1,5-Biaryl pyrrole derivatives as EP1 receptor antagonists. Structure-activity relationships of 6-substituted and 5,6-disubstituted benzoic acid derivatives

Article abstract of DOI:10.1016/j.bmcl.2006.11.059

Herein we describe the SAR of 1,5-biaryl pyrrole derivatives, with substituents in the 6-position of the benzoic acid moiety, as EP₁ receptor antagonists. Substitution at this position was well tolerated and led to the identification of several analogues with high affinity for the EP₁ receptor that displayed good efficacy in the established FCA model of inflammatory pain. Furthermore, several analogues were prepared which combined substitution at the 5- and 6-positions as well as derivatives with an aromatic ring fused to the 5- and 6-positions.

Full text of DOI:10.1016/j.bmcl.2006.11.059

Bioorganic & Medicinal Chemistry Letters 17 (2007) 916–920
1,5-Biaryl pyrrole derivatives as EP1 receptor antagonists.
Structure–activity relationships of 6-substituted and 5,6-
disubstituted benzoic acid derivatives
Adrian Hall,* Susan H. Brown, Iain P. Chessell, Anita Chowdhury, Nicholas M. Clayton,
Tanya Coleman, Gerard M. P. Giblin, Beverley Hammond, Mark P. Healy,
Matthew R. Johnson, Ann Metcalf, Anton D. Michel, Alan Naylor,
Riccardo Novelli, David J. Spalding, Jennifer Sweeting and Lisa Winyard
Neurology and Gastrointestinal Centre of Excellence for Drug Discovery, GlaxoSmithKline, New Frontiers Science Park,
Third Avenue, Harlow, Essex CM19 5AW, UK
Received 26 October 2006; revised 17 November 2006; accepted 18 November 2006
Available online 22 November 2006
Abstract—Herein we describe the SAR of 1,5-biaryl pyrrole derivatives, with substituents in the 6-position of the benzoic acid moi-
ety, as EP1 receptor antagonists. Substitution at this position was well tolerated and led to the identification of several analogues
with high affinity for the EP1 receptor that displayed good efficacy in the established FCA model of inflammatory pain. Further-
more, several analogues were prepared which combined substitution at the 5- and 6-positions as well as derivatives with an aromatic
ring fused to the 5- and 6-positions.
Ó 2006 Elsevier Ltd. All rights reserved.
The EP1 receptor is one of four 7-transmembrane recep-
tor subtypes, known as EP1-4,1 that is activated by pros-
taglandin E2 (PGE2). The importance of PGE2 as an
inflammatory mediator has been demonstrated by the
correlation between PGE2 blood concentration and
analgesia for cyclooxygenase (COX) inhibitors.2 Evi-
dence that the pro-inflammatory actions of PGE2 are
mediated by the EP1 receptor has been provided by
studies with knock-out (KO) mice and tool compounds.
KO mice have implicated the EP1 receptor subtype in
the sensation of PGE2-mediated inflammatory pain.3
In conjunction with this, EP1 receptor antagonists have
shown efficacy in preclinical models of postoperative
pain,4 neuropathic pain5 and allodynia.6 We have previ-
ously shown that EP1 antagonists are efficacious in pre-
clinical models of inflammatory pain.7
There have been several reports of EP1 antagonists in
the recent literature.8 We disclosed the structure–activity
relationships (SAR), in vivo rat pharmacokinetