.

354 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3
Ta ble 3. Cyclopyrazolopyrimidines of Formula 10
Almansa et al.
a-e See footnotes a-e of Table 2.
Ta ble 4. 6,7-Dimethylpyrazolopyrimidines of Formulas 23 and
26
pyridine. On the other hand, the R-bromo ketone 39,
obtained from ketone 5a upon treatment with bromine
in CH2Cl2/AcOH, was reacted with the corresponding
arylamines 40, 42, and 44 in DMF at 60 °C26 to give
imidazo[1,2-b]pyridazines 41, imidazo[1,2-a]pyrazines
43, and imidazo[1,2-a]pyrimidines 45. Amines 40, 42,
and 44 were commercially available or were prepared
according to reported procedures (see Experimental
Section). The Glaxo-reported bicycle 46 was also pre-
pared from 39 as described.17
Resu lts a n d Discu ssion
The new compounds were tested in vitro for their
ability to inhibit COX-1 and COX-2 activity in a human
whole blood (HWB) assay.27 Percent inhibition at a 10
µM concentration was initially determined, and active
compounds were also tested at 1 µM (Tables 1-5). All
compounds showing more than 60% inhibition at 10 µM
were tested in vivo in the rat carrageenan-induced paw
edema assay (CPE)28 at a 30 mg/kg dose; some of them
were also tested in the carrageenan-induced air-pouch
model29 to estimate PG production at a 1 mg/kg dose.
The main SAR was carried out with the 3-(4-fluoro-
phenyl)-2-(4-methylsulfonylphenyl) derivatives 10, which
are characterized by a 4-methylsulfonylphenyl group
vicinal to a phenyl ring similarly to other COX-2-
selective inhibitors. The gradual introduction of methyl
groups into the poorly active parent unsubstituted
compound (10a ) was initially explored. In this initial
series the activity in human cell lines expressing COX-1
a
Physicochemical properties described in the Experimental
b
Section. See footnote c of Table 1.
Other bicycles (Table 6) were prepared as described
in Schemes 5 and 6. Imidazo[4,5-c]pyridines 38 were
obtained from the corresponding 4-hydroxy-3-nitropy-
ridines 34 upon treatment with POCl3, followed by
nucleophilic displacement with 4-fluoroaniline, reduc-
tion to the amine 37 with SnCl2 in EtOH, and final
cyclization with 4-methylsulfonylbenzoyl chloride in

New Series of COX-2-Selective Inhibitors
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3 355
Sch em e 5a
Ta ble 5. Pyrazolotriazines of Formulas 29 and 33
a
(i) POCl3, DMF, toluene, 110 °C, 7 h; (ii) 4-fluoroaniline, NEt3,
CHCl3, 67 °C, 48 h; (iii) SnCl2, EtOH, 78 °C, 45 min; (iv)
4-methylsulfonylbenzoyl chloride, pyridine, 115 °C, 18 h.
Sch em e 6a
a,b
See footnotes a,b of Table 4.
Sch em e 4a
a
(i) NaNO2, H2O, 0 °C, 1 h; (ii) 1. KOH, H2O, 0 °C, 24 h, 2.
a
(i) Br2, CH2Cl2, AcOH, 20 °C, 18 h; (ii) DMF, 60 °C, 18-36 h;
(iii) H2, Pd/C, TEA, 200 psi, 18 h.
NaOAc, HCl, H2O, 0-20 °C, 1 h; (iii) AcOH, 20 °C, 18 h; (iv) reflux,
18 h.
related quite well with those obtained in the whole blood
assay. The most potent compounds in the whole cell
assay, 10c,f, gave also the highest inhibitions in HWB.
When their IC50 in HWB were calculated (10c: COX-1
) 10 µM, COX-2 ) 0.6 µM; 10f: COX-1 > 10 µM, COX-2
) 0.08 µM) the superior activity shown at 1 µM by 10f
was confirmed. This compound had excellent in vitro
activity, which compared well with celecoxib (2), whether
in whole cells (Table 1) or in HWB (2 IC50: COX-1 ) 13
µM, COX-2 ) 0.6 µM).
and COX-230 was also evaluated (Table 1). While
substitution at the 5- or 6-position provided some
increase in whole cell activity, a substantial gain was
observed for the 7-methyl derivative 10c. Its activity
was maintained with a second methyl group in position
6 (10f) and impaired by introduction of additional
methyl groups at the 5 (10e) or 5,6 (10h ) positions.
These results, together with the poor activity of the 5,6-
disubstituted compound 10g, indicated the need for
substitution at position 7, which then tolerated a second
substituent at position 6. The whole cell results cor-
Among the methylated derivatives only compound 10f
showed some in vivo activity. In the air-pouch model it

356 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3
Almansa et al.
Ta ble 6. Heterocycle Variations: Compounds of Schemes 5
showed 74% inhibition of PGE2 production at 1 mg/kg,
approximately one-third of that shown by 2 (93.4% and
71.3% inhibition at 1 and 0.3 mg/kg, respectively). In
the CPE model 10f provided around 20% of edema
inhibition at 30 as well as at 10 mg/kg, in comparison
to 43% shown by 2 at 10 mg/kg. In initial pharmacoki-
netic evaluation in the rat, compound 10f had a short
half-life (1 h) and poor bioavailability.
and 6
With the aim of improving these results and keeping
the 7-position substituted, we introduced further varia-
tion. Changing the methyl of 10c for CF3 (10i) or
lengthening to propyl (10j) or phenyl (10k ) clearly
diminished activity. Replacing the 5-methyl of 10e by
CF3, COOEt, or COOH (10l-n ) was clearly detrimental,
while lengthening it to butyl provided the more active
albeit less selective compound 10o. Substitution of the
7-methyl of 10e by butyl, hydroxy, methoxy, or chloro
(10p -t) produced a complete loss in activity. Next, some
6,7-disubstituted analogues of 10f were prepared and
evaluated. Replacing the 6-methyl group by ethyl (10u ),
acetyl (10v), CF3 (10z), or Br (10a b) maintained activity
but provided some decrease in selectivity, whereas
replacement by COOH, COOEt, CH2OH, or CH2F
(10w -y,a a ) was more detrimental. Elongation of the
7-methyl group to ethyl provided a loss in selectivity
(10a c), while the 7-phenyl (10a e) completely lost po-
tency, as in the case of 10k . The 7-hydroxymethyl
derivative, which was prepared together with 10y as a
possible metabolite of 10f, retained its potency. Among
these derivatives only the ethyl ester 10w and the
monohydroxylated 10y showed some in vivo activity in
the CPE test (20.2 and 17.3% inhibition at 30 mg/kg,
respectively), somewhat inferior to that of 10f.
a,b
See footnotes a,b of Table 4.
The HWB activity of the 6,7-dimethyl-1,3,5-triazine 29a
was inferior to that of 10f, although in this case it was
improved upon lengthening to the 6-ethyl derivative
29c. However, a second ethyl group (29f) diminished
selectivity. The 7-propyl 29b was more active than its
pyrimidine counterpart 10j, but further lengthening
(29e) or introduction of a methyl group (29d ) failed to
improve this result. On the other hand, the 1,3,4-
triazines 33 lost activity in relation to their pyrimidine
counterparts. Among these compounds only 29a ,b
showed in vivo activity, but again neither of them
improved the potency of 10f.
