Metabolism-based identification of a potent thrombin receptor antagonist

Article abstract of DOI:10.1021/jm061043e

The metabolism of our prototypical thrombin receptor antagonist 1, K _i = 2.7 nM, was studied and three major metabolites (2, 4, and 5) were found. The structures of the metabolites were verified independently by synthesis. Compound 4 was shown

Full text of DOI:10.1021/jm061043e

J. Med. Chem. 2007, 50, 129-138
129
Metabolism-Based Identification of a Potent Thrombin Receptor Antagonist
Martin C. Clasby,*,† Samuel Chackalamannil,† Michael Czarniecki,† Dario Doller,† Keith Eagen,† William Greenlee,†
Grace Kao,† Yan Lin,† Hsingan Tsai,† Yan Xia,† Ho-Sam Ahn,‡ Jacqueline Agans-Fantuzzi,‡ George Boykow,‡
Madhu Chintala,‡ Carolyn Foster,‡ April Smith-Torhan,‡ Kevin Alton,§ Matthew Bryant,§ Yunsheng Hsieh,§ Janice Lau,§ and
Jairam Palamanda§
Central NerVous System and CardioVascular Chemical Research, CardioVascular and Metabolic Disease Research, and Drug Metabolism and
Pharmacokinetics Research, Schering-Plough Research Institute, 2015 Galloping Hill Road, Kenilworth, New Jersey 07033
ReceiVed August 30, 2006
The metabolism of our prototypical thrombin receptor antagonist 1, Ki ) 2.7 nM, was studied and three
major metabolites (2, 4, and 5) were found. The structures of the metabolites were verified independently
by synthesis. Compound 4 was shown to be a potent antagonist of the thrombin receptor with a Ki ) 11
nM. Additionally, compound 4 showed a 3-fold improvement in potency with respect to 1 in an agonist-
induced ex-vivo platelet aggregation assay in cynomolgus monkeys after oral administration; this activity
was sustained with 60% inhibition observed at 24 h post-dose. Compound 4 was highly active in functional
assays and showed excellent oral bioavailability in rats and monkeys. Compound 4 showed a superior rat
enzyme induction profile relative to compound 1, allowing it to replace compound 1 as a development
candidate.
Scheme 1. Thrombin Receptor Antagonists Based on
(+)-Himbacine
Introduction
Thrombin is a multifunctional protease involved in hemostasis
and wound healing.1 Thrombin plays a key role in the coagula-
tion cascade by converting fibrinogen to fibrin which is then
cross-linked to form a clot by trapping aggregated platelets, red
blood cells, and other plasma particles;2 the clot is then stabilized
by factor XIIIa (fibrin-stabilizing factor), which itself is activated
by thrombin. Additionally, thrombin promotes clot formation
by upregulating its own production through activation of blood
Factors V, VIII, and XI. In addition to its important roles in
the coagulation cascade, thrombin activates a number of cell
types such as platelets, leukocytes, endothelial cells, and vascular
smooth muscle cells.3,4 Thrombin is the most potent activator
of platelets which normally has a beneficial effect on essential
clot formation under normal physiological conditions. However,
under pathophysiological conditions, thrombin-mediated platelet
activation plays a major role in arterial thrombosis.5 In addition,
cellular activation of thrombin is also known to play pro-
inflammatory and proliferative roles in vascular disorders such
as atherosclerosis and restenosis.
The cellular activity of thrombin is mediated via proteolytic
activation of specific cell surface receptors known as protease
activated receptors (PARa).6-10 There are four receptor subtypes
known, PAR-1, PAR-3, and PAR-4, which are activated by
thrombin, and PAR-2 which is activated by tryptase.11 PAR-1
(also known as the thrombin receptor) is the most important of
these receptors and is the most prevalent on human and primate
platelets; PAR-4 is also present on human platelets but is
activated only at very high thrombin concentrations and is
hypothesized to be a rescue receptor that becomes activated in
the event of a serious vascular lesion.
The thrombin receptor (PAR-1) belongs to the super family
of seven transmembrane G-protein coupled receptors. The
mechanism by which thrombin activates PAR-1 (discovered by
Coughlin’s group in 1991) is unique. Thrombin binds to PAR-1
through its exo-anion binding site. Cleavage of the extracellular
domain at Arg41-Ser42 reveals an amino terminus that then binds
intramolecularly to the receptor.12-14 Thrombin receptor activat-
ing peptides (TRAPs),15 designed to mimic the amino terminus
of the activated receptor, have been shown to elicit functional
agonist responses such as platelet aggregation.
By virtue of the cellular action of thrombin, it has been
hypothesized that a thrombin receptor antagonist may be useful
in the treatment of disorders such as arterial thrombosis,
atherosclerosis, and restenosis. Because a thrombin receptor
(PAR-1) antagonist would only target the cellular effects of
thrombin, while sparing its fibrin-generating property, such an
agent could have a significant advantage in safety with regard
to bleeding side effects over current antithrombotic therapies.16-18
* Corresponding author. Tel: 908-740-2244. Fax: 908-740-7164. E-
mail: martin.clasby@spcorp.com.
We have recently reported the discovery of a potent thrombin
receptor antagonist 1 (Sch 205831, Scheme 1)19 based on the
natural product himbacine. Compound 1 showed a Ki value of
2.7 nM against the thrombin receptor and was highly active in
several functional assays such as human platelet aggregation
inhibition assay, calcium transient assay, and thymidine incor-
† Central Nervous System and Cardiovascular Chemical Research.
‡ Cardiovascular and Metabolic Disease Research.
§ Drug Metabolism and Pharmacokinetics Research.
a Abbreviations: ADP, adenosine diphosphate; hCASMC, human coro-
nary artery smooth muscle cells; haTRAP, high-affinity thrombin receptor
activating peptide; PAR, protease activated receptors; TRAP, thrombin
receptor activating peptide.
10.1021/jm061043e CCC: $37.00 © 2007 American Chemical Society
Published on Web 12/13/2006

130 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1
Scheme 2. C-Ring Hydroxylated Derivatives of 1
Clasby et al.
Table 1. Blood Levels of Compound 1 and Its Monohydroxylated
Metabolites after Single and Multiple Oral Dosing in Rats at 300 mg/kg
in 0.4% Methylcellulose
enzyme after 10 days of dosing, and the CYP1A level was also
modestly increased (3.6-fold). The elevation of hepatic cyto-
chrome P450 (CYP) enzymes was associated with a 3-fold
increase in the level of the 8 â-OH metabolite 2 on day 10
compared with day 1. A concomitant reduction in the level of
parent between day 1 and day 10 was noted (AUC ) 35 µg·
h/mL on day 1 versus 21 µg·h/mL on day 10) which strongly
suggested an autoinduction pattern for the hepatic enzymes. At
100 mg/kg, a small induction of CYP1A (1.8-fold) and CYP2B
(2.5-fold) was noted. However, based on the plasma level of
the drug at the efficacious dose of 3 mg/kg in cynomolgus
monkey, the 100 mg/kg rat pharmacokinetic measurements gave
an exposure multiple of only 5. Therefore, the development of
1 as a thrombin receptor antagonist was suspended due to
expectation that adequate plasma exposure multiples of com-
pound 1 would not be maintained in rodents in long term
toxicological studies.
Identification of a Replacement for Compound 1. In an
effort to identify a replacement candidate for compound 1
without enzyme induction liability, we adopted a 2-fold ap-
proach. The first approach was to explore C-ring hydroxylated
derivatives (Scheme 2), including the known hydroxy metabo-
lites. We reasoned that the enzyme induction was, in part, due
to the lack of appropriate functional groups for conjugation and
clearance. There have been several instances in the history of
drug discovery research where a metabolite has served as an
improved replacement for the initial drug candidate.23-26 It was
also important that such functional groups be situated at
sterically unencumbered parts of the molecule to facilitate
conjugation. The accumulation of 8-hydroxy metabolite 2 in
rat plasma during the 10-day enzyme induction study suggested
a slow clearance rate for this metabolite, perhaps due to steric
hindrance around the hydroxyl group. A second approach was
to selectively block the readily metabolized positions of the
tricylic unit so that the molecule would undergo in vivo
oxidation at alternate sites where it might be more easily
conjugated and cleared.
Synthesis of Targets. The synthesis of the target molecules
was carried out in a fashion similar to our earlier synthesis of
tricyclic thrombin receptor antagonists,19 as shown in Scheme
3. A notable exception in this case was that a protected ketone
functionality was incorporated into the dienoic acid unit at an
early stage of the synthesis for eventual elaboration to the
hydroxy- or difluoro-substituted compounds. The other building
blocks 11 and 15 were similar to the intermediates reported
previously. Phosphonate 11 was synthesized in three steps from
2-methyl-5-hydroxypyridine 8 as shown in Scheme 3. Triflate
formation followed by Suzuki coupling gave 10 which was
converted into 11 by way of deprotonation followed by treatment
with diethyl chlorophosphate. This synthesis of 11 can be
applied to most 5-aryl 2-methylpyridines, and we have synthe-
sized a number of phosphonates using this method. Alcohol 15
compound
day
AUC (0-24 h) µg·h/mL
Cmax µg/mL
1
1
10
1
10
1
10
1
35
21
49
150
93
41
29
17
29
79
58
25
11
47
2
4
5
18
81
10
poration assay. This antagonist was highly selective for the
PAR-1 receptor, had excellent oral bioavailability in rat and
monkey models, and showed complete and sustained inhibition
of platelet aggregation after oral administration in an ex-vivo
cynomolgus monkey model.20,21 Due to its excellent profile,
compound 1 was selected for further development.
Metabolic Profile of 1. During the course of our biological
3
profiling of 1, the metabolism of H-1 was evaluated in rat
hepatocytes and cynomolgus monkeys following oral admin-
istration at 3 mg/kg in 0.4% methylcellulose. Plasma samples
were taken from the monkeys at 6 and 24 h. In both species,
one major and two minor hydroxylated metabolites were
observed by LC-MS/MS analysis along with smaller amounts
of dihydroxylated metabolites. In order to further characterize
these metabolites, milligram quantities were generated by
incubation of compound 1 with pregnenolone 16R-carbonitrile
(PCN)-induced rat liver microsomes.22 Subsequent NMR studies
revealed that the major monohydroxylated metabolite was 2 (8â-
OH-1), and the minor monohydroxylated metabolites were 4
(7R-OH-1) and 5 (7â-OH-1), the structures of which are shown
in Scheme 2. The plasma concentration of 2 in monkeys was
160 ng/mL at 24 h and exceeded that of the parent (35 ng/mL
of 1 at 24 h). The presence of high plasma levels of hydroxylated
metabolites was a concern for us because this could indicate
liver enzyme induction in addition to a prolonged plasma half-
life. A 10-day enzyme induction study in rat was undertaken at
daily oral doses of 100 mg/kg and 300 mg/kg. Fed male Sprague
Dawley rats (four rats/ group) were dosed with compound 1
daily at 300 mg/kg and 100 mg/kg in 0.4% methylcellulose for
10 consecutive days. On day one and day ten, plasma concen-
trations of 1, 2, 4, and 5 were measured at 1, 2, 4, 8, 12, and 24
h (the data is shown in Table 1). As we had seen earlier in
monkeys, the level of compound 2 exceeded that of the parent
1, and the in vivo levels of all three hydroxylated metabolites
were high. The 100 and the 300 mg/kg group also showed a
10% and 34% increase in the liver/body weight ratio, respec-
tively. Spectral cytochrome P450 (CYP) relative to the control
group was increased by 80% and 99%, respectively, at the 100
and 300 mg/kg dose. Additionally, the 300 mg/kg group was
found to have a 21-fold increase in the levels of the CYP2B

Thrombin Receptor Antagonist
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1 131
Scheme 3. Synthesis of Derivatives of 1a
a Reagents and conditions: (a) Tf2O, pyridine, 83%; (b) ArB(OH)2, K2CO3, Pd(Ph3P)4, PhMe/ EtOH/H2O, 82%; (c) LDA, (EtO)2POCl, -78 °C, 85%;
(d) DHP, TsOH, 100%; (e) BuLi, BnCOCl, THF, -78 °C; (f) TsOH, MeOH, 71%, two steps; (g) Lindlar catalyst, H2 (1 atm), THF, 93%; (h) methyl
acrylate, (Ph3P)2PdCl2, DMF, 75 °C, 71%; (i) NaOH, MeOH, THF, 99%; (j) DCC, PPY, CH2Cl2, 77%; (k) xylene, 215 °C, 48%; (l) DBU, THF, 99%; (m)
10% Pd(C), H2 (1 atm) EtOAc, 99%; (n) PtO2, H2; (1 atm), EtOAc, 99%; (o) SOCl2, PhMe, 75 °C; (p) Bu3SnH, Pd(Ph3P)4, PhMe, 0 °C, 48% two steps;
(q) 11, BuLi, THF, 0 °C then 23, 83%; (r) HCl, acetone, 99%; (s) NaBH4, EtOH, 4, 70%, 5, 15%; (t) DAST, DCE, 80 °C, 45%.
was synthesized in four steps. Protection of 12 with dihydro-
pyran gave 13, and deprotonation of the terminal alkyne and
treatment with benzyl chloroformate followed by acid hydrolysis
gave 14 which was reduced to 15 by hydrogenation over Lindlar
catalyst. The absolute chirality of 15 was derived from use of
enantiopure (R)-butynol 12.27 Dienoic acid 18 was synthesized
in two steps from 16.28,29 Heck reaction of 16 with methyl
acrylate gave 17 which was hydrolyzed to give 18. DCC-
mediated ester formation of 18 with 15 in the presence of
4-pyrrolidinopyridine gave 19 which was heated at 185 °C to
give the intramolecular Diels-Alder (IMDA) adduct 20. The
absolute chirality of the stereogenic center of the IMDA
precursor 19 is diastereospecifically transmitted to C3a of the
tricyclic product 20 during Diels-Alder reaction by virtue of
an allylic strain-induced facial selectivity of the dienophile in
the IMDA transition state.19 The remainder of the stereogenic
centers in the middle ring of the IMDA adduct 20 can be
envisioned as formed relative to C3a. For example, the relative
stereochemistry between C-3a and C-9a is a function of the exo-
selective transition state of the IMDA. The relative stereochem-
istry between C-3a and C-4 is imparted due to the cis-geometry
of the dienophile. Epimerization of C9a in 20 with DBU
produces the cis-lactone. At this point the tricyclic portion of
the molecule was assembled and required only minor manipula-
tion. The benzyl protecting group was removed by hydrogenoly-
sis using 10% palladium on carbon as the catalyst, and
diastereoselective hydrogenation of the internal double bond
using Adams catalyst generates the C-8a stereogenecity in the
required absolute and relative chirality. The key aldehyde 23
was synthesized in two steps from 22 via reduction of its acid
chloride with tributyltin hydride in the presence of tetrakistri-
phenylphosphine palladium(0).30 The phosphonate 11 was then
coupled to the aldehyde 23 using Horner-Emmons conditions
to give the ketal 24. Eventual deprotection of the ketal and
stereoselective reduction of the resulting ketone would produce
the required C7 alcohol. The ketal moiety was removed under
acidic hydrolysis conditions and the resulting ketone 25 reduced
with sodium borohydride to give the 7-hydroxy metabolites 4
and 5 which were readily separable using standard silica gel
chromatography. Additionally ketone 25 was converted to the

132 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1
Scheme 4. Synthesis of 6- and 8-Substituted Targetsa
Clasby et al.
a Reagents and conditions: (a) methyl acrylate, (Ph3P)2PdCl2, DMF, 75 °C, 48%; (b) NaBH4, EtOH, 2, 27%, 3, 55%; (c) DAST, DCE, 80 °C; (d) methyl
acrylate, (Ph3P)2PdCl2, DMF, 75 °C, 89%; (e) NaBH4, EtOH.
Table 2. Binding Data of Derivatives of 1
7,7-difluoro derivative 26 via treatment with diethylaminosulfur
trifluoride (DAST) (Scheme 3).
compound
IC50 (nM) ( SEMa
Similarly ketal protected precursors 2731 and 31 would
eventually lead to the C-8 hydroxy derivatives 2 and 3 and the
C-6 hydroxy compounds 6 and 7, respectively (Scheme 4). The
8,8-difluoro compound 30 was synthesized in a manner similar
to that of compound 25. Comparison of the synthetic samples
of 2, 4, and 5 with samples obtained from the in Vitro
metabolism studies gave independent confirmation of the
structures of the three monohydroxylated metabolites.
A systematic study of the heteroaryl region of the monhy-
droxylated and gem-difluroinated thrombin receptor antagonists
was also carried out. Toward this goal, Horner-Emmons
reactions using appropriately substituted heteroaryl phosphonates
were carried out on tricyclic aldehyde 23.
2
3
4
5
6
7
26
30
41 ( 3.8 (n ) 3)
28 ( 3 (n ) 4)
17 ( 1.3 (n ) 8)
23 ( 2.5 (n ) 4)
481 ( 107 (n ) 4)
845 ( 227 (n ) 4)
37 ( 13 (n ) 2)
53 ( 15 (n ) 5)
a PAR-1 binding assay ligand: [3H]haTRAP, 10 nM (Kd ) 15 nM).
the C-6 hydroxy compounds 6 and 7 were essentially inactive.
Among the gem-difluorinated analogues tested the 8,8-difluoro
derivative 30 had similar PAR-1 binding to the C-8 hydroxylated
compounds (2 and 3); however, the 7,7-difluoro analogue 26
showed marginally more favorable PAR-1 binding.
An SAR study of the phenyl substitution in the C-7 hydroxy
series and the 7,7-difluoro series confirmed our previous finding
that ortho- and meta-substituted phenyl groups are favored over
the para-substituted phenyl ring and that meta was similar to
ortho. Selected SAR is shown in Table 3. Replacement of the
3-trifluoromethyl group in 4 with either chlorine (37) or fluorine
(34) led to a slight loss in potency (IC50 ) 38 and 28 nM,
respectively), whereas replacement with a methyl group (38)
led to a substantial loss in potency (147 nM). In the 7,7-
difluorinated series the chlorinated derivatives 45 and 46 proved
to be the most potent (IC50 ) 12, 11 nM, respectively).
Results and Discussion
The hydroxyl and difluoro derivatives of 1 were screened in
the in Vitro binding assay using purified human platelet
membranes as a PAR-1 source and [3H]haTRAP as the ligand
as previously described.32 The PAR-1 binding results for the
hydroxylated and fluorinated derivatives of 1 are given in Table
2. Among the hydroxylated derivatives, the C-7 hydroxy
compound 4 was the most potent (IC50 ) 17 nM). The C-8
hydroxylated compounds (2 and 3) and the C-7 hydroxy
compound 5 showed slightly lower PAR-1 binding affinity, and

Thrombin Receptor Antagonist
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1 133
Table 3. SAR of the Heteroaryl Region of Compounds 4 and 26
compound
Ar
IC50 (nM), n ) 2 ( SEM
34
35
36
37
38
39
40
41
42
43
44
45
46
47
3-fluorophenyl
2-fluorophenyl
4-fluorophenyl
3-chlorophenyl
3-methylphenyl
2-methylphenyl
4-methylphenyl
2,5-dichlorophenyl
2,3-dichlorophenyl
3-fluorophenyl
2-fluorophenyl
3-chlorophenyl
2-chlorophenyl
3-methylphenyl
28 ( 2
72 ( 28
829 ( 430
38.5 ( 8.5
147 ( 5
70 ( 17
1374 ( 23
97 ( 1.5
16.5 ( 1.5
15 ( 5
Figure 2. Blood level of 4 at various time points after single dose of
4 (1 mg/ kg, 20% HPBCD), in cynomolgus monkeys.
complete inhibition only at the 3 mg/kg dose. Interestingly, we
noted that 5 underwent extensive metabolic transformation to
4 and the corresponding ketone 25 in ViVo in cynomolgus
monkeys, presumably an oxidation/reduction cycle via the
ketone 25. On the other hand, compound 4 was only minimally
metabolized to 5 and 25 under the same experimental conditions
in cynomolgus monkeys. The ketone 25 was equipotent to
alcohols 4 and 5 in the cynomolgus monkey ex ViVo model.
