Discovery and SAR of 2-(1-propylpiperidin-4-yl)-1H-benzimidazole-4-carboxamide: A potent inhibitor of poly(ADP-ribose) polymerase (PARP) for the treatment of cancer

Article abstract of DOI:10.1016/j.bmc.2008.05.044

We have developed a series of cyclic amine-containing benzimidazole carboxamide poly(ADP-ribose)polymerase (PARP) inhibitors, with good PARP-1 enzyme potency, as well as cellular potency. These efforts led to the identification of a lead preclinical candi

Full text of DOI:10.1016/j.bmc.2008.05.044

Bioorganic & Medicinal Chemistry 16 (2008) 6965–6975
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Discovery and SAR of 2-(1-propylpiperidin-4-yl)-1H-benzimidazole-
4-carboxamide: A potent inhibitor of poly(ADP-ribose) polymerase
(PARP) for the treatment of cancer
a
a
a
a
Thomas D. Penning a, , Gui-Dong Zhu , Viraj B. Gandhi , Jianchun Gong , Sheela Thomas ,
Wilfried Lubisch b, Roland Grandel b, Wolfgang Wernet b, Chang H. Park c, Elizabeth H. Fry c,
Xuesong Liu a, Yan Shi a, Vered Klinghofer a, Eric F. Johnson a, Cherrie K. Donawho a,
David J. Frost a, Velitchka Bontcheva-Diaz a, Jennifer J. Bouska a, Amanda M. Olson a,
Kennan C. Marsh d, Yan Luo a, Saul H. Rosenberg a, Vincent L. Giranda a
*
a Cancer Research, GPRD, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064, USA
b Abbott Discovery Research, Neuroscience, D-67008 Ludwigshafen, Germany
c Structural Biology, GPRD, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064, USA
d Pharmacokinetics, GPRD, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
We have developed
a series of cyclic amine-containing benzimidazole carboxamide poly(ADP-
Received 16 April 2008
Revised 20 May 2008
Accepted 21 May 2008
Available online 27 May 2008
ribose)polymerase (PARP) inhibitors, with good PARP-1 enzyme potency, as well as cellular potency.
These efforts led to the identification of a lead preclinical candidate, 10b, 2-(1-propylpiperidin-4-yl)-
1H-benzimidazole-4-carboxamide (A-620223). 10b displayed very good potency against both the
PARP-1 enzyme with a Ki of 8 nM and in a whole cell assay with an EC50 of 3 nM. 10b is aqueous soluble,
orally bioavailable across multiple species, and demonstrated good in vivo efficacy in a B16F10 subcuta-
neous murine melanoma model in combination with temozolomide (TMZ) and in an MX-1 breast xeno-
graph model in combination with cisplatin.
Keywords:
Poly(ADP-ribose)polymerase
PARP inhibitor
Ó 2008 Elsevier Ltd. All rights reserved.
A-620223
Benzimidazole carboxamide
1. Introduction
benefit of several cytotoxic chemotherapeutics, as well as ionizing
radiation.3–7 A significant number of potent PARP inhibitors have
Poly(ADP-ribose) polymerase-1 (PARP-1) is a key DNA damage-
sensing enzyme that facilitates the repair of DNA. All of the PARP
family of enzymes share a catalytic PARP homology domain, but
only PARP-1 and PARP-2 contain DNA-binding domains which
localize them to the sites of DNA damage.1 When triggered by
DNA damage, PARP-1 catalyzes the transfer of ADP-ribose units
from intracellular nicotinamide adenine dinucleotide (NAD+) to
nuclear acceptor proteins, leading to the formation of protein-
bound ADP-ribose polymers. This cellular ADP-ribose transfer pro-
cess is pivotal for the repair of DNA caused by DNA damaging che-
motherapeutic agents or radiation. Thus PARP-1 and PARP-2
contribute to the resistance that often develops after cancer ther-
apy.2 In vivo inhibition of PARP has been shown to block this intra-
cellular DNA repair process and increase the maximum therapeutic
been described in recent years, most containing a primary amide
or lactam functionality.4–8 These inhibitors generally bind to the
nicotinamide binding site of the PARP enzyme and structurally mi-
mic the binding mode of nicotinamide.9 Utilizing a benzimidazole
carboxamide scaffold with relatively high intrinsic potency (i.e., 1),
we have developed a series of cyclic amine-substituted benzimid-
azole analogs leading to a lead preclinical candidate 10b (A-
620223). This compound displayed very good potency against both
PARP-1 and PARP-2, along with oral efficacy in a number of pre-
clinical murine tumor models, potentiating the efficacy of radiation
and several cytotoxic agents such as temozolomide (TMZ) and cis-
platin. 10b has excellent pharmaceutical properties, with signifi-
cant aqueous solubility, moderate protein binding, and good oral
bioavailability in multiple species.
2. Chemistry
* Corresponding author at present address: Abbott Laboratories, Dept. R47S/
AP10-3, 100 Abbott Park Road, Abbott Park, IL 60064-6101, USA. Tel.: +1 847 938
6707; fax: +1 847 935 5165.
A nine-step large-scale synthesis of 10b has been recently de-
scribed.10 The compounds described herein were synthesized
E-mail address: thomas.penning@abbott.com (T.D. Penning).
0968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.05.044

6966
T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
CO2Et
NH2
CO2Et
CONHNH2
N
N
i)
j)
OHC
+
N
R1
N
H
NH2
N
R1
N
H
N
R1
23
24
25
26
k)
CONH2
NH2
2HCl
NH2
CONH2
CONH2
N
NH2
b)
a)
HO2C
O
+
N
R1
N
H
N
R1
N
H
15
16
17
18
N
R1
c) if R1 = BOC
d) if R1 = CBZ
CONH2
N
CONH2
N
h)
e) or f)
g)
N
R5
N
H
N
N
CONH2
N
CHR2R3
CHR2R3
22
20
N
H
N
H
CONH2
N
19
N
H
N
SO2R4
21
Scheme 1. General synthesis of benzimidazole carboxamide analogs. a) CDI, DMF, pyridine or EtO2CCl, THF; b) HOAc, heat; c) TFA, CH2Cl2; d) 10% Pd/C, H2, MeOH; e)
R2R3C(O), NaBH3CN, MeOH; f) R2R3C(O), NaBH(OAc)3, DCE, MeOH, AcOH; g) R4SO2Cl; h) NaH, R5X, DMF; i) Cu(OAc)2, MeOH, H2O, heat; j) NH2NH2, EtOH, heat; k) Raney Ni,
DMF, H2O, heat.
using closely-related procedures outlined in Scheme1. 2,3-Diamin-
obenzamide dihydrochloride 15 was coupled to a cyclic amino acid
16, under CDI or ethyl chloroformate conditions, to give amide 17.
Heating in acetic acid provided the corresponding substituted
benzimidazole 18. In analogs where the cyclic amine was protected
with a BOC- or a CBZ-group, the protecting group was removed
using trifluoroacetic acid or catalytic hydrogenation, respectively,
to provide secondary amines 19. Alkylation of 19 under sodium
cyanoborohydride or sodium triacetoxyborohydride-mediated
reductive amination conditions with aldehydes or ketones pro-
vided tertiary amines 20. Alternately, amine 19 was sulfonylated
using a sulfonyl chloride to provide sulfonamides 21. Selected ana-
logs could also be selectively alkylated at N-1 of the benzimidazole
to provide alkylated compounds 22. An alternate route has also
been employed to generate benzimidazole analogs. Ethyl 2,3-
diaminobenzoate 2311 was heated with a cyclic amino aldehyde
24 and cupric acetate in methanol and water to directly provide
benzimidazole 25. The ester was converted to the hydrazide 26
with hydrazine, followed by Raney nickel reduction to give 18.
The cupric acetate mediated benzimidazole formation with an
aldehyde could also be employed with amide 15 to give 18 in a
one-step process. However, overall, yields were inconsistent and
purity of crude product generally not as high.
3. Results and discussion
The benzimidazole carboxamide series of PARP inhibitors is a
relatively small scaffold with high intrinsic potency due to an
intramolecular hydrogen-bonded conformation that closely mim-
ics the nicotinamide binding interactions in the PARP-1 active site
Gly-863
N
O
NAD+
H
H
H
N
Ser-904
O
N
O
NH2
N
H
N
H
H
O
N
N
O
P
O
P
N+
O
O
N
N
H
O
O
O- O-
HO
HO
OH
1 Ki: 240 nM
OH
OH
Tyr-907
Figure 1. NAD+ interactions with the PARP-1 enzyme.

T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
6967
Table 2
SAR of tertiary cyclic amines
Table 1
SAR of secondary cyclic amines
CONH2
N
CONH2
N
R
R
N
H
N
H
Compound
R
PARP-1 (Ki,
l
M)a
Cellular (EC50
—
, l
M)a,b
Compound
R
PARP-1 (Ki,
l
M)a
Cellular (EC50
—
, l
M)a
1
–H
0.24
3a
0.11
N
N
2a
2b
3
0.044
0.017
0.034
0.006
0.007
0.21
OMe
0.075
0.20
3b
0.18
—
N
H
4a
4b
5a
0.017
0.015
0.008
0.016
0.018
0.022
N
N
4
0.61
NH
5
0.018
N
H
N
6
0.030
0.016
N
H
6a
5b
0.026
0.021
0.076b
0.012
N
N
7
0.004
0.037
0.041
0.011
0.005
0.005
0.16
0.029
0.014
0.024
>1
NH
8a
8b
9
HN
HN
6b
0.020
0.015
N
NH
NH
10
7a
7b
7c
0.008
0.009
0.005
0.003
0.004
0.002
N
N
11
0.031
NH
a
Mean of two or more independent determinations.
N
N
(Fig. 1). Contributing to this scaffold’s potency are key hydrogen-
bond interactions between the amide and two residues in the PARP
active site, Gly-863 and Ser-904, along with a
p-stacking interac-
tion with Tyr-907.9 The benzimidazole carboxamide scaffold has
been previously disclosed by the University of Newcastle12 in
which a series of phenyl-substituted analogs were described.
Although these were relatively potent inhibitors of the PARP-1 en-
zyme, many of these analogs suffered from poor cellular potency.
