Bioorganic & Medicinal Chemistry Letters 10 (2000) 963±966
Design, Synthesis, and In Vitro Biological Activity of
Benzimidazole Based Factor Xa Inhibitors
Zuchun (Spring) Zhao,* Damian O. Arnaiz, Brian Griedel, Steven Sakata,
Jerry L. Dallas, Marc Whitlow, Lan Trinh, Joseph Post, Amy Liang,
Michael M. Morrissey and Kenneth J. Shaw
Discovery Research, Berlex Biosciences, 15049 San Pablo Avenue, PO Box 4099, Richmond, CA 94804-0099, USA
Received 13 December 1999; accepted 22 February 2000
AbstractÐInhibitors based on the benzimidazole scaold showed subnanomolar potency against Factor Xa with 500±1000-fold
selectivity against thrombin and 50±100-fold selectivity against trypsin. The 2-substituent on the benzimidazole ring had a strong
impact on the FXa inhibitory activity. Crystallography studies suggest that the 2-substituent may have a conformational eect
favoring the extended binding conformation. # 2000 Elsevier Science Ltd. All rights reserved.
Factor Xa (FXa) is a trypsin-like serine protease that
plays a central role in the blood coagulation cascade
linking the intrinsic and extrinsic pathways to the ®nal
common pathway of coagulation. The primary role of
FXa is the proteolytic activation of thrombin (FIIa),
after combining with factor Va and calcium on a phos-
pholipid membrane to form the prothrombinase complex.
Prothrombinase catalyzes the formation of thrombin
from prothrombin. Thrombin, in turn, promotes blood
clot formation by catalyzing the formation of poly-
merizable ®brin from ®brinogen, as well as strongly
inducing platelet aggregation.1 Since direct inhibition of
thrombin has demonstrated a tendency to undesirably
prolong bleeding, the development of FXa inhibitors has
emerged as a alternative focus for the treatment and
prevention of thrombotic disorders.2 This is supported
by the recent reports of the low molecular weight FXa
inhibitors that have been found to be ecacious in animal
models of thrombosis, and oer the potential for improved
treatment of a variety of thrombotic disorders.3
anticoagulants as measured by their prothrombin based
clotting time (PT) in plasma. The poor anticoagulant
activity was thought to be due to the low aqueous solu-
bility and high lipophilicity of these inhibitors that
could lead to extensive protein binding in plasma,
therefore the central scaold was modi®ed in an attempt
to increase the hydrophilicity. In this communication,
we describe our initial series of inhibitors based on the
benzimidazole scaold with improved potency, selectiv-
ity and anticoagulant activity.
A general synthesis of the benzimidazole based FXa
inhibitors described in Table 1 is shown in Scheme 1.5
The commercially available 4-amino-3-nitrophenol 1
was reduced to the bis-aniline, acylated with the appro-
priate acid chloride and then cyclized by re¯uxing in
concentrated hydrochloric acid to aord the various 2-
substituted benzimidazole derivatives 2. BOC protection
followed by Mitsunobu6 reaction with N-t-butoxy-
carbonyl-4-hydroxyl-piperidine and then selective removal
of the benzimidazole BOC group with methanolic
ammonia aorded intermediate 4. Alkylation of 4 with
sodium hydride and 7-cyano-2-bromomethylnapthalene
5 aorded a 1:1 mixture of 5 and 6 regioisomers (6).
This regioisomeric mixture was transformed to the cor-
responding amidines by sequential treatment with HCl
and ammonia gas in ethanol with concomitant depro-
tection of the Boc group.7 The regioisomeric mixtures
were separable by reverse-phase HPLC, but were gen-
erally carried on as a mixture and treated with ethyl
acetimidoimidate and triethylamine to aord the dibasic
benzimidazole analogues 7 and 8. The regioisomers
In a previous communication we identi®ed a series of
inhibitors based on indole and carbazole scaolds.4
These studies identi®ed the 2-amidinonapthyl and 4-
iminoethylpiperidonyl groups as optimal substituents to
bind in the S1 and S4 pockets of Factor Xa, similar to
the reported studies with DX9065.3a Compounds from
these series were potent inhibitors of FXa when measured
against free FXa in vitro, however were weak functional
*Corresponding author. Tel.: +1-510-669-4457; fax: +1-510-669-4310;
e-mail: Spring_Zhao@berlex.com
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00139-6