Therefore, it is apparent that compound 4 is metabolically more
stable in primates than its epimer 5. However, this profile was
found to be reversed in rats. When the alcohol 4 was dosed in
rats, it underwent extensive metabolic conversion to 5 as
measured by the plasma levels, whereas similar studies using 5
showed little conversion to 4. Thus, the metabolic interconver-
sion of 4 and 5 follows symmetrically opposite pathways in
primates and rodents. This species-dependent metabolism was
a concern for us; in particular we needed to know how these
compounds would be metabolized in humans. In order determine
this, we carried out a comparative study of compounds 4 and 5
in rat, monkey, and human hepatocytes. Upon incubation of 4
and 5 in rat and monkey hepatocytes, we obtained similar results
to those seen in ViVo; in cynomolgus monkeys, 4 was the
preferred epimer while in rats 5 was the preferred epimer. A
similar human hepatocyte incubation study also was undertaken
to assess the potential for conversion of 4 to 5. Upon incubation
of compounds 4 and 5 in human hepatocytes, we obtained results
similar to the monkey hepatocytes with 4 undergoing very little
conversion to 5, and 5 undergoing substantial interconversion
to 4. This correlation between the human and cynomolgus
monkey metabolism gave us further assurance to the validity
of using a cynomolgus monkey ex-vivo platelet aggregation
assay as the pharmacodynamic model.
Profile of 4. Since compound 4 showed the best character-
istics in terms of PAR-1 affinity, ex-vivo potency, and phar-
macokinetics, it was selected for further studies. It binds
competitively to PAR-1 on human platelet membranes with a
Ki of 11 nM against [3H]haTRAP. Scatchard plots of saturation
binding of [3H]haTRAP in the presence and absence of 4 as
consistent with a competitive inhibition of the radioligand
binding to PAR-1. Compound 4 appears to dissociate very
slowly from the receptor; compound 4 was preincubated with
platelet membranes, and once saturation binding was achieved,
the excess compound was washed out. Inhibition of binding of
[3H]haTRAP was observed for 3 h after the washout, thus
indicating slow dissociation of compound 4 from the receptor.
Slow dissociation from the receptor is thought to be critical in
determining antagonist efficacy.16,18,19
28 ( 9
12 ( 2.8
11 ( 0.5
80 ( 20
a PAR-1 binding assay ligand: [3H]haTRAP, 10 nM (Kd ) 15 nM).
Figure 1. Ex-vivo platelet aggregation response after single dose (1
mg/kg, 20% HPBCD) of 4.
In order to evaluate the in ViVo activity of the difluorinated
derivatives, representative compounds were selected for test-
ing in the ex-vivo platelet aggregation inhibition assay in
cynomolgus monkeys. Both compounds 26 and 45 were active
only at a dose of 3 mg/kg in 20% HPâCD for 6 h (compared
to 24 h for compound 1). The 8,8-difluoro compound 30, not
surprisingly, was inactive when dosed. In general this series of
compounds showed only moderate plasma levels in the rat
(typically AUC(0-6) ) 1200-1500 ng·h/mL @ 10 mg/kg in 20%
HPâCD). This is presumably due to poor oral absorption due
to the low solubility of these compounds.33
From the in vitro data it can be seen that the 7-R-hydroxy
metabolite 4 has potency (IC50 )17 nM) similar to that of the
parent compound 1 which had an IC50 of 11 nM. In the
pharmacokinetic studies conducted in rat, metabolites (2, 4, and
5) showed high plasma levels after oral administration (AUC(0-6)
) 5755, 6130, and 3148 ng·h/mL, respectively). In the ex-vivo
platelet aggregation assay in cynomolgus monkeys, compounds
4 and 5 were the most potent and showed complete inhibition
of haTRAP-induced platelet aggregation at 1 mg/kg for 6 h,
whereas compound 4 still showed 60% inhibition at the 24 h
time point (Figure 1), additionally compound 4 showed blood
levels that were consistent with this data (Figure 2). This is a
3-fold improvement over the parent compound 1, which showed

134 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1
Clasby et al.
Compound 4 has been tested for its ability to inhibit thrombin-
mediated activation of calcium transients in human coronary
smooth muscle cell (hCASMC). Intracellular calcium concentra-
tion was measured using a fluorescent calcium dye (Fluo-3)
and a fluoremetric imaging plate reader. Thrombin elevated the
intracellular calcium concentration in a dose-dependent manner
with an EC50 of 0.9 nM. Compound 4 completely inhibited this
effect in a dose-dependent manner with a Ki ) 85 nM. When
the smooth muscle cells were treated with 4, no elevation of
calcium levels was observed, indicating that 4 does not possess
agonist activity.
PAR-1 activation is known to be mitogenic in smooth muscle
cells.16,18 Arterial smooth muscle cell proliferation is a key event
in the formation of arterial lesions and restenosis that often
occurs following percutaneous coronary artery interventions
(PCI). The potential utility of a PAR-1 antagonist to inhibit these
processes can be assessed in Vitro by measuring thrombin-
induced incorporation of thymidine in hCASMC. Compound 4
completely inhibited the thrombin-stimulated [3H]thymidine
incorporation with an apparent Ki of 22 nM. This data suggests
that 4 should inhibit arterial smooth muscle cell proliferation
and therefore has potential utility as an antirestinosis agent in
addition to its promising antiplatelet effects.
To characterize the antiplatelet effects of antagonist 4, it was
preincubated for 1 h with washed human platelets obtained from
healthy subjects who had been aspirin-free for 7 days. The
preincubation is required because, in addition to a slow
disassociation-rate, 4 has a slow association rate. Antagonist 4
completely inhibited the aggregation response of the washed
platelets to 0.3 µM of haTRAP with an IC50 of 60 ( 10 nM.
Additionally, 4 does not block the aggregation response of
platelets to ADP, indicating that this activity is specific to
inhibition of PAR-1. As mentioned previously, 4 showed
complete inhibition of platelet aggregation in the ex-vivo model
in cynomolgus monkeys for up to 6 h with 60% inhibition
observed at 24 h. This was a 3-fold improvement in potency in
activity in this model over 1 when dosed similarly.
induction. A synthetic method was developed that allowed for
the incorporation of 6-, 7-, and 8-hydroxy substitution as well
as difluoro substitution. In addition we developed a method to
allow variation in the biaryl moiety via palladium(0) chemistry.
The 7-hydroxy metabolite 4 is highly potent with a Ki of 11
nM and showed 3-fold greater oral poptency in a cynomolgus
monkey ex-vivo platelet aggregation inhibition model than its
prototype 1. In a 10-day multiple dosing rat enzyme induction
model, compound 4 and its active metabolites showed excellent
exposure multiples based on the plasma levels in cynomolgus
monkey at the efficacious dose of 1 mg/kg. Additionally, these
compounds showed a slow dissociation rate from PAR-1
receptor which is beneficial in competing against the tethered
ligand mechanism.16 Compound 4 did not affect clotting
parameters, confirming that its mode of mechanism is not by
active site inhibition of thrombin.
Experimental Section
General Comments. Melting points were taken on a Thomas-
Hoover or Mel-Temp II melting point apparatus and are uncorrected.
Chromatography was performed over Universal Scientific or Selecto
Scientific flash silica gel (particle size 32-63 µm). 1H NMR spectra
were determined on a Gemini 400 MHz instrument using either
Me4Si or residual solvent signal as internal standards. Rotations
were determined on a Rudolph Autopol III or Perkin-Elmer 243B
Polarimeter with concentration expressed in milligrams per 1 mL.
Mass spectra were obtained on VG-ZAB-SE, Extrel-401, HP-MS
Engine, JEOL HX-110, Sciex API 100, or Sciex API 150 mass
spectrometer. Elemental analyses were determined by the Physical-
Analytical Department of Schering-Plough Research Institute using
either CEC 240-HA, CEC CE-440, or Fisons EA 1108 CHNS
elemental analyzers and are within 0.4% of the theoretical value
unless otherwise noted. The conditions for the LC/MS analyses in
the preparations and examples below are as follows: 5 min gradient
from 10% f 95% CH3CN/H2O with 0.05% TFA, then 2 min
isocratic at 95% CH3CN/H2O with 0.05% TFA, 1.0 mL/min flow
rate on a MAC-MOD ACE5 C18 column (4.6 × 50 mm). (R)-
Butynol (ee > 97%) was purchased from DSM Fine Chemicals,
217 Rt. 46W, Saddle Brook, NJ 07663.
Compound 4 shows good fasted oral absorption (92%) and
bioavailability (89%) in cynomolgus monkeys. The bioavail-
ability was not affected by the presence of food. The major
metabolite in cynomolgus monkeys was the glucoronide, which
was primarily excreted in bile. Compound 4 did not inhibit
cytochrome P450’s 2D6, 3A4, 2C9, or 2C19 in human liver
microsomes at concentrations up to 10 µM. Enzyme induction
studies of 4 in rats over 8 days showed a weak to moderate
amount of enzyme induction of cytocrome P450s 1A and 2B
at daily doses of 10 and 30 mg/kg. At both 10 and 30 mg/kg
dose the change in body weight relative to control was not
significant. We observed improved multiples for the parent and
active metabolites in cynomolgus monkeys. This multiple was
21-fold at the 10 mg/kg dose and 36-fold at the 10 mg/kg.
Additionally, contrary to compound 1, no progressive reduction
of the plasma level of the parent was noted at the 30 mg/kg
dose (21.7 µg·h/mL on day 1 vs 21.3 µg·h/mL on day 8). This
study indicates that it would be feasible to conduct a chronic
drug safety study in rats. Other studies34 have shown that 4 did
not induce changes in gastrointestinal, renal, cardiovascular,
respiratory, or central nervous system function.
Trifluoromethanesulfonic Acid 6-Methylpyridin-3-yl Ester
(9). Triflic anhydride (46 mL, 0.275 mol) was added dropwise to
a stirred solution of 6-methylpyridin-3-ol 8 (10 g, 0.092 mol) in
pyridine (200 mL) at 0 °C and stirred at 0 °C to room temperature
for 16 h. The mixture was poured into ice-water (300 mL) and
extracted with ether. The ether layer was washed with water (2 ×
150 mL) and brine, dried (MgSO4), and concentrated in Vacuo to
give 9 (18.7 g, 83%) as a brown oil. 1H NMR (400 MHz, CDCl3)
δ 2.67 (s, 3H), 7.32 (d, 1H, J ) 8.5 Hz), 7.57 (dd, 1H, J ) 8.6, 2.8
Hz), 8.53 (d, 1H, J ) 2.8 Hz); MS (ESI) m/z 242 (MH+, 100%).
