Synthesis and evaluation of indenopyrazoles as cyclin-dependent kinase inhibitors. 3. Structure activity relationships at C3

Article abstract of DOI:10.1021/jm0201722

The identification of indeno[1,2-c]pyrazol-4-ones as inhibitors of cyclin-dependent kinases (CDKs) has led to the discovery of a series of novel and potent compounds. Herein, we report the effects of substitutions at C3 of the indeno[1,2-c]pyrazol-4-one core with alkyls, heterocycles, and substituted phenyls. Substitutions at the para position of the phenyl ring at C3 were generally well-tolerated; however, larger groups were generally inactive. For alkyls directly attached to C3, longer chain substituents were not tolerated; however, shorter alkyl groups and cyclic alkyls were acceptable. In general, the heterocycles at C3 gave the most potent analogues. One such heterocycle, 24j, was examined in detail and was determined to have a biological profile consistent with CDK inhibition. An X-ray crystal structure of one of the alkyl compounds, 13q, complexed with CDK2 was determined and showed the inhibitor residing in the adenosine 5′-triphosphate pocket of the enzyme.

Full text of DOI:10.1021/jm0201722

J . Med. Chem. 2002, 45, 5233-5248
5233
Syn th esis a n d Eva lu a tion of In d en op yr a zoles a s Cyclin -Dep en d en t Kin a se
In h ibitor s. 3. Str u ctu r e Activity Rela tion sh ip s a t C31,2
Eddy W. Yue,* C. Anne Higley, Susan V. DiMeo, David J . Carini, David A. Nugiel, Carrie Benware,
Pamela A. Benfield, Catherine R. Burton, Sarah Cox, Robert H. Grafstrom, Diane M. Sharp, Lisa M. Sisk,
J ohn F. Boylan, J odi K. Muckelbauer, Angela M. Smallwood, Haiying Chen, Chong-Hwan Chang,
Steven P. Seitz, and George L. Trainor
Bristol-Myers Squibb Company, Experimental Station, P.O. Box 80500, Wilmington, Delaware 19880-0500
Received April 22, 2002
The identification of indeno[1,2-c]pyrazol-4-ones as inhibitors of cyclin-dependent kinases
(CDKs) has led to the discovery of a series of novel and potent compounds. Herein, we report
the effects of substitutions at C3 of the indeno[1,2-c]pyrazol-4-one core with alkyls, heterocycles,
and substituted phenyls. Substitutions at the para position of the phenyl ring at C3 were
generally well-tolerated; however, larger groups were generally inactive. For alkyls directly
attached to C3, longer chain substituents were not tolerated; however, shorter alkyl groups
and cyclic alkyls were acceptable. In general, the heterocycles at C3 gave the most potent
analogues. One such heterocycle, 24j, was examined in detail and was determined to have a
biological profile consistent with CDK inhibition. An X-ray crystal structure of one of the alkyl
compounds, 13q, complexed with CDK2 was determined and showed the inhibitor residing in
the adenosine 5′-triphosphate pocket of the enzyme.
In tr od u ction
nones,15 pyridopyrimidinones,16 and aminothiazoles17
(Figure 1). GW8510 is a representative of the indolino-
nes with good potency in CDK2 and with selectivity
against CDK1 and CDK4. The pyridopyrimidinone
PD172803 and the aminothiazole AG12286 both showed
good potency against CDK4/D and CDK2/A.
We have previously disclosed our investigations into
the novel indenopyrazole class of CDK inhibitors, which
centered around the optimization of the C5 aniline
substituent18,19 (Figure 2). Earlier analogues employed
a 4-methoxyphenyl group at C3, and we were particu-
larly interested in the effects of alternately substituted
phenyl groups as well as heterocycles and alkyls in this
position. In this paper, we present our findings on the
impact of a variety of substitutions at C3.
Cyclin-dependent kinase (CDK) inhibitors have gar-
nered attention recently for their potential as anticancer
therapeutics.3 CDKs are cellular kinases, which play a
crucial role in the regulation of progression through the
different phases of the cell cycle. As their name implies,
each of these kinases requires the association with a
cyclin in order for it to become active. Several CDK/
cyclin pairs have been identified, which have key roles
in the cell cycle machinery including CDK4/D1, CDK2/
E, CDK2/A, and CDK1/B.4-6 Because cell cycle dysregu-
lation is a characteristic of transformed cells, it has been
theorized that inhibition of certain CDK/cyclin com-
plexes would be of value in the treatment of proliferative
diseases.
The search for small molecule inhibitors of CDKs has
already led to the discovery of several classes of com-
pounds including butyrolactone I,7 the flavonoids rep-
resented by flavopiridol,8,9 and purines such as olomou-
cine,10,11 roscovitine,12,13 and purvalanol14 (Figure 1).
Butyrolactone I is an inhibitor of CDK1 and CDK2 with
selectivity over other non-CDK kinases. Flavopiridol has
activity for several CDKs including CDK1, CDK2, and
CDK4 and has shown efficacy in animal tumor models.
Flavopiridol is currently undergoing clinical trials as a
cell cycle inhibitor for the treatment of cancer. Olomou-
cine has modest activity for CDK1 and CDK2 but is
inactive against CDK4. Roscovitine, purvalanol A, and
purvalanol B are more potent purine analogues with
selectivity for CDK2 and CDK5 vs CDK4 and several
other kinases. The last four examples suggest that some
selectivity within the CDK family can be achieved.
More recent reports of CDK inhibitors have included
several novel classes of compounds including the indoli-
Ch em istr y
The synthesis of indenopyrazoles substituted at C3
involved two different routes. For C3 para-substituted
phenyls, the chemistry previously described19 was uti-
lized. Condensation of diester 2 with an acetophenone
appropriately substituted at the para position gave the
triketones 4 in good yield (Scheme 1). Cyclization of the
triketones using hydrazine provided the desired inde-
nopyrazoles with a C5 acetamide (5). Glycinamides at
C5 were prepared by removal of the acetamide (5 f 6),
formation of the R-chloroacetamide (6 f 7), and con-
densation with the appropriate amine (7 f 8).
The conditions developed for the synthesis of C3 aryl
analogues were incompatible with the C3 alkyl sub-
strates. Attempts to synthesize the triketone using 2
and the 1,1-dimethylhydrazone of 2-butanone also failed.
The synthesis of triketones with C3 alkyl groups,
however, was accomplished using a procedure developed
by Rotberg and Oshkaya.20 Condensation of 3-nitro-
phthalic anhydride with symmetrical 1,3-diones in
acetic anhydride and triethylamine gave the 4-nitro-1,3-
* To whom correspondence should be addressed. Tel: (302)467-6054.
Fax: (302)467-6951. E-mail address: eddy.yue@bms.com.
10.1021/jm0201722 CCC: $22.00 © 2002 American Chemical Society
Published on Web 10/23/2002

5234 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24
Yue et al.
20 (R ) alkyl group to be transferred and R′ ) methyl
or trifluoromethyl), it was discovered that selective
elimination of the acetyl or trifluoroacetyl group could
be achieved (Scheme 3). This simplified the synthesis
of analogues to the preparation of unsymmetrical 1,3-
diketones 20, which were easily prepared from the
corresponding ketones (19).
In an attempt to generate a late-stage intermediate
to facilitate the derivatization of the C5 aniline, the
formation of the indenopyrazole core from the nitrot-
riketone (10) intermediate was investigated. Cyclization
of nitrotriketone 10a (R ) Me) with hydrazine in
refluxing ethanol did not produce any desired pyrazole
but afforded exclusively the hydrazone 14 (Scheme 2).
This phenomenon seemed to be confined to the alkyl
analogues since cyclization of the substituted phenyls
and heterocycles all proceeded in ethanol. To induce
cyclization, elevated temperatures were tried. When the
triketone 10a as well as the hydrazone 14 were sub-
jected to the same conditions using higher boiling
n-butanol, the cyclized pyrazole was formed but as the
undesired 8-nitro regioisomer 15. This was in contrast
to the previously reported cyclizations of acetamide-
bearing substrates, which gave the desired 5-position
substitution.19 The regiochemistry of this 8-nitro com-
pound was confirmed by converting the nitro group to
the 8-acetamide (17) via the aniline (16) and comparing
this compound with that of the desired 5-acetamide
compound 13a .
Although disappointing, the regiochemistry of the
cyclization was not surprising since the nitro group
would be expected to enhance the electrophilic nature
of the carbonyl ortho to the nitro group (14 f 15). In
the case of the acetamide, the carbonyl ortho to the
acetamide functionality would have reduced reactivity
due to its conjugation with the aniline as a vinylogous
amide (Figure 3) and therefore gives the desired cyclized
product (21 f 13a ).
Glycinamides at the 5-position in the C3 alkyl series
were prepared using the chemistry described for the
para-substituted phenyl analogues.19 Formation of the
chloroacetamide 18 proceeded in good yield (Scheme 2).
Displacement of the chlorine with 4-aminomethylpip-
eridine or isonipecotamide gave the desired triketones,
which were cyclized to the pyrazole analogues.
F igu r e 1. Structures of CDK inhibitors.
Ureas at the 5-position for both the C3 alkyl and the
heterocycle series were prepared according to the previ-
ously disclosed indenopyrazole chemistry.19 Treatment
of the aniline 11 with phenylchloroformate gave the
carbamate 22 in good yield (Scheme 4). Addition of
ammonium hydroxide to the carbamate generated the
desired urea triketones 23, which were cyclized under
standard conditions to the pyrazole 24.
F igu r e 2. Structures of indenopyrazole 1.
indanediones 10 in good yields (Scheme 2). Reduction
of the nitro group with zinc and calcium chloride
provided the desired aniline 11 with no sign of overre-
duced materials. Acetylation of the aniline produced the
triketone precursor 12, which was cyclized to give the
desired pyrazole 13 with alkyl substituents at C3.
The use of symmetrical 1,3-diones for the phthalic
anhydride condensation was efficient for smaller alkyl
groups, but as the substitutions increased in size and
complexity, the yields for the reaction decreased. To
circumvent this problem, the use of unsymmetrical 1,3-
diones was explored. For unsymmetrical ketones of type
Resu lts a n d Discu ssion s
C3 pa r a -P h en yl Su bstitu en ts. The optimization at
C3 began with an examination of the para position of
the C3 phenyl group. A number of substituents were
tolerated at the para position (Table 1). With the alkyl-
substituted analogues, the activity in both CDK4/D1
and CDK2/E steadily decreased from methyl and ethyl
to longer chain alkyls such as propyl and butyl. The
same pattern was also observed with the oxygenated
variants; methoxy was tolerated, but ethoxy, phenoxy,

Indenopyrazoles as Inhibitors
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5235
Sch em e 1a
a
Reagents: (a) NaH, DMF, 90 °C. (b) H2NNH2, EtOH, reflux. (c) Concentrated HCl, MeOH, reflux. (d) ClCH2COCl, 2.5 M Na2CO3(aq),
dioxane, 50 °C. (e) 2° amine, EtOH, reflux.
Sch em e 2a
a
Reagents: (a) RC(dO)CH2C(dO)R′ (20), Ac2O, Et3N, 25 °C. (b) Zn, CaCl2, EtOH, H2O, reflux. (c) AcCl, Na2CO3, acetone, 50 °C. (d)
H2NNH2, BuOH, reflux. (e) H2NNH2, EtOH, reflux. (f) BuOH, reflux. (g) H2NNH2, BuOH, reflux. (h) ClCH2C(dO)Cl, Na2CO3, acetone, 50
°C. (i) 4-Aminomethylpiperidine or isonipecotamide, EtOH, 100 °C sealed tube.
and benzyloxy all gave decreased activity in both CDK4/
D1 and CDK2/E. In both of these series of compounds,
the smaller para substituents, methyl (5b) and methoxy
(5g), were clearly favored.
analogues as compared with the parent acetamide 5j
(Table 2). In CDK4/D1, however, the results were
dependent on the groups used. The morpholino (8a ),
hydroxy piperidine (8b), and methyl piperazine (8d )
glycinamides were slightly more active than the aceta-
mide 5j, but the aminomethylpiperidine (8c) gave a
significant improvement in CDK4/D1 activity. The
p-morpholinophenyl glycinamides were either the same
(8e) or slightly worse (8f-h ) against CDK4/D1 and
CDK2/E as compared with the p-dimethylaminophenyl
derivatives (Table 2). The aminomethylpiperidine ana-
logue 8g, however, again proved to be a good inhibitor
of CDK4/D1 with similar activity in CDK2/E. These
results were consistent with previous work, which
showed that a p-methoxyphenyl at C3 combined with
the aminomethylpiperidine glycinamide at C5 gave a
very potent CDK4/D1 inhibitor with balanced CDK2/E
activity (CDK4/D1 IC50 ) 13 nM, CDK2/E IC50 ) 14
nM).19
In the aniline series, the dimethylamino analogue 5j
had potency similar to the methoxy group and again
there was a slight loss in activity with larger groups at
that position. The piperidino (5k ) and morpholino (5l)
analogues were both less active in CDK4/D1. Interest-
ingly, although 5k was also less active in CDK2/E as
compared with 5j, 5l was essentially unchanged and
suggested that cyclic groups could be tolerated at that
position. The thio derivative 5m followed the trend of
the other small groups with activity similar to that of
the methyl compound 5b.
Although the p-dimethylaminophenyl (5j) and p-
morpholinophenyl (5l) analogues were similar in activity
to p-methoxyphenyl (5g), it was reasoned that these
compounds might provide improved solubility; therefore,
these analogues were further pursued. A series of
glycinamides were synthesized and for those containing
p-dimethylaminophenyl at C3 (8a -d ), there was a
marked increase in activity in CDK2/E for all of the
C3 Alk yl Su bstitu en ts. Intrigued by the results of
the para-substituted phenyl compounds, the phenyl
group was removed and analogues that had an alkyl
group directly attached to the C3 position were next

