Structure - activity relationship studies leading to the identification of (2E)-3-[l-[(2,4-dichlorophenyl)methyl]-5-fluoro-3-methyl-lH-indol-7-yl]-N-[(4, 5-dichloro-2-thienyl)sulfonyl]-2-propenamide (DG-041), a potent and selective prostanoid EP3 receptor antagonist, as a novel antiplatelet agent that does not prolong bleeding

Article abstract of DOI:10.1021/jm9005912

The EP₃ receptor on the platelet mediates prostaglandin E ₂ potentiation of thrombogenic coagonists including collagen and adenosine diphosphate (ADP). A pharmacophore driven approach led to the identification of diverse peri-substituted heterocycles as potent and selective EP₃ receptor antagonists. A simultaneous chemical optimization and druglike assessment of prioritized molecules converged on a lead compound 50 (DG-041) that displayed favorable in vitro and functional activities as an inhibitor of human platelet aggregation. This agent is currently in human clinical trials for the treatment of atherothrombosis.

Full text of DOI:10.1021/jm9005912

18 J. Med. Chem. 2010, 53, 18–36
DOI: 10.1021/jm9005912
Structure-Activity Relationship Studies Leading to the Identification of (2E)-3-[l-[(2,4-
Dichlorophenyl)methyl]-5-fluoro-3-methyl-lH-indol-7-yl]-N-[(4,5-dichloro-2-thienyl)sulfonyl]-
2-propenamide (DG-041), a Potent and Selective Prostanoid EP3 Receptor Antagonist,
as a Novel Antiplatelet Agent That Does Not Prolong Bleeding
Jasbir Singh,*,† Wayne Zeller,† Nian Zhou,† Georgeta Hategan,† Rama K. Mishra,† Alex Polozov,† Peng Yu,†
†
†
Emmanuel Onua, Jun Zhang, Jose L. Ramı
´
rez,‡ Gudmundur Sigthorsson,‡ Margret Thorsteinnsdottir,‡ Alex. S. Kiselyov,†
ꢀ
David E. Zembower, Thorkell Andresson, and Mark E. Gurney
†
‡
†,‡
ꢀ
†deCODE Chemistry, Inc., 2501 Davey Road, Woodridge, Illinois 60517, and ‡deCODE Genetics, Sturlgata 8, IS-101, Reykjavik, Iceland
Received May 7, 2009
The EP3 receptor on the platelet mediates prostaglandin E2 potentiation of thrombogenic coagonists including
collagen and adenosine diphosphate (ADP). A pharmacophore driven approach led to the identification of
diverse peri-substituted heterocycles as potent and selective EP3 receptor antagonists. A simultaneous
chemical optimization and druglike assessment of prioritized molecules converged on a lead compound
50 (DG-041) that displayed favorable in vitro and functional activities as an inhibitor of human platelet
aggregation. This agent is currently in human clinical trials for the treatment of atherothrombosis.
Introduction
platelet function. Irreversible COX-1 inhibitors (e.g., aspirin)
or irreversible P2Y12 ADP receptor antagonists (e.g., clopido-
grel, prasugrel), reduce atherothrombosis but also negatively
impact hemostasis by reducing platelet aggregation in re-
sponse to vascular breech.6 Since neither platelets nor the
healthy arterial wall produces PGE2, inhibition of the PGE2-
EP3 signaling is not expected to affect bleeding.2,4 Thus, EP3
antagonists have the potential to lower risk of atherothrom-
bosis without increasing bleeding risk. Recently, we reported
the identification and pharmacology of 50, a potent and
selective EP3 receptor antagonist, that is currently in phase
II clinical trials as a novel antiplatelet agent.7 In this report, we
describe both discovery and the structure-activity relation-
ship (SAR) studies of peri-substituted indole derivatives that
yielded 50.
The EP3 receptor is one of eight G-protein-coupled recep-
tors (GPCRs) that belong to the prostanoid receptor family.1
Prostanoids are ubiquitous autocrine mediators involved in
numerous physiological and pathological processes including
inflammation. PGE2, the natural ligand for the EP3 receptor,
is synthesized from arachidonic acid by sequential action of
COX-1a/COX-2 and PGE synthase. PGE2 is known to act
through four GPCR receptors referred to as EP1-4. Among
these four, the EP3 receptor is unique, as it signals through Gi
(inhibitory G-protein). The other three EP receptors (EP1,
EP2, and EP4) are coupled via Gs (stimulatory G-protein).
In the blood compartment, EP3 receptors are expressed on
platelets. Their activation results in lowering intracellular
cAMP levels. This step is associated with an increased sensitiv-
ity of platelets to a coagonist such as collagen or ADP, resulting
in platelet activation and subsequent aggregation.2,3 Studies of
EP3-null platelets, harvested from EP3-gene knockout mice,
suggest its role in the lesion-specific atherothrombosis over
atherosclerotic plaque.4 Plaque rupture exposes the collagen-
rich subendothelial matrix to platelets in the context of PGE2
produced by the inflamed plaque, thereby triggering thrombo-
sis. Thrombosis is blocked, however, if the platelets lack EP3.
Standard antiplatelet medications used to reduce risk of
heart attack and stroke increase risk of severe or fatal bleed-
ing.5 This is because such medications have a global impact on
Results and Discussion
Ligand-Based Design of EP3 Receptor Antagonists. Con-
sidering structural information for both the endogenous
ligand PGE2 (1) and the reported EP3 antagonist acylsulfo-
namides (3),8 we developed a pharmacophore hypothesis
and identified respective molecules to fit this putative model
(Figure 1). Our initial medicinal chemistry effort focused on
the synthesis of 1,3-disubstituted five-membered hetero-
cycles, which appeared to fit this initial pharmacophore
model. Unfortunately, in our hands these molecules featured
weak EP3 binding activity and no apparent structure-
activity relationship (SAR) with the IC50 values in the low
micromolar range. Representative examples (4-9) of this
series are shown in Table 1.
*To whom correspondence should be addressed. Phone: 1-630-904-
2218. E-mail: JSingh@decode.com.
a Abbreviations: ADP, adenosine diphosphate; CHO, Chinese hamster
ovary; COX-1, cyclooxygenase 1; COX-2, cyclooxygenase 2; PAA, platelet
aggregation; PGE, prostaglandin E; P2Y12, adenosine diphosphate G-
protein-coupled receptor; clopidrogel, methyl (2S)-(2-chlorophenyl)(6,7-
dihydrothieno[3,2-c]pyridin-5(4H)-yl)acetate; prasugrel, 5-[2-cyclopropyl-
1-(2-fluorophenyl)-2-oxoethyl]-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl
acetate.
Further comparison of PGE2 (1) and sulprostone (2), an
EP3 selective synthetic agonist, suggested tolerance for
structural variations in the acyclic portion of PGE2 for
binding to EP3 . We reasoned that the proper spatial arrange-
ment of peripheral pharmacophores should provide more
r
pubs.acs.org/jmc
Published on Web 12/03/2009
2009 American Chemical Society

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 19
Figure 1. Known EP3 agonists and antagonists.
Table 1. Representative Examples of 1,3-Disubstituted Heterocycles
Figure 3. Initial hit from rational design.
Table 2. Indole-Core-Derived 1,7-Disubstituted Analoguesa
compd
Ar1
R
hEP3 IC50 (nM)
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
phenyl
H
98
3,4-dichlorophenyl
2,4-dichlorophenyl
2,6-dichlorophenyl
2,4-dichlorophenyl
3,4-dimethylphenyl
3,5-dimethoxyphenyl
3,4-OCH2O-phenyl
3-CF3-phenyl
CH3
H
14.2
2
H
420
5
CH3
H
71
88
H
H
14
25
H
2-CF3-phenyl
2-biphenyl
H
42
H
326
34
4-methoxy phenyl
3-pyridyl
3-pyridyl
H
H
4044
2838
10091
CH3
CH3
2-pyridyl
a Ar2 = 2-thiophenyl.
Figure 2. Knowledge of ligand conformation to aid design of small
molecule antagonists: (a) energy minimized structure of PGE2;8 (b)
representation of deCODE’s hypothesis and its overlay on PGE2.
Figure 4. 1,7- vs 3,4-indole derivatives.
Table 3. 3,4-Disubstituted Indole Analogues (Z = CH2)
potent EP3 binders. Initial data for derivatives (3) also
supported this notion. Energy minimization studies of
PGE2 (Figure 2a) and our structures were in agreement with
the literature data.9 Two main assumptions were applied to
the design of new cores: (i) in order to ascertain optimal
spatial orientation of the substituents, these were to project
key binding elements, namely, polar and hydrophobic bind-
ing groups through nonvicinal arrangement, and (ii) the key
anchor atoms of the core that would bear acidic and hydro-
phobic pharmacophoric features would most likely overlap
with the C7/C13 or C8/C14 atoms of PGE2 (Figure 2b).
compd
X-Y
R
hEP3 IC50 nM
26
27
28
29
30
CHdCH
CHdCH
CH2-CH2
CH2-CH2
O-CH2
H
2.5
2.7
2.4
2.5
3.0
CH3
H
CH3
H
In order to provide an element of preorganization for
the final molecule, we selected peri-substituted bicyclic
heterocycles as our initial templates. Gratifyingly, the

20 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Singh et al.
indole analogues (10) and (11) featured good activity in a
human (h) EP3 receptor-binding assay featuring nanomo-
lar IC50 values (Figure 3). Pharmacologically, compounds
10 and 11 displayed full antagonist activity in a follow-up
functional assay (45 and 119 nM, respectively, in CHO
cells).
Following this initial success, we prepared a number of
indole analogues to explore both electronic and steric re-
quirements of this binding mode.10 Compounds 11-25
(Table 2; hEP3 binding IC50 values are shown) highlight
Figure 5. Generic pharmacophore highlighting vectors with solid
blue arrows.
Table 4. Diverse Cores Provided Active Analogues with hEP3 IC50 < 100 nMa
a Ar1a = 2-naphthyl. Ar1b = 2,4-dichlorophenyl. Ar1c = 3,4-dichlorophenyl. Ar2a = 2-thiophenyl. Ar2b = 2,3-dichloro-5-thiophenyl.

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 21
Table 5. Metabolic Stability and hEP3 Receptor Binding Assay (IC50
representative SAR for the N-substituted derivatives of
indole. Compared to the parent N-benzyl analogue 11, more
lipophilic meta and para substituents (examples 12-15 and
18) provided good activity, 2,4-dichlorobenyl derivative 13
being the most active in these series. Incorporation of methyl
group at C3 position of the indole core did not affect activity,
as analogues 13 and 15 were essentially equally active.
Piperonyl (18) and 4-methoxy (22) molecules featured better
potency than the respective 3,5-dimethoxy ether 17. Ortho
substituents (see 20, 21, and 14) afforded less potent com-
pounds. More polar pyridyl derivatives 23-25 were consid-
erably less active.
Considering the lipophilic nature of molecules 11-25, we
routinely determined hEP3 IC50 values in the presence of
varying concentrations of human serum albumin (HSA)
and/or 10% human serum (HS). Notably, the 2,3-dichloro-
thiophene sulfonamide substituent consistently featured low
fold shift (<10ꢀ) in the presence of plasma protein. Conse-
quently, this sulfonamide moiety was favored in our SAR
studies.
We further hypothesized that structural variations in the
bicyclic core portion of the molecule should be tolerated as
long as preorganization of the key binding elements, namely,
“acidic proton and lipophilic tail”, was maintained (Figure 4).
For instance, 3,4- versus 1,7-substitution patterns provided
similar trajectory for the key binding elements (Figure 4).
Indeed, 3,4-disubstituted indole analogues showed similar or
better potency compared to the respective 1,7-disubstituted
molecules (26-30) in the binding assay (Table 3).
Utilizing the SAR gathered thus far, we further refined our
pharmacophore hypothesis by studying torsional effects for
the key vectors tethering the core to the putative binding
elements via linkers L1 and L2 (Figure 5). A resulting
pharmacophore model11 yielded diverse [6.5], [6.6], and
[5.5] bicyclic scaffolds as hits. The most active molecules in
these series featuring IC50 < 100 nM in the hEP3 binding
assay are summarized in Table 4.
Lead Optimization. In order to further prioritize our lead
candidates, we investigated their in vitro metabolic stability
in multiple species including rat, dog, and human liver micro-
somes. Several peri-substituted 1,7-disubstituted indoles had
low metabolic stability in the assay. This effect was irrespec-
tive of the N-indole substituents and was likely derived from
the indole scaffold. As a confirmation, we incubated a
representative molecule 11 with rat liver microsomes
(RLM) followed by LC/MS/MS analysis of its metabolic
profile. Studies revealed two major metabolites that matched
mono- and bis-oxidative products most likely derived from
the C3 and C5 positions of the parent indole core. The
observed metabolic liability was successfully addressed by
incorporating a methyl group at C3 carbon and a fluorine
atom at C5 of the parent template. The resulting compounds
43 and 44 (Table 5) retained affinity against EP3 and
displayed adequate stability in the rat liver microsome assay.
Several 3,4-disubstituted indoles also displayed considerable
metabolic instability, although the phenomenon was species-
dependent. For example, compound 30 was stable upon
incubation with mouse and rat liver microsomes (71% and
68% parent remaining after 30 min). However, it was rapidly
metabolized (2% parent remaining after 30 min) in the pre-
sence of human liver microsomes (HLM) to yield an inactive
molecule, a product of oxidation at the benzylic methylene
carbon (Figure 4, Z = CO, X-Y = O-CH2, 45). The
structure of 45 was confirmed by an independent synthesis.
)
% parent remaininga
hEP3 IC50
(nM)
compd
Ar
R1 R2
15 min
30 min
10
13
43
44
2-naphthyl
2,4-dichlorophenyl H
H
H
H
F
4.5
1.9
14
3
8
0
22.2
88
77
4.2
82
56
2-naphthyl
2,4-dichlorophenyl CH3
CH3
F
a Following incubation with rat liver microsomes.
We subsequently conducted an extensive SAR study with
the metabolically stable 1,7-disubstituted 3-methyl-5-fluoro-
indole core varying Ar1 and Ar2 substituents. The data
(Table 6) indicated that more lipophilic meta and para
monosubstituted, ortho/para disubstituted, and 2-naphthyl
analogues as Ar1 provided improved activity when com-
pared to the unsubstituted N-benzyl analogue (11).
Although several analogues exhibited good activity in
the normal buffer, several of these showed relatively poor
IC50 in the presence of 10% human serum (Table 7),
indicating high plasma protein binding (PPB). As pre-
sented in Table 7, the analogues 30, 50, and 51, containing
4,5-dichloro thiophene as Ar2, provided low PPB poten-
tial.
The analogues derived from diverse cores containing peri-
substituents in general showed good to excellent selectivity
when profiled against a panel of other EP receptors and the
IP receptor. The selectivity data for a number of potent EP3
antagonists from the 1,7-indole, 3,4-indole, and 1,7-benzi-
midazoles series are shown in Table 8. The 2,4-dichloroben-
zyl derivatives (13, 30, 34, and 50) from different structural
series consistently yielded >100-1000 selectivity. The ana-
logue 51 that contained 2-naphthyl substituent displayed
lower selectivity for EP1. Overall, molecule 50 afforded the
best combination of in vitro activity and selectivity across
receptor panels along with low potential for PPB (Tables 6, 7,
and 8).
In the next optimization step, we varied the tether length
for the acidic portion of the metabolically stable 1,7-disub-
stituted indole core. The molecule 44 (Table 5) featuring a
two atom spacer CdC between the core and the acylsulfo-
namide group showed good EP3 activity, metabolic stability,
and low potential for PPB.7
Lead Candidate. Compound 50 was found to be a non-
competitive antagonist of PGE2 binding to EP3. In addition,
preincubation of EP3 receptor with 50 followed by exten-
sive washing completely abolished [3H]PGE2 binding
(Figure 6A). This behavior was common for other potent
EP3 antagonists, including 30. We speculated that the ana-
logues bearing the indole core behave as quasi-irreversible
EP3 antagonists. In order to differentiate slow Koff rate
from the covalent interaction of 50 with the receptor, we
conducted a series of displacement binding experiments.
Specifically, a solution of 1 mM dithiothreitol (DTT) as
reducing agent did not affect the displacement curve

22 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Table 6. SAR of 1,7-Disubstituted 3-Methyl-5-fluoroindole Series
Singh et al.
(Figure 6C), ruling out the possibility that 50 acts as a
Michael acceptor for the thiolate group of DTT or for an
-SH group on EP3.12 Notably, molecule 30, lacking an olefin
moiety, also displayed quasi-irreversible antagonism against
EP3 presumably due to a slow Koff. The similar behavior of the
analogues 66 and 67 (Figure 6B)13 further supported this
notion.
tail vein, thromboembolism in the lung and conse-
quent mortality were greatly reduced.4 We therefore ex-
plored the protection afforded by our lead compounds
in this assay. Adult mice were gavaged with 30, 50, and
51 at 30 and 100 mg/kg single dose. All molecules
were delivered as a soluble formulation in HPβCD. Com-
pound 50 afforded roughly 70% protection in the assay
(Figure 7). Although efficacious in the thromboembolism
test, 30 was unstable upon incubation with human liver
Studies with EP3 gene knockout mice had shown that
when animals were injected with arachidonic acid via the

