Discovery of imidazo[1,5-c]imidazol-3-ones: Weakly basic, orally active factor Xa inhibitors

Article abstract of DOI:10.1021/jm701548u

The coagulation enzyme factor Xa (FXa) has been recognized as a promising target for the development of new antithrombotic agents. We previously found compound 1 to be an orally bioavailable FXa inhibitor in fasted monkeys; however, 1 showed poor bioavail

Full text of DOI:10.1021/jm701548u

3422
J. Med. Chem. 2008, 51, 3422–3436
Discovery of Imidazo[1,5-c]imidazol-3-ones: Weakly Basic, Orally Active Factor Xa Inhibitors
Yasuhiro Imaeda,* Takanobu Kuroita, Hiroki Sakamoto, Tetsuji Kawamoto, Mamoru Tobisu,§ Noriko Konishi, Katsuhiko Hiroe,
Masaki Kawamura, Toshimasa Tanaka, and Keiji Kubo
Pharmaceutical Research DiVision, Takeda Pharmaceutical Co., Ltd., 2-17-85, Jusohonmachi, Yodogawa-ku, Osaka 532-8686, Japan
ReceiVed December 12, 2007
The coagulation enzyme factor Xa (FXa) has been recognized as a promising target for the development of
new antithrombotic agents. We previously found compound 1 to be an orally bioavailable FXa inhibitor in
fasted monkeys; however, 1 showed poor bioavailability in rats and fed monkeys. To work out the
pharmacokinetic problems, we focused our synthetic efforts on the chemical conversion of the 4-(imidazo[1,2-
a]pyridin-5-yl)piperazine moiety of 1 to imidazolylpiperidine derivatives (fused and nonfused), which resulted
in the discovery of the weakly basic imidazo[1,5-c]imidazol-3-one 3q as a potent and selective FXa inhibitor.
Compound 3q showed favorable oral bioavailability in rats and monkeys under both fasted and fed conditions
and antithrombotic efficacy in a rat model of venous thrombosis after oral administration, without a significant
increase in bleeding time (unlike warfarin). On the basis of these promising properties, compound 3q was
selected for further evaluation.
Introduction
the prothrombinase complex with calcium ion and factor Va,
and the complex amplifies the procoagulant action of FXa. FXa
acts at an earlier stage in the coagulation cascade than thrombin.
Inactivation of FXa by specific inhibitors does not influence
preformed thrombin but does effectively prevent the generation
of thrombin. Extensive preclinical and clinical proof-of-principal
data show that inhibition of FXa is effective in both venous
and arterial thrombosis.3 In animal models, a better therapeutic
index (antithrombotic efficacy vs antihemostatic effects, such
as bleeding) has been shown for direct FXa inhibitors compared
with those for direct thrombin inhibitors and warfarin.4 On the
basis of these data, a cleaner side effect profile would be
expected for a FXa inhibitor relative to a thrombin inhibitor in
the clinical setting.3,5
Abnormal intravascular clot formation is an important factor
in a number of cardiovascular diseases such as deep vein
thrombosis, pulmonary embolism, myocardial infarction, un-
stable angina, and ischemic stroke. Agents inhibiting the
coagulation cascade, such as heparins, low-molecular-weight
heparin, fondaparinux, and vitamin K antagonists (warfarin,
acenocoumarol, and fluindione), have been available for the
treatment and prevention of these thromboembolic diseases.
Vitamin K antagonists prevent the synthesis of vitamin K
dependent coagulation factors in the liver and inhibit the
activation of blood coagulation leading to thrombus formation.
As the only three oral anticoagulants used in a clinical setting,
warfarin, acenocoumarol, and fluindione are widely used for
the prophylaxis and treatment of venous thromboembolic
disease. However, warfarin therapy necessitates a routine blood
coagulation test and dose adjustment for each individual because
it has substantial drawbacks such as narrow therapeutic dose
window, slow onset and offset of action, and interaction with
many drugs and food.1 Despite careful medical treatment, it is
difficult to protect patients from the risk of bleeding. Therefore,
there is a clear need to develop improved anticoagulants that
are orally active, have long duration of action, and have useful
in vivo potency without unpredictable or serious bleeding
complications.
A variety of direct and low-molecular-weight FXa inhibitors
have been described,6 and orally active FXa inhibitors have been
extensively explored.5d–h For example, some nonamidine FXa
inhibitors such as rivaroxaban7 and apixaban8 have been reported
to be orally active FXa inhibitors and are under clinical study.
As part of our FXa inhibitor program, we have previously
reported that 4-(imidazo[1,2-a]pyridin-5-yl)piperazine 19 sig-
nificantly inhibited human FXa with an IC50 of 0.0097 µM, and
its in vitro anticoagulant activity PT2 (the concentration of
compound required to double the clotting time in a human
prothrombin time (PT) assay) was 1.4 µM. Compound 1
demonstrated good selectivity for FXa versus other serine
proteinases and favorable oral bioavailability (BA) in fasted
monkeys (BA ) 30.5%). Unfortunately, compound 1 showed
poor oral bioavailability in fed monkeys, fasted rats, and fed
rats. We hoped that the pharmacokinetic behavior might be
improved by the modification of the 4-(imidazo[1,2-a]pyridin-
5-yl)piperazine moiety of compound 1 as the S4 binding
element.6c,8,10
Factor Xa (FXaa) is a key serine protease in the coagulation
cascade and essential in the formation of a thrombin, a key
mediator of both fibrin formation and platelet activation. It plays
an important role in the coagulation network at the common
pathway that connects both the tissue factor-activated extrinsic
pathway and the surface-activated intrinsic pathway.2 FXa forms
* To whom correspondence should be addressed. Phone: +81-6-6300-
6458. Fax: +81-6-6300-6306. E-mail: Imaeda_Yasuhiro@takeda.co.jp.
Our binding model of compound 1 in FXa9 demonstrated that
the 2-hydroxymethylimidazo[1,2-a]pyridine moiety formed no
salt bridge or hydrogen bond with any amino acid residue in
the S4 site, but it did form hydrophobic contacts with the
aromatic rings of Tyr99, Phe174, and Trp215 in “the S4 aryl
binding site”.11 In particular, the imidazo[1,2-a]pyridine ring
extends across the face of Phe174 phenyl ring with the favorable
π-π stacking interaction. This result suggests that the 4-(imi-
§
Current address: Osaka University, 2-1 Yamadaoka, Suita-shi, Osaka
565-0871, Japan.
a Abbreviations: FXa, factor Xa; PT, prothrombin time; PT2, the
concentration of compound required to double the clotting time in a human
prothrombin time assay; BA, bioavailability; WSC, 1-[3-(dimethylamino)-
propyl]-3-ethylcarbodiimide hydrochloride; HOBt, 1-hydroxybenzotriazole
hydrate; TFA, trifluoroacetic acid; CDI, 1,1′-carbonyldiimidazole; DBU,
1,8-diazabicyclo[5.4.0]-7-undecene; PDB, Protein Data Bank; CYP, cyto-
chrome P450; t-PA, tissue plasminogen activator.
10.1021/jm701548u CCC: $40.75
2008 American Chemical Society
Published on Web 05/29/2008

DiscoVery of Imidazo[1,5-c]imidazol-3-ones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12 3423
Table 1. In Vitro Activities of Imidazo[1,2-a]pyridines 1 and 2
nitrogen atom in the imidazole rings of 3b and 3c, subsequent
formation of the quaternary imidazolium salt using iodomethane,
and finally removal of the trityl group under acidic conditions.16
4-(Imidazol-1-yl)piperidines 3f-i were prepared by the
method outlined in Scheme 3. The displacement reaction of
mesylate 11 with imidazoles 12a and 12b gave piperidines 13a
and 13b. Removal of the Boc group from piperidines 13a-d17
followed by coupling with carboxylic acid 8 afforded amides
3f-i.
The synthesis of 4-(imidazol-2-yl)piperidines 3j and 3k is
depicted in Scheme 4. Removal of the Boc group from
piperidines 1417 and 15, prepared by methylation of 14, followed
by coupling with carboxylic acid 8 gave the desired amides 3j
and 3k.
Alkylene-linked imidazoles 3l-o were synthesized as de-
scribed in Scheme 5. Intermediates 17a-d were prepared from
2-methylimidazole by alkylation with 4-(bromoalkyl)piperidine
16a-d18 in the presence of sodium hydride. Removal of the
Boc group from 17a-c followed by condensation with car-
boxylic acid 8 and treatment with 4 M HCl in EtOAc gave the
desired amides 3l-n. Removal of the Boc group from 17d
followed by coupling with carboxylic acid 8 gave amide 3o.
The synthesis of alkylene-linked imidazole 3p was shown in
Scheme 6. Alkylation of 2-methylimidazole with 1819 in the
presence of sodium hydride gave amide 19. Treatment with
trifluoroacetic acid (TFA) and subsequent reduction of the amide
bond with lithium aluminum hydride afforded piperazine 20,
which was condensed with carboxylic acid 8 to give the desired
amide 3p.
Imidazo[1,5-c]imidazol-3-ones 3q and 3r were synthesized
as follows (Scheme 7). Reductive amination of imidazolecar-
baldehydes 22a and 22b with 4-aminopiperidine 21 gave
aminomethylimidazole 23a and 23b. Cyclization of 23a or 23b
using 1,1′-carbonyldiimidazole (CDI) and 1,8-diazabicyclo[5.4.0]-
7-undecene (DBU) afforded imidazo[1,5-c]imidazol-3-one 24a
or 24b. Removal of the Boc group from 24a and 24b with
concentrated HCl and successive condensation with 8 provided
the corresponding amides 3q and 3r.
The synthesis of imidazo[1,5-c]imidazol-3-one 3s, a pipera-
zine analogue of 3q, is depicted in Scheme 8. Hydrazone 26,
prepared from hydrazine 2518 and 22a, was reduced with
borane-tetrahydrofuran complex and subsequently treated with
hydrochloric acid for decomposition of aminopiperazine-borane
complex to give hydrazine 27. Cyclization of 27 with CDI
afforded imidazo[1,5-c]imidazol-3-one 28, which was converted
into piperazine 29 by palladium-catalyzed debenzylation with
ammonium formate. Coupling of carboxylic acid 8 with
piperazine 29 gave the desired amide 3s.
a Inhibitory activity against human FXa. IC50 values shown are the mean
of duplicate measurements. The 95% confidence intervals are shown in
parentheses.
dazo[1,2-a]pyridin-5-yl)piperazine moiety might not be essential
for inhibitory activity, but the hydrophobic contacts at the S4
aryl binding site might be important, so we hypothesized that
the imidazo[1,2-a]pyridine moiety of 1 as the S4 binding
element could be replaced with other basic heteroaromatic rings
to improve the pharmacokinetic properties without loss of
inhibitory activity. Thus, we initially attempted to replace the
piperazine ring with a piperidine ring, shift the substitution
position of piperidine ring on the imidazo[1,2-a]pyridine ring,
and remove the 2-hydroxymethyl group to modulate the
physiochemical profiles. This initial effort resulted in the
discovery of 4-(imidazo[1,2-a]pyridin-3-yl)piperidine 2 as an
interesting new lead (Table 1). The quite similar FXa inhibitory
activities between 1 and 2 (IC50 ) 0.0097 µM for 1 vs 0.012
µM for 2) opened up the possibility to explore imidazolylpip-
eridine derivatives (fused and nonfused). There are no reports
on FXa inhibitors bearing an imidazolylpiperidine moiety.
Therefore, we were prompted to design and synthesize new
imidazole and bicyclic imidazole derivatives 3 (Figure 1) in an
attempt to further improve the pharmacokinetic properties.
In this paper, we describe the synthesis and structure-activity
relationships for 3 and show that weakly basic imidazo[1,5-
c]imidazol-3-one 3q is a potent and selective direct FXa inhibitor
with excellent in vivo activity and favorable oral bioavailability
unaffected by food intake, thus making it a promising drug
candidate for further evaluation.12
Chemistry
The synthesis of imidazo[1,2-a]pyridine 2 and tetrahydroimi-
dazo[1,2-a]pyridine 3a is shown in Scheme 1. The key intermediate
piperidine 7a was prepared by condensation of bromoacetaldehyde
Results and Discussion
4
13 and 2-aminopyridine 5 followed by removal of the Boc group.
The compounds thus synthesized were evaluated in vitro for
inhibitory potency against human FXa, expressed as IC50 values,
and for their activity in the prolongation of human PT, expressed
as PT2. To estimate the oral bioavailability of these compounds,
the ex vivo PT prolonging activity was also determined 1 h
after oral administration to mice at a dose of 30 mg/kg and
expressed as ratios of the PT of the compound in treatment mice
to that of the untreated group.
The activities of 4-(imidazol-5(4)-yl)piperidine derivatives are
shown in Table 2. Tetrahydroimidazo[1,2-a]pyridine 3a exhib-
ited slightly less potent FXa inhibitory activity (IC50 ) 0.023
µM for 3a vs 0.012 µM for 2) compared to 2, while 3a showed
more potent in vitro anticoagulant activity compared to 2 (PT2
) 0.48 µM for 3a vs 2.2 µM for 2). This result suggests that
Catalytic hydrogenation of imidazo[1,2-a]pyridine 6 using PtO2
followed by deprotection of the Boc group gave tetrahydroimi-
dazo[1,2-a]pyridine 7b. Piperidines 7a and 7b were coupled with
3-(6-chloronaphthalen-2-yl)sulfonylpropionic acid 814 using
1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride
(WSC) and 1-hydroxybenzotriazole hydrate (HOBt) and suc-
cessively treated with concentrated HCl to give the desired
amides 2 and 3a.
4-(Imidazol-5(4)-yl)piperidines 3b-e were synthesized as
shown in Scheme 2. Piperidines 9a and 9b15,16 were coupled
with carboxylic acid 8 to give the desired amides 3b and 3c.
Regioselective N-methylation to obtain 1-methylimidazoles 3d
and 3e was carried out by N-tritylation on the less hindered