From our own work, most simple cycloalkyl and phenyl analogs
such as 2a and 2b (Table 1) generally showed a profile of good en-
zyme potency with relatively poor cellular penetration. We found
that incorporation of a basic amine into the 2-substituent of the
benzimidazole ring system in many cases demonstrated a marked
improvement in cellular potency, while maintaining good enzyme
potency. In addition, pharmacokinetic properties also improved
significantly. In this report, we describe the exploration of a series
of cyclic amines, ranging from 4- to 8-membered rings. Table 1
outlines a variety of cyclic, secondary amines.
7d
8c
0.009
0.029
0.006
0.030
N
N
8d
9a
0.045
0.030
0.013
0.015
N
N
9b
0.022
0.019
0.012
0.009
All compounds in this Table had reasonable PARP-1 enzyme
potencies, with Ki values ranging from 5 to 41 nM. However,
although all contained an unsubstituted cyclic amine, there were
profound differences in their ability to inhibit PARP-1 in a whole
cell system. Azetidines 3 and 4, as well as 3-pyrrolidine analog 7
all showed poor cellular potency, while 2-pyrrolidines 5 and 6
10a
N
(continued on next page)

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T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
Table 2 (continued)
Table 3
SAR of modified cyclic amines
Compound
R
PARP-1 (Ki,
M)a
Cellular (EC50
l
,
l
M)a,b
CONH2
N
R
10b
0.008
0.007
0.003
0.004
N
N
N
H
10c
Compound
R
PARP-1 (Ki,
M)a
Cellular (EC50
l
,
l
M)a,b
10d
10e
0.006
0.003
0.005
0.021
N
N
6c
0.016
0.059
0.027
0.028
0.031
0.026
0.18
N
H
O
CN
O
10f
10g
10h
10i
12
—
N
11a
0.005
0.004
0.006
0.001
N
>1
N SO2Ph
0.17
N
N
SO2NMe2
11b
N
0.094b
a
Mean of two or more determinations.
Single determination.
NEt2
b
HN
N
>1
demonstrated relatively good potency. Both 2- and 3-piperidine
analogs 8a, 8b and 9, as well as azepane analog 11 showed reason-
able cellular potency, however, 4-piperidine analog 10 did not.
Since none of the analogs in Table 1 met both our enzyme and
cellular potency criteria (both Ki and EC50 of 610 nM), a variety
of N-alkyl analogs were prepared and the results are highlighted
in Table 2. N-Alkylated 2-azetidines showed a dramatic decrease
in enzyme potency (3a, 3b), while alkylated 3-azetidine analogs
4a and 4b maintained good enzyme potency, while showing a
>30-fold increase in cellular potency. 2-Pyrrolidine analogs 5a,
5b, 6a, and 6b showed a similar enzyme and cellular potency pro-
file as parent analogs 5 and 6. Generally, little difference was noted
between individual enantiomers. However, N-alkylated 3-pyrroli-
dine analogs 7a–7d, showed an exceptional improvement in cellu-
lar potency, improving from a cellular EC50 of 160 nM for parent 7,
to 66 nM for all alkylated analogs. In addition, enzyme potency
was maintained at <10 nM. As demonstrated by compound 7d, sig-
nificant steric bulk was tolerated on the nitrogen substituent, con-
sistent with the presence of a large pocket in the PARP-1 active
site.13 As with the 2-pyrrolidine series, the alkylated 2- and 3-
piperidine series showed very little improvement in either enzyme
and cellular potencies (8c, 8d, 9a, 9b) relative to parent molecules
8a/b and 9. Alkylated 4-piperidine analogs, on the other hand,
demonstrated a dramatic increase in cellular potency (10a–10e)
relative to parent 10, while maintaining good enzyme potency.
Only analog 10e showed cellular EC50 of >20 nM, possibly due to
the reduced basicity of the amine. Alkylated 4-azepane analogs
11a and 11b also showed a very good enzyme and cellular potency
profile. Analogs with somewhat more diverse modifications are
highlighted in Table 3. Reduction in amine basicity by amide or
sulfonamide formation (6c, 10f–10h) resulted in a moderate de-
crease in enzyme potency, but significantly decreased the cellular
potency. Addition of a second basic amine (10i, 12, 12a) was also
detrimental to potency. Small substituents such as chlorine at
the 6-position of the benzimidazole ring were tolerated (i.e., 13),
but larger substituents at this position were generally detrimental
to enzyme potency (data not shown). Alkylation at the 1-position
of the benzimidazole ring (14) resulted in a moderate decrease in
enzyme and cellular potency, consistent with the importance of a
water-mediated hydrogen bond of the NH with Glu-988. Selected
NH
N
12a
13
0.035
0.018
0.097
0.009
CONH2
N
N
N
H
Cl
CONH2
N
14
0.034
0.021
N
N
a
Mean of two or more determinations.
Single determination.
b
compounds were also tested against the closely-related PARP-2 en-
zyme and all had similar Ki values (i.e., 11, 3, 5, 2 and 24 nM for 4,
6c, 10b, 10c and 10g, respectively). Generally, the majority of com-
pounds within the benzimidazole class tested against both PARP-1
and PARP-2 showed similar potency against both enzymes.
Although it would be expected that most PARP-1 inhibitors would
also inhibit PARP-2 to some extent, there have been very limited
reports detailing such selectivity data.16
Mouse (CD-1) pharmacokinetics for selected compounds was
also assessed and the data is highlighted in Table 4. All of these
analogs, with the exception of 5a, showed excellent mouse PK,
with oral bioavailabilities ranging from 26% to 100%, half-lives of
0.4 to 2 h and oral exposures from 0.8 to 3.4 lg-h/mL with a
10 mg/kg dose. Although alkylated 3-pyrrolidine, 4-piperidine
and 4-azepane analogs all showed an acceptable potency profile
and, in addition, several analogs showed a good mouse pharmaco-
kinetic profile, 4-piperidine analog 10b was selected for further
characterization due to its overall profile.
An X-ray co-crystal structure of PARP-1 with 10b is shown in
Figure 2. The key interactions in the PARP-1 active site, consistent

T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
6969
Table 4
Mouse pharmacokinetics
species, including human. In addition, the compound showed min-
imal inhibition of seven cytochrome P450s (<10%) at a concentra-
tion of 10 lg/mL.
Compounda
% F
t1/2 (iv)b
AUC (po)c
Cmax (po)d
In vivo, 10b showed excellent potentiation of cytotoxic agents
in two subcutaneous, murine tumor models: with temozolomide
in B16F10, a syngeneic melanoma model, and with cisplatin in
MX-1, a human breast xenograft model. The B16F10 model
(Fig. 3A), while relatively resistant to most chemotherapeutics,
is moderately sensitive to TMZ and this sensitivity can be en-
hanced with PARP inhibitors.5,7 Compound 10b was adminis-
tered sc via osmotic minipump (OMP) on days 1–14 at doses
of 1, 12.5 and 25 mg/kg/day, while TMZ was administered at
50 mg/kg/day, po, qd, on days 3–7. 10b significantly potentiated
the efficacy of TMZ in a dose-dependent manner. Significant
potentiation was observed as early as day 13 with TGI (tumor
growth inhibition) values (vs vehicle control) of 83%, 79% and
74% for the 25, 12.5 and 1 mg/kg/day 10b combination groups,
respectively, compared to a 62% TGI for TMZ alone. All three
dosing groups continued to differentiate from the TMZ alone
group out to day 17, with TGI values (vs TMZ control) of 82%,
70% and 32% for the 25, 12.5 and 1 mg/kg/day 10b combination
groups, respectively. The 10b/TMZ combinations were well toler-
ated, with maximum body weight loss for all combination
groups similar to the TMZ monotherapy group (5–10%).
5
100
25
89
26
100
63
1.2
0.1
0.8
2
0.4
1.2
1.4
3.4
0.04
1.3
1.0
1.2
1.2
1.7
1.8
0.03
0.7
0.2
0.8
0.4
0.3
5a
7d
9
9b
10b
13
100
a
10 mg/kg po (2.5% EtOH, 1 drop TW-80, 25% PEG400, PBS), 3 mg/kg iv (2.5%
EtOH, 5% DMSO, 1 drop TW-80, 25% PEG400, PBS).
b
h.
c
l
l
g h/mL.
g/mL.
d
In the MX-1 model in female SCID mice (Fig. 3B), 10b was
dosed via sc OMP at doses of 5 and 25 mg/kg/day for 7 days
starting on day 16 post-tumor inoculation, while cisplatin was
given as a single dose on day 18 at 6 mg/kg, ip. The 25 mg/kg/
day 10b/cisplatin combination group was significantly different
than cisplatin alone, with a TGI of 78% at day 48. The 5 mg/kg/
day dose of 10b, however, was not significantly different than
cisplatin alone. The 10b/cisplatin combinations were well toler-
ated, with maximum body weight loss for all combination
groups of 2% compared to 8% loss with the cisplatin mono-
therapy group. Mice quickly regained weight after cessation of
treatment. Although 10b was dosed by OMP in both efficacy
models, similar results were obtained when the compound was
dosed by oral administration (data not shown).
Figure 2. X-ray co-crystal structure of PARP-1 and 10b.
with previous literature reports,9 are highlighted. Both Ser-904 and
Gly-863 are involved in key hydrogen-bond interactions with the
carboxamido group of 10b. There is also a
p-stacking interaction
4. Conclusion
between the benzimidazole ring and Tyr-907. In addition, Glu-
988 is involved in a water-mediated hydrogen bond with the
1-NH of the benzimidazole ring system. These same interactions
are also displayed in the binding of NAD+ (Fig. 1) to the PARP-1 ac-
tive site.
The pharmacokinetics of 10b was further assessed in several
other species and the results are shown in Table 5. Compound
10b was well-absorbed in all species, with 32–82% oral bioavail-
ability and terminal elimination half-lives in the 1.2–2.7 h range.
Clearance and equilibrium volume of distribution values were rel-
atively high. 10b has excellent water solubility of 6 mg/mL at pH
7.8 and moderate plasma protein binding of 57–62% across various
In summary, we have described a potent benzimidazole carbox-
amide series of PARP-1 inhibitors that, due to the presence of an
appropriately substituted cyclic amino group, also demonstrated
good PARP inhibition in whole cells. In addition, several com-
pounds demonstrated a good pharmacokinetic profile in mice.