5236 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24
Yue et al.
Sch em e 3a
Ta ble 1. C3 para-Phenyl, C5 Acetamide SAR
IC50 (nM)
compd
R
CDK4/D1a
CDK2/Ea
5a
5b
5c
5d
5e
5f
5g
5h
5i
H
Me
Et
n-Pr
n-Bu
OH
OMe
OEt
OPh
840
490
640
240
160
280
490
>560
530
1900
>2800
2300
450
>580
>2500
310
270
>400
>2500
320
5j
NMe2
5k
5l
5m
piperidino
morpholino
SMe
680
860
340
900
370
120
a
Reagents: (a) F3CCO2Et, Na, EtOH, 25 °C. (b) Ac2O, Et3N,
25 °C.
a
Values correspond to n ) 2.
Ta ble 2. C3 para-Phenyl, C5 Glycinamide SAR
F igu r e 3. Proposed pathways for the cyclization of the 5-nitro
and 5-acetamido triketones to the indenopyrazole core.
Sch em e 4a
a
Values correspond to n ) 2.
strained c-hexyl was also active, which led to the
conclusion that for straight chain analogues, a two
carbon length was optimal; however, larger groups could
also be accommodated if they have restricted conforma-
tions. These results were largely in line with the para-
substituted phenyl series, which also showed a prefer-
ence for smaller or cyclic substituents. It was clear,
though, that the phenyl spacer did provide an increase
in activity especially against CDK4/D1.
a
Reagents: (a) PhOC(dO)Cl, Na2CO3, acetone, 50 °C. (b)
NH4OH, DMSO, 90 °C. (c) H2NNH2, EtOH, reflux.
Next to be profiled were aliphatic C3 substituents
with the more potent C5 glycinamides. The nipecota-
mide glycinamides provided a large increase in potency
for the i-propyl (13k ), c-propyl (13l), and c-hexyl (13m )
compounds in CDK2/E (Table 4). Similar improvements
were seen with the 4-aminomethylpiperidine glycina-
mides. In addition, the solubility of these compounds
at pH 7.4 (i-propyl (13o) ) 1.23 mg/mL, c-propyl (13p )
examined. A series of C3 alkyl substituents with C5-
acetamides were all inactive against CDK4/D1 (Table
3). CDK2/E was significantly more tolerant of aliphatic
substituents. A large increase in activity was observed
in going from methyl to ethyl with further improvement
when branching was added to give the isopropyl ana-
logue. Longer chain compounds such as the n-butyl and
i-butyl lost inhibitory activity. The larger but con-

Indenopyrazoles as Inhibitors
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5237
Ta ble 5. C3 Alkyl, C5 Primary Urea SAR
Ta ble 3. C3 Alkyl, C5 Acetamide SAR
IC50 (nM)
CDK4/D1a
IC50 (nM)
CDK4/D1a
CDK2/Ea
compd
R
CDK2/Ea
compd
R
13a
13b
13c
13d
13e
13f
13g
13h
13i
Me
Et
>4200
>3900
>3700
>3700
>3700
>3500
>3500
>3500
>3200
11000
1500
>3700
840
24a
24b
24c
24d
Et
i-Pr
c-Pr
c-Hx
>2000
>1900
1400
290
84
140
140
n-Pr
i-Pr
c-Pr
i-Bu
n-Bu
t-Bu
c-Hx
>810
480
a
Values correspond to n ) 2.
Ta ble 6. C3 Heterocycle, C5 Acetamide SAR
>3500
>3500
980
1100
a
Values correspond to n ) 2.
Ta ble 4. C3 Alkyl, C5 Glycinamide SAR
IC50 (nM)
compd
R
CDK4/D1a
CDK2/Ea
13r
13s
13t
3-pyridyl
4-pyridyl
2-thienyl
>1600
1300
4500
770
76
180
13u
13v
13w
2-thienyl, 3-Me
3-thienyl, 2,5-diCl
2-furanyl
>3100
>2600
>3500
260
>2600
>1700
a
Values correspond to n ) 2.
Ta ble 7. C3 Heterocycle, C5 Primary Urea SAR
IC50 (nM)
compd
R
CDK4/D1a
CDK2/Ea
a
Values correspond to n ) 2.
24e
24f
24g
24h
24i
24j
24k
24l
2-thienyl
150
350
53
300
67
11
17
13
130
9
13
61
86
26
2-thienyl, 3-Me
2-thienyl, 5-Me
2-thienyl, 5-CO2Et
3-thienyl
3-thienyl, 5-Cl
3-thienyl, 2,5-diMe
2-furanyl
> 0.87 mg/mL, and c-hexyl (13q) > 0.6 mg/mL) was
quite good as compared with those of the C3 phenyl
analogues. Comparison of 13o with 8g (0.01 mg/mL) and
8c (0.001 mg/mL) shows an increase in solubility of more
than 100-fold.
57
>1500
840
130
24m
3-pyrrolyl, 1-Me
From previous work, it was suggested that further
improvements in activity could be made by incorporat-
ing ureas at C5.19 Even though the primary ureas (24a -
d ) showed a marked improvement in activity as com-
pared with the 5-acetamides, they were in general less
potent than the substituted acetamides (Table 5). In
addition, these ureas (24a -d ) were all quite insoluble
at pH 7.4 (<0.01 mg/mL).
C3 Heter ocycle Su bstitu en ts. The next series of
compounds to be examined were the C3 heterocycles
(Table 6). With an acetamide at the 5-position, all of
the examples were less active in CDK4/D1 as compared
with the unsubstituted phenyl derivative 5a . In CDK2/
E, the 3-pyridyl analogue (13r ) was less potent than the
phenyl analogue (5a ) but the 4-pyridyl compound (13s)
a
Values correspond to n ) 2.
was more active suggesting the presence of a specific
binding interaction, which was optimal at the para
position. The 2-thienyl compound proved a little better
than phenyl for CDK2/E. The 3-methyl-2-thienyl ana-
logue was about equipotent in CDK2/E as compared
with phenyl, but the 2,5-dichloro-3-thienyl and 2-furanyl
were both inactive.
Encouraged by these results, primary ureas at C5
were examined. The 2-thienyl compound 24e was a
potent inhibitor in CDK2/E with modest activity in
CDK4/D1 (Table 7). There was a slight loss in activity
with 2-furanyl (24l) in CDK2/E and CDK4/D1, and the
1-methyl-3-pyrrolyl analogue (24m ) was about the same

5238 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24
Yue et al.
F igu r e 5. Superposition of 13q and ATP27 complexed to
CDK2. The CDK2/13q carbons are colored white, and the
CDK2/ATP carbons are colored purple. The magnesium atom
from the CDK2/ATP structure27 is colored green.
structure of 13q folds back onto itself in a U shape
conformation, which brings the C3 cyclohexyl and the
C5 glycinamide groups in close proximity. This confor-
mation results in the cyclohexyl and glycinamide groups
pointing toward the exterior of the protein and hence
are partially exposed to solvent. It also results in the
formation of a hydrogen bond between N17 of the
glycinamide group and the side chain oxygen of Asp 86
(Figure 4). The conformation of the C5 amide is coplanar
with the carbonyl at C4, which allows the hydrogen of
N9 to form an intramolecular hydrogen bond to the
carbonyl (2.89 Å). This is consistent with the proton shift
of the N9 hydrogen in the NMR spectra of 13q (δ 11.19)
as well as in other analogues. The indenopyrazole core
occupies the same region as the adenine ring of ATP,
and the hydrogen bond between Leu83 and N1 of 13q
mimics the hydrogen bond between the same backbone
nitrogen and the adenine ring of ATP (Figure 5).27,28
Other residues within 3.5 Å of 13q are Ile10, Lys 33,
Lys 89, Gln 131, and Asp145.
Although this X-ray structure does not include bound
cyclin, the differences in the ATP binding site of CDK2
are minimal and, therefore, provide useful information
for the understanding of how these inhibitors bind in
the active site. The crystal structure of 13q indicates
that there is space to accommodate groups at C3 of the
indenopyrazole core. This is consistent with the CDK2/E
enzyme binding data, which showed a variety of differ-
ent substituents gave good activity; however, straight
chain alkyl groups directly attached to the indenopy-
razole core at C3 were not good binders. Because the
C3 substituents point to the exterior of the protein,
bulkier groups that fill the pocket and maximize van
der Waals interactions might be less exposed to solvent
as compared with linear alkyl groups. This was also
evident in the para-substituted phenyl series in which
the activity decreased from methyl (5b) to longer chain
alkyls (5c-e). The p-phenoxy phenyl analogue (5i) had
poor activity, which suggests large hydrophobic groups
that extend too far out into the solvent-exposed areas
F igu r e 4. Structure of 13q in the CDK2 ATP binding pocket.
Protein carbons are colored light green, and inhibitor carbons
are colored white.
as the 2-thienyl compound. With the 2-thienyl as the
starting point, substitutions around the thiophene ring
were examined. When a methyl substituent was added
to the 3-position of the 2-thienyl group, the potency
dropped with a slight loss in both CDK2/E and CDK4/
D1 (24f). A 5-methyl group gave comparable activity for
CDK2/E, but it also showed a slight improvement in
activity for CDK4/D1 (24g). There seemed to be some
requirements for the groups that can be accommodated
in the 5-position of the thiophene. For example, the
5-ethyl ester (24h ) decreased the activity in both
CDK2/E and CDK4/D1 as compared with the 5-methyl
group (24g).
In going from a 2-thienyl to a 3-thienyl system, an
improvement in CDK4/D1 was again noticed. Compound
24i gave a slight increase in activity for CDK4/D1 as
compared with the 2-thienyl (24e) whereas the CDK2/E
remained the same. The 5-chloro-3-thienyl compound
(24j) was also prepared, and it was comparable to the
3-thienyl. Interestingly, the 2,5-dimethyl-3-thienyl de-
rivative (24k ) lost most of the CDK4/D1 activity while
maintaining CDK2/E potency.
X-r a y Cr ysta l Str u ctu r e of 13q. To determine the
binding mode of these inhibitors, an X-ray crystal
structure of 13q complexed to CDK2 was obtained (1.85
Å resolution). Compound 13q binds into the adenosine
5′-triphosphate (ATP) binding pocket of CDK2 and
shares the same bidentate hydrogen bond with Leu83
as seen in other CDK2 inhibitors.12,14,16,21-26 Both ni-
trogens of the indenopyrazole core of 13q are involved
in the hydrogen bonds with Leu83. The distance of N1
with the backbone nitrogen of Leu83 is 3.04 Å and that
of N2 with the backbone oxygen of Leu83 is 2.65 Å. The