3424 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12
Imaeda et al.
Figure 1
Scheme 1a
a Reagents and conditions: (a) EtOH, reflux, 60%; (b) concd HCl, 91%; (c) (1) H2, PtO2, (2) concd HCl, 95%; (d) (1) WSC, HOBt, DBU, Et3N, (2) concd
HCl, 81-82%.
Scheme 2a
a Reagents and conditions: (a) 8, WSC, HOBt, DBU, Et3N, 50-91%; (b) TrCl, Et3N, 60-100%; (c) (1) MeI, (2) AcOH, 48-76%.
Scheme 3a
Scheme 4a
a Reagents and conditions: (a) MeI, NaH, 100%; (b) (1) concd HCl, (2)
8, WSC, HOBt, DBU, Et3N, 54-70%.
a Reagents and conditions: (a) K2CO3 or NaH, 3-11%; (b) (1) concd
HCl, (2) 8, WSC, HOBt, DBU, Et3N, 66-98%.
human FXa inhibitory potency and in vitro anticoagulant
activity. On the other hand, compounds 3b and 3c (R1 ) H)
are slightly less potent than 3d and 3e. The basicity and the
lipophilicity of 3c (pKa ) 7.9, log D ) 2.0 at pH 7.4) are similar
to those of 3d and 3e (i.e., pKa ) 8.2, log D ) 2.0 at pH 7.4
for 3e). This result suggests that the existence of a free hydrogen
atom on either nitrogen atom in the imidazole rings of 3b and
3c might interrupt the interaction between the S4 site of FXa
the improvement in anticoagulant potency of 3a may be
attributed to lower lipophilicity (log D ) 2.2 for 3a, 2.5 for 2
at pH 7.4)20 and increased basicity (pKa ) 8.2 for 3a, 5.9 for
2).21 Thus, we speculated that the lipophilicity and basicity of
the S4 binding element influenced the in vitro anticoagulant
activities of these compounds.22 The 1-methylimidazoles 3d and
3e are equipotent to tetrahydroimidazo[1,2-a]pyridine 3a in

DiscoVery of Imidazo[1,5-c]imidazol-3-ones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12 3425
Scheme 5a
was found to be available to accommodate structural modifica-
tions and the imidazole rings appeared able to shift into the
deeper S4 aryl binding site as shown in Figure 2 (yellow arrow).
Thus, we designed our next set of compounds to incorporate
varied alkylene chains between the imidazole ring and the
piperidine ring, hoping to enhance the interaction with this site.
The in vitro anti-FXa and PT prolonging activities obtained
for the compounds thus synthesized, of type 3i, are listed in
Table 4. Ethylene-linked imidazole 3m (FXa IC50 ) 0.037 µM)
was found to be more potent than the other alkylene-linked
imidazoles 3l, 3n, 3o, and original compound 3i. The in vitro
anticoagulant activity of 3m was equipotent to that of 3i (PT2
) 0.91 for 3m vs 0.99 µM for 3i). The in vitro activity of
piperazine 3p, which has an ethylene linker, was approximately
equal to that of piperidine 3m. A molecular modeling study of
3i, 3m, and 3p was carried out using the same model described
above (Figure 3). In this model, the imidazole ring of 3i (orange)
and the imidazolylethyl groups of 3m (white) and 3p (green)
are located at the S4 aryl binding site of FXa (yellow circle).
In this site, the imidazolylethyl groups of 3m and 3p appeared
to occupy the deeper S4 aryl binding site more than the
imidazole ring of 3i. Because 3m and 3p were found to be more
potent FXa inhibitors than 3i, we speculated that the imida-
zolylethyl group in 3m and 3p might contribute to the
enhancement of interaction with the S4 site of FXa compared
to the imidazolyl group of 3i.
a Reagents and conditions: (a) NaH, 7-100%; (b) (1) concd HCl, (2) 8,
WSC, HOBt, DBU, Et3N, (3) 4 M HCl/EtOAc, 53-74%; (c) (1) concd
HCl, (2) 8, WSC, HOBt, DBU, Et3N, 15%.
Scheme 6a
Compound 3m was considered to have poor oral bioavail-
ability in mice, as discussed in more detail below, which might
be ascribed to the flexible ethylene linker. We expected that
lowering the flexibility of the ethylene linker by cyclization with
the imidazole ring might improve oral bioavailability because
of a decrease in the number of rotatable bonds.26 Moreover,
we attempted to lower the basicity of 3m by introduction of an
electron-withdrawing carbonyl group onto the imidazole nitro-
gen atom as a component for cyclization because the basicity
of compound 1 (pKa ) 5.6), which showed favorable oral
bioavailability in mice and fasted monkeys, was lower than for
compounds that had poor oral bioavailability in mice (pKa >
7.1 for 3a, 3d, 3e, and 3m), as discussed in more detail below.27
Hence, we designed and synthesized the weakly basic bicyclic
imidazo[1,5-c]imidazol-3-one moiety as a novel S4 binding
element by introduction of an electron-withdrawing carbonyl
group on the imidazole nitrogen atom as a component for
cyclization. The imidazo[1,5-c]imidazol-3-one derivatives have
not been reported previously; however, the chemical synthesis
is simple and robust. The results of imidazo[1,5-c]imidazol-3-
ones 3q-s (pKa ) 5.0-5.4) are shown in Table 5. Interestingly,
compounds 3q and 3r showed more potent FXa inhibitory
activity than imidazolylethylpiperidine 3m, with IC50 values in
the single-digit nanomolar range (FXa IC50 ) 0.0048-0.0053
µM for 3q and 3r vs 0.037 µM for 3m). Piperazine 3s showed
potent FXa inhibitory activity (IC50 ) 0.0069 µM) comparable
to that of the corresponding piperidine 3q. Surprisingly, in vitro
anticoagulant activities of 3q-s were similar to that of 3m (PT2
) 0.61-1.2 µM for 3q-s vs 0.91 µM for 3m) despite the
unambiguous difference in their FXa inhibitory activities. The
reason for these results could not be explained in terms of
lipophilicity (log D ) 2.1-2.5 for 3q-s, 2.2 for 3m at pH 7.4);
however, it might be attributed to the difference in basicity (pKa
) 5.0-5.4 for 3q-s, 7.9 for 3m) caused by the existence of a
carbonyl group on the imidazo[1,5-c]imidazol-3-one ring.
a Reagents and conditions: (a) 2-methylimidazole, NaH, 68%; (b) (1)
TFA, (2) LiAlH4, 40%; (c) 8, WSC, HOBt, Et3N, 2%.
and the imidazole ring. As a result, the 1-methylimidazole
moiety of 3d and 3e was found to be a new effective S4 binding
element.
To investigate the influence of the substitution position of
the 4-piperidine moiety on the imidazole ring, some 4-(imidazol-
1-yl)piperidines (3f-i) and 4-(imidazol-2-yl)piperidines (3j and
3k) were examined and the results are shown in Table 3. Among
them, the 2-methyl (3i) and 2-ethyl (3f) derivatives showed the
most potent anticoagulant activities. The FXa inhibitory activity
of 4-(2-methylimidazol-1-yl)piperidine 3i was slightly weaker
than that of the regioisomer 4-(1-methylimidazol-5-yl)piperidine
3d. Compound 3j, lacking an N-substituent, exhibited notably
less potent in vitro activity. In comparison to 3i, 4-(1-methy-
limidazol-2-yl)piperidine 3k inhibited FXa slightly more po-
tently, whereas its anticoagulant activity was slightly less potent.
The FXa inhibitory activity, basicity, and lipophilicity of 3k
(IC50 ) 0.056 µM, pKa ) 6.9, log D ) 2.1 at pH 7.4) are similar
to those of 3d (IC50 ) 0.056 µM, pKa ) 7.2, log D ) 2.0), but
3k was nevertheless less potent with respect to in vitro
anticoagulant activity than 3d (PT2 ) 2.1 for 3k vs 0.49 µM
for 3d). These data suggested that the position of the imidazole
ring nitrogen might be more important for in vitro anticoagulant
activity than basicity or lipophilicity for these inhibitors.
4-(Imidazol-5-yl)piperidines 3d and 3e were found to be the
most potent FXa inhibitors in our 4-(imidazolyl)piperidine series.
In our effort to achieve further enhancement of FXa potency,
a molecular modeling study of 2 and 3a was carried out using
the program GOLD23 and the X-ray structure of FXa reported
by Sanofi-Aventis for an inhibitor complex (PDB code 1EZQ)24
(Figure 2). In this model, the imidazo[1,2-a]pyridine ring of 2
(orange) and the tetrahydroimidazo[1,2-a]pyridine ring of 3a
(white) are located at the S4 aryl binding site of FXa.25 In this
site, an open space around the imidazole rings (yellow circle)
The ex vivo PT prolongation activity after oral administration
in mice and cytochrome P450 (CYP) 3A428 inhibitory activities
of the more potent compounds described above are shown in

3426 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12
Imaeda et al.
Scheme 7a
a Reagents and conditions: (a) NaBH(OAc)3, AcOH, 100%; (b) CDI, DBU, 77-97%; (c) (1) concd HCl; (2) 8, WSC, HOBt, Et3N, 50-73%.
Scheme 8a
a Reagents and conditions: (a) 22a, 88% (b) (1) BH3 ·THF, (2) 6 M HCl, 100%; (c) CDI, DBU, 65%; (d) HCO2NH4, 10% Pd/C, 93%; (e) 8, WSC, HOBt,
Et3N, 76%.
Table 2. In Vitro Activities of 4-(Imidazol-5(4)-yl)piperidines 3a-e
human FXa,
human PT,
b
d
compd
R1
R2
IC50 (µM)a
PT2 (µM)
log Dc
pKa
2
0.012 (0.0099-0.014)
0.023 (0.022-0.024)
0.17 (0.15-0.20)
2.2
0.48
3.0
2.5
0.49
0.56
2.5
2.2
2.0
2.0
2.0
2.0
5.9
8.2
6.9
7.9
7.2
8.2
3a
3b
3c
3d
3e
- (CH2)4 -
H
H
Me
Me
H
Me
H
0.12 (0.10-0.14)
0.056 (0.052-0.059)
0.031 (0.028-0.034)
Me
a Inhibitory activity against human FXa. IC50 values shown are the mean of duplicate measurements. The 95% confidence intervals are shown in parentheses.
b PT (prothrombin time) is defined as the concentration of compound required to double the time to clot formation in the PT assay. PT2 values shown are
the mean of duplicate measurements. c Measured at pH 7.4. d Measured using a titration method.
Table 6. Tetrahydroimidazo[1,2-a]pyridine 3a, imidazolylpip-
eridine derivatives 3d, 3e, and 3i, and imidazolylethyl deriva-
tives 3m and 3p exhibited low ex vivo mouse PT ratios, and so
these compounds were judged to have poor oral bioavailability
in mice. On the other hand, imidazo[1,5-c]imidazol-3-one
derivatives 3q-s showed significant mouse PT prolonging
activities and thus appeared to have favorable oral bioavailability
in mice. These results support our assumption that lowering the
flexibility and basicity of the S4 binding element could
dramatically improve oral bioavailability. 5-Methylimidazo[1,5-
c]imidazol-3-ones 3q and 3s showed considerably weak CYP3A4
inhibitory activity at 10 µM, whereas 5,7-dimethylimidazo[1,5-
c]imidazol-3-one 3r inhibited CYP3A4 significantly. On the
basis of their ex vivo PT prolongation activities in mice and
their weak CYP3A4 inhibitory potencies, compounds 3q and
3s were selected for evaluation in cynomolgus monkeys.
In order to estimate the oral bioavailability in fasted and fed
monkeys, the ex vivo PT prolonging activities in cynomolgus
monkeys were determined for 3q and 3s. After a 3 mg/kg oral
dose, both compounds prolonged PT significantly with long
duration of action under fasted conditions (Figure 4A). Under
fed conditions, compound 3q showed as potent a PT prolonging
activity as under fasted conditions, whereas compound 3s
showed prolonged PT to a lesser extent following oral dosing
(Figure 4B).
Since many trypsin-like proteases have essential physiological
functions,29 an important consideration for the development of
FXa inhibitors is the ability of the molecule to bind selectively.
Compound 3q displayed >3000-fold selectivity for FXa versus
several coagulation (thrombin), fibrinolytic (plasmin, tissue
plasminogen activator (t-PA)), and digestive enzymes (trypsin)
(Table 7).

DiscoVery of Imidazo[1,5-c]imidazol-3-ones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12 3427
Table 3. In Vitro Activities of 4-(Imidazol-1-yl)piperidines 3f-i and
4-(Imidazol-2-yl)piperidines 3j and 3k
compd type
R
human FXa, IC50 (µM)a human PT, PT2 (µM)b
3d
3f
3g
3h
3i
3j
3k
0.056 (0.052-0.059)
0.16 (0.14-0.17)
0.23 (0.19-0.28)
1.4 (1.3-1.6)
0.17 (0.16-0.18)
1.6 (1.4-1.8)
0.056 (0.053-0.060)
0.49
1.0
2.0
A
A
A
A
B
B
Et
Pr
H
Me
H
7.0
0.99
>10
2.1
Me
a Inhibitory activity against human FXa. IC50 values shown are the mean
of duplicate measurements. The 95% confidence intervals are shown in
parentheses. b PT (prothrombin time) is defined as the concentration of
compound required to double the time to clot formation in the PT assay.
PT2 values shown are the mean of duplicate measurements.
Figure 3. Superimposed binding model of compounds 3i (orange),
3m (white), and 3p (green) in FXa.
has favorable oral bioavailability in rats and monkeys without
significant influence from food intake. Furthermore, 3q showed
considerably weak inhibition of CYP isoforms at 10 µM
(CYP2C8, 35% inhibition;30 CYP2C9, 0% inhibition;30 CYP2D6,
5% inhibition31).
In order to evaluate the efficacy of compound 3q as an oral
anticoagulant agent, it was tested in a rat model of venous
thrombosis (Figure 5). Compound 3q at doses of 1, 3, and 10
mg/kg po reduced thrombus formation by 32%, 53%, and 86%,
respectively. On the other hand, warfarin, an orally active
antithrombotic agent, reduced thrombus formation by 82% at 1
mg/kg po; however, its antithrombotic effect was reduced
remarkably at 0.3 mg/kg po.
The bleeding times for compound 3q and warfarin were
determined in rats and are shown in Table 10. Compound 3q
at 30 mg/kg po had little effect on the bleeding time, and a
higher dose (100 mg/kg, po) tended to prolong bleeding time
to 2.3 times the control value. Meanwhile, warfarin at 1 mg/kg
po (minimal dose preventing thrombus formation in rat model)
showed significant prolongation of bleeding time to 4.4 times
the control value. These results suggest that compound 3q could
achieve antithrombotic efficacy without a significant increase
in bleeding time in rats, unlike warfarin. On the basis of its
potency, selectivity, pharmacokinetic profile, and in vivo
efficacy, compound 3q was selected for further evaluation.
To predict the binding mode of compound 3q, a molecular
modeling study was carried out using the program GOLD and
the X-ray structure of FXa reported by Sanofi-Aventis for an
inhibitor complex (PDB code 1EZQ)24 (Figure 6). In the model,
the imidazo[1,5-c]imidazol-3-one moiety forms no salt bridge
with the carboxyl group of Asp189 at the bottom of the S1
pocket, while the 6-chloronaphthyl group is deeply buried inside
the hydrophobic S1 region with the chlorine atom pointing
toward the center of the Tyr228 aromatic ring.7,11b,22b,32
Compound 3q represents a related example in FXa inhibition,
wherein a basic group thought to be essential in forming a
crucial salt bridge with Asp1896d,24 can be eliminated and
replaced by a neutral group capable of providing high affinity
for the enzyme. A hydrogen bonding interaction between the
amide carbonyl group in 3q and the backbone amide nitrogen
of Gly219 is observed. Recently, other researchers have reported
that the important interaction for high affinity to FXa is not an
ionic interaction with Asp189 but hydrogen bonding interactions
with the carbonyl groups in the amide linkages of Gly216 and/
or Gly219.24,32c,33 The imidazo[1,5-c]imidazol-3-one moiety is
Figure 2. Superimposed binding model of compounds 2 (orange) and
3a (white) in FXa.
Table 4. In Vitro Activities of Alkylene-Linked Imidazoles 3l-p
compd
n
X
human FXa, IC50 (µM)a human PT, PT2 (µM)b
3i
3l
3m
3n
3o
3p
0
1
2
3
4
2
CH
CH
CH
CH
CH
N
0.17 (0.16-0.18)
0.91 (0.85-0.98)
0.037 (0.036-0.039)
0.46 (0.42-0.49)
0.30 (0.29-0.32)
0.054 (0.051-0.057)
0.99
7.7
0.91
>10
>10
0.99
a Inhibitory activity against human FXa. IC50 values shown are the mean
of duplicate measurements. The 95% confidence intervals are shown in
parentheses. b PT (prothrombin time) is defined as the concentration of
compound required to double the time to clot formation in the PT assay.
PT2 values shown are the mean of duplicate measurements.
The pharmacokinetic profile of compound 3q was evaluated
after single intravenous and oral administration to male
Sprague-Dawley rats and male cynomolgus monkeys. As
illustrated in Table 8, 3q displayed favorable oral bioavailability
after oral administration at a dose of 3 mg/kg in fed rats (24%)
and monkeys (46%), respectively. As shown in Table 9, the
values of Cmax in fasted animals were higher than those in the
fed state, whereas the values of AUC0-24h in the fasted and fed
states were almost the same. These results indicated that 3q

3428 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12
Table 5. In Vitro Activities of Imidazo[1,5-c]imidazol-3-ones 3q-s
Imaeda et al.
d
compd
R1
R2
X
human FXa, IC50 (µM)a
human PT, PT2 (µM)b
log Dc
pKa
3m
3q
3r
0.037 (0.036-0.039)
0.91
1.0
1.2
2.2
2.3
2.5
2.1
7.9
5.0
5.4
5.0
Me
Me
Me
H
Me
H
CH
CH
N
0.0048 (0.0046-0.0051)
0.0053 (0.0048-0.0059)
0.0069 (0.0061-0.0078)
3s
0.61
a Inhibitory activity against human FXa. IC50 values shown are the mean of duplicate measurements. The 95% confidence intervals are shown in parentheses.
b PT (prothrombin time) is defined as the concentration of compound required to double the time to clot formation in the PT assay. PT2 values shown are
the mean of duplicate measurements. c Measured at pH 7.4. d Measured using a titration method.
Table 6. In Vitro and Ex Vivo Activities of Potent FXa Inhibitors
in vitro
ex vivo,
CYP3A4,
% inhibition at 10 µM
a
compd
human PT PT2 (µM)a
mouse PT PT2 (µM)
mouse PT Ratiob
3a
3d
3e
3i
3m
3p
3q
3r
3s
0.48
0.49
0.56
0.99
0.91
0.99
1.0
3.0
2.7
3.4
3.3
1.7
2.1
2.0
2.5
0.9
1.1
1.2
1.2
1.1
1.1
1.1
1.7
2.0
2.8
42
82
25
34
38
53
23
62
38
1.2
0.61
a PT (prothrombin time) is defined as the concentration of compound required to double the time to clot formation in the PT assay. PT2 values shown are
the mean of duplicate measurements. b Ex vivo mouse PT prolonging activity was determined 1 h after oral administration to mice at a dose of 30 mg/kg
and expressed as ratios of the PT of the compound in treatment mice with that of the untreated group (n ) 3).
Figure 4. Ex vivo PT prolonging activities of compounds 3q and 3s in cynomolgus monkeys (n ) 3) at 3 mg/kg, po, under fasted conditions (A)
and fed conditions (B).
located deeply in the hydrophobic S4 aryl binding site. The S4
pocket is a narrow hydrophobic channel defined by the aromatic
rings of Tyr99, Phe174, and Trp215, and hydrophobic interac-
Table 8. Pharmacokinetic Parameters for 3q in Fed Rats and Monkeysa
iv (0.3 mg/kg)
rat
monkey
tion with the three aromatic rings in the S4 site has been reported
for many potent FXa inhibitors.24,32c,33 The imidazo[1,5-
c]imidazol-3-one is sandwiched between the aromatic rings of
Tyr99 and that of Phe174 and forms π-π interactions with the
aromatic rings of Tyr99 and Phe174 and a CH-π interaction
with the indole ring of Trp215. The carbonyl group of the
C5min (ng/mL)
AUC0-24h (ng·h/mL)
MRT (h)
Vd(ss) (mL/kg)
CLtotal (mL/h/kg)
291 ( 45
245 ( 38
612 ( 98
565 ( 94
1.0 ( 0.2
556 ( 38
541 ( 90
0.7 ( 0.1
888 ( 149
1245 ( 189
po (3 mg/kg)
rat
monkey
Table 7. Selectivity Profile for Compound 3q
Cmax (ng/mL)
Tmax (h)
AUC0-24h (ng·h/mL)
MRT (h)
145 ( 26
1.2 ( 0.8
597 ( 159
3.6 ( 0.6
24 ( 8
269 ( 79
2.3 ( 1.5
2592 ( 580
7.0 ( 0.6
46 ( 6
serine protease
FXa
Ki (µM)a 0.0020 ( 0.0010 6.2 ( 0.4
a Data are expressed as the mean ( SEM of three determinations.
thrombin
trypsin plasmin t-PA
>60 >60 >60
BA (%)
a Data are expressed as the mean ( SD of three determinations.