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 23
microsomes (vide supra), so we prioritized 50 for further
development.
In ADME-PK studies, 50 showed similar metabolic pro-
files in mouse, rat, dog, monkey, and human liver microsome
assays (Table 9). It featured weak to moderate inhibition of
hCYP3A4, 2D6, and 2C19 isozymes but showed strong
inhibition of hCYP2C9 (Table 10) with no potential to cause
its metabolism-dependent inhibition. Although potentially
of concern, drug-drug interaction studies conducted in
healthy human volunteers showed that 50 does not inhibit
CYP3A4 or CYP2C9, likely because of the relatively low free
drug concentration in the presence of human plasma pro-
teins (∼0.7%).
Further efficacy assessement of compound 50 was carried
out by measuring inhibition of collagen and sulprostone
induced platelet aggregation (PAA) using whole human
blood. Compound 50 furnished similar IC50 (218 nM) for
PAA inhibition in blood derived from both male and female
subjects (Figure 8).
Table 7. Fold-Shift in IC50 for Selected Analogues in the Presence of
Plasma Proteins (Human Serum, HS)
Below is a summary of the ADME/PK study results for
compound 50. It exhibited high potential for GI permeability
in Caco2 assay (Papp = 11.2 ꢀ 10-6 cm/s) with no significant
efflux. The agent was marginally water-soluble (23 μg/mL,
pH7) as measured by HPLC (shake flask method). We
speculated that the limited solubility was rate-limiting for
absorption through the intestinal wall. As a result, bioavail-
ability (F, %) varied with formulation. Reduction in particle
size (e.g., micronization) or utilization of solubility enhan-
cing excipients increased F.14 Following intravenous and
oral dosing, compound 50 formulated with hydroxypropyl
β-cyclodextran (HPβCD) showed favorable pharmaco-
kinetic profiles in preclinical species (10 mg/kg oral dose;
F of 27% and 54%, t1/2 of 4.1 and 3.1 h, Cmax of 2.7 and
18.1 μM for rat and dog, respectively).
fold-shift in
IC50 in presence
of 10% HS
hEP3 IC50 (nM)
buffer þ 10% HSb
378
compd
hEP3 IC50 (nM)a
10
11
12
13
16
18
19
20
22
30
31
32
33
34
35
36
37
38
41
42
43
44
47
48
49
50
51
57
58
59
60
61
62
4.5
98
84
6129
7155
357
623
504
178
60
14.2
2
71
14
4223
2208
4048
2710
5568
5
158
162
65
25
42
34
164
1.7
52
3
22
1146
2730
635
2.2
3
1241
212
72
Compound 50 was well tolerated in preclinical safety
pharamacology and 3-month chronic toxicity studies in rat
(up to 150 mg/kg) and dog (up to 20 mg/kg) following
oral dosing. It was not genotoxic in AMES or in in vitro
micronucleus assays. The compound 50 currently is in
1.1
8.9
36
79
2751
1552
7260
1683
880
309
43
27
269
61
27.6
4.9
14.6
14
180
154
44
2252
619
Table 8. Prostanoid Receptor Selectivitya across Series in [3H]Ligand
Displacement Binding Assays
3
299
466
100
126
90
IC50 (nM)
3.7
6.6
1.3
4.6
7
595
682
compd
hEP3
hEP1
8104
hEP2
hEP4
hIP
mEP3
524
1.3
5.1
46
13
30
34
50
51
2
>10000
12261
4832
8178
6498
10172
8039
3647
ND
ND
28
4
6
36
3
6354
7959
1.1
4.6
7
5875
14414
8823
4
5.7
0.9
14.5
2.3
46.4
3.6
261
147
>20000
360
4169
32921
11
5
163
330
170
150
568
4790
393
a Experimental IC50 values ((SEM) from displacement bind-
ing analysis with a minimum of five experiments per value. Displace-
ment binding was assessed with [3H]PGE2 for human EP1-4 receptors
and the mouse EP3 receptor, with [3H]iloprost for the human
IP receptor.
6939
2045
a Normal assay buffer. b Assay buffer containing 10% human serum.
Figure 6. (A) Quasi-irreversibility of 50 and its structural analogues for binding to hEP3 receptor. Membranes were washed three times for 1 h
at room temperature and incubated with 2 nM [3H]PGE2 for an additional hour at 30 °C. Quantified binding is expressed as percentage of
control (buffer only) membranes and shown as the mean ( SEM of three independent experiments. Statistical analysis was performed by
impaired nonparametric Student t test using GraphPad Prism software: (a) p < 0.01 vs control; (b) p < 0.01 vs PGE2-incubated membranes.
(B) Structural features of probe analogues 50, 66, and 67. All analogues have Ar1 = 2,4-dichlorobenzyl and 3-methyl-5-fluoroindole as the
core. (C) Effect of reducing agent dithiothreitol (DTT) on binding of 50 to hEP3 DR.

24 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Singh et al.
human PhII clinical trials as a novel antiplatelet agent in
cardiovascular disease.
Synthesis. Peri-substituted bicyclic analogues described
here were synthesized as outlined in Schemes 1-6. Reaction
of commercially available 7-formylindole (68) with triethyl
phosphonoacetate followed by base hydrolysis of the ethyl
ester 69 provided the acid 70. Coupling of acid 70 with 2-
thiophenesulfonamide using EDCI/HOBt provided the key
intermediate 71, which, following alkylation of the indole
nitrogen, provided 1,7-disubstitued indole analogues 10, 11,
13-22. As shown in Scheme 2, the C3 methylindole inter-
mediate 73 was prepared by Heck coupling of the 3-methyl-
7-bromoindole (72) with methyl acrylate. Base hydrolysis
followed by formation of the acylsulfonamide 75 and indole
nitrogen alkylation, as shown in Scheme 1, provided analo-
gues 12, 15, 24, and 25.
The 3,4-disubstituted indole analogues were prepared start-
ing from 4-bromoindole (76, Scheme 3). C3-Acylation of 76
was performed using EtMgBr/ZnCl2 protocol, analogous to
the procedure of Bergman and Venemalm.15 Reduction of
ketone 77 with borane followed by Heck coupling of the
bromoaryl 78 provided the key 3,4-disubstituted intermedi-
ate 79. Subsequent transformation of the ester via acid 80
provided the target compound 26. N-Methylation of 79
afforded 81, and subsequent derivatization of the correspond-
ing acid 82 provided analogue 27. Hydrogenation of 79
Figure 7. Protection from pulmonary thromboembolism by 30, 50,
and 51. Test compounds were given orally formulated in HPβCD.
Thirty minutes after dosing, mice were challenged by injection of
arachidonic acid (30 mg/kg) into the lateral tail vein.
Table 9. In Vitro Metabolic Stability of Compound 50a
% parent remaining at 30 min
mouse
rat
dog
monkey
human
72.0 ( 4.7
77.7 ( 0.4
88.6 ( 1.4
88.4 ( 3.6
80.9 ( 0.5
a 5 μM compound was incubated with mouse, rat, dog, monkey, and
human liver microsomes, and % parent remaining was determined by
LC/MS/MS.
Table 10. hCYP Inhibition IC50 (μM) for Compound 50
Figure 8. Inhibition of human platelet aggregation by compound
50. Platelet aggregation was triggered by collagen combined with
sulprostone,1 a selective EP3 agonist.
1A2
2C19
6.6
2C9
0.03
2D6
>10
3A4 (DBF)
7.6
3A4 (BFC)
1.5
>10
Scheme 1a
a (a) Triethylphosphonoacetate, NaH, THF, 78 °C, 14 h, 68%; (b) NaOH, EtOH, H2O, 78 °C, 4 h, 86%; (c) 2-thiophenesulfonamide, DMAP, EDCI,
CH2Cl2, room temp, 72 h, 56%; (d) NaH, DMF, Ar/(R)-CH2Br(Cl).
Scheme 2a
a (a) Methyl acrylate, Et3N, Pd(OAc)2, tri-o-tolylphosphine, 100 °C, 4 h, sealed tube, 82%; (b) aq NaOH, THF, CH3OH, room temp, 16 h, 100%;
(c) 2-thiophenesulfonamide, EDCI, DMAP, CH2Cl2, DMSO, room temp, 16 h, 58%; (d) NaH, DMF, Ar/R-CH2Br(Cl).

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 25
Scheme 3a
a (a) CH3MgBr, ZnCl2, 2-naphthoyl chloride, CH2Cl2, rt, 16 h, 74%; (b) BH3, THF, 0 °C to rt, 1.5 h; (c) methyl acrylate, Pd(OAc)2, (o-tolyl)3P, Et3N,
sealed tube, 100 °C, 2.5 h, 65%; (d) 1 N aqueous NaOH, THF, CH3OH, 60 °C, 4 h, 100%; (e) 4,5-dichlorothiophene-2-sulfonamide, DMAP, EDCI,
CH2Cl2, rt, 16 h, 42%; (f) NaH, THF, CH3I, 0 °C to rt, 16 h; (g) H2, 10% Pd/C, CH3OH, 40 °C, 2 h, rt, 16 h, 86%; (h) 1 N aq NaOH, THF, CH3OH,
60 °C, 4 h, 90%; (i) 4,5-dichlorothiophene-2-sulfonamide, DMAP, EDCI, CH2Cl2, rt, 16 h.
Scheme 4a
a (a) Ac2O, pyridine, 25-32 °C, 15 min, 98%; (b) 2-naphthoyl chloride, CH3MgBr, ZnCl2, rt, 3 h, 65%; (c) BH3, THF, 62%; (d) methyl 2-
bromoacetate, K2CO3, DMF, rt, 17 h, 75%; (e) 2 N aq NaOH, THF, CH3OH, rt, 1 h, 92%; (f) 4,5-dichlorothiophene-2-sulfonamide, DMAP, EDCI,
CH2Cl2, rt, 16 h, 71%.
provided the saturated ester 83, which was carried through
a synthetic sequence analogous to synthesis of olefin analo-
gues 26 and 27, providing the R = NH and N-methyl
saturated acylsulfonamide analogues 28 and 29, respectively
(Scheme 3).
removal of the acetate protecting group provided 4-hydroxy-
3-(2-naphthylmethyl)indole (90). O-Alkylation of 90 with
methyl bromoacetate followed by base hydrolysis provided
the acid 92. EDCI-mediated coupling with 4,5-dichloro-2-
thiophenesulfonamide provided the analogue 30.
The synthesis of compound 30 is outlined in Scheme 4. 4-
Hydroxyindole (87) was protected as the acetate 88,
and subsequent Freidel-Crafts acylation with 2-naphthoyl
chloride followed by reduction of ketone 89 with concomitant
For 1,7-disubstituted analogues derived from the 5-
fluoro-3-methylindole core, the 7-bromoindole intermedi-
ate 93 was prepared according to the method of Dobbs,16
Bartoli et al.,17or Zegar et.al.18 Following the synthetic

26 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Singh et al.
Scheme 5a
a (a) (o-Tol)3P, methyl acrylate, Pd(OAc)2; (b) aq MeOH, NaOH, 87%; (c) EDCI, DMAP, sulfonamide; (d) DMF, NaH, aryl bromide.
Scheme 6a
a (a) (o-Tol)3P, methyl acrylate, Pd(OAc)2, 98%; (b) aq MeOH, NaOH; (c) EDCI, DMAP, sulfonamide, 18%; (d) DMF, NaH, aryl bromide, 64%.
protocols described in Scheme 5, the target compounds
43 and those shown in Table 6 were obtained. For the
synthesis of analogue 63, the indole 93 was first N-alkylated
to provide 97. Heck coupling was carried out to pro-
vide, after base hydrolysis, the acid 99 which upon coupl-
ing with the trifluoromethanesulfonamide provided the
analogue 63. As shown in Scheme 6, the acrylic acid inter-
mediate 95 was hydrogenated and the resulting saturated
acid 100 was subsequently functionalized analogous to
a two-step sequence used in Scheme 5 to provide ana-
logue 66. Preparation of compound 50 has been reported
in ref 18.
Experimental Section
Chemistry. General Methods. All reagents and anhy-
drous solvents were obtained from commercial sources
and used without further purification unless otherwise noted.
NMR spectra were recorded at 400 or 500 MHz (Varian
Instruments) in the solvent indicated, and TMS was used as
an internal reference. ACDLabs NMR software was used to
process FIDs to generate spectral parameters (ppm/Hz).
Coupling constants (J) are given in Hz. Mass spectra were
obtained using either APCI or electrospray ionization (PE-
SCIEX single-quad or Agilent mixed-mode units). Elemental
analyses were carried out by Galbraith Laboratories, Inc.
(Knoxville, TN) or Midwest Microlab, LLC (Indianapolis,
IN). Column chromatography was carried out in the solvents
indicated with silica gel (MP EcoChrom, 32-63D, 60 A).
Purity of all final products was determined by LC-MS and/
or analytical HPLC to be g95%. HPLC purity of compounds
was measured with a reverse phase HPLC (Phenomenex
Prodigy C18 column, 4.6 mm ꢀ 150 mm, 5 μm, 254 nm) with
two diverse solvent systems. In system 1, compounds were
eluted using a gradient elution of 95/5 to 5/95 A/B over 20 min
at a flow rate of 1.0 mL/min, where solvent A was aqueous
0.05% TFA and solvent B is acetonitrile (0.05% TFA). In
system 2, compounds were eluted using a gradient of 95/5 to
5/95 A/C over 20 min at a flow rate of 1.0 mL/min, where
solvent A was aqueous 0.05% TFA and solvent C is methanol.
For HPLC data, peak area percent and retention time (tR in
minutes) are provided. The following compounds were pur-
chased from commercial soureces and used as such, compounds
Conclusion
In summary, on the basis of the structure of an endogenous
ligand (PGE2), we generated a pharmacophore hypothesis
and devised multiple series of potent and selective EP3 recep-
tor antagonists. Subsequent refinement of this model follow-
ing several rounds of SARculminated inthe identificationof a
lead compound 50 that currently is in phase II clinical trials.
This agent featured high affinities for both hEP3 (IC50=4.6
nM) and mEP3 (IC50=5.3 nM) receptors in the absence of
serum proteins. Compound 50 showed inhibitory activity
against human platelet aggregation (IC50 = 218 nM). The
compound displayed favorable preclinical PK and safety
pharmacology profiles in rats and dogs. The in vivo profile
of 50 is reported elsewhere.7

