1,2-disubstituted indole, azaindole and benzimidazole derivatives possessing amine moiety: A novel series of thrombin inhibitors

Article abstract of DOI:10.1016/S0960-894X(00)00454-6

A novel series of 1,2-disubstituted indole, azaindole and benzimidazole derivatives possessing an amine moiety was identified as thrombin inhibitors. An indole with basic diamine moieties (12a) was the most potent thrombin inhibitor in the series with K(ass) = 197 x 10⁶ L/mol. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0960-894X(00)00454-6

Bioorganic & Medicinal Chemistry Letters 10 (2000) 2347±2351
1,2-Disubstituted Indole, Azaindole and Benzimidazole Derivatives
Possessing Amine Moiety: A Novel Series of Thrombin Inhibitors
Kumiko Takeuchi,* Jolie A. Bastian, Donetta S. Gi€ord-Moore, Richard W. Harper,
Shawn C. Miller, Je€rey T. Mullaney, Daniel J. Sall, Gerald F. Smith, Minsheng Zhangy
and Matthew J. Fisher
Lilly Research Laboratories, Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN 46285, USA
Received 16 May 2000; accepted 8 August 2000
AbstractÐA novel series of 1,2-disubstituted indole, azaindole and benzimidazole derivatives possessing an amine moiety was
identi®ed as thrombin inhibitors. An indole with basic diamine moieties (12a) was the most potent thrombin inhibitor in the series
with Kass=197 Â 106 L/mol. # 2000 Elsevier Science Ltd. All rights reserved.
Thrombin, a trypsin-like serine protease, catalyzes ®brin
formation and activates platelets, thereby playing a
pivotal role in the development of thrombotic diseases.1
Recently we have reported the discovery of speci®c,
active site directed thrombin inhibitors that are based on
the 2,3-disubstituted benzo[b]thiophenes (e.g., 1a,b).2 7
Within that e€ort, we observed by an X-ray analysis of a
crystal of the ternary complex between a-thrombin,
hirugen, and a compound from this series that the benzo-
[b]thiophene nucleus binds into the hydrophobic S1
speci®city pocket of thrombin.2,3 Since this binding
interaction is thought to be critical for good thrombin
Chemistry
inhibitory potency, we elected to examine this relation-
ship in detail. During the initial evaluation of this series,
the role of the heterocyclic core was investigated by repla-
cing the benzo[b]thiophene nucleus with other heteroaro-
matic rings such as benzofuran and indole. The initial brief
structure±activity relationship (SAR) of the 2,3-di-
substituted heteroaromatic compounds seemed to indicate
that benzo[b]thiophene was the preferred heteroaromatic
ring (1b±3, Table 1).2 However, we found that the di€er-
ential positioning of the two side chains profoundly
in¯uenced the thrombin inhibitory activity of the nitro-
gen-containing heteroaromatic compounds. In this report
we describe the SAR studies of novel 1,2-disubstituted
indole, azaindole and benzimidazole compounds that
exhibited unexpected thrombin inhibitory activity.
Synthesis of the compounds 1a,b has previously been
reported2,6 and the compounds 2 and 3 were prepared in a
similar manner. In those previous SAR studies for the
benzo[b]thiophene-derived thrombin inhibitors2 7 exten-
sive modi®cation of the 2,3-disubstituted heteroaromatic
moiety proved to be a labor intensive undertaking due
to the two carbon±carbon bond formations. A 1,2-di-
substituted indole (or similar nitrogen heterocycles)
would allow us to avoid one of the carbon±carbon bond
forming reactions for the attachment of the two required
side chains. Such substitution pattern would then facil-
itate rapid SAR development towards the desired
thrombin inhibitory potency. Schemes 1±3 depict gen-
eral syntheses of benzimidazole, azaindole, and indole
compounds. Generally, 2-substituted heteroaromatic ring
formation, N1-alkylation with 4-carbomethoxybenzyl
bromide, followed by modi®cation of the carbomethoxy
group to appropriate amine moiety furnished target
molecules. In the case of benzimidazoles, N-alkylation
preceded the ring formation.
*Corresponding author. Tel.: +1-317-276-6771; fax: +1-317-433-0715;
e-mail: ktak@lilly.com
yCurrent address: Bristol-Myers Squibb Pharmaceutical Research
Institute, Princeton, NJ 08543.
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00454-6