Indenopyrazoles as Inhibitors
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5239
Ta ble 8. Cellular Activity of Selected Compounds
IC50 (nM)
compd
HCT116a
CDK4/D1a
CDK2/Ea
8c
8g
74
110
290
120
41
140
29
360
130
64
7
18
15
26
21
26
11
17
13
9
13k
13l
24e
24f
24g
24i
24j
24m
>1300
>99
150
350
53
67
57
130
13
26
F igu r e 6. Treatment of asynchronous AG1523 with 1.45 µM
24j results in Rb dephosphorylation.
a
Values correspond to n ) 2.
Ta ble 9. Enzymatic and Cellular Activity of 24j
First, 24j was tested for its inhibitory activity against
a broader panel of kinase targets (Table 9). Compound
24j showed good selectivity (>1000-fold) for CDK targets
as compared with non-CDK targets. Good inhibitory
activity (IC50 < 100 nM) was observed against each of
the three major cell cycle CDK targets tested with
maximum potency being recorded against CDK2/E (IC50
) 13 nM). The compound was approximately 4-fold less
potent on CDK4/D1 (IC50 ) 57 nM) and CDK1/B (IC50
) 44 nM).
Second, 24j was tested for inhibitory potency against
a broader panel of human and murine tumor cell lines.
The compound showed good inhibitory activity against
each of the additional six cell lines tested with IC50
values in the 200-690 nM range. One exception was
the HT29 human colon carcinoma cell line, which
appeared unresponsive to 24j treatment (IC50 > 2900
nM).
IC50 (nM)a
CDK1/B
c-abl
PKA
44
>60000
>58000
>58000
PKC
AG1523 (arrested)
B16-F1
HT29
EC50 > 29000
200
>2900
490
HT1080
MiaPaCa2
NCI-H460
PC3
260
240
690
a
Values correspond to n ) 2.
lead to poor binding. The similarly sized p-piperidino
(5k ) and p-morpholino (5l) derivatives are more hydro-
philic and, therefore, interact more favorably as evi-
denced by their increased binding (Table 1).
The enzyme data also showed that substituted gly-
cinamides at C5 usually gave better potency than the
smaller acetamides. The close proximity of the C5
glycinamide group in 13q with the C3 cyclohexyl group
may add to the stability of the complex by increasing
the number of van der Waals interactions while de-
creasing exposure to solvent. In addition, the greater
steric bulk of the glycinamides may contribute to
increased binding by occupying more of the pocket. As
can be seen from the CDK2/E enzyme binding data,
there was not much difference in activity between the
glycinamides as they showed similar potencies (Table
2). Primary ureas at C5 also had good activity in the
CDK2/E enzyme assay even though their size resembles
the acetamides more than the glycinamides.
Cellu la r Activity. The most active compounds in
CDK4/D1 and CDK2/E were assayed in an HCT116
colon carcinoma-derived cell line. The translation of the
enzymatic activity into cellular potency was quite good
(Table 8). In general, the heterocycles were more potent
than the substituted phenyls and alkyls and several
compounds showed interesting activity against HCT116.
Compound 24j represented one of the more potent
inhibitors, and it appeared to have differential activity
on tumor cells as compared to arrested normal human
fibroblasts. As shown in Table 9, the difference in
activity of 24j against HCT116 vs the normal human
fibroblast AG1523 is greater than 230-fold. This large
gap in activity for proliferating tumor cells vs arrested
normal cells was deemed important as it might lead to
a potentially useful therapeutic window in vivo. For this
reason, the cellular properties of 24j were examined in
more detail.
Compound 24j was further tested to determine if it
had cell cycle effects consistent with its profile of CDK
inhibition. These experiments were performed in normal
human fibroblasts (AG1523) to avoid confounding effects
of loss of checkpoint control in tumor-derived cell lines.
Treatment of subconfluent, asynchronously proliferating
AG1523 with 1.45 µM 24j resulted in loss of phospho-
rylated retinoblastoma (Rb), a CDK substrate, as indi-
cated by a slight increase in mobility on polyacrylamide
gel electrophoresis (PAGE). Loss of phosphorylated
substrate was apparent within 24 h of treatment and
complete by 48 h (Figure 6). Cell cycle progression
experiments were designed to examine synchronized cell
populations through one transit of the cell cycle (AG1523
doubling time is approximately 60 h, data not shown).
The impact of 24j on G1 progression in these cells was
tested by treatment of cells released from a G1 block.
The cells were arrested in G1 by growing to confluence.
Cells were stimulated to reenter the cell cycle by
subculture at lower density and followed by FACS
analysis for a 48 h period. At no time during the 48 h
observation period were cells arrested through contact
inhibition. Within 24 h of serum treatment, significant
progression of cells into G2/M had occurred in vehicle-
treated control cells as judged by FACS analysis (Figure
7, panel 4). This cell cycle progression is almost com-
pletely inhibited in cells treated with 348 nM 24j and
is totally blocked in cells treated with 10-fold higher
levels of compound. This is the expected response of cells
to a compound capable of inhibiting CDK4/D1 and
CDK2/E. The concentration of 24j at which cell cycle
progression is inhibited (approximately 350 nM) is

5240 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24
Yue et al.
F igu r e 7. FACS diagram showing the effect of 24j in G1-arrested AG1523.
F igu r e 8. FACS diagram showing the effect of 24j in G2-synchronized AG1523.
consistent with the in vitro potency of the compound
on CDK2/E (IC50 ) 13 nM).
CDK1/B as indicated in its CDK inhibition profile (Table
9). The higher concentration of compound needed to
completely block G2/M progression as compared to that
needed to block exit from a G1 block may reflect the
lower inhibitory potency of 24j against CDK1/B (44 nM)
as compared with that against CDK2/E (13 nM). When
HCT116 cells were treated with 24j and analyzed by
FACS analysis, the appearance of a sub-G1 peak oc-
curred within 24 h of treatment (data not shown).
HCT116 cells treated with 24j appeared to lift off the
culture plate and die. This observation, in combination
with the appearance of a sub-G1 peak during FACS
Similarly, the ability of 24j to impact cell cycle
progression through G2/M was tested by exposing cells
synchronized in G2/M to compound (Figure 8). When
these G2/M-synchronized cells were exposed to vehicle
alone, they progressed back into G1 within about 10 h.
However, cells treated with 348 nM 24j took longer (24-
48 h) to progress into G1 indicative of a slowing through
G2/M. Cells treated with 3.5 µM 24j remained com-
pletely blocked in G2/M. This result is biologically
consistent with the activity of a compound able to inhibit

Indenopyrazoles as Inhibitors
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5241
F igu r e 9. Compound 24j induces apoptosis in asynchronous
HCT116 tumor cells.
analysis, led us to examine the ability of 24j to cause
apoptosis in HCT116 cells in greater detail (Figure 9).
HCT116 cells were treated with 580 nM 24j and
samples of attached cells, and floaters were taken as a
function of time, harvested, and analyzed separately.
After 24 h of 24j treatment, inhibition of phosphoryla-
tion of the CDK substrate Rb was observed as would
be predicted. This loss of phosphorylated Rb was
observed predominantly in the floating population.
Furthermore, these floating cells also contained the 85
kDa cleaved form of PARP and the 20 kDa and 11 kDa
cleaved fragments of caspase 3. Cleavage of PARP and
caspase 3 occurs as a result of activation of upstream
proapoptotic caspases. Collectively, these results suggest
that in HCT116 cells treated with 24j, CDK inhibition
results in inhibition of Rb phosphorylation and activa-
tion of the apoptotic machinery. This leads to detach-
ment of the cells from the tissue culture dish and death.
As judged by cleavage of apoptotic markers, significant
death occurs within 24 h of treatment with 24j. Cleav-
age of apoptotic markers did not occur when 24j was
evaluated in the normal human fibroblast, AG1523,
under identical conditions (data not shown).
F igu r e 10. Graph of arrested AG1523 vs proliferating
HCT116 for 24j.
Exp er im en ta l Section
All reactions were run under an atmosphere of dry nitrogen.
All solvents were used without purification as acquired from
commercial sources. NMR spectra were obtained using either
a Varian (Palo Alto, CA) Unity-300, Inova-300, VXR-400, or
Inova-400 spectrometer. Chemical shifts are reported in parts
per million relative to tetramethylsilane (TMS) as internal
standard. Microanalyses were performed by Quantitative
Technologies Inc. and were within 0.4% of the calculated
values. For compounds where microanalysis was not obtained,
high-performance liquid chromatography (HPLC) purity was
determined to be >98%. Mass spectra were obtained using
either a Finnigan MAT8230 sector or MAT95S sector with API
interface system. Flash chromatography was done using EM
Science silica gel 60. HPLC purifications were performed on
a Rainin Dynamax SD300 instrument using a C18 reverse
phase column with acetonitrile/water (containing 0.05% trif-
luoroacetic acid (TFA)) as a mobile phase. HPLC purity values
were obtained using a Rainin Dynamax SD300 instrument
system with a photodiode array detector. Melting points were
determined in open glass capillaries using a Thomas-Hoover
UniMelt melting point apparatus and are uncorrected. Solu-
bilities were determined by HPLC analysis of supernatants
from a pH 7.4, 0.05 M potassium phosphate buffer-based assay.
CDK4/D1 En zym e In h ibition . CDK4/D1 complexes were
expressed in insect cells following dual infection by baculovirus
vectors containing each of the components, and extracts of
these cells were then prepared as described previously.29
CDK4/D1 kinase activity was measured in 96 well polypro-
pylene microtiter plates using a GST-60 kDa Rb fusion protein
(comprising amino acids 378 to 928 of Rb) and γ-32P-ATP and
capturing the 32P-labeled reaction products on GSH-Sepharose
beads.30 Briefly, each reaction (50 µL) contained 50 mM Tris-
HCl (pH 7.6), 10 mM MgCl2, 10% dimethyl sulfoxide (DMSO),
1 mM dithiothreitol (DTT), 50 µM ATP, 10 µg/mL bovine serum
albumin (BSA), 1% glycerol, 6 µg GST-Rb (60 kDa Rb), and
0.5 µCi [γ-32P]ATP. Kinase reactions were initiated by addition
of 4 units of CDK4/D1. One unit of CDK4/D1 kinase activity
results in the transfer of 1 pmol of 32P per minute at room
temperature to the GST-Rb fusion protein. After 15 min, 50
µL of stop buffer (phosphate-buffered saline (PBS) with 100
mM Na2EDTA, 10 mM ATP, 200 µg/mL BSA, and 0.2% NP-
40) was added to the reaction. Half of the reaction was then
transferred to a Millipore MHVB N45 filter plate containing
50 µL GSH-Sepharose beads (25% slurry in PBS with 0.8%
NP40 and 80 mM Na2EDTA) to capture the GST-Rb. The
plates were incubated at room temperature for 90 min with
The likelihood that the biology of 24j reflects CDK
target inhibition was further examined by evaluating
the activity of 24j on arrested normal fibroblasts where
cell cycle CDK activity is low or absent. Whereas 24j
had potent growth inhibitory activity on HCT116 cells
reflecting cell cycle inhibition and death, it lacked
activity in arrested fibroblasts (Figure 10). This lack of
activity is consistent with 24j killing through a CDK
inhibition mechanism.
Con clu sion
We have made many new compounds that explored
the structure-activity relationships (SAR) of the C3
position of our indenopyrazole series of CDK inhibitors.
We have shown that C3 can be substituted with
heterocycles and alkyls in addition to other phenyl
groups. A crystal structure was determined, which
identifies interactions necessary for good inhibition. As
expected, these compounds bind in the ATP pocket.
Compound 24j was found to be a unique inhibitor with
cell cycle effects expected from inhibitors of this type.
In addition, 24j was found to be active in a variety of
cancer cell lines. These data coupled with the inactivity
in the normal cell line would suggest that compounds
with this profile would provide an advantageous thera-
peutic window in vivo. The structural information and
SAR data will allow us to continue to generate CDK
inhibitors with better biological profiles.