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 27
68, 76, and 87. Synthesis of compounds 50, 93, 94, 95, and 96b
have been reported previously.18
Thiophene-2-sulfonic Acid {(E)-3-[1-(2,4-Dichlorobenzyl)-1H-
indol-7-y l]acryloyl}amide (13). Compound 13 was prepared
from 71 using general procedure B. Yield: 51% (36% after
crystallization). 1H NMR (500 MHz, acetone-d6) δ 5.64 (s, 2H),
6.26 (d, J = 8.5 Hz, 1H), 6.40 (d, J = 15.5 Hz, 1H), 6.65 (d, J =
3.5 Hz, 1H), 7.09-7.44 (m, 6H), 7.72 (d, J = 8.0 Hz, 1H), 7.90
(d, J = 4.0 Hz, 1H), 7.96 (d, J = 15.5 Hz, 1H), 8.00 (d, J = 3.5
Hz, 1H). MS (ESI-): mass calcd for C22H16Cl2N2O3S2, 490.0;
m/z found 489.4 (M - 1). LCMS 97%.
Thiophene-2-sulfonic Acid {(E)-3-[1-(2,6-Dichlorobenzyl)-1H-
indol-7-y l]acryloyl}amide (14). Compound 14 was prepared
from 71 using general procedure B. Yield: 14%. 1H NMR
(500 MHz, acetone-d6) δ 5.32 (s, 2H), 6.37-6.48 (m, 2H), 6.64
(d, J = 3.0 Hz, 1H), 6.88-7.22 (m, 3H), 7.30-7.55 (m, 3H), 7.64
(d, J = 6.0 Hz, 1H), 7.88-7.96 (m, 2H), 8.68 (d, J = 15.0 Hz,
1H). MS (ESI-): mass calcd for C22H16Cl2N2O3S2, 490.0; m/z
found 489.3 (M - 1). LCMS 97%.
Preparation of Acylsulfonamides. General Procedure A. To a
round-bottom flask (500 mL) that contained a solution of
aryl(heteroaryl)sulfonamide (6 mmol), 4-dimethyaminopyri-
dine (DMAP, 1.56 g, 13 mmol), and 1-[3-(dimethyamino)-
propyl]-3-ethylcarbodiimide hydrochloride (EDCI, 2.4 g, 13
mmol) in CH2Cl2 (150 mL) was added the carboxylic acid
(6 mmol) at room temperature. The resulting mixture was
allowed to stir at room temperature for 72 h, then was cooled
to 5 °C and was acidified with addition of aqueous HCl (10%)
until pH 1 was obtained, which was followed by extraction with
CH2Cl2/MeOH (9/1, 3 ꢀ 100 mL). The combined organic layers
were dried over anhydrous Na2SO4 and the solvent was con-
centrated in vacuo to provide the crude product which was
purified by flash chromatography (silica gel) to afford aryl-
(heteroaryl)acylsulfonamide.
Thiophene-2-sulfonic Acid {(E)-3-[1-(2,4-Dichlorobenzyl)-3-
methyl-1H-indol-7-yl]acryloyl}amide (15). Compound 15 was
prepared from 75 using general procedure B. Yield: 41%. H
N-Alkylation of Indoles. General Procedure B. As an example,
to a suspension of NaH (60% in mineral oil, 5 mg, 0.11 mmol) in
DMF (5 mL) was added thiophene-2-sulfonic acid (3-1H-indol-
7-ylacryloyl)amide, 71 (20 mg, 0.066 mmol), at 0 °C. [Except
when the Ar/R-CH2Br(Cl) was used in the form of a salt (e.g.,
HCl salt), an additional equiv of NaH was used.] The resulting
mixture that resulted was allowed to warm to room temperature
and was stirred for 2 h and then cooled to 0 °C. To this solution,
ArCH2Br (or ArCH2Cl) (0.072 mmol, 1.1 equiv) was added at
0 °C, and the resulting reaction mixture was allowed to warm
to room temperature and was stirred for 16-48 h. The reac-
tion mixture was cooled to 5 °C and was acidified with
addition of aqueous HCl (10%) until pH 1 was obtained,
which was followed by extraction with CH2Cl2/MeOH (9/1, 3 ꢀ
10 mL). The combined organic layers were dried over anhydrous
Na2SO4 and filtered, and the solvent was concentrated in vacuo
to provide the crude product, which was purified by flash
chromatography (silica gel, gradient elution, CH2Cl2 and then
EtOAc/hexane, 1:8 to 1:2), recrystallization, or trituration from
ethyl ether to afford the desired N-alkylated thiophene-2-sulfo-
nic acid (3[CH2R]-1H-indol-7-ylacryloyl)amides.
Thiophene-2-sulfonic Acid [(E)-3-(1-Naphthalen-2-ylmethyl-
1H-indol-7-yl)acryloyl]amide (10). Compound 10 was prepared
from 71 using general procedure B. Yield: 61% (36% after
recrystallization). 1H NMR (500 MHz, CDCl3) δ 5.63 (s, 2H),
6.13 (d, J = 15.0 Hz, 1H), 6.64 (d, J = 3.5 Hz, 1H), 7.06-7.10
(m, 2H), 7.19 (t, J = 7.5 Hz, 2H), 7.23 (d, J = 3.5 Hz, 1H),
7.41-7.45 (m, 3H), 7.64-7.78 (m, 5H), 7.89 (dd, J = 4.0, 1.0 Hz,
1H), 8.29 (d, J = 15.0 Hz, 1H). 13C NMR (500 MHz, CDCl3) δ
53.5, 102.4, 118.3, 119.3, 120.0, 121.9, 124.0, 124.1, 125.3, 126.1,
126.4, 127.4, 127.7, 128.1, 129.1, 130.9, 131.5, 132.9, 133.4,
133.7, 133.9, 134.9, 135.0, 139.2, 144.0, 162.5. MS (ESI-), mass
calcd for C26H20N2O3S2, 472.1; m/z found 471.5 (M - 1).
LCMS 98%.
1
NMR (500 MHz, DMSO-d6) δ 2.28 (d, J = 1.5 Hz, 3H), 5.52 (s,
2H), 6.14 (d, J = 8.5 Hz, 1H), 6.26 (d, J = 15.0 Hz, 1H), 7.11 (t,
J = 7.5 Hz, 1H), 7.20-7.30 (m, 4H), 7.52 (d, J = 1.5 Hz, 1H),
7.65 (dd, J = 7.5, 1.5 Hz, 1H), 7.76 (d, J = 15.0 Hz, 1H), 7.82 (m,
1H), 8.09 (m, 1H), 12.26 (br s, 1H). MS (ESI-): mass calcd for
C23H18Cl2N2O3S2, 504.0; m/z found 503.3 (M - 1). LCMS
100%. HPLC system 1: 99%, tR = 16.73 min. Anal. C, H, N.
Thiophene-2-sulfonic Acid {(E)-3-[1-(3,4-Dimethylbenzyl)-
1H-indol-7-y l]acryloyl}amide (16). Compound 16 was prepared
from 71 using general procedure B. Yield: 44%. 1H NMR (500
MHz, acetone-d6) δ 2.28 (s, 6H), 5.51 (s, 2H), 6.45 (d, J = 15.0
Hz, 1H), 6.62 (d, J = 3.0 Hz, 1H), 6.92-6.96 (m, 1H), 7.03-7.09
(m, 3H), 7.22-7.29 (m, 2H), 7.33 (d, J = 3.0 Hz, 1H), 7.71 (d,
J = 7.5 Hz, 1H), 7.87-7.89 (m, 1H), 7.98-8.02 (m, 1H), 8.02 (d,
J = 15.0 Hz, 1H), 10.75 (br s, 1H). LCMS (ESI-): mass calcd for
C24H22N2O3S2, 450.1; m/z found 449.4 (M - 1). LCMS 96%.
HPLC system 1: 95.4%, tR = 15.61 min.
Thiophene-2-sulfonic Acid {(E)-3-[1-(3,5-Dimethoxybenzyl)-
1H-indol-7-yl]acryloyl}amide (17). Compound 17 was prepared
from 71 using general procedure B. Yield: 10%. 1H NMR (500
MHz, CD3OD) δ 3.65 (s, 6H), 5.48 (s, 2H), 6.11 (d, J = 2.5 Hz,
2H), 6.28-6.32 (m, 2H), 6.57 (d, J = 3.5 Hz, 1H), 7.04 (t, J = 7.5
Hz, 1H), 7.14-7.18 (m, 1H), 7.27 (d, J = 7.5 Hz, 1H), 7.31 (d,
J = 3.0 Hz, 1H), 7.64 (d, J = 7.5 Hz, 1H), 7.81-7.84 (m, 2H),
8.25 (d, J = 15.0 Hz, 1H). MS (API-): mass calcd for
C24H22N2O5S2, 482.1; m/z found 480.8 (M - 1).
Thiophene-2-sulfonic Acid [(E)-3-(1-Benzo[1,3]dioxol-5-yl-
methyl-1H-indol-7-yl)acryloyl]amide (18). Compound 18 was
prepared from 71 using general procedure B. Yield: 85%. H
1
NMR (500 MHz, acetone-d6) δ 5.56 (s, 2H), 5.94 (s, 2H), 6.45 (s,
1H), 6.51(d, J = 15.0 Hz, 1H), 6.58-6.67 (m, 3H), 7.08 (t, J =
7.5 Hz, 1H), 7.27-7.31 (m, 2H), 7.47 (d, J = 3.5 Hz, 1H), 7.70
(d, J = 7.5 Hz, 1H), 7.95 (dd, J = 3.0, 1.5 Hz, 1H), 8.02 (dd, J =
4.5, 1.5 Hz, 1H), 8.35 (d, J = 15.0 Hz, 1H), 10.80 (br s, 1H). MS
(ESI-): mass calcd for C23H18N2O5S2, 466.1; m/z found 465.3
(M - 1). LCMS 97%.
Thiophene-2-sulfonic Acid {(E)-3-[1-(3-Trifluoromethylben-
zyl)-1H-indol-7-yl]acryloyl}amide (19). Compound 19 was pre-
pared from 71 using general procedure B. Yield: 85%. 1H NMR
(500 MHz, acetone-d6) δ 5.75 (2H, s), 6.47 (d, J = 15.0 Hz, 1H),
6.65 (d, J = 3.0 Hz, 1H), 7.08 (t, J = 7.5 Hz, 1H), 7.24-7.34 (m,
4H), 7.45 (t, J = 8.0 Hz, 1H), 7.52-7.57 (m, 2H), 7.70 (d, J = 8.0
Hz, 1H), 7.92 (m, 1H), 8.00 (m, 1H), 8.23 (d, J = 15.0 Hz, 1H).
MS (ESI-): mass calcd for C23H17F3N2O3S2, 490.1; m/z found
489.4 (M - 1). LCMS 96%.
Thiophene-2-sulfonic Acid [(E)-3-(1-Benzyl-1H-indol-7-yl)acry-
loyl]amide (11). Compound 11 was prepared from 71 using
general procedure B. Yield: 42%. 1H NMR (500 MHz, CDCl3)
δ 5.49 (s, 2H), 6.14 (d, J = 15.0 Hz, 1H), 6.60 (d, J = 3.0 Hz,
1H), 7.00 (d, J = 7.5 Hz, 2H), 7.09 (t, J = 7.5 Hz, 1H),
7.13-7.16 (m, 2H), 7.20-7.28 (m, 5H), 7.69-7.71 (m, 2H),
7.93 (dd, J = 4.0, 1.0 Hz, 1H), 8.16 (d, J = 15.0 Hz, 1H). MS
(ESI-): mass calcd for C22H18N2O3S2, 422.1; m/z found 421.3
(M - 1). LCMS 98%.
Thiophene-2-sulfonic Acid {(E)-3-[1-(3,4-Dichlorobenzyl)-3-
methyl-1H-indol-7-yl]acryloyl}amide (12). Compound 12 was
prepared from 75 using general procedure B. H NMR (500
1
MHz, CDCl3) δ 2.33 (d, J = 1.5 Hz, 3H), 5.34 (s, 2H), 6.26 (d,
J = 15.0 Hz. 1H), 6.88, (s, 1H), 6.90 (dd, J = 8.0, 4.0 Hz, 1H),
7.05-7.10 (m, 2H), 7.11 (dd, J = 5.0, 4.0 Hz, 1H), 7.22-7.30 (m,
3H), 7.62 (d, J = 7.5 Hz, 1H), 7.67 (dd, J = 4.5, 1.5 Hz, 1H), 7.91
(dd, J = 4.0, 1.5 Hz, 1H), 8.17 (d, J = 15.0 Hz. 1H). MS (ESI-):
mass calcd for C23H18Cl2N2O3S2, 504.0; m/z found 503.3
(M - 1). LCMS 96%. HPLC system 2: 95%, tR = 17.71 min.
Thiophene-2-sulfonic Acid {(E)-3-[1-(2-Trifluoromethylben-
zyl)-1H-indol-7-yl]acryloyl}amide (20). Compound 20 was pre-
pared from 71 using general procedure B. Yield: 17%. 1H NMR
(500 MHz, CDCl3) δ 5.64 (2H, s), 6.18 (d, J = 15.0 Hz, 1H), 6.49
(d, J = 8.0 Hz, 1H), 6.65 (d, J = 3.0 Hz, 1H), 7.07 (d, J = 3.0 Hz,

28 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Singh et al.
1H), 7.11-7.14 (m, 2H), 7.21 (d, J = 8.0 Hz, 1H), 7.31-7.38 (m,
2H), 7.69 (dd, J = 5.0, 1.0 Hz, 1H), 7.72 (t, J = 7.5 Hz, 1H), 7.82
(d, J = 15.0 Hz, 1H). MS (ESI-): mass calcd for C23H17F3-
N2O3S2, 490.1; m/z found 489.4 (M - 1). LCMS 95%.
Thiophene-2-sulfonic Acid [(E)-3-(1-Biphenyl-2-ylmethyl-1H-
indol-7-yl)acryloyl]amide (21). Compound 21 was prepared
from 71 using general procedure B. Yield: 17%. 1H NMR
(500 MHz, CD3OD) δ 5.36 (2H, s), 6.18 (d, J = 15.0 Hz, 1H),
6.82 (d, J = 8.0 Hz, 1H), 7.04 (t, J = 8.0 Hz, 1H), 7.11-7.37 (m,
13H), 7.82-7.91 (m, 2H), 7.92 (d, J = 15.0 Hz, 1H). MS (ESI-):
mass calcd for C28H22N2O3S2, 498.1; m/z found 497.6 (M - 1).
LCMS 96%.
Thiophene-2-sulfonic Acid {(E)-3-[1-(4-Methoxybenzyl)-1H-
indol-7-yl]acryloyl}amide (22). Compound 22 was prepared
from 71 using general procedure B. Yield: 77%. 1H NMR
(500 MHz, acetone-d6); δ 3.74(s, 3H), 5.58 (s, 2H), 6.45 (d,
J = 15.0 Hz, 1H), 6.61 (d, J = 3.0 Hz, 1H), 6.68 (dd, J = 6.5, 2.5
Hz, 2H), 6.96 (d, J = 9.0 Hz, 2H), 7.07 (t, J = 7.5 Hz, 1H),
7.25-7.30 (m, 2H), 7.47 (d, J = 3.0 Hz, 1H), 7.70 (d, J = 7.5 Hz,
1H), 7.95 (dd, J = 4.0, 1.5 Hz, 1H), 8.04 (dd, J = 5.0, 1.5 Hz,
1H), 8.37 (d, J = 15.0 Hz, 1H), 10.80 (br s, 1H). MS (ESI-):
mass calcd for C23H20N2O4S2, 452.1; m/z found 451.3 (M - 1).
LCMS 97%.
calcd for C26H18Cl2N2O3S2, 540.0; m/z found 539.6 (M - 1).
LCMS 96%. HPLC system 2: 95%, tR = 17.64 min.
4,5-Dichlorothiophene-2-sulfonic Acid [(E)-3-(1-Methyl-3-
naphthalen-2-ylmethyl-1H-indol-4-yl)acryloyl]amide (27). To a
suspension of the acid 82 (42 mg, 0.123 mmol) in CH2Cl2 (1 mL)
was sequentially added 4,5-dichlorothiophene-2-sulfonamide
(29 mg, 0.123 mmol), DMAP (30 mg, 0.246 mmol), and EDCI
(47 mg, 0.246 mmol). The mixture was stirred at room tempera-
ture overnight. The solution was acidified with 10% aqueous
HCl and extracted with EtOAc (2 ꢀ 5 mL). The organic layer
was washed with water, dried over MgSO4, and concentrated in
vacuo to afford 50 mg of residue. Purification of this residue by
column chromatography using CH2Cl2 gave 18 mg (26%) of the
compound 27. 1H NMR (500 MHz, DMSO-d6) δ 3.78 (s, 3H),
4.33 (s, 2H), 6.41 (d, J = 16.0 Hz, 1H), 7.15-7.18 (t, J = 8.0 Hz,
1H), 7.25 (d, J = 7.5 Hz, 1H), 7.348 (s, 1H), 7.37-7.43 (m, 4H),
7.51 (d, J = 8.0 Hz, 1H), 7.66 (d, J = 8.5 Hz, 1H), 7.71 (d, J =
8.5 Hz, 1H), 7.79 (d, J = 8.5 Hz, 1H), 7.8 (s, 1H), 8.38 (d, J =
16.0 Hz, 1H), 12.7 (br s, 1H). LCMS 98%. MS (ESI-) m/z 553
(M - 1). HPLC system 2: 98.6%, tR = 17.84 min.
4,5-Dichlorothiophene-2-sulfonic Acid [3-(3-Naphthalen-2-yl-
methyl-1H-indol-4-yl)propionyl]amide (28). To a suspension of
the propionic acid 84 (126 mg, 0.383 mmol) in CH2Cl2 (2 mL)
was sequentially added 4,5-dichlorothiophene-2-sulfonamide
(89 mg, 0.383 mmol), DMAP (93.5 mg, 0.765 mmol), and EDCI
(147 mg, 0.765 mmol). The mixture was stirred at room tem-
perature overnight. The solution was diluted with EtOAc (5 mL)
and acidified with 10% aqueous HCl. The organic layer was
washed with water, dried over anhydrous MgSO4, filtered, and
concentrated in vacuo. The resulting residue was purified by a
column chromatography using CH2Cl2 as eluent to provide 65
mg (31%) of the acylsulfonamide 28. 1H NMR (500 MHz,
DMSO-d6) δ 2.49 (t, J = 8.0 Hz, 2H), 2.97 (t, J = 8.0 Hz, 2H),
4.29 (s, 2H), 6.56 (d, J = 7.0 Hz, 1H), 6.87 (t, J = 7.0 Hz, 1H),
7.08 (d, J = 2.0 Hz, 1H), 7.20 (d, J = 8.0 Hz, 1H), 7.31 (dd, J =
8.5, 1.5 Hz, 1H), 7.42-7.44 (m, 2H), 7.55 (s, 1H), 7.71 (m, 1H),
7.75 (d, J = 8.5 Hz. 1H), 7.83 (m, 1H), 7.85 (s, 1H), 10.96 (s, 1H),
12.56 (v br s, 1H). MS (ESI-): mass calcd for C26H20Cl2N2O3S2,
542.0; m/z found 541.6 (M - 1). LCMS 98.5%. HPLC system 1:
99.8%, tR = 12.16 min.
4,5-Dichlorothiophene-2-sulfonic Acid [3-(1-Methyl-3-naph-
thalen-2-ylmethyl-1H-indol-4-yl)propionyl]amide (29). To a sus-
pension of the propionic acid 86 (120 mg, 0.34 mmol) in CH2Cl2
(2 mL) was sequentially added 4,5-dichlorothiophene-2-sulfon-
amide (78 mg, 0.34 mmol), DMAP (85 mg, 0.69 mmol), and
EDCI (132 mg, 0.69 mmol). The mixture was stirred at room
temperature overnight. The solution was acidified with 10%
aqueous HCl and extracted with EtOAc (2 ꢀ 5 mL). The organic
layer was washed with water, dried over anhydrous MgSO4,
filtered, and concentrated in vacuo to afford 110 mg of a residue.
This residue was triturated with MeOH to provide 80 mg (42%)
of the compound 29. 1H NMR (500 MHz, DMSO-d6) δ 2.52 (t,
J = 8.0 Hz, 2H), 3.00 (t, J = 8.0 Hz, 2H), 3.71 (s, 3H), 4.28 (s,
2H), 6.62 (d, J = 7.0 Hz, 1H), 6.95 (t, J = 7.5 Hz, 1H), 7.01 (s,
1H), 7.23 (d, J = 8.5 Hz, 1H), 7.33 (d, J = 8.5 Hz, 1H), 7.44 (m,
2H), 7.57 (s, 1H), 7.73 (m, 1H), 7.77 (d, J = 8.5 Hz, 1H), 7.85 (m,
2H), 12.6 (br s, 1H). MS (ESI-): mass calcd for C27H22Cl2-
N2O3S2, 556.0; m/z found 554.9 (M - 1). LCMS 96.4%.
4,5-Dichlorothiophene-2-sulfonic Acid [2-(3-Naphthalen-2-yl-
methyl-1H-indol-4-yloxy)acetyl]amide (30). To the solution of
the acid 92 (2.87 g, 8.65 mmol, 1 equiv) in CH2Cl2 (80 mL) was
added DMAP (2.11 g, 17.3 mmol, 2 equiv), 4,5-dichloro-2-
thiophenesulfonamide (2.11 g, 9.1 mmol, 1.05 equiv), and EDCI
(3.22 g, 17.3 mmol, 2 equiv). The mixture was stirred at room
temperature for 16 h and then quenched with 10% aqueous HCl
(5 mL) followed by dilution with EtOAc (100 mL). The aqueous
layer was extracted with EtOAc (2 ꢀ 50 mL). The combined
organic layers were washed with saturated aqueous NH4Cl
(100 mL), brine (2 ꢀ 100 mL), and then dried over anhydrous
MgSO4, filtered and concentrated in vacuo to afford a residue
Thiophene-2-sulfonic Acid [(E)-3-(1-Pyridin-3-yl)-1H-indol-7-
yl)acryloyl]amide (23). Compound 23 was prepared from 71
using general procedure B. Yield: 68%. H NMR (400 MHz,
1
DMSO-d6), δ 5.63 (s, 2H), 6.28 (d, J = 15.2 Hz, 1H), 6.61 (d,
J = 2.8 Hz, 1H), 7.08 (t, J = 7.6 Hz, 1H), 7.19 (m, 3H), 7.31 (m,
1H), 7.59 (d, J = 3.2 Hz, 1H), 7.68 (d, J = 7.6 Hz, 1H), 7.80 (m,
1H), 8.03 (m, 1H), 8.08 (d, J = 15.2 Hz, 1H), 8.12 (m, 1H), 8.40
(m, 1H), 12.38 (br s, 1H). MS (APCIþ): mass calcd for
C21H17N3O3S2, 423.1; m/z found 424.1 (M þ 1). LCMS 97%.
HPLC system 1: 95%, tR = 9.57 min.
Thiophene-2-sulfonic Acid [(E)-3-(3-Methyl-1-pyridin-3-yl-
methyl-1H-indol-7-yl)acryloyl]amide (24). Compound 24 was
prepared from 75 using general procedure B. H NMR (500
1
MHz, CDCl3) δ 2.34 (s, 3H), 5.47 (s, 2H), 6.23 (d, J = 15.0 Hz,
1H), 6.92 (s, 1H), 7.12 (m, 2H), 7.24 (m, 1H), 7.46 (d, J = 8.0 Hz,
1H), 7.60 (d, J = 5.0 Hz, 1H), 7.64 (d, J = 7.5 Hz, 1H), 7.69 (m,
1H), 7.92 (d, J = 3.5 Hz, 1H), 8.23 (d, J = 15.0 Hz, 1H), 8.24 (s,
1H), 8.48 (d, J = 3.5 Hz, 1H). MS (ESI-): mass calcd for
C22H19N3O3S2, 437.1; m/z found 436.4 (M - 1). LCMS 97%.
HPLC system 1: 96%, tR = 10.16 min.
Thiophene-2-sulfonic Acid [(E)-3-(3-Methyl-1-pyridin-2-yl-
methyl-1H-indol-7-yl)acryloyl]amide (25). Compound 25 was
prepared from 75 using general procedure B. Yield: 58%. H
1
NMR (400 MHz, CDCl3) δ 2.27 (s, 3H), 5.55 (s, 2H), 6.26 (d,
J = 15.2 Hz, 1H), 6.77 (d, J = 7.6 Hz, 1H), 7.08 (t, J = 7.6 Hz,
1H), 7.15-7.21 (m, 2H), 7.24 (dd, J = 4.8, 4.4 Hz, 1H), 7.28 (s,
1H), 7.56-7.63 (m, 2H), 7.81 (dd, J = 4.0, 1.6 Hz, 1H), 8.07 (dd,
J = 5.2, 1.6 Hz, 1H), 8.12 (d, J = 15.2 Hz, 1H), 8.39 (m, 1H),
12.25 (br s, 1H). MS (APþ): mass calcd for C22H19N3O3S2,
437.1; m/z found 438.1 (M þ 1). LCMS 97.5%.
4,5-Dichlorothiophene-2-sulfonic Acid [(E)-3-(3-Naphthalen-
2-ylmethyl-1H-indol-4-yl)acryloyl]amide (26). To a suspension
of acid 80 (26.3 mg, 0.08 mmol) in CH2Cl2 (1 mL) was sub-
sequently added 4,5-dichlorothiophene-2-sulfonamide (19 mg,
0.082 mmol), DMAP (20 mg, 0.16 mmol), and EDCI (31 mg,
0.16 mmol). The mixture was stirred at room temperature
overnight. The solution was diluted with EtOAc (5 mL) and
acidified with 10% aqueous HCl. The organic layer was washed
with water, dried over ahydrous MgSO4, filtered, and concen-
trated in vacuo. The residue was purified by a column chromato-
graphy using 15-25% EtOAc/hexanes gradient elution to yield
18 mg (42%) of acylsulfonamide 26. 1H NMR (500 MHz,
DMSO-d6) δ 4.34 (s, 2H), 6.4 (d, J = 16.0, 1H), 7.11 (t, J = 7.5
Hz, 1H), 7.21 (d, J = 7.0 Hz, 1H), 7.36-7.42 (m, 4H), 7.46 (d,
J = 8.0 Hz, 1H), 7.63 (d, J = 7.0 Hz, 1H), 7.68 (d, J = 8.5 Hz,
1H), 7.77 (d, J = 7.0 Hz, 1H), 7.79 (s, 1H), 7.94 (s, 1H), 8.40 (d,
J = 16.0 Hz, 1H), 11.20 (s, 1H), 12.5 (br s, 1H). MS (ESI-): mass