Based on the overall profile, N-propyl-4-piperidinyl analog 10b
was identified for further pharmacological characterization. 10b
showed good pharmaceutical properties, including pharmacoki-
netics across multiple species. In addition, 10b demonstrated
excellent potentiation of two cytotoxic agents, temozolomide and
cisplatin, in both a mouse melanoma and a breast cancer model.
Compound 10b was identified as a lead preclinical candidate for
the treatment of human cancer in combination with both radiation
and various cytotoxic agents.
Table 5
Multispecies pharmacokinetics for 10b
e
Species
Dosea
% F
t1/2 (iv)b
AUC (po)c
Cmax (po)d
Vss
CLf
5. Experimental
5.1. Chemistry
Mouse
Rat
Dog
10
63
48
82
32
1.2
2.6
2.7
1.7
1.2
0.7
3.7
0.7
0.4
0.8
1
9.1
10.7
3.6
5.3
2.9
0.9
2
4.1
4.1
4.1
Monkey
0.2
4.9
a
mg/kg.
h.
5.1.1. General
b
c
d
e
f
1H NMR spectra were obtained on Varian M-300, Bruker
AMX-400, Varian U-400, and Varian Unity Inova 500 magnetic
resonance spectrometers with indicated solvent and an internal
standard. Chemical shifts are given in delta (d) values and
l
l
g-h/mL.
g/mL.
L/kg.
L/h/kg.

6970
T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
1400
1200
1000
800
600
400
200
0
A
10b / TMZ (25 / 50 mkd)
10b / TMZ (12.5 / 50 mkd)
10b / TMZ (1 / 50 mkd)
10b / TMZ (0 / 50 mkd)
Combination vehicle
0
5
10
15
20
Temozolomide
PO, QD d3-7
Days Post Tumor Inoculation
10b, SC, OMP, d1-14
compd
dose
(mg/kg/day)
tumor volumea
(day 13)
%TGI vehicleb
(day 13)
tumor volumea
(day 17)
%TGI cytotoxicc
(day 17)
10b/TMZ
25/50
12.5/50
1/50
161 16
196 13
246 23
364 32
955 120
83*
79*
74*
62*
--
233 32
378 45
853 72
1250 83
--
82*
70*
32*
--
vehicle/TMZ
0/50
combination vehicle
0/0
--
* p<0.0001; a Mean (mm3) SEM of 9-10 mice/group; b %TGI vs. vehicle control; c %TGI vs. TMZ control.
2500
B
10b / cisplatin (25 / 6 mkd)
10b / cisplatin (5 / 6 mkd)
10b / cisplatin (0 / 6 mkd)
Combination vehicle
2000
1500
1000
500
0
0
7
14
21
28
35
42
49
56
63
Cisplatin, IP, d18
Days Post Tumor Inoculation
10b, SC
OMP
d16-23
compd
dose
(mg/kg/day)
tumor volumea
(day 29)
%TGIb
(day 29)
tumor volumea
(day 48)
%TGIb
(day 48)
10b/cisplatin
25/6
5/6
0/6
0/0
106 12
164 40
32
0
299 67
1100 289
1389 300
--
78*
21
--
vehicle/cisplatin
156 28
--
--
combination vehicle
1730 429
--
* p< 0.001; a Mean (mm3) SEM of 9-10 mice/group; b %TGI vs. cisplatin control.
Figure 3. (A) In vivo efficacy of 10b with TMZ in the B16F10 model. (B) In vivo efficacy of 10b with cisplatin in the MX-1 model.
coupling constants (J) in Hertz (Hz). The following peak multi-
plicity abbreviations are used: s, singlet; d, doublet; t, triplet;
q, quartet; m, multiplet; br, broadened. Mass spectra were
performed as follows: ESI (electrospray ionization) was per-
formed on a Finnigan SSQ7000 MS run as a flow injection
acquisition; DCI (desorption chemical ionization) was performed
on a Finnigan SSQ7000 MS using a direct exposure probe with
ammonia gas; APCI (atmospheric pressure chemical ionization)
was performed on a Finnigan Navigator MS run as flow injec-
tion acquisition. Elemental analyses were performed by Quantita-
tive Technologies Inc. Whitehouse, NJ. All manipulations were
performed under nitrogen atmosphere unless otherwise indicated.

T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
6971
All solvents and other reagents were obtained from commercial
5.2.3. MX-1 tumor model
sources and used without further purification, except where
noted.
A 0.2 cc of 1:10 MX-1 tumor brei was injected subcutaneously
into the flank of female SCID mice (Charles River Labs) on study
day 0. On day 15, tumors were size matched and animals placed
into study groups (N = 10 mice/group). PARP inhibitor therapy be-
gan on day 16, with cisplatin treatment starting on day 18. At var-
ious intervals following tumor inoculation, the individual tumor
dimensions were serially measured using calibrated microcalipers
and the tumor volumes calculated according to the formula
V = L ꢀ W2/2 (V: volume, L: length, W: width). Mice were humanely
euthanized when the tumor volumes reached a predetermined
size.
5.1.2. X-ray crystallography data
A GST fusion of h-PARP (654–1014) expressed in Escherichia
coli was concentrated to 60 mg/mL (0.874 mM) in 50 mM pH
7.5 Tris buffer containing 150 mM NaCl and 1.5 mM DTT. To
make the PARP–10b complex, 2 mM 10b in DMSO was added
to the protein solution and incubated for 4 h on ice. Crystals
were grown at 17 °C in hanging drops after 2–5 d. The well
solution contained 1.3 M ammonium sulfate and 1.3 M NaCl.
X-ray diffraction data were collected at Advanced Photon
Source Beamline 32-ID. The cryoprotectant solution of 1.2 M
ammonium sulfate, 1.6 M NaCl and 20% ethylene glycol was
used to collect the data at 110 K. The space group is P321
5.2.4. B16F10 tumor model
For B16F10 syngeneic studies, 6 ꢀ 104 cells were mixed with
50% matrigel (BD Biosciences, Bedford, MA) and inoculated by sc
injection into the flank of 6–8 week old female C57BL/6 mice,
20 g (Charles River Laboratories, Wilmington, MA). Mice were
injection-order allocated to treatment groups and PARP inhibitor
therapy was initiated on day 1 following inoculation, with tem-
ozolomide treatment starting on day 3. All animal studies were
conducted in a specific pathogen-free environment in accordance
to the Internal Institutional Animal Care and Use Committee,
accredited by the American Association of Laboratory Animal Care
under conditions that meet or exceed the standards set by the Uni-
ted States Department of Agriculture Animal Welfare Act, Public
Health Service policy on humane care and use of animals, and
the NIH guide on laboratory animal welfare.
with cell dimensions, a = b = 94.17, c = 68.38 Å,
a = 90,
c
= 120 °. There is one PARP–10b complex molecule per asym-
metric unit. The crystal diffracted up to 2.8 Å and 8321 unique
reflections were obtained using HKL2000. The overall Rsym(I) is
0.08 and I/r(I) is 13.8. The overall completeness is 92%. The
complex structure was solved using CCP4 molrep program with
a search model of PDB entry 1UK0 and was refined with CNX
2002. The conventional and free R-factors after refinement are
0.234 and 0.317. The coordinates of the PARP–10b complex
molecule are deposited in Protein Data Bank with the code
2RCW.
5.2. Biology
5.3. Chemistry
5.2.1. PARP enzyme assay4
Enzyme assay was conducted in buffer containing 50 mM Tris,
pH 8.0, 1 mM DTT, and 4 mM MgCl2. PARP reactions contained
5.3.1. Procedure A: 2-(Azetidin-2-yl)-1H-benzimidazole-4-
carboxamide (3)
1.5
l
M [3H]NAD+ (1.6
lCi/mmol), 200 nM biotinylated histone
A solution of 1-(tert-butoxycarbonyl)azetidine-2-carboxylic acid
(2.5 g, 12.4 mmol) in a mixture of pyridine (15 mL) and DMF (15 mL)
was treated with1,10-carbonyldiimidazole (CDI, 2.2 g, 13.6 mmol) at
45 °C for 2 h. 2,3-Diaminobenzamide dihydrochloride10 (2.8 g,
12.4 mmol) was added and the mixture stirred at ambient tempera-
ture overnight. Solvents were removed and the residue heated in
10 mL of acetic acid at 100 °C for 30 min. After concentration, the
residuewas partitionedbetweenethylacetateand saturatedsodium
bicarbonate solution and the organic phase washed with water and
concentrated. The residue was purified by flash chromatography
using ethyl acetate to give the BOC-protected intermediate. This
was treated with trifluoroacetic acid (5 mL) in dichloromethane
(20 mL) for 1 h and concentrated. The residue was purified by HPLC
(Zorbax, C-18, 0–100% gradient of 0.1% TFA in H2O/0.1% TFA in
CH3CN) to afford 670 mg (12%) of the title compound as the trifluo-
roacetate salt. 1H NMR (CD3OD) d 2.90–3.17 (m, 2H), 4.10–4.26 (m,
1H), 4.26–4.41 (m, 1H), 5.87 (t, J = 8.4 Hz, 1H), 7.42 (m, 1H), 7.69–
7.83 (m, 1H), 7.89–8.09 (m, 1H). HRMS m/z 217.10781 (calcd for
H1, 200 nM slDNA, and 1 nM PARP-1 or 4 nM PARP-2 enzyme.
Auto reactions utilizing SPA bead-based detection were carried
out in 100 mL volumes in white 96-well plates. Reactions were
initiated by adding 50 ml of 2ꢀ NAD+ substrate mixture to
50 ml of 2ꢀ enzyme mixture containing PARP and DNA. These
reactions were terminated by the addition of 150 ml of 1.5 mM
benzamide (ꢁ1000-fold over its IC50). One hundred and seventy
microliters of the stopped reaction mixtures were transferred to
streptavidin-coated Flash Plates, incubated for 1 h, and counted
using a TopCount microplate scintillation counter. Ki data was
determined from inhibition curves at various substrate
concentrations.
5.2.2. Cellular PARP assay
C41 cells were treated with test compound for 30 min in a
96-well plate. PARP was activated by damaging DNA with
1 mM H2O2 for 10 min. Cells were washed with ice-cold PBS
once and fixed with pre-chilled methanol/acetone (7:3) at
ꢂ20 °C for 10 min. After air-drying, plates were rehydrated with
PBS and blocked 5% non-fat dry milk in PBS-Tween (0.05%)
(blocking solution) for 30 min at room temperature. Cells were
incubated with anti-PAR antibody 10H (1:50) in blocking solu-
tion at room temperature for 60 min followed by washing with
PBS-Tween 20 five times, and incubation with goat anti-mouse
fluorescein 5(6)-isothiocyanate (FITC)-coupled antibody (1:50)
C11H13N4O (M+H), 217.10839).
5.3.2. 2-(Azepan-4-yl)-1H-benzimidazole-4-carboxamide (11)
The title compound was prepared from 1-(tert-butoxycar-
bonyl)azepane-4-carboxylic acid14 according to procedure A. 1H
NMR (CD3OD) d 1.99–2.15 (m, 2H), 2.16–2.28 (m, 1H), 2.36–2.54
(m, 3H), 3.33–3.41 (m, 2H), 3.41–3.49 (m, 1H), 3.53–3.62 (m,
1H), 3.61–3.73 (m, 1H), 7.58 (t, J = 7.5 Hz, 1H), 7.91 (d, J = 7.5 Hz,
1H), 7.98–8.03 (d, J = 7.4 Hz, 1H). HRMS m/z 259.15500 (calcd for
and 1 l
g/ml 40,6-diamidino-2-phenylindole (DAPI) in blocking
solution at room temperature for 60 min. After washing with
PBS-Tween 20 five times, analysis was performed using an fmax
Fluorescence Microplate Reader set at the excitation and emis-
sion wavelength for FITC or the excitation and emission wave-
length for DAPI. PARP activity (FITC signal) was normalized
with cell numbers (DAPI).
C14H19N4O (M+H), 259.15534).
5.3.3. 2-(1-(4-cyanophenyl)piperidin-4-yl)-1H-benzimidazole-
4-carboxamide (10e)
The title compound was prepared from 1-(4-cyano-
phenyl)piperidine-4-carboxylic acid according to procedure
A

6972
T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
(6.5% yield). 1H NMR (DMSO-d6) d 1.89 (dd, J = 12.5, 3.1 Hz, 2H),
2.12 (s, 2H), 3.10 (t, J = 11.7 Hz, 2H), 4.05 (d, J = 13.1 Hz, 3H), 7.08
(d, J = 9.4 Hz, 3H), 7.25 (s, 1H), 7.56–7.65 (m, 4H), 7.78 (s, 1H),
9.28 (s, 1H). Anal. calcd for C20H19N5Oꢃ0.9 H2O: C: 66.43, H: 5.80,
N: 19.37. Found: C: 66.67, H: 5.69, N: 19.21.
J = 6.4 Hz, 1H), 9.22 (br s, 1H). Anal. calcd for C12H14N4O: C, 62.59;
H, 6.13; N, 24.33. Found: C, 62.25; H, 5.99; N, 23.97.
5.3.8. 2-(Pyrrolidin-3-yl)-1H-benzimidazole-4-carboxamide (7)
The title compound was prepared from 1-(benzyloxycar-
bonyl)pyrrolidine-3-carboxylic acid according to procedure B (57%
yield). 1H NMR (D2O) d 2.45–2.55 (m, 1H), 2.73–2.81 (m, 1H),
3.54–3.61 (m, 1H), 3.69–3.74 (m, 1H), 3.74–3.80 (m, 1H), 4.01 (dd,
J = 12.1, 8.4 Hz, 1H), 4.21–4.31 (m, 1H), 7.58 (t, J = 8.0 Hz, 1H),
7.87–7.93 (m, 2H). Anal. calcd for C12H14N4Oꢃ2.4HCl: C, 45.36; H,
5.20; N, 17.63. Found: C, 45.75; H, 4.82; N, 16.95.
5.3.4. 6-Chloro-2-(1-isopropylpiperidin-4-yl)-1H-benzimida-
zole-4-carboxamide (13)
5.3.4.1. Step 1: Preparation of 2-amino-5-chloro-3-nitrobenza-
mide. To
a
solution of 2-amino-3-nitrobenzamide10 (4.0 g,
22.08 mmol) in acetonitrile (1250 mL) was added N-chlorosuc-
cinimide (3.1 g, 23.18 mmol) and the mixture stirred at 60 °C
overnight. After cooling, the solid was collected by filtration,
washed with acetonitrile, and dried to give 2.95 g of the title
compound. The mother liquor was concentrated and the residue
recrystallized from acetonitrile to give 800 mg of a second crop
of solid (79% yield). 1H NMR (DMSO-d6) d 7.75 (br s, 1H), 8.00
(d, J = 2.4 Hz, 1H), 8.19 (d, J = 2.7 Hz, 1H), 8.25 (br s, 1H), 8.47
(br s, 2H). MS (DCI/NH3) m/z 216 (M+H)+.
5.3.9. (R)-2-(Piperidin-2-yl)-1H-benzimidazole-4-carboxamide
(8a)
The title compound was prepared from (R)-(+)-1-(carbobenzyl-
oxy)-2-piperidine carboxylic acid according to procedure B. 1H
NMR (CD3OD) d 1.75–1.84 (m, 2H), 1.88–1.96 (m, 2H), 1.99–2.06
(m, 1H), 2.33–2.46 (m, 1H), 3.08–3.20 (m, 1H), 3.43–3.54 (m,
1H), 4.53 (dd, J = 11.5, 3.22 Hz, 1H), 7.34–7.39 (m, 1H), 7.74 (d,
J = 8.0 Hz, 1H), 7.94 (d, J = 7.7 Hz, 1H). HRMS m/z 245.13952 (calcd
for C13H17N4O (M+H), 245.13969).
5.3.4.2. Step 2: Preparation of 6-Chloro-2-(1-isopropylpi-
peridin-4-yl)-1H-benzimidazole-4-carboxamide. The title com-
pound was prepared from the compound from step 1 and 1-isopro-
pylpiperidine-4-carboxylic acid hydrochloride15 according to
procedure A (66% yield). 1H NMR (DMSO-d6) d 1.45 (d, J = 6.7 Hz,
6H), 2.39–2.55 (m, 4H), 3.31–3.37 (m, 2H), 3.57–3.82 (m, 4H),
7.98 (d, J = 1.8 Hz, 1H), 8.10 (d, J = 1.8 Hz, 1H). Anal. calcd for
5.3.10. (S)-2-(Piperidin-2-yl)-1H-benzimidazole-4-carboxamide
(8b)
The title compound was prepared from (S)-(ꢂ)-1-(carbobenzyl-
oxy)-2-piperidine carboxylic acid according to procedure B. 1H
NMR (CD3OD): d 1.73–1.85 (m, 2H), 1.87–1.97 (m, 2H), 1.99–2.07
(m, 1H), 2.39 (dd, J = 12.9, 2.8 Hz, 1H), 3.09–3.19 (m, 1H), 3.48 (d,
J = 11.7 Hz, 1H), 4.54 (dd, J = 11.3, 3.4 Hz, 1H), 7.35–7.41 (m, 1H),
7.75 (dd, J = 8.0, 0.9 Hz, 1H), 7.95 (dd, J = 7.5, 1.1 Hz, 1H). HRMS
m/z 245.13939 (calcd for C13H17N4O (M+H), 245.13969).
C
16H21ClN4Oꢃ2.4 HCl: C, 47.06; H, 5.78; N, 13.72. Found: C, 46.98;
H, 5.50; N, 13.41.
5.3.5. Procedure B: 2-(Azetidin-3-yl)-1H-benzimidazole-4-
carboxamide (4)
A solution of (1.2 g, 5.0 mmol) in a mixture of pyridine (10 mL)
and DMF (10 mL) was treated with CDI (0.9 g, 5.5 mmol) at 45 °C
for 2 h. 2,3-Diaminobenzamide dihydrochloride10 (1.1 g, 5.0 mmol)
was added and the mixture stirred at ambient temperature over-
night. Solvents were removed and the residue heated in acetic acid
(10 mL) at 100 °C for 30 min. After concentration, the residue was
purified by flash chromatography using ethyl acetate to afford the
CBZ-protected intermediate. This was dissolved in methanol
(10 mL), and treated with 10% palladium on carbon (100 mg) under
hydrogen for 6 h. Solid material was filtered off and the filtrate
concentrated. The residue was purified by HPLC (Zorbax C-8, 0.1%
trifluoroacetic acid/acetonitrile/water) to provide 820 mg (54%) of
the title compound. 1H NMR (CD3OD) d 4.51–4.58 (m, 4H), 7.42
(t, J = 7.9 Hz, 1H), 7.79 (d, J = 8.2 Hz, 1H), 7.94 (d, J = 7.6 Hz, 1H).
HRMS m/z 217.10796 (calcd for C11H13N4O (M+H), 217.10839).
5.3.11. 2-(Piperidin-3-yl)-1H-benzimidazole-4-carboxamide (9)
The title compound was prepared from 1-(benzyloxycar-
bonyl)piperidine-3-carboxylic acid according to procedure B. 1H
NMR (CD3OD) d 1.62–1.73 (m, 1H), 1.77–1.86 (m, 1H), 1.87–1.98
(m, 1H), 2.19–2.28 (m, 1H), 2.64–2.77 (m, 1H), 2.93–3.02 (m,
1H), 3.03–3.10 (m, 1H), 3.10–3.20 (m, 1H), 3.37 (s, 1H), 7.29 (t,
J = 7.8 Hz, 1H), 7.66 (dd, J = 7.8, 1.0 Hz, 1H), 7.86 (dd, J = 7.8,
1.0 Hz, 1H). MS (DCI/NH3) m/z 245 (M+H)+.
5.3.12. 2-(Piperidin-4-yl)-1H-benzimidazole-4-carboxamide
(10)
The title compound was prepared from 1-(benzyloxycar-
bonyl)piperidine-4-carboxylic acid according to procedure B. 1H
NMR (DMSO-d6): d 1.63–1.71 (m, 1H), 1.71–1.78 (m, 1H), 1.90–
2.00 (m, 2H), 2.55–2.66 (m, 2H), 2.94–2.99 (m, 1H), 3.00–3.08 (m,
2H), 3.26 (br s, 1H), 7.24 (t, J = 7.8 Hz, 1H), 7.62 (d, J = 7.1 Hz, 1H),
7.64 (br s, 1H), 7.78 (d, J = 7.5 Hz, 1H), 9.34 (br s, 1H), 12.61 (br s,
1H). HRMS m/z 245.13979 (calcd for C13H17N4O (M+H), 245.13969).