2348
K. Takeuchi et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2347±2351
Results and Discussion
was 150 times more potent than the 2,3-disubstituted
indole 3. It was also twofold more potent than the 2,3-
disubstituted benzo[b]thiophene 1b. 1,2-Disubstituted
benzimidazole 5 showed eightfold better thrombin inhibi-
tory activity than the 2,3-disubstituted indole 3. In contrast,
it was 10-fold weaker than the 2,3-disubstituted benzo-
[b]thiophene 1b. Among the 2,3-disubstituted hetero-
cycles the benzo[b]thiophene 1b was clearly superior to
the benzofuran 2 and the indole 3 (1b >2 >> 3). These
results seemed to indicate that the nitrogen-containing
heteroaromatic core with the 1,2-disubstition pattern
might have a salient feature that would be very di€erent
from the 2,3-disubstituted benzo[b]thiophene ring. 1,2-Dis-
ubstituted indoles and other heteroaromatic compounds
could be viable candidates for developing structurally
unique, potent thrombin inhibitors. Further SAR stu-
dies of 1,2-disubstituted indoles, benzimidazoles, and
azaindoles were therefore warranted.
There was a stark contrast in the e€ects of the disubstitu-
tion pattern of the heterocycles on the thrombin inhibitory
activity as shown in Table 1. 1,2-Disubstituted indole 4
Table 1. SAR on the heteroaromatic core
Compound
A
B
Kass (Â106 L/mol)8
1b
2
3
4
5
S
O
NH
C
C
C
C
N
N
3.43
0.974
0.042
6.27
The results of SAR studies on the benzimidazole series
are shown in Tables 2±4. Table 2 shows the substituent
N
0.332
Scheme 1. Synthesis of benzimidazole derivatives.
Scheme 2. Synthesis of azaindole derivatives.9

K. Takeuchi et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2347±2351
2349
Scheme 3. Synthesis of indole derivatives.
e€ects on the thrombin inhibitory activity of the benz-
imidazoles. Our extensive SAR studies of benzo[b]thio-
phene derived thrombin inhibitors revealed that the C30-
methoxy substituent4 on the C3-phenyl ring and the
contracted C40-side chain by a methylene linker5 7 were
extremely bene®cial to the thrombin inhibitory activity.
In the case of 1,2-disubstituted indole analogues, the N1
side chain could be considered equivalent to the C3-side
chain of the benzo[b]thiophene in its molecular orienta-
tion, and therefore the same substituent pattern at C30
and C40 positions of the C3-phenyl ring was applied to
the N1 side chain of benzimidazole 5. This modi®cation
proved to be bene®cial and improved the thrombin
inhibitory activity of benzimidazole 36-fold (from 5 to 6a).
6-Hydroxy substituent (7a) did not have any bene®cial
e€ect on the activity in contrast to the benzo[b]thiophene
Table 2. Substituent e€ects on the thrombin inhibitory activity of
benzimidazoles versus benzo[b]thiophenes
series4 which showed about eightfold increase from Kass
=
126 to 815 Â 106 L/mol. Longer chain substituents such
as methyleneamine (8) and methylenehydroxy group (9)
were detrimental to the inhibitory activity as were seen
with benzo[b]thiophene series.2
Kass (Â106 L/mol)8
Table 3 shows the e€ects of the C40-amine substituents.
The basicity of the cyclic amine was not critical to the
inhibitory activity (7a±d) in contrast to the benzo-
[b]thiophene derivatives where a basic amine side chain
was required.2,4 The benzylamine 7e showed moderate
improvement on the activity. These results indicate that
benzimidazole compounds behave di€erently than the
benzo[b]thiophene series. We also examined the e€ect of
C2-side chain modi®cation (Table 4). As in the case of
benzo[b]thiophene,2 the C2-side chain preferred a basic
1,2-Disubstituted
benzimidazole
(A, B=N)
2,3-Disubstituted
benzo[b]thiophene
(A=S; B=C)
Compound
Y
6a
7a
8
H
OH
CH2NH2
CH2OH
11.9
9.72
4.49
1.93
126
815
Ð
9
Ð
Table 3. Modi®cation of C40-amine
Table 4. Modi®cation of C2-side chain
Compound
NR1R2
Kass (Â106 L/mol)8
7a
7b
7c
7d
7e
Pyrrolidin-1-yl
Imidazol-1-yl
Triazol-1-yl
Pyrazol-1-yl
Benzylamino
9.72
9.60
10.2
12.0
16.5
Compound
NR3R4
Kass (Â106 L/mol)8
6a
6b
6c
Pyrrolidin-1-yl
Pyrrolid-2-on-1-yl
Pyrazol-1-yl
11.9
1.50
1.46