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 29
(3.31 g, 63%). This residue was triturated with MeOH (12 mL),
heated to reflux, then cooled to 0 °C and filtered to provide
2.97 g (71%) of the sulfonamide 30. 1H NMR (500 MHz,
DMSO-d6) δ 4.33 (s, 2H), 4.68 (s, 2H), 6.18 (d, J = 7.5 Hz,
1H), 6.87 (t, J = 8.0 Hz, 1H), 6.94 (d, J = 8.5 Hz, 1H), 7.01 (d,
J = 2.5 Hz, 1H), 7.41 (m, 2H), 7.46 (dd, J = 8.5, 1.5 Hz, 1H),
7.72 (d, J = 8.5 Hz, 2H), 7.79-7.81 (m, 2H), 7.89 (s, 1H), 10.92
(br s, 1H). MS (ESI-): mass calcd for C25H18Cl2N2O4S2, 544.0;
m/z found 543.3 (M - 1). HPLC system 1: 97.2%, tR = 16.65
min. Anal. C, H, N, S.
Thiophene-2-sulfonic Acid [(E)-3-(5-Fluoro-3-methyl-1-naph-
thalen-2-ylmethyl-1H-indol-7-yl)acryloyl]amide (43). Compound
43 was prepared from intermediate compound 96a using
general procedure B. To a solution of compound 96a (45 mg,
0.012 mmol) in DMF (3 mL), NaH (60% in oil, 15 mg, 0.370 mmol)
was added at 0 °C and stirred at room temperature for 1 h
followed by addition of 2-(bromomethyl)naphthalene (53 mg,
0.24 mmol). The reaction mixture was stirred at room tempera-
ture overnight and diluted with CH2Cl2 (12 mL). The reaction
mixture was washed with dilute aqueous HCl (2 ꢀ 8 mL), water (4
ꢀ 8 mL), brine, dried over anhydrous Na2SO4, and filtered. The
solvent was concentrated in vacuo, and the residue was washed
with Et2O/hexane to give 55 mg (90%) of the compound 43. 1H
NMR (500 MHz, DMSO-d6) δ 2.28 (s, 3H), 5.65 (s, 2H), 6.28 (d,
J = 15.5 Hz, 1H), 6.99 (dd, J = 10.0, 2.5 Hz, 1H), 7.09 (dd, J =
8.5, 1.5 Hz, 1H), 7.26 (dd, J = 5.0, 4.0 Hz, 1H), 7.40-7.48 (m,
4H), 7.51 (s, 1H), 7.60 (dd, J = 9.0, 1.5 Hz, 1H), 7.73 (d, J = 8.0
Hz, 1H), 7.81 (d, J = 7.5 Hz, 1H), 7.86 (dd, J = 3.5, 1.5 Hz, 1H),
8.10 (dd, J = 5.0, 1.0 Hz, 1H), 8.19 (d, J = 15.5Hz, 1H), 12.30 (br
s, 1H). MS (ESI-): mass calcd for C27H21FN2O3S2, 504.1; m/z
found 503.4 (M - 1). LCMS 96%. HPLC system 2: 96.7%, tR =
17.67 min.
4,5-Dichlorothiophene-2-sulfonic Acid {(E)-3-[1-(3-Cyanob-
enzyl)-5-fluoro-3-methyl-1H-indol-7-yl]acryloyl}amide (47). Com-
pound 96b (250 mg, 0.58 mmol) was alkylated using NaH (60%)
(69.2 mg, 1.73 mmol) and 3-(bromomethyl)benzonitrile (192 mg,
0.98 mmol) in anhydrous THF (2.8 mL) in a manner analogous to
the preparation of compound 43. After filtration, 186 mg (59%) of
1
compound 47 was isolated as a lime-green solid. H NMR (400
MHz, DMSO-d6) δ 2.26 (s, 3H), 5.59 (s, 2H), 6.32 (d, J = 15.6 Hz,
1H), 7.05 (dd, J = 10.0, 2.4 Hz, 1H), 7.22 (d, J = 7.6 Hz, 1H), 7.37
(br s, 1H), 7.40-7.47 (m, 3H), 7.69 (d, J = 7.6 Hz, 1H), 7.90
(s, 1H), 7.94 (d, J=15.6 Hz, 1H). MS (ESI-): mass calcd for
C24H16Cl2FN3O3S2, 547.0; m/z found 546.2 (M - 1). HPLC
system 2: 99.4%, tR = 17.81 min.
4,5-Dichlorothiophene-2-sulfonic Acid {(E)-3-[1-(4-Cyano-
benzyl)-5-fluoro-3-methyl-1H-indol-7-yl]acryloyl}amide (48). Com-
pound 96b (250 mg, 0.58 mmol) was alkylated using NaH
(60%) (69.2 mg, 1.73 mmol) and 4-(bromomethyl)benzonitrile
(192 mg, 0.98 mmol) in anhydrous THF (2.8 mL) in a manner
analogous to the preparation of compound 43. After filtration,
90.8 mg (29%) of compound 49 was isolated as a lime-green
solid. 1H NMR (400 MHz, DMSO-d6) δ 2.26 (s, 3H), 5.64
(s, 2H), 6.22 (d, J = 15.2 Hz, 1H), 7.03 (m, 3H), 7.43 (m, 2H),
7.62(d,J= 8.8 Hz, 2H), 7.91-7.95 (m, 2H). MS (ESI-):masscalcd
for C24H16Cl2FN2O3S2, 547.0; m/z found 546.2 (M - 1). HPLC
system 1: 98.3%, tR = 16.85 min. HPLC system 2: 99.3%, tR
=
17.73 min.
4,5-Dichlorothiophene-2-sulfonic Acid [(E)-3-(5-Fluoro-1-
imidazo[1,2-a]pyridin-2-ylmethyl-3-methyl-1H-indol-7-yl)acry-
loyl]amide (49). Compound 49 was prepared through the alkyla-
tion of indole derivative 96b with 2-chloromethylimidazo[1,2-
1
a]pyridine according to general procedure B. Yield: 72%. H
NMR (400 MHz, DMSO-d6) δ 2.22 (s, 3H), 5.49 (s, 2H), 6.24 (d,
J = 15.6 Hz, 1H), 6.87 (ddd, J = 6.8, 6.8, 1.2 Hz, 1H), 7.04 (dd,
J = 10.4, 2.4 Hz, 1H), 7.18-7.27 (m, 2H), 7.36 (s, 1H),
7.45-7.50 (m, 2H), 8.22 (d, J = 15.6 Hz, 1H), 8.27 (s, 1H),
8.42 (m, 1H). MS (ESI-): mass calcd for C24H17Cl2FN4O3S2,
Thiophene-2-sulfonic Acid {(E)-3-[1-(2,4-Dichlorobenzyl)-5-
fluoro-3-methyl-1H-indol-7-yl]acryloyl}amide (44). Compound
44 was prepared from 96a by a procedure similar to that
described for the synthesis of compound 43. Yield 87% 1H
NMR (500 MHz, DMSO-d6) δ 2.25 (s, 3H), 5.52 (s, 2H), 6.15 (d,
J = 8.0 Hz, 1H), 6.25 (d, J = 15.0 Hz, 1H), 7.02 (dd, J = 10.5,
2.0 Hz, 1H), 7.22 (m, 2H), 7.33 (s, 1H), 7.43 (dd, J = 10.0, 2.0
Hz, 1H), 7.51 (m, 1H), 7.66 (d, J = 15.0 Hz, 1H), 7.76 (m, 1H),
8.01 (d, J = 4.5 Hz, 1H), 12.4 (br s, 1H). MS (ESI-): mass calcd
for C23H17Cl2N2O3S2, 522.0; m/z found 521.6 (M - 1). HPLC
system 2: 99.4%, tR = 17.76 min.
562.0; m/z found 561.1 (M - 1). HPLC system 1: 95.1%, tR
22.32 min
=
4,5-Dichlorothiophene-2-sulfonic Acid [(E)-3-(5-Fluoro-3-
methyl-1-naphthalen-2-ylmethyl-1H-indol-7-yl)acryloyl]amide (51).
Compound 96b (52 mg, 0.12 mmol) was alkylated using NaH
(60% in oil, 15 mg, 037 mmol) and 2-(bromomethyl)-
naphthalene (53 mg, 0.24 mmol) in anhydrous DMF (3 mL),
in a manner analogous to the preparation of compound 43. The
crude product was purified by column chromatography on silica
gel with 1-5% MeOH/CH2Cl2 as eluent to give 44 mg (64%) of
compound 51. 1H NMR (400 MHz, DMSO-d6) δ 2.29 (s, 3H),
5.68 (s, 2H), 6.28 (d, J = 15.2 Hz, 1H), 7.02 (dd, J = 10.0, 2.0
Hz, 1H), 7.10 (m, 1H), 7.40-7.50 (m, 3H), 7.54 (m, 2H), 7.64 (d,
J = 7.2 Hz, 1H), 7.74 (d, J = 8.8 Hz, 1H), 7.82 (d, J = 8.0 Hz,
1H), 7.94 (s, 1H), 8.21 (d, J = 15.2 Hz, 1H), 12.50 (br s, 1H). MS
(ESI-): mass calcd for C27H19Cl2FN2O3S2, 572.0; m/z found
571.3 (M - 1). LCMS 98%. HPLC system 2: 99.6%, tR = 17.81
min.
4,5-Dichlorothiophene-2-sulfonic Acid {2-[3-(Naphthalene-2-
carbonyl)-1H-indol-4-yloxy]acetyl}amide (45). To an 8 mL vial
equipped with a magnetic stir bar and a screw cap was added
compound 30 (58.5 mg, 0.107 mmol) followed by anhydrous
THF (1 mL) and water (0.050 mL). To this stirring light-yellow
solution was added DDQ (73 mg, 0.322 mmol) in one portion,
immediately turning the solution black. The mixture was al-
lowed to stir at room temperature overnight. The solution was
concentrated to dryness and purified via preparative TLC
utilizing 7:3 hexanes/EtOAc as eluent to give 9 mg (15%) of
1
an off-white solid. H NMR (400 MHz, DMSO-d6) δ 4.61 (s,
2H), 6.66 (d, J = 7.2 Hz, 1H), 7.12-7.20 (m, 2H), 7.56-7.70 (m,
3H), 7.90 (d, J = 1.2 Hz, 1H), 7.94 (dd, J = 8.8, 1.2 Hz, 1H), 8.02
(d, J = 8.4 Hz, 1H), 8.05 (d, J = 8.8 Hz, 1H), 8.09 (d, J = 7.6 Hz,
1H), 8.41 (s, 1H), 12.17 (br s, 1H). MS (APþ) mass calcd for
C25H16Cl2N2O5S2, 558.0; m/z found 559.0 (M þ 1). LCMS
98.0%.
4,5-Dichlorothiophene-2-sulfonic Acid {(E)-3-[5-Fluoro-1-(4-
fluorobenzyl)-3-methyl-1H-indol-7-yl]acryloyl}amide (46). The
target compound was prepared from 96b using general proce-
dure A. Yield 57%. 1H NMR (400 MHz, DMSO-d6) δ 2.25 (d,
J = 0.8 Hz, 3H), 5.51 (s, 2H), 6.28 (d, J = 15.0 Hz, 1H),
7.00-6.91 (m, 5H), 7.03 (dd, J = 10.0, 2.5 Hz, 1H), 7.41 (m, 2H),
7.90 (s, 1H), 8.03 (d, J = 15.0 Hz, 1H). MS (ESI-) mass calcd for
C23H16Cl2F2N2O3S2, 540.0; m/z found 539.4 (M - 1). LCMS
97%. HPLC system 1: 98.3%, tR = 17.79 min. HPLC system 2:
98.9%, tR = 17.81 min.
N-{(E)-3-[1-(2,4-Dichlorobenzyl)-5-fluoro-3-methyl-1H-indol-
7-yl]acryloyl}-3,5-difluorobenzenesulfonamide (52). Compound
52 was synthesized from 96c following general procedure B.
Yield: 55%. 1H NMR (500 MHz, acetone-d6) δ 2.30 (s, 3H), 5.54
(s, 2H), 6.28 (d, J = 8.0 Hz, 1H), 6.41 (d, J = 15.5 Hz, 1H), 6.86
(br s, 1H), 7.03 (dd, J = 10.5, 2.5 Hz, 1H), 7.17 (dd, J = 8.0, 2.0
Hz, 1H), 7.28 (s, 1H), 7.23-7.39 (m, 2H), 7.49-7.53 (m, 2H),
7.68-7.70 (m, 1H), 7.84 (d, J = 15.5 Hz, 1H). MS (ESI-): mass
calcd for C25H17Cl2FN2O3S, 552.0; m/z found 551.3 (M - 1).
HPLC system 1: 96.1%, tR=17.68 min. HPLC system 2: 97.2%,
t
R = 17.8 min.
N-{(E)-3-[1-(3,4-Difluorobenzyl)-5-fluoro-3-methyl-1H-indol-
7-yl]acryloyl}-3,5-difluorobenzenesulfonamide (53). Compound
53 was synthesized from 96c following general procedure B.
1
Yield: 73%, as a yellow solid. H NMR (400 MHz, CDCl3) δ
2.29 (d, J = 1.2 Hz, 3H), 5.34 (s, 2H), 6.11 (d, J = 15.2 Hz, 1H),