2-Methyl-5-(3-trifluoromethylphenyl)pyridine (10). To a solu-
tion of pyridine 9 (8.5 g, 34.5 mmol) and 3-trifluoromethylphenyl-
boronic acid (10 g, 55 mmol) in toluene (100 mL) were added EtOH
(25 mL), K2CO3 (14.3 g, 104 mmol) in H2O (50 mL), and Pd(PPh3)4
(400 mg, 0.345 mmol). The mixture was heated in a closed pressure
tube under argon at 120 °C for 16 h. The mixture was diluted with
EtOAc, washed with 5% NaOH and brine, dried (MgSO4), and
concentrated in Vacuo. Flash chromatography of the residue on a
silica gel column with EtOAc-hexane (10:90 then 20:80) as eluent
gave 10 (6.7 g, 82%) as yellow solids. 1H NMR (400 MHz, CDCl3)
δ 2.68 (s, 3H), 7.32 (d, 1H, J ) 8 Hz), 7.62-7.90 (m, 5H), 8.79
(d, 1H, J ) 2 Hz).
[5-(3-Trifluoromethylphenyl)pyridin-2-ylmethyl]phosphon-
ic Acid Diethyl Ester (11). Compound 10 (4 g, 0.0146 mol) and
diisopropylamine (2.28 mL, 1.1 equiv) were dissolved in THF (73
mL) and cooled to -78 °C with stirring. n-Butyllithium (12.28 mL
of a 2.5 M solution in hexanes, 2.1 equiv) was added dropwise,
and after 20 min diethyl chlorophosphate (2.11 mL, 1 equiv) was
added. After a further 20 min, the mixture was allowed to warm to
rt. Ammonium chloride solution (saturated) was added and the
Summary
In summary, we have used a metabolism-based approach to
identify compound 4 as an excellent replacement candidate for
the prototypical thrombin receptor antagonist 1 which encoun-
tered development issues due to rat specific CYP-2B enzyme

Thrombin Receptor Antagonist
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1 135
mixture extracted with ethyl acetate. The organic extracts were dried
(magnesium sulfate), concentrated, and chromatographed (SiO2, 1:1
hexane/ethyl acetate to 100% ethyl acetate) to give 11 (3.9 g, 71%)
as a tan oil. 1H NMR (400 MHz, CDCl3) δ 1.36 (t, 6H, J ) 7 Hz),
3.56 (d, 2H, J ) 22 Hz), 4.19 (dq, 4H, J ) 7, 7 Hz), 7.58-7.96
(m, 6H), 8.84 (d, 1H, J ) 2 Hz); MS (FAB) m/z 374 (MH+, 100%);
(2R)-2-(1-Methylprop-2-ynyloxy)tetrahydropyran (13). (2R)-
3-butynol 12 (15 mL, 0.204 mol) and 3,4-dihydro-2H-pyran (26.1
mL, 1 equiv) were stirred at 0 °C. To this mixture was added
p-toluenesulfonic acid (monohydrate) (0.38 g, 5 mol %) and the
mixture stirred for a further 2 h. EtOAc (319 mL) and NaHCO3
(1.6 g) were added, and after another 1 h the mixture was filtered
and concentrated. Chromatography (SiO2, 19:1 hexane/EtOAc) gave
0 °C for 2 h and at rt for 1 h. The mixture was then filtered and
concentrated in Vacuo, and column chromatography (9:1 to 4:1
hexane/EtOAc) gave 27 g of 19. Compound 19 was dissolved in
xylene (300 mL) and heated at 215 °C for 7 h. Column chroma-
tography (9:1 to 4:1 to 2:1 hexane/EtOAc) gave 13.2 g of 20.
Compound 20 was dissolved in THF (264 mL), and DBU (4.9 mL,
0.033 mol) was added. The mixture was stirred for 1 h, diluted
with EtOAc (500 mL), washed with NH4Cl(sat.), dried (MgSO4),
concentrated in Vacuo, filtered through a pad (1 in.) of SiO2 (eluting
with EtOAc), and concentrated in Vacuo to give 21 (13 g, 48%
1
from 18). H NMR (CDCl3) δ 1.10 (d, J ) 6.0 Hz, 3H), 1.2 (m,
1H), 1.65-1.85 (m, 2H), 1.92 (m, 1H), 2.35 (m, 1H), 2.47 (m, 1H),
2.59 (dd, J ) 10.75, 4.0 Hz, 1H), 2.70 (m, 1H), (q, J ) 2.5 Hz,
1H), 3.85-4.0 (m, 5H), 4.45 (m, 1H), 5.15 (AB quartet, J ) 12.0,
10.5 Hz, 2H), 5.36 (br s, 1H), 7.35 (s, 5H).
1
31.49 g (100%) of 13 as a mixture of diastereomers. H NMR
(major diastereomer) (400 MHz, CDCl3) δ 1.54 (d, J ) 7.5 Hz,
3H), 1.55-2.0 (m, 6H), 2.42 (s, 1H), 3.56 (m, 1H), 3.88 (m, 1H),
4.60 (br q, J ) 7.5 Hz, 1H), 5.00 (t, J ) 5.0 Hz, 1H).
(1′R,3′aR,4′aR,8′aR,9′S,9′aS)-Decahydro-1′-methyl-3′-oxo-
spiro[1,3-dioxolane-2,6′(3′H)-naphtho[2,3-c]furan]-9′-carbox-
ylic Acid (22). Ester 21 (4.92 g, 0.0123 mol) was dissolved in
EtOAc (250 mL), 10% palladium on carbon (492 mg) was added,
and the mixture was stirred under 1 atm of H2(g) for 1 h. The
mixture was filtered through a pad of celite. PtO2 (492 mg) was
added to the filtrate and the mixture stirred for 16 h under 1 atm
H2(g). The mixture was then filtered and concentrated in Vacuo to
(2R)-4-Hydroxypent-2-ynoic Acid Benzyl Ester (14). A solu-
tion of compound 13 (31.49 g, 0.204 mol) in THF (1 L) was cooled
to -78 °C with stirring. n-BuLi (97.8 mL, 2.5 M, 1.2 equiv) was
added dropwise. After stirring for 20 min, benzyl chloroformate
(35.1 mL, 1.2 equiv) was added and the reaction mixture was stirred
at -78 °C for 2 h. The mixture was allowed to warm to room
temperature, a saturated solution of NH4Cl was added, and the
mixture was extracted with EtOAc. The organic extracts were dried
over anhydrous MgSO4, concentrated under reduced pressure, and
then dissolved in methanol (2 L). DOWEX 50WX8-100 ion-
exchange resin (60 g, prewashed with MeOH) was added, and the
mixture was stirred at room temperature overnight. The mixture
was filtered, concentrated, and chromatographed (SiO2, 9:1 to 4:1
1
give 3.81 g (99%) of 22. H NMR (CDCl3) δ 1.25 (m, 2H), 1.35
(d, J ) 6.5 Hz, 3H), 1.3-1.5 (m, 3H), 1.6 (m, 1H), 1.7-1.95 (m,
3H), 2.5 (m, 1H), 2.58 (m, 1H), 2.68 (m, 1H), 3.95 (m, 5H), 4.69
(m, 1H).
(1′R,3′aR,4′aR,8′aR,9’S,9′aS)-Decahydro-1′-methyl-3′-oxo-
spiro[1,3-dioxolane-2,6′(3′H)-naphtho[2,3-c]furan]-9′-carbox-
aldehyde (23). Acid 22 (1 g, 0.0032 mol) was dissolved in toluene
(20 mL), thionyl chloride (1.25 mL) was added, and the mixture
was heated at 80 °C for 16 h. The mixture was then concentrated
in Vacuo, dissolved in fresh toluene (16 mL) and cooled to 0 °C.
Pd(Ph3P)4 (186 mg) was added, followed by tributyltinhydride (1.3
mL, 0.0048 mol). The mixture was stirred for 3 h, chromatographed
(4:1 to 2.5:1 hexane/EtOAc) to give 450 mg (48%) of 23. 1H NMR
(CDCl3) δ 1.24 (d, J ) 6.5 Hz, 3H), 1.0-1.9 (m, 10H), 2.48 (m,
1H), 2.6-2.7 (m, 2H), 3.87 (m, 4H), 4.54 (m, 1H), 9.70 (br s, 1H).
(1′R,3a′R,4a′R,8a′R,9′S,9a′S)-Decahydro-1′-methyl-9′-[(E)-2-
[5-[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]-spiro[1,3-di-
oxolane-2,6′(3′H)-naphtho[2,3-c]furan]-3′-one (24). Phosphonate
11 (1.14 g, 0.0030 mol) was dissolved in THF (10 mL) and cooled
to 0 °C. n-BuLi (1.9 mL of 2.5 M solution in hexanes, 0.0029 mol)
was added and the mixture stirred for 10 min. This solution was
then added to a solution of aldehyde 23 (450 mg, 0.00153 mol) in
THF (10 mL) at 0 °C. This mixture was stirred for 2 h and then
NH4Cl (sat.) was added. The mixture was extracted (EtOAc), dried
(MgSO4), concentrated in Vacuo, and then chromatographed (60/
40 hexane/EtOAc) to give 650 mg (83%) of 24. 1H NMR (CDCl3)
δ 1.12-1.55 (m, 6H), 1.43 (d, J ) 6 Hz, 3H), 1.78 (m, 2H), 1.79
(m, 1H), 1.96 (dd, J ) 6.5, 3.0 Hz, 1H), 2.9 (m, 2H), 2.70 (quintet,
J ) 6. 5 Hz, 1H), 3.95 (m, 4H), 4.76 (m, 1H), 6.55 (d, J ) 15 5
Hz, 1H), 6.65 (m, 1H), 7.29 (d, J ) 8.0 Hz, 1H), 7.60 (t, J ) 7.5
Hz, 1H), 7.66 (d, J ) 7.5 Hz, 1H), 7.75 (d, J ) 7.5 Hz, 1H), 7.80
(s, 1H), 7.86 (d, J ) 8.0 Hz, 1H), 8.79 (s, 1H).
1
hexane/EtOAc) to give 29.9 g (71%) of 14. H NMR (400 MHz,
CDCl3) δ 1.55 (d, J ) 7.5 Hz, 3H), 4.70 (q, J ) 7.5 Hz, 1H), 5.27
(s, 2H), 7.44 (br s, 5H).
(2R)-4-Hydroxypent-(Z)-2-enoic Acid Benzyl Ester (15). Com-
pound 14 (23.28 g, 0.114 mol) was dissolved in THF (232 mL).