5.3.6. (R)-2-(Pyrrolidin-2-yl)-1H-benzimidazole-4-carboxamide
(5)
The title compound was prepared from (R)-1-(benzyloxycar-
bonyl)pyrrolidine-2-carboxylic acid according to procedure
B
5.3.13. (S)-2-(5-Oxopyrrolidin-2-yl)-1H-benzimidazole-4-
(57% yield). 1H NMR (DMSO-d6) d 1.69–1.83 (m, 2H), 1.88–2.05
(m, 1H), 2.12–2.29 (m, 1H), 2.96 (t, J = 6.6 Hz, 2H), 3.31 (br s, 1H),
4.44 (dd, J = 8.0, 5.9 Hz, 1H), 7.23 (t, J = 7.6 Hz, 1H), 7.61 (d,
J = 1.0 Hz, 1H), 7.64 (br s, 1H), 7.77 (dd, J = 7.8, 1.0 Hz, 1H), 9.22
(br s, 1H), 12.22 (br s, 1H). Anal. calcd for C12H14N4O: C, 62.59;
H, 6.13; N, 24.33. Found: C, 62.32; H, 5.93; N, 24.43.
carboxamide (6c)
The title compound was prepared from Z-D-pyroglutamic acid
according to procedure B. 1H NMR (DMSO-d6) d 2.14–2.19 (m,
1H), 2.27–2.36 (m, 2H), 2.53–2.60 (m, 1H), 4.99 (dd, J = 8.0,
5.3 Hz, 1H), 7.27–7.34 (m, 1H), 7.69 (d, J = 7.1 Hz, 2H), 7.82 (d,
J = 7.1 Hz, 1H), 8.25 (br s, 1H), 9.20 (br s, 1H), 12.97 (brs, 1H). Anal.
calcd for C12H12N4O2: C, 59.01; H, 4.95; N, 22.94. Found: C, 58.87;
H, 4.79; N, 22.79.
5.3.7. (S)-2-(Pyrrolidin-2-yl)-1H-benzimidazole-4-carboxamide
(6)
The title compound was prepared from (S)-1-(benzyloxycar-
bonyl)pyrrolidine-2-carboxylic acid according to procedure B. 1H
NMR (DMSO-d6) d 1.71–1.82 (m, 2H), 1.89–2.02 (m, 1H), 2.14–2.26
(m, 1H), 2.96 (t, J = 6.8 Hz, 2H), 4.44 (dd, J = 8.1, 6.1 Hz, 1H), 7.23 (t,
J = 7.8 Hz, 1H), 7.61 (br s, 1H), 7.63 (dd, J = 1.4, 7.0 Hz, 1H), 7.77 (d,
5.3.14. Procedure C: 2-(N-Acetylpiperidin-4-yl)-1H-benzimida-
zole-4-carboxamide (10f)
5.3.14.1. Step 1: Preparation of Methyl 2-(N-acetylpiperidin-4-
yl)-1H-benzimidazole-4-carboxylate. To a solution of ethyl 2,3-
diaminobenzoate11 (3.3 g, 19.9 mmol) in methanol (100 mL) was

T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
6973
added a solution of N-acetylpiperidine-4-carboxaldehyde (4.0 g,
25.8 mmol) in methanol (100 mL) and the mixture stirred at ambi-
ent temperature for 10 min. A solution of copper(II) acetate (5.2 g,
25.8 mmol) in water (100 mL) was added and the mixture stirred
at reflux for 30 min. After cooling, concentrated hydrochloric acid
(25 mL) was added and the mixture warmed to reflux. A solution
of sodium sulfide nonahydrate (7.15 g, 29.8 mmol) in water
(100 mL) was added and the mixture refluxed for 10 min. After
cooling, the mixture was concentrated and the residue stirred in
water and filtered. The filtrate was made basic with aqueous so-
dium bicarbonate and extracted with ethyl acetate. The organic
phase was washed with water, dried and concentrated to provide
4.5 g of the title compound.
J = 8.1, 0.9 Hz, 1H), 8.06 (dd, J = 7.6, 0.9 Hz, 1H). Anal. calcd for
C
14H19N5Oꢃ2.5HClꢃ2H2O: C, 41.98; H, 6.42; N, 17.49. Found: C,
42.21; H, 6.53; N, 17.46.
5.3.19. 2-Piperazin-2-yl-1H-benzimidazole-4-carboxamide (12)
5.3.19.1. Step 1: Preparation of 2-(1,4-dibenzylpiperazin-2-yl)-
1H-benzimidazole-4-carboxamide. The title compound was pre-
pared from 1,4-dibenzylpiperazine-2-carboxaldehyde according to
Procedure C. 1H NMR (DMSO-d6) d 2.95–3.7 (7H), 3.8–4.9 (4H), 7.1–
7.55 (8H), 7.65 (2H), 7.85 (2H), 7.94 (1H), 8.7 (br), 12.2 (br).
5.3.19.2. Step 2: Preparation of 2-Piperazin-2-yl-1H-benzimi-
dazole-4-carboxamide. A solution of the compound from step 1
(1.83 g, 3.67 mmol) and 10% palladium on carbon (1 g) in methanol
(250 mL) was stirred under hydrogen until starting material was
consumed. The catalyst was filtered off and the filtrate concen-
trated. To a solution of the residue in isopropanol (20 mL) was
added 20% hydrogen chloride in isopropanol (50 mL) and the pre-
cipitate filtered to obtain 1.1 g of the title compound. 1H NMR
(CD3OD) d 3.39–3.72 (m, 5H), 3.97 (dd, J = 13.6, 3.4 Hz, 1H), 5.08
(dd, J = 10.2, 3.7 Hz, 1H), 7.47 (t, J = 7.9 Hz, 1H), 7.86 (dd, J = 8.1,
1.0 Hz, 1H), 7.99 (dd, J = 7.5, 1.0 Hz, 1H). HRMS m/z 246.13497
(calcd for C12H16N5O (M+H), 246.13494).
5.3.14.2. Step 2: Preparation of 2-(N-acetylpiperidin-4-yl)-1H-
benzimidazole-4-carbohydrazide. A solution of the compound
from step 1 (4.3 g, 14.9 mmol) and hydrazine hydrate (3.7 g, 74.
3 mmol) in ethanol (100 mL) was stirred at reflux for 2.5 h. The
mixture was concentrated and the crude product used without fur-
ther purification.
5.3.14.3. Step 3: Preparation of 2-(N-Acetylpiperidin-4-yl)-1H-
benzimidazole-4-carboxamide. A solution of the compound from
step 2 in water was added to a solution of Raney nickel (5 g) in
DMF (100 mL) and water (50 mL) and the mixture heated at
100 °C for 2 h. After cooling, the mixture was filtered and the fil-
trate concentrated. The residue was dissolved in dichloromethane
and diethyl ether added to provide a precipitate. Filtration gave
3.2 g of the title compound. 1H NMR (DMSO-d6) d 1.60–1.95 (m,
4H), 2.04 (s, 3H), 2.76 (m, 3H), 3.92 (d, J = 13.9 Hz, 1H), 4.42 (d,
J = 13.1 Hz, 1H), 7.26 (t, J = 7.2 Hz, 1H), 7.64 (br s, d, J = 7.5 Hz,
2H), 7.80 (d, J = 7.2 Hz, 1H), 8.5 (br), 9.29 (br, 1H), 12.70 (br, 1H).
Anal. calcd for C15H18N4O2ꢃ1.4H2O: C, 57.83; H, 6.73; N, 17.98.
Found: C, 57.83; H, 6.34; N, 18.00.
5.3.20. Procedure D: 2-(1-Isopropylazetidin-2-yl)-1H-
benzimidazole-4-carboxamide (3a)
A solution of 3 (100 mg, 0.22 mmol) in methanol (10 mL) was
stirred with formaldehyde (37 wt% in water, 163 lL, 2.2 mmol) at
ambient temperature overnight. Sodium cyanoborohydride
(117 mg, 2.2 mmol) was added and the solution stirred at ambient
temperature for 3 h. After concentration, the residue was purified
by HPLC (Zorbax C-8, 0.1% trifluoroacetic acid/acetonitrile/water)
to give the title compound (28 mg, 23%). 1H NMR (CD3OD) d 1.25
(d, J = 6.4 Hz, 6H), 2.92 (d, J = 8.3 Hz, 1H), 3.00–3.14 (m, 1H),
3.66–3.79 (m, 1H), 4.19–4.29 (m, 2H), 5.84 (t, J = 9.0 Hz, 1H),
7.40–7.47 (m, 1H), 7.80–7.84 (m, 1H), 8.00 (dd, J = 7.7, 1.2 Hz,
5.3.15. 2-(N-Methylpiperidin-3-yl)-1H-benzimidazole-4-
carboxamide (9a)
1H). HRMS m/z 259.15523 (calcd for
C14H19N4O (M+H),
The title compound was prepared from 1-methylpiperidine-3-
carboxaldehyde according to Procedure C. 1H NMR (DMSO-d6) d
1.75 (m, 1H), 2.04 (m, 2H), 2.24 (m, 1H), 2.83 (d, J = 4.7 Hz, 3H),
3.44 (m, 2H), 3.60–3.95 (m, 3H), 7.36 (t, J = 7.8 Hz, 1H), 7.76 (d,
J = 7.8 Hz, 1H), 7.87 (d, J = 6.8 Hz, 1H), 8.80 (br, 1H), 10.62 (br,
1H). Anal. calcd for C14H18N4Oꢃ2HClꢃ1H2O: C, 48.15; H, 6.35; N,
16.04. Found: C, 47.85; H, 6.40; N, 15.87.
259.15534).
