Structure-activity relationship of biaryl acylsulfonamide analogues on the human EP3 prostanoid receptor

Article abstract of DOI:10.1016/S0960-894X(02)00518-8

Potent and selective ligands for the human EP3 prostanoid receptor are described. Biaryl compounds bearing a tethered ortho substituted acidic moiety were identified as potent EP3 antagonists based on the SAR described herein. The binding affinity of key compounds on all eight human prostanoid receptors is reported.

Full text of DOI:10.1016/S0960-894X(02)00518-8

Bioorganic & Medicinal Chemistry Letters 12 (2002) 2583–2586
Structure–Activity Relationship of Biaryl Acylsulfonamide
Analogues on the Human EP3 Prostanoid Receptor
M. Gallant,* M. C. Carriere, A. Chateauneuf, D. Denis, Y. Gareau, C. Godbout,
G. Greig, H. Juteau, N. Lachance, P. Lacombe, S. Lamontagne, K. M. Metters,
C. Rochette, R. Ruel, D. Slipetz, N. Sawyer, N. Tremblay and M. Labelle
´
Merck Frosst Centre for Therapeutic Research, PO Box 1005, Pointe Claire-Dorval, Quebec, Canada H9R 4P8
Received 11 April 2002; accepted 19 June 2002
Abstract—Potent and selective ligands for the human EP3 prostanoid receptor are described. Biaryl compounds bearing a tethered
ortho substituted acidic moiety were identified as potent EP3 antagonists based on the SAR described herein. The binding affinity
of key compounds on all eight human prostanoid receptors is reported. # 2002 Elsevier Science Ltd. All rights reserved.
In response to various extracellular stimuli, pros-
taglandins are rapidly generated through the con-
secutive action of the cyclo-oxygenases and distinct
synthases on free arachidonic acid and exert their action
in close proximity to the site of their synthesis. To date,
eight prostanoid receptors have been cloned and char-
acterized. These receptors are members of the growing
class of G-protein-coupled receptors. PGE2 will bind
preferentially to the EP1, EP2, EP3, and EP4 receptors,
PGD2 to the DP and FP receptors, PGF2a to the FP
and EP3 receptors, PGI2 to the IP receptor and TXA2 to
the TP receptor.1,2 PGE2 binding to the EP3 receptor
has been found to play a key role in the regulation of
ion transport, smooth muscle contraction of the GI
tract, acid secretion, uterine contraction during fertili-
zation and implantation, fever generation and PGE2-
mediated hyperalgesia. The EP3 receptor has been
detected in many organs such as the kidney, the gastro-
intestinal tract, the uterus and the brain.3 The molecular
characterization of these receptors has generated
renewed interest in this field as the selectivity of prosta-
noid ligands can now be determined. Thus far, PGE2 or
prostanoid-like compounds have been utilized to study
the pharmacological role of the EP3 receptor.4 Since
these compounds behave as agonists and cross react
with other prostanoid receptors, the search for selective
EP3 antagonists has become a critical issue in the phar-
macological characterization of the EP3 receptor.
Figure 1. Lead optimization through combinatorial library.
All compounds described herein behave as full
antagonist.
Our search for a selective EP3 receptor ligand was initi-
ated by screening a large number of compounds in our
in vitro binding assays.5 The biaryl acylsulfonamide6
(1), an angiotensin II antagonist, was found to be a
submicromolar ligand for the EP4 receptor (Fig. 1,
Table 1). The lead structure was explored by preparing
a combinatorial library of 750 compounds leading to
the identification of potent and selective EP3 (3 and 4)
and EP1 (2) ligands. The resulting SAR clearly established
that affinity for the EP3 receptor, was improved by intro-
ducing an ortho substituted acidic moiety, consisting of a
two carbon tethered acyl sulfonamide, relative to the
biaryl scaffold.
*Corresponding author. Fax: +1-514-428-4900; e-mail: michel_gallant@
merck.com
0960-894X/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(02)00518-8

