Novel bicyclic lactam inhibitors of thrombin: Highly potent and selective inhibitors

Article abstract of DOI:10.1016/S0960-894X(02)00131-2

The potency and selectivity of a previous series of low molecular weight thrombin inhibitors were improved through modifications of the P1 and P3 residues. Introduction of diphenyl substituted sulfonamides in the P3 moiety led to highly efficacious compou

Full text of DOI:10.1016/S0960-894X(02)00131-2

Bioorganic & Medicinal Chemistry Letters 12 (2002) 1181–1184
Novel Bicyclic Lactam Inhibitors of Thrombin: Highly Potent
and Selective Inhibitors
Yves St-Denis,a Sophie Levesque,a,* Benoit Bachand,a Jeremy J. Edmunds,b
Lorraine Leblond,a Patrice Preville,a Micheline Tarazi,a Peter D. Winocoura
and M. Arshad Siddiquia
a
´
Shire BioChem., 275 Armand-Frappier Blvd., Laval, Quebec, Canada H7V 4A7
bPfizer Global Research and Development, Ann Arbor, MI 48105, USA
Received 16 January 2002; accepted 6 February 2002
Abstract—The potency and selectivity of a previous series of low molecular weight thrombin inhibitors were improved through
modifications of the P1 and P3 residues. Introduction of diphenyl substituted sulfonamides in the P3 moiety led to highly efficacious
compounds. By correctly selecting the combination of P1 and P3 residues, high levels of potency, selectivity and in vivo efficacy
were obtained. # 2002 Elsevier Science Ltd. All rights reserved.
The development of low molecular weight thrombin
inhibitors has been an active area of research due to the
many limitations of anticoagulants that are approved
for the treatment of thrombosis and related diseases. An
ideal inhibitor should be potent, highly selective and
bioavailable. It should also involve minimum levels of
bleeding and provide predictable levels of coagulation,
which would translate into a reduced need to monitor
patients.
levels of oral absorption (Fꢀ0.5%)2 which was a key
requirement of the program. In this paper, we report
further optimization of inhibitors using novel P3 and P1
residues (Table 1), in an attempt to optimize for
improved bioavailability.
The syntheses of the bicyclic template 63 and of P1
moieties P, Q, S, V, W and Z,1 as well as X and Y,4
(Table 1) have been published elsewhere. P3 moieties A
and B are commercially available as acids. The coupling
methods of the P3 and P1 moieties to 6 have already
been reported,1,3 except for those described below.
We have recently reported a series of bicyclic lactam
based low molecular weight thrombin inhibitors dis-
playing low nanomolar potency, high selectivity and in
vivo efficacy (1 and 2).1 Despite the attractive biological
profile of these inhibitors, they did not display high
The diphenylethylsulfonamide C, diphenylmethyl-sulfo-
namide D, and phenylethylsulfonamide E were prepared
according to Scheme 1, eqs 1, 2, and 3, respectively.
Thus, 2,2-diphenylethanol 3 was first converted to
thioacetate 4 by a Mitsunobu reaction. The thioacetate
was then hydrolyzed and oxidized in one pot using 30%
H2O2 in acetic acid. The sulfonic acid thus obtained was
converted to the sulfonyl chloride 5 using thionyl chlo-
ride in DMF (Scheme 1, eq 1). A similar approach was
attempted to prepare the diphenylmethylsulfonyl
chloride, but failed to yield the desired product. An
alternative, shorter sequence was used, based on the
method of Kloosterziel et al.5 Diphenyldiazomethane6
was mixed with bicyclic amine 6 in ether, and SO2
was bubbled through the solution to give sulfonamide
7 in good yield (Scheme 1, eq 2). The purity of the
*Corresponding author. Tel.: +1-450-978-7786; fax: +1-450-978-
7777; e-mail: slevesque@ca.shire.com
0960-894X/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(02)00131-2

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Y. St-Denis et al. / Bioorg. Med. Chem. Lett. 12 (2002) 1181–1184
Table 1. In vitro and in vivo data
Compda
P3
P1
Kithr (nM)
Selb
Rat arterial thrombosis modelc
aPTTe
% Bio
MOTd
TTf
21
A
B
B
P
P
Q
P
Q
S
V
X
Y
P
180
80
25
8
2
5
ndg
nd
nd
29Æ2
nd
85Æ10
22a
22b
23a
23b
23c
23d
23e
23f
24a
24b
24c
24d
24e
24f
24g
24h
24i
125
306
471
921
>2174
24
137
354
38
469
2000
65
38
1214
11
9
189
37
21Æ1
35Æ16
36Æ10
45Æ14
37Æ14
>60
23Æ2
63Æ8
C
C
C
C
C
C
D
D
D
D
D
D
D
D
D
D
E
35Æ2
26Æ3
35Æ4
74Æ14
19Æ1
nd
28Æ1
28Æ7
44Æ2
54Æ7
42Æ4
27Æ2
nd
214Æ25
148Æ14
150Æ19
636Æ100
51Æ1
2.2
0.7
1.5
1
35
48
9
2
5
30Æ18
nd
nd
52Æ9
46Æ17
>60
154Æ5
230Æ45
216Æ13
631Æ123
354Æ73
102Æ7
nd
Q
S
1.9
V
W
Y
Z
U
R
T
1
2
51Æ11
>60
>60
10
75
135
2
75
14
nd
nd
nd
54Æ8
nd
724Æ94
44Æ10
0.1
2.7
24j
25
nd
49Æ13
nd
25Æ1
nd
146Æ36
Q
200
aAll new target compounds were characterized by 1H NMR, reverse-phase HPLC and mass spectroscopy.
bSelectivity: Ki trypsin/Ki thrombin
.
cDose: intravenous bolus dose (0.75 mg/kg) followed by an infusion (50 mg/kg/min).
dMean occlusion time in min (control: 17 to 19 min).
eActivated partial thromboplastin time in seconds (control: 20 to 22 s).
fThrombin time in seconds (control: 40 to 45 s).
gnd, Not determined.
diphenyldiazomethane was very important for the reac-
tion to proceed in high yield.7 Finally, phenylethyl
mercaptan 8 was converted to phenylethylsulfonyl
chloride 9 by oxidation with SO2Cl2 and KNO3
(Scheme 1, eq 3).8
amidine 13 through a three-step sequence involving the
formation of the amidoxime, the hydrogenolysis of its
O-acetate 9 and the removal of the BOC group (Scheme 2,
eq 1). In eq 2, bromo-indanone 14 was converted to
cyanoalcohol 15 by treatment with CuCN, and sub-
sequent reduction of the resulting ketone to the alcohol
with NaBH4. Alcohol 15 was mesylated followed by
displacement with NaN3 to form azide 16. The nitrile
group was further transformed into the amidine,10 and
the azide was reduced to amine 17 through hydro-
genolysis. Aminohistamine 20 was prepared by the
diazotization of histamine acetate.11 Azo compound 19
We describe in Scheme 2 the preparation of P1 moieties
R, T, and U (eqs 1, 2 and 3, respectively). Protected
methylaminocyclohexyl-carboxylic acid 10 was first
converted into an amide, which was then dehydrated to
nitrile 11. The CBZ protecting group was then exchan-
ged for a BOC group, and nitrile 12 was converted into

