Potent, low molecular weight thrombin receptor antagonists.

Article abstract of DOI:10.1016/S0960-894X(01)00555-8

Several benzimidazole derivatives have been identified as potent thrombin receptor (PAR-1) antagonists as represented by compound 1h, which showed an IC(50) of 33 nM.

Full text of DOI:10.1016/S0960-894X(01)00555-8

Bioorganic & Medicinal Chemistry Letters 11 (2001) 2851–2853
Potent, Low Molecular Weight Thrombin Receptor Antagonists
Samuel Chackalamannil,* Darıo Doller, Keith Eagen, Michael Czarniecki,
Ho-Sam Ahn, Carolyn J. Foster and George Boykow
Schering-Plough Research Institute, 2015 Galloping Hill Rd., Kenilworth, NJ07033, USA
Received 20 March 2001; accepted 6 August 2001
Abstract—Several benzimidazole derivatives have been identified as potent thrombin receptor (PAR-1) antagonists as represented
by compound 1h, which showed an IC50 of 33 nM. # 2001 Elsevier Science Ltd. All rights reserved.
Thrombin is a serine proteinase that plays a key role in
hemostasis and wound healing.1 In addition to its pivo-
tal role in hemostasis, thrombin also stimulates potent
proliferative and inflammatory processes in a variety of
cell types by direct cellular activation.2 Cellular actions
of thrombin are mediated by the activation of specific
cell surface receptors known as protease activated
receptors (PAR) that comprise a small family of G-
protein coupled receptors.3 It has been postulated that
thrombin binds to the cellular receptor through its
anion binding exocyte and subsequently cleaves the
peptide at Arg41-Ser42. The newly unmasked serine
amino terminus acts as a ‘tethered ligand’ binding
intramolecularly to the proximal receptor which elicits
trans-membrane signaling. The prototype of this new
class of receptors, known as protease activated receptor-
1 (PAR-1) or the thrombin receptor, is present in a vari-
ety of human cell types including platelets, fibroblasts,
endothelial cells, and cardiac myocytes.4
available antithrombotic treatment.5 Several peptide
agonists and antagonists of PAR-1 based on the amino
acid sequence of the ‘tethered ligand’ have been repor-
ted.6 Recently, there have been reports of nonpeptide
PAR-1 antagonists.7 However, these compounds, in
general, have only modest affinity for the thrombin
receptor. Herein we wish to report the identification and
structure activity relationship studies of 2-aminobenzi-
midazole derivatives that are potent thrombin receptor
antagonists. Benzimidazole derivative 1h, with an IC50
value of 33 nM, is the most potent nonpeptide thrombin
receptor antagonist reported todate.
The benzimidazole derivative 1b (Table 1) was identified
as a thrombin receptor antagonist lead through high
throughput screening. This sparked our interest in the
SAR exploration of this class of compounds. The
required 1,3-disubstituted benzimidazole derivatives
represented by structure 1 were readily prepared
according to the literature procedure8 by sequential N-1
and N-3 alkylation of 2-aminobenzimdazole (2) in
methanol or acetone using commercially available
phenacyl bromides and alkyl halides (Scheme 1). In the
first step, use of the alkylating agent as the limiting
reagent minimized dialkylation. The second alkylation
often required refluxing conditions and the product
could be easily isolated by crystallization from the
reaction mixture. IC50 determinations were carried out
on PAR-1 receptors on human platelets according to
the published procedure.9 The structure–activity rela-
tionship data are presented in Table 1.
The potential pathophysiological role of the thrombin
receptor in thrombosis, atherosclerosis, and restenosis
has been well recognized.4 As such, a thrombin receptor
antagonist may have considerable therapeutic utility in
the treatment of these disorders, especially restenosis for
which no effective treatment is currently available. Since
a thrombin receptor antagonist is specific for the cel-
lular actions of thrombin and does not interfere with the
coagulation cascade, such agents are likely to confer
added safety margin with regard to hemorrhagic side
effects which is a complicating factor in the currently
The monosubstituted benzimidazole derivative 1a
showed an IC50 of 1500 nM. Systematic variation of the
N-3 substitution pattern revealed that a lower alkyl or
*Corresponding author. Tel.: +1-908-740-3474; fax: +1-908-740-
7152; e-mail: samuel.chackalamannil@spcorp.com
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00555-8