Since 6,7-disubstitution seemed to provide the best
activity, several 6,7-cyclo derivatives (Table 3) were
prepared. The naked cyclohexyl (10a h ) and cyclopentyl
(10a j) derivatives had a profile similar to that of 10f,
while substitution by heteroatoms was detrimental.
Despite this low in vitro potency, the cyclohexanone
10a g and cyclopentanone 10a i were the only ones
displaying activity in the CPE test (22.4 and 26.3%
inhibition at 30 mg/kg, respectively).
Finally, other bicyclic frameworks, which eventually
could provide the desired in vivo properties, were
evaluated (Table 6). As in the pyrazolopyrimidine series,
the unsubstituted imidazopyridine compound 38a was
inactive, but in this case the introduction of two methyl
groups at the 6- and 7-positions (38b) impaired activity
even further. As far as the imidazopyrazines were
concerned, the poor activity of the parent 43a could not
be improved upon introduction of alkyl groups, and
compounds 43b,c were much less potent than their
pyrazolopyrimidine counterparts 10c,e. Moreover, around
10 different substituted imidazopyrimidines were pre-
pared (results not shown), and as in the case of 45, none
of them showed significant COX-2 inhibitory activity.
These results showed a restricted SAR around the
pyrimidine substituents, which precluded their further
optimization. Maintaining the 6,7-dimethyl groups, we
decided to introduce some changes in the nature of the
aromatic ring and explore changing the SO2Me for an
SO2NH2 group, a switch that in some instances has been
shown to improve in vivo activity through an improve-
ment of bioavailability.14 In this case, the sulfonamide
26a was equipotent to its sulfone counterpart 10f in the
whole cell assay (IC50: COX-1 > 10 µM, COX-2 ) 0.019
µM), although in the HWB assay 26a showed less
potency than expected. This result contrasted with that
generally observed in the diaryl C5-heterocycle class,
where this group usually provides more potency albeit
less selectivity.14 Although the 4-methyl and difluoro
derivatives 23b-d and 26b,c were highly potent in the
HWB assay, none of them showed improved in vivo
results and only compound 26b showed some activity
in the CPE test (22.8% inhibition at 10 mg/kg).
Among these modifications only the imidazopyri-
dazine framework 41 showed any activity. The presence
of the two methyl groups was also important in this
case, since 41c was much more active than the parent
41a . However, none of these derivatives showed any in
vivo activity. For purposes of comparison, the imida-
zopyridine 46 described in the patent literature17 was
prepared but exhibited less potency both in vitro and
in vivo than its counterpart 10f.
Altogether these results indicate that a nitrogen atom
at position 4 (10f vs 46) increases activity, whereas it
is merely tolerated at position 5 (29). On the contrary,
To improve in vivo results, the introduction of a
second nitrogen atom in the pyrimidine ring was envis-
aged and triazines 29 and 33 were prepared (Table 5).

New Series of COX-2-Selective Inhibitors
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3 357
tetramethoxypropane (12; 0.164 mL, 1 mmol) in EtOH (0.4
mL) was added. After 1 h at reflux the solution thus obtained
was poured over ice and basified with NH4OH. The mixture
was extracted with AcOEt and the organic phase dried and
concentrated to a crude product. This was chromatographed
on silica gel (hexane-AcOEt mixtures) to afford 10a as a white
solid (0.34 g, 76%): mp 184-185 °C; 1H NMR (CDCl3 δ TMS)
3.06 (s, 3 H), 6.93 (dd, J ) 4 Hz, J ) 7 Hz, 1 H), 7.13 (m, 2 H),
7.48 (m, 2 H), 7.86 (d, J ) 7 Hz, 2 H), 7.95 (d, J ) 7 Hz, 2 H),
8.49 (dd, J ) 1.8 Hz, J ) 4 Hz, 1 H), 8.69 (dd, J ) 1.8 Hz, J
) 7 Hz, 1 H). Anal. (C19H14FN3O2S‚0.25H2O) C, H, N, S.
its presence in position 6 (compounds 33, 38, and 43)
or 7 (45) is clearly detrimental.
Con clu sion
In summary, the tight SAR around the pyrazolo[1,5-
a]pyrimidine nucleus indicated that 6,7-disubstitution
provided the best activity and led to identification of the
6,7-dimethylated derivative 10f as one of the most
potent and selective COX-2 inhibitor in this series.
While 10f exhibited a remarkable in vitro potency and
selectivity profile in relation to reference compounds,
it was less active in vivo due to low oral bioavailability.
In vivo activity could not be improved by introducing
more polar groups in the side chains (Tables 1, 2),
changing the methylsulfonyl to sulfonamide groups
(Table 4), or introducing more nitrogen atoms to the
nucleus (Table 6). The pyrazolo[1,5-a]pyrimidine and
imidazo[1,2-b]pyridazine were the best bicyclic frame-
works among those studied, indicating that the presence
of a nitrogen atom at position 4 of the central core was
positive, at position 5 was tolerated, and at position 6
or 7 was clearly detrimental. Further efforts to discover
novel orally active COX-2-selective inhibitors related to
these structures are underway and will be reported in
due course.
3-(4-F lu or op h en yl)-5,7-d im eth yl-2-(4-m eth ylsu lfon yl-
p h en yl)p yr a zolo[1,5-a ]p yr im id in e (10e). Meth od B. A
mixture of 9a (0.51 g, 1.7 mmol), acetylacetone (11, R1, R3
)
Me, R2 ) H; 0.176 g, 1.7 mmol), 1 drop of piperidine and EtOH
(8 mL) was refluxed for 18 h. The solvent was concentrated
and the residue was chromatographed on silica gel (hexane-
AcOEt mixtures) to afford 10e as a white solid (0.48 g, 80%):
1
mp 220 °C; H NMR (CDCl3 δ TMS) 2.59 (s, 3 H), 2.81 (s, 3
H), 3.07 (s, 3 H), 6.67 (s, 1 H), 7.07 (m, 2 H), 7.47 (m, 2 H),
7.87 (d, J ) 7 Hz, 2 H), 7.93 (d, J ) 7 Hz, 2 H). Anal. (C21H18
FN3O2S‚0.25H2O) C, H, N, S.
-
Using the same procedure but starting from 4-ethoxy-3-
buten-2-one (13, R3 ) Me), a 1:4 mixture of 10b,c was obtained.
1
10b: 11% yield; mp 209-216 °C; H NMR (CDCl3 δ TMS)
2.63 (s, 3 H, 5-Me), 3.09 (s, 3 H, SO2Me), 6.78 (d, J ) 7.2 Hz,
1 H, H6), 7.09 (m, 2 H, C6H4F), 7.48 (m, 2 H, C6H4F), 7.82 (d,
J ) 7 Hz, 2 H, C6H4SO2Me), 7.94 (d, J ) 7 Hz, 2 H, C6H4SO2-
Me), 8.56 (d, J ) 7.2 Hz, 1 H, H7); 13C NMR (CDCl3 δ TMS)
26.51 (CH3, 5-Me), 45.90 (CH3, SO2Me), 109.44 (C), 110.64
(CH), 117.11 (CH, d, J F-C ) 20 Hz), 128.54 (C), 129.73 (CH),
131.03 (CH), 133.19 (CH, d, J F-C ) 8 Hz), 135.52 (CH, C7),
140.14 (C), 141.59 (C), 147.88 (C), 152.97 (C), 161.43 (C),
163.45 (C, d, J F-C ) 245 Hz). Anal. (C20H16FN3O2S‚0.25H2O)
C, H, N, S.