DiscoVery of Imidazo[1,5-c]imidazol-3-ones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12 3429
Table 9. Pharmacokinetic Parameters for 3q after Oral Administration
6.83 (1H, t, J ) 6.4 Hz), 7.16 (1H, t, J ) 8.0 Hz), 7.41 (1H, s),
7.62 (1H, d, J ) 9.2 Hz), 7.97 (1H, d, J ) 6.8 Hz).
at a Dose of 3 mg/kg in the Fed and Fasted Statesa
3-(Piperidin-4-yl)imidazo[1,2-a]pyridine Dihydrochloride
(7a). A solution of 6 (9.04 g, 30.0 mmol) in concentrated HCl (49
mL) was stirred at room temperature for 10 min. The reaction
mixture was diluted with EtOH and concentrated in vacuo. The
precipitated powder was collected by filtration and washed with
EtOH and Et2O to give 7a (7.92 g, 91%) as a colorless powder. 1H
NMR (200 MHz, D2O) δ: 1.94-2.22 (2H, m), 2.39-2.59 (2H, m),
3.23-3.48 (2H, m), 3.52-3.78 (3H, m), 7.48-7.61 (1H, m), 7.86
(1H, s), 7.98-8.04 (2H, m), 8.71 (1H, d, J ) 7.0 Hz).
3-(Piperidin-4-yl)-5,6,7,8-tetrahydroimidazo[1,2-a]pyridine Di-
hydrochloride (7b). A mixture of 6 (1.51 g, 5.01 mmol) and
platinum dioxide (114 mg) in acetic acid (15 mL) was hydrogenated
at atmospheric pressure and room temperature for 16 h. The reaction
mixture was filtered through a Celite pad, and the filtrate was
concentrated in vacuo. The residue was diluted with water, basified
with 8 M NaOH to pH 12, and extracted with CHCl3. The extract
was washed with brine, dried over anhydrous MgSO4, and
concentrated in vacuo to give tert-butyl 4-(5,6,7,8-tetrahydroimi-
dazo[1,2-a]pyridin-3-yl)piperidine-1-carboxylate (1.53 g, quantita-
tive) as a pale-yellow amorphous powder.
fed state
rat
monkey
Cmax (ng/mL)
Tmax (h)
MRT (h)
145 ( 26
1.2 ( 0.8
3.6 ( 0.6
597 ( 159
269 ( 79
2.3 ( 1.5
7.0 ( 0.6
2592 ( 580
AUC0-24h (ng·h/mL)
fasted state
rat
monkey
Cmax (ng/mL)
Tmax (h)
MRT (h)
570 ( 90
0.3 ( 0.0
1.5 ( 0.4
417 ( 10
1204 ( 451
0.8 ( 0.3
1.6 ( 0.2
2122 ( 1030
AUC0-24h (ng·h/mL)
a Data are expressed as the mean ( SD of three determinations.
imidazo[1,5-c]imidazol-3-one does not appear to interact directly
with FXa but rather effects a planarization of the imidazo[1,5-
c]imidazol-3-one ring and brings it into a perpendicular ar-
rangement with respect to the piperidine ring. The sulfone
oxygen is involved in a hydrogen bond with the Gln192 main
chain nitrogen.6d
A mixture of tert-butyl 4-(5,6,7,8-tetrahydroimidazo[1,2-a]py-
ridin-3-yl)piperidine-1-carboxylate (1.53 g, 5.01 mmol) and con-
centrated HCl (8.20 mL) was stirred at room temperature for 10
min. The reaction mixture was diluted with 2-propanol and
concentrated in vacuo. The precipitate was collected by filtration
and washed with 2-propanol and Et2O to give 7b (1.32 g, 95%) as
a colorless powder. 1H NMR (200 MHz, D2O) δ: 1.74-2.19 (6H,
m), 2.20-2.37 (2H, m), 2.95-3.31 (5H, m), 3.50-3.66 (2H, m),
4.11 (2H, t, J ) 5.4 Hz), 7.20 (1H, s).
Conclusion
In order to address the pharmacokinetic liabilities of our FXa
inhibitor 1, chemical modification of its 4-(imidazo[1,2-a]py-
ridin-5-yl)piperazine moiety to other basic substituents led to
the discovery of potent FXa inhibitor 4-(imidazo[1,2-a]pyridin-
3-yl)piperidine 2 as an interesting new lead. Further modification
focused on shifting the imidazole ring into the deeper S4 site
and lowering the flexibility and basicity resulting in the
discovery of the imidazo[1,5-c]imidazol-3-ones 3q, 3r, and 3s
as highly potent FXa inhibitors. As a result of further investiga-
tion, 3q showed favorable oral bioavailability in rats and
monkeys without significant influence from food intake, good
enzyme selectivity, and weak inhibition of CYP3A4. Oral
administration of 3q prevented thrombus formation dose-
dependently in rats without a significant increase in bleeding
time, unlike warfarin. On the basis of its excellent in vitro and
in vivo efficacy and a favorable pharmacokinetic profile,
compound 3q was selected for further evaluation.
3-(1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}piperidin-
4-yl)imidazo[1,2-a]pyridine Hydrochloride (2). To a solution of
8 (0.74 g, 2.48 mmol) and HOBt (0.57 g, 3.72 mmol) in acetonitrile
(15 mL) was added WSC (0.72 g, 3.75 mmol), and the mixture
was stirred at room temperature for 15 min. A solution of 7a (0.82
g, 2.99 mmol), DBU (0.90 mL, 6.02 mmol), and Et3N (1.05 mL,
7.50 mmol) in acetonitrile (5 mL) was added, and the resulting
mixture was stirred at room temperature for 3 h. The reaction
mixture was concentrated in vacuo, and the residue was diluted
with water and extracted with CHCl3. The extract was washed with
brine, dried over anhydrous MgSO4, and concentrated in vacuo.
The residue was purified by silica gel chromatography (EtOAc/
EtOH ) 10/1). To a solution of this product in EtOH (5 mL) was
added concentrated HCl (0.38 mL, 4.60 mmol), and the resulting
mixture was concentrated in vacuo. The residue was diluted with
EtOH and Et2O, and the precipitate was collected by filtration to
Experimental Section
1
Melting points were determined with a Yanagimoto melting point
apparatus or a Bu¨chi melting point apparatus B-545 and are
uncorrected. H NMR spectra were obtained at 300 or 200 MHz
give 2 (1.14 g, 81%) as a colorless powder, mp 224-225 °C. H
NMR (300 MHz, DMSO-d6) δ: 1.28-1.39 (1H, m), 1.39-1.68
(1H, m), 1.93-2.11 (2H, m), 2.62-2.84 (3H, m), 3.12-3.28 (1H,
m), 3.32-3.57 (1H, m), 3.61-3.72 (2H, m), 3.88-4.01 (1H, m),
4.31-4.45 (1H, m), 7.54 (1H, t, J ) 6.6 Hz), 7.73 (1H, dd, J )
7.4 and 6.0 Hz), 7.93-8.04 (4H, m), 8.19 (1H, d, J ) 8.7 Hz),
8.26-8.31 (2H, m), 8.68 (1H, s), 9.02 (1H, d, J ) 6.9 Hz). Anal.
(C25H24ClN3O3S·HCl·H2O) C, H, N.
1
on a Varian Ultra-300 or a Varian Gemini-200 spectrometer.
Chemical shifts are given in δ values (ppm) using tetramethylsilane
as the internal standard. Reactions were followed by TLC on silica
gel 60 F 254 precoated TLC plates (E. Merck) or NH TLC plates
(Fuji Silysia Chemical Ltd.). Chromatographic separations were
carried out on silica gel 60 (0.063-0.200 or 0.040-0.063 mm, E.
Merck) or basic silica gel (ChromatorexNH, 100-200 mesh, Fuji
Silysia Chemical Ltd.) using the indicated eluents. Yields are
unoptimized. Chemical intermediates were characterized by 1H
NMR.
tert-Butyl 4-(Imidazo[1,2-a]pyridin-3-yl)piperidine-1-carboxy-
late (6). A solution of 4 (13.7 g, 44.7 mmol) and 5 (2.63 g, 27.9
mmol) in EtOH (100 mL) was refluxed for 3 h. The reaction mixture
was concentrated in vacuo, and the residue was partitioned between
EtOAc and water. The aqueous layer was separated, basified with
8 M NaOH to pH 9, and extracted with EtOAc. The extract was
washed with brine, dried over anhydrous Na2SO4, and concentrated
in vacuo. The residue was purified by silica gel chromatography
(EtOAc/EtOH ) 10/1) to give 6 (8.11 g, 60%) as a pale-yellow
solid. 1H NMR (200 MHz, CDCl3) δ: 1.49 (9H, s), 1.59-1.85 (2H,
m), 1.98-2.17 (2H, m), 2.83-3.10 (3H, m), 4.18-4.38 (2H, m),
3-(1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}piperidin-
4-yl)-5,6,7,8-tetrahydroimidazo[1,2-a]pyridine Hydrochloride
(3a). Compound 3a was prepared in a manner similar to that
described for 2 in 82% yield as a colorless powder, mp 213-214
°C. 1H NMR (200 MHz, DMSO-d6) δ: 1.12-1.58 (2H, m),
1.76-2.08 (6H, m), 2.48-2.68 (2H, m), 2.75 (2H, t, J ) 7.4 Hz),
2.84-3.19 (3H, m), 3.64 (2H, t, J ) 7.0 Hz), 3.34-3.99 (1H, m),
4.00-4.12 (2H, m), 4.26-4.40 (1H, m), 7.34 (1H, s), 7.74 (1H,
dd, J ) 8.8 and 2.2 Hz), 8.01 (1H, dd, J ) 8.8 and 1.8 Hz),
8.17-8.32 (3H, m), 8.67 (1H, s). Anal. (C25H28ClN3O3S·HCl ·H2O)
C, H, N.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(1H-
imidazol-4-yl)piperidine (3b). To a mixture of 8 (0.60 g, 2.00
mmol) and HOBt (0.46 g, 3.00 mmol) in acetonitrile (15 mL) was
added WSC (0.58 g, 3.00 mmol), and the mixture was stirred at
room temperature for 15 min. A solution of 9a (0.46 g, 2.00 mmol),