5.3.21. 2-(1-Cyclopentylazetidin-2-yl)-1H-benzimidazole-4-
carboxamide (3b)
The title compound was prepared from 3 and cyclopentanone
according to procedure D (37% yield). 1H NMR (CD3OD) d 1.50–
1.85 (m, 6H), 1.98–2.21 (m, 2H), 2.88–3.02 (m, 1H), 3.02–3.19
(m, 1H), 4.03 (s, 1H), 4.16–4.35 (m, 2H), 5.82 (t, J = 8.9 Hz, 1H),
7.44 (t, J = 7.8 Hz, 1H), 7.83 (d, J = 8.0 Hz, 1H), 7.95–8.03 (m,
5.3.16. 2-(N-Methylpiperidin-4-yl)-1H-benzimidazole-4-
carboxamide (10a)
1H). HRMS m/z 285.17066 (calcd for
C16H21N4O (M+H),
The title compound was prepared from 1-methylpiperidine-4-
carboxaldehyde according to Procedure C. HRMS m/z 259.15584
(calcd for C14H19N4O (M+H), 259.15534).
285.17099).
5.3.22. 2-(1-Propylazetidin-3-yl)-1H-benzimidazole-4-
carboxamide (4a)
5.3.17. 2-(1-(2-(Diethylamino)ethyl)piperidin-4-yl)-1H-
benzimidazole-4-carboxamide (10i)
The title compound was prepared from 4 and propionaldehyde
according to procedure D (28% yield). 1H NMR (CD3OD) d 1.03 (t,
J = 7.5 Hz, 3H), 1.62–1.71 (m, 2H), 3.31–3.35 (m, 2H), 4.42–4.73
(m, 4H), 7.39 (t, J = 7.9 Hz, 1H), 7.77 (d, J = 7.9 Hz, 1H), 7.93 (d,
J = 7.6 Hz, 1H). HRMS m/z 259.15507 (calcd for C14H19N4O (M+H),
259.15534).
The title compound was prepared from 1-(2-diethylaminoeth-
yl)piperidine-4-carboxaldehyde according to Procedure C. 1H
NMR (CD3OD) d 1.37 (t, J = 7.1 Hz, 6H), 1.82–2.38 (m, 6H), 2.65–
3.54 (m, 11H), 7.39 (t, J = 7.9 Hz, 1H), 7.75 (d, J = 7.9 Hz, 1H), 7.90
(d, J = 7.5 Hz, 1H). Anal. calcd for C19H29N5Oꢃ2.3HClꢃ1H2O: C,
51.24; H, 7.54; N, 15.73. Found: C, 51.35; H, 7.61; N, 15.49.
5.3.23. 2-(1-Cyclopentylazetidin-3-yl)-1H-benzimidazole-4-
carboxamide (4b)
5.3.18. 2-(1,4-Dimethylpiperazin-2-yl)-1H-benzimidazole-4-
carboxamide (12a)
The title compound was prepared from 1,4-dimethylpiperazine-
2-carboxaldehyde according to Procedure C. 1H NMR (CD3OD) d
2.38 (s, 3H), 2.94 (m, 2H), 3.01 (s, 3H), 3.39 (m, 2H), 3.68 (m,
1H), 3.82 (m, 1H), 4.44 (m, 1H), 7.63 (t, J = 8.0 Hz, 1H), 7.95 (dd,
The title compound was prepared from 4 and cyclopentanone
according to procedure D (27% yield). 1H NMR (CD3OD) d 1.58–
1.72 (m, 2H), 1.72–1.88 (m, 4H), 2.03–2.20 (m, 2H), 3.92–4.07
(m, 1H), 4.38–4.51 (m, 1H), 4.58–4.64 (m, 4H), 7.39 (t, J = 6.8 Hz,
1H), 7.77 (d, J = 6.8 Hz, 1H), 7.93 (d, J = 6.44 Hz, 1H). HRMS m/z
285.17078 (calcd for C16H21N4O (M+H), 285.17099).

6974
T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
5.3.24. (R)-2-(1-methylpyrrolidin-2-yl)-1H-benzimidazole-4-
carboxamide (5a)
5.3.31. 2-(1-Isopropylpiperidin-2-yl)-1H-benzimidazole-4-
carboxamide (8d)
The title compound was prepared from 5 and formaldehyde
according to procedure D (67% yield). 1H NMR (CD3OD) d 2.30–
2.46 (m, 2H), 2.46–2.61 (m, 1H), 2.78–2.88 (m, 1H), 3.09 (s, 3H),
3.45–3.57 (m, 1H), 3.87–4.00 (m, 1H), 5.00–5.13 (m, 1H), 7.53 (t,
J = 7.8 Hz, 1H), 7.89 (d, J = 8.1 Hz, 1H), 8.04 (d, J = 7.5 Hz, 1H). MS
(DCI/NH3) m/z 245 (M+H)+.
The title compound was prepared as described for 8c using ace-
tone in place of formaldehyde. 1H NMR (CD3OD) d 1.30 (d,
J = 6.6 Hz, 3H), 1.36 (d, J = 6.9 Hz, 3H), 1.74–1.88 (m, 1H), 1.98–
2.06 (m, 2H), 2.09–2.15 (m, 1H), 2.28–2.35 (m, 2H), 3.17–3.24
(m, 1H), 3.36–3.46 (m, 1H), 3.67 (d, J = 11.8 Hz, 1H), 4.74–4.80
(m, 1H), 7.43 (t, J = 8.0 Hz, 1H), 7.82 (d, J = 9.0 Hz, 1H), 7.99 (d,
J = 7.8 Hz, 1H). Anal. calcd for C16H22N4Oꢃ1.9TFA: C, 47.28; H,
4.79; N, 11.14. Found: C, 47.49; H, 4.54; N, 11.24.
5.3.25. (S)-2-(1-Methylpyrrolidin-2-yl)-1H-benzimidazole-4-
carboxamide (6a)
The title compound was prepared from 6 and formaldehyde
according to procedure D (67% yield). 1H NMR (CD3OD) d 2.34–
2.41 (m, 2H), 2.52 (m, 1H), 2.83 (m, 1H), 3.09 (s, 3H), 3.49 (m, 1H),
3.94 (m, 1H), 5.06 (br s, 1H), 7.53 (t, J = 8.0 Hz, 1H), 7.89 (d,
J = 8.1 Hz, 1H), 8.03 (d, J = 7.5 Hz, 1H). MS (APCI) m/z 245 (M+H)+.
5.3.32. 2-(1-Isopropylpiperidin-3-yl)-1H-benzimidazole-4-
carboxamide (9b)
The title compound was prepared from 9 and acetone according
to procedure D. 1H NMR (CD3OD) d 1.43 (d, J = 6.5 Hz, 6H), 1.89–
1.99 (m, 1H), 2.00–2.10 (m, 1H), 2.21 (d, J = 14.3 Hz, 1H), 2.33–
2.41 (m, 1H), 3.11–3.20 (m, 1H), 3.46 (t, J = 12.2 Hz, 1H), 3.57 (d,
J = 11.8 Hz, 1H), 3.61–3.72 (m, 2H), 3.89 (d, J = 11.5 Hz, 1H), 7.41
(t, J = 7.8 Hz, 1H), 7.78 (d, J = 7.8 Hz, 1H), 7.93 (d, J = 7.5 Hz, 1H).
Anal. calcd for C16H22N4Oꢃ2.2TFA: C, 45.61; H, 4.54; N, 10.43.
Found: C, 45.43; H, 4.85; N, 10.53.
5.3.26. (R)-2-(1-Isopropylpyrrolidin-2-yl)-1H-benzimidazole-4-
carboxamide (5b)
The title compound was prepared from 5 and acetone accord-
ing to procedure D (60% yield). 1H NMR (CD3OD) d 1.37 (d,
J = 6.4 Hz, 3H), 1.39 (d, J = 6.4 Hz, 3H), 2.23–2.32 (m, 2H),
2.33–2.44 (m, 1H), 2.66–2.77 (m, 1H), 3.46–3.58 (m, 1H), 3.75–
3.90 (m, 2H), 5.08–5.19 (m, 1H), 7.42 (t, J = 7.8 Hz, 1H), 7.81
(d, J = 8.0 Hz, 1H), 7.98 (d, J = 7.4 Hz, 1H). Anal. calcd for
5.3.33. 2-(1-Isopropylpiperidin-4-yl)-1H-benzimidazole-4-
carboxamide (10c)
The title compound was prepared from 10 and acetone accord-
ing to procedure D. 1H NMR (D2O) d 1.30 (d, J = 6.8 Hz, 6H), 2.08–
2.23 (m, 2H), 2.45 (br d, J = 14.2 Hz, 2H), 3.11–3.26 (m, 2H),
3.47–3.66 (m, 4H), 7.48 (t, J = 8.0 Hz, 1H), 7.78 (d, J = 7.7 Hz, 1H),
7.81 (d, J = 8.1 Hz, 1H). Anal. calcd for C16H22N4Oꢃ1.8 HClꢃ1.3 H2O:
C, 51.19; H, 7.09; N, 14.92, Cl, 17.00. Found: C, 51.03; H, 7.15; N,
14.69, Cl, 16.88.
C
15H20N4Oꢃ1.6TFA: C, 48.07; H, 4.79; N, 12.32. Found: C, 48.26;
H, 4.85; N, 12.55.
5.3.27. (S)-2-(1-Isopropylpyrrolidin-2-yl)-1H-benzimidazole-4-
carboxamide (6b)
The title compound was prepared from 6 and acetone according
to procedure D. 1H NMR (CD3OD) d 1.42 (d, J = 5.9 Hz, 6H), 2.33 (m,
2H), 2.43–2.50 (m, 1H), 2.72–2.83 (m, 1H), 3.57 (m, 1H), 3.81–3.89
(m, 2H), 5.26 (m, 1H), 7.51 (t, J = 7.8 Hz, 1H), 7.89 (d, J = 8.1 Hz, 1H),
8.02 (d, J = 7.5 Hz, 1H). Anal. calcd for C15H20N4Oꢃ1.7 HCl: C, 53.89;
H, 6.54; N, 16.76. Found: C, 53.76; H, 6.59; N, 16.67.