5242 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24
Yue et al.
Ta ble 10. Crystallographic Statistics for CDK2/13q
continuous mixing on a plate shaker. After they were filtered,
each well was washed 5 times with 200 µL of PBS containing
0.5% NP-40. The samples were dried, and 50 µL of Microscint
scintillation fluid was added to each well. The 32P radioactivity
in each sample was determined using the Packard Top Count
scintillation counter. In a typical experiment, less than 10%
of the GST-Rb was phosphorylated and greater than 95% of
the radioactivity was found in Rb by sodium dodecyl sulfate
(SDS)-PAGE analysis.
CDK2/E En zym e In h ibition . CDK2/his-cyclinE was pre-
pared as previously described.31 CDK2/E kinase reactions were
carried out in 50 µL reactions containing 50 mM Tris-HCl (pH
7.6), 10 mM MgCl2, 10% DMSO, 1 mM DTT, 50 µM ATP, 0.05
µCi [γ-32P]ATP (2000 Ci/mmol, NEN Life Science Products),
and 6 µM GST-Rb fusion. After they were incubated at room
temperature, the reactions were terminated by addition of an
equal volume of cold PBS containing 100 mM ethylenedi-
aminetetraacetic acid (EDTA), 10 mM ATP, 200 µg/mL BSA,
and 0.2% NP40. Aliquots of the stopped reactions were
processed as described previously.30
space group
P212121
cell dimensions (Å)
maximal resolution (Å)
total observations
unique reflections
completeness (%)
a ) 71.98, b ) 72.20, c ) 53.52
1.70
329,259
31,367
99.9
0.065
27.1
20.0-1.85
0.212
0.256
31.1
a
Rmerge
mean I/σ (I)
refinement resolution (Å)
b
Rfactor
Rfree
c
avg protein temp factor (Å)
avg inhibitor temp factor (Å)
rms bonds (Å)
40.5
0.0067
1.60
rms angles (°)
a
Rmerge ) ∑h∑j|Ihj - hIh|/∑h∑j|Ihj|, where Ihj is the jth observation
b
of reflection h. Rfactor ) ∑h||Fobs| - |Fcalcd||/∑h|Fobs|, where Fobs and
Fcalcd are the observed and the calculated structure factor ampli-
c
tudes, respectively, for reflection h. Rfree ) Rfactor for a 9.2% subset
of reflections not used in the refinement.
P u r ifica t ion a n d Cr yst a lliza t ion of H u m a n CDK2.
CDK2 protein was prepared and purified as previously de-
scribed32 except for the addition of 10% (v/v) glycerol during
the SP-sepharose and ATP-agarose column steps. Protein was
concentrated to 6 mg/mL using a Collodion concentrator
against 10 mM HEPES, pH 7.4, 15 mM NaCl. Crystals were
grown at 18 °C from sitting drops containing premixed and
filtered solutions of 3.0 mg/mL CDK2, 32.5 mM HEPES, pH
7.4, 11.3 mM sodium chloride, 12.5 mM ammonium acetate, 2
mM DTT, and 2-4% PEG 4000 against 100 mM HEPES, pH
7.4, 50 mM ammonium acetate, 2 mM DTT, and 4-14% PEG
4000. Crystals appeared in 1 day and grew for 1 week to
approximately 0.25 mm × 0.25 mm in size.
bated for 5 (HCT116, NCI-H460, PC-3, MiaPaCa-s, HT-29, HT-
1080, and B16-F0) or 6 (AG1523) days at 37 °C in 5% CO2. A
50 µL amount of cold 50% TCA was gently added to each well,
and plates were placed at 4 °C for 1 h. Plates were decanted,
rinsed 5 times with cold tap water, and allowed to air dry. A
50 µL amount of 0.4% SRB in 1% acetic acid was added, and
the plates were incubated at room temperature for 15 min.
Plates were washed four times with 1% acetic acid and again
allowed to air-dry. A 150 µL amount of 10 mM Tris base was
added, and plates were agitated on a plate shaker for 5 min
before reading optical densities at 570 nm using a BIORAD
3550 plate reader.
Cell Cycle An a lysis. Normal human fibroblasts derived
from human foreskin (AG1523) were grown to confluence and
maintained at confluence for a minimum of 5 days. Cells were
trypsinized and subcultured at 8000 cells/cm2 in growth
medium containing 10% fetal calf serum, and the indicated
concentrations of compound were added either immediately
(G1 release) or 24 h later (G2 synchronized). Cells were
incubated in the presence of compound for up to 48 h, and
samples were harvested at the indicated time points after
compound addition. Cells were trypsinized and fixed in ice cold
ethanol for a minimum of 24 h. After ethanol was removed,
RnaseA (Sigma Chemicals) was added at 180 µg/mL for 30 min
at room temperature followed by the addition of propidium
iodide (Sigma Chemicals) at a final concentration of 50 µg/
mL. Samples were run on a Becton Dickinson FACScan flow
Cr ysta l P r ep a r a tion , Da ta Collection , a n d P r ocessin g.
To prevent the crystals from cracking when soaked with
inhibitor, CDK2 crystals were cross-linked in a solution of 0.1%
gluteraldehyde, 10 mM sodium phosphate, pH 7.5, and 15 mM
sodium chloride for 30 min. The cross-linked crystals were
soaked for 6 days in inhibitor solution (0.5 mM 13q, 0.5%
DMSO, 10 mM HEPES, pH 7.4, 15 mM sodium chloride) and
then briefly transferred into cryo-protectant (10 mM HEPES,
pH 7.4, 15 mM sodium chloride, 25% MPD) and flash frozen
in liquid nitrogen. Data were collected at the DND-CAT beam
line, Advanced Photon Source, Argonne National Laboratories.
Data were processed, scaled, and merged with the program
HKL.33
Str u ctu r e Solu tion a n d Refin em en t. Molecular replace-
ment using the program EPMR34 was used to generate an
initial phasing model from a previously determined in-house
structure of CDK2. After five cycles of refinement (positional
refinement, simulated annealing, and B-factor refinement)
using the program CNX,35 the electron density maps clearly
showed the location of inhibitor inside the ATP binding site.
Iterative cycles of refinement and model building were per-
formed resulting in a final model, which includes residues
1-35, 43-147,163-298, 13q, and 104 water molecules. Data
and refinement statistics are presented in Table 10.
cytometer equipped with
a 488 nM argon-ion laser and
analyzed using CELLQUEST software.
West er n Blot An a lysis. Asynchronous subconfluent
HCT116 or AG1523 cells were treated with compound or
vehicle control as indicated. Samples were taken at the
indicated time points with attached cells and floaters har-
vested separately. Floaters, when present, were obtained by
collection of the culture medium, washing of the monolayers,
and centrifugation of the pooled medium and wash at 1000
rpm. Cells were processed by lysing in an SDS-based buffer
containing phosphatase and proteinase cocktails and subjected
to analysis by SDS gel electrophoresis on appropriate (7.5 or
10% depending on protein to be detected) polyacrylamide gels.
Gel loading was normalized for total protein as determined
by BioRad’s DC protein analysis kit, and 5 µg of protein was
loaded per lane. Gels were blotted onto nitrocellulose mem-
branes. Membranes were probed with the following antibod-
ies: retinoblastoma (pRb14001A, Pharmingen); poly-ADP
ribose polymerase (PARP 8192-1, Clontech); and caspase 3
65906E (Pharmingen). The presence of bound antibody was
detected using the following enhanced chemiluminescence
procedure as follows: a species specific secondary antibody
conjugated to biotin was allowed to react with blots previously
probed with protein specific primary antibodies. After exten-
sive washes, blots were incubated with biotinylated horserad-
ish peroxidase (HRP) and avidin (Vector Laboratories Vectastain
Cellu la r Gr ow th In h ibition Assa ys. Effects of compounds
on growth of the following transformed cell lines was evalu-
ated; HCT116 (human colon carcinoma, ATCC), NCI-H460
(human lung carcinoma, ATCC), PC-3 (human prostate ad-
enocarcinoma, ATCC), MiaPaCa-2 (human pancreatic carci-
noma, ATCC), HT-29 (human colon adenocarcinoma, ATCC),
HT-1080 (human colon fibrosarcoma, ATCC) and B16-F0
(murine melanoma, ATCC). In addition, death effects on
normal human fibroblasts (AG1523; Coriell Institute for
Medical Research) were evaluated in a colorimetric assay using
sulforhodamine B (SRB).36 Briefly, exponentially growing cells
were seeded in wells of a 96 well microtiter plate at a
concentration to allow for 3-5 doublings before obtaining 85%
confluence (transformed cell lines) or at
a concentration
resulting in confluence and contact inhibition of growth
(AG1523). Eighteen hours later, graded concentrations of test
compounds were added to the cell plates. Plates were incu-

Indenopyrazoles as Inhibitors
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5243
ABC Kit). Following extensive washes, the HRP substrate
luminol (NEN Renaissance reagent) was added and allowed