3430 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12
Imaeda et al.
Figure 5. Antithrombotic effect of 3q and warfarin in a rat venous thrombosis model. Thrombus formation was stimulated by the combination of
endothelial damage and blood stagnation. Data are expressed as the mean ( SEM: (/) p ) 0.025 vs vehicle (a one-tailed Williams test), n ) 5-6.
Table 10. Effect of 3q and Warfarin on Bleeding Time in Ratsa
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(2-meth-
yl-1H-imidazol-4-yl)piperidine (3c). Compound 3c was prepared
dose (mg/kg, po)
bleeding time (s)
in a manner similar to that described for 3b in 91% yield as a
vehicle
3q
460 ( 60
490 ( 90
1040 ( 296
1
colorless powder, mp 190-192 °C. H NMR (200 MHz, CDCl3)
30
100
δ: 1.41-1.60 (2H, m), 1.93-2.10 (2H, m), 2.39 (3H, s), 2.59-2.71
(2H, m), 2.83-2.91 (2H, m), 3.07-3.21 (1H, m), 3.53-3.60 (2H,
m), 3.83-3.90 (1H, m), 4.47-4.54 (1H, m), 6.56 (1H, s), 7.58
(1H, dd, J ) 9.0 and 2.0 Hz), 7.88-7.97 (4H, m), 8.48 (1H, br s).
Anal. (C22H24ClN3O3S·0.2H2O) C, H, N.
vehicle
warfarin
360 ( 35
445 ( 87
0.3
1
1590 ( 158b
a Data are expressed as the mean ( SEM. b p e 0.025 vs vehicle (a
one-tailed Shirley-Williams’ test). n ) 6.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(1-tri-
tyl-1H-imidazol-4-yl)piperidine (10a). To a solution of 3b (0.35
g, 0.81 mmol) and Et3N (0.10 g, 1.00 mmol) in DMF (10 mL) was
added trityl chloride (0.25 g, 0.90 mmol) at 0 °C, and the resulting
mixture was stirred at 0 °C for 1 h and then at room temperature
for 15 h. The reaction mixture was concentrated in vacuo, and the
residue was diluted with saturated aqueous NaHCO3 solution and
extracted with EtOAc. The extract was washed with brine, dried
over anhydrous Na2SO4, and concentrated in vacuo. The residue
was purified by basic silica gel chromatography (EtOAc) to give
10a (0.54 g, quantitative) as a colorless amorphous powder. 1H
NMR (200 MHz, CDCl3) δ: 1.40-1.50 (2H, m), 1.92-2.10 (2H,
m), 2.58-2.87 (4H, m), 3.04-3.17 (1H, m), 3.51-3.59 (2H, m),
3.78-3.84 (1H, m), 4.41-4.47 (1H, m), 6.49 (1H, s), 7.09-7.35
(16H, m), 7.57 (1H, dd, J ) 8.8 and 1.8 Hz), 7.92-8.02 (4H, m),
8.46 (1H, s).
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(2-meth-
yl-1-trityl-1H-imidazol-4-yl)piperidine (10b). Compound 10b was
prepared in a manner similar to that described for 10a in 60% yield
as a colorless amorphous powder. 1H NMR (300 MHz, CDCl3) δ:
1.33-1.59 (2H, m), 1.90-2.06 (2H, m), 2.57-2.74 (2H, m),
2.82-2.89 (2H, m), 3.05-3.13 (1H, m), 3.52-3.57 (2H, m),
3.80-3.84 (1H, m), 4.45-4.49 (1H, m), 6.34 (1H, s), 7.09-7.12
(5H, m), 7.30-7.35 (10H, m), 7.58 (1H, dd, J ) 8.7 and 1.8 Hz),
7.91-7.96 (4H, m), 8.47 (1H, s).
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(1-meth-
yl-1H-imidazol-5-yl)piperidine (3d). A solution of 10a (0.54 g,
0.80 mmol) and iodomethane (0.10 mL, 1.61 mmol) in DMF (5
mL) was stirred at room temperature for 15 h. The reaction mixture
was concentrated in vacuo, and the residue was dissolved in acetic
acid (5 mL), water (5 mL), and MeOH (2 mL). After being stirred
at 95 °C for 2 h, the reaction mixture was concentrated in vacuo.
The residue was basified with saturated aqueous NaHCO3 solution
and extracted with EtOAc. The extract was washed with saturated
aqueous NaHCO3 solution, dried over anhydrous Na2SO4, and
concentrated in vacuo. The residue was purified by basic silica gel
chromatography (EtOAc to 10/1 EtOAc/MeOH) to give 3d (0.27
Figure 6. Binding model of imidazo[1,5-c]imidazol-3-one 3q in FXa.
DBU (0.61 g, 4.00 mmol), and Et3N (0.61 g, 6.00 mmol) in
acetonitrile (15 mL) was added to the mixture, and the resulting
mixture was stirred at room temperature for 15 h. The reaction
mixture was concentrated in vacuo, and the residue was diluted
with aqueous K2CO3 solution and extracted with EtOAc. The extract
was washed with brine, dried over anhydrous Na2SO4, and
concentrated in vacuo. The residue was purified by basic silica gel
chromatography (EtOAc) to give 3b (0.43 g, 50%) as a colorless
amorphous powder. 1H NMR (300 MHz, CDCl3 + CD3OD) δ:
1.40-1.60 (2H, m), 1.95-2.11 (2H, m), 2.66-2.91 (5H, m),
3.13-3.20 (1H, m), 3.54-3.61 (2H, m), 3.86-3.90 (1H, m),
4.47-4.51 (1H, m), 6.70 (1H, s), 7.53 (1H, d, J ) 1.2 Hz), 7.59
(1H, dd, J ) 8.4 and 2.4 Hz), 7.93-7.98 (4H, m), 8.48 (1H, s).
Anal. (C21H22ClN3O3 ·0.5H2O) C, H, N.

DiscoVery of Imidazo[1,5-c]imidazol-3-ones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12 3431
1
g, 76%) as a colorless amorphous powder. H NMR (200 MHz,
CDCl3) δ: 1.43-1.70 (2H, m), 1.88-2.04 (2H, m), 2.60-2.78 (2H,
m), 2.87-2.95 (2H, m), 3.09-3.22 (1H, m), 3.53-3.61 (2H, m),
3.60 (3H, s), 3.91-3.98 (1H, m), 4.55-4.62 (1H, m), 6.76 (1H,
s), 7.38 (1H, s), 7.59 (1H, dd, J ) 8.8 and 1.8 Hz), 7.93-7.97
(4H, m), 8.48 (1H, s). Anal. (C22H24ClN3O3S·0.5H2O) C, H, N.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(1,2-di-
methyl-1H-imidazol-5-yl)piperidine (3e). Compound 3e was
prepared in a manner similar to that described for 3d in 48% yield
as a colorless amorphous powder. 1H NMR (200 MHz, CDCl3) δ:
1.40-1.56 (2H, m), 1.87-2.00 (2H, m), 2.36 (3H, s), 2.60-2.71
(2H, m), 2.87-2.95 (2H, m), 3.09-3.21 (1H, m), 3.45 (3H, s),
3.53-3.61 (2H, m), 3.90-3.96 (1H, m), 4.54-4.60 (1H, m), 6.60
(1H, s), 7.60 (1H, dd, J ) 8.8 and 1.8 Hz), 7.93-7.97 (4H, m),
8.49 (1H, s). Anal. (C23H26ClN3O3S·H2O) C, H, N.
m), 3.14-3.26 (1H, m), 3.54-3.63 (2H, m), 4.01-4.14 (2H, m),
4.71-4.78 (1H, m), 6.80 (1H, d, J ) 1.4 Hz), 6.97 (1H, d, J ) 1.0
Hz), 7.60 (1H, dd, J ) 8.8 and 1.8 Hz), 7.89-7.98 (4H, m), 8.49
(1H, s). Anal. (C24H28ClN3O3S·0.5H2O) C, H, N.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(2-meth-
yl-1H-imidazol-1-yl)piperidine (3i). Yield 83%, colorless amor-
1
phous powder. H NMR (300 MHz, CDCl3) δ: 1.68-1.84 (2H,
m), 1.97-2.09 (2H, m), 2.42 (3H, s), 2.61-2.69 (1H, m), 2.91-2.98
(2H, m), 3.15-3.24 (1H, m), 3.54-3.62 (2H, m), 4.02-4.13 (2H,
m), 4.73-4.77 (1H, m), 6.81 (1H, d, J ) 1.5 Hz), 6.94 (1H, d, J
) 1.5 Hz), 7.60 (1H, dd, J ) 8.9 and 2.0 Hz), 7.91-7.97 (4H, m),
8.50 (1H, s). Anal. (C22H24ClN3O3S·H2O) C, H, N.
tert-Butyl 4-(1-Methyl-1H-imidazol-2-yl)piperidine-1-carbox-
ylate (15). To a solution of 14 (0.25 g, 1.00 mmol) in DMF (3
mL) was added NaH (60% in oil, 40 mg, 1.00 mmol) at 0 °C, and
the mixture was stirred at 0 °C for 30 min. Iodomethane (0.07 mL,
1.12 mmol) was added to the mixture, and then the resulting mixture
was stirred at 0 °C for 2 h. The reaction mixture was concentrated
in vacuo, and the residue was diluted with aqueous NaHCO3
solution and extracted with EtOAc. The extract was dried over
anhydrous Na2SO4 and concentrated in vacuo. The residue was
purified by silica gel chromatography (CHCl3/MeOH ) 10/1) to
tert-Butyl 4-(2-Ethyl-1H-imidazol-1-yl)piperidine-1-carboxy-
late (13a). A mixture of 11 (10.0 g, 35.8 mmol), 12a (4.13 g, 43.0
mmol), and K2CO3 (4.95 g, 35.8 mmol) in DMF (80 mL) was stirred
at 100 °C for 72 h. The reaction mixture was concentrated in vacuo,
and the residue was diluted with aqueous NaHCO3 solution and
extracted with EtOAc. The extract was washed with saturated
aqueous NaHCO3 solution, dried over anhydrous Na2SO4, and
concentrated in vacuo. The residue was purified by basic silica gel
chromatography (EtOAc to 5/1 EtOAc/MeOH) to give 13a (1.10
g, 11%) as a yellow oil. 1H NMR (300 MHz, CDCl3) δ: 1.31-1.39
(3H, m), 1.49 (9H, s), 1.70-1.95 (4H, m), 2.67-2.87 (4H, m),
3.96-4.01 (1H, m), 4.29-4.33 (2H, m), 6.86 (1H, d, J ) 1.5 Hz),
6.97 (1H, d, J ) 1.5 Hz).
tert-Butyl 4-(2-Propyl-1H-imidazol-1-yl)piperidine-1-carboxy-
late (13b). Compound 13b was prepared in a manner similar to
that described for 13a in 3% yield as a colorless amorphous powder.
1H NMR (300 MHz, CDCl3) δ: 0.98-1.04 (3H, m), 1.49 (9H, s),
1.71-1.94 (6H, m), 2.59-2.69 (2H, m), 2.79-2.87 (2H, m),
3.96-4.04 (1H, m), 4.28-4.33 (2H, m), 6.85 (1H, d, J ) 1.2 Hz),
6.97 (1H, d, J ) 1.5 Hz).
1
give 15 (0.27 g, quantitative) as a pale-yellow oil. H NMR (200
MHz, CDCl3) δ: 1.46 (9H, s), 1.80-1.91 (4H, m), 2.70-2.96 (3H,
m), 3.61 (3H, s), 4.18-4.25 (2H, m), 6.79 (1H, d, J ) 1.0 Hz),
6.94 (1H, d, J ) 1.0 Hz).
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(1H-
imidazol-2-yl)piperidine (3j). Compound 3j was prepared in a
manner similar to that described for 3h in 54% yield as a colorless
amorphous powder. 1H NMR (300 MHz, CDCl3) δ: 1.60-1.85 (2H,
m), 1.98-2.14 (2H, m), 2.72-3.04 (4H, m), 3.15-3.24 (1H, m),
3.49-3.61 (2H, m), 3.90-3.95 (1H, m), 4.46-4.50 (1H, m), 6.96
(1H, br s), 7.00 (1H, br s), 7.58 (1H, dd, J ) 2.1 and 9.0 Hz),
7.90-7.97 (4H, m), 8.48 (1H, d, J ) 0.9 Hz), 8.88 (1H, br s).
Anal. (C21H22ClN3O3S) C, H, N.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(1H-
imidazol-1-yl)piperidine (3h). A mixture of 13c (0.28 g, 1.11
mmol) in concentrated HCl (4 mL) was stirred at room temperature
for 3 h. The reaction mixture was concentrated in vacuo, and the
residue was azeotroped with EtOH. The residue, DBU (0.33 mL,
2.23 mmol), and Et3N (0.47 mL, 3.36 mmol) were dissolved in
acetonitrile (15 mL). The solution was added to a mixture of 8
(0.33 g, 1.10 mmol), HOBt (0.26 g, 1.70 mmol), and WSC (0.32
g, 1.70 mmol) in acetonitrile (15 mL), and the resulting mixture
was stirred at room temperature for 15 h. The reaction mixture
was concentrated in vacuo, and the residue was diluted with aqueous
K2CO3 solution and extracted with EtOAc. The extract was dried
over anhydrous Na2SO4 and concentrated in vacuo. The residue
was purified by basic silica gel chromatography (EtOAc) to give
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(1-meth-
yl-1H-imidazol-2-yl)piperidine (3k). Compound 3k was prepared
in a manner similar to that described for 3h in 70% yield as a
colorless amorphous powder. 1H NMR (200 MHz, CDCl3) δ:
1.60-2.00 (4H, m), 2.70-2.94 (4H, m), 3.11-3.26 (1H, m),
3.52-3.61 (2H, m), 3.61 (3H, s), 3.93-4.00 (1H, m), 4.45-4.52
(1H, m), 6.79 (1H, d, J ) 1.2 Hz), 6.92 (1H, d, J ) 1.4 Hz), 7.58
(1H, dd, J ) 2.0 and 8.8 Hz), 7.89-7.98 (4H, m), 8.48 (1H, s),
8.88 (1H, br s). Anal. (C22H24ClN3O3S·0.5H2O) C, H, N.
tert-Butyl 4-[(2-Methyl-1H-imidazol-1-yl)methyl]piperidine-
1-carboxylate (17a). To a solution of 16a (8.50 g, 30.6 mmol)
and 2-methylimidazole (1.80 g, 21.9 mmol) in DMF (100 mL) was
added NaH (60% in oil, 0.87 g, 21.8 mmol) at room temperature.
After the mixture was stirred at 80 °C for 12 h, the reaction mixture
was concentrated in vacuo. The residue was diluted with CHCl3,
washed with aqueous K2CO3 solution, dried over anhydrous
Na2SO4, and concentrated in vacuo. The residue was purified by
silica gel chromatography (EtOAc to 5/1 EtOAc/MeOH) to give
1
3h (0.40 g, 83%) as a yellow gum. H NMR (300 MHz, CDCl3)
δ: 1.70-1.92 (2H, m), 2.09-2.22 (2H, m), 2.64-2.72 (1H, m),
2.90-2.97 (2H, m), 3.17-3.25 (1H, m), 3.54-3.61 (2H, m),
4.00-4.09 (1H, m), 4.08-4.21 (1H, m), 4.69-4.73 (1H, m), 6.93
(1H, t, J ) 1.2 Hz), 7.08 (1H, d, J ) 1.2 Hz), 7.54 (1H, s), 7.60
(1H, dd, J ) 8.7 and 2.1 Hz), 7.93-7.97 (4H, m), 8.49 (1H, d, J
) 1.2 Hz). Anal. (C21H22ClN3O3S·0.5H2O) C, H, N.
The following compounds 3f, 3g, and 3i were prepared in a
manner similar to that described for 3h.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(2-eth-
1
17a (0.43 g, 7%) as a brown oil. H NMR (300 MHz, CDCl3) δ:
1.13-1.26 (2H, m), 1.46 (9H, s), 1.68 (1H, m), 2.37 (3H, s), 2.67
(2H, t, J ) 11.4 Hz), 3.86 (2H, t, J ) 7.5 Hz), 4.11 (2H, br s),
6.80 (1H, d, J ) 1.5 Hz), 6.91 (1H, d, J ) 1.5 Hz).
The following compounds 17b-d were prepared in a manner
similar to that described for 17a.
yl-1H-imidazol-1-yl)piperidine (3f). Yield 98%, colorless amor-
phous powder. H NMR (300 MHz, CDCl3) δ: 1.36 (3H, t, J )
tert-Butyl 4-[2-(2-Methyl-1H-imidazol-1-yl)ethyl]piperidine-
1-carboxylate (17b). Yield 80%, brown oil. 1H NMR (300 MHz,
CDCl3) δ: 1.13-1.28 (2H, m), 1.46 (9H, s), 1.57-1.63 (4H, m),
1.82 (1H, m), 2.37 (3H, s), 2.65 (2H, t, J ) 12.9 Hz), 3.71 (2H, d,
J ) 7.5 Hz), 4.16 (2H, br s), 6.78 (1H, d, J ) 1.5 Hz), 6.91 (1H,
d, J ) 1.5 Hz).
tert-Butyl 4-[3-(2-Methyl-1H-imidazol-1-yl)propyl]piperidine-
1-carboxylate (17c). Yield quantitative, brown oil. 1H NMR (300
MHz, CDCl3) δ: 1.03-1.35 (4H, m), 1.45 (9H, s), 1.59-1.82 (5H,
m), 2.37 (3H, s), 2.66 (2H, t, J ) 12.4 Hz), 3.81 (2H, t, J ) 7.0
Hz), 4.06 (2H, br s), 6.80 (1H, d, J ) 1.4 Hz), 6.90 (1H, d, J ) 1.4
Hz).
1
7.4 Hz), 1.65-1.86 (2H, m), 1.96-2.05 (2H, m), 2.61-2.68 (1H,
m), 2.71 (2H, q, J ) 7.5 Hz), 2.91-2.98 (2H, m), 3.16-3.24 (1H,
m), 3.54-3.62 (2H, m), 4.01-4.13 (2H, m), 4.73-4.77 (1H, m),
6.81 (1H, d, J ) 1.8 Hz), 6.98 (1H, d, J ) 1.5 Hz), 7.60 (1H, dd,
J ) 8.7 and 1.8 Hz), 7.94-7.97 (4H, m), 8.49 (1H, d, J ) 0.6 Hz).
Anal. (C23H26ClN3O3S·0.5H2O) C, H, N.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-(2-pro-
pyl-1H-imidazol-1-yl)piperidine (3g). Yield 66%, colorless amor-
1
phous powder. H NMR (200 MHz, CDCl3) δ: 1.02 (3H, t, J )
7.5 Hz), 1.68-2.05 (6H, m), 2.62-2.69 (3H, m), 2.90-2.99 (2H,