Exp er im en ta l Section
Melting points were determined with a Mettler FP 80
central processor melting point apparatus and are uncorrected.
Infrared spectra were recorded on a Perkin-Elmer 983 spec-
trophotometer. 1H (80 MHz) and 13C (20.1 MHz) NMR spectra
1
1
were recorded on a Bru¨cker AC 80 spectrometer and H (300
10c: 44% yield; mp 192 °C; H NMR (CDCl3 δ TMS) 2.88
MHz) NMR spectra were recorded on a Brucker Avance DPX-
300 spectrometer. They are reported in ppm on the δ scale,
from the indicated reference. Combustion analyses were
performed with a Carlo Erba 1106 analyzer. Liquid chroma-
tography was performed with a forced flow (flash chromatog-
raphy) of the indicated solvent system on SDS silica gel
Chromagel 60 ACC (230-400 mesh). Analytical thin-layer
chromatography (TLC) was performed with Macherey-Nagel
0.25-mm silica gel SIL G-25 plates.
(s, 3 H, 7-Me), 3.09 (s, 3 H, SO2Me), 6.80 (d, J ) 4 Hz, 1 H,
H6), 7.10 (m, 2 H, C6H4F), 7.48 (m, 2 H, C6H4F), 7.89 (d, J )
7 Hz, 2 H, C6H4SO2Me), 7.96 (d, J ) 7 Hz, 2 H, C6H4SO2Me),
8.46 (d, J ) 4 Hz, 1 H, H5); 13C NMR (CDCl3 δ TMS) 18.53
(CH3, 7-Me), 45.90 (CH3, SO2Me), 110.16 (CH), 110.49 (C),
117.22 (CH, d, J F-C ) 20 Hz), 128.75 (C), 128.96 (CH), 131.14
(CH), 133.05 (CH, d, J F-C ) 8 Hz), 140.24 (C), 141.61 (C),
147.58 (C), 148.59 (C), 150.85 (CH, C5), 152.35 (C), 163.65 (C,
d, J F-C ) 245 Hz). Anal. (C20H16FN3O2S‚H2O) C, H, N, S.
3-Ch lor o-2-(4-flu or oph en yl)-3-(4-m eth ylsu lfon ylph en yl)-
p r op en en itr ile (7a ). A mixture of DMF (33.5 mL, 40 mmol)
and POCl3 (14.25 mL, 155 mmol) was stirred at 0 °C under
argon for 15 min. To this solution, 5a 19(15.2 g, 52 mmol) in
CHCl3 (310 mL) was added dropwise and the mixture was
heated at 80 °C overnight. The mixture was cooled at 0 °C
and NH2OH‚HCl (10.5 g, 154 mmol) in DMF (29.25 mL) was
added and stirred for 4 h at room temperature. The suspension
was poured over saturated NaHCO3 solution and extracted
with CHCl3. The organic phase was dried and concentrated
to a crude product which was used directly in the next step
(20 g): 1H NMR (CDCl3 δ TMS) 3.10 (s, 3 H), 7.06 (m, 2 H),
7.23 (m, 2 H), 7.50 (d, J ) 7 Hz, 2 H), 7.90 (d, J ) 7 Hz, 2 H).
3-Am in o-4-(4-flu or oph en yl)-5-(4-m eth ylsu lfon ylph en yl)-
p yr a zole (9a ). To a solution of crude 7a (20 g, 52 mmol) in
EtOH (546 mL), NH2NH2‚H2O (5.4 mL, 104 mmol) was added
and the mixture was stirred at reflux overnight. The solvent
was concentrated and the residue poured over a mixture of
H2O and AcOEt. The organic phase was extracted with AcOEt
and the combined organic extracts were dried and concentrated
to a crude product. This was chromatographed on silica gel
(hexane-AcOEt mixtures) to afford 9a as a white solid (9.0 g,
52%): mp 192-193 °C; 1H NMR (CDCl3 δ TMS) 3.06 (s, 3 H),
3.92 (s, 3 H), 7.05 (m, 2 H), 7.20 (m, 2 H), 7.52 (d, J ) 7 Hz,
2 H), 7.80 (d, J ) 7 Hz, 2 H).
Compound 10b was also obtained by the following proce-
dure: A mixture of 10s (0.13 g, 0.35 mmol), EtOH (10 mL),
Pd/C (5%) (0.075 g) and NaOAc (0.070 g, 0.85 mmol) was
hydrogenated at atmospheric pressure for 1 h. The suspension
thus obtained was filtered over Celite, concentrated, and
recrystallized twice from AcOEt to give 10b as a white solid
(0.098 g, 71%).
Compound 10c was also obtained on decarboxylation of 10x
using the conditions described below for 10j.
3-(4-F lu or op h en yl)-6,7-d im eth yl-2-(4-m eth ylsu lfon yl-
p h en yl)p yr a zolo[1,5-a ]p yr im id in e (10f). Meth od C. A
mixture of 9a (0.4 g, 1.2 mmol), 3-methyl-4-methoxy-3-buten-
2-one21 (19, R2 ) R3 ) Me; 0.176 g, 1.2 mmol), HCl (0.1 mL)
and EtOH (8 mL) was refluxed for 4 h. The solvent was
concentrated and the residue was chromatographed on silica
gel (hexane-AcOEt mixtures) to afford 10f as a white solid
(0.330 g, 70%): mp 202 °C; 1H NMR (CDCl3 δ TMS) 2.42 (s, 3
H, 6-Me), 2.85 (s, 3 H, 7-Me), 3.08 (s, 3 H, SO2Me), 7.12 (m, 2
H, C6H4F), 7.48 (m, 2 H, C6H4F), 7.88 (d, J ) 7 Hz, 2 H, C6H4-
SO2Me), 7.94 (d, J ) 7 Hz, 2 H, C6H4SO2Me), 8.36 (s, 1 H,
H5). Anal. (C21H18FN3O2S‚0.25H2O) C, H, N, S. Additionally,
0.039 g of 10f was obtained (8%).
The same procedure was used for the preparation of 10d
and 23b-d starting from 9b-d respectively. 10d : mp 234 °C;
Anal. (C20H16FN3O2S‚0.25H2O) C, H, N, S. 23b: mp 244 °C;
Anal. (C22H21N3O2S‚0.25H2O) C, H, N, S. 23c: mp 230-232
°C; Anal. (C21H17F2N3O2S‚0.5H2O) C, H, N, S. 23d : mp 189-
194 °C; Anal. (C21H17F2N3O2S) C, H, N, S.
3-(4-Flu or oph en yl)-2-(4-m eth ylsu lfon ylph en yl)pyr azolo-
[1,5-a ]p yr im id in e (10a ). Meth od A. A mixture of 9a (0.41
g, 1.33 mmol), EtOH (1 mL), concentrated HCl (2.6 mL) and
ZnCl2 (0.08 g, 0.57 mmol) was heated to reflux and 1,1,3,3-

358 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3
Almansa et al.