to react for 1 min. Membranes were lightly hand-blotted dry,
exposed to Kodak X-OMAT Blue autoradiography film, and
developed.
1 H), 6.85 (d, J ) 9.0 Hz, 2 H), 3.04 (s, 6 H), 2.20 (s, 3 H).
HRMS (CI) calcd for C20H19N4O2 (M + H)+, 347.1508; found,
347.1496. Anal. (C20H18N4O2) C, H, N.
3-(4-P iper idin oph en yl)-5-(acetam ido)in den o[1,2-c]pyr a-
1
zol-4-on e (5k ). mp 289-291 °C. H NMR (300 MHz, DMSO-
d6): δ 10.26 (s, 1 H), 8.24 (d, J ) 8.0 Hz, 1 H), 8.07 (d, J ) 8.5
Hz, 2 H), 7.49 (t, J ) 8.0 Hz, 1 H), 7.21 (d, J ) 8.0 Hz, 1 H),
7.06 (d, J ) 8.5 Hz, 2 H), 3.30 (m, 4 H), 2.20 (s, 3 H), 1.60 (m,
6 H). HRMS (CI) calcd for C23H23N4O2 (M + H)+, 387.1821;
found, 387.1801. Anal. (C23H22N4O2) C, H, N.
Compounds 5a -m and 8a -h were prepared as illustrated
in Scheme 1 using the procedures as previously described19
with the appropriate starting materials and reagents.
3-P h en yl-5-(acetam ido)in den o[1,2-c]pyr azol-4-on e (5a).
1
mp >300 °C. H NMR (300 MHz, DMSO-d6): δ 10.18 (s, 1 H),
3-(4-Mor p h olin op h en yl)-5-(a cet a m id o)in d en o[1,2-c]-
p yr a zol-4-on e (5l). mp >300 °C. 1H NMR (300 MHz, DMSO-
d6): δ 10.23 (s, 1 H), 8.24 (d, J ) 9.0 Hz, 1 H), 8.11 (d, J ) 9.0
Hz, 2 H), 7.49 (t, J ) 9.0 Hz, 1 H), 7.21 (d, J ) 9.0 Hz, 1 H),
7.10 (d, J ) 9.0 Hz, 2 H), 3.76 (m, 4 H), 3.27 (m, 4 H), 2.20 (s,
3 H). HRMS (CI) calcd for C22H21N4O3 (M + H)+, 389.1614;
found, 389.1607. Anal. (C22H20N4O3) C, H, N.
3-(4-Meth ylth ioph en yl)-5-(acetam ido)in den o[1,2-c]pyr a-
zol-4-on e (5m ). mp 280-283 °C. 1H NMR (300 MHz, DMSO-
d6): δ 10.21 (s, 1 H), 8.22 (d, J ) 8.5 Hz, 1 H), 8.15 (d, J ) 8.5
Hz, 2 H), 7.49 (t, J ) 8.5 Hz, 1 H), 7.44 (d, J ) 8.5 Hz, 2 H),
7.20 (d, J ) 8.5 Hz, 1 H), 2.55 (s, 3 H), 2.20 (s, 3 H). HRMS
(CI) calcd for C19H16N3O2S (M + H)+, 350.0963; found, 350.0956.
Anal. (C19H15N3O2S) C, H, N.
8.25-8.21 (m, 3 H), 7.61-7.49 (m, 4 H), 7.23 (d, J ) 7.0 Hz, 1
H), 2.20 (s, 3 H). HRMS (CI) calcd for C18H14N3O2 (M + H)+,
304.1086; found, 304.1065. Anal. (C18H13N3O2‚0.25H2O) C, H,
N.
3-(4-Meth ylph en yl)-5-(acetam ido)in den o[1,2-c]pyr azol-
4-on e (5b). mp >300 °C. 1H NMR (300 MHz, DMSO-d6): δ
13.78 (s, 1 H), 10.18 (s, 1 H), 8.23 (d, J ) 8.8 Hz, 1 H), 8.11 (d,
J ) 8.1 Hz, 2 H), 7.50 (dd, J ) 8.0, 7.7 Hz, 1 H), 7.39 (d, J )
8.1 Hz, 2 H), 7.22 (d, J ) 7.3 Hz, 1 H), 2.39 (s, 3 H), 2.20 (s, 3
H). HRMS (CI) calcd for C19H16N3O2 (M + H)+, 318.1243;
found, 318.1222. Anal. (C19H15N3O2) C, H, N.
3-(4-Eth ylp h en yl)-5-(a ceta m id o)in d en o[1,2-c]p yr a zol-
1
4-on e (5c). mp 292-294 °C. H NMR (300 MHz, DMSO-d6):
3-(4-(N,N-Dim eth yla m in o)p h en yl)-5-(m or p h olin oa ce-
δ 10.19 (s, 1 H), 8.24 (d, J ) 8.5 Hz, 1 H), 8.14 (d, J ) 8.0 Hz,
2 H), 7.51 (t, J ) 8.5 Hz, 1 H), 7.43 (d, J ) 8.0 Hz, 2 H), 7.23
(d, J ) 8.5 Hz, 1 H), 2.69 (q, J ) 7.0 Hz, 2 H), 2.20 (s, 3 H),
1.22 (t, J ) 7.0 Hz, 3 H). HRMS (CI) calcd for C20H18N3O2 (M
+ H)+, 332.1399; found, 332.1388. Anal. (C20H17N3O2‚0.18H2-
NNH2) C, H, N.
1
ta m id o)in d en o[1,2-c]p yr a zol-4-on e (8a ). mp >300 °C. H
NMR (300 MHz, DMSO-d6): δ 11.56 (s, 1 H), 8.41 (d, J ) 8.5
Hz, 1 H), 8.08 (d, J ) 9.0 Hz, 2 H), 7.49 (t, J ) 8.5 Hz, 1 H),
7.22 (d, J ) 8.5 Hz, 1 H), 6.85 (d, J ) 9.0 Hz, 2 H), 3.82 (m, 4
H), 3.21 (s, 2 H), 3.03 (s, 6 H), 2.59 (m, 4 H). HRMS (ESI)
calcd for C24H26N5O3 (M + H)+, 432.2036; found, 432.2020.
Anal. (C24H25N5O3‚1.0TFA) C, H, N.
3-(4-n -P r op ylp h en yl)-5-(a ceta m id o)in d en o[1,2-c]p yr a -
1
zol-4-on e (5d ). mp 266-269 °C. H NMR (300 MHz, DMSO-
3-(4-(N,N-Dim eth yla m in o)p h en yl)-5-(4-h yd r oxyp ip er i-
d in oa ceta m id o)in d en o[1,2-c]p yr a zol-4-on e (8b). mp 264-
d6): δ 10.19 (s, 1 H), 8.24 (d, J ) 8.5 Hz, 1 H), 8.13 (d, J ) 8.0
Hz, 2 H), 7.51 (t, J ) 8.5 Hz, 1 H), 7.42 (d, J ) 8.0 Hz, 2 H),
7.23 (d, J ) 8.5 Hz, 1 H), 2.63 (t, J ) 7.5 Hz, 2 H), 2.20 (s, 3
H), 1.64 (sextet, J ) 7.5 Hz, 2 H), 0.92 (t, J ) 7.5 Hz, 3 H).
HRMS (CI) calcd for C21H20N3O2 (M + H)+, 346.1556; found,
346.1555. Anal. (C21H19N3O2) C, H, N.
1
267 °C. H NMR (300 MHz, DMSO-d6): δ 11.52 (s, 1 H), 8.40
(d, J ) 8.0 Hz, 1 H), 8.08 (d, J ) 9.0 Hz, 2 H), 7.49 (t, J ) 8.0
Hz, 1 H), 7.21 (d, J ) 8.0 Hz, 1 H), 6.83 (d, J ) 9.0 Hz, 2 H),
3.60 (m, 1 H), 3.16 (s, 2 H), 3.02 (s, 6 H), 2.80 (m, 2 H), 2.36
(m, 2 H), 1.83 (m, 2 H), 1.68 (m, 2 H). HRMS (ESI) calcd for
C
25H28N5O3 (M + H)+, 446.2192; found, 446.2206. Anal.
3-(4-n -Bu tylph en yl)-5-(acetam ido)in den o[1,2-c]pyr azol-
(C25H27N5O3) C, H, N.
1
4-on e (5e). mp 256-259 °C. H NMR (300 MHz, DMSO-d6):
3-(4-(N,N-Dim eth yla m in o)p h en yl)-5-(4-(a m in om eth yl)-
p ip er id in oa ceta m id o)in d en o[1,2-c]p yr a zol-4-on e (8c). 1H
NMR (300 MHz, DMSO-d6): δ 11.74 (s, 1 H), 8.38 (d, J ) 8.0
Hz, 1 H), 8.10 (d, J ) 9.0 Hz, 2 H), 7.48 (t, J ) 8.0 Hz, 1 H),
7.19 (d, J ) 8.0 Hz, 1 H), 6.80 (d, J ) 9.0 Hz, 2 H), 3.15 (s, 2
H), 3.02 (s, 6 H), 2.87 (m, 2 H), 2.59 (d, J ) 6.5 Hz, 2 H), 2.23
(m, 2 H), 1.71 (m, 2 H), 1.63-1.54 (m, 2 H), 1.33 (m, 1 H).
HRMS (ESI) calcd for C26H31N6O2 (M + H)+, 459.2508; found,
459.2508.
3-(4-(N,N-Dim et h yla m in o)p h en yl)-5-((4-m et h ylp ip er -
azin o)acetam ido)in den o[1,2-c]pyr azol-4-on e (8d). mp >300
°C. 1H NMR (300 MHz, DMSO-d6): δ 11.65 (s, 1 H), 8.40 (d, J
) 8.0 Hz, 1 H), 8.13 (d, J ) 9.0 Hz, 2 H), 7.49 (t, J ) 8.0 Hz,
1 H), 7.21 (d, J ) 8.0 Hz, 1 H), 6.81 (d, J ) 9.0 Hz, 2 H), 3.18
(s, 2 H), 3.02 (s, 6 H), 2.57 (m, 8 H), 2.28 (s, 3 H). HRMS (ESI)
calcd for C25H29N6O2 (M + H)+, 445.2352; found, 445.2359.
Anal. (C25H28N6O2‚0.25H2O) C, H, N.
3-(4-Mor p h olin op h en yl)-5-(m or p h olin oa ceta m id o)in -
d en o[1,2-c]p yr a zol-4-on e (8e). mp 257-258 °C. 1H NMR
(300 MHz, DMSO-d6): δ 11.56 (s, 1 H), 8.40 (d, J ) 8.0 Hz, 1
H), 8.11 (d, J ) 9.0 Hz, 2 H), 7.50 (t, J ) 8.0 Hz, 1 H), 7.22 (d,
J ) 8.0 Hz, 1 H), 7.11 (d, J ) 9.0 Hz, 2 H), 3.82 (m, 4 H), 3.75
(m, 4 H), 3.29 (m, 4 H), 3.20 (s, 2 H), 2.59 (m, 4 H). HRMS
(ESI) calcd for C26H28N5O4 (M + H)+, 474.2141; found, 474.2151.
3-(4-Mor p h olin op h en yl)-5-(4-h yd r oxyp ip er id in oa ceta -
m id o)in d en o[1,2-c]p yr a zol-4-on e (8f). mp 241-245 °C. 1H
NMR (300 MHz, DMSO-d6): δ 11.52 (s, 1 H), 8.41 (d, J ) 8.5
Hz, 1 H), 8.11 (d, J ) 9.0 Hz, 2 H), 7.50 (t, J ) 8.5 Hz, 1 H),
7.22 (d, J ) 8.5 Hz, 1 H), 7.09 (d, J ) 9.0 Hz, 2 H), 3.77 (m, 4
H), 3.60 (m, 1 H), 3.28 (m, 4 H), 3.16 (s, 2 H), 2.80 (m, 2 H),
2.33 (m, 2 H), 1.84 (m, 2 H), 1.70 (m, 2 H). HRMS (ESI) calcd
for C27H30N5O4 (M + H)+, 488.2298; found, 488.2290. Anal.
(C27H29N5O4‚0.25H2O) C, H, N.
δ 13.80 (s, 1 H), 10.19 (s, 1 H), 8.23 (d, J ) 8.5 Hz, 1 H), 8.13
(d, J ) 8.0 Hz, 2 H), 7.51 (t, J ) 8.5 Hz, 1 H), 7.42 (m, 3 H),
7.22 (d, J ) 8.5 Hz, 1 H), 2.65 (t, J ) 7.0 Hz, 2 H), 2.20 (s, 3
H), 1.59 (quintet, J ) 7.0 Hz, 2 H), 1.33 (sextet, J ) 7.0 Hz, 2
H), 0.91 (t, J ) 7.0 Hz, 3 H). HRMS (CI) calcd for C22H22N3O2
(M + H)+, 360.1712; found, 360.1701.
3-(4-Hyd r oxyp h en yl)-5-(a ceta m id o)in d en o[1,2-c]p yr a -
1
zol-4-on e (5f). mp 289 °C. H NMR (300 MHz, DMSO-d6): δ
13.60 (s, 1 H), 10.21 (s, 1 H), 8.24 (d, J ) 8.4 Hz, 1 H), 8.09 (d,
J ) 8.8 Hz, 2 H), 7.51 (dd, J ) 8.4, 7.3 Hz, 1 H), 7.23 (d, J )
7.0 Hz, 1 H), 6.95 (d, J ) 8.8 Hz, 2 H), 6.55 (s, 1 H), 2.21 (s, 3
H). HRMS (CI) calcd for C18H14N3O3 (M + H)+, 320.1035;
found, 320.1030. Anal. (C18H13N3O3) C, H, N.
3-(4-Eth oxyph en yl)-5-(acetam ido)in den o[1,2-c]pyr azol-
1
4-on e (5h ). mp 287-288 °C. H NMR (300 MHz, DMSO-d6):
δ 10.21 (s, 1 H), 8.24 (d, J ) 8.5 Hz, 1 H), 8.17 (d, J ) 9.0 Hz,
2 H), 7.50 (t, J ) 8.5 Hz, 1 H), 7.22 (d, J ) 8.5 Hz, 1 H), 7.13
(d, J ) 9.0 Hz, 2 H), 4.12 (q, J ) 7.5 Hz, 2 H), 2.20 (s, 3 H),
1.37 (t, J ) 7.5 Hz, 3 H). HRMS (CI) calcd for C20H18N3O3 (M
+ H)+, 348.1348; found, 348.1325. Anal. (C20H17N3O3) C, H,
N.
3-(4-P h en oxyp h en yl)-5-(a ceta m id o)in d en o[1,2-c]p yr a -
1
zol-4-on e (5i). mp >300 °C. H NMR (300 MHz, DMSO-d6):
δ 10.18 (s, 1 H), 8.24 (m, 3 H), 7.54-7.43 (m, 3 H), 7.25-7.11
(m, 6 H), 2.19 (s, 3 H). HRMS (CI) calcd for C24H18N3O3 (M +
H)+, 396.1348; found, 396.1320. Anal. (C24H17N3O3‚0.25H2O)
C, H, N.
3-(4-(N,N-Dim eth ylam in o)ph en yl)-5-(acetam ido)in den o-
1
[1,2-c]p yr a zol-4-on e (5j). mp >300 °C. H NMR (300 MHz,
DMSO-d6): δ 10.28 (s, 1 H), 8.23 (d, J ) 8.5 Hz, 1 H), 8.09 (d,
J ) 9.0 Hz, 2 H), 7.48 (t, J ) 8.5 Hz, 1 H), 7.20 (d, J ) 8.5 Hz,