Lindlar’s hydrogenation catalyst was added (3.48 g). The mixture
was then placed under 1 atm pressure of H2(g) and stirred for 2.5
h. The mixture was filtered and concentrated in Vacuo to give 15
1
(22 g, 93%). H NMR (CDCl3) δ 1.32 (d, J ) 6.5 Hz, 3H), 5.09
(m, 1H), 5.17 (s, 2H), 5.86 (d, J ) 11.7 Hz, 1H), 6.30 (dd, J )
11.7, 7.0 Hz, 1H), 7.38 (s, 5H).
3-(1,4-Dioxaspiro[4.5]dec-7-en-7-yl)acrylic Acid Ethyl Ester
(17). 7-Bromo-1,4-dioxaspiro[4.5]dec-7-ene 1628,29 (27.5 g, 0.1255
mol) was dissolved in DMF (400 mL), and methylacrylate (23 mL,
0.251 mol), triethylamine (52.25 mL, 0.3765 mol), and Pd(Ph3P)2Cl2
(4.37 g, 5 mol %) were added successively. The mixture was heated
at 75 °C for 16 h. The reaction was worked up by the addition of
NH4Cl (sat.), extracted with ether, and dried (MgSO4). The extracts
were concentrated in Vacuo, and the residue was chromatographed
1
(9:1 to 4:1 hexane/EtOAc) to give 20 g (71%) of 17. H NMR
(CDCl3) δ 1.78 (t, J ) 6.5 Hz, 2H), 2.38 (s, 2H), 2.44 (m, 2H),
3.74 (s, 3H), 4.0 (s, 4H), 5.73 (d, J ) 15 Hz, 1H), 6.17 (br s, 1H),
7.36 (d, J ) 15 Hz, 1H).
3-(1,4-Dioxaspiro[4.5]dec-7-en-7-yl)acrylic Acid (18). Com-
pound 17 (20 g, 0.089 mol) was dissolved in a 1:1 mixture of THF/
MeOH (520 mL total). 1 M NaOH solution (260 mL) was added
slowly. The mixture was stirred for 4 h then water was added. The
mixture was washed with ether, the aqueous layer was then acidified
to pH 1 and extracted with EtOAc (×3), the combined extracts
were dried (MgSO4), and the solution was concentrated in Vacuo
to give 19 g (99%) of 18. 1H NMR (CDCl3) δ 1.79 (t, J ) 6.5 Hz,
2H), 2.40 (s, 2H), 2.46 (m, 2H), 4.01 (m, 4H), 5.73 (d, J ) 15.7
Hz, 1H), 6.23 (s, 1H), 7.41 (d, J ) 15.7 Hz, 1H).
(1′R,3′a R,8′aS,9’S,9′aR)-1′,3′a,5′,7′,8′,8′a,9′,9′a-Octahydro-1′-
methyl-3′-oxo-spiro[1,3-dioxolane-2,6′(3′H)-naphtho[2,3-c]furan]-
9′-carboxylic Acid Phenylmethyl Ester (21). Acid 18 (18 g,
0.0856 mol) was dissolved in CH2Cl2 (350 mL) and cooled to 0
°C. 1,3-Dicyclohexylcarbodiimide (23.23 g, 0.112 mol) was added,
followed by 4-pyrrolidinopyridine (1.39 g, 0.0094 mol). After 5
min of stirring, a solution of 15 (22 g, 0.1067 mol) in CH2Cl2 (127
mL) was added over a 10 min period. The mixture was stirred at
(3R,3aS,4S,4aR,8aR,9aR)-Octahydro-3-methyl-4-[(E)-2-[5-
[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]naphtho[2,3-c]-
furan-1,7(3H,4H)-dione (25). Compound 24 (650 mg, 0.00126
mol) was dissolved in acetone (7.5 mL), and HCl (7.5 mL of a 1
M solution) was added. The mixture was heated at 50 °C for 16 h.
NaHCO3(sat.) was added and the mixture extracted with EtOAc.
The combined extracts were dried (MgSO4), concentrated in Vacuo,
and chromatographed (1:1 hexane/EtOAc) to give 590 mg (99%)
1
of 25. H NMR (CDCl3) δ 1.2-1.5 (m, 2H), 1.47 (d, J ) 7.0 Hz,
3H), 1.65 (m, 2H), 2.08 (m, 2H), 2.10 (m, 2H), 2.3-2.5 (m, 4 H),
2.74 (quintet, J ) 6.5 Hz, 1H), 4.80 (m, 1H), 6.59 (d, J ) 6.5 Hz,
1H), 6.72 (m, 1H), 7.28 (d, J ) 8.0 Hz, 1H), 7.61 (t, J ) 7.5 Hz,
1H), 7.66 (d, J ) 7.5 Hz, 1H), 7.76 (d, J ) 7.5 Hz, 1H), 7.81 (s,
1H), 7.87 (d, J ) 8.0 Hz, 1H), 8.80 (s, 1H); MS (CI) m/z 470
20
(MH+, 100%); [R]D -15.7 (c 7.13 mg/mL, MeOH); Anal.
(C27H26F3NO3•HCl) C, H, N.

136 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1
Clasby et al.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]naph-
tho[2,3-c]furan-1(3H)-one (4) and (3R,3aS,4S,4aR,7S,8aR,9aR)-
Decahydro-7-hydroxy-3-methyl-4-[(E)-2-[5-[3-(trifluoromethyl)phenyl]-
2-pyridinyl]ethenyl]naphtho[2,3-c]furan-1(3H)-one (5). Compound
25 (100 mg, 0.000213 mol) was dissolved in EtOH (8 mL), and
NaBH4 (30 mg) was added. After 5 min, NaHCO3 (sat.) was added
and the mixture extracted with EtOAc. The extracts were dried
(MgSO4) and concentrated in Vacuo. Purification by preparative
TLC (47.5:47.5:5 hexane/EtOAc/MeOH) gave two isomers. The
least polar isomer 5 (15 mg, 15%): 1H NMR (CDCl3) δ 1.15-1.4
(m, 4H), 1.43 (d, J ) 6.0 Hz, 3H), 1.5-1.7 (m, 3H), 1.75-1.95 (m,
3H), 2.35-2.5 (m, 2H), 2.72 (quintet, J ) 6.6 Hz, 1H), 4.16 (br s,
1H), 4.75 (m, 1H), 5.46, J ) 15.5 Hz, 1H), 6.65 (m, 1H), 7.29 (d,
J ) 8.0 Hz, 1H), 7.60 (t, J ) 8 Hz, 1H), 7.66 (d, J ) 7.5 Hz, 1H),
7.76 (d, J ) 7.5 Hz, 1H), 7.80 (s, 1H), 7.85 (d, J ) 8.0 Hz, 1H),
8.79 (s, 1H); MS (CI) m/z 472 (MH+, 100%); Anal. (C27H28F3NO3·
0.3H2O) C, H, N. The most polar isomer 4 (70 mg, 70%): 1H NMR
(CDCl3) δ 0.93 (m, 1H), 1.06-1.4 (m, 5H), 1.43 (d, J ) 6.0 Hz,
3H), 1.6 (m, 1H), 1.85-2.05 (m, 4H), 2.40 (m, 2H), 2.70 (quintet,
J ) 6.5 Hz, 1H), 3.64 (m, 1H), 4.75 (m, 1H), 6.55 (d, J ) 15.5
Hz, 1H), 6.64 (m, 1H), 7.29 (d, J ) 8.0 Hz, 1H), 7.60 (t, J ) 7.75
Hz, 1H), 7.65 (d, J ) 7.5 Hz, 1H), 7.75 (d, J ) 7.5 Hz, 1H), 7.80
(s, 1H), 7.85 (d, J ) 8.0 Hz, 1H), 8.79 (s, 1H); MS (CI) m/z 472
8.85 (s, 1H), LCMS (MH+ ) 472.3) purity ) 95%, and 550 mg of
1
3 (55%), H NMR (CDCl3) δ 0.82-0.95 (m, 1H), 1.24-1.40 (m,
3H), 1.48 (d, J ) 6 Hz, 3H), 1.70-1.95 (m, 6H), 2.38-2.48 (m,
2H), 2.76 (q, J ) 6.6 Hz, 1H), 3.95 (br s, 1H), 4.85 (m, 1H), 6.58-
6.72 (m, 2H), 7.36 (d, J ) 8.3 Hz, 1H), 7.65-7.75 (m, 2H), 7.82
(d, J ) 7.6 Hz, 1H), 7.87 (s, 1H), 7.90 (d, J ) 8.5 Hz, 1H), 8.85
(s, 1H), LCMS (MH+ ) 472.3) purity ) 100%.
Trifluoromethanesulfonic Acid 1,4-Dioxa-spiro[4.5]dec-7-en-
8-yl Ester (31). To a solution of 1,4-cyclohexanedione mono-
ethylene ketal (10 g, 64 mmol) and 2,6-di-tert-butyl-4-methyl-
pyridine (21 g, 102 mmol) in CH2Cl2 (350 mL) at room temperature
was added triflic anhydride (16 mL, 96 mmol) and stirred for 16
h. The mixture was washed with NaHCO3 (sat.). The organic layer
was dried (MgSO4) and concentrated in Vacuo. Flash chromatog-
raphy of the residue on a silica gel column with EtOAc-hexane
(5-95 then 10-90) as eluent gave 31 (13.4 g, 72%) as a clear oil.
1H NMR (CDCl3) δ 1.96 (t, J ) 6.6 Hz, 2H), 2.37 (m, 2H), 2.59
(m, 2H), 4.05 (m, 4H), 5.72 (m, 1H).
3-(1,4-Dioxa-spiro[4.5]dec-7-en-8-yl)acrylic Acid Methyl Ester
(32). To a solution of 31 (13 g, 46 mmol) in DMF (150 mL) were
added methyl acrylate (8.4 mL, 92 mmol), Et3N (19 mL, 138
mmol), and Pd(PPh3)2Cl2 (1.62 g, 2.3 mmol). The mixture was
stirred at 75 °C for 10 h. The mixture was diluted with NH4Cl
(sat.) and extracted with ether. The organic layer was washed with
brine, dried (MgSO4), and concentrated in Vacuo. Flash chroma-
tography of the residue on a silica gel column with EtOAc-hexane
20
(MH+, 100%); [R]D +18.6 (c 3.38 mg/mL, MeOH); Anal.