5.3.34. 2-(1-Cyclopentylpiperidin-4-yl)-1H-benzimidazole-4-
carboxamide (10d)
The title compound was prepared from 10 and cyclopentanone
according to procedure D. 1H NMR (CD3OD) d 1.67–1.81 (m, 4H),
1.81–1.91 (m, 2H), 2.20–2.26 (m, 3H), 2.26–2.32 (m, 1H), 2.49 (d,
J = 14.0 Hz, 2H), 3.21 (t, J = 12.0 Hz, 2H), 3.44–3.52 (m, 1H), 3.55–
3.65 (m, 1H), 3.81 (d, J = 12.8 Hz, 2H), 7.45 (t, J = 8.0 Hz, 1H), 7.80
(d, J = 7.8 Hz, 1H), 7.94 (d, J = 7.8 Hz, 1H). HRMS m/z 313.20172
(calcd for C18H25N4O (M+H), 313.20229).
5.3.28. 2-(1-Isopropylazepan-4-yl)-1H-benzimidazole-4-
carboxamide (11a)
The title compound was prepared from 11 and acetone accord-
ing to procedure D (23% yield). 1H NMR (CD3OD) d 1.41 (d,
J = 6.9 Hz, 6H), 2.06–2.22 (m, 2H), 2.20–2.31 (m, 1H), 2.34–2.43
(m, 1H), 2.53 (d, J = 5.3 Hz, 2H), 3.34–3.56 (m, 2H), 3.56–3.65 (m,
2H), 3.65–3.79 (m, 2H), 7.59 (t, J = 8.0 Hz, 1H), 7.93 (d, J = 7.5 Hz,
1H), 8.00 (d, J = 7.5 Hz, 1H). HRMS m/z 301.20161 (calcd for
5.3.35. Procedure E: 2-(1-Propylpyrrolidin-3-yl)-1H-
benzimidazole-4-carboxamide (7a)
To a solution of 7 (200 mg, 0.87 mmol) in dichloroethane
(8 mL) and methanol (1 mL) was added propionaldehyde
C17H25N4O (M+H), 301.20229).
(127
ature for 1 h. Sodium triacetoxyborohydride (369 mg, 1.74 mmol)
and acetic acid (100 L, 1.74 mmol) were added and the mixture
lL, 1.74 mmol) and the solution stirred at ambient temper-
5.3.29. 2-(1-Cyclopentylazepan-4-yl)-1H-benzimidazole-4-
carboxamide (11b)
l
The title compound was prepared from 11 and cyclopentanone
according to procedure D (25% yield). 1H NMR (CD3OD) d 1.63–1.93
(m, 6H), 2.03-2.28 2.03–2.28 (m, 5H), 2.33–2.41 (m, 1H), 2.42–2.52
(m, 2H), 3.36–3.49 (m, 1H), 3.55–3.73 (m, 3H), 3.72–3.83 (m, 2H),
7.53 (t, J = 8.0 Hz, 1H), 7.88 (d, J = 8.0 Hz, 1H), 7.97 (d, J = 7.4 Hz, 1H).
HRMS m/z 327.21744 (calcd for C19H27N4O (M+H), 327.21794).
stirred at ambient temperature overnight. After concentration,
the residue was purified by HPLC (Zorbax, C-18, 0.1% TFA/
CH3CN/H2O). The trifluoroacetate salt was dissolved in dichloro-
methane/methanol, treated with 1N HCl in ether and concen-
trated to provide 159 mg (48%) of the hydrochloride salt. 1H
NMR (D2O) d 1.03 (t, J = 7.5 Hz, 3H), 1.78–1.87 (m, 2H), 2.50–
2.56 (m, 0.5H), 2.66–2.71 (m, 0.5H), 2.83 (dd, J = 14.0, 7.6 Hz,
0.5H), 2.90–2.96 (m, 0.5H), 3.33–3.47 (m, 2H), 3.54–3.68 (m,
1H), 3.91 (t, J = 11.3 Hz, 0.5H), 3.96–4.01 (m, 1H), 4.02–4.07 (m,
0.5H), 4.16 (dd, J = 12.5, 7.0 Hz, 0.5H), 4.31–4.37 (m, 1H), 4.50–
4.55 (m, 0.5H), 7.65 (t, J = 7.9 Hz, 1H), 7.95 (d, J = 8.1 Hz, 1H),
7.97 (d, J = 8.1 Hz, 1H). MS (APCI) m/z 273 (M+H)+.
5.3.30. 2-(1-Methylpiperidin-2-yl)-1H-benzimidazole-4-
carboxamide (8c)
The title compound was prepared from ( )-1-(benzyloxycar-
bonyl)-2-piperidine carboxylic acid according to procedure B, fol-
lowed by procedure D using formaldehyde. 1H NMR (CD3OD) d
1.71–1.87 (m, 1H), 1.90–2.12 (m, 3H), 2.14–2.26 (m, 1H), 2.29–
2.39 (m, 1H), 2.81 (s, 3H), 3.32–3.36 (m, 1H), 3.73 (d, J = 13.2 Hz,
1H) 4.52–4.67 (m, 1H), 7.41–7.45 (m, 1H), 7.82 (d, J = 8.0 Hz, 1H),
7.99 (d, J = 7.7 Hz, 1H). Anal. calcd for C14H18N4Oꢃ1.5 TFA: C,
47.56; H, 4.58; N, 13.05. Found: C, 47.84; H, 4.52; N, 13.23.
5.3.36. 2-(1-Isopropylpyrrolidin-3-yl)-1H-benzimidazole-4-
carboxamide (7b)
The title compound was prepared from 7 and acetone according
to procedure E (54% yield). 1H NMR (D2O) d 1.45 (d, J = 6.4 Hz, 6H),

T. D. Penning et al. / Bioorg. Med. Chem. 16 (2008) 6965–6975
6975
2.44–2.51 (m, 0.5H), 2.57–2.64 (m, 0.5H), 2.73–2.80 (m, 0.5H), 2.83–
2.91 (m, 0.5H), 3.42–3.48 (m, 0.5H), 3.56–3.62 (m, 0.5H), 3.62–3.70
(m, 1H), 3.88–3.96 (m, 1H), 4.09 (dd, J = 11.9, 6.4 Hz, 0.5H), 4.23
(m, 1H), 4.38 (m, 0.5H), 7.57 (t, J = 8.1 Hz, 1H), 7.89 (d, J = 8.1 Hz,
1H), 7.90 (d, J = 8.1 Hz, 1H). Anal. calcd for C15H20N4Oꢃ 2.7 HCl: C,
48.59; H, 6.17; N, 15.11. Found: C, 48.37; H, 6.24; N, 14.79.
and the mixture stirred at ambient temperature for 1 h. Iodometh-
ane (52 mg, 0.37 mmol) was added and the mixture stirred at
ambient temperature for 16 h and concentrated. The residue was
partitioned between ethyl acetate and brine and the organic phase
was washed with brine and concentrated. Purification by flash
chromatography on silica gel using 20% methanol in dichlorometh-
ane afforded 70 mg (67%) of the title compound. 1H NMR (DMSO-
d6) d 0.95 (t, J = 7.5 Hz, 3H), 1.65–1.77 (m, 2H), 2.10–2.26 (m, 4H),
3.01–3.17 (m, 4H), 3.18–3.25 (m, 1H), 3.29 (s, 3H), 3.38–3.49 (m,
1H), 3.57–3.69 (m, 1H), 7.36 (t, J = 7.8 Hz, 1H), 7.77 (dd, J = 8.1,
0.9 Hz, 1H), 7.86 (d, J = 7.5 Hz, 1H), 9.07 (br s, 1H), 9.22 (br s,
5.3.37. 2-(1-Cyclopentylpyrrolidin-3-yl)-1H-benzimidazole-4-
carboxamide (7c)
The title compound was prepared from 7 and cyclopentanone
according to procedure E (54% yield). 1H NMR (D2O) d 1.67–1.87
(m, 6H), 2.18–2.26 (m, 2H), 2.46–2.55 (m, 0.5H), 2.63–2.69 (m,
0.5H), 2.80 (dd, J = 14.5, 7.8 Hz, 0.5H), 2.88–2.94 (m, 0.5H), 3.42–
3.50 (m, 0.5H), 3.55–3.62 (m, 0.5H), 3.64–3.70 (m, 0.5H), 3.77–
3.84 (m, 1H), 3.91–3.97 (m, 1H), 4.01 (dd, J = 10.4, 7.9 Hz, 0.5H),
4.13 (dd, J = 12.7, 6.9 Hz, 0.5H), 4.30 (q, J = 7.1 Hz, 1H), 4.45–4.53
(m, 0.5H), 7.63 (t, J = 8.1 Hz, 1H), 7.93 (d, J = 7.6 Hz, 1H), 7.95 (d,
J = 7.6 Hz, 1H). Anal. calcd for C17H22N4Oꢃ2.9 HCl: C, 50.53; H,
6.21; N, 13.86. Found: C, 50.83; H, 6.20; N, 13.49.
1H). HRMS m/z 301.20211 (calcd for
C17H25N4O (M+H),
301.20229).
References and notes
1. Virág, L.; Szabó, C. Pharmacol. Rev. 2002, 54, 375–429.
2. (a) Munoz-Gamez, J. A.; Martin-Oliva, D.; Aguilar-Quesada, R.; Canuelo, A.;
Nunez, M. I.; Valenzuela, M. T.; Ruiz de Almodovar, J. M.; de Murcia, G.; Oliver,
J. A. Biochem. J. 2005, 386, 119–125; (b) Tentori, L.; Graziani, G. Pharmacol. Res.
2005, 52, 25–33; (c) Shirou, S.; Nomura, F.; Tomonaga, T.; Sunaga, M.; Noda, M.;
Ebara, M.; Saisho, H. Oncol. Rep. 2004, 12, 821–825; (d) Griffin, R. J.; Curtin, N. J.;
Newell, D. R.; Golding, B. T.; Durkacz, B. W.; Calvert, A. H. Biochemie 1995, 77,
408–422.