5244 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24
Yue et al.
3-(4-Mor p h olin op h en yl)-5-(4-(a m in om et h yl)p ip er id i-
3-Eth yl-5-(a ceta m id o)in d en o[1,2-c]p yr a zol-4-on e (13b).
mp 242-244 °C. H NMR (300 MHz, DMSO-d6): δ 13.1 (brs,
1 H), 10.10 (s, 1 H), 8.16 (d, J ) 8.4 Hz, 1 H), 7.43 (dd, J )
7.9, 7.9 Hz, 1 H), 7.12 (d, J ) 7.0 Hz, 1 H), 2.67 (q, J ) 7.6 Hz,
2 H), 2.14 (s, 3 H), 1.29 (t, J ) 7.7 Hz, 3 H). HRMS (ESI) calcd
for C14H14N3O2 (M + H)+, 256.1086; found, 256.1090. Anal.
(C14H13N3O2‚0.5TFA) C, H, N.
3-n -P r op yl-5-(a cet a m id o)in d en o[1,2-c]p yr a zol-4-on e
(13c). mp 200-202 °C. 1H NMR (300 MHz, DMSO-d6): δ 13.12
(brs, 1 H), 10.11 (s, 1 H), 8.18 (d, J ) 8.5 Hz, 1 H), 7.46 (dd, J
) 8.4, 7.3 Hz, 1 H), 7.14 (d, J ) 6.6 Hz, 1 H), 2.65 (t, J ) 7.3
Hz, 2 H), 2.16 (s, 3 H), 1.78 (sextet, J ) 7.5 Hz, 2 H), 0.92 (t,
J ) 7.5 Hz, 3 H). HRMS (CI) calcd for C15H16N3O2 (M + H)+,
270.1243; found, 270.1240. Anal. (C15H15N3O2‚0.25TFA) C, H,
N.
1
n oa ceta m id o)in d en o[1,2-c]p yr a zol-4-on e (8g). mp 237-
1
240 °C. H NMR (300 MHz, DMSO-d6): δ 11.73 (s, 1 H), 8.41
(d, J ) 8.5 Hz, 1 H), 8.13 (d, J ) 9.0 Hz, 2 H), 7.50 (t, J ) 8.5
Hz, 1 H), 7.21 (d, J ) 8.5 Hz, 1 H), 7.07 (d, J ) 9.0 Hz, 2 H),
3.76 (m, 4 H), 3.27 (m, 4 H), 3.16 (s, 2 H), 2.89 (m, 2 H), 2.57
(d, J ) 6.5 Hz, 2 H), 2.22 (m, 2 H), 1.70 (m, 2 H), 1.62-1.53
(m, 2 H), 1.31 (m, 1 H). HRMS (ESI) calcd for C28H33N6O3 (M
+ H)+, 501.2614; found, 501.2619.
3-(4-Mor p h olin op h en yl)-5-((4-m eth ylp ip er a zin o)a ceta -
m id o)in d en o[1,2-c]p yr a zol-4-on e (8h ). mp 256-258 °C. 1H
NMR (300 MHz, DMSO-d6): δ 13.53 (brs, 1 H), 11.60 (s, 1 H),
8.41 (d, J ) 8.5 Hz, 1 H), 8.15 (d, J ) 9.0 Hz, 2 H), 7.50 (t, J
) 8.5 Hz, 1 H), 7.22 (d, J ) 8.5 Hz, 1 H), 7.08 (d, J ) 9.0 Hz,
2 H), 3.76 (m, 4 H), 3.27 (m, 4 H), 3.18 (s, 2 H), 2.57 (m, 8 H),
2.27 (s, 3 H). HRMS (ESI) calcd for C27H31N6O3 (M + H)+,
487.2458; found, 487.2447.
3-Isop r op yl-5-(a cet a m id o)in d en o[1,2-c]p yr a zol-4-on e
1
(13d ). mp >250 °C. H NMR (300 MHz, DMSO-d6): δ 13.08
(s, 1 H), 10.10 (s, 1 H), 8.14 (d, J ) 8.4 Hz, 1 H), 7.41 (dd, J )
8.5, 7.3 Hz, 1 H), 7.09 (d, J ) 7.4 Hz, 1 H), 2.98 (septet, J )
6.9 Hz, 1 H), 2.12 (s, 3 H), 1.28 (d, J ) 7.0 Hz, 6 H). HRMS
(ESI) calcd for C15H16N3O2 (M + H)+, 270.1243; found, 270.1258.
Anal. (C15H15N3O2‚0.15H2O) C, H, N.
2-Acetyl-4-n itr o-1H-in d en e-1,3(2H)-d ion e (10a ). A solu-
tion of 3-nitrophthalic anhydride (4.2 g, 20 mmol) and 2,4-
pentanedione (2.0 mL, 20 mmol) in acetic anhydride (11.3 mL,
120 mmol) at 25 °C was treated dropwise with triethylamine
(5.6 mL, 40 mmol). The reaction mixture was stirred at 25 °C
for 30 min and quenched with 1 M HCl (45 mL). The solid
that precipitated was filtered and washed with ether (2 × 25
mL) and hexane (3 × 25 mL) to give the nitrotrione 10a (3.7
g, 80%) as a brown solid; mp 144-145 °C. 1H NMR (300 MHz,
DMSO-d6): δ 8.05 (dd, J ) 7.0, 1.5 Hz, 1 H), 7.96-7.87 (m, 2
H), 2.49 (s, 3 H). HRMS (CI) calcd for C11H8NO5 (M + H)+,
234.0402; found, 234.0395. Anal. (C11H7NO5) C, H, N.
2-Cyclop r op ylca r b on yl-4-n it r o-1H-in d en e-1,3(2H)-d i-
on e (10e). A solution of sodium (2.3 g, 0.1 mol) in ethanol (200
mL) was treated dropwise with cyclopropyl methyl ketone (9.9
mL, 0.1 mol) and ethyl trifluoroacetate (11.9 mL, 0.1 mol). The
reaction mixture was stirred at room temperature for 16 h,
diluted with 1 M HCl (50 mL), and extracted with ethyl
acetate. The organic extract was dried (MgSO4), filtered, and
concentrated in vacuo to give the dione 20e (9.1 g, 50%) as a
pale yellow oil. This was used without further purification.
Dione 20e (2.0 g, 11 mmol) was treated with the conditions
described for the preparation of 10a to give trione 10e (2.1 g,
2-Acetyl-4-a m in o-1H-in d en e-1,3(2H)-d ion e (11a ). A so-
lution of 10a (700 mg, 3.0 mmol), zinc dust (6.4 g, 99 mmol),
and calcium chloride (220 mg, 2.0 mmol) in 4:1 ethanol/water
(22 mL) was refluxed for 1.5 h. The reaction mixture was
filtered hot over Celite and washed with hot 4:1 ethanol/water
(50 mL). The filtrate was concentrated in vacuo, the crude
residue was dissolved in methanol (10 mL), and the solid was
filtered. The filtrate was concentrated in vacuo, and the solid
was triturated with ether (10 mL) and hexane (20 mL) to give
the anilinetrione 11a (500 mg, 82%) as an orange solid; mp
1
74%) as a bright yellow solid; mp 158-160 °C. H NMR (300
MHz, DMSO-d6): δ 7.80 (dd, J ) 6.2, 2.6 Hz, 1 H), 7.73-7.69
(m, 2 H), 3.28-3.20 (m, 1 H), 0.88-0.84 (m, 2 H), 0.79-0.75
(m, 2 H). HRMS (CI) calcd for C13H10NO5 (M + H)+, 260.0559;
found, 260.0554.
3-Cyclop r op yl-5-(a cet a m id o)in d en o[1,2-c]p yr a zol-4-
on e (13e). mp 220-221 °C. 1H NMR (300 MHz, DMSO-d6): δ
13.20 (brs, 1 H), 10.05 (s, 1 H), 8.13 (d, J ) 8.8 Hz, 1 H), 7.39
(dd, J ) 8.5, 7.4 Hz, 1 H), 7.07 (d, J ) 7.3 Hz, 1 H), 2.11 (s, 3
H), 1.94-1.90 (m, 1 H), 1.10 (d, J ) 6.6 Hz, 4 H). HRMS (CI)
calcd for C15H13N3O2 (M+), 267.1008; found, 267.1011. Anal.
(C15H13N3O2‚0.2TFA) C, H, N.
1
>265 °C. H NMR (300 MHz, DMSO-d6): δ 7.22 (dd, J ) 8.1,
7.3 Hz, 1 H), 6.75 (d, J ) 8.1 Hz, 1 H), 6.68 (d, J ) 6.9 Hz, 1
H), 6.29 (brs, 2 H), 2.31 (s, 3 H). HRMS (CI) calcd for C11H10
-
NO3 (M + H)+, 204.0661; found, 204.0685.
N-(2-Acetyl-1,3-d ioxo-2,3-d ih yd r o-1H-in d en -4-yl)a ceta -
m id e (12a ). A solution of 11a (450 mg, 2.2 mmol) in acetic
anhydride (10 mL) was refluxed for 2 h. The reaction mixture
was cooled to room temperature, and the solid was filtered.
The filtrate was diluted with heptane and concentrated in
vacuo to give the acetamide 12a (390 mg, 72%) as a tan solid;
3-Isobu tyl-5-(acetam ido)in den o[1,2-c]pyr azol-4-on e (13f).
mp 209-211 °C. 1H NMR (300 MHz, DMSO-d6): δ 13.12 (brs,
1 H), 10.12 (s, 1 H), 8.18 (d, J ) 8.4 Hz, 1 H), 7.46 (dd, J )
8.4, 7.7 Hz, 1 H), 7.14 (d, J ) 7.0 Hz, 1 H), 2.55 (d, J ) 7.4 Hz,
2 H), 2.19-2.11 (m, 1 H), 2.16 (s, 3 H), 0.91 (d, J ) 6.6 Hz, 6
H). HRMS (ESI) calcd for C16H18N3O2 (M + H)+, 284.1399;
found, 284.1413. Anal. (C16H17N3O2‚0.25TFA) C, H, N.
3-n -Bu tyl-5-(acetam ido)in den o[1,2-c]pyr azol-4-on e (13g).
1
mp >265 °C. H NMR (300 MHz, DMSO-d6): δ 10.72 (s, 1 H),
8.41 (d, J ) 8.1 Hz, 1 H), 7.49 (dd, J ) 8.1, 7.3 Hz, 1 H), 7.14
(d, J ) 7.0 Hz, 1 H), 2.31 (s, 3 H), 2.18 (s, 3 H). HRMS (CI)
calcd for C13H12NO4 (M + H)+, 246.0766; found, 246.0758.
1
mp 187-189 °C. H NMR (300 MHz, DMSO-d6): δ 13.10 (s, 1
3-Meth yl-5-(acetam ido)in den o[1,2-c]pyr azol-4-on e (13a).
A solution of 12a (300 mg, 1.2 mmol), hydrazine (77 µL, 2.4
mmol), and p-toluenesulfonic acid (12 mg, 60 µmol) in n-
butanol (20 mL) was refluxed for 4.5 h. Additional hydrazine
(19 µL, 0.6 mmol and 77 µL, 2.4 mmol) was added at 4 and 2
h, respectively. The reaction mixture was cooled and concen-
trated in vacuo to give a crude residue. Purification by reverse
phase HPLC gave the indenopyrazole 13a (19 mg, 6%) as a
H), 10.09 (s, 1 H), 8.16 (d, J ) 8.8 Hz, 1 H), 7.43 (dd, J ) 7.7,
7.7 Hz, 1 H), 7.11 (d, J ) 7.4 Hz, 1 H), 2.65 (t, J ) 7.7 Hz, 2
H), 2.14 (s, 3 H), 1.72 (quintet, J ) 7.5 Hz, 2 H), 1.30 (sextet,
J ) 7.4 Hz, 2 H), 0.88 (t, J ) 7.4 Hz, 3 H). HRMS (CI) calcd
for C16H18N3O2 (M + H)+, 284.1399; found, 284.1398. Anal.
(C16H17N3O2‚1.0TFA) C, H, N.
3-t-Bu tyl-5-(acetam ido)in den o[1,2-c]pyr azol-4-on e (13h ).
mp 234-236 °C. 1H NMR (300 MHz, DMSO-d6): δ 12.98 (brs,
1 H), 10.12 (s, 1 H), 8.16 (d, J ) 8.4 Hz, 1 H), 7.41 (dd, J )
8.4, 7.3 Hz, 1 H), 7.11 (d, J ) 7.3 Hz, 1 H), 2.13 (s, 3 H), 1.33
(s, 9 H). HRMS (CI) calcd for C16H18N3O2 (M + H)+, 284.1399;
found, 284.1395. Anal. (C16H17N3O2‚1.0TFA) C, H, N.
3-Cycloh e xyl-5-(a ce t a m id o)in d e n o[1,2-c]p yr a zol-4-
on e (13i). mp >250 °C. 1H NMR (300 MHz, DMSO-d6): δ
13.07 (brs, 1 H), 10.11 (s, 1 H), 8.15 (d, J ) 8.4 Hz, 1 H), 7.41
(dd, J ) 8.8, 7.0 Hz, 1 H), 7.10 (d, J ) 7.3 Hz, 1 H), 2.75-2.60
(m, 1 H), 2.13 (s, 3 H), 1.95-1.85 (m, 2 H), 1.80-1.50 (m, 5
H), 1.40-1.15 (m, 3 H). HRMS (ESI) calcd for C18H20N3O2 (M
+ H)+, 310.1566; found, 310.1564. Anal. (C18H19N3O2‚1.0H2O)
C, H, N.
1
tan solid; mp >250 °C. H NMR (400 MHz, DMSO-d6, 30 °C):
δ 13.06 (brs, 1 H), 10.08 (s, 1 H), 8.16 (d, J ) 8.3 Hz, 1 H),
7.44 (dd, J ) 8.5, 7.3 Hz, 1 H), 7.12 (d, J ) 7.3 Hz, 1 H), 2.33
(s, 3 H), 2.15 (s, 3 H). HRMS (CI) calcd for C13H12N3O2 (M +
H)+, 242.0930; found, 242.0928. Anal. (C13H11N3O2‚1.0TFA) C,
H, N.
Compounds 13b-i,r -w were prepared as illustrated in
Scheme 2 using the procedure described for 13a using the
appropriate starting materials and reagents. Compounds
13j-q were prepared as illustrated in Scheme 2 using the
procedures as previously described19 with the appropriate
starting materials and reagents.