3-(4-F lu or op h en yl)-5,6-d im eth yl-2-(4-m eth ylsu lfon yl-
p h en yl)p yr a zolo[1,5-a ]p yr im id in e (10g). Meth od D. A
mixture of 9a (0.30 g, 0.9 mmol), 4-hydroxy-3-methyl-3-buten-
2-one (21, R2 ) R3 ) Me; 0.11 g, 0.9 mmol) and toluene (2 mL)
was refluxed for 2 h. The solvent was concentrated and the
residue was chromatographed on silica gel (hexane-AcOEt
mixtures) to afford 10g as a white solid (0.22 g, 62%): mp 242-
243 °C; 1H NMR (CDCl3 δ TMS) 2.37 (s, 3 H, 6-Me), 2.60 (s, 3
H, 5-Me), 3.10 (s, 3 H, SO2Me), 7.12 (m, 2 H, C6H4F), 7.48 (m,
2 H, C6H4F), 7.84 (d, J ) 7 Hz, 2 H, C6H4SO2Me), 7.95 (d, J )
7 Hz, 2 H, C6H4SO2Me), 8.42 (s, 1 H, H7). Anal. (C21H18FN3O2S‚
CHCl3) C, H, N, S. Additionally, 0.024 g of 10f was obtained
(6%).
with CHCl3 and the organic phase was dried and concentrated
to a crude product, which was chromatographed on silica gel
(hexane-AcOEt mixtures) to afford 10j as a white solid (0.03
1
g, 27%): mp 194 °C; H NMR (CDCl3 δ TMS) 1.15 (t, J ) 7
Hz, 3 H), 2.02 (m, 2 H), 3.10 (s, 3 H), 3.26 (t, J ) 7 Hz, 2 H),
6.80 (d, J ) 4.5 Hz, 1 H), 7.14 (m, 2 H), 7.49 (m, 2 H), 7.90 (d,
J ) 7 Hz, 2 H), 7.96 (d, J ) 7 Hz, 2 H), 8.49 (d, J ) 4.5 Hz, 1
H). Anal. (C22H20FN3O2S‚0.5H2O) C, H, N, S.
3-(4-F lu or op h en yl)-5-m eth yl-2-(4-m eth ylsu lfon ylp h en -
yl)-7-m eth oxyp yr a zolo[1,5-a ]p yr im id in e (10r ). A mixture
of 10q (0.51 g, 1.36 mmol), acetone (4.2 mL), Me2SO4 (0.13
mL, 1.36 mmol), K2CO3 (0.186 g, 1.36 mmol) and KI (0.042 g,
0.2 mmol) was refluxed for 2 h. The solvent was concentrated
and the residue dissolved in a mixture CHCl3-H2O. The
organic phase was extracted with CHCl3 and the combined
organic extracts were dried and concentrated to a crude
product. This was chromatographed on silica gel (hexane-
AcOEt mixtures) to afford 10r as a white solid (0.18 g, 25%):
The same procedure was used for the preparation of 10h :
mp 202 °C; Anal. (C22H20FN3O2S‚0.5H2O) C, H, N, S.
Compound 10g was also obtained on hydrogenation of 10t
over Pd/C/NaOAc as described above for the preparation of
10b.
1
mp 213 °C; H NMR (CDCl3 δ TMS) 2.40 (s, 3 H), 3.14 (s, 3
6-Acetyl-3-(4-flu or op h en yl)-7-m eth yl-2-(4-m eth ylsu lfo-
n ylp h en yl)p yr a zolo[1,5-a ]p yr im id in e (10v). Meth od E. A
mixture of 9a (0.30 g, 0.9 mmol), 3-dimethylaminomethylene-
2,4-pentanedione (0.13 g, 0.9 mmol; 20, R3 ) Me, R2 ) COMe,
obtained by reaction of 2,4-pentanedione and 1.6 equiv of
dimethylformamide dimethylacetal in THF at room temper-
ature overnight) and AcOH (1.5 mL) was refluxed for 1 h. The
solvent was eliminated and the residue was dissolved in CHCl3
and washed with saturated NaHCO3 and NaCl solution. The
organic phase was dried and concentrated to a crude product.
This was chromatographed on silica gel (hexane-AcOEt
mixtures) to afford 10v as a white solid (0.14 g, 37%): mp 238
°C; 1H NMR (CDCl3 δ TMS) 2.73 (s, 3 H), 3.09 (s, 3 H), 3.24 (s,
3 H), 7.14 (m, 2 H), 7.48 (m, 2 H), 7.90 (d, J ) 7 Hz, 2 H), 7.97
(d, J ) 7 Hz, 2 H), 8.90 (s, 1 H). Anal. (C22H18FN3O3S) C, H,
N, S.
H), 3.88 (s, 3 H), 6.10 (s, 1 H), 7.03 (m, 2 H), 7.37 (m, 2 H),
7.61 (d, J ) 7 Hz, 2 H), 8.04 (d, J ) 7 Hz, 2 H). Anal. (C21H18
FN3O3S‚0.75H2O) C, H, N, S.
-
7-Ch lor o-3-(4-flu or op h en yl)-5-m et h yl-2-(4-m et h ylsu l-
fon ylp h en yl)p yr a zolo[1,5-a ]p yr im id in e (10s). A mixture
of 10q (0.33 g, 0.9 mmol) and POCl3 (1 mL) was refluxed for
1 h. After concentration the residue was dissolved in ice-H2O
and basified with NH4OH. The mixture was extracted with
AcOEt and the organic phase was dried and concentrated to
a crude product. This was chromatographed on silica gel
(hexane-AcOEt mixtures) to afford 10s as a white solid (0.04
1
g, 11%): mp 230-232 °C; H NMR (CDCl3 δ TMS) 2.65 (s, 3
H), 3.09 (s, 3 H), 6.96 (s, 1 H), 7.13 (m, 2 H), 7.48 (m, 2 H),
7.87 (d, J ) 7 Hz, 2 H), 7.96 (d, J ) 7 Hz, 2 H). Anal. (C20H15
ClFN3O2S‚0.25H2O) C, H, N, S.
-
Using the same procedure from the corresponding hydroxy
derivative [3-(4-fluorophenyl)-5,6-dimethyl-2-(4-methylsulfo-
nylphenyl)pyrazolo[1,5-a]pyrimidin-7-ol, obtained from 9a and
15 (R1, R2 ) Me) under conditions of method D: mp 293-298
°C; Anal. (C21H18FN3O3S‚H2O) C, H, N, S] 10t was obtained
in 88% yield.
3-(4-F lu or op h en yl)-7-m eth yl-2-(4-m eth ylsu lfon ylp h en -
yl)p yr a zolo[1,5-a ]p yr im id in e-6-ca r boxylic Acid (10x). A
mixture of 10w (0.23 g, 0.54 mmol), KOH (0.13 g, 1.4 mmol),
H2O (1 mL) and EtOH (10 mL) is heated at reflux overnight.
The solvent was concentrated and the residue poured over a
mixture of H2O and AcOEt. The aqueous phase was washed
with AcOEt and acidified with 6 N HCl. This was extracted
6-Br om o-3-(4-flu or op h en yl)-7-m eth yl-2-(4-m eth ylsu lfo-
n ylp h en yl)p yr a zolo[1,5-a ]p yr im id in e (10a b). Meth od F .
A mixture of 9a (0.4 g, 1.2 mmol), 3-bromo-4-diethylamino-3-
buten-2-one (20, R2 ) Br, R3 ) Me, obtained by bromination
with Br2 in CHCl3 of 4-diethylamino-3-buten-2-one; 0.340 g,
1.6 mmol), HBr (0.1 mL) and EtOH (4 mL) was refluxed for 1
h. The solvent was concentrated and the residue was chro-
matographed on silica gel (hexane-AcOEt mixtures) to afford
10a b as a white solid (0.170 g, 31%): mp 255-256 °C; 1H NMR
(CDCl3 δ TMS) 3.01 (s, 3 H), 3.08 (s, 3 H), 7.13 (m, 2 H), 7.47
(m, 2 H), 7.89 (d, J ) 7 Hz, 2 H), 7.94 (d, J ) 7 Hz, 2 H), 8.52
(s, 1 H). Anal. (C25H15BrFN3O2S‚0.5H2O) C, H, N, S.