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K. Takeuchi et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2347±2351
amine group for potent thrombin inhibitory activity.
Less basic pyrrolidone 6b and pyrrazole 6c were eight-
fold less active than the pyrrolidine 6a.
Table 6 shows the SAR results of indole derivatives. In
contrast to the benzimidazole series, 1,2-disubstituted
indole 12a showed potent thrombin inhibitory activity,
even better than the benzo[b]thiophene series (6a benzo-
[b]thiophene). Within the indole series, the compound
12a containing optimal side chains by the 1,2-substitu-
tion pattern was over 30-fold more potent than 4 and
almost 4700 times more potent than 3. As in the case of
benzimidazole (Table 4), a basic amine moiety on the
C2-side chain was preferred to maintain potent thrombin
inhibitory activity for both the indole and benzo[b]thio-
phene series. The inhibitory activity decreased from six-
to 30-fold with the indole series and from 15- to 32-fold
with the benzo[b]thiophene series as the basicity of the
C2-side chain was reduced (12b±d). Among these com-
pounds, the indole series exhibited better inhibition
overall than the benzo[b]thiophene series. In contrast,
the benzimidazole series lost activity considerably, 10-
to 80-fold when compared with benzo[b]thiophene
(Table 2) and 17-fold when compared with indole (6a
versus 12a).
Table 5 shows the thrombin inhibitory activity of 5- or
6-azaindole derivatives. Interestingly, the more rigid
and less basic amide moiety of the N1 side chain (10a)
showed modestly better activity than the methylene
amine linkage (10b) in the azaindole series. The aza-
indole 10b exhibited respectable activity with over sixfold
increase from 6a (benzimidazole), but about twofold
less potency than 6a (benzo[b]thiophene) and threefold
less than 12a (indole, Table 6). 5-Azaindole 10a showed
better activity than 6-azaindole 11. This was possibly
due to the better positioning of the ring nitrogen at the
5- rather than the 6-position to interact with the carboxy
terminus of Asp189 of thrombin, assuming the orienta-
tion of the molecule being the same as that of the 2,3-
disubstituted benzo[b]thiophene in the thrombin speci-
®city pocket. These results suggest that the positioning
of the ring nitrogen in¯uences the thrombin inhibitory
activity of disubstituted nitrogen-containing heteroaro-
matic compounds much more signi®cantly than the
basicity of an amine moiety at the C40-phenyl ring.
Results in Tables 1±3, 5, and 6 suggest that the nitrogen-
containing heteroaromatic derivatives might be inter-
acting with thrombin very di€erently from the benzo-
[b]thiophene series of thrombin inhibitors. In general, a
hydrophobic ring was favored in the narrow hydro-
phobic cavity of the S1 speci®c pocket of thrombin, at
the end of which was exposed the carboxy group of
Asp189.10 Among the nitrogen-containing heteroaro-
matic compounds studied, the most hydrophobic indole
ring with one nitrogen was more favored than azaindole
or benzimidazole with two nitrogens in the ring in this
order. 5-Azaindole was more favored than 6-azaindole
possibly due to the better positioning of the ring nitrogen
at the 5- rather than the 6-position for its interaction with
the Asp189 carboxy terminus. The more hydrophilic
benzimidazole ring was less favored possibly due to the
electrostatic repulsion by the N3 in the narrow and
hydrophobic cavity of the S1 pocket, which might have
prevented the inhibitor from intercalating deep into the
cavity. This might explain why the bene®cial e€ect of
the hydroxy group observed with the benzo[b]thiophene
series11 appeared to be nulli®ed in the case of benzimi-
dazole, where the hydroxy group could not reach deep
enough to interact with the Asp189 carboxy terminus of
thrombin. The preliminary results of indole and aza-
indole derivatives presented here suggest that these
compounds may provide a novel class of thrombin
inhibitors with unique features, di€erent from the benzo-
[b]thiophene series. The indole analogues could o€er
advantages of synthetic ease and possess desirable throm-
bin inhibitory potency without a hydroxy substituent,
which was required for the benzo[b]thiophene series.
Table 5. Azaindole derivatives
Compound
X
Y
Z
Kass (Â106 L/mol)8
10a
10b
11
C
C
N
N
N
C
O
H,H
O
83.7
73.9
59.7
Table 6. 1,2-Disubstituted indoles versus 2,3-disubstituted benzo-
[b]thiophene
Kass (Â106 L/mol)8
1,2-Disubstituted 2,3-Disubstituted
Indole
(A=C; B=N)
Benzo[b]thiophene
(A=S; B=C)
Compd. X
R
In summary, we identi®ed 1,2-disubstituted indole,
azaindole and benzimidazole derivatives possessing
basic amine moieties as novel series of active site direc-
ted thrombin inhibitors with unique features versus the
benzo[b]thiophene derived thrombin inhibitors. In contrast
to the benzo[b]thiophenes, the 2,3-disubstituted indoles did
not show thrombin inhibitory activity. The preference of
1,2-disubstituted nitrogen-containing heteroaromatic
12a
H
197
126
12b
12c
12d
H
H
H
33.7
24.7
6.41
14.9
7.68
3.91