3432 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12
Imaeda et al.
tert-Butyl 4-[4-(2-Methyl-1H-imidazol-1-yl)butyl]piperidine-
1-carboxylate (17d). Yield 47%, brown oil. 1H NMR (300 MHz,
CDCl3) δ: 1.03-1.35 (4H, m), 1.45 (9H, s), 1.59-1.82 (7H, m),
2.37 (3H, s), 2.66 (2H, t, J ) 12.4 Hz), 3.81 (2H, t, J ) 7.0 Hz),
4.06 (2H, br s), 6.80 (1H, d, J ) 1.4 Hz), 6.90 (1H, d, J ) 1.4 Hz).
(3H, s), 3.44-3.50 (6H, m), 3.60-3.71 (2H, m), 4.67 (2H, s), 6.80
(1H, d, J ) 1.6 Hz), 6.94 (1H, d, J ) 1.6 Hz).
1-[2-(2-Methyl-1H-imidazole-1-yl)ethyl]piperazine (20). 19
(2.80 g, 9.08 mmol) was dissolved in TFA (15 mL), and the
resulting mixture was stirred at room temperature for 1 h. The
reaction mixture was concentrated in vacuo, diluted with saturated
aqueous NaHCO3 solution, and extracted with CHCl3. The extract
was dried over anhydrous Na2SO4 and concentrated in vacuo. The
residue was dissolved in THF (30 mL), and lithium aluminum
hydride (0.46 g, 12.1 mmol) was added to the solution at 0 °C.
After being stirred at 60 °C for 1 h, the reaction mixture was cooled
to 0 °C. The reaction mixture was quenched with saturated
ammonium chloride and extracted with EtOAc. The extract was
dried over anhydrous Na2SO4 and concentrated in vacuo to give
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-[(2-
methyl-1H-imidazol-1-yl)methyl]piperidine Hydrochloride (3l).
17a (0.43 g, 1.54 mmol) was dissolved in concentrated HCl (4 mL)
and diluted with EtOH (20 mL). The solution was concentrated in
vacuo, and the residue was dissolved in acetonitrile (20 mL) with
Et3N (0.63 mL, 4.51 mmol) and DBU (0.46 mL, 3.26 mmol). The
solution was added to a mixture of 8 (0.45 g, 1.50 mmol), WSC
(0.43 g, 2.24 mmol), and HOBt (0.35 g, 2.29 mmol) in acetonitrile
(10 mL), and then the resulting mixture was stirred at room
temperature for 12 h. The reaction mixture was concentrated in
vacuo, diluted with saturated aqueous NaHCO3 solution, and
extracted with CHCl3. The extract was dried over anhydrous Na2SO4
and concentrated in vacuo. The residue was purified by silica gel
chromatography (EtOAc), and the product was dissolved in EtOAc.
An amount of 4 M HCl in EtOAc (0.30 mL, 1.20 mmol) was added,
and then the mixture was stirred at room temperature for 5 min.
The resulting mixture was concentrated in vacuo, and the residue
was triturated with Et2O to give 3l (0.55 g, 74%) as a beige
amorphous powder. 1H NMR (200 MHz, DMSO-d6) δ: 1.04-1.25
(2H, m), 1.63-1.73 (2H, m), 1.91 (1H, m), 2.37 (3H, s), 2.52 (1H,
m), 2.82-3.04 (3H, m), 3.51-3.60 (2H, m), 3.70 (2H, d, J ) 7.4
Hz), 3.88 (1H, d, J ) 13.4 Hz), 4.56 (1H, d, J ) 13.4 Hz), 6.76
(1H, d, J ) 1.4 Hz), 6.92 (1H, d, J ) 1.4 Hz), 7.60 (1H, dd, J )
2.0 and 8.8 Hz), 7.87-7.97 (4H, m), 8.48 (1H, s). Anal.
(C23H26ClN3O3S·HCl·H2O·0.25Et2O) C, H, N.
1
20 (0.70 g, 40%) as a colorless oil. H NMR (200 MHz, DMSO-
d6) δ: 2.27 (3H, s), 2.49-2.69 (8H, m), 3.60 (2H, t, J ) 9.1 Hz),
3.89 (2H, t, J ) 9.1 Hz), 6.69 (1H, d, J ) 1.6 Hz), 7.02 (1H, d, J
) 1.6 Hz).
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-[2-(2-
methyl-1H-imidazol-1-yl)ethyl]piperazine (3p). To a solution of
8 (0.90 g, 3.01 mmol), 20 (0.58 g, 3.01 mmol), Et3N (1.26 mL,
9.03 mmol), and HOBt (0.69 g, 4.52 mmol) in CH2Cl2 (30 mL)
was added WSC (0.87 g, 4.52 mmol) at room temperature. After
being stirred at room temperature for 16 h, the reaction mixture
was concentrated in vacuo, diluted with saturated aqueous NaHCO3
solution, and extracted with CHCl3. The extract was dried over
anhydrous Na2SO4 and concentrated in vacuo. The residue was
purified by silica gel chromatography (CHCl3/MeOH ) 40/1 to
10/1) to give 3p (0.021 g, 2%) as a brown amorphous powder. 1H
NMR (200 MHz, CDCl3) δ: 2.39 (3H, s), 2.41-2.46 (4H, m),
2.61-2.67 (2H, m), 2.82-2.89 (2H, m), 3.42-3.60 (6H, m), 3.93
(2H, t, J ) 9.3 Hz), 6.85 (1H, d, J ) 2.1 Hz), 6.90 (1H, d, J ) 2.1
Hz), 7.59 (1H, dd, J ) 2.4 and 13.2 Hz), 7.92-7.94 (4H, m), 8.47
(1H, s). Anal. (C23H27ClN4O3S·H2O) C, H, N.
The following compounds 3m and 3n were prepared in a manner
similar to that described for 3l.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-[2-(2-
methyl-1H-imidazol-1-yl)ethyl]piperidine Hydrochloride (3m).
Yield 69%, beige amorphous powder. 1H NMR (200 MHz, DMSO-
d6) δ: 1.00-1.24 (2H, m), 1.48 (1H, m), 1.63-1.77 (4H, m), 2.37
(3H, s), 2.49 (1H, t, J ) 13.2 Hz), 2.83-2.90 (2H, m), 2.98 (1H,
t, J ) 13.2 Hz), 3.47-3.60 (2H, m), 3.80-3.88 (3H, m), 4.49 (1H,
d, J ) 11.4 Hz), 6.79 (1H, s), 6.91 (1H, s), 7.59 (1H, dd, J ) 1.8
and 9.0 Hz), 7.90-7.96 (4H, m), 8.47 (1H, s). Anal.
(C24H28ClN3O3S·HCl·0.25H2O) C, H, N.
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-[3-(2-
methyl-1H-imidazol-1-yl)propyl]piperidine Hydrochloride (3n).
Yield 53%, beige amorphous powder. 1H NMR (200 MHz, CDCl3)
δ: 1.00-1.31 (4H, m), 1.52 (1H, m), 1.65-1.88 (4H, m), 2.37 (3H,
s), 2.49 (1H, t, J ) 11.1 Hz), 2.83-2.90 (2H, m), 3.00 (1H, t, J )
11.1 Hz), 3.47-3.58 (2H, m), 3.84 (1H, d, J ) 14.1 Hz), 4.02
(2H, t, J ) 6.9 Hz), 4.51 (1H, d, J ) 14.1 Hz), 7.05 (1H, s), 7.05
(1H, s), 7.28 (1H, s), 7.61 (1H, dd, J ) 2.1 and 9.0 Hz), 7.89-7.97
(4H, m), 8.47 (1H, s). Anal. (C25H30ClN3O3S·HCl·H2O) C, H, N.
tert-Butyl4-{[(2-Methyl-1H-imidazol-5-yl)methyl]amino}piperidine-
1-carboxylate (23a). A mixture of 21 (2.00 g, 10.0 mmol), 22a
(1.10 g, 10.0 mmol), acetic acid (0.69 mL, 12.0 mmol), and sodium
triacetoxyborohydride (5.30 g, 25.0 mmol) in MeOH (40 mL) was
stirred at room temperature for 2 h. The reaction mixture was
concentrated in vacuo, and the residue was basified with aqueous
K2CO3 solution. The solution was extracted with EtOAc, and the
extract was dried over anhydrous Na2SO4 and concentrated in vacuo
1
to give 23a (2.94 g, quantitative) as a pale-yellow oil. H NMR
(300 MHz, CDCl3) δ: 1.26 (2H, t, J ) 7.2 Hz), 1.45 (9H, s), 1.86
(2H, d, J ) 12.6 Hz), 2.40 (3H, s), 2.61-2.86 (3H, m), 3.74 (2H,
s), 3.86-4.18 (2H, m), 6.75 (1H, s).
tert-Butyl 4-{[(2,4-Dimethyl-1H-imidazol-5-yl)methyl]am-
ino}piperidine-1-carboxylate (23b). Compound 23b was prepared
in a manner similar to that described for 23a in quantitative yield
1
as a pale-yellow oil. H NMR (300 MHz, CDCl3) δ: 1.28-1.40
(2H, m), 1.45 (9H, s), 1.87 (2H, d, J ) 12.3 Hz), 2.14 (3H, s),
2.33 (3H, s), 2.67-2.80 (3H, m), 3.71 (2H, s), 4.06 (2H, br s),
6.06 (1H, s).
1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}-4-[4-(2-
methyl-1H-imidazol-1-yl)butyl]piperidine (3o). Compound 3o
was prepared in a manner similar to that described for 3h in 15%
tert-Butyl 4-(5-Methyl-3-oxo-1H-imidazo[1,5-c]imidazol-2(3H)-
yl)piperidine-1-carboxylate (24a). To a solution of compound 23a
(2.94 g, 10.0 mmol) in acetonitrile (20 mL) was added DBU (1.50
mL, 10.0 mmol) and CDI (1.62 g, 10.0 mmol), and the mixture
was stirred at room temperature for 15 h. The reaction mixture
was concentrated in vacuo, diluted with aqueous K2CO3 solution,
and extracted with EtOAc. The extract was dried over anhydrous
MgSO4 and concentrated in vacuo. The residue was purified by
silica gel chromatography (EtOAc to 10/1 EtOAc/MeOH) to give
24a (2.45 g, 77%) as a pale-yellow solid. 1H NMR (300 MHz,
CDCl3) δ: 1.47 (9H, s), 1.63-1.73 (2H, m), 1.75-1.98 (2H, m),
2.61 (3H, s), 2.83 (2H, t, J ) 12.7 Hz), 4.10 (1H, tt, J ) 12.1 and
4.1 Hz), 4.28 (4H, s), 6.71 (1H, t, J ) 1.5 Hz).
1
yield as a beige amorphous powder. H NMR (200 MHz, CDCl3)
δ: 0.91-1.08 (2H, m), 1.22-1.43 (5H, m), 1.61-1.74 (4H, m),
2.37 (3H, s), 2.40-2.54 (1H, m), 2.81 -3.02 (3H, m), 3.52-3.60
(2H, m), 3.78-3.85 (3H, m), 4.26 (1H, d, J ) 13.2 Hz), 6.79 (1H,
d, J ) 1.4 Hz), 6.90 (1H, d, J ) 1.4 Hz), 7.58 (1H, dd, J ) 2.0
and 8.8 Hz), 7.92-7.97 (4H, m), 8.47 (1H, s). Anal.
(C26H32ClN3O3S·0.5H2O) C, H, N.
tert-Butyl 4-[(2-Methyl-1H-imidazol-1-yl)acetyl]piperazine-
1-carboxylate (19). To a solution of 2-methylimidazole (1.10 g,
13.3 mmol) in DMF was added NaH (60% in oil, 0.62 g, 15.5
mmol) at 0 °C. After the mixture was stirred at 0 °C for 30 min,
18 (4.09 g, 13.3 mmol) was added to the mixture, and then the
resulting mixture was stirred at 50 °C for 1 h. The reaction mixture
was concentrated in vacuo, diluted with aqueous K2CO3 solution,
and extracted with EtOAc. The extract was dried over anhydrous
Na2SO4 and concentrated in vacuo to give 19 (2.80 g, 68%) as a
tert-Butyl 4-(5,7-Dimethyl-3-oxo-1H-imidazo[1,5-c]imidazol-
2(3H)-yl)piperidine-1-carboxylate (24b). Compound 24b was
prepared in a manner similar to that described for 24a in 97% yield
as a pale-yellow oil. 1H NMR (200 MHz, CDCl3) δ: 1.47 (9H, s),
1.57-1.72 (2H, m), 1.79-1.84 (2H, m), 2.15 (3H, s), 2.57 (3H,
1
colorless oil. H NMR (200 MHz, CDCl3) δ: 1.48 (9H, s), 2.33