(C27H28F3NO3·HCl·0.1H2Ã) C, H, N.
1
(3R,3aS,4S,4aR,8aR,9aR)-7,7-Difluoro-decahydro-3-methyl-
4-[(E)-2-[5-[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]naph-
tho[2,3-c]furan-1(3H)-one (26). Compound 25 (0.63 g, 0.00134
mol) was dissolved in dichloroethane (6 mL), Diethylaminosulfur
trifluoride (DAST) (0.383 mL, 0.0029 mol, 2.16 equiv) was added
and the mixture stirred at 80 °C for 1 h. Saturated NaHCO3 solution
was added and the mixture extracted with EtOAc. The organic layer
was dried (MgSO4) and concentrated and the residue purified by
silica gel chromatography (1:4 EtOAc in hexanes) to give 300 mg
(15-85) as eluent gave 32 (9.15 g, 89%) as a clear oil. H NMR
(CDCl3) δ 1.90 (t, J ) 6.5 Hz, 2H), 2.44 (m, 2H), 2.51 (m, 2H),
3.80 (s, 3H), 4.05 (s, 4H), 5.85 (d, J ) 15 Hz, 1H), 6.11 (br s,
1H), 7.36 (d, J ) 15 Hz, 1H).
(-)-(3R,3aS,4S,4aR,6R,8aS,9aR)-Decahydro-6-hydroxy-3-meth-
yl-4-[(E)-2-[5-[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]-
1
naphtho[2,3-c]f uran-1(3H)-one (6). H NMR (CDCl3) δ 0.88-
2.78 (m, 13H), 1.48 (d, J ) 6.0 Hz, 3H), 3.67 (m, 1H), 4.78 (m,
1H), 6.64 (m, 2H), 7.33 (d, J ) 8.0 Hz, 1H), 7.64-7.92 (m, 5H),
1
20
8.85 (s, 1H); MS (FAB) m/z 472 (MH+, 100%); [R]D -11.1 (c
of compound 26 (45%). H NMR (CDCl3) δ 1.16-1.77 (m, 6H),
1.44 (d, J ) 6.0 Hz, 3H), 1.89-2.19 (m, 4H), 2.35-2.47 (m, 2H),
2.73 (q, J ) 6.8 Hz, 1H), 4.71-4.77 (m, 1H), 6.53-6.68 (m, 2H),
7.25-7.29 (m, 1H), 7.57-7.68 (m, 2H), 7.75 (d, J ) 7.6 Hz, 1H),
7.80 (s, 1H), 7.84-7.88 (m, 1H), 8.80 (m, 1H), MS (CI) m/z 492
4.13 mg/ mL, MeOH); Anal. (C27H28F3NO3·HCl·0.6CH2Cl2) C, H,
N. (calcd 59.31, 5.45, 2.51; found 59.03, 5.72, 2.68).
(+)-(3R,3aS,4S,4aR,6S,8aS,9aR)-Decahydro-6-hydroxy-3-meth-
yl-4-[(E)-2-[5-[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]-
naphtho[2,3-c]furan-1(3H)-one (7). 1H NMR (CDCl3) δ 1.08-2.78
(m, 13H), 1.48 (d, J ) 6.0 Hz, 3H), 4.21 (m, 1H), 4.85 (m, 1H),
6.63 (m, 2H), 7.37 (d, J ) 8.0 Hz, 1H), 7.64-7.92 (m, 5H), 8.83 (s,
1H); MS (FAB) m/z 472 (MH+, 100%); [R]D20 +41.2 (c 4.05 mg/
mL, MeOH); Anal. (C27H28F3NO3·HCl·0.75CH2Cl2) C, H, N. (calcd
58.30, 5.38, 2.45; found 58.23, 5.80, 2.45).
(3R,3aS,4S,4aS,8aS,9aR)-8,8-Difluoro-decahydro-3-methyl-4-
[(E)-2-[ 5-[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]naph-
tho[2,3-c]furan-1(3H)-one (30). 1H NMR (CDCl3) δ 0.84-0.94 (m,
1H), 1.44 (d, J ) 6.8 Hz, 3H), 1.46-1.81 (m, 6H), 1.86-1.93 (m,
1H), 2.14-2.25 (m, 1H), 2.31-2.40 (m, 2H), 2.43-2.49 (m, 1H), 2.70
(q, J ) 6.0 Hz, 1H), 4.72-4.79 (m, 1H), 6.55-6.66 (m, 2H), 7.29
(d, J ) 8 Hz, 1H), 7.59-7.68 (m, 2H), 7.76 (d, J ) 8 Hz, 1H), 7.81
(s, 1H), 7.83-7.89 (m, 1H), 8.80 (s, 1H), MS (CI) m/z 492, Anal.
(C27H26F5NO2·HCl·1.25H2O) C, H, N.
20
(MH+, 100%); [R]D +26.6 (c 6.05 mg/mL, MeOH); Anal.
(C27H26F5NO2·HCl·1.2H2O) C, H, N.
3-(1,4-Dioxaspiro[4.5]dec-6-en-6-yl)acrylic Acid Methyl Ester
(28). Compound 2731 (33 g, 0.151 mol) was dissolved in DMF
(400 mL), and methyl acrylate (28 mL, 0.31 mol, 2 equiv),
triethylamine (65.2 mL, 0.468 mol, 3 equiv), and Pd(Ph3P)2Cl2 (5.5
g, 5 mol %) were added successively. The mixture was heated at
80 °C for 16 h. The reaction was worked up by the addition of
NH4Cl (sat), extracted with ether, and dried (MgSO4). The extracts
were concentrated in Vacuo and the residue chromatographed (15%
EtOAc in hexanes) to give 16 g of 28 (48%). 1H NMR (CDCl3) δ
1.72-1.82 (m, 4H), 2.18-2.27 (m, 2H), 3.73 (s, 3H), 4.03-4.12 (m,
4H), 6.06 (d, J ) 15.2 Hz, 1H), 6.45 (m, 1H), 7.26 (d, J ) 15.2
Hz, 1H).
(3R,3aS,4S,4aS,8S,8aS,9aR)-Decahydro-8-hydroxy-3-methyl-
4-[(E)-2-[5-[3-(trifluoromethyl)phenyl]-2-pyridinyl]ethenyl]-
naphtho[2,3-c]furan-1(3H)-one (2) and (3R,3aS,4S,4aS,8R,
8aS,9aR)-Decahydro-8-hydroxy-3-methyl-4-[(E)-2-[5-[3-(trifluoro-
methyl)phenyl]-2-pyridinyl]ethenyl]naphtho[2,3-c]furan-1(3H)-one
(3). Compound 29 (1 g, 0.0021 mol) was dissolved in 1:1 MeOH/
THF (160 mL), and NaBH4 (161 mg, 0.0042 mol, 2 equiv) was
added. After 10 min, NH4Cl(sat.) was added and the mixture
extracted with EtOAc. The extracts were dried (MgSO4) and
concentrated in Vacuo. Purification by silica gel chromatography
(40% EtOAc/hexanes) gave the following in order of elution: 270
mg of 2 (27%), 1H NMR (CDCl3) δ 0.85-0.95 (m, 1H), 1.10-1.28
(m, 2H), 1.30-1.45 (m, 3H), 1.50 (d, J ) 6 Hz, 3H), 1.80-1.90 (m,
2H), 2.04-2.12 (m, 1H), 2.41-2.45 (m, 1H), 2.47-2.53 (m, 1H), 2.61-
2.66 (m, 1H), 2.72-2.77 (m, 1H), 3.35 (br m, 1H), 4.80 (m, 1H),
6.58-6.72 (m, 2H), 7.34 (d, J ) 8 Hz, 1H), 7.65-7.75 (m, 2H),
7.82 (d, J ) 7.6 Hz, 1H), 7.87 (s, 1H), 7.90 (d, J ) 8 Hz, 1H),
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(3-fluorophenyl)-2-pyridinyl]ethenyl]naphtho[2,3-c]-
furan-1(3H)-one (34). 1H NMR (CD3OD) δ 0.95 (m, 1H), 1.0-1.6
(m, 6H), 1.38 (d, J ) 6.0 Hz, 3H), 1.75-2.0 (m, 3H), 2.40 (m, 2H),
2.73 (quintet, J ) 6.25 Hz, 1H), 3.56 (m, 1H), 4.75 (m, 1H), 6.62
(m, 2H), 7.1-7.2 (m, 1H), 7.44 (d, J ) 13.2 Hz, 1H), 7.48-7.52 (m,
2H), 7.56 (d, J ) 8.4 Hz, 1H), 8.04 (dd, J) 8.4, 2.4 Hz, 1H), 8.74
20
(d, J ) 2.4 Hz, 1H); MS (CI) m/z 422 (MH+, 100%); [R]D +31
(c 4.36 mg/mL, MeOH); Anal. (C26H28FNO3·HCl) C, H, N.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(2-fluorophenyl)-2-pyridinyl]ethenyl]naphtho[2,3-c]-
1
furan-1(3H)-one. (35). H NMR (CDCl3) δ 0.97 (m, 1H), 1.14-
1.41 (m, 5H), 1.49 (d, J ) 6.0 Hz, 3H), 1.69 (m, 1H), 1.9-2.1 (m,
4H), 2.40 (m, 2H), 2.75 (quintet, J ) 6.4 Hz, 1H), 3.7 (m, 1H),
4.80 (m, 1H), 6.55-6.70 (m, 2H), 7.22-7.33 (m, 3H), 7.40-7.46 (m,
1H), 7.50 (t, J ) 7.7 Hz, 1H), 7.89 (d, J) 8.0 Hz, 1H), 8.79 (s,

Thrombin Receptor Antagonist
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1 137
20
1H); MS (CI) m/z 422 (MH+, 100%); [R]D +33.4 (c 1.95 mg/
(3R,3aS,4S,4aR,8aR,9aR)-7,7-Difluoro-decahydro-3-methyl-
4-[(E)-2-[ 5-(3-fluorophenyl)-2-pyridinyl]ethenyl]naphtho[2,3-
c]furan-1(3H)-one (43). H NMR (CDCl3) δ 1.16-1.77 (m, 6H),
mL, MeOH); Anal. (C26H28FNO3·H2O) C, H, N.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(4-fluorophenyl)-2-pyridinyl]ethenyl]naphtho[2,3-c]-
1
1.44 (d, J ) 6.0 Hz, 3H), 1.89-2.19 (m, 4H), 2.37-2.46 (m, 2H),
2.73 (q, J ) 6.4 Hz, 1H), 4.74 (m, 1H), 6.54-6.65 (m, 2H), 7.07-
7.12 (m, 1H), 7.24-7.28 (m, 2H), 7.34-7.38 (m, 1H), 7.41-7.47 (m,
1H), 7.82 (dd, J ) 8, 2 Hz, 1H), 8.76 (m, 1H), MS (CI) m/z 442
1
furan-1(3H)-one (36). H NMR (CD3OD) δ 0.97 (m, 1H), 1.11
(m, 1H), 1.2-1.4 (m, 4H), 1.37 (d, J ) 6.4 Hz, 3H), 1.79 (m, 1H),
1.87 (dd, J ) 11.2, 4.8, 1H), 1.93 (br d, J ) 12 Hz, 2H), 2.44-2.48
(m, 1H), 2.45-2.49 (m, 1H), 2.77 (quintet, J ) 6.0 Hz, 1H), 3.59
(m, 1H), 4.80 (m, 1H), 6.80 (d, J ) 16 Hz, 1H), 7.13 (dd, J ) 16,
10.4 Hz, 1H), 7.33 (t, J ) 8.8 Hz, 2H), 7.85 (m, 2H), 8.32 (d, J )
8.8 Hz, 1H), 8.77 (dd, J ) 8.8, 1.6 Hz, 1H), 8.95 (s, 1H); MS (CI)
m/z 422 (MH+, 100%); [R]D20 +30.6 (c 3.7 mg/mL, MeOH); Anal.