3. (a) Plummer, E. R. Curr. Opin. Pharmacol. 2006, 6, 364–368; (b) Horváth, E. M.;
Szabó, C. Drug News Perspect. 2007, 20, 171–181; (c) Ratnam, K.; Low, J. A. Clin.
Cancer Res. 2007, 13, 1383–1388.
4. Donawho, C. K.; Luo, Y.; Luo, Y.; Penning, T. D.; Bauch, J. L.; Bouska, J. J.;
Bontcheva-Diaz, V. D.; Cox, B. F.; DeWeese, T. L.; Dillehay, L. E.; Ferguson, D. C.;
Ghoreishi-Haack, N. S.; Grimm, D. R.; Guan, R.; Han, E. K.; Holley-Shanks, R.;
Hristov, B.; Idler, K. B.; Jarvis, K.; Johnson, E. F.; Kleinberg, L. E.; Klinghofer, V.;
Lasko, L. M.; Liu, X.; Marsh, K. C.; McGonigal, T. P.; Meulbroek, J. A.; Olson, A.
M.; Palma, J. P.; Rodriguez, L. E.; Shi, Y.; Stavropoulos, J. A.; Tsurutani, A. C.; Zhu,
G.-D.; Rosenberg, S. H.; Giranda, V. L.; Frost, D. J. Clin. Cancer Res. 2007, 13,
2728–2737.
5.3.38. 2-(1-Phenethylpyrrolidin-3-yl)-1H-benzimidazole-4-
carboxamide (7d)
The title compound was prepared from 7 and phenylacetalde-
hyde according to procedure E (45% yield). 1H NMR (CD3OD) d
2.48–2.56 (m, 1H), 2.73 (m, 1H), 3.11–3.15 (m, 2H), 3.59–3.64
(m, 2H), 3.66 (m, 2H), 3.93–4.04 (m, 2H), 4.13–4.20 (m, 1H),
7.26–7.38 (m, 5H), 7.40 (t, J = 8.0 Hz, 1H), 7.76 (d, J = 8.1 Hz, 1H),
7.93 (d, J = 8.1 Hz, 1H). Anal. calcd for C20H22FN4Oꢃ2.25 TFA: C,
49.79; H, 4.14; N, 9.48. Found: C, 49.96; H, 4.19; N, 9.54.
5.3.39. 2-(1-(Phenylsulfonyl)piperidin-4-yl)-1H-benzimidazole-
4-carboxamide (10g)
5. Lapidus, R. G.; Tentori, L.; Graziani, G.; Leonetti, C.; Scarsella, M.; Vergati, M.;
Muzi, A.; Zhang, J. J. Clin. Oncol. 2005, 23, 3136.
To a solution of 10 (100 mg, 0.41 mmol) in pyridine (5 mL) was
added benzenesulfonyl chloride (109 mg, 0.62 mmol) and the mix-
ture stirred at ambient temperature for 16 h. The mixture was con-
centrated, stirred in water and methanol for 30 min and filtered.
The solid was washed with water and methanol and dried to give
90 mg (57%) of the title compound. 1H NMR (DMSO-d6) d 1.78–1.95
(m, 2H), 2.06–2.19 (m, 2H), 2.53–2.59 (m, 2H), 2.93–3.06 (m, 1H),
3.74 (d, J = 11.9 Hz, 2H), 7.26 (t, J = 7.6 Hz, 1H), 7.60–7.63 (m, 1H),
7.63–7.66 (m, 1H), 7.67 (t, J = 1.9 Hz, 1H), 7.69–7.70 (m, 1H), 7.72–
7.75 (m, 1H), 7.76–7.78 (m, 1H), 7.78 (br s, 1H), 7.79–7.82 (m, 1H),
9.18 (br s, 1H), 12.68 (br s, 1H). Anal. calcd for C19H20N4O3S: C,
59.36; H, 5.24; N, 14.57. Found: C, 59.23; H, 5.20; N, 14.64.
6. Calabrese, C. R.; Almassy, R.; Barton, S.; Batey, M. A.; Calvert, A. H.; Canan-Koch,
S.; Durkacz, B. W.; Hostomsky, Z.; Kumpf, R. A.; Kyle, S.; Li, J.; Maegley, K.;
Newell, D. R.; Notarianni, E.; Stratford, I. J.; Skalitsky, D.; Thomas, H. D.; Wang,
L.-Z.; Webber, S. E.; Williams, K. J.; Curtin, N. J. J. Natl. Cancer Inst. 2004, 96, 56–
67.
7. Tentori, L.; Leonetti, C.; Scarsella, M.; d’Amati, G.; Vergati, M.; Portarena, I.; Xu,
W.; Kalish, V.; Zupi, G.; Zhang, J.; Graziani, G. Clin. Cancer Res. 2003, 9, 5370–
5379.
8. (a) Li, J.-H.; Zhang, J. IDrugs 2001, 4, 804–812; (b) Cockcroft, X.; Dillon, K. J.;
Dixon, L.; Drzewiecki, J.; Kerrigan, F.; Loh, V. M.; Martin, N. M. B.; Menear, K. A.;
Smith, G. C. M. Bioorg. Med. Chem. Lett. 2006, 16, 1040–1044.
9. (a) Ferraris, D.; Ficco, R. P.; Dain, D.; Ginski, M.; Lautar, S.; Lee-Wisdom, K.;
Linag, S.; Lin, Q.; Lu, M. X.-C.; Morgan, L.; Thomas, B.; Williams, L. R.; Zhang, J.;
Zhou, Y.; Kalish, V. J. Bioorg. Med. Chem. 2003, 11, 3695–3707; (b) Canan-Koch,
S. S.; Thoresen, L. H.; Tikhe, J. G.; Maegley, K. A.; Yu, X.-H.; Zook, S. E.; Kumpf, R.
A.; Zhang, C.; Boritzki, T. J.; Mansour, R. N.; Zhang, K. E.; Ekker, A.; Calabrese, C.
R.; Curtin, N. J.; Kyle, S.; Thomas, H. D.; Wang, L.-Z.; Calvert, A. H.; Golding, B. T.;
Griffin, R. J.; Newell, D. R.; Webber, S. E.; Hostomsky, Z. J. Med. Chem. 2002, 45,
4961–4974; (c) Costatino, G.; Macchiarulo, A.; Camaioni, E.; Pellicciari, R.
J. Med. Chem. 2001, 44, 3786–3794.
5.3.40. 2-(1-(N,N-dimethylsulfamoyl)piperidin-4-yl)-1H-
benzimidazole-4-carboxamide (10h)
To a solution of 10 (150 mg, 0.614 mmol) in dichloromethane
10. Barkalow, J. H.; Breting, J.; Gaede, B. J.; Haight, A. R.; Henry, R.; Kotecki, B.; Mei,
J.; Pearl, K. B.; Tedrow, J. S.; Viswanath, S. K. Org. Process Res. Dev. 2007, 11,
693–698.
(8 mL) and methanol (1 mL) was added triethylamine (171
lL,
1.23 mmol) and dimethylsulfamoylchloride (79 L, 0.737 mmol)
l
11. Lubisch, W.; Kock, M.; Hoeger, T.; Grandel, R.; Schult, S.; Mueller, R. Cyclo-alkyl
substituted benzimidazoles and their use as PARP inhibitors. U.S. Patent
6,737,421, 2004.
12. White, A. W.; Curtin, N. J.; Eastman, B. W.; Golding, B. T.; Hostomsky, Z.; Kyle,
S.; Li, J.; Maegley, K. A.; Skalitzky, D. J.; Webber, S. E.; Yu, X.-H.; Griffin, R. J.
Bioorg. Med. Chem. Lett. 2004, 14, 2433–2437.
13. Hattori, K.; Kido, Y.; Yamamoto, H.; Ishida, J.; Kamijo, K.; Murano, K.; Ohkubo,
M.; Kinoshita, T.; Iwashita, A.; Mihara, K.; Yamazaki, S.; Matsuoka, N.;
Teramura, Y.; Miyake, H. J. Med. Chem. 2004, 47, 4151–4154.
14. Okamoto, Y.; Kubota, H.; Sato, I.; Hattori, K.; Kanayama, T.; Yokoyama, K.; Terai,
Y.; Takeuchi, M. 2-Aminopyridine derivative. PCT Int. Appl. WO 2005100341,
2005.
and the mixture stirred at ambient temperature overnight. The solu-
tion was purified by flash chromatography on silica gel using meth-
anol in dichloromethane to afford 165 mg (77%) of the title
compound. 1H NMR (DMSO-d6) d 1.81–1.93 (m, 2H), 2.12 (t,
J = 12.6 Hz, 2H), 2.79 (s, 6H), 3.03 (t, J = 11.5 Hz, 2H), 3.15 (t,
J = 11.2 Hz, 1H), 3.65–3.70 (m, 2H), 7.29 (t, J = 7.5 Hz, 1H), 7.66 (d,
J = 7.7 Hz, 1H), 7.67 (s, 1H), 7.83 (d, J = 7.1 Hz, 1H), 9.31 (br s, 1H),
12.74 (br s, 1H). HRMS m/z 352.14350 (calcd for C15H22N5O3S
(M+H), 352.14379).
15. Lubisch, W.; Kock, M.; Höger, T.; Schult, S.; Grandel, R.; Müller, R.
Substituted benzimidazoles and their use as PARP inhibitors. U.S. Patent
6,448,271, 2002.
16. Ishida, J.; Yamamoto, H.; Kido, Y.; Kamijo, K.; Murano, K.; Miyake, H.; Ohkubo,
M.; Kinoshita, K.; Warizaya, M.; Iwashita, A.; Mihara, K.; Matsuoka, N.; Hattori,
K. Bioorg. Med. Chem. 2006, 14, 1378–1390.
5.3.41. 1-Methyl-2-(1-propylpiperidin-4-yl)-1H-benzimidazole-
4-carboxamide (14)
To a solution of 10b (100 mg, 0.35 mmol) in DMF (5 mL) was
added sodium hydride (60% in mineral oil, 0.35 mmol, 14 mg)