DiscoVery of Imidazo[1,5-c]imidazol-3-ones
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12 3433
s), 2.82 (2H, t, J ) 12.7 Hz), 4.03-4.15 (1H, m), 4.20 (2H, s),
4.29 (1H, br s).
one (3s). Compound 3s was prepared in a manner similar to that
described for 3p in 76% yield as a colorless solid, mp 230-231
°C. 1H NMR (300 MHz, CDCl3) δ: 2.60 (3H, s), 3.13 (2H, t, J )
5.0 Hz), 3.23 (2H, t, J ) 5.0 Hz), 3.54-3.59 (4H, m), 3.62 (2H, t,
J ) 5.0 Hz), 3.69 (2H, t, J ) 5.0 Hz), 4.42 (2H, s), 6.70 (1H, t, J
) 1.5 Hz), 7.59 (1H, dd, J ) 8.6 and 2.3 Hz), 7.89-7.97 (4H, m),
8.47 (1H, s). Anal. (C23H24ClN5O4S) C, H, N.
2-(1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}piperidin-
4-yl)-5-methyl-1,2-dihydro-3H-imidazo[1,5-c]imidazol-3-one (3-
q). 24a (1.18 g, 3.69 mmol) was dissolved in concentrated HCl (4
mL), and the resulting mixture was stirred at room temperature for
30 min. The reaction mixture was concentrated in vacuo. The
residual solid, 8 (1.00 g, 3.35 mmol), HOBt (0.62 g, 4.02 mmol),
and Et3N (1.68 mL, 12.1 mmol) were suspended in DMF (7 mL).
WSC (0.77 g, 4.02 mmol) was added to the suspension, and then
the resulting mixture was stirred at room temperature for 5 h. The
reaction mixture was cooled to 0 °C and diluted with water (14
mL). After the mixture was stirred at 0 °C for 2 h, the precipitate
was collected by filtration and washed with water. The solid was
recrystallized from EtOH-water to give 3q (1.23 g, 73%) as a
colorless crystal, mp 195 °C. 1H NMR (200 MHz, CDCl3) δ:
1.58-1.72 (2H, m), 1.86-1.99 (2H, m), 2.61 (3H, s), 2.63-2.68
(1H, m), 2.81-3.01 (2H, m), 3.14-3.23 (1H, m), 3.46-3.65 (2H,
m), 3.97-4.02 (1H, m), 4.13-4.22 (1H, m), 4.25 (2H, s), 4.69-4.74
(1H, m), 6.70 (1H, d, J ) 3.3 Hz), 7.59 (1H, dd, J ) 8.7 and 2.1
Hz), 7.89-7.96 (4H, m), 8.47 (1H, d, J ) 1.5 Hz). Anal.
(C24H25ClN4O4S) C, H, N.
2-(1-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}piperidin-
4-yl)-5,7-dimethyl-1,2-dihydro-3H-imidazo[1,5-c]imidazol-3-
one (3r). Compound 3r was prepared in a manner similar to that
described for 3q in 50% yield as a colorless solid, mp 196-197
°C. 1H NMR (200 MHz, CDCl3) δ: 1.62-1.79 (2H, m), 1.83-1.99
(2H, m), 2.15 (3H, s), 2.57 (3H, s), 2.64-2.71 (1H, m), 2.86-2.99
(2H, m), 3.31-3.26 (1H, m), 3.49-3.63 (2H, m), 3.97-4.18 (4H,
m), 4.75-4.96 (1H, m), 7.61 (1H, dd, J ) 8.8 and 1.8 Hz),
7.89-7.99 (4H, m), 8.49 (1H, s). Anal. (C25H27ClN4O4S) C, H, N.
4-Benzyl-N-[(2-methyl-1H-imidazol-5-yl)methylene]piperazin-
1-amine (26). A suspension of 25 (10.0 g, 52.3 mmol) and 22a
(5.80 g, 52.7 mmol) in MeOH (200 mL) was refluxed for 3 h. The
reaction mixture was cooled to room temperature and concentrated
in vacuo. The residue was triturated with Et2O to give 26 (13.0 g,
88%) as a pale-yellow solid. 1H NMR (200 MHz, CDCl3) δ: 2.40
(3H, s), 2.62 (4H, t, J ) 5.2 Hz), 3.09 (4H, t, J ) 5.2 Hz), 3.56
(2H, s), 6.97 (1H, s), 7.24-7.34 (5H, m), 7.43 (1H, s), 9.62 (1H,
br s).
In Vitro Assays for the Inhibition of Human FXa. Human
FXa (0.3 U/mL) was obtained from Roche Diagnostics. Chromoge-
nic substrate, S-2765 (Chromogenix-Instrumentation Laboratory)
was used for the measurement of the inhibition of FXa. Anti-FXa
activity was assayed in a buffer containing 50 mM Tris-HCl, 145
mM NaCl, and 2 mM CaCl2 at pH 8.3. Enzyme assay was carried
out in 96-well microtiter plates. Test compounds were diluted in
DMSO. Compound dilutions and 10 µL of enzyme solution were
added to the well containing buffer and preincubated. The enzymatic
reactions were initiated with the addition of 10 µL of 3 mM
substrate, and the mixture was incubated for 10 min at 37 °C. The
reaction was terminated with the addition of 25 µL of 50% acetic
acid. The color development from the release of p-nitroanilide from
each chromogenic substrate was measured at 405 nm on a microtiter
plate reader (Multiscan Ascent, Dainippon Sumitomo Pharmaceuti-
cal Co.). Each absorbance [T] was calculated by subtracting the
absorbance measured without the substrate. The control [C] was
performed using DMSO solution in place of test compound.
Inhibitory effect (%) was calculated according to the equation (1
- [T]/[C]) × 100. IC50 values were calculated by nonlinear
regression analysis of the concentration-response curve. Data for
new compounds were compared to positive control DX-9065a6a
(FXa IC50 ) 0.13 µM). The 95% confidence interval for the IC50
of DX-9065a was 0.12-0.15 µM (duplicate).
In Vitro PT Assays. The assay of plasma clotting time was
performed using an automatic coagulometer (STA Compact,
Diagnostica Stago). PT was measured with STA PT reagents
(Roche Diagnostics). An amount of 1.5 µL of compound
dilutions in DMSO was added to 48.5 µL of human normal
plasma (fresh human plasma, FFP, Sekisui Chemical Co.), and
the mixture was preincubated at 37 °C for 4 min. Coagulation
was initiated with the addition of 100 µL of thromboplastin,
and coagulation time was measured. Coagulation time prolonging
ratio (%) was calculated on the basis of coagulation time when
DMSO was added instead of test compound. The plasma clotting
time doubling concentration (PT2) was calculated from the regres-
sion line based on the method of least squares. Data for new
compounds were compared to positive control DX-9065a6a (PT2
) 0.81 ( 0.029 µM (mean ( SEM, n ) 3))
Measurement of CYP Inhibition Activity. Inhibition activity
of test compounds of CYP3A4 was evaluated by incubating 100
µM testosterone with 10 nM CYP3A4 derived from CYP3A4-
expressing human B-lymphoblastoid cells (BD Biosciences) in the
presence of 10 µM test compound. The incubation mixture was
allowed to stand for 30 min at 37 °C. The concentration of 6ꢀ-
hydroxytestosterone was measured by HPLC system equipped with
a UV detector.
Measurement of ex Vivo PT in Mouse after Oral Adminis-
tration. Male ICR mice (30-35 g, Clea Japan Inc.) fasting for
more than 12 h were employed. Test compounds were orally
administered to these animals. An hour after administration, 800
µL of blood was collected from artery abdominal aorta using 3.8%
sodium citrate (whole blood/sodium citrate solution ) 9:1, v/v),
under anesthesia with pentobarbital (50 mg/kg, ip). The citrated
blood was centrifuged at 1000g for 10 min to obtain platelet poor
plasma (PPP). PT was measured with an automatic coagulometer.
An amount of 50 µL of PPP was preincubated at 37 °C for 4 min.
The PPP was mixed with 100 µL of thromboplastin solution, and
then its coagulation time was measured. Test compounds were
suspended in 0.5% methyl cellulose (Metolose cp100, Shin-Etsu
Chemical), and as control, 0.5% methylcellulose was administered
instead of test compounds. Activity of each test compound was
shown as a ratio (%) determined by comparing the clotting time of
compound-treatment mice with that of control group. We have
4-Benzyl-N-[(2-methyl-1H-imidazol-5-yl)methyl]piperazin-1-
amine (27). To a solution of 26 (13.0 g, 45.9 mmol) in THF (100
mL) was added a solution of borane-THF complex (1 M in THF,
140 mL, 140 mmol) at room temperature. After being stirred at
room temperature for 15 h, the reaction mixture was concentrated
in vacuo. The residue was dissolved in 6 M HCl, and then the
mixture was stirred at 100 °C for 2 h. After cooling to 0 °C, the
mixture was basified to pH 12 with 6 M NaOH and extracted with
CHCl3. The extract was dried over anhydrous Na2SO4 and
concentrated in vacuo to give 27 (14.7 g, quantitative) as a brown
1
oil. H NMR (200 MHz, CDCl3) δ: 2.37 (3H, s), 2.53 (4H, br s),
2.75 (4H, br s), 3.52 (2H, s), 3.90 (2H, s), 6.76 (1H, s), 7.29-7.32
(5H, m).
2-(4-Benzylpiperazin-1-yl)-5-methyl-1,2-dihydro-3H-imida-
zo[1,5-c]imidazol-3-one (28). Compound 28 was prepared in a
manner similar to that described for 24a in 65% yield as a colorless
1
solid. H NMR (200 MHz, CDCl3) δ: 2.60-2.65 (7H, m), 3.14
(4H, t, J ) 4.6 Hz), 3.55 (2H, s), 4.43 (2H, s), 6.70 (1H, t, J ) 2.0
Hz), 7.25-7.34 (5H, m).
5-Methyl-2-(piperazin-1-yl)-1,2-dihydro-3H-imidazo[1,5-c]im-
idazol-3-one (29). A suspension of 28 (8.00 g, 25.7 mmol),
ammonium formate (16.0 g, 254 mmol), and palladium on carbon
(1.60 g) in MeOH (100 mL) was refluxed for 6 h. The mixture
was cooled to room temperature, and the insoluble material was
filtered off through a Celite pad. The filtrate was concentrated in
vacuo, and the residue was triturated with EtOAc and Et2O to give
29 (5.30 g, 93%) as a pale-yellow solid. 1H NMR (300 MHz,
CDCl3) δ: 2.61 (3H, s), 2.98-3.03 (4H, m), 3.10-3.15 (4H, m),
4.44 (2H, s), 6.70 (1H, s).
2-(4-{3-[(6-Chloronaphthalen-2-yl)sulfonyl]propanoyl}pipe-
razin-1-yl)-5-methyl-1,2-dihydro-3H-imidazo[1,5-c]imidazol-3-