Indenopyrazoles as Inhibitors
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5245
3-Meth yl-8-n itr oin den o[1,2-c]pyr azol-4-on e (15). Meth od
A. A solution of 10a (100 mg, 0.4 mmol), hydrazine (27 µL,
0.9 mmol), and p-toluenesulfonic acid (4 mg, 22 µmol) in
ethanol (1 mL) was refluxed for 1 h. The reaction mixture was
cooled to room temperature, and the solid was filtered and
washed with cold ethanol (10 mL) to give the hydrazone 14
(75 mg, 71%) as an orange-yellow solid. 1H NMR (300 MHz,
DMSO-d6): δ 7.87 (dd, J ) 6.2, 2.2 Hz, 1 H), 7.77-7.71 (m, 2
H), 5.59 (s, 2 H), 2.52 (s, 3 H). A solution of hydrazone 14 (75
mg, 0.3 mmol) was treated with the conditions described for
the preparation of 13a to give 15 (46 mg, 68%) as a solid.
Meth od B. A solution of 10a was treated with the condi-
tions described for the preparation of 13a to give 15. mp 224-
225 °C. 1H NMR (300 MHz, DMSO-d6): δ 13.41 (brs, 1 H),
8.16 (d, J ) 7.3 Hz, 1 H), 7.83 (d, J ) 6.9 Hz, 1 H), 7.55 (dd,
J ) 8.1, 7.3 Hz, 1 H), 2.36 (s, 3 H). HRMS (CI) calcd for
(400 MHz, DMSO-d6, 120 °C): δ 11.14 (s, 1 H), 8.28 (d, J )
8.6 Hz, 1 H), 7.65 (brs, 2 H), 7.52 (dd, J ) 8.3, 7.3 Hz, 1 H),
7.14 (d, J ) 7.1 Hz, 1 H), 3.33 (s, 2 H), 3.00 (d, J ) 10.5 Hz,
2 H), 2.78 (d, J ) 6.9 Hz, 2 H), 2.72 (q, J ) 7.6 Hz, 2 H), 2.44-
2.40 (m, 2 H), 1.82 (d, J ) 13.4 Hz, 2 H) 1.74-1.68 (m, 1 H),
1.61-1.51 (m, 2 H), 1.34 (t, J ) 7.6 Hz, 3 H). HRMS (ESI)
calcd for C20H26N5O2 (M + H)+, 368.2086; found, 368.2078.
Anal. (C20H25N5O2‚2.0TFA) C, H, N.
3-Isop r op yl-5-(4-(a m in om eth yl)p ip er id in oa ceta m id o)-
in d en o[1,2-c]p yr a zol-4-on e (13o). mp 187-189 °C. 1H NMR
(400 MHz, DMSO-d6, 120 °C): δ 11.40 (s, 1 H), 8.33 (d, J )
8.5 Hz, 1 H), 7.42 (dd, J ) 8.6, 7.1 Hz, 1 H), 7.11 (dd, J ) 7.3,
7.1 Hz, 1 H), 3.12 (s, 2 H), 3.10-3.01 (m, 2 H), 2.90-2.85 (m,
2 H), 2.56 (d, J ) 6.4 Hz, 2 H), 2.33-2.21 (m, 2 H), 1.75-1.65
(m, 2 H), 1.60-1.45 (m, 2 H), 1.36 (d, J ) 6.8 Hz, 6 H). HRMS
(ESI) calcd for C21H28N5O2 (M + H)+, 382.2243; found, 382.2254.
Anal. (C21H27N5O2‚2.0TFA) C, H, N.
3-Cyclop r op yl-5-(4-(a m in om eth yl)p ip er id in oa ceta m i-
d o)in d en o[1,2-c]p yr a zol-4-on e (13p ). mp 138-140 °C. 1H
NMR (400 MHz, DMSO-d6, 120 °C): δ 11.24 (s, 1 H), 8.30 (d,
J ) 8.5 Hz, 1 H), 7.65 (brs, 2 H), 7.43 (dd, J ) 8.5, 7.3 Hz, 1
H), 7.11 (d, J ) 7.1 Hz, 1 H), 3.22 (s, 2 H), 2.95 (app. d, J )
11.5 Hz, 2 H), 2.79 (d, J ) 6.8 Hz, 2 H), 2.35 (app. t, J ) 11.2
Hz, 2 H), 2.05-1.95 (m, 1 H), 1.81 (app. d, J ) 14.2 Hz, 2 H),
1.70-1.62 (m, 1 H), 1.59 (dd, J ) 11.4, 3.6 Hz, 1 H), 1.53 (dd,
J ) 10.9, 3.8 Hz, 1 H), 1.20-1.05 (m, 4 H). HRMS (ESI) calcd
for C21H26N5O2 (M + H)+, 380.2086; found, 380.2079. Anal.
(C21H25N5O2‚2.0TFA) C, H, N.
3-Cycloh exyl-5-(4-(am in om eth yl)piper idin oacetam ido)-
in d en o[1,2-c]p yr a zol-4-on e (13q). mp 196-198 °C. 1H NMR
(400 MHz, DMSO-d6, 120 °C): δ 11.19 (s, 1 H), 8.26 (d, J )
8.5 Hz, 1 H), 7.83-7.63 (brs, 2 H), 7.44 (dd, J ) 8.5, 8.0 Hz, 1
H), 7.13 (d, J ) 7.1 Hz, 1 H), 3.38 (s, 2 H), 3.06-3.01 (m, 1 H),
2.79-2.75 (m, 2 H), 2.01-1.95 (m, 4 H), 1.85-1.78 (m, 4 H),
1.77-1.57 (m, 5 H), 1.51-1.29 (m, 6 H). HRMS (ESI) calcd
for C24H32N5O2 (M + H)+, 422.2556; found, 422.2540. Anal.
(C24H31N5O2‚1.0TFA) C, H, N.
C
11H8N3O3 (M + H)+, 230.0566; found, 230.0574. Anal.
(C11H7N3O3‚0.1H2O) C, H, N.
3-Meth yl-8-a m in oin d en o[1,2-c]p yr a zol-4-on e (16). A so-
lution of 15 was treated with the conditions described for the
1
preparation of 11a to give 16 as a solid; mp 186-187 °C. H
NMR (300 MHz, DMSO-d6): δ 7.01 (dd, J ) 8.0, 7.4 Hz, 1 H),
6.77 (d, J ) 7.7 Hz, 1 H), 6.73 (d, J ) 7.3 Hz, 1 H), 2.28 (s, 3
H). HRMS (ESI) calcd for C11H10N3O (M + H)+, 200.0824;
found, 200.0831.
3-Meth yl-8-(acetam ido)in den o[1,2-c]pyr azol-4-on e (17).
A solution of 16 (10 mg, 50 µmol) in dioxane (1 mL) was treated
with saturated sodium bicarbonate (1 drop) and acetyl chloride
(1 drop) and stirred at room temperature for 1 h. The reaction
mixture was concentrated in vacuo and purified by reverse
phase HPLC to give the indenopyrazole 17 (5 mg, 42%) as a
pale yellow solid; mp 239-241 °C. 1H NMR (400 MHz, DMSO-
d6, 30 °C): δ 13.03 (brs, 1 H), 9.18 (brs, 1 H), 7.79 (br s, 1 H),
7.31-7.26 (m, 2 H), 2.34 (s, 3 H), 2.10 (s, 3 H). HRMS (CI)
calcd for C13H12N3O2 (M + H)+, 242.0930; found, 242.0933.
3-Eth yl-5-((4-ca r ba m oylp ip er id in o)a ceta m id o)in d en o-
[1,2-c]p yr a zol-4-on e (13j). mp 208-210 °C. 1H NMR (400
MHz, DMSO-d6, 140 °C): δ 10.85 (s, 1 H), 8.20 (d, J ) 8.3 Hz,
1 H), 7.45 (dd, J ) 8.5, 7.4 Hz, 1 H), 7.16 (d, J ) 7.0 Hz, 1 H),
6.57 (brs, 2 H), 4.09 (brs, 1 H), 3.58 (s, 2 H), 3.16 (d, J ) 10.7
Hz, 2 H), 2.73 (q, J ) 7.6 Hz, 2 H), 2.67-2.65 (m, 2 H), 2.31-
2.25 (m, 1 H), 1.99-1.86 (m, 4 H), 1.34 (t, J ) 7.6 Hz, 3 H).
HRMS (ESI) calcd for C20H24N5O3 (M + H)+, 382.1879; found,
382.1886. Anal. (C20H23N5O3‚1.0TFA) C, H, N.
3-Isop r op yl-5-((4-ca r b a m oylp ip er id in o)a cet a m id o)-
in d en o[1,2-c]p yr a zol-4-on e (13k ). mp 213-215 °C. 1H NMR
(400 MHz, DMSO-d6, 30 °C): δ 13.04 (brs, 1 H), 11.34 (s, 1
H), 8.35 (d, J ) 8.5 Hz, 1 H), 7.44 (dd, J ) 8.1, 7.5 Hz, 1 H),
7.13 (d, J ) 7.3 Hz, 1 H), 6.68 (brs, 2 H), 3.12 (s, 2 H), 3.07-
3.00 (m, 1 H), 2.89-2.86 (m, 2 H), 2.21-2.16 (m, 2 H), 2.12-
2.08 (m, 1 H), 1.89-1.79 (m, 2 H), 1.74-1.72 (m, 2 H), 1.31 (d,
J ) 6.9 Hz, 6 H). HRMS (ESI) calcd for C21H26N5O3 (M + H)+,
396.2036; found, 396.2045. Anal. (C21H25N5O3‚1.7TFA) C, H,
N.
3-Cyclop r op yl-5-((4-ca r ba m oylp ip er id in o)a ceta m id o)-
in d en o[1,2-c]p yr a zol-4-on e (13l). mp 178-180 °C. 1H NMR
(400 MHz, DMSO-d6, 120 °C): δ 10.92 (brs, 1 H), 8.21 (d, J )
8.3 Hz, 1 H), 7.44 (dd, J ) 8.3, 7.3 Hz, 1 H), 7.14 (d, J ) 6.9
Hz, 1 H), 6.62 (brs, 2 H), 3.60-3.42 (m, 2 H), 3.17-3.04 (m, 2
H), 2.68-2.51 (m, 2 H), 2.30-2.21 (m, 1 H), 2.03-1.97 (m, 1
H), 1.96-1.84 (m, 4 H), 1.19-1.01 (m, 4 H). HRMS (ESI) calcd
for C21H24N5O3 (M + H)+, 394.1879; found, 394.1876. Anal.
(C21H23N5O3‚1.0TFA) C, H; N: calcd, 13.80; found, 14.51.
3-Cycloh exyl-5-((4-ca r ba m oylp ip er id in o)a ceta m id o)-
in d en o[1,2-c]p yr a zol-4-on e (13m ). mp 229-231 °C. 1H NMR
(400 MHz, DMSO-d6, 120 °C): δ 11.60 (brs, 1 H), 8.16 (d, J )
8.3 Hz, 1 H), 7.46 (dd, J ) 8.3, 7.3 Hz, 1 H), 7.17 (d, J ) 7.1
Hz, 1 H), 6.67 (brs, 2 H), 3.82-3.68 (m, 2 H), 3.31-3.18 (m, 2
H), 2.79-2.74 (m, 3 H), 2.38-2.26 (m, 1 H), 1.99-1.91 (m, 6
H), 1.82-1.78 (m, 2 H), 1.72-1.64 (m, 3 H), 1.41-1.30 (m, 3
H). HRMS (ESI) calcd for C24H30N5O3 (M + H)+, 436.2356;
found, 436.2349. Anal. (C24H29N5O3‚1.25TFA) C, H, N.
3-E t h yl-5-(4-(a m in om e t h yl)p ip e r id in oa ce t a m id o)-
in d en o[1,2-c]p yr a zol-4-on e (13n ). mp 174-176 °C. 1H NMR
P h en yl 1,3-d ioxo-2-p r op ion yl-2,3-d ih yd r o-1H-in d en -4-
ylca r ba m a te (22a ). A solution of aniline 11b (1.1 g, 5.1
mmol), made using the chemistry described for 10a and 11a ,
and sodium carbonate (2.2 g, 200 wt %) in acetone (20 mL)
was treated with phenylchloroformate (0.76 mL, 6.1 mmol).
The reaction mixture was stirred at 50 °C for 30 min, cooled
to room temperature, and diluted with water (20 mL) and ethyl
acetate (20 mL). The organic phase was dried (MgSO4),
filtered, and concentrated in vacuo to give a solid. Trituration
of the solid with ether gave the carbamate 22a (1.2 g, 71%) as
1
a tan solid; mp 146-148 °C. H NMR (300 MHz, DMSO-d6):
δ 10.06 (s, 1 H), 8.22 (d, J ) 8.1 Hz, 1 H), 7.74 (dd, J ) 8.1,
7.7 Hz, 1 H), 7.45-7.40 (m, 3 H), 7.29-7.24 (m, 3 H), 2.90 (q,
J ) 7.7 Hz, 2 H), 1.15 (t, J ) 7.5 Hz, 3 H). HRMS (ESI) calcd
for C19H16NO5 (M + H)+, 338.1028; found, m/z 338.1020.
N-(1,3-Dioxo-2-p r op ion yl-2,3-d ih yd r o-1H -in d en -4-yl)-
u r ea (23a ). A solution of carbamate 22a (340 mg, 1.0 mmol)
and ammonium hydroxide (0.28 mL, 2.0 mmol) in methylsul-
foxide (5 mL) was heated at 90 °C for 4 h. The reaction mixture
was concentrated in vacuo to give a crude residue. Purification
by flash column chromatography (silica, ethyl acetate:hexane
1:1) gave the urea 23a (190 mg, 71%) as a light yellow solid;
mp >250 °C. 1H NMR (300 MHz, DMSO-d6): δ 8.45 (d, J )
8.5 Hz, 0.6 H), 8.40 (d, J ) 8.5 Hz, 0.4 H), 7.53-7.46 (m, 1 H),
7.12 (d, J ) 7.4 Hz, 1 H), 6.72 (brs, 2 H), 2.89-2.80 (m, 2 H),
1.22-1.15 (m, 3 H).
3-E t h yl-5-(ca r b a m oyla m in o)in d en o[1,2-c]p yr a zol-4-
on e (24a ). Trione 23a was treated with the conditions
described for the preparation of 13a ; mp 250 °C. 1H NMR (300
MHz, DMSO-d6): δ 9.27 (s, 1 H), 8.19 (d, J ) 8.8 Hz, 1 H),
7.35 (dd, J ) 8.7, 7.2 Hz, 1 H), 6.97 (d, J ) 7.3 Hz, 1 H), 6.75
(brs, 2 H), 2.79 (q, J ) 7.6 Hz, 2 H), 1.33 (t, J ) 7.7 Hz, 3 H).
HRMS (CI) calcd for C13H13N4O2 (M + H)+, 257.1039; found,
257.1033. Anal. (C13H12N4O2) C, H, N.
Compounds 24b-m were prepared as illustrated in Scheme
4 using the procedure described for 24a using the appropriate
starting materials and reagents.