2584
M. Gallant et al. / Bioorg. Med. Chem. Lett. 12 (2002) 2583–2586
Table 1. Affinity of lead compounds on prostanoid E2 receptors
aKi (mM)
Table 3. SAR on the lipophilic residue
Entry
EP1
EP2
EP3
EP4
1
2
3
4
>10
0.083
>10
7.0
0.046
1.5
1.0
orthob
metab
parab
3.8
2.0
2.7
0.51
0.027
0.046
0.82
1.7
1.1
aKi determinations are averages of at least two experiments.
bPosition of phenethylthiomethyl ether moiety relative to biaryl bond.
The nature of the acid group was firstly investigated.
Acidic surrogates (Table 2) to the acylsulfonamide
group, such as carboxylic acid (5), tetrazole (6), inverse
acylsulfonamide (7) and sulfonimide (8) failed to
improved potency on the EP3 receptor. Interestingly,
the introduction of a tetrazole moiety (6) modified the
receptor specificity to give an EP2 selective ligand.
Attempt to modify the tether length, as exemplified by 9
and 10 did not improve selectivity nor potency toward
the EP3 receptor.
aKi (mM)
Entry
EP1
13
4.6
49
27
EP2
EP3
EP4
11
12
13
14
15
4.9
2.9
16
0.016
0.053
0.12
0.24
5.9
7.1
1.6
19
31
>100
25
>100
>100
aKi determinations are averages of at least two experiments.
The lipophilic residue was examined next. As demon-
strated by the selected examples in Table 3, large lipo-
philic group are well tolerated. Compound, such as 11,
bearing a long lipophilic chain exhibit high affinity for
the EP3 receptor but are found to be metabolically
unstable. The truncated analogue 12 and the quinoline
13, though metabolically more robust, are less potent on
the EP3 receptor. The relatively small methylthioether
14 is surprisingly only 7.5-fold less potent than 11 while
the corresponding polar methylsulfone 15 analogue is a
poor ligand. Following characterization of the major
metabolites of 11, we discovered that the acylsulfon-
amide hydrolysis, was the major degradation pathway
problem we prepared a small number of compounds in
which the acylsulfonamide moiety was sterically hin-
dered or electronically enriched. Compounds 16 and 17
(Table 4) in which an a-methyl groups were introduced,
are stable in rat plasma. However, they show a 6-fold
reduction in affinity for the EP3 receptor. The introduc-
tion of an oxygen atom( 18) resulted in a rat plasma
stable compound with a 2.2-fold shift in affinity. In
contrast, the corresponding nitrogen analogue 19 is
readily cleaved in rat plasma.
In fact, these acylsulfonamides derivatives of phenyl-
propionic acid are readily cleaved in an ex vivo rat
plasma assay. Compounds 12 and 13 were incubated for
1.5 h in rat plasma at 37 ꢀC. This resulted in partial
hydrolysis to their corresponding inactive carboxylic
acid in 20 and 66%, respectively. To alleviate this
Ultimately, the metabolic instability of the phenyl-
propionic acylsulfonamide class of compounds was
solved by introducing a double bond in the tether. The
cinnamic acylsulfonamide 20 (Table 5) is stable in rat
plasma and is equipotent to its saturated analogue 13
on the EP3 receptor with Ki of 0.11 and 0.12 mM,
Table 2. SAR on the acidic moiety; acylsulfonamide surrogates and
tether length
Table 4. SAR on the metabolic stability of the acylsulfonamide
moietyb
aKi (mM)
aKi (mM)
Entry
R
EP1
EP2
EP3
EP4
5
6
7
8
9
10
(CH2)2COOH
(CH2)2-tetrazole
(CH2)2SO2NHCOThb
(CH2)2SO2NHSO2Thb
CH2CONHSO2Thb
(CH2)3CONHSO2Thb
8.6
4.9
2.8
7.5
3.7
3.0
0.14
0.063
2.5
3.0
0.59
1.8
0.74
1.0
0.48
0.61
0.33
0.20
4.4
2.6
0.69
1.1
0.85
0.70
Entry
R
EP1
EP2
EP3
EP4
16
17
18
19
CH2CH(CH3)CONHSO2Th
CH2C(CH3)2CONHSO2Th
CH2OCONHSO2Th
5.8
1.8
2.8
15
2.9
2.9
2.1
8.6
0.34
0.44
0.12
0.15
1.1
1.0
0.73
2.9
CH2NHCONHSO2Th
aKi determinations are averages of at least two experiments.
bTh=2-thienyl.
aKi determinations are averages of at least two experiments.
bTh=2-thienyl.