3434 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 12
Imaeda et al.
validated the ex vivo PT assay for the evaluation of oral bioavail-
ability in mice by use of the positive control compound DX-9065a.6a
DX-9065a prolonged mouse PT by 1.1-fold at a dose of 30 mg/kg
and by 1.4-fold at a dose of 100 mg/kg.
HCO2NH4 (adjusted to pH 3.0 with HCO2H)/acetonitrile ) 6/4;
flow rate, 0.2 mL/min; column temperature, 40 °C.
Rat Venous Thrombosis Model. Male Sprague-Dawley rats
(7 weeks old) were anesthetized with sodium pentobarbital. A 10
mL length of inferior vena cava from the distal region staring the
left renal vein was isolated, and any side branches were ligated. A
balloon catheter was introduced from the left femoral vein to inferior
vena cava to injure the endothelium. Denudation of the endothelium
was performed by three passes of the inflated balloon catheter. In
order to trigger the thrombus formation at the injured region, a
silk thread was tied around the vena cava caudal to the left renal
vein with a blunt needle (26 gauge), followed by the removal of
the needle. Thirty minutes after the starting of partial stasis, the
thrombus formed in the vena cava was removed and the wet weight
was measured. Blood samples were collected for the measurement
of blood coagulation parameters just after the removal of thrombus.
Compound 3q and warfarin were orally administered 30 min and
18 h, respectively, before inducing the thrombus formation to
examine their antithrombotic potency at the points that give maximal
anticoagulating effects. Compound 3q was given after the animals
had fasted for more than 12 h.
Measurement of Bleeding Time in Rats after Oral Admin-
istration. Male Sprague-Dawley rats (7 weeks old) were anes-
thetized with sodium pentobarbital. The tail was transected at 2-3
mm proximal site from the tip. Blood was blotted every 30 s with
filter paper until either bleeding had stopped or 30 min had elapsed.
Compound 3q and warfarin were orally administered 30 min and
18 h before the measurement of bleeding time, respectively.
Docking Studies. The coordinates of FXa were retrieved from
the Protein Data Bank (accession code 1EZQ).24 Compounds 2,
3a, 3i, 3m, 3p, and 3q were docked into the FXa protein crystal
structure using GOLD23 (version 2.0, the Cambridge Crystal-
lographic Data Centre, U.K.) with the standard default settings. The
initial structures of the compounds-complexed FXa models were
energy-minimized using the MMFF94s force field in MOE (version
2003.02, Chemical Computing Group, Montreal, Canada) to obtain
the final docking models. During the minimization procedure, the
following conditions were adopted. The dielectric constant was set
to 4r, where r is the distance between two interacting atoms. The
residues, which are 8 Å away from each compound, were fixed. A
harmonic force constraint against the initial atomic positions of the
backbone was added, using 10 kcal/Å2 as a force constant. And
atomic charges for the protein and the compounds were set
according to the AMBER99 force field and the AM1-BCC method,
respectively.
Measurement of ex Vivo PT in Monkey after Oral Admin-
istration. Male cynomolgus monkeys (3.5-4 kg, NAFOVANNY)
fasting for more than 12 h were employed. Test compounds were
orally administered to these animals. After administration, 1.5 mL
of blood was collected from the femoral vein using 3.8% sodium
citrate (whole blood/sodium citrate solution ) 9:1, v/v) at several
time points. The citrated blood was centrifuged at 1000g for 10
min to obtain PPP. Plasma clotting time and anticoagulating activity
of drug were determined as described above.
Enzyme Inhibition Assays. Enzyme assays using chromogenic
substrates were performed as follows. Human FXa was obtained
from Roche diagnostics. Human thrombin was obtained from Sigma
Chemical Co. Human trypsin was obtained from Atheus Research
and Technology, Inc. Human plasmin was purchased from BioPur
AG. Human t-PA was obtained from Kyowa Hakko Kogyo Co.
The chromogenic substrates used were S-2222, S-2366, S-2222,
S-2366, S-2302, and S-2288 for FXa, thrombin, trypsin, plasmin,
and t-PA, respectively, and obtained from Chromogenix-Instru-
mentation Laboratory. All enzyme assays were performed at 37
°C in 96-well microtiter plates. The final enzyme concentrations
were 0.024 U/mL, 0.080 U/mL, 0.040 µg/mL, 0.040 U/mL, 0.020
U/mL, and 4000 U/mL for FXa, thrombin, trypsin, plasmin, and
t-PA, respectively. Compound dilutions were added to the wells
containing buffer and enzyme and incubated for 30 min. The
enzyme reactions were initiated by the addition of substrate, and
the color developed from the release of p-nitroanilide from each
chromogenic substrate was monitored continuously for 5 min at
405 nm on a microtiter plate reader. The Ki values were determined
from a Lineweaver-Burk plot, when the optical densities were
measured with different concentrations of substrates.
Pharmacokinetic Analysis in Rats. Test compound was
administered to fasted or fed SD (IGS) rats (male, 8 weeks old,
n ) 3) intravenously (0.3 mg/kg, DMA/PEG400) and orally (3
mg/kg, 0.5% methylcellulose suspension). At 5, 10, 15, and 30
min and 1, 2, 4, 8, 24 h after intravenous administration or at
15 and 30 min and 1, 2, 4, 8, 24 h after oral administration,
blood samples were collected from tail vein. The blood samples
were centrifuged to obtain the plasma fraction. The plasma
samples were deproteinized with acetonitrile. After centrifugation,
the supernatant obtained was diluted with 0.01 mol/L HCO2NH4
(adjusted to pH 3.0 with HCO2H) and centrifuged again. The
compound concentration in the supernatant was measured by LC/
MS/MS with an API3000 triple quadrupole mass spectrometer
(Perkin-Elmer Sciex). The mass spectrometer was equipped with
a turbo ionspray source and operated in positive ion mode. The
HPLC conditions were as follows: column, an L-column ODS (2.1
mm × 150 mm); mobile phase, 0.01 mol/L HCO2NH4 (adjusted to
pH 3.0 with HCO2H)/acetonitrile ) 6/4; flow rate, 0.2 mL/min;
column temperature, 40 °C.
Pharmacokinetic Analysis in Cynomolgus Monkeys. Test
compound was administered to fasted or fed cynomolgus
monkeys (male, n ) 3) intravenously (0.3 mg/kg, DMA/PEG400)
and orally (3 mg/kg, 0.5% methylcellulose suspension). At 5,
10, 15, 30 min and 1, 2, 4, 8, 24 h after intravenous
administration or at 15 and 30 min and 1, 2, 4, 8, 24 h after oral
administration, blood samples were collected from femoral vein.
The blood samples were centrifuged to obtain the plasma
fraction. The plasma samples were deproteinized with acetoni-
trile. After centrifugation, the supernatant obtained was diluted
with 0.01 mol/L HCO2NH4 (adjusted to pH 3.0 with HCO2H) and
centrifuged again. The compound concentration in the supernatant
was measured by LC/MS/MS with an API3000 triple quadrupole
mass spectrometer (Perkin-Elmer Sciex). The mass spectrometer
was equipped with a turbo ionspray source and operated in positive
ion mode. The HPLC conditions were as follows: column, an
L-column ODS (2.1 mm × 150 mm); mobile phase, 0.01 mol/L
Acknowledgment. The authors thank Terufumi Takagi for
his docking model study of compounds 3a and 3q and helpful
discussions; Dr. Teruaki Okuda and Shin-ichi Niwa for carrying
out CYP3A4 inhibition measurements for the compounds
highlighted in Table 6; Masashi Yamaguchi, Teruki Hamada,
and Atsutoshi Furuta for obtaining rat and monkey pharmaco-
kinetic data for compound 3q in Tables 8 and 9; and Dr. Satoru
Asahi, Miyako Sudo, and Naohiro Kawaguchi for carrying out
CYP2C8, 2C9, and 2D6 inhibition measurements for com-
pound 3q.
Supporting Information Available: Elemental analysis data and
the binding model of the protonated 3a in FXa. This material is
available free of charge via the Internet at http://pubs.acs.org.
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