Discovery and gram-scale synthesis of BMS-593214, a potent, selective FVIIa inhibitor

Article abstract of DOI:10.1016/j.bmcl.2013.02.013

A 6-amidinotetrahydroquinoline screening hit was driven to a structurally novel, potent, and selective FVIIa inhibitor through a combination of library synthesis and rational design. An efficient gram-scale synthesis of the active enantiomer BMS-593214 was developed, which required significant optimization of the key Povarov annulation. Importantly, BMS-593214 showed antithrombotic efficacy in a rabbit arterial thrombosis model. A crystal structure of BMS-593214 bound to FVIIa highlights key contacts with Asp 189, Lys 192, and the S2 pocket.

Full text of DOI:10.1016/j.bmcl.2013.02.013

Bioorganic & Medicinal Chemistry Letters 23 (2013) 2432–2435
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery and gram-scale synthesis of BMS-593214, a potent,
selective FVIIa inhibitor
a
b
b
b
b
E. Scott Priestley a, , Indawati De Lucca , Jinglan Zhou , Jiacheng Zhou , Eddine Saiah , Robert Stanton ,
Leslie Robinson b, Joseph M. Luettgen a, Anzhi Wei a, Xiao Wen a, Robert M. Knabb a, Pancras C. Wong a,
Ruth R. Wexler a
⇑
a Bristol–Myers Squibb Pharmaceutical Research Institute, Experimental Station, PO Box 80500, Wilmington, DE 19880, USA
b DuPont Pharmaceuticals Research Laboratories, 4570 Executive Drive, Suite 400, San Diego, CA 92121, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A 6-amidinotetrahydroquinoline screening hit was driven to a structurally novel, potent, and selective
FVIIa inhibitor through a combination of library synthesis and rational design. An efficient gram-scale
synthesis of the active enantiomer BMS-593214 was developed, which required significant optimization
of the key Povarov annulation. Importantly, BMS-593214 showed antithrombotic efficacy in a rabbit arte-
rial thrombosis model. A crystal structure of BMS-593214 bound to FVIIa highlights key contacts with
Asp 189, Lys 192, and the S2 pocket.
Received 21 November 2012
Revised 30 January 2013
Accepted 1 February 2013
Available online 14 February 2013
Keywords:
FVIIa
Ó 2013 Elsevier Ltd. All rights reserved.
Coagulation factor VIIa
Inhibitor
Thrombosis
Antithrombotic
Anticoagulant
Tetrahydroquinoline
Povarov annulation
Benzamidine
Coagulation factor VIIa (FVIIa) is the key enzyme which initiates
the coagulation cascade and thrombus formation in vivo. FVIIa is
synthesized in the liver and circulates in the bloodstream. Its enzy-
matic activity is greatly enhanced by tissue factor (TF), its mem-
brane anchored glycoprotein cofactor, which is expressed on cells
surrounding the vascular epithelium and in the core of atheroscle-
rotic plaques. Upon vessel injury or plaque rupture, circulating
FVIIa binds to the exposed TF, and the resulting TF/FVIIa complex
activates its zymogen substrates FIX and FX. These enzymes to-
gether generate a local burst of thrombin, resulting in platelet acti-
vation, fibrin generation, and ultimately thrombus formation.1
Initial reports of the robust antithrombotic efficacy and limited
bleeding liability of inactivated FVIIa generated significant interest
in FVIIa inhibition as a target for antithrombotic therapy.2,3 These
initial results are supported by studies of small molecule FVIIa
inhibitors in a variety of preclinical thrombosis models, which con-
sistently show strong efficacy with low bleeding side effects.4–9
Herein, we disclose our efforts toward identification of a structur-
ally novel, potent, and selective FVIIa inhibitor suitable for evalua-
tion in a preclinical thrombosis model.
Compound 1 was identified as a modestly potent FVIIa inhibitor
(FVIIa Ki = 8.8 lM) by screening a library of amidine substituted
tetrahydroquinolines originally synthesized as thrombin inhibi-
tors. Subsequent libraries resulted in rapid potency enhancement,
as exemplified by key compounds 2 and 3. Tetrahydroquinoline 2
(FVIIa Ki = 1.8 lM), which lacks the quaternary methyl and incor-
⇑
Corresponding author. Tel.: +1 609 818 5756.
E-mail address: scott.priestley@bms.com (E.S. Priestley).
Scheme 1. Synthesis of 6-amidinotetrahydroquinolines using the Povarov reaction.
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.02.013