3-(4-F lu or op h en yl)-6,7,8,9-tetr a h yd r o-2-(4-m eth ylsu l-
fon ylp h en yl)-6-oxop yr a zolo[1,5-a ]q u in a zolin e (10a g).
Meth od E. Following the same procedure described for the
preparation of 10v but starting from 1,3-cyclohexanedione,
compound 10a g was obtained as a sole isomer in 30% yield:
with CHCl3, which was dried and concentrated to afford 10x
1
as a white solid (78%): mp 287-292 °C; H NMR (CDCl3
+
CD3OD δ TMS) 3.09 (s, 3 H), 3.25 (s, 3 H), 4.28 (s, 1 H + H2O),
7.07 (m, 2 H), 7.42 (m, 2 H), 7.89 (d, J ) 7 Hz, 2 H), 7.91 (d,
J ) 7 Hz, 2 H), 8.97 (s, 1 H). Anal. (C21H16FN3O4S‚1.25CHCl3)
C, H, N, S.
1
mp 145-149 °C; H NMR (CDCl3 δ TMS) 2.41 (m, 2 H), 2.78
(m, 2 H), 3.10 (s, 3 H), 3.58 (m, 2 H), 7.14 (m, 2 H), 7.47 (m, 2
H), 7.89 (d, J ) 7 Hz, 2 H), 7.97 (d, J ) 7 Hz, 2 H), 9.05 (s, 1
H).
The same procedure was used for the preparation of 10n
from 10m .
3-(4-F lu or op h en yl)-6,7,8,9-tetr a h yd r o-2-(4-m eth ylsu l-
fon ylp h en yl)p yr a zolo[1,5-a ]qu in a zolin e (10a h ). Meth od
G. A mixture of 9a (0.5 g, 1.5 mmol), 2-hydroxymethylene-
cyclohexanone31 (21; 0.189 g, 1.5 mmol), p-toluensulfonic acid
(0.011 g) and toluene (19 mL) was refluxed for 1 h. The solvent
was concentrated and the residue chromatographed on silica
gel (hexane-AcOEt mixtures) to afford 10a h as a white solid
(0.27 g, 43%), together with its regioisomer 3-(4-fluorophenyl)-
6,7,8,9-tetrahydro-2-(4-methylsulfonylphenyl)pyrazolo[5,1-b]-
quinazoline (0.26 g, 42%): mp 193-194 °C; 1H NMR (CDCl3 δ
TMS) 2.0 (m, 4 H), 2.85 (m, 2 H), 3.07 (s, 3 H), 3.21 (m, 2 H),
7.10 (m, 2 H), 7.47 (m, 2 H), 7.87 (d, J ) 7 Hz, 2 H), 7.93 (d,
J ) 7 Hz, 2 H), 8.32 (s, 1 H). Anal. (C23H20FN3O2S‚0.75H2O)
C, H, N, S.
3-(4-F lu or op h en yl)-6-h yd r oxym et h yl-7-m et h yl-2-(4-
m eth ylsu lfon ylp h en yl)p yr a zolo[1,5-a ]p yr im id in e (10y).
To a solution of 10w (0.2 g, 0.44 mmol) in THF (4 mL) at -78
°C, a solution of DIBAL (1.24 mL, 1.24 mmol) 1 M in THF
was added and the mixture stirred at -78 °C for 2 h and at
room temperature overnight. H2O was added and the solution
extracted with AcOEt. The organic phase was dried and
concentrated to a crude product, which was chromatographed
on silica gel (hexane-AcOEt mixtures) to afford 10y as a
1
yellow solid (0.05 g, 28%): mp 268-270 °C; H NMR (CDCl3
+ CD3OD δ TMS) 2.95 (s, 3 H), 3.11 (s, 3 H), 3.49 (s, 1 H +
H2O), 4.79 (s, 2 H), 7.13 (m, 2 H), 7.46 (m, 2 H), 7.89 (d, J )
7 Hz, 2 H), 7.94 (d, J ) 7 Hz, 2 H), 8.51 (s, 1 H). Anal. (C21H18
FN3O3S‚0.75H2O) C, H, N, S.
-
3-(4-F lu or op h en yl)-2-(4-m eth ylsu lfon ylp h en yl)-7-p r o-
p ylp yr a zolo[1,5-a ]p yr im id in e (10j). A mixture of 10a f
(0.124 g, 0.27 mmol) and 40% H2SO4 (1.2 mL) was refluxed
for 18 h. After cooling, the mixture was poured over H2O and
basified with 2 N NaOH. The aqueous phase was extracted
3-(4-F lu o r o p h e n y l)-6-flu o r o m e t h y l-7-m e t h y l-2-(4-
m eth ylsu lfon ylp h en yl)p yr a zolo[1,5-a ]p yr im id in e (10a a ).
A mixture 10y (0.2 g, 0.48 mmol), DAST (0.067 g, 0.52 mmol)
and CH2Cl2 (8 mL) was stirred at room temperature for 2 h.
The solution thus obtained was washed with H2O and the

New Series of COX-2-Selective Inhibitors
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3 359
organic phase was dried and concentrated to a crude product.
This was chromatographed on silica gel (hexane-AcOEt
mixtures) to afford 10a a as a white solid (0.15 g, 75%): mp
218-219 °C; 1H NMR (CDCl3 δ TMS) 2.99 (d, J ) 3 Hz, 3 H),
3.09 (s, 3 H), 3.57 (d, J ) 48 Hz, 2 H), 7.13 (m, 2 H), 7.47 (m,
2 H), 7.89 (d, J ) 7 Hz, 2 H), 7.94 (d, J ) 7 Hz, 2 H), 8.50 (s,
1 H). Anal. (C21H17F2N3O2S‚0.5H2O) C, H, N, S.
and H2O. The phases were separated and the aqueous phase
was extracted with AcOEt. The combined organic phases were
dried and concentrated to a crude product. This was chro-
matographed on silica gel (hexane-AcOEt mixtures) to afford
26a as a white solid (0.083 g, 30%): mp 228-229 °C; 1H NMR
(CDCl3 + CD3OD δ TMS) 2.39 (s, 3 H), 2.81 (s, 3 H), 4.09 (s,
2 H + H2O), 7.07 (m, 2 H), 7.40 (m, 2 H), 7.74 (d, J ) 7 Hz, 2
H), 7.85 (d, J ) 7 Hz, 2 H), 8.30 (s, 1 H). Anal. (C20H17FN4O2S‚
0.75H2O) C, H, N, S.
3-(4-F lu or op h en yl)-7-h yd r oxym et h yl-6-m et h yl-2-(4-
m eth ylsu lfon ylp h en yl)p yr a zolo[1,5-a ]p yr im id in e (10a d ).