K. Takeuchi et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2347±2351
2351
compounds for thrombin inhibitory activity was in the
order: indole (12a) > azaindole (10b) > benzimidazole
(6a). These results suggest that positioning of the second
nitrogen in the heteroaromatic ring may profoundly
in¯uence and change biological activity of these novel
thrombin inhibitors. Synthetic ease of these compounds
would enable further SAR exploration.
5. Zhang, M.; Bailey, D. L.; Bastian, J. A.; Briggs, S. L.;
Chirgadze, N. Y.; Clawson, D. K.; Denney, M. L.; Gi€ord-
Moore, D. S.; Harper, R.; Johnson, L. M.; Klimkowski, V. J.;
Kohn, T. J.; Lin, H.-S.; McCowan, J. R.; Richett, M. E.; Sall,
D. J.; Smith, A. J.; Smith, G. F.; Snyder, D. W.; Takeuchi, K.;
Utterback, B. G.; Yan, S.-C. B. Bioorg. Med. Chem. Lett.
1999, 9, 775.
6. Sall, D. J.; Bailey, D. L.; Bastian, J. A.; Buben, J. A.; Chir-
gadze, N. Y.; Clemens-Smith, A. J.; Denney, M. L.; Fisher, M.
J.; Giera, D. D.; Gi€ord-Moore, D. S.; Harper, R.; Johnson, L.
M.; Klimkowski, V. J.; Kohn, T. J.; Lin, H.-S.; McCowan, J.
R.; Richett, M. E.; Smith, G. F.; Snyder, D. W.; Takeuchi, K.;
Toth, J. E.; Zhang, M. J. Med. Chem. 2000, 43, 649.
References and Notes
1. (a) Machovich, R. In The Thrombin; Machovich, R., Ed.;
CRC: Boca Raton, 1984; Vol. 1, pp 1±22. (b) Hirsh, J.; Salzman,
E. W.; Marder, V. J.; Colman, R. W. In Hemostasis and Throm-
bosis: Basic Principles and Clinical Practice, 2nd ed.; Colman,
R. W., Hirsh. J., Marder, V. J., Salzman, E. W., Eds.; Lippincott:
Philadelphia, 1987; pp 1063±1072. (c) Hirsh, J.; Salzman,
E. W. Ibid, pp 1199±1207. (d) Barnett, H. J. M. Ibid, pp 1301±
1315.
2. Sall, D. J.; Bastian, J. A.; Briggs, S. L.; Buben, J. A.; Chir-
gadze, N. Y.; Clawson, D. K.; Denney, M. L.; Gierra, D. D.;
Gi€ord-Moore, D. S.; Harper, R. W.; Hauser, K. L.; Klim-
kowski, V. J.; Kohn, T. J.; Lin, H.-S.; McCowan, J. R.; Palk-
owitz, A. D.; Smith, G. F.; Takeuchi, K.; Thrasher, K. J.;
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8. Kass represents the apparent association constant as mea-
sured by the methods of Smith, G. F.; Gi€ord-Moore, D.;
Craft, T. J.; Chirgadze, N.; Ruterbories, K. J.; Lindstrom, T.
D.; Satterwhite, J. H. In New Anticoagulants for the Cardiovas-
cular Patient; Pifarre, R., Ed.; Hanley & Belfus: Philadelphia,
1997; pp 265±300. Kass values reported here are preliminary
results with the variation in the assay in the range of Æ10%.
9. Hands, D.; Bishop, B.; Cameron, M.; Edwards, J. S.; Cot-
trell, I. F.; Wright, S. H. B. Synthesis 1996, 877.
10. Speci®c shape requirements of S1 pocket for accepting
hydrophobic groups based upon X-ray crystallographic stud-
ies have been discussed in ref 2.
11. We had examined the substituent e€ects of the benzo[b]thio-
phene nucleus on the binding interaction and discovered that the
6-hydroxy substituent improved its binding anity possibly via
hydrogen-bond interaction with the carboxy terminus of Asp189
which was exposed at the crevice of the S1 pocket.2,4
3. Bastian, J. A.; Chirgadze, N. Y.; Denney, M. L.; Gi€ord-
Moore, D. S.; Sall, D. J.; Smith, G. F.; Wikel, J. H. Bioorg.
Med. Chem. Lett. 1999, 9, 363.
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