30 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Singh et al.
6.69 (ddd, J = 9.6, 7.2, 2.0 Hz, 1H), 6.80 (m, 1H), 6.95 (s, 1H),
6.97 (dd, J = 9.6, 2.4 Hz, 1H), 7.03 (ddd, J = 10.0, 8.0, 8.0 Hz,
1H), 7.12 (dddd, J = 10.8, 8.4, 2.4, 2.4 Hz, 1H), 7.29 (dd, J =
8.4, 2.4 Hz, 1H), 7.64-7.70 (m, 2H), 8.09 (dd, J = 15.2 Hz, J =
1.2 Hz, 1H), 8.23 (br s, 1H). MS (ESI-): mass calcd for
C25H17F5N2O3S, 520.1; m/z found 519.7. LCMS 98%. HPLC
system 2: 99%, tR = 17.72 min.
58 was prepared via the alkylation of indole derivative 96d with
3-methoxybenzyl bromide according to general procedure B.
Yield: 19%. 1H NMR (400 MHz, DMSO-d6) δ 2.25 (s, 3H), 3.66
(s, 3H), 5.43 (s, 2H), 6.37 (dd, J = 11.2, 4.0 Hz, 2H), 6.51 (dd,
J = 2.0, 1.6 Hz, 1H), 6.74 (dd, J = 8.4, 2.0 Hz, 1H), 7.03 (dd,
J = 10.4, 2.4 Hz, 1H), 7.07 (dd, J = 8.0, 7.6 Hz, 1H), 7.40 (s,
1H), 7.43 (dd, J = 8.8, 2.4 Hz, 1H), 7.76 (m, 1H), 7.86 (m, 1H),
7.97-8.03 (m, 2H), 12.50 (br s, 1H). LCMS 97%. MS (ESI-):
mass calcd for C26H21F3N2O4S, 514.1; m/z found 513.7 (M - 1).
HPLC system 1: 98.8%, tR = 16.37 min.
N-{(E)-3-[1-(3,5-Dimethylisoxazol-4-ylmethyl)-5-fluoro-3-
methyl-1H-indol-7-yl]acryloyl}-3,4-difluorobenzenesulfonamide (59).
Compound 59 was prepared through the alkylation of indole
derivative 96e with 4-bromomethyl-3,5-dimethylisoxazole accord-
ing to general procedure B, except KOtBu was used as the base.
Yield: 31%. 1H NMR (400 MHz, DMSO-d6) δ 1.71 (s, 3H), 1.77
(s, 3H), 2.21 (s, 3H), 5.29 (s, 2H), 6.38 (d, J = 15.2 Hz, 1H), 7.08
(dd, J = 10.4, 2.4 Hz, 1H), 7.21 (s, 1H), 7.43 (dd, J = 8.8, 2.8 Hz,
1H), 7.77 (m, 1H), 7.88 (m, 1H), 8.00 (d, J = 2.8 Hz, 1H), 8.03
(m, 1H), 12.22 (s, 1H). MS (ESI-): mass calcd for C24H20F3N3O4S,
N-{(E)-3-[1-(2,4-Dichlorobenzyl)-5-fluoro-3-methyl-1H-indol-
7-yl]acryloyl}-2,4,5-trifluorobenzenesulfonamide (54). In an
8 mL vial equipped with a stir bar was placed (2E)-3-[1-(2,4-
dichlorobenzyl)-5-fluoro-3-methyl-1H-indol-7-yl]prop-2-enoic
acid (41 mg, 0.11 mmol), 2,4,5-trifluorobenzenesulfonamide
(23.2 mg, 0.11 mmol), DMAP (31.6 mg, 0.26 mmol), anhydrous
CH2Cl2 (2 mL), and EDCI (51.8 mg, 0.27 mmol) at room
temperature. The vial was sealed with a cap and allowed to
react for 22 h at room temperature. The contents of the vial were
washed with 1 M aqueous HCl (5 mL), water (3 ꢀ 5 mL) and
then brine (5 mL). The organic layer was dried over anhydrous
MgSO4 and concentrated in vacuo to a light-greenish solid. This
crude material was dissolved in boiling Et2O (1 mL) followed by
the addition of hexanes (1 mL). The resulting solid was collected
by suction filtration to provide 23.9 mg (39%) of compound 55
503.1; m/z found 502.4 (M - 1). HPLC system 1: 98.2%, tR
15.19 min.
=
1
as a lime-green solid. H NMR (400 MHz, CDCl3) δ 2.29 (s,
3,4-Difluoro-N-[(E)-3-(5-fluoro-1-imidazo[1,2-a]pyridin-2-yl-
methyl-3-methyl-1H-indol-7-yl)acryloyl]benzenesulfonamide (60).
Compound 60 was prepared through the alkylation of indole
derivative 96d and 2-chloromethylimidazo[1,2-a]pyridine ac-
cording to general procedure B. Yield: 12%. 1H NMR (400
MHz, DMSO-d6) δ 2.25 (s, 3 H), 5.79 (s, 2 H), 6.34 (d, J = 16 Hz,
1 H), 7.08 (dd, J = 10.0, 2.0 Hz, 1 H), 7.38 (s, 1 H), 7.43 (t, J = 2
Hz, 1 H), 7.49 (dd, J = 9.0, 2.0 Hz, 1 H), 7.72-8.02 (m, 7 H),
8.69 (d, J = 7.0 Hz, 1 H). MS (ESI-): mass calcd for C26H19-
F3N4O3S, 524.1; m/z found 523.6 (M - 1). LCMS 97%. HPLC
system 1: 97.6%, tR = 14.96 min.
4-Fluoro-N-[(E)-3-(5-fluoro-1-imidazo[1,2-a]pyridin-2-yl-
methyl-3-methyl-1H-indol-7-yl)acryloyl]benzenesulfonamide (61).
Compound 61 was prepared through the alkylation of indole
derivative 96f with 2-chloromethylimidazo[1,2-a]pyridine ac-
cording to general procedure B. Yield: 67%. 1H NMR (400
MHz, DMSO-d6) δ 2.24 (s, 3 H), 5.74 (s, 2 H), 6.39 (d, J =
16.0 Hz, 1 H), 7.07 (dd, J = 10.0, 2.0 Hz, 1 H), 7.32-7.36 (m, 1
H), 7.38 (s, 1 H), 7.45-7.52 (m, 3 H), 7.79 (d, J = 3.0 Hz, 2 H),
7.81 (s, 1 H), 8.00-8.07 (m, 3 H), 8.63 (d, J = 7.0 Hz, 1 H). MS
(ESI-): mass calcd for C26H20F2N4O3S, 506.1; m/z found 506.0.
(M - 1). LCMS 96%.
3,5-Difluoro-N-[(E)-3-(5-fluoro-1-imidazo[1,2-a]pyridin-2-yl-
methyl-3-methyl-1H-indol-7-yl)acryloyl]benzenesulfonamide (62).
Compound 62 was prepared through the alkylation of indole
derivative 96c and 2-chloromethylimidazo[1,2-a]pyridine ac-
cording to general procedure B. Yield: 68%. 1H NMR (400
MHz, DMSO-d6) δ 2.24 (s, 3 H), 5.77 (s, 2 H), 6.44 (d, J = 15 Hz,
1 H), 7.09 (dd, J = 10.0, 2.0 Hz, 1 H), 7.34-7.37 (m, 1 H), 7.40
(s, 1 H), 7.47 (dd, J = 9.0, 2.0 Hz, 1 H), 7.62-7.65 (m, 2 H),
7.72-7.80 (m, 3 H), 7.86 (s, 1 H), 8.07 (d, J = 15 Hz, 1 H), 8.66
(d, J = 7 Hz, 1 H). MS (ESI-): mass calcd for C26H19F3N4O3S,
3H), 5.37 (s, 2H), 6.18 (d, J = 15.2 Hz, 1H), 6.31 (d, J = 8.4 Hz,
1H), 6.87 (s, 1H), 6.82 (dd, J = 10.0, 2.8 Hz, 1H), 7.04 (dd, J =
2.0, 8.4 Hz, 1H), 7.07-7.13 (m, 1H), 7.31 (dd, J = 8.8, 2.8 Hz,
1H), 7.41 (d, J = 2.4 Hz, 1H), 7.82 (d, J = 14.8 Hz, 1H),
7.98-8.04 (m, 1H). MS (ESI-): mass calcd for C25H16Cl2F4-
N2O3S, 570.0; m/z found 569.4 (M - 1). LCMS 97%. HPLC
system 1: 97.9%, tR =17.64 min. HPLC system 2: 99.6%, tR
=
17.77 min.
N-{(E)-3-[1-(3,4-Difluorobenzyl)-5-fluoro-3-methyl-1H-indol-
7-yl]acryloyl}-3,4-difluorobenzenesulfonamide (55). Compound
55 was prepared via the alkylation of indole derivative 96d with
3,4-difluorobenzyl bromide according to general procedure B.
Yield: 73%. 1H NMR (500 MHz, CDCl3) δ 2.29 (s, 3H), 5.33 (s,
2H), 6.09 (d, J = 15.0 Hz, 1H) 6.71 (m, 1H), 6.77 (m, 1H),
6.94-6.97 (m, 2H), 7.03 (dd, J = 10.0, 8.5 Hz, 1H), 7.29 (dd, J =
8.0, 2.5 Hz, 1H), 7.36 (dd, J = 16.5, 9.0 Hz, 1H), 7.92 (m, 1H),
7.99 (m, 1H), 8.07 (d, J = 15.0 Hz, 1H), 8.40 (br s, 1H). MS
(ESI-): mass calcd for C25H17F5N2O3S, 520.1; m/z found 519.8
(M - 1). HPLC system 1: 97.9%, tR = 17.80 min. HPLC system
2: 97.8%, tR = 17.84 min.
Synthesis of 4,5-Dichlorothiophene-2-sulfonic Acid {(E)-
3-[1-(3,4-Difluorobenzyl)-5-fluoro-3-methyl-1H-indol-7-yl]acry-
loyl}amide (56). Compound 96b (70 mg, 0.16 mmol) was alky-
lated using NaH (60%) (8.5 mg, 0.36 mmol) and 3,4-difluoroben-
zyl bromide (50 mg, 0.24 mmol) in anhydrous DMF (2.8 mL) in a
manner analogous to the preparation of compound 43. After
filtration and column chromatography, 50 mg (55%) of com-
1
pound 56 was isolated as a yellow solid . H NMR (400 MHz,
DMSO-d6) δ 2.25 (d, J = 0.8 Hz, 3H), 5.51 (s, 2H), 6.28 (d, J =
15.6 Hz, 1H), 6.66 (m, 1H), 6.99 (ddd, J = 10.0, 7.6, 2.0 Hz, 1H),
7.04 (dd, J = 10.0, 2.4 Hz, 1H), 7.23 (ddd, J = 10.8, 8.2, 8.2 Hz,
1H), 7.41 (s, 1H), 7.43 (dd, J = 8.8, 2.4 Hz, 1H), 7.87 (s, 1H), 7.98
(d, J = 15.6 Hz, 1H). MS (ESI-): mass calcd for C23H15Cl2F3-
N2O3S, 558.0; m/z found 557.2 (M - 1). LCMS 96.2%.
524.1; m/z found 523.6 (M - 1). HPLC system 1: 98.1%, tR
10.96 min.
=
N-{(E)-3-[1-(2,3-Dihydrobenzo[1,4]dioxin-6-ylmethyl)-5-fluoro-
3-methyl-1H-indol-7-yl]acryloyl}-C,C,C-trifluoromethanesulfo-
namide (63). A mixture of the acid 99 (0.100 g, 0.27 mmol),
EDCI (0.104 g, 0.54 mmol), DMAP (0.066 g, 0.54 mmol), and
CF3SO2NH2 (0.049 g, 0.33 mmol) in anhydrous CH2Cl2 (10 mL)
was stirred overnight at room temperature. The reaction mix-
ture was diluted with CH2Cl2 (10 mL). The solution was
successively washed with 1 N HCl (2 ꢀ 5 mL), brine, dried over
anhydrous Na2SO4, and concentrated in vacuo. The residue was
chromatographed by flash silica gel (4 g) and eluted with 1%
MeOH in CH2Cl2 to yield 0.050 g (37%) of compound 63. 1H
NMR (400 MHz, DMSO-d6) δ 2.23 (s, 3H), 4.16 (m, 4H), 4.70
(br s, 1H, w /water peak), 5.34 (s, 2H), 6.28 (d, J = 15.6 Hz, 1H),
6.38 (d, J = 2.0 Hz, 1H), 6.61 (dd, J = 8.4, 2.0 Hz, 1H), 6.74 (d,
2,4,5-Trifluoro-N-{(E)-3-[5-fluoro-1-(3-methoxybenzyl)-3-
methyl-1H-indol-7-yl]acryloyl}benzenesulfonamide (57). Com-
pound 57 was prepared via the alkylation of indole derivative
96d with 3-methoxybenzyl bromide according to general proce-
dure B. Yield: 30%. 1H NMR (400 MHz, DMSO-d6) δ 2.25 (s,
3H), 3.64 (s, 3H), 5.42 (s, 2H), 6.34 (s, 1H), 6.38 (d, J = 15.2 Hz,
1H), 6.49 (t, J = 2.0 Hz, 1H), 6.73 (dd, J = 8.4, 2.8 Hz, 1H),
7.02-7.09 (m, 2H), 7.40 (s, 1H), 7.43 (dd, J = 8.8, 2.4 Hz, 1H),
7.92-8.07 (m, 3H). MS (ESI-): mass calcd for C26H20F4N2O4S,
532.1; m/z found 531.5 (M - 1). HPLC system 2: 98.8%, tR
=
17.63 min.
3,4-Difluoro-N-{(E)-3-[5-fluoro-1-(3-methoxybenzyl)-3-meth-
yl-1H-indol-7-yl]acryloyl}benzenesulfonamide (58). Compound

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 31
J = 8.4 Hz, 1H), 7.08 (dd, J = 10.4, 2.4 Hz, 1H), 7.26-7.34 (m,
2H), 7.94 (d, J = 15.6 Hz, 1H). MS (APCI-): mass calcd for
C22H18F4N2O5S, 498.1; m/z found 497.0 (M - 1). LCMS
98.6%. HPLC system 1: 98.7%, tR = 15.57 min.
(ESIþ): mass calcd for C13H13NO2, 215.1; m/z found 216.3 (M þ
1). LCMS 97%. HPLC system 1: 98%, tR = 14.32 min.
3-(1H-Indol-7-yl)acrylic Acid (70). To a round-bottom flask
(500 mL) that contained a solution of NaOH (1.2 g, 30 mmol) in
EtOH (100 mL) and H2O (30 mL) was added the ester 69 (3.2 g,
15 mmol) at 5 °C. The resulting mixture was allowed to warm to
room temperature and stir for 10 min, then heated to 78 °C and
stirred for 4 h. The reaction mixture was cooled to 5 °C and was
acidified with addition of aqueous HCl (10%) until pH 1, which
was followed by extraction with CH2Cl2/MeOH (95/5, 3 ꢀ 150
mL). The combined organic layers were washed with brine (2 ꢀ
20 mL), dried over anhudrous Na2SO4, filtered, and the solvent
was removed in vacuo to provide a residue which was purified by
recrystallization from acetone/EtOAc/hexane to yield 2.4 g
4,5-Dichlorothiophene-2-sulfonic Acid {(E)-3-[1-(2,3-Dihy-
drobenzo[1,4]dioxin-6-ylmethyl)-5-fluoro-3-methyl-1H-indol-7-
yl]acryloyl}amide (64). Compound 64 was prepared through the
alkylation of indole derivative 96b with 6-bromomethyl-2,3-
dihydrobenzo[1,4]dioxine according to general procedure B.
1
Yield: 45%. H NMR (400 MHz, DMSO-d6) δ 2.23 (s, 3H),
4.14 (m, 4H), 5.36 (s, 2H), 6.32 (d, J = 0.8 Hz, 1H), 6.34 (d, J =
15.6 Hz, 1H), 6.45 (dd, J = 8.4, 2.0 Hz, 1H), 6.65 (d, J = 9.2 Hz,
1H), 7.0 (dd, J = 10.0, 2.4 Hz, 1H), 7.36 (s, 1H), 7.41 (dd, J =
8.8, 2.4 Hz, 1H), 7.90 (s, 1H), 8.08 (d, J = 15.6 Hz, 1H). MS
(APCI-): mass calcd for C25H19Cl2FN2O5S2, 580.0; m/z
found 578.9 (M - 1). LCMS 96.9%. HPLC system 1: 97.7%,
1
(86%) of the acrylic acid 70 as a white solid. H NMR (500
MHz, DMSO-d6) δ 6.58 (dd, J = 3.5, 1.5 Hz, 1H), 6.62 (d, J =
16.0 Hz, 1H), 7.06 (t, J = 7.5 Hz, 1H), 7.39 (t, J = 3.0 Hz, 1H),
7.51 (d, J = 7.5 Hz, 1H), 7.64 (d, J = 7.5 Hz, 1H), 8.12 (d, J =
16.5 Hz, 1H), 11.56 (s, 1H), 12.38 (s, 1H). 13C NMR (125 MHz,
acetone-d6) δ 103.3, 118.6, 119.3, 120.5, 122.1, 123.9, 126.6,
130.4, 135.7, 142.1, 168.4. MS (APCI-): mass calcd for
C11H9NO2, 187.1; m/z found 186.2 (M - 1). LCMS 99%. HPLC
system 1: 99%, tR = 11.35 min.
t
R =17.35 min. HPLC system 2: 99.3%, tR = 17.81 min.
Piperidine-1-sulfonic Acid {(E)-3-[1-(2,4-Dichlorobenzyl)-5-
fluoro-3-methyl-1H-indol-7-yl]acryloyl}amide (65). Compound
65 was prepared analogous to synthesis of 54, piperidine-1-
sulfonamide was used according to general procedure A. Yield:
49%. 1H NMR (400 MHz, DMSO-d6) δ 1.53 (m, 6H), 2.26 (s, 3
H), 3.17 (m, 4H), 5.57 (s, 2H), 6.15 (d, J = 8.4 Hz, 1H), 6.32 (d,
J = 15.6 Hz, 1H), 7.03 (dd, J = 10.4, 2.4 Hz, 1 H), 7.27 (dd, J =
8.4, 2.0 Hz, 1H), 7.37 (s, 1H), 7.46 (dd, J = 9.2, 2.4 Hz, 1H), 7.65
(d, J = 2.4 Hz, 1H), 7.72 (d, J = 15.6 Hz, 1H), 11.43 (s, 1H). MS
(ESI-): mass calcd for C24H24Cl2FN3O3S, 523.1; m/z found
Synthesis of Thiophene-2-sulfonic Acid ((E)-3-1H-Indol-7-yl-
acryloyl)amide (71). To a round-bottom flask (500 mL) that
contained a solution of 2-thiophene sulfonamide (1.05 g, 6 mmol),
DMAP (1.56 g, 13 mmol) and EDCI (2.4 g, 13 mmol) in CH2Cl2
(150 mL) was added the acrylic acid 70 (1.2 g, 6 mmol) at room
temperature. The resulting mixture was allowed to stir at room
temperature for 72 h, then was cooled to 5 °C and was acidified
with aqueous HCl (10%) until pH 1, which was followed by
extraction of CH2Cl2/MeOH (9/1, 3 ꢀ 100 mL). The combined
organic layers were dried over anhydrous Na2SO4 and the
solvent was removed in vacuo to provide the crude product
which was purified by flash chromatography (silica gel, CH2Cl2/
EtOAc/hexane = 1:10, 20:20:10) to afford 1.2 g, (56%) of the
acylsulfonamide 71 as a white solid. 1H NMR (500 MHz,
DMSO-d6) δ 6.47 (m, 1H), 6.67 (d, J = 16.0 Hz, 1H), 7.06 (t,
J = 7.5 Hz, 1H), 7.18 (t, J = 3.5 Hz, 1H), 7.37-7.39 (m, 2H),
7.63 (d, J = 7.5 Hz, 1H), 7.80 (d, J = 3.5 Hz, 1H), 7.98 (d, J =
4.5 Hz, 1H), 8.11 (d, J = 15.5 Hz, 1H), 11.54 (s, 1H). MS (ESI-):
mass calcd for C15H12N2O3S2, 332.0; m/z found 331.1 (M - 1).
LCMS 99%. HPLC system 1: 99%, tR = 12.95 min.
522.0 (M - 1). LCMS 99.1%. HPLC system 1: 99.3%, tR
17.88 min. HPLC system 2: 98.9%, tR = 17.47 min.
=
4,5-Dichlorothiophene-2-sulfonic Acid {3-[1-(2,4-Dichloro-
benzyl)-5-fluoro-3-methyl-1H-indol-7-yl]propionyl}amide (66).
To a solution of acid 101 (0.16 mmol, 1.0 equiv), 4,5-dichloro-
2-thiophenesulfonamide (0.16 mmol, 1.0 equiv), and DMAP
(0.38 mmol, 2.4 equiv) in CH2Cl2 was added EDCI (0.39 mmol,
2.5 equiv). The reaction mixture was stirred at room tempera-
ture for 16 h and acidified to pH 1-2 with 1 N HCl. The organic
layer was washed with successive portions of water and brine
and dried over anhudrous MgSO4. The filtrate was concentrated
in vacuo and the residue purified by flash chromato-
graphy on silica gel, using MeOH in CH2Cl2 as eluent, to afford
the title compound 66 in 64% yield. 1H NMR (400 MHz,
DMSO-d6) δ 2.22 (s, 3H), 2.49 (m, 2H), 2.76 (m, 2H), 5.49 (s,
2H), 6.46 (d, J = 7.6 Hz, 1H), 6.66 (dd, J = 10.8, 2.4 Hz, 1H),
7.14 (dd, J = 9.2, 2.8 Hz, 1H), 7.25 (s, 1H), 7.27 (dd, J = 8.4, 2.0
Hz, 1H), 7.52 (d, J = 2.0 Hz, 1H), 7.80 (s, 1H), 7.96 (s, 1H). MS
(ESI-): mass calcd for C23H17Cl4FN2O3S2, 594.3; m/z found
593.2 (M - 1). LC/MS 95.7%.
3-(1H-Indol-7-yl)acrylic Acid Ethyl Ester (69). To a round-
bottom flask (100 mL) that contained a suspension of NaH
(60% in mineral oil, 320 mg, 8 mmol) in THF (20 mL) was added
triethyl phosphonoacetate (1.5 g, 6.6 mmol) at 0 °C. The
resulting mixture was allowed to warm to room temperature,
stirred for 2 h, and then cooled to 0 °C. To this solution, indole-
7-carboxaldehyde, 68 (450 mg, 3 mmol), was added at 0 °C. The
reaction mixture was allowed to warm to room temperature,
stirred for 2 h, then heated and stirred at 78 °C for 14 h. The
reaction mixture was cooled to 5 °C, quenched with the addition
of saturated aqueous NH4Cl solution (15 mL) followed by
extraction with EtOAc (3 ꢀ 30 mL). The combined organic
layers were washed with brine (2 ꢀ 20 mL), dried over anhy-
drous Na2SO4, and filtered, and the solvent was removed in
vacuo. The residue was purified by flash chromatography with
gradient elution (silica gel, EtOAc/hexanes, 1:20 to 1:8) to afford
450 mg (68%) of compound 69 as a white solid. 1H NMR (500
MHz, CDCl3) δ 1.35 (t, J = 7.5 Hz, 3H), 4.30 (q, J = 7.5 Hz,
2H), 6.55 (d, J = 16.0 Hz, 1H), 6.58 (dd, J = 3.5, 2.0 Hz, 1H),
7.13 (t, J = 7.5 Hz, 1H), 7.19 (t, J = 3.0 Hz, 1H), 7.41 (d, J = 7.0
Hz, 1H), 7.69 (d, J = 7.5 Hz, 1H), 8.10 (d, J = 16.0 Hz, 1H), 8.97
(s, 1H). 13C NMR (125 MHz, CDCl3) δ 14.3, 60.5, 103.1, 117.9,
118.3, 120.0, 122.0, 123.3, 124.9, 128.9, 134.3, 141.2, 167.5. MS
7-Bromo-3-methyl-1H-indole (72). 2-Bromonitrobenzene was
reacted with allylmagnesium bromide according to the literature
1
procedure16 to provide compound 72 in 43% yield. H NMR
(400 MHz, CDCl3) δ 2.32 (d, J = 1.2 Hz, 3H), 7.00 (t, J = 7.6
Hz, 1H), 7.03 (m, 1H), 7.34 (ddd, J = 7.2, 0.4, 0.4 Hz, 1H), 7.52
(ddd, J = 7.2, 0.8, 0.8 Hz, 1H), 8.07 (br s, 1H). MS (ESIþ): mass
calcd for C9H8BrN, 209.0; m/z found 130.1 (M - Br). LCMS
99.2%.
(E)-3-(3-Methyl-1H-indol-7-yl)acrylic Acid Methyl Ester (73).
To a mixture of compound 72 (300 mg, 1.42 mmol) and methyl
acrylate (183 mg, 2.13 mmol) in Et3N (1 mL), palladium(II)
acetate (31 mg, 0.14 mmol) and tri-o-tolylphosphine (129 mg,
0.42 mmol) were added under argon at room temperature. The
reaction mixture was stirred at 100 °C for 4 h in a sealed pressure
tube and then cooled to room temperature. The reaction mixture
was diluted with CH2Cl2 (50 mL), washed with water (3 ꢀ 30
mL), brine, and dried over anhydrous Na2SO4. After removal of
solvent in vacuo, the residue was purified by column chromato-
graphy on silica gel with EtOAc/hexane as an eluent to give 250
1
mg (82%) of compound 73 as a yellow solid. H NMR (500
MHz, CDCl3) δ 2.34 (s, 3H), 3.84 (s, 3H), 6.49 (d, J = 16.0 Hz,
1H), 7.03 (s, 1H), 7.15 (t, J = 7.5 Hz, 1H), 7.41 (d, J = 7.5 Hz,
1H), 7.64 (d, J = 7.5 Hz, 1H), 8.01 (d, J = 16.0 Hz, 1H), 8.40 (br
s, 1H). MS (AP-): mass calcd for C13H13NO2, 215.1; m/z found
214.1 (M - 1). LCMS 99.6%.
(E)-3-(3-Methyl-1H-indol-7-yl)acrylic Acid (74). To a solu-
tion of compound 73 (180 mg, 0.84 mmol) in THF (5 mL) and

32 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Singh et al.
MeOH (4 mL), 2 N aqueous NaOH (4 mL) was added at room
temperature. The reaction mixture was stirred at room tempera-
ture overnight, and then the pH was adjusted to acidic by adding
2 N aqueous HCl. The reaction mixture was extracted with
EtOAc (2 ꢀ 30 mL). The combined organic phase was washed
with water and brine and dried over anhydrous Na2SO4. After
removal of solvent, 170 mg (100%) of compound 74 was
obtained. 1H NMR (500 MHz, DMSO-d6) δ 2.26 (s, 3H), 6.59
(d, J = 16 Hz, 1H), 7.05 (m, 1H), 7.15 (s, 1H), 7.50 (d, J = 8 Hz,
1H), 7.57 (d, J = 8 Hz, 1H), 8.08 (d, J = 16 Hz, 1H), 11.2 (s, 1H),
12.3 (br s, 1H). MS (AP-): mass calcd for C12H11NO2, 201.1;
m/z found 200.1 (M - 1). LCMS 95.3%.
Thiophene-2-sulfonic Acid [(E)-3-(3-Methyl-1H-indol-7-
yl)acryloyl]amide (75). A mixture of the acid 74 (170 mg, 0.84
mmol), 2-thiophenesulfonamide (163 mg, 1 mmol), DMAP (207
mg, 1.7 mmol) and EDCI (325 mg, 1.7 mmol) in CH2Cl2 (20
mL), and DMSO (0.5 mL) was stirred at room temperature
overnight. The solution was diluted with CH2Cl2, washed with
diluted aqueous HCl, water, brine, and dried over anhydrous
Na2SO4. After removal of solvent in vacuo, the residue was
purified by column chromatography on silica gel with MeOH/
CH2Cl2 as an eluent to give 170 mg (58%) of compound 75. 1H
NMR (500 MHz, DMSO-d6) δ 2.32 (s, 3H), 6.60 (d, J = 15.5 Hz,
1H), 7.02 (s, 1H), 7.08-7.14 (m, 2H), 7.38 (d, J = 8.0 Hz, 1H),
7.65 (m, 2H), 7.92 (m, 1H), 8.11 (d, J = 15.5 Hz, 1H), 8.54 (br s,
1H). MS (ESI-): mass calcd for C16H14N2O3S2, 346.0; m/z
found 345.1 (M - 1). LCMS 97.6%.
(4-Bromo-1H-indol-3-yl)naphthalen-2-ylmethanone (77). To a
solution of 4-bromoindole (Aldrich Chemical Co.) (76, 5.0 g,
25.5 mmol) in 100 mL of anhydrous CH2Cl2 was added 3 M
MeMgBr in Et2O (8.95 mL, 26.7 mmol) dropwise at 20 °C. A
slightly exothermic reaction was observed, and the temperature
of the mixture rose to 28 °C. The resulting orange solution was
stirred for 10 min at room temperature. Then 1 M ZnCl2
solution in Et2O (76.5 mL, 765 mmol) was added via an addition
funnel. The reaction mixture was stirred for 0.5 h. Then a
solution of 2-naphthoyl chloride (5.1 g, 26.7 mmol) in 25 mL
of CH2Cl2 was added. During the addition, the color changed
from light-orange to dark-red. The resulting mixture was stirred
at room temperature overnight. TLC (30% EtOAc/hexanes)
showed the reaction was complete, and then the mixture was
quenched with saturated solution of NH4Cl (100 mL). The
suspension was stirred for 15 min. The solid was filtered off
and washed several times with CH2Cl2. The filtrate was washed
with saturated NH4Cl, water, brine, dried over anhydrous
MgSO4, and concentrated in vacuo to afford 7 g of a residue.
The solid was taken into 10% aqueous HCl and was extracted
with EtOAc. The organic layer was washed with water, brine,
dried over anhydrous MgSO4, and concentrated in vacuo to give
about 500 mg of additional residue. The combined residues
(7.5 g) were washed with 15 mL of MTBE. Then the solvent was
decanted and then washed with 10 mL MTBE/hexane (1:1), and
the suspension was filtered to afford 4.61 g of 77. The filtrate was
concentrated and the residue was purified by column chromato-
graphy using EtOAc/hexane (20-50% gradient) to give 2 g of
pure 77. A total of 6.61 g (74%) of product 77 was obtained. 1H
NMR (500 MHz, DMSO-d6) δ 7.17-7.203 (t, J = 8.5 Hz, 1H),
7.38 (dd, J = 8, 0.5 Hz, 1H), 7.58 (dd, J = 8.0, 0.5 Hz, 1H),
7.57-7.68 (m, 2H), 7.94 (s, 1H), 7.96 (dd, J = 8.5, 1.5 Hz, 1H),
8.00-8.09 (m, 3H), 8.04 (s, 1H), 12.2 (s, 1H).
MHz, DMSO-d6) δ 4.48 (s, 2H), 6.98 (t, J = 8 Hz, 1H), 7.14 (dd,
J = 7.5, 0.5 Hz, 1H), 7.20 (d, J = 2.5 Hz, 1H), 7.40-7.45 (m,
4H), 7.63 (s, 1H), 7.76-7.86 (m, 4H), 11.29 (s, 1H).
(E)-3-(3-Naphthalen-2-ylmethyl-1H-indol-4-yl)acrylic Acid Me-
thyl Ester (79). To a pressure resistant tube containing 1.5 g of 78
(4.46 mmol) in 2 mL of Et3N were added Pd(OAc)2 (50 mg, 0.22
mmol), tri-o-tolylphosphine (270 mg, 0.88 mmol), and methyl
acrylate (500 mg, 5.8 mmol). Mixture was heated to 100 °C for
2.5 h. A bulky precipitate formed. TLC (20% EA/hexane)
indicated the reaction completion. The reaction mixture was
cooled to room temperature, the suspension was diluted with
water and extracted with CH2Cl2, the combined organic layers
were washed with water, brine, dried over anhydrous MgSO4,
concentrated in vacuo to give 1.6 g of a residue, which was
purified by column chromatography using 10-25% EtOAc/
hexane gradient elution to provide 1 g (65%) of compound 79.
1H NMR (500 MHz, DMSO-d6) δ 3.68 (s, 3H), 4.37 (s, 2H), 6.35
(d, J = 16 Hz, 1H), 7.08-7.11 (t, J = 7.5 Hz, 1H), 7.37 (t, J = 7
Hz, 1H), 7.39-7.48 (m, 6H), 7.73 (s, 1H), 7.74-7.84 (m, 3H),
8.29 (d, J = 16 Hz, 1H), 11.22 (s, 1H).
(E)-3-(3-Naphthalen-2-ylmethyl-1H-indol-4-yl)acrylic Acid
(80). To a solution of the methyl ester 79 (1 g, 2.9 mmol) in a
mixture of THF/MeOH (3:2, 50 mL) was added 20 mL of 1 N
aqueous NaOH,. The mixture was stirred at 60 °C for 4 h. TLC
(EtOAc/hexane, 1:1) indicated the reaction completion. The
reaction mixture was concentrated in vacuo, and to this residue
a solution of 10% aqueous HCl was added to bring the mixture
to pH 2. A gummy dark-brown solid formed. The mixture was
stirred at room temperature for 0.5 h when solid started crashing
out. The suspension was filtered off to afford, after washing with
water and drying, 0.95 g (100%) of the compound 80. 1H NMR
(500 MHz, DMSO-d6) δ 4.36 (s, 2H), 6.28 (d, J = 16 Hz, 1H),
7.09-7.105 (t, J = 7.5 Hz, 1H), 7.33 (s, 1H), 7.34 (d, J = 7 Hz,
1H), 7.40-7.45 (m, 4H), 7.73 (s, 1H), 7.75-7.84 (m, 4H), 8.33 (d,
J = 16 Hz, 1H), 11.18 (s, 1H), 12.22 (s, 1H).
(E)-3-(1-Methyl-3-naphthalen-2-ylmethyl-1H-indol-4-yl)acry-
lic Acid Methyl Ester (81). To a suspension of NaH (60%
dispersion in mineral oil, 47 mg, 1.17 mmol) in 3 mL of
anhydrous THF at 0 °C was added the methyl ester 79 (200
mg, 0.58 mmol). The mixture was stirred at room temperature
for 1 h, then cooled to 0 °C, and CH3I (0.180 mL, 2.9 mmol) was
added. The reaction mixture was stirred at room temperature
overnight. The mixture was quenched with 10% aqueous HCl
and extracted with CH2Cl2. The organic layer was washed with
brine, dried over anhydrous MgSO4, and concentrated in vacuo
to afford 200 mg of compound 81. This material was used for
next step without purification. 1H NMR (400 MHz, CDCl3) δ
3.78 (s, 3H), 4.35 (s, 2H), 6.32 (d, J = 16 Hz, 1H), 7. 16 (t, J = 7.5
Hz, 1H), 7.27 (s, 1H), 7.37-7.45 (m, 4H), 7.50 (d, J = 8 Hz, 1H),
7.76-7.85 (m, 4H), 8.33 (d, J = 16 Hz, 1H).
(E)-3-(1-Methyl-3-naphthalen-2-ylmethyl-1H-indol-4-yl)acry-
lic Acid (82). To a solution of the methyl ester 81, (200 mg, 0.448
mmol) in a mixture of THF/MeOH (3:2) was added 2 mL of 1 N
aqueous NaOH. The mixture was stirred at 60 °C for 4 h. TLC
(EtOAc/hexanes, 1:4) indicated the reaction completion. The
reaction mixture was concentrated in vacuo, and to this residue
a solution of 10% aqueous HCl was added to give a mixture of
pH 2. The oily mixture was extracted with EtOAc (2 ꢀ 10 mL).
The combined organic layers were washed with water, brine,
dried over anhydrous MgSO4, filtered, and concentrated in
vacuo to provide 180 mg (90%, for two steps from 80) of the
acid 82. 1H NMR (500 MHz, DMSO-d6) δ 3.78 (s, 3H), 4.35 (s,
2H), 6.31 (d, J = 16 Hz, 1H), 7.15-7.18 (t, J = 8 Hz, 1H), 7.27
(s, 1H), 7.37-7.44 (m, 4H), 7.50 (d, J = 8 Hz, 1H), 7.76-7.85
(m, 4H), 8.33 (d, J = 16 Hz, 1H), 12.22 (s, 1H).
4-Bromo-3-naphthalen-2-ylmethyl-1H-indole (78). To a 100
mL round-bottomed flask containing 2.1 g (6 mmol) of 77 and
50 mL of anhydrous THF, under N2 at 0 °C, was added
dropwise 18 mL (18 mmol, 3 equiv) of BH3 THF (1 M in THF).
3
The cooling bath was removed, and the reaction mixture was
stirred at room temperature for 1.5 h. The reaction mixture was
cooled to 0 °C, and 10 mL of MeOH was added dropwise (with
gas evolution). After 0.5 h at room temperature, the mixture was
concentrated in vacuo to obtain 2 g of 78 as a light-yellow oil
that was used without purification for next step. 1H NMR (500
3-(3-Naphthalen-2-ylmethyl-1H-indol-4-yl)propionic Acid Me-
thyl Ester (83). To a solution of the acrylate 79, (500 mg, 1.46
mg) in 30 mL of MeOH under N2 was added 200 mg of 10% Pd/
C. The mixture was stirred at 40 °C under H2 for 2 h, then at
room temperature overnight. The reaction mixture was filtered