A mixture of 10f (0.2 g, 0.5 mmol), 6 N H2SO4 (0.6 mL) and
CHCl3 (0.5 mL) was added to a suspension of KMnO4 (0.079
g, 0.5 mmol), NEt3 (0.07 g, 0.5 mmol), H2O (0.1 mL) and CHCl3
(2 mL) and it was stirred at room temperature for 3 days. The
mixture was filtered over a silica gel/CaCl2 (2.5 g/0.5 g) mixture
and washed thoroughly with CHCl3. The organic phase was
washed with a saturated NaHCO3 solution, dried and concen-
trated to a crude product. This was chromatographed on silica
gel (hexane-AcOEt mixtures) to afford 10a d as a white solid,
which was recrystallized from CH2Cl2 (0.07 g, 33%): mp 226-
The same procedure was used to obtain 26b: mp 281 °C;
Anal. (C21H20N4O2S‚0.25H2O) C, H, N, S. 26c: mp 250-252
°C; Anal. (C20H16F2N4O2S‚0.5H2O) C, H, N, S.
8-(4-F lu or op h en yl)-3,4-d im eth yl-7-(4-m eth ylsu lfon yl-
p h en yl)p yr a zolo[5,1-c][1,2,4]tr ia zin e (29a ). A mixture of
ethyl 2-methylacetoacetate (0.21 mL, 1.5 mmol) and KOH (0.13
g, 1.9 mmol) in H2O (1 mL) was stirred at 0 °C for 24 h. After
adjusting the pH to 6 with concentrated HCl, a suspension of
27 [prepared from 9a (0.5 g, 1.5 mmol) by reaction at 0 °C
with NaNO2 (0.104 g, 1.5 mmol) in H2O (0.27 mL)] in HCl (0.37
mL) and H2O (0.27 mL) was added under the surface followed
by AcONa (0.38 g, 4.6 mmol). The mixture was stirred at 0 °C
for 1 h and allowed to reach room temperature. CHCl3 was
added and the phases were separated. The aqueous phase was
extracted with CHCl3 and the combined organic phases were
dried and concentrated to a crude product. This was chro-
matographed on silica gel (hexane-AcOEt mixtures) to afford
29a as a white solid (0.100 g, 17%): mp 219 °C; 1H NMR
(CDCl3 δ TMS) 2.91 (s, 3 H), 2.93 (s, 3 H), 3.11 (s, 3 H), 7.17
(m, 2 H), 7.63 (m, 2 H), 7.94 (d, J ) 7 Hz, 2 H), 7.99 (d, J ) 7
Hz, 2 H). Anal. (C20H17FN4O2S‚2H2O) C, H, N, S.
1
227 °C; H NMR (CDCl3 δ TMS) 1.55 (s, 1 H), 2.25 (s, 3 H),
3.09 (s, 3 H), 5.20 (s, 2 H), 7.13 (m, 2 H), 7.47 (m, 2 H), 7.89
(m, 4 H), 8.42 (s, 1 H). Anal. (C21H18FN3O3S‚0.5CH2Cl2) C, H,
N, S.
3-(4-F lu or op h en yl)-2-(4-m eth ylsu lfon ylp h en yl)-7-p r o-
p ylp yr a zolo[1,5-a ]p yr im id in e-6-ca r boxylic Acid (10a f). A
mixture of ethyl 3-(4-fluorophenyl)-2-(4-methylsulfonylphenyl)-
7-propylpyrazolo[1,5-a]pyrimidine-6-carboxylate [obtained from
9a and 16 (R3 ) Pr) under conditions of method C; 0.26 g, 0.54
mmol], KOH (0.13 g, 1.4 mmol), H2O (1 mL) and EtOH (10
mL) was heated at reflux overnight. The solvent was concen-
trated and the residue poured over a mixture of H2O and
AcOEt. The aqueous phase was washed with AcOEt and
acidified with 6 N HCl. This was extracted with CHCl3, which
was dried and concentrated to afford 10a f as a white solid
(70%): mp: 257 °C; 1H NMR (CDCl3 δ TMS) 1.20 (t, J ) 7
Hz, 3 H), 1.98 (m, 2 H), 3.12 (s, 3 H), 3.85 (t, J ) 7 Hz, 2 H),
7.17 (m, 2 H), 7.48 (m, 2 H), 7.91 (d, J ) 7 Hz, 2 H), 8.00 (d,
The same procedure was used to obtain 29b: mp 164 °C;
Anal. (C21H19FN4O2S) C, H, N, S. 29c: mp 213-214 °C; Anal.
(C21H19FN4O2S) C, H, N, S. 29d : mp 221-224 °C; Anal.
(C22H21FN4O2S) C, H, N, S. 29e: mp 170-176 °C; Anal.
(C22H21FN4O2S‚0.5H2O) C, H, N, S. 29f: mp 186-190 °C; Anal.
(C22H21FN4O2S‚0.25H2O) C, H, N, S.
8-(4-F lu or op h en yl)-2,4-d im eth yl-7-(4-m eth ylsu lfon yl-
p h en yl)p yr a zolo[1,5-a ][1,3,5]tr ia zin e (33a ). A mixture of
ethyl acetimidate hydrochloride (30, R1 ) Me; 0.25 g, 2 mmol),
K2CO3 (0.28 g, 2 mmol) and CH3CN (5 mL) was stirred for 5
min and the suspension thus obtained was filtered. To the
resulting solution a mixture of 9a (0.3 g, 0.9 mmol) in AcOH
(0.05 mL) was added and stirring continued overnight. The
suspension thus obtained was filtered and the solvent was
concentrated to give 31a (R1 ) Me; 0.3 g). This compound was
dissolved in ethyl orthoacetate (32, R2 ) Me; 1 mL) and heated
at reflux overnight. The solvent was concentrated to a crude
product, which was chromatographed on silica gel (hexane-
AcOEt mixtures) to afford 33a as a white solid (0.075 g, 21%):
J ) 7 Hz, 2 H), 9.09 (s, 1 H), 10.63 (s, 1 H). Anal. (C23H20
FN3O4S‚0.5H2O) C, H, N, S.
-
4-[3-(4-F lu or op h e n yl)-6,7-d im e t h ylp yr a zolo[1,5-a ]-
p yr im id in -2-yl]ben zen esu lfon a m id e (26a ). To a solution
of 22a (0.37 g, 1.0 mmol, obtained from 8a as described for
10f) in CH2Cl2 (2.5 mL) was added m-chloroperbenzoic acid
(0.32 g, 1.0 mmol) at 0 °C and the mixture was stirred for 2 h
at room temperature. The suspension was washed with 1 N
NaOH and the aqueous phase was extracted with CH2Cl2. The
combined organic phases were dried and concentrated to a
crude product. This was chromatographed on silica gel (hex-
ane-AcOEt mixtures) to afford 3-(4-fluorophenyl)-6,7-di-
methyl-2-(4-methylsulfinylphenyl)pyrazolo[1,5-a]pyrimidine as
a white solid (0.32 g, 82%): 1H NMR (CDCl3 δ TMS) 2.41 (s,
3 H), 2.75 (s, 3 H), 2.84 (s, 3 H), 7.10 (m, 2 H), 7.46 (m, 2 H),
7.67 (d, J ) 7 Hz, 2 H), 7.83 (d, J ) 7 Hz, 2 H), 8.35 (s, 1 H).
1
mp 190 °C; H NMR (CDCl3 δ TMS) 2.71 (s, 3 H), 3.02 (s, 3
H), 3.11 (s, 3 H), 7.13 (m, 2 H), 7.45 (m, 2 H), 7.86 (d, J ) 7
Hz, 2 H), 7.99 (d, J ) 7 Hz, 2 H). Anal. (C20H17FN4O2S) C, H,
N, S.