2852
S. Chackalamannil et al. / Bioorg. Med. Chem. Lett. 11 (2001) 2851–2853
benzyl group yielded optimum activity. For example,
the N-3-butyl derivative 1i gave an IC50 value of 359 nM
whereas the N-3 methyl derivative (1b) and N-3 heptyl
derivative (1j) were less active. Compound 1c bearing 2-
(1-piperidinyl)ethyl group was somewhat less potent
(IC50=2866 nM) whereas the corresponding morpho-
line derivative 1f was quite potent (IC50=98 nM).
Compound 1g bearing N-3 benzyl substituent gave an
IC50 of 65 nM. Phenyl substituent effect on the N-3
benzyl group was also studied (1d–1e and 1h). Among
the various N-3 benzyl derivatives examined, compound
1h, bearing a p-tolyl group, showed the best activity
(IC50=33 nM).
Finally, the requirement of the 2-imino group was
established by examining a series of compounds repre-
sented by the structure 6 (Scheme 1). Compound 6 as
well as a number of its close analogues that we examined
(not shown) were uniformly inactive.
Several compounds in the benzimidazole series were
also tested in functional platelet aggregation assay
employing either the PAR-1 selective agonist, high affi-
nity thrombin receptor agonist peptide (ha-TRAP), or
thrombin to induce aggregation.10 The results for com-
pounds 1f–1i are shown in Table 3. These compounds
were effective inhibitors of ha-TRAP-induced platelet
aggregation, although the IC50 values for blocking the
aggregation were consistently higher than those
observed in the binding assay.11 The compounds also
inhibited thrombin-induced platelet aggregation. The
rank order of potency paralleled that for ha-TRAP
inhibition, but higher concentrations of the compounds
were needed to produce inhibition.
The effect of substituent at N-1 was also examined. A
representative example of the variations employed is
presented in Table 2. Presence of the 3,5-bis-tert-butyl-
4-hydroxy phenacylmethyl substitution at the N-1 ring
nitrogen was found to be essential for thrombin recep-
tor antagonist property. All the compounds represented
by the structure 8, devoid of this motif, were inactive.
The 1,3-bis-tert-butyl phenol derivative 7 per se did
not show any inherent thrombin receptor inhibition.
In conclusion, we have identified a new series of benzi-
midazole derivatives as potent thrombin receptor
antagonists. Optimal activity was seen when the two
ring nitrogen atoms of benzimidazole were substituted,
respectively, by a 3,5-bis-tert-butyl-4-hydroxy phena-
cylmethyl group and a lower alkyl or benzyl group.
Table 1. Structure–activity relationship of substituted benzimida-
zoles (1)
Table 2.
Entry
R
PAR-1
IC50 (nM)
Entry
R
PAR-1
IC50 (nM)
1a
H
1500
1f
98
1b
1c
1d
1e
Me
900
2866
734
1g
1h
1i
Benzyl
65
33
R1
R2
3-OH
2-OMe
4-Me
2-Me
4-CN
4-Br
4-OH
5-OMe
H
4-Me
H
n-C4H9
359
964
1324
1j
n-C7H15
H
2-Cl
3-Me
4-FH
4-Cl
4-Cl
Table 3. Inhibition of human platelet aggregation
Entry
PAR-1 IC50 (nM)
Platelet aggregation IC50 (nM)10
ha-TRAP
Thrombin
1f
1g
1h
1i
98
65
33
359
2000
265
575
825
>10,000
600
1500
Scheme 1. Reagents and conditions: (a) (i) BrCH2(CO)Ar (3) (1
equiv), acetone, 18 h; (ii) R-X, EtOH, reflux; (b) NaH, DMF, 3.
10,000

S. Chackalamannil et al. / Bioorg. Med. Chem. Lett. 11 (2001) 2851–2853
2853
Acknowledgements
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ziano for helpful discussions.
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