Y. St-Denis et al. / Bioorg. Med. Chem. Lett. 12 (2002) 1181–1184
1183
Scheme 1. (a) Ph3P, DIAD, CH3COSH, THF, 82%; (b) H2O2,
CH3CO2H, 70 ꢁC, 100% (CAUTION: a shield must be used at all
times during the reaction and evaporative workup); (c) SOCl2, DMF,
50%; (d) diphenyldiazomethane, SO2, Et2O, 85%; (e) SO2Cl2, KNO3,
MeCN, quant.
Scheme 2. (a) isopropyl chloroformate, NMM, THF, À20 ꢁC, 1 h
then NH3 45 min, 100%; (b) SOCl2, NMM, DMF, 0 ꢁC, 90%; (c)
H2, Pd/C 10%, MeOH; (d) (BOC)2O, CH2Cl2, 88% (two steps);
.
(e) NH2OH HCl, Na2CO3, EtOH, Á, 13%; (f) Ac2O, H2, Pd/C 10%,
AcOH, 97%; (g) 4 N HCl in dioxane, 100%; (h) CuCN, N-methyl-
pyrrolidone, 150 ꢁC, 18 h, 59%; (i) NaBH4, MeOH, THF, À78 ꢁC (1
h), then rt (1 h), 84%; (j) SOCl2, CH2Cl2, 0 ꢁC, 50%; (k) NaN3, DMF,
60 ꢁC, 71%; (l) HCl gaz, EtOH, 0 ꢁC, 48 h, then NH3 gaz, EtOH, 18 h;
(m) H2, Pd/C 10%, MeOH, 100% (two steps); (n) NaNO2, 2.3 N HCl,
0 ꢁC; (o) N-Ac-histamine, Na2CO3, then 1 N NaOH, 30%; (p) H2,
PtO2, EtOH, 81%; (q) 6 N HCl, Á, quant.
was then hydrogenolyzed, and the acetate group was
hydrolyzed with 6 N HCl (Scheme 2, eq 3). The two
amidino-amines 13 and 17, as well as aminoimidazole-
amine 20, were coupled using the previously reported
procedure1,3 to the diphenyl-sulfonyl bicyclic template
without any protection of their basic functional groups.
Inhibition of the amidolytic activity of thrombin (Ki)
and in vivo coagulation parameters in the rat arterial
thrombosis model, such as the mean occlusion time
(MOT), the activated partial thromboplastin time
(aPTT) and the thrombin time (TT), were measured
according to an already published procedure.12 Oral
bioavailability (%F) was determined in conscious rats
by comparing the areas under the curves for TT values
versus time following administration of compounds by
oral (30 mg/kg) and iv bolus (0.15 mg/kg) routes.
improved the binding of inhibitors 22b, 23b, and 24b by
a factor of four, and also demonstrated a good efficacy
in vivo. Residue W was prepared in order to regain the
selectivity lost with V. By adding a more hydrophobic
substituent to the phenyl ring, it was hoped that
favorable interactions would be gained in the S1 subsite
of thrombin. However, even with the more bulky
moiety T (24j), the selectivity over trypsin was not
comparable to the compounds with the cyclohexyl or
piperidine residues. Other less basic aromatic sub-
stituents, such as U and Z (24g and 24h, respectively),
were not potent enough and did not exhibit a significant
increase in selectivity to be considered for in vivo
testing. However, compounds 23e, 23f, and 24f showed
a high level of selectivity. Indeed, the size of these two
residues should be comparable (if not smaller) to the
phenylamidines.
We have previously demonstrated that the replacement
of the P3 acetamides with the sulfonamides yielded
generally a 2- to 3-fold increase in potency.3 The addi-
tion of a second phenyl ring in P3 also improved the
potency by 2-fold (cf., 21 with 22a in Table 1). It is
interesting to note that the S3 subsite of thrombin is
capable of accommodating large hydrophobic groups.
The above results prompted us to explore the possibility
of preparing diphenyl substituted sulfonamides. Inhibi-
tors 23a and 24a were prepared and found to be con-
siderably potent. These two compounds not only
exhibited good binding to thrombin but also displayed
good efficacy in vivo, with a significant extension of the
mean occlusion times.
Selected potent compounds were tested for oral bio-
availability. Unfortunately, only modest levels of
absorption were observed.
We have demonstrated that the S3 subsite of thrombin
can accommodate large substituents such as the diphe-
nyl carboxamides and sulfonamides in this series of
inhibitors. By carefully choosing the combination of P1
and P3 residues attached to the central bicyclic tem-
plate, high levels of potency, selectivity and in vivo effi-
cacy could be obtained. Attempts to reduce the overall
polarity of the molecules by increasing the hydro-
phobicity of the P3 moiety and reducing the basicity of
the P1 moiety failed to render the inhibitors orally
bioavailable.
We then decided to turn our attention to the P1 moiety,
keeping both sulfonamides C and D constant. The goal
was to reduce the basicity of the guanidine of P1 moiety
P, while retaining its ability to form H-bonds with
Asp189 at the bottom of the S1 subsite of thrombin.
Substituents Q, S, V, W, and Z had already been used
in our initial exploration.1 As expected, substituent Q

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Y. St-Denis et al. / Bioorg. Med. Chem. Lett. 12 (2002) 1181–1184
4. Feng, D.-M.; Bock, M. G.; Freidinger, R. M.; Vacca, J. P.;
Dorsey, B. D. WO9631504, 1996.
Acknowledgements
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6. Prepared according to: Adamson, J. R.; Bywood, R.; East-
lick, D. T.; Gallagher, G.; Walker, D.; Wilson, E. M. J. Chem.
Soc., Perkin Trans. 1 1975, 20, 2030.
7. We found that it was very important to eliminate the
unreacted benzophenone hydrazone in the preparation of diphe-
nyldiazomethane. This could be done through crystallization of
the hydrazone at low temperature.
The authors would like to acknowledge Ms. Brigitte
Grouix and Ms. Quan Yang for providing binding
constants, Ms. Isabelle Deschenes and Ms. Chantal
Boudreau for conducting in vivo experiments, and Ms.
Lyne Marcil for her help in the preparation of this
manuscript.
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