The previous compound was treated with Ac2O (2.6 mL) and
AcONa (0.26 g, 3.2 mmol) at reflux for 8 h. The solvent was
eliminated and the crude 2-[4-(acetoxymethylthio)phenyl]-3-
(4-fluorophenyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidine (24a )
thus obtained was used directly in the next step.
A solution of the previous compound in CH2Cl2 (2.6 mL) and
MeOH (1.7 mL) at 0 °C was treated with hexahydrated
magnesium monoperoxyphthalate (0.58 g, 0.97 mmol) and the
mixture stirred at room temperature overnight. Saturated
NaHCO3 solution was added and the mixture extracted with
CH2Cl2. The solvent was eliminated and the residue dissolved
in THF (2.6 mL) and MeOH (1.3 mL) and cooled to 0 °C. 1 N
NaOH (0.84 mL) was added and the mixture stirred at room
temperature for 1 h. The solvent was eliminated and the
residue vacuum-dried to afford crude sodium 4-[3-(4-fluorophe-
nyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidin-2-yl]benzenesulfi-
nate (25a ; 0.28 g).
The same procedure was used to obtain 33b: mp 177-183
°C; Anal. (C23H23FN4O2S‚1.25H2O) C, H, N, S. 33c: mp 170-
173 °C; Anal. (C22H21FN4O2S‚0.25H2O) C, H, N, S.
4-Ch lor o-2,6-d im eth yl-3-n itr op yr id in e (35b). A mixture
of 2,6-dimethyl-4-hydroxy-3-nitropyridine (34b;32 4.6 g, 27.3
mmol), POCl3 (2.5 mL), DMF (2 mL) and toluene (20 mL) was
heated at 110 °C for 7 h. After cooling to 0 °C, the mixture
was basified with Na2CO3 and the solid thus obtained filtered
and dried to afford 35b as an oil (3.54 g, 69%): 1H NMR (CDCl3
δ TMS) 2.56 (s, 3 H), 2.57 (s, 3 H), 7.18 (s, 1 H).
2,6-Dim e t h yl-4-(4-flu or op h e n yla m in o)-3-n it r op yr i-
d in e (36b). To a solution of 35b (3.54 g, 19 mmol) in CHCl3
(3 mL) was added, at 0 °C under argon, a solution of
4-fluoroaniline (2.2 mL, 23 mmol) and NEt3 (3.7 mL) in CHCl3
(6.5 mL) and the mixture was refluxed for 48 h. The solvent
was eliminated and the residue was dissolved with AcOEt and
H2O. The phases were separated and the aqueous phase was
extracted with AcOEt. The combined organic phases were
dried and concentrated to a crude product. This was chro-
matographed on silica gel (hexane-AcOEt mixtures) to afford
36b as a yellow solid (4.0 g, 81%): mp 128 °C; 1H NMR (CDCl3
A mixture of the previous crude, H2O (3.6 mL), AcONa
(0.063 g, 0.76 mmol) and hydroxylamine O-sulfonic acid (0.086
g, 0.76 mmol) was stirred at room temperature overnight. The
suspension thus obtained was filtered and washed with AcOEt

360 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3
Almansa et al.
δ TMS) 2.35 (s, 3 H), 2.75 (s, 3 H), 6.52 (s, 1 H), 7.20 (m, 4 H),
8.60 (s, 1 H).
ro-3,5-dimethylpyrazine in a manner similar to that described
in ref 34), 2-amino-3-butylpyrazine (42c)35 and 2-amino-4-
methylpyrimidine (44). 43b: mp 239-242 °C. Anal. (C21H18
-
3-Am in o-2,6-d im et h yl-4-(4-flu or op h en yla m in o)p yr i-
d in e (37b). A mixture of 36b (2.0 g, 7.6 mmol), SnCl2 (7.2 g,
38 mmol) and EtOH (54 mL) was refluxed for 45 min. The
solvent was eliminated and the residue was dissolved with H2O
and basified with 25% NaOH. The aqueous phase was ex-
tracted with CHCl3 and the organic phase was dried and
concentrated to afford 37b as an oil (1.0 g, 56%): 1H NMR
(CDCl3 δ TMS) 2.34 (s, 3 H), 2.44 (s, 3 H), 3.19 (s, 2 H), 5.91
(s, 1 H), 6.63 (s, 1 H), 7.04 (m, 4 H).
4,6-Dim et h yl-1-(4-flu or op h en yl)-2-(4-m et h ylsu lfon yl-
p h en yl)im id a zo[4,5-c]p yr id in e (38b). A mixture of 37b (1.0
g, 4.0 mmol), 4-methylsulfonylbenzoyl chloride (0.98 g, 4.5
mmol) and pyridine (16 mL) was refluxed overnight. The
solvent was eliminated and the residue was dissolved with H2O
and extracted with CHCl3. The organic phase was washed with
0.1 N NaOH, dried and concentrated to afford a crude product.
This was chromatographed on silica gel (hexane-AcOEt
mixtures) to afford 38b as a white solid (0.46 g, 29%): mp 259
°C; 1H NMR (CDCl3 δ TMS) 2.62 (s, 3 H), 2.94 (s, 3 H), 3.06 (s,
3 H), 6.85 (s, 1 H), 7.28 (m, 4 H), 8.76 (d, J ) 7 Hz, 2 H), 7.91
(d, J ) 7 Hz, 2 H). Anal. (C21H18FN3O2S‚0.25H2O) C, H, N, S.
The same procedure was used to obtain 38a : mp 218-221
°C; Anal. (C19H14FN3O2S‚0.25H2O) C, H, N, S.
FN3O2S‚0.25H2O) C, H, N, S. 43c: Anal. (C23H22FN3O2S‚
0.25H2O) C, H, N, S. 45: mp 272-273 °C; Anal. (C20H16FN3O2S)
C, H, N, S.
In h ibition of COX-1 a n d COX-2 Activities in HWB.27
The tests were performed with heparinized human whole blood
from healthy donors who had not taken NSAIDs for 1 week
nor alcohol or xanthines within 24 h of blood collection. COX-1
activity was measured in spontaneously clotting human blood
for 5 h at 37 °C and stimulated with A23187 (5 µM) for 30
min longer. The reaction was stopped at 0 °C with EGTA and
MeOH. After centrifugation the supernatant was removed and
stored at -70 °C. TXB2 levels were determinated by enzyme
immunoassay kit (RPN 220, Amersham). COX-2 activity was
stimulated by adding lipopolysaccharide 026:B6 (25 µL/mL)
at 37 °C for 24 h. Aspirin (20 µg/mL) was used to inhibit COX-1
activity. After centrifugation, the samples were stored at -70
°C. PGE2 levels were determinated by enzyme immunoassay
kit (RPN 222, Amersham).