(C26H28FNO3·HCl) C, H, N.
20
(MH+, 100%); [R]D +25.1 (c 5.2 mg/mL, MeOH); Anal.
(C26H26F3NO2·HCl·1.3H2O) C, H, N.
(3R,3aS,4S,4aR,8aR,9aR)-7,7-Difluoro-decahydro-3-methyl-
4-[(E)-2-[ 5-(2-fluorophenyl)-2-pyridinyl] ethenyl]naphtho[2,3-
1
c]furan-1(3H)-one (44). H NMR (CDCl3) δ 1.17-1.75 (m, 6H),
1.45 (d, J ) 6.0 Hz, 3H), 1.88-2.18 (m, 4H), 2.38-2.46 (m, 2H),
2.73 (q, J ) 6.8 Hz, 1H), 4.74 (m, 1H), 6.54-6.65 (m, 2H), 7.16-
7.28 (m, 3H), 7.34-7.40 (m, 2H), 7.84 (d, J ) 8.4 Hz, 1H), 8.674
(m, 1H), MS (CI) m/z 442 (MH+, 100%); [R]D20 +24.4 (c 4.5 mg/
mL, MeOH); Anal. (C26H26F3NO2·HCl·1.3H2O) C, H, N.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(3-chlorophenyl)-2-pyridinyl]ethenyl]naphtho[2,3-c]-
furan-1(3H)-one (37). 1H NMR (CDCl3) δ 0.9-1.03 (m, 2H), 1.1-
1.4 (m, 4H), 1.49 (d, J ) 6.4 Hz, 3H), 1.7 (br s, 1H), 1.8-2.1 (m,
4H), 2.43 (m, 2H), 2.75 (quintet, J ) 6.4 Hz, 1H), 3.7 (m, 1H),
4.81 (m, 1H), 6.57-6.7 (m, 2H), 7.32 (d, J ) 8 Hz, 1H), 7.41-7.53
(m, 3H), 7.61 (s, 1H), 7.88 (dd, J ) 8.1, 2.2 Hz, 1H), 8.8 (s, 1H);
MS (CI) m/z 438 (MH+, 100%); Anal. (C26H28ClNO3·1.5H2O) C,
H, N.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(3-methylphenyl)-2-pyridinyl]ethenyl]naphtho[2,3-
c]furan-1(3H)-one (38). H NMR (CDCl3) δ 0.92 (m, 1H), 1.05-
1.35 (m, 5H), 1.45 (d, J ) 6.2 Hz, 3H), 1.54 (m, 1H), 1.88-2.1 (m,
4H), 2.3-2.4 (m, 2H), 2.43 (s, 3H), 2.70 (quintet, J ) 6.7 Hz, 1H),
3.65 (m, 1H), 4.75 (m, 1H), 6.55 (m, 2H), 7.2-7.26 (m, 2H), 7.36-
7.38 (m, 3H), 7.82 (dd, J ) 8.2, 2.3 Hz, 1H), 8.77 (d, J ) 2 Hz,
1H); MS (CI) m/z 418 (MH+, 100%); [R]D20 +28.6 (c 4.8 mg/mL,
MeOH); Anal. (C27H31NO3·H2O) C, H, N.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(2-methylphenyl)-2-pyridinyl]ethenyl]naphtho[2,3-
c]furan-1(3H)-one (39). H NMR (CD3OD) δ 0.92 (m, 1H), 1.1
(m, 1H), 1.15-1.37 (m, 4H), 1.39 (d, J ) 6 Hz, 3H), 1.82-1.86 (m,
2H), 1.93-1.96 (m, 2H), 2.26 (s, 3H), 2.37-2.4 (m, 2H), 2.70
(quintet, J ) 6.4 Hz, 1H), 3.55 (m, 1H), 4.85 (m, 1H), 6.6 (m,
2H), 7.2-7.31 (m, 4H), 7.53 (d, J ) 8 Hz, 1H), 7.74 (dd, J) 8, 2
Hz, 1H), 8.42 (s, 1H); MS (CI) m/z 418 (MH+, 100%); [R]D20 +27.6
(c 3.94 mg/ mL, MeOH); Anal. (C27H31NO3·HCl) C, H, N.
(3R,3aS,4S,4aR,8aR,9aR)-7,7-Difluoro-decahydro-3-methyl-
4-[(E)-2-[ 5-(3-chlorophenyl)-2-pyridinyl] ethenyl]naphtho[2,3-
1
c]furan-1(3H)-one (45). H NMR (CDCl3) δ 1.16-1.76 (m, 6H),
1.44 (d, J ) 6.0 Hz, 3H), 1.88-2.19 (m, 4H), 2.37-2.45 (m, 2H),
2.73 (q, J ) 6.8 Hz, 1H), 4.74 (m, 1H), 6.53-6.65 (m, 2H), 7.25
(d, J ) 7.2 Hz, 1H), 7.35-7.46 (m, 4H), 7.55 (m, 1H), 7.81 (dd, J
) 8, 2.8 Hz, 1H), MS (CI) m/z 458 (MH+, 100%); [R]D20 +23.6 (c
4.2 mg/mL, MeOH); Anal. (C26H26ClF2NO2·HCl·1.5H2O) C, H, N.
(3R,3aS,4S,4aR,8aR,9aR)-7,7-Difluoro-decahydro-3-methyl-
4-[(E)-2-[ 5-(2-chlorophenyl)-2-pyridinyl] ethenyl]naphtho[2,3-
1
1
c]furan-1(3H)-one (46). H NMR (CDCl3) δ 1.17-1.72 (m, 6H),
1.46 (d, J ) 6.0 Hz, 3H), 1.90-2.19 (m, 4H), 2.38-2.46 (m, 2H),
2.73 (q, J ) 6.4 Hz, 1H), 4.75 (m, 1H), 6.55-6.65 (m, 2H), 7.26
(m, 1H), 7.34 (m, 3H), 7.49-7.51 (m, 1H), 7.77 (dd, J ) 8, 2 Hz,
1H), 8.63 (d, J ) 2.4 Hz, 1H), MS (CI) m/z 458 (MH+, 100%);
20
[R]D +18.6 (c 4.8 mg/mL, MeOH); Anal. (C26H26ClF2NO2·HCl·
1.2H2O) C, H, N.
1
(3R,3aS,4S,4aR,8aR,9aR)-7,7-Difluoro-decahydro-3-methyl-
4-[(E)-2-[ 5-(3-methylphenyl)-2-pyridinyl] ethenyl]naphtho[2,3-
1
c]furan-1(3H)-one (47). H NMR (CDCl3) δ 1.0-1.76 (m, 6H),
1.47 (d, J ) 5.6 Hz, 3H), 1.93-2.19 (m, 4H), 2.30 (s, 3H), 2.40-
2.45 (m, 2H), 2.73 (q, J ) 6.8 Hz, 1H), 4.75 (m, 1H), 6.55-6.61
(m, 2H), 7.20-7.32 (m, 5H), 7.61-7.64 (m, 1H), 8.54 (d, J ) 1.2
Hz, 1H), MS (CI) m/z 438 (MH+, 100%); Anal. (C27H29F2NO2·
HCl·1.6H2O) C, H, N.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(4-methylphenyl)-2-pyridinyl]ethenyl]naphtho[2,3-
c]furan-1(3H)-one (40). H NMR (CDCl3) δ 0.97 (m, 1H), 1.11
1
Acknowledgment. The authors thank Drs. Catherine Strader,
John Piwinski, Michael Graziano, Ashit Ganguly, John Hunter,
Ronald White, and Ted Sybertz for helpful discussions, Dr. Jesse
Wong for scale-up of 22, Drs. Birendra Pramanik and Pradip
Das for mass spectral data, and Drs. T.-M. Chan and Mohindar
Puar for NMR data.
(m, 1H), 1.22-1.4 (m, 4H), 1.37 (d, J ) 6 Hz, 3H), 1.75-1.98 (m,
4H), 2.42 (s, 3H), 2.45-2.49 (m, 1H), 2.57 (m, 1H), 2.76 (quintet,
J ) 6 Hz, 1H), 3.58 (m, 1H), 4.91 (m, 1H), 6.81 (d, J ) 16 Hz,
1H), 7.11 (dd, J ) 16, 9.2 Hz, 1H), 7.4 (m, 2H), 7.7 (m, 2H), 8.29
(d, J ) 8.4 Hz, 1H), 8.76 (d, J) 8.4 Hz, 1H), 8.93 (s, 1H); MS
20
(CI) m/z 418 (MH+, 100%); [R]D +35 (c 3.15 mg/mL, MeOH);
Anal. (C27H31NO3·HCl·0.2H2O) C, H, N.