M. Gallant et al. / Bioorg. Med. Chem. Lett. 12 (2002) 2583–2586
Table 5. Affinity of compounds 21 on all eight prostanoid receptors
2585
aKi (mM)
Entry
EP1
EP2
EP3
EP4
DP
FP
17
IP
TP
20
21
41
8.2
70
27
0.11
0.025
28
3.7
0.83
7.3
0.77
aKi determinations are averages of at least two experiments.
Scheme 1. (a) PdCl2(dppf)2, pinacoldiborane, KOAc, 80 ꢀC; (b)
Pd(PPh3)4, Na2CO3, DME, 80 ꢀC; (c) TFA/CH2Cl2 for t-Bu, LiOH/
MeOH/THF for Et; (d) EDCI, DMAP, thiophene sulfonamine,
DMF.
respectively. Following this observation we pursued the
SAR of the lipophilic residue. Previous results had
shown that the presence of a large lipophilic group was
not essential for potency on the EP3 receptor (e.g., 14,
Table 3). Consequently, we identified the 3,4-dichlor-
ophenyl analogue 21, with a Ki of 25 nM on the EP3
receptor and an acceptable selectivity profile (>30-fold)
against all eight prostanoid receptors. Compound 21
behaves as a full antagonist with a Kb of 130 nM in our
EP3 functional assay.7
Lead optimization through combinatorial library is a
powerful tool to initiate SAR. It allowed us to address
the spatial characteristics of a good ligand for the EP3
receptor. Conventional SAR allowed the optimization
of the binding affinity of our antagonists but more
importantly to address the metabolic stability of this
series of compounds. We ultimately identified the potent
and selective ligand 21 which pharmacologic properties
will be disclosed elsewhere.
Scheme 2. (a) (i) N-(Thiophene-2-carbonyl) methanesulfonamide/n-
BuLi, THF, À78 ꢀC, (ii) benzyl bromide À78 ꢀC to rt; (b) (i) N-(thio-
phene-2-sulfonyl) methanesulfonamide/n-BuLi, THF, À78 ꢀC (ii) ben-
zyl bromide À78 ꢀC to rt; (c) (i) cyclohexyl-isopropyl amine/n-BuLi,
THF, À78 ꢀC, (ii) MeI À78 ꢀC to rt.
Generally, biaryl acylsulfonamide are prepared in three
steps (Scheme 1). Suzuki coupling between a properly
substituted phenylboronic acid (23) and the corre-
sponding 2-bromophenyl propionate (24) affords the
biaryl ester 25. Subsequent deprotection followed by an
EDCI/DMAP coupling8 with a sulfonamine generates
the desired acylsulfonamide 26. Occasionally, the boro-
nic acid 23 could not be prepared or was not commer-
cially available. In such cases, the pinacol-boronate 28
was prepared fromthe corresponding aryl bromide 24
by a palladiumcatalyzed coupling with pinacol diboron
ester followed by a Suzuki coupling with the aryl bro-
mide 26. The latter procedure can not be performed on
the phenyl cinnamic analogues because of addition of
the pinacol diboron ester on the electron poor double
bond.
The mono a-methyl substituted analogue 32 were pre-
pared (Scheme 2) by first, formation of the enolate of
the tert-butyl ester 24 using the lithiumsalt of cyclo-
hexyl-isopropyl amine10 followed by addition of methyl
iodide. The dimethyl analogue was synthesized by
repeating the procedure.
The carbamate sulfonamide 35 and urea sulfonamide 37
were synthesized starting fromthe benzyl alcohol 34
(Scheme 3). Addition of thiophene sulfonyl-isocyanate11
to 34 led to the desired compound. The alcohol 34 was
Inverse acylsulfonamides such as 30 and sulfonimides
31 were prepared in a similar fashion as exemplified in
Scheme 2. This was achieved by displacement of the
benzylic bromide 29 by the dianion generated fromthe
deprotonation of N-(thiophene-2-carbonyl) methane-
sulfonamide or N-(thiophene-2-sulfonyl) methanesulfo-
namide using 2 equiv of n-BuLi at low temperature.9
Scheme 3. (a) Thiophene sulfonyl-isocyanate, Et2O, rt; (b) AcOH/
HBr, heat; (c) potasiumphthalimide, DMF, rt; (d) hydrazine/EtOH,
reflux.