5246 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24
Yue et al.
3-Isop r op yl-5-(ca r ba m oyla m in o)in d en o[1,2-c]p yr a zol-
4-on e (24b). mp >250 °C. H NMR (300 MHz, DMSO-d6): δ
3-(3-Meth ylth ien -2-yl)-5-(ca r ba m oyla m in o)in d en o[1,2-
c]p yr a zol-4-on e (24f). mp 270 °C (dec). H NMR (300 MHz,
1
1
13.00 (brs, 1 H), 9.27 (s, 1 H), 8.16 (d, J ) 8.7 Hz, 1 H), 7.31
(dd, J ) 8.8, 7.3 Hz, 1 H), 6.93 (d, J ) 7.0 Hz, 1 H), 6.70 (brs,
2 H), 2.98 (quintet, J ) 6.9 Hz, 1 H), 1.28 (d, J ) 6.6 Hz, 6 H).
HRMS (CI) calcd for C14H15N4O2 (M + H)+, 271.1195; found,
271.1196. Anal. (C14H14N4O2‚0.5TFA) C, H; N: calcd, 17.12;
found, 10.23.
3-Cyclop r op yl-5-(ca r ba m oyla m in o)in d en o[1,2-c]p yr a -
zol-4-on e (24c). mp 252-253 °C. 1H NMR (400 MHz, DMSO-
d6, 120 °C): δ 12.80 (brs, 1 H), 9.20 (s, 1 H), 8.14 (d, J ) 8.6
Hz, 1 H), 7.31 (dd, J ) 8.6, 7.4 Hz, 1 H), 6.93 (d, J ) 7.2 Hz,
1 H), 6.31 (brs, 2 H), 2.01-1.94 (m, 1 H), 1.18-1.08 (m, 4 H).
HRMS (CI) calcd for C14H12N4O2 (M+)+, 268.0960; found,
268.0972. Anal. (C14H12N4O2‚0.15TFA) C, H, N.
3-Cycloh exyl-5-(car bam oylam in o)in den o[1,2-c]pyr azol-
4-on e (24d ). mp 178-179 °C. 1H NMR (300 MHz, DMSO-d6):
δ 12.98 (brs, 1 H), 9.30 (s, 1 H), 8.16 (d, J ) 8.4 Hz, 1 H), 7.31
(dd, J ) 8.5, 7.3 Hz, 1 H), 6.93 (d, J ) 7.0 Hz, 1 H), 6.70 (brs,
2 H), 2.70-2.62 (m, 1 H), 1.95-1.88 (m, 2 H), 1.75-1.53 (m, 4
H), 1.36-1.10 (m, 4 H). HRMS (ESI) calcd for C17H19N4O2 (M
+ H)+, 311.1508; found, 311.1500. Anal. (C17H18N4O2‚0.25TFA)
C, H, N.
DMSO-d6): δ 13.41 (brs, 1 H), 9.25 (brs, 1 H), 8.25 (d, J ) 8.8
Hz, 1 H), 7.71 (brs, 1 H), 7.41 (dd, J ) 8.8, 7.3 Hz, 1 H), 7.21-
7.15 (m, 2 H), 6.85-6.73 (m, 2 H), 2.41 (s, 3 H). HRMS (ESI)
calcd for C16H13N4O2S (M + H)+, 325.0759; found, 325.0744.
Anal. (C16H12N4O2S‚0.3TFA‚0.2CH3CN) C, H, N.
3-(5-Meth ylth ien -2-yl)-5-(ca r ba m oyla m in o)in d en o[1,2-
c]p yr a zol-4-on e (24g). mp >280 °C. 1H NMR (300 MHz,
DMSO-d6): δ 13.65 (brs, 1 H), 9.31 (brs, 1 H), 8.24 (d, J ) 8.1
Hz, 1 H), 7.79 (brs, 1 H), 7.40 (m, 1 H), 7.05-7.00 (m, 1 H),
7.00-6.92 (m, 1 H), 6.85-6.75 (m, 2 H) 2.53 (s, 3 H). HRMS
(ESI) calcd for C16H13N4O2S (M + H)+, 325.0759; found,
325.0767. Anal. (C16H12N4O2S‚0.2TFA) C, H, N.
3-(5-Ca r boeth oxyth ien -2-yl)-5-(ca r ba m oyla m in o)in d e-
n o[1,2-c]p yr a zol-4-on e (24h ). mp >280 °C. 1H NMR (300
MHz, DMSO-d6): δ 9.27 (brs, 1 H), 8.18 (d, J ) 8.1 Hz, 1 H),
7.85 (brs, 2 H), 7.33 (dd, J ) 8.1, 7.0 Hz, 1 H), 6.96 (d, J ) 7.5
Hz, 1 H), 6.75 (brs, 2 H), 4.29 (q, J ) 7.3 Hz, 2 H), 1.28 (t, J
) 6.8 Hz, 3 H). HRMS (ESI) calcd for C18H15N4O4S (M + H)+,
383.0814; found, 383.0788. Anal. (C18H14N4O4S‚0.5TFA) C, H,
N.
3-(Th ien -3-yl)-5-(ca r ba m oyla m in o)in d en o[1,2-c]p yr a -
3-(P yr id -3-yl)-5-(a ce t a m id o)in d e n o[1,2-c]p yr a zol-4-
on e (13r ). mp >300 °C. 1H NMR (300 MHz, DMSO-d6): δ
10.09 (s, 1 H), 9.34 (d, J ) 1.8 Hz, 1 H), 8.66 (d, J ) 4.7, 1.4
Hz, 1 H), 8.46 (ddd, J ) 8.1, 8.1, 1.8 Hz, 1 H), 8.19 (d, J ) 8.4
Hz, 1 H), 7.59 (dd, J ) 8.0, 4.7 Hz, 1 H), 7.49 (dd, J ) 8.5, 7.3
Hz, 1 H), 7.43 (d, J ) 8.1 Hz, 0.5 H) 7.20 (d, J ) 6.9 Hz, 1 H),
7.07 (d, J ) 8.0 Hz, 0.5 H), 2.16 (s, 3 H). HRMS (CI) calcd for
1
zol-4-on e (24i). mp >280 °C. H NMR (300 MHz, DMSO-d6):
δ 13.65 (brs, 1 H), 9.29 (brs, 1 H), 8.32 (s, 1 H), 8.21 (d, J )
8.8 Hz, 1 H), 7.83 (d, J ) 4.8 Hz, 1 H), 7.77-7.75 (m, 1 H),
7.37 (dd, J ) 8.1, 7.7 Hz, 1 H), 7.03 (d, J ) 7.3 Hz, 1 H), 6.76
(brs, 2 H). HRMS (ESI) calcd for C15H11N4O2S (M + H)+,
311.0603; found, 311.0610. Anal. (C15H10N4O2S‚0.5TFA‚0.01-
DMF) C, H, N.
C
17H13N4O2 (M + H)+, 305.1039; found, 305.1048. Anal.
3-(5-Ch lor oth ien -3-yl)-5-(ca r ba m oyla m in o)in d en o[1,2-
c]p yr a zol-4-on e (24j). mp >300 °C. 1H NMR (400 MHz,
DMSO-d6, 30 °C): δ 13.70 (brs, 1 H), 9.29 (brs, 1 H), 8.25 (d,
J ) 8.4 Hz, 1 H), 8.19 (d, J ) 1.5 Hz, 1 H), 7.85 (d, J ) 1.7 Hz,
1 H), 7.41 (dd, J ) 8.5, 8.5 Hz, 1 H), 7.07 (d, J ) 6.8 Hz, 1 H),
6.75 (brs, 2 H). HRMS (ESI) calcd for C15H10N4O2SCl (M +
H)+, 345.0213; found, 345.0209.
(C17H12N4O2) C, H, N.
3-(P yr id -4-yl)-5-(a ce t a m id o)in d e n o[1,2-c]p yr a zol-4-
on e (13s). mp >300 °C. 1H NMR (300 MHz, DMSO-d6): δ
10.08 (s, 1 H), 8.76 (dd, J ) 4.6, 1.7 Hz, 2 H), 8.18 (d, J ) 8.5
Hz, 1 H), 8.06 (dd, J ) 4.6, 1.7 Hz, 2 H), 7.49 (dd, J ) 8.4, 7.3
Hz, 1 H), 7.19 (d, J ) 7.3 Hz, 1 H), 2.16 (s, 3 H). HRMS (CI)
calcd for C17H13N4O2 (M + H)+, 305.1039; found, 305.1046.
Anal. (C17H12N4O2) C, H, N.
3-(Th ien -2-yl)-5-(a cet a m id o)in d en o[1,2-c]p yr a zol-4-
on e (13t). mp >300 °C. 1H NMR (300 MHz, DMSO-d6): δ
10.13 (s, 1 H), 8.22 (d, J ) 8.7 Hz, 1 H), 7.98 (d, J ) 3.1 Hz,
1 H), 7.83 (d, J ) 4.8 Hz, 1 H), 7.49 (dd, J ) 8.1, 8.0 Hz, 1 H),
7.27 (m, 1 H), 7.20 (d, J ) 7.0 Hz, 1 H), 2.07 (s, 3 H). HRMS
(CI) calcd for C16H12N3O2S (M + H)+, 310.0650; found, 310.0641.
Anal. (C16H11N3O2S) C, H, N.
3-(2,5-Dim eth ylth ien -3-yl)-5-(ca r ba m oyla m in o)in d en o-
[1,2-c]p yr a zol-4-on e (24k ). mp >280 °C. 1H NMR (300 MHz,
DMSO-d6): δ 13.80 (brs, 1 H), 9.26 (s, 1 H), 8.20 (d, J ) 8.4
Hz, 1 H), 7.35 (dd, J ) 8.4, 7.3 Hz, 1 H), 7.11 (brs, 1 H), 7.00
(d, J ) 7.3 Hz, 1 H), 6.74 (brs, 2 H), 2.52 (s, 3 H), 2.38 (s, 3 H).
HRMS (ESI) calcd for C17H15N4O2S (M + H)+, 339.0916; found,
339.0905. Anal. (C17H15N4O2S‚0.5TFA) C, H, N.
3-(F u r a n -2-yl)-5-(ca r ba m oyla m in o)in d en o[1,2-c]p yr a -
zol-4-on e (24l). mp 278 °C (dec). 1H NMR (300 MHz, DMSO-
d6): δ 13.29 (brs, 1 H), 9.24 (brs, 1 H), 8.21 (d, J ) 8.8 Hz, 1
H), 7.94 (brs, 1 H), 7.37 (dd, J ) 8.4, 7.3 Hz, 1 H), 7.21 (d, J
) 3.3 Hz, 1 H), 7.02 (d, J ) 6.9 Hz, 1 H), 6.74 (brs, 2 H). HRMS
(ESI) calcd for C15H11N4O3 (M + H)+, 295.0831; found, 295.0838.
Anal. (C15H10N4O3‚1.0TFA) C, H, N.
3-(1-Met h ylp yr r ol-3-yl)-5-(ca r b a m oyla m in o)in d en o-
[1,2-c]p yr a zol-4-on e (24m ). mp >300 °C. 1H NMR (300 MHz,
DMSO-d6): δ 13.20 (brs, 1 H), 9.38 (brs, 1 H), 8.21 (d, J ) 8.8
Hz, 1 H), 7.61 (dd, J ) 1.9, 1.8 Hz, 1 H), 7.36 (dd, J ) 8.8, 8.8
Hz, 1 H), 7.02 (d, J ) 7.3 Hz, 1 H), 6.88 (dd, J ) 2.6, 2.2 Hz,
1 H), 6.75 (brs, 2 H), 6.71 (dd, J ) 2.6, 1.8 Hz, 1 H), 3.71 (s, 3
H). HRMS (ESI) calcd for C16H14N5O2 (M + H)+, 308.1148;
found, 308.1166.
3-(3-Meth ylth ien -2-yl)-5-(a ceta m id o)in d en o[1,2-c]p yr a -
1
zol-4-on e (13u ). mp 275 °C. H NMR (300 MHz, DMSO-d6):
δ 13.45 (brs, 1 H), 10.06 (brs, 1 H), 8.19 (d, J ) 8.5 Hz, 1 H),
7.72-7.68 (m, 1 H), 7.47 (dd, J ) 8.4, 7.7 Hz, 1 H), 7.19 (d, J
) 7.0 Hz, 1 H), 7.06 (d, J ) 5.1 Hz, 1 H), 2.36 (s, 3 H), 2.14 (s,
3 H). HRMS (ESI) calcd for C17H14N3O2S (M + H)+, 324.0807;
found, 324.0811.
3-(2,5-Dich lor oth ien -3-yl)-5-(a ceta m id o)in d en o[1,2-c]-
p yr a zol-4-on e (13v). 1H NMR (300 MHz, DMSO-d6): δ 13.68
(brs, 1 H), 10.08 (brs, 1 H), 8.25 (d, J ) 8.7 Hz, 1 H), 7.68 (brs,
1 H), 7.54 (dd, J ) 8.1, 7.6 Hz, 1 H), 7.27 (brs, 1 H), 2.21 (s, 3
H). HRMS (ESI) calcd for C16H8N3O2SCl2 (M - H)+, 375.9714;
found, 375.9698.
3-(F u r a n -2-yl)-5-(a cet a m id o)in d en o[1,2-c]p yr a zol-4-
on e (13w ). mp >300 °C. 1H NMR (300 MHz, DMSO-d6): δ
13.90 (s, 1 H), 10.11 (s, 1 H), 8.22 (d, J ) 8.4 Hz, 1 H), 8.00 (s,
1 H), 7.50 (dd, J ) 8.5, 7.3 Hz, 1 H), 7.29 (s, 1 H), 7.23 (d, J )
6.9 Hz, 1 H), 2.19 (s, 3 H). HRMS (CI) calcd for C16H12N3O3
(M + H)+, 294.0879; found, 294.0878. Anal. (C16H11N3O3‚
0.26TFA‚0.34acetone‚0.45H2O) C, H, N.
Ack n ow led gm en t. We thank Ann Klemm and
Marge M. Stafford for their technical expertise. We
thank Michael Haas, Wayne Daneker, and Karl Blom
for their mass spectroscopic assistance, Gregory Nemeth
and Ernest Schubert for their NMR assistance, and
Gerry Everlof, Peter Crawford, Mei Li, and Wayne
Rankin for their optical spectroscopic assistance. We
also thank the staff at DND-CAT, Advanced Photon
Source, Argonne National Laboratories for their as-
sistance in X-ray crystallographic data collection.
3-(Th ien -2-yl)-5-(ca r ba m oyla m in o)in d en o[1,2-c]p yr a -
1
zol-4-on e (24e). mp 214 °C. H NMR (300 MHz, DMSO-d6):
δ 13.75 (brs, 1 H), 9.28 (brs, 1 H), 8.22 (d, J ) 8.0 Hz, 1 H),
7.93 (dd, J ) 3.7, 1.1 Hz, 1 H), 7.83-7.77 (m, 1 H), 7.37 (dd,
J ) 8.8, 8.8 Hz, 1 H), 7.25-7.20 (m, 1 H), 7.05-7.00 (m, 1 H),
6.77 (brs, 2 H). HRMS (ESI) calcd for C15H11N4O2S (M + H)+,
311.0603; found, 311.0611.

Indenopyrazoles as Inhibitors
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 24 5247
(14) Gray, N. S.; Wodicka, L.; Thunnissen, A.-M. W. H.; Norman, T.
C.; Kwon, S.; Espinoza, F. H.; Morgan, D. O.; Barnes, G.;
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G. Exploiting Chemical Libraries, Structure, and Genomics in
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