E. S. Priestley et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2432–2435
2433
Table 1
porates a methoxy group ortho to the phenol, is fivefold more po-
tent than 1. Addition of a bridging methylene to generate tetracy-
clic analog 3 (FVIIa Ki = 0.16 M), resulted in a 55-fold potency
increase relative to compound 1.
Enzyme inhibition data for compounds 3–6a,b
l
Compd FVII Ki, (nM) FXa Ki, (nM) Thrombin Ki, (nM) Trypsin Ki, (nM)
The initial library and subsequent inhibitors were prepared as
shown in Scheme 1, using the Povarov multicomponent tetrahy-
droquinoline synthesis10 with lanthanide triflate catalysis as dis-
closed by Kobayashi.11,12 The compounds were obtained as
mixtures of cis and trans diastereomers, which were separated by
HPLC. They were initially assayed as racemates.
3
4
5
6
160
0.62
8.1
1.9
>500
16
1500
1300
ꢀ300
35
330
ꢀ300
7.5
6900
>8000
1800
a
Compounds 3–6 are racemates; relative stereochemistry is indicated.
Ki values were averaged from two experiments. FVIIa assays were performed
b
with recombinant human enzyme, while other enzyme assays were performed with
purified human enzymes.
Recognition of potential structural overlap of 3 with a series of
biphenyl carboxylate FVIIa inhibitors13,14 led to preparation of
biphenyl carboxylate compounds 4–6 (Table 1).
The addition of the phenylcarboxylate group resulted in very
potent FVIIa inhibitors. ortho-Carboxylate 4 had subnanomolar
FVIIa potency, but very poor selectivity. Compound 6, containing
the para-carboxylate, had the most attractive potency and selectiv-
ity profile, and was also active in the prothrombin time clotting as-
Because of its excellent potency and selectivity, 6 was selected
for in vivo characterization in rabbit thrombosis and bleeding
models, requiring its preparation in gram quantities. The synthetic
route described in Scheme 1 was unsuitable for scale up, as it pro-
duced the compound in modest yield as a mixture of diastereomers
say (IC2Â = 16 l
M).15
Table 2
Optimization of the Povarov reaction for the gram scale synthesis of compound 6
Entry
R
X
Y
Conditions
Yield (trans/cis)
1
2
3
4
5
6
7
8
9
CN
CN
CN
Br
Br
Br
Br
OAc
OBn
OH
Aa
Aa
Bb
Bb
B
A
B
A
A
100 (<5:95)
98 (<5:95)
52 (<5:95)
48, Quinoline product
19 (1:2)
33 (2:7)
47 (1:1)
55 (1:2)
51 (1:4)
5-Me-1,2,4-oxadiazol-3-yl
5-Me-1,2,4-oxadiazol-3-yl
5-Me-1,2,4-oxadiazol-3-yl
5-Me-1,2,4-oxadiazol-3-y
CN
CN
CN
CN
OH
4-BnOOC-phenyl
4-BnOOC-phenyl
4-BnOOC-phenyl
4-BnOOC-phenyl
4-BnOOC-phenyl
4-BnOOC-phenyl
4-BnOOC-phenyl
OBn
OBn
OH
OBn
OAc
OH
10
11
A
B
71 (2:1)
51 (1:1)
OH
Reaction conditions: (A) preform imine with 5 mol % ZnCl2 in toluene under Dean–Stark conditions, then indene (10 equiv), In(OTf)3 (1 equiv), CH3CN, 70 °C; (B) one pot
reaction: indene (10 equiv), In(OTf)3 (1 equiv), CH3CN, rt.
a
0.3 equiv In(OTf)3, rt.
1 equiv indene, 70 °C.
b