In h ibition of COX-1 a n d COX-2 Activities in Hu m a n
Cell Lin es. The production of PGE2 after AA stimulation by
lines expressing h-COX-1 (U-937 from human histiocytic
lymphoma) and h-COX-2 (143.98.2 from human osteosarcoma)
was used as a cell-based assay to evaluate COX-1 and COX-2
inhibition, respectively. Briefly, osteosarcoma cells were cul-
tured in 1 mL of media in 24-well multidishes until confluent
(2 × 105 cells/well during 24 h). U-937 cells were grown in
flasks and resuspended to a final density of 3 × 106 cells/mL/
2-Br om o-2-(4-flu or oph en yl)-1-(4-m eth ylsu lfon ylph en yl)-
eth a n on e (39). To a solution of 5a (5 g, 17.1 mmol) in CH2-
Cl2 (44 mL) was added a solution of Br2 (0.98 mL, 19.2 mmol)
in AcOH (107 mL) and the mixture was stirred at room
temperature overnight. The solvent was eliminated and the
residue was dried to afford 39 as a white solid (6.35 g, 100%):
well. The cells were resuspended in HBSS without Ca2+/Mg2+
.
1
10 µL of a DMSO solution of test compound or DMSO vehicle
was added. The samples were then incubated for 15 min at
37 °C (5% CO2 and 95% humidity) before the addition of AA
(final concentration: 10 µM) for 10 min. The reactions were
stopped by the addition of indomethacine (8 mM, 30 µL). PGE2
levels in supernatant were determined by specific EIA (Am-
ersham). All assays were performed in triplicate.
mp 113-116 °C; H NMR (CDCl3 δ TMS) 3.13 (s, 3 H), 6.34
(s, 1 H), 7.22 (m, 2 H), 7.58 (m, 2 H), 8.09 (d, J ) 7 Hz, 2 H),
8.21 (d, J ) 7 Hz, 2 H).
3-Am in o-6-ch lor o-4,5-d im eth ylp yr id a zin e (40b). A mix-
ture of 3,6-dichloro-4,5-dimethylpyridazine33 (0.5 g, 2.8 mmol),
30% aqueous NH3 (20 mL) and THF (5 mL) was stirred in a
pressure vessel at 150 °C for 36 h. The solvent was eliminated
and the residue was chromatographed on silica gel (hexane-
AcOEt mixtures) to afford 40b as an oil (0.22 g, 50%): 1H NMR
(CDCl3 δ TMS) 2.14 (s, 3 H), 2.33 (s, 3 H), 4.81 (s, 2 H).
6-Ch lor o-7,8-d im eth yl-3-(4-flu or op h en yl)-2-(4-m eth yl-
su lfon ylp h en yl)im id a zo[1,2-b]p yr id a zin e (41b). A mixture
of 40b (0.22 g, 1.4 mmol), 39 (0.52 g, 1.4 mmol) and DMF (3
mL) was kept at 60 °C overnight. The solvent was eliminated
and the residue was dissolved with H2O and extracted with
CHCl3. The organic phase was washed with saturated NaHCO3
solution, dried and concentrated to afford a crude product. This
was chromatographed on silica gel (hexane-AcOEt mixtures)
to afford 41b as a white solid (0.46 g, 29%): mp 192-194 °C;
H NMR (CDCl3 δ TMS) 2.44 (s, 3 H), 2.74 (s, 3 H), 3.05 (s, 3
In h ibition of Ca r r a geen a n -In d u ced Ra t P a w Ed em a .
Male Sprague-Dawley rats (175-200 g) were used. Edemas
were produced by injecting 0.1 mL of a solution of 1%
λ-carrageenan in the hindpaw. Paw volumes were measured
by water displacement with a plethysmometer (UGO BASILE)
before and 1, 2, 3, 4 and 5 h after treatment. The compounds
were administered orally as a suspension in carboxymethyl
cellulose and Tween-80 at 1% (10 mL/kg), 0.5 h before
carrageenan injection and after being hydrated with H2O (5
mL). The percentages of inhibition were calculated by compar-
ing the areas under the curve of treated and control animals.
Air -P ou ch Mod el of In fla m m a tion : Deter m in a tion of
P GE2 in Exu d a te a n d Stom a ch . Male Lewis rats (175-200
g) were used. Air cavities were produced by a subcutaneous
injection of sterile air (20 mL) into the intracapsular area.
Every 2 days air (10 mL) was injected into the cavity to keep
the space open. 7 days after the first injection, λ-carregeenan
(Sigma) in saline (2 mL of a 1% solution) was injected into
the air pouch to produce an inflammatory reaction. The
compounds were administered by oral route as a suspension
in carboxymethyl cellulose and Tween-80 at 1% (10 mL/kg),
0.5 h before carrageenan injection. The animals were killed 6
h later and the exudate volume was measured. Cells were
pelleted by centrifugation at 1200g for 5 min at 4 °C, and PGE2
was determined in the supernatant by specific ELISA. Im-
mediately after the exudates were collected, the stomachs were
excised and frozen at -70 °C. On the day of analysis, stomachs
were homogenized in 70% ethanol. The homogenates were
centrifuged (1000g, 10 min, 4 °C) and the supernatants were
dried under N2 stream and resuspended in ELISA buffer for
PGE2 determination.
H), 7.22 (m, 2 H), 7.54 (m, 2 H), 7.94 (s, 4 H). Anal. (C21H17
-
ClFN3O2S‚0.25H2O) C, H, N, S.
The same procedure was used to obtain 41a : mp 216 °C;
Anal. (C19H14FN3O2S‚0.25H2O) C, H, N, S.
7,8-Dim et h yl-3-(4-flu or op h en yl)-2-(4-m et h ylsu lfon yl-
p h en yl)im id a zo[1,2-b]p yr id a zin e (41c). A mixture of 41b
(0.15 g, 0.3 mmol), 10% Pd/C (0.03 g), NEt3 (0.14 mL, 1 mmol)
and THF (12 mL) was hydrogenated at 200 psi overnight. The
mixture was filtered over Celite and the solvent was elimi-
nated to afford a crude product. This was chromatographed
on silica gel (hexane-AcOEt mixtures) to afford 41c as a white
solid (0.095 g, 69%): mp 216-217 °C; H NMR (CDCl3 d TMS)
2.38 (s, 3 H), 2.68 (s, 3 H), 3.05 (s, 3 H), 7.22 (m, 2 H), 7.54 (m,
2 H), 7.90 (m, 4 H), 8.10 (s, 1 H). Anal. (C21H18FN3O2S‚H2O)
C, H, N, S.
3-(4-Flu or oph en yl)-2-(4-m eth ylsu lfon ylph en yl)im idazo-
[1,2-a ]p yr a zin e (43a ). The same procedure described for 41b
starting from 2-aminopyrazine (42a ) was used to obtain 43a :
1
mp 229-231 °C; H NMR (CDCl3 d TMS) 3.04 (s, 3 H), 7.33
Ack n ow led gm en t . We thank M. Carmen Torres,
Concepcio´n Gonza´lez, Consol Ferreri, Guadalupe Mar-
t´ınez, Nuria Recasens, Teresa Gamero, J ose´ Antonio
Garc´ıa, Ana Ester Sanahuja, and Assumpta Oliveras
(m, 2 H), 7.47 (m, 2 H), 8.87 (m, 6 H), 9.19 (s, 1 H). Anal.
(C19H14FN3O2S‚0.5H2O) C, H, N, S.
Analogously, 43b,c and 45 were obtained using, respec-
tively, 2-amino-3,5-dimethylpyrazine (42b, obtained from 2-chlo-

New Series of COX-2-Selective Inhibitors
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 3 361
Kennedy, B.; Leblanc, Y.; Le´ger, S.; Mancini, J .; O′Neill, G. P.;
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for their excellent technical assistance and J ordi Belloc
for the GC, MS, and HPLC analyses.
Su p p or tin g In for m a tion Ava ila ble: Combustion analy-
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