Supporting Information Available: Microanalytical data. This
material is available free of charge via the Internet at http://
pubs.acs.org.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(2,5-dichlorophenyl)-2-pyridinyl]ethenyl]naphtho-
[2,3-c]furan-1(3H)-one (41). H NMR (CDCl3) δ 0.91 (m, 1H),
1
References
1.06-1.36 (m, 5H), 1.44 (d, J ) 5.6 Hz, 3H), 1.86-2.04 (m, 4H),
2.32-2.41 (m, 2H), 2.70 (quintet, J ) 6.4 Hz, 1H), 3.65 (m, 1H),
4.75 (m, 1H), 6.54 (d, J ) 15.2 Hz, 1H), 6.63 (dd, J ) 15.2, 9.6
Hz, 1H), 7.25-7.33 (m, 3H), 7.43 (d, J ) 8.4 Hz, 1H), 7.73 (dd,
J) 8, 2 Hz, 1H), 8.6 (m, 1H); MS (CI) m/z 472 (MH+, 100%);
(1) Coleman, R. W.; Marder, V. J.; Salzman, E. W.; Hirsh, J. Overview
of hemostasis. In Hemostasis and Thrombosis: Basic Principles and
Clinical Practice; Coleman, R. W., Hirsch, J., Marder, V. J., Salzman,
E. W., Eds.; Lippincott, Williams and Wilkins: Philadelphia, PA,
1994; pp 3-17.
(2) Davie, E. W.; Fujikawa, K.; Kisiel, W. The coagulation cascade:
initiation, maintenance, and regulation. Biochemistry 1991, 30,
10363-10370.
(3) De Caterina, R.; Sicari, R. Cellular effects of thrombin: Pharmacol-
ogy of the receptor(s) in various cell types and possible development
of receptor antagonists. Pharmacol. Res. 1993, 27, 1-19.
(4) Shuman, M. A. Thrombin-cellular interactions. Ann. N.Y. Acad. Sci.
1986, 485, 228-239.
(5) Harker, L. A. Pathogenesis of thrombosis. In Hematology; Williams,
W. J., Beutler, E., Erslev, A. J., Lictman, M. A., Eds.; McGraw
Hill: New York, 1990; pp 1559-1568.
(6) Coughlin, S. R. Protease-activated Receptors. In Handbook of Cell
Signaling; Bradshaw, R. A.; Dennis, E. A., Eds.; Elsevier: San Diego,
2004; Vol. 1, pp 167-171.
20
[R]D +3.7 (c 13.9 mg/ mL, MeOH); Anal. (C26H27Cl2NO3·HCl·
0.3H2O) C, H, N.
(3R,3aS,4S,4aR,7R,8aR,9aR)-Decahydro-7-hydroxy-3-methyl-
4-[(E)-2-[5-(2,3-dichlorophenyl)-2-pyridinyl]ethenyl]naphtho-
[2,3-c]furan-1(3H)-one (42). H NMR (CDCl3) δ 0.89 (m, 1H),
1.04-1.34 (m, 5H), 1.43 (d, J ) 6 Hz, 3H), 1.85-2.02 (m, 4H),
2.30-2.39 (m, 2H), 2.68 (quintet, J ) 6.8 Hz, 1H), 3.63 (m, 1H),
4.74 (m, 1H), 6.53 (d, J ) 15.2 Hz, 1H), 6.60 (dd, J ) 15.2, 8.8
Hz, 1H), 7.2-7.29 (m, 3H), 7.49 (d, J ) 8 Hz, 1H), 7.71 (dd, J)
8.4, 2.4 Hz, 1H), 8.57 (m, 1H); MS (CI) m/z 472 (MH+, 100%);
[R]D20 +21.6 (c 2.62 mg/ mL, MeOH); Anal. (C26H27Cl2NO3·HCl·
0.5H2O) C, H, N.
1

138 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 1
Clasby et al.
(7) Coughlin, S. R. Protease-Activated Receptors in the Cardiovascular
System. Cold Spring Harbor Symp. Quant. Biol. 2002, 67, 197-
208.
(8) Coughlin, S. R. Protease-Activated Receptors in Vascular Biology.
Thromb. Haemostasis 2001, 86, 298-307.
(9) Grand, R. J. A.; Turnell, A. S.; Grabham, P. W. Cellular consequences
of thrombin-receptor activation. Biochem. J. 1996, 313, 353-368.
(10) Coughlin, S. R. Protease-activated receptors in hemostasis, thrombosis
and vascular biology. J. Thromb. Haemostasis 2005, 3, 1800-1814.
(11) Coughlin, S. R. How the protease thrombin talks to cells. Proc. Natl.
Acad. Sc. U.S.A. 1999, 96, 11023-11027.
A.; Scarborough, R. M.; Maryanoff, B. E. Discovery and Optimiza-
tion of A Novel Series of Thrombin Receptor (PAR-1) Antagonists:
Potent, Selective Peptide Mimetics Based on Indole and Indazole
Templates. J. Med. Chem. 2001, 44, 1021-1024.
(22) Kim, H.; Ahn, H.; Bryant, M.; Nomeir, A.; Chan, T.-M. Isolation
and identification of the circulating major metabolites of the thrombin
receptor antagonist SCH 205831 from rat liver microsomal incuba-
tions and rat bile using HPLC, LC/MS and LC/NMR. Manuscript in
preparation.
(23) McClellan, K.; Jarvis, B. Desloratadine. Drugs 2001, 61, 789-
796.
(12) Vu, T-K. H.; Hung, D. T.; Wheaton, V. I.; Coughlin, S.R. Molecular
cloning of a functional thrombin receptor reveals a novel proteolytic
mechanism of receptor activation. Cell 1991, 64, 1057-1068.
(13) Hung, D. T.; Vu, T.-H.; Nelken, N. A.; Coughlin, S. R. Thrombin-
induced events in non-platelet cells are mediated by the unique
proteolytic mechanism established for the cloned platelet thrombin
receptor. J. Cell Biol. 1992, 116, 827-832.
(24) Markham, A.; Wagstaff, A. J. Fexofenadine. Drugs 1998, 55, 269-
274.
(25) Singh, S. S. Preclinical pharmacokinetics: an approach towards safer
and efficacious drugs. Curr. Drug Metab. 2006, 7, 165-182.
(26) Nassar A-E.; F.; Kamel, A. M.; Clairmont, C. Improving the decision-
making process in the structural modification of drug candidates:
enhancing metabolic stability. Drug DiscoVery Today 2004, 9, 1020-
1028.
(27) (R)-Butynol was purchased from DSM Fine Chemicals, 217 Rt. 46W.,
Saddle Brook, NJ 07663. This material had an enantiomeric excess
greater than 97%. As described previously, due to the stereocontrol
of the intramolecular Diels-Alder reaction,19 we obtained compounds
that were enantiopure (>99%) by HPLC (CHIRALPAK AD,
CHIRALCEL OD).
(28) Lambert, J. B.; Liu, X. The â -effect of silicon in the orthogonal
geometry. J. Organomet. Chem. 1996, 521, 203-210.
(29) Shih, C.; Swenton, J. S. Use of protected â-bromocyclopentenones
and â-bromocyclohexenones as â-acylvinyl anion equivalents. J. Org.
Chem. 1982, 47, 2825-2832.
(30) Four, P.; Guibe, F. Palladium-catalyzed reaction of tributyltin hydride
with acyl chlorides. A mild, selective, and general route to aldehydes.
J. Org. Chem. 1981, 46, 4439-4445.
(14) Vu, T.-K. H.; Wheaton, V. I.; Hung, D. T.; Charo, I.; Coughlin, S.
R. Domains specifying thrombin-receptor interaction. Nature 1991,
353, 674-677.
(15) Nanevicz, T.; Ishii, M.; Wang, L.; Chen, M.; Chen, J.; Turck, C.
W.; Cohen, F. E.; Coughlin, S. R. Mechanisms of thrombin receptor
agonist specificity. Chimeric receptors and complementary mutations
identify an agonist recognition site. J. Biol. Chem. 1995, 270, 21619-
21625.
(16) Chackalamannil, S. Thrombin Receptor (Protease-Activated Receptor-
1) Antagonists as Potent Antithrombotic Agents with Strong Anti-
platelet Effects. J. Med. Chem. 2006, 49, 5389-5403.
(17) Seiler, S. M.; Bernatowicz, M. S. Peptide-derived protease-activated
receptor-1 (PAR-1) antagonists. Curr. Med. Chem.; CardioVasc.
Hemat. Agents 2003, 1, 13.
(18) Chackalamannil, S.; Xia, Y. Thrombin receptor (PAR-1) antagonists
as novel antithrombotic agents. Expert Opin. Ther. Pat. 2006, 16,
493-505.
(19) Chackalamannil, S.; Xia, Y.; Greenlee, W. J.; Clasby, M.; Doller,
D.; Tsai, H.; Asberom, T.; Czarniecki, M.; Ahn, H.-S.; Boykow, G.;
Foster, C.; Agans-Fantuzzi, J.; Bryant, M.; Lau, J.; Chintala, M.
Discovery of Potent Orally Active Thrombin Receptor (Protease-
Activated Receptor 1) Antagonists as Novel Antithrombotic Agents.
J. Med. Chem. 2005, 48, 5884-5887.
(20) Chackalamannil, S.; Asberom, T.; Xia, Y.; Doller, D.; Clasby, M.
C.; Czarniecki, M. F. Preparation of himbacine analogs as thrombin
receptor antagonists. U.S. Patent 6,063,847, May 16, 2000.
(21) Zhang, H.-C.; Derian, C. K.; Andrade-Gordon, P.; Hoekstra, W. J.;
McComsey, D. F.; White, K. B. Poulter, B. L.; Addo, M. F.; Cheung,
W.-M.; Damiano, B. P.; Oksenberg, D.; Reynolds, E. E.; Pandey,
(31) Shih, C.; Fritzen, E. L.; Swenton, J. S. Photocycloaddition chemistry
of 2-(trimethylsilyl)cyclopentenone and 5-(trimethylsilyl)uracil. The
utility of a trimethylsilyl group as a removable directing group in
photochemistry. J. Org. Chem. 1980, 45, 4462-4471.
(32) Ahn, H.-S.; Foster, C.; Boykow, G.; Arik, L.; Smith-Torhan, A.; Hesk,
D.; Chatterjee, M. Mol. Pharm. 1997, 51, 350-356. Assays were
carried out in duplicate. Compounds of high interest were assayed
multiple times (n > 5, SEM ( 20%).
(33) Kinetic solubilities were measured for these compounds and were
found to be ∼5 µM
(34) Data not shown.
JM061043E