2434
E. S. Priestley et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2432–2435
Figure 1. Crystal structure of BMS-593214 bound to FVIIa at 1.8 Å resolution.
give the desired enantiomer (+)-13 in 21% yield overall. Tetrahy-
droquinoline nitrile (+)-13 was reacted with hydroxylamine in
DMSO to give the amidoxime. Subsequent acylation and hydroge-
nation gave compound (+)-6 (BMS-593214) in 78% yield for the fi-
nal three steps.
With gram quantities of BMS-593214 in hand, it was tested in
the rabbit electric current arterial thrombosis model. It was found
to have an EC50 of 570 nM and showed no significant prolongation
of the rabbit cuticle bleeding time (see Ref. 15 for a complete ac-
count of the in vitro and in vivo profile of BMS-593214).
We also obtained a crystal structure of BMS-593214 bound to
FVIIa (Fig. 1).16 As expected, the amidine group forms a salt bridge
with the Asp 189 side chain, and the tetrahydroquinoline NH forms
a weak hydrogen bond with the catalytic Ser195. The methoxy
substituent fills the small, hydrophobic S2 pocket defined by
Thr99 and His57, while the phenol forms a hydrogen bond with
a structural water bridging the Thr98 carbonyl and Tyr94 side
Scheme 2. Gram-scale synthesis of compound 6. (a) Ac2O, DMAP, 40 °C, 88%; (b)
Br2, NaOAc, AcOH, H2O, 40 °C, 54%; (c) BnBr, KHCO3, DMF, 48%; (d) Pd(OAc)2, P(o-
tol)3, NaHCO3, DME, H2O, K2CO3, 89%; (e) 4-cyanoaniline, ZnCl2, toluene, reflux;
indene (10 equiv), In(OTf)3 (1 equiv), CH3CN, reflux over mol. sieves, chiral HPLC,
Chiralcel OG column, 3:7 ethanol/hexanes, 21%; (f) H2NOH, DMSO, 70 °C; Ac2O,
CH2Cl2; H2, 10% Pd/C, MeOH; 54%.
favoring the undesired cis isomer. We therefore sought to optimize
the chemistry for scale up as described in Table 2. In order to re-
duce the polarity of the synthetic intermediates, we investigated
4-cyanoaniline and 4-(5-methyl-1,2,4-oxadiazol-3-yl)aniline as
amidine precursors. For the aldehyde component, we varied both
the phenol protecting group (Bn, Ac, or none) and the ortho substi-
tuent (Br or 4-BnOOC-phenyl), reasoning that modifying electronic
and/or steric properties might impact the diastereoselectivity of
the reaction. Initial experiments with the ortho-bromo aldehydes
(X = Br in Table 2) gave good yields, but favored the undesired cis
product (entries 1–3). For the oxadiazole substituted aniline and
unprotected phenol (entry 4), only oxidized quinoline product
was observed. Next, the oxadiazole aniline was studied with sev-
eral biphenyl aldehydes, giving modest yields of cis–trans mix-
tures. Finally, 4-cyanoaniline was reacted with the same biphenyl
aldehydes, and the cyclization yields were significantly improved.
With an unprotected biphenyl phenol (entry 10), we obtained a
71% yield of a 2:1 mixture favoring the desired trans diastereomer.
These conditions were deemed suitable for gram-scale synthesis of
6. The yield and diastereoselectivity dropped significantly when
imine formation and cyclization were run in one pot (entry 11).
After identifying Povarov reaction conditions which provided
the desired trans diastereomer as the major annulation product,
the synthesis of compound 6 was carried out on gram-scale as
shown in Scheme 2. Vanillin, 7, was acetylated and then bromi-
nated to afford 2-bromobenzaldehyde 9. Commercially available
4-carboxyphenylboronic acid 10 was benzylated and then coupled
to 9 under Suzuki–Miyaura conditions to give biaryl aldehyde 12 in
89% yield. The optimized Povarov reaction conditions were then
employed to generate tetrahydroquinoline 13 from biaryl aldehyde
12, 4-cyanoaniline, and indene. The diastereomeric products were
separated by flash chromatography, followed by chiral HPLC to
chain. The 4-carboxyphenyl group forms
a salt bridge with
Lys192, accounting for its significant impact on potency. Addition-
ally, the crystal structure enabled assignment of the absolute ste-
reochemistry of BMS-593214. The three contiguous chiral centers
on the tetrahydroisoquinoline ring all have the (R)-configuration.
In summary, we have reported the discovery and gram-scale
synthesis of a structurally novel, highly potent, and selective FVIIa
inhibitor, BMS-593214. Importantly, this compound shows efficacy
in a rabbit model of arterial thrombosis, with no significant prolon-
gation of cuticle bleeding time. Moreover, its novel tetracyclic
structure makes it an intriguing lead FVIIa inhibitor.
Acknowledgments
The authors thank Steven Sherriff for helping to prepare the
coordinates and data for PDB deposition, Joel Renick for resynthesis
of compounds 4 and 6, and Earl Crain and Carol Watson for techni-
cal assistance with the in vivo studies.
Supplementary data
Supplementary data associated with this article can be found, in
the
online
version,
at
http://dx.doi.org/10.1016/
j.bmcl.2013.02.013.
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