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 33
through a Celite plug using MeOH and CH2Cl2. The solvent was
removed and residue was purified by column chromatography
using 10-20% EtOAc/hexanes to afford 435 mg (86%) of the
compound 83. 1H NMR (500 MHz, DMSO-d6) δ 2.37 (t, J = 8.5
Hz, 2H), 3.03 (t, J = 8.5, 2H), 3.51 (s, 3H), 4.35 (s, 2H), 6.69 (d,
J = 7.0 Hz, 1H), 6.95-6.98 (t, J = 8.0 Hz, 1H), 7.11 (d, J = 2.0
Hz, 1H), 7.22 (d, J = 7 Hz, 1H), 7.36 (dd, J = 8.0, 1.5 Hz, 1H),
7.42-7.44 (m, 2H), 7.61 (s, 1H), 7.75 (m, 1H), 7.81 (d, J = 8.5
Hz, 1H), 7.86 (m, 1H), 10.9 (s, 1H).
CDCl3) δ 2.39 (s, 3H), 6.43 (br s, 1H), 6.86 (d, J = 16.0 Hz,
1H), 7.14 (m, 2H), 7.24 (m, 1H), 8.20 (br s, 1H). MS (AP-): mass
calcd for C10H9NO2, 175.1; m/z found 174.2 (M - 1).
Acetic Acid 3-(Naphthalene-2-carbonyl)-1H-indol-4-yl Ester
(89). MeMgBr (26 mL, 78 mmol, 1.05 equiv, 3.0 M solution in
Et2O) was added slowly (over 10 min) via syringe to a solution of
the acetate 88 (13 g, 74 mmol, 1 equiv) in anhydrous CH2Cl2
(260 mL) at room temperature. After the mixture was stirred for
5 min at room temperature, ZnCl2 (223 mL, 3.0 equiv, 1.0 M
solution in Et2O) was added via syringe over 5 min at room
temperature. After the mixture was stirred for 10 min at room
temperature, a solution of 2-naphthoyl chloride (14.85 g, 1.05
equiv) in anhydrous CH2Cl2 (87 mL) was added over 3 min. The
reaction mixture was stirred for 3 h at room temperature and
then was poured into saturated aqueous NH4Cl (866 mL), and
the mixture was diluted with CH2Cl2 (200 mL). The aqueous
layer was extracted with CH2Cl2 (100 mL). The combined
organic layers were washed with water (2 ꢀ 500 mL), brine
(500 mL), dried over anhydrous MgSO4, filtered, and concen-
trated in vacuo to afford 24.4 g (100%) of brown solid. MTBE
(100 mL) was added to the solid, heated to reflux, cooled to
30-40 °C, and filtered. The residue was washed on filter with
MTBE (50 mL, 25 mL, 2 ꢀ 5 mL), then with Et2O (2 ꢀ 5 mL) to
afford 16.0 g (65%) of 89 as a light-brown solid. 1H NMR (500
MHz, CDCl3) δ 2.41 (s, 3H), 6.89 (d, 1H), 7.03 (d, J = 16.5 Hz,
1H), 7.09 (m, 2H), 7.52 (t, J = 15.0 Hz, 1H), 7.56 (t, J = 15.0 Hz,
1H), 7.79 (m, 3H), 7.88 (d, J = 16.0 Hz, 1H), 8.12 (br s, 1H), 9.35
(br s, 1H). MS (ESI-): mass calcd for C21H15NO3, 329.3; m/z
found 328.1 (M - 1).
3-(3-Naphthalen-2-ylmethyl-1H-indol-4-yl)propionic Acid (84).
To a solution of the methyl ester 83 (130 mg, 0.380 mmol)
in a mixture of THF/MeOH (3:2) was added 2 mL of 1 N aqueous
NaOH. The mixture was stirred at 60 °C for 4 h. TLC
(EtOAc/hexanes, 1:1) indicated the reaction completion. The
reaction mixture was concentrated in vacuo, and to this residue
a solution of 10% aqueous HCl was added bringing the solution
to pH 2. The oily mixture was extracted with EtOAc (2 ꢀ 10 mL).
The combined organic layers were washed with water, brine,
dried over anhydrous MgSO4, and concentrated in vacuo to give
1
126 mg (quantitative) of compound 84. H NMR (500 MHz,
DMSO-d6) δ 2.41 (t, J = 8.0 Hz, 2H), 3.14 (t, J = 8.0 Hz, 2H),
4.35 (s, 2H), 6.71 (d, J = 7.0 Hz, 1H), 6.96 (t, J = 8.0 Hz, 1H),
7.05 (d, J = 2.5 Hz, 1H), 7.22 (d, J = 8.5 Hz, 1H), 7.36 (dd, J =
8.0, 1.5 Hz, 1H), 7.42-7.44 (m, 2H), 7.62 (s, 1H), 7.74-7.76 (m,
1H), 7.81 (d, J = 8.5 Hz, 1H), 7.86 (m, 1H), 10.9 (s, 1H), 12.05
(br s, 1H).
3-(1-Methyl-3-naphthalen-2-ylmethyl-1H-indol-4-yl)propionic
Acid Methyl Ester (85). To a suspension of NaH (60% disper-
sion in mineral oil, 35 mg, 0.87 mmol) in 3 mL of anhydrous
THF at 0 °C was added the methyl ester 83 (200 mg, 0.58 mmol).
The mixture was stirred at room temperature for 1 h, then cooled
to 0 °C, and CH3I (0.180 mL, 2.9 mmol) was added. The reaction
mixture was stirred at room temperature overnight. The mixture
was quenched with 10% aqueous HCl and extracted with
CH2Cl2. The organic layer was washed with brine, dried over
anhydrous Na2SO4, and concentrated in vacuo to afford 160 mg
(77%) of the compound 85. 1H NMR (500 MHz, CDCl3) δ 2.52
(t, J = 8.0 Hz, 2H), 3.24 (t, J = 8.0 Hz, 2H), 3.58 (s, 3H), 3.71 (s,
3H), 4.41 (s, 2H), 6.66 (s, 1H), 6.85 (d, J = 7.0 Hz, 1H),
7.12-7.19 (m, 2H), 7.38-7.43 (m, 3H), 7.64 (s, 1H), 7.73 (m,
1H), 7.77 (d, J = 8.5 Hz, 1H), 7.80 (m, 1H).
3-(1-Methyl-3-naphthalen-2-ylmethyl-1H-indol-4-yl)propio-
nic Acid (86). To a solution of the methyl ester 85 (160 mg, 0.448
mmol) in a mixture of THF/MeOH (3:2) was added 2 mL of 1 N
aqueous NaOH. The mixture was stirred at 60 °C for 4 h. TLC
(EtOAc/hexanes, 1:4) indicated the reaction completion. The
reaction mixture was concentrated in vacuo, and to this residue
a solution of 10% aqueous HCl was added to obtain pH 2. The
oily mixture was extracted with EtOAc (2 ꢀ 10 mL). The
combined organic layers were washed with water, brine, dried
over anhydrous MgSO4, and concentrated in vacuo to give 120
mg (78%) of compound 86. 1H NMR (500 MHz, DMSO-d6) δ
2.4 (t, J = 8.0 Hz, 2H), 3.06 (t, J = 8.0 Hz, 2H), 3.71 (s, 3H), 4.34
(s, 2H), 6.78 (d, J = 8.5 Hz, 1H), 6.99 (s, 1H), 7.04 (t, J = 8.0 Hz,
1H), 7.25 (d, J = 8.5 Hz, 1H), 7.38- 7.45 (m, 3H), 7.64 (s, 1H),
7.76 (m, 1H), 7.82 (d, J = 8.5 Hz, 1H), 7.85 (m, 1H), 12.07 (br s,
1H).
Acetic Acid 1H-Indol-4-yl Ester (88). Ac2O (16.75 mL, 170
mmol, 1.5 equiv) was added slowly (over 7 min) to a solution of
4-hydroxyindole (87), 15 g, 110 mmol, 1 equiv) in pyridine (113
mL) at room temperature. Temperature rose from 25 to 32 °C.
After beirng stirred for an additional 5 min, the reaction mixture
was cooled in an ice bath, and 10% aqueous HCl (340 mL)
followed by EtOAc (565 mL) was added. The organic layer was
separated, and the aqueous layer was extracted with EtOAc (100
mL). The combined organic layers were washed with 10%
aqueous HCl (2 ꢀ 50 mL), water (2 ꢀ 300 mL), brine (300
mL) and dried over anhydrous MgSO4. Filtration and evapora-
tion of the solvent in vacuo afforded 19.3 g (98%) of the
compound 88 as a light-brown solid. 1H NMR (500 MHz,
3-Naphthalen-2-ylmethyl-1H-indol-4-ol (90). BH3 THF (1 M
3
in THF, 120 mL, 120 mmol, 3.3 equiv) was added slowly to a
solution of 89 (12.0 g, 36.4 mmol, 1 equiv) at 0 °C under a
nitrogen atmosphere. The reaction mixture was allowed to
warm to room temperature and stir overnight. The reaction
mixture was quenched by slow addition of MeOH (120 mL),
evaporated, and coevaporated with MeOH (3 ꢀ 120 mL). The
residue was purified by silica gel chromatography (150 g) using
CH2Cl2 as eluent to afford 6.2 g (62%) of the compound 90. 1H
NMR (DMSO-d6) δ 4.35 (s, 2H), 6.30 (d, J = 7.0 Hz, 1H), 6.79
(m, 2H), 6.87 (d, J = 1.5 Hz, 1H), 7.43 (m, 2H), 7.49 (dd, J =
8.5, 1.5 Hz, 1H), 7.73 (s, 1H), 7.78 (d, J = 8.5 Hz, 1H), 7.83 (d,
J = 7.0 Hz, 1H), 9.2 (s, 1H), 10.67 (s, 1H). MS (ESI-): mass
calcd for C19H15NO, 273.1; m/z found 271 (M - 2).
(3-Naphthalen-2-ylmethyl-1H-indol-4-yloxy)acetic Acid Me-
thyl Ester (91). A solution of methyl bromoacetate (3.10 g, 1.9
mL, 20.3 mmol, 1.05 equiv) in DMF (20 mL) was added slowly
to the mixture of 90 (5.29 g, 19.4 mmol, 1 equiv) and K2CO3
(3.21 g, 23.2 mmol, 1.2 equiv) in DMF (51 mL). Reaction
mixture was stirred at room temperature overnight (17 h).
Water-brine (5:1, 180 mL) was added, and the resulting solu-
tion was extracted with EtOAc (180 mL, 140 mL, 70 mL). The
organic layer was washed with brine (2 ꢀ 210 mL), dried over
anhydrous MgSO4, filtered, and concentrated in vacuo to afford
7.8 g of a solid. This residue was washed with Et2O (20 mL) and
filtered to afford 5.0 g (75%) of the compound 91 as an off-white
solid. 1H NMR (DMSO-d6) δ 3.70 (s, 3H), 4.38 (s, 2H), 4.80 (s,
2H), 6.34 (d, J = 7.5, 1H), 6.90-6.97 (m, 2H), 6.99 (d, J = 1.5
Hz, 1H), 7.42 (m, 2H), 7.50 (dd, J = 8.5, 1.5 Hz, 1H), 7.76-7.83
(m, 4H), 10.88 (s, 1H). MS (ESIþ): mass calcd for C22H19NO3,
345.4; m/z found 346.5 (M þ 1).
(3-Naphthalen-2-ylmethyl-1H-indol-4-yloxy)acetic Acid (92).
A solution 2 N aqueous NaOH (9.5 mL, 19 mmol, 2 equiv) was
added to a solution of 91 (3.3 g, 9.55 mmol, 1 equiv) in
THF-MeOH, 2:1 (132 mL). The reaction mixture was stirred
at room temperature for 1 h. The mixture was concentrated to
∼20 mL, and 10% aqueous HCl (10 mL) was added followed by
water (50 mL). The mixture was extracted with EtOAc (250 mL),
and the organic phase was dried over anhydrous MgSO4,
filtered, and concentrated in vacuo to afford 2.95 g (92%)
of the acid 92 as an off-white solid. 1H NMR (500 MHz,

34 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Singh et al.
DMSO-d6) δ 4.38 (s, 2H), 4.70 (s, 2H), 6.34 (d, J = 6.5 Hz, 1H),
6.89-6.95 (m, 2H), 6.97 (d, J = 2.0 Hz, 1H), 7.42 (m, 2H), 7.52
(dd, J = 8.5, 1.5 Hz, 1H), 7.76 (d, J = 8.5 Hz, 1H), 7.78 (d, J =
7.5 Hz, 1H), 7.80 (d, J = 7.5 Hz, 1H), 7.85 (s, 1H), 10.85 (d, J =
1.0 Hz, 1H), 12.75 (br s, 1H).
(100 mL). The organic layer was separated, washed with brine,
dried over anhydrous Na2SO4, filtered, and concentrated in
vacuo. The residue was chromatographed on flash silica (145 g),
eluting with 5% EtOAc in hexanes to yield 6.56 g (80%) of
compound 97. 1H NMR (400 MHz, CDCl3) δ 2.23 (d, J = 0.8
Hz, 3H), 4.21 (s, 4 H), 5.59 (s, 2H), 6.50-6.54 (m, 2H), 6.77 (d, J
= 8.0 Hz, 1H), 6.89 (s, 1H), 7.14 (m, (overlapping dd), 2H).
(E)-3-[1-(2,3-Dihydrobenzo[1,4]dioxin-6-ylmethyl)-5-fluoro-3-
methyl-1H-indol-7-yl]acrylic Acid Methyl Ester (98). Compound
97 (2.163 g, 5.75 mmol), methyl acrylate (1.56 mL, 17.24 mmol),
tri-o-tolylphosphine (0.525 g, 1.73 mmol), Pd(OAc)2 (0.130 g,
0.58 mmol), and Et3N (4.00 mL, 29.00 mmol) were heated in a
sealed tube under nitrogen atmosphere for 4 h at 100 °C. The
mixture was cooled to room temperature, and the solids were
filtered off and washed on with EtOAc. The combined filtrates
were diluted with EtOAc (200 mL), washed successively with 1
N aqueous HCl (2 ꢀ 30 mL) and brine (2 ꢀ 30 mL), dried over
anhydrous Na2SO4, and concentrated in vacuo. The residue was
dissolved in CH2Cl2 (8 mL), and the solution was diluted with
hexanes (200 mL). The precipitate was filtered off and dried to
yield 1.49 g (68%) of compound 98. 1H NMR (400 MHz,
CDCl3) δ 2.28 (s, 3H), 3.79 (s, 3H), 4.21 (s, 4 H), 5.30 (s, 2H),
6.22 (d, J = 15.6 Hz, 1H), 6.50 (d, J = 2.0 Hz, 1 H), 6.56 (dd,
J = 8.4, 2.0 Hz, 1H), 6.80 (d, J = 8.4 Hz, 1H), 6.93 (s, 1H), 7.01
(dd, J = 10.0, 2.4 Hz, 1H), 7.23 (dd, J = 8.4, 2.4 Hz, 1H), 8.12
(d, J = 15.6 Hz, 1H). MS (APCIþ): mass calcd for C22H20-
FNO4, 381.4; m/z found 382.0 (M þ 1).
Thiophene-2-sulfonic Acid [(E)-3-(5-Fluoro-3-methyl-1H-
indol-7-yl)acryloyl]amide (96a). A mixture of the acid 95 (100
mg, 0.46 mmol), 2-thiophenesulfonamide (90 mg, 0.55 mmol),
DMAP (112 mg, 0.92 mmol), and EDCI (176 mg, 0.92 mmol) in
CH2Cl2 (10 mL) was stirred at room temperature overnight. The
solution was diluted with CH2Cl2 and washed with diluted
aqueous HCl and water. The resulting solid was filtered washed
with water and CH2Cl2 to give 55 mg of compound 96a. The
remaining CH2Cl2 mother liquid was purified by column chro-
matography on silica gel with CH2Cl2/MeOH as an eluent to
give 45 mg of acylsulfonamide 96a. A total of 100 mg (60%) of
compound 96a was obtained. 1H NMR (500 MHz, DMSO-d6) δ
2.22 (s, 3H), 6.69 (d, J = 15.0 Hz, 1H), 7.19 (dd, J = 10.0, 2.0
Hz, 1H), 7.22-7.25 (m, 2H), 7.37 (dd, J = 9.0, 2.0 Hz, 1H), 7.86
(m, 1H), 8.06 (m, 1H), 8.10 (d, J = 15.0 Hz, 1H), 11.35 (s, 1H),
12.42 (br s, 1H).).
3,5-Difluoro-N-[(E)-3-(5-fluoro-3-methyl-1H-indol-7-yl)acryl-
oyl]benzenesulfonamide (96c). Compound 96c was prepared
through the coupling of acid 95 with the 3,5-difluorobenzene-
sulfonamide according to general procedure A. Yield: 51%. 1H
NMR (400 MHz, DMSO-d6) δ 2.22 (s, 3 H), 6.70 (d, J = 16.0
Hz, 1 H), 7.21 (dd, J = 10.0, 2.0 Hz, 1 H), 7.25 (s, 1 H), 7.38 (dd,
J = 10.0, 2.0 Hz, 1 H), 7.57-7.77 (m, 3 H), 8.10 (d, J = 16.0 Hz,
1 H), 11.33 (s, 1 H), 12.56 (bs, 1 H). MS (ESI-): mass calcd for
C18H13F3N2O3S, 394.4; m/z found 393.5 (M - 1). LCMS 98%.
3,4-Difluoro-N-[(E)-3-(5-fluoro-3-methyl-1H-indol-7-yl)acry-
loyl]benzenesulfonamide (96d). Compound 96d was prepared
through the coupling of 95 with 3,4-difluorobenzenesulfona-
mide according to general procedure A. Yield: 41%. 1H NMR
(400 MHz, DMSO-d6) δ 2.22 (s, 3 H), 6.69 (d, J = 16.0 Hz, 1 H),
7.20 (dd, J = 10.0, 2.0 Hz, 1 H), 7.25 (s, 1 H), 7.37 (dd, J = 10.0,
2.0 Hz, 1 H), 7.75 (m, 1 H), 7.88 (m, 1 H), 8.04 (m, 1 H), 8.10 (s,
1 H), 11.32 (s, 1 H), 12.47 (br s, 1 H). MS (ESI-): mass calcd for
C18H13F3N2O3S, 394.4; m/z found 393.5 (M - 1). LCMS 99%.
2,4,5-Trifluoro-N-[(E)-3-(5-fluoro-3-methyl-1H-indol-7-yl)acry-
loyl]benzenesulfonamide (96e). Compound 96e was prepared
through the coupling of 95 with 2,4,5-trifluorobenzenesulfona-
mide according to general procedure A, and the material was
used as such for the next step. Yield: 49%. 1H NMR (500 MHz,
DMSO-d6) δ 2.22 (s, 3H), 6.72 (d, J = 15.5 Hz, 1H), 7.20 (dd,
J = 10.0, 2.0 Hz, 1H), 7.25 (s, 1H), 7.38 (dd, J = 9.5, 2.0 Hz,
1H), 7.92 (m, 1H), 8.07 (m, 1H), 8.09 (d, J = 15.5 Hz, 1H), 11.31
(s, 1H), 12.85 (br s, 1H). MS (ESI-): mass calcd for
C18H12F4N2O3S, 412.1; m/z found 411.4 (M - 1). LCMS 96%.
4-Fluoro-N-[(E)-3-(5-fluoro-3-methyl-1H-indol-7-yl)acryloyl]-
benzenesulfonamide (96f). Compound 96f was prepared through
the coupling of 95 with 4-fluorobenzenesulfonamide according
to general procedure A. Yield: 53%. 1H NMR (400 MHz,
DMSO-d6) δ 2.22 (s, 3 H), 6.69 (d, J = 16 Hz, 1 H), 7.18 (dd,
J = 10.0, 2.0 Hz, 1 H), 7.24 (s, 1 H), 7.37 (dd, J = 10.0, 2.0 Hz, 1
H), 7.48-7.52 (m, 2 H), 8.03 (s, 1 H), 8.04-8.09 (m, 2 H), 11.31
(s, 1 H), 12.35 (br s, 1 H). MS (ESI-): mass calcd for
C18H14F2N2O3S, 376.4; m/z found 375.5 (M - 1). LCMS 96%.
7-Bromo-1-(2,3-dihydrobenzo[1,4]dioxin-6-ylmethyl)-5-fluoro-
3-methyl-1H-indole (97). A solution of bromoindole 93 (5.00 g,
21.90 mmol) in anhydrous DMF (100 mL) was cooled to 5 °C.
NaH (60% in mineral oil, 1.314 g, 32.85 mmol) was added
portionwise over a period of 15 min. The cooling bath was
removed, and the mixture was allowed to warm to room
temperature and stirred for 1.5 h. The mixture was recooled
to 5 °C, and 6-(bromomethyl)-2,3-dihydro-1,4-benzodioxine
(6.024 g, 26.30 mmol) was added dropwise. The cooling bath
was removed, and the mixture was stirred overnight at room
temperature. The reaction mixture was diluted with EtOAc
(300 mL) followed by addition of saturated aqueous NH4Cl
(E)-3-[1-(2,3-Dihydrobenzo[1,4]dioxin-6-ylmethyl)-5-fluoro-3-
methyl-1H-indol-7-yl]acrylic Acid (99). To a solution of methyl
acrylate 98 (1.43 g, 3.75 mmol) in a mixture of MeOH (30 mL)
and THF (30 mL) was added 2 N aqueous solution of NaOH
(30 mL). The mixture was stirred overnight at room tempera-
ture. Most volatiles were removed under reduced pressure. The
residue was diluted with water (20 mL), and the pH was adjusted
to 2 with concentrated HCl. The precipitate was filtered off,
washed on funnel with water, and dried in air to yield 1.20 g
(87%) of the acid 99. 1H NMR (400 MHz, DMSO-d6) δ 2.24 (s,
3H), 4.16 (t, J = 4.8 Hz, 4 H), 5.37 (s, 2H), 6.31 (d, J = 15.6 Hz,
1H), 6.42 (d, J = 2.0 Hz, 1 H), 6.47 (dd, J = 8.4, 2.0 Hz, 1H),
6.74 (d, J = 8.4 Hz, 1H), 7.20 (dd, J = 10.4, 2.4 Hz, 1H), 7.35 (s,
1H), 7.38 (dd, J = 8.8, 2.4 Hz, 1H), 8.05 (d, J = 15.6 Hz, 1H),
12.40 (br s, 1H). LCMS 100%. MS (APCI-) m/z 366.1 (M - H).
MS (APCI-): mass calcd for C21H18FNO4, 367.4; m/z found
366.1 (M - 1). LCMS 100%.
3-(5-Fluoro-3-methyl-1H-indol-7-yl)propionic Acid (100). To
a solution of the acid 95 in EtOH was added 10% by weight of
10% Pd/C. The reaction vessel was degassed, the atmosphere
replaced with hydrogen, and the process repeated 3 times. The
suspension was stirred under 1 atm of hydrogen for 16 h at room
temperature. An additional 10% by weight of 10% Pd/C was
added, and the reaction was allowed to proceed an additional
16 h at room temperature. The reaction was filtered through a
pad of Celite and the cake washed with EtOH. The filtrate was
concentrated in vacuo to afford compound 100 in 98% yield. 1H
NMR (400 MHz, DMSO-d6): 2.20 (s, 3H), 2.62 (dd, J = 7.6 Hz,
2H), 3.04 (dd, J = 7.6 Hz, 2H), 6.76 (dd, J = 10.4, 2.4 Hz, 1H),
7.05 (dd, J = 9.6, 2.4 Hz, 1H), 7.15 (s, 1H), 10.86 (s, 1H), 12.15
(s, 1H).
3-[1-(2,4-Dichlorobenzyl)-5-fluoro-3-methyl-1H-indol-7-yl]-
propionic Acid (101). To a solution of 100 (2.03 mmol, 1.0 equiv)
in anhydrous THF at 0-5 °C was added 60% NaH (6.1 mmol,
3.0 equiv) over 3 min. The resultant mixture was allowed to
warm to room temperature and stir for 30 min. The reaction
mixture was then cooled to 0-5 °C, and 2,4-dichlorobenzyl
chloride (4.1 mmol, 2.0 equiv) was added. The reaction mixture
was allowed to warm naturally to room temperature and stirred
for 16 h, then cooled to 0-5 °C. Additional 60% NaH (2.2
mmol, 1.1 equiv) was added, and the mixture was warmed to
room temperature and stirred for 30 min before cooling again to
0-5 °C. Additional 2,4-dichlorobenzyl chloride (4.0 mmol,

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 35
2.0 equiv) was added, and the mixture again was allowed to
warm to room temperature and stir for 16 h. The mixture was
diluted with EtOAc and acidified to pH 1-2 with 1 N HCl. The
organic portion was washed with successive portions of water,
and brine and dried over anhydrous MgSO4. The filtrate was
concentrated in vacuo and the residue purified by flash chro-
matography on silica gel using acetone in CH2Cl2 as eluent to
afford compound 101 in 18% yield. 1H NMR (400 MHz,
DMSO-d6) δ 2.23 (s, 3H), 2.45 (dd, J = 8.4, 7.2 Hz, 2H), 2.84
(dd, J = 8.0, 7.6 Hz, 2H), 5.56 (s, 2H), 6.53 (d, J = 8.4 Hz, 1H),
6.77 (dd, J = 10.4, 2.4 Hz, 1H), 7.16 (dd, J = 9.2, 2.4 Hz, 1H),
7.26 (s, 1H), 7.28 (dd, J = 8.4, 2.4 Hz, 1H), 7.68 (d, J = 2.0 Hz,
1H), 12.19 (s, 1H) .
Biological Assays. Displacement Binding Assays. The pri-
mary binding assays were established using membranes pre-
pared from cell lines expressing the hEP3 and other various
prostanoid receptors. An amount of 10 μg of membrane protein
expressing hEP3 receptor was incubated for 60 min at 30 °C with
binding buffer in the presence of various concentrations of the
test compound (dose response of 20 ꢀ 10-6 to 10-9 M, dilution
factor 3ꢀ) and 2 nM [3H]PGE2 radioligand. For hIP and hFP
displacement binding experiments, [3H]iloprost (one tritium-
labeled analogue of PGI2, 10-20 Ci/mmol, Amersham
Biosciences) and [5,6,8,9,11,12,14,15(n)-3H]PGF2R (160-240
Ci/mmol, Amersham Biosciences) were used as radioligand,
respectively. Nonspecific binding was determined by adding,
in specific wells, 10 μM unlabeled PGE2 (Cayman Chemicals,
Inc.) in the incubation mixture. Unlabeled PGI2 or PGF2R
(Cayman Chemicals, Inc.) was used for binding experiment
using hIP or hFP receptors, respectively. The specific binding
was calculated by subtracting the nonspecific binding
(membranes incubated with cold and labeled ligand) from the
total binding (membranes incubated only with tritiated radi-
oligand). In all binding studies, reactions were terminated by
filtering the mixture through a glass fiber filter (Packard Bio-
sciences, PerkinElmer, MA). Filters were then washed three
times with binding buffer, and the radioactivity associated with
filters was quantified in 7 mL of scintillation liquid (Ultima
Gold, PerkinElmer, Boston, MA) using a liquid scintillation
counter (TRI-CARB 2100TR, Packard Biosciences, Perkin-
Elmer, MA). IC50 values were calculated using GraphPad Prism
for Windows (GraphPad Software, San Diego, CA).
Functional Assay. Stably transfected CHO-K1 cells expres-
sing the hEP3 receptor were plated into 96-well plates at a cell
density of 105 cells/well and cultured overnight at 37 °C and 5%
CO2 in culture media supplemented with 10% FIBS, 2% PS
(penicillin-streptomycin, GIBCO), and 1 mg/mL geneticin
(GIBCO). Cells were washed once with phosphate-buffered
saline (PBS) and preincubated in fresh serum- and antibiotic-
free medium containing 1 mM IBMX (3-isobutyl-1-methyl-
xanthine, Sigma) for 30 min at 37 °C. After preincubation, fresh
medium was added without aspiration, containing PGE2 at the
appropriate concentration (dose response from 10-4 to 10-13
M) in the presence of 5 μM forskolin (Sigma). Similarly, cells
were challenged with the test compound at the appropriate
concentrations (dose response of 10-4-10-13 M) in the presence
of 5 μM forskolin (Sigma) and 5 nM PGE2. Cells were then
incubated for an additional 10 min at 37 °C. Reactions were
terminated by aspiration of medium and addition of 200 μL of
lysis buffer 1B (cAMP EIA system kit, Amersham). cAMP levels
were determined using a commercially available cAMP EIA
system kit (Amersham). Raw OD values, measured using a
Spectra MAX190 plate reader (Molecular Devices), were trans-
formed into an amount of cAMP (fmol/well) using GraphPad
Prism for Windows (GraphPad Software, San Diego, CA). For
IC50 calculations in a dose-response experiment, sigmoidal
nonlineal regressions were performed.
challenge was done 30 min following a single oral dose of the test
compounds. The test compounds were formulated in 40%
hydroxylpropyl-β-cyclodextrin (HPβCD) in 50 mM phosphate
buffer, pH 7.4, containing PVP (0.25% w/v) and lysine hydro-
chloride (50 mM) solution at 5 mg/mL. The data are presented
as percent survival.
Platelet Aggregation Assays. These assays were perfomed
with human platelet rich plasma (PRP). Blood was taken by
venipuncture from nonsmoking volunteers of both sexes after
overnight fasting into 3.2% sodium citrate tubes. Individual
experiments were performed with blood from a single subject.
Platelet-rich plasma (PRP) was obtained by centrifugation of
whole blood at 100g for 15 min at 25 °C. Platelet aggregation
was measured by light absorbance using a platelet aggregometer
(model 490, Chronolog Corp., Havertown, PA) according to the
manufacturer’s instructions using collagen (0.25 μg/mL) and
sulprostone (100 nM) to stimulate platelet aggregation.
Acknowledgment. The authors acknowledge technical
contributions of Jenny Lin for providing guidance and overall
analytical support, Denise Anderson for running mass spec-
tral analyses, Dr. Nelson Zhao for acquiring NMR spectra,
Miriam Goldsmith for preparing compounds 30 and 45, Hua
Caofor preparing compounds63and 64, Matthew O’ Connell
for preparing compounds 57-59, James Burgeson for pre-
paring compounds 49 and 60-62, and Drs. Livia Enecha, Lei
Zhao, and Kris Pupek for preparing compounds 40, 46, and
65, respectively. The authors also thank Dr. Kari Stefansson
for helpful discussions and encouragement during the course
of this work.
Supporting Information Available: Synthetic schemes and
experimental and supporting data for compounds 31-33,
35-37, and 39-42. This material is available free of charge
via the Internet at http://pubs.acs.org.
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