2586
M. Gallant et al. / Bioorg. Med. Chem. Lett. 12 (2002) 2583–2586
Acknowledgements
The authors would like to thank Dr Daniel Dube
proofreading this manuscript.
´
for
References and Notes
1. Abramovitz, M.; Adam, A.; Boie, Y.; Godbout, C.;
Lamontagne, S.; Rochette, C.; Sawyer, N.; Tremblay, N. M.;
Belley, M.; Gallant, M.; Dufresne, C.; Gareau, Y.; Ruel, R.;
Juteau, H.; Labelle, M.; Ouimet, N.; Metters, K. M. Biochim.
Biophys. Acta 2000, 285, 1483.
2. Kiryiama, A.; Ushikubi, F.; Kobayashi, T.; Hirata, M.;
Sugimoto, Y.; Narumiya, S. Br. J. Pharmacol. 1997, 122, 217.
3. (a) Sugimoto, Y.; Narumiya, S.; Ichikawa, A. Prog. Lipid.
Res. 2000, 39, 289. (b) Narumiya, S.; Sugimoto, Y.; Ushikubi,
F. Physiol. Rev. 1999, 79, 1193. (c) Coleman, R. A.; Smith,
W. L.; Narumiya, S. Pharmacol. Rev. 1994, 46, 205.
4. Shimazaki, Y.; Kameo, K.; Tanami, T.; Tanaka, H.; Ono,
N.; Kiuchi, Y.; Okamoto, S.; Sato, F.; Ichikawa, A. Bioorg.
Med. Chem. 2000, 8, 353.
Scheme 4. (a) (i) NaH, DMF, (ii) BrCH2CH2CH2OH, (iii) AcOH/
HBr, heat; (b) 3-methylphenol, NaH, DMF; (c) NBS, AIBN, CCl4; (d)
PhCH2CH2SH, NaH, DMF; (e) n-BuLi, À78 ꢀC, B(Oi-Pr)3, À78 ꢀC to
rt, then HCl; (f) (7-chloro-quinolin-2-ylmethyl) triphenylphosphonium
bromide, t-BuOK, THF.
5. See ref 1 for stable expression of prostanoid receptors in the
human embryonic kidney (HEK) 293 cell line and also for
prostanoid receptor binding assays. The EP3-III subtype was
used for the binding assay.
6. Ashton, W. T.; Chang, L. L.; Flanagan, K. L.; Hutchins,
S. M.; Naylor, E. M.; Chakravarty, P. K.; Patchett, A. A.;
Greenlee, W. J.; Chen, T. B. J. Med. Chem. 1994, 37, 2808.
7. EP3 antagonist assays were performed in human ery-
throleukemia cells (ATCC HEL 92.1.7) that endogenously
express the EP3 receptor. cAMP accumulation assays were
performed in a total of 0.2 mL HBSS containing 100 mM RO-
20174 and 15 mM Forskolin. 21 (0–3 Â10À6 M) was added to
the incubation mixture in DMSO (0.5% final) and pre-
incubated with the cells (2Â105 cells per incubation) for 10 min
at 37 ꢀC. The reaction was initiated with the EP3 selective
agonist sulprostone (Biomol) (0–3 Â10À5 M in 0.5% DMSO),
and incubated at 37 ꢀC for 10 min. The reaction was termi-
nated by boiling the samples for 3 min and cAMP was mea-
sured by cAMP SPA assay (Amersham).
8. Pelletier, J. C.; Hesson, D. P. Synlett 1995, 11, 1141.
9. Belletire, J. L.; Spletzer, E. G. Tetrahedron Lett. 1986, 27,
131.
10. Rathke, M. W.; Linbert, A. J. Am. Chem. Soc. 1971, 93,
2318.
11. Friesen, R. W.; Giroux, A. Tetrahedron Lett. 1993, 34,
1867.
12. Weinstein, B.; Ho, T. N. S.; Fukura, R. T.; Angell, E. C.
Synth. Commun. 1976, 6, 17.
converted in three steps to the corresponding benzylic
amine 36. Formation of the benzyl bromide (AcOH/
HBr) followed by nucleophilic displacement with the
potassiumphthalimide and finally, deprotection using
hydrazine gave the amine 36. Addition of thiophene sul-
fonyl-isocyanate12 afforded the desired urea sulfonamide.
The aryl boronic acid 41 bearing the lipophilic chain
was prepared by addition of 3-bromopropanol on the
sodiumsalt of 3-bromophenol followed by a bromina-
tion to give 38. Displacement with 3-methylphenol,
radical bromination and displacement using the sodium
salt 2-phenyl ethanethiol gave the aryl bromide 40.
Lithiumhalogen exchange at low temperature followed
by addition of tri-isopropyl boronate and subsequent
acid work up led to the desired boronic acid. The quin-
oline ester 43 was prepared by a Wittig reaction using
the ylide generated form(7-chloro-quinolin-2-ylmethyl)
triphenylphosphoniumbromide and potassium
butoxide followed by addition of the aldehyde 42
tert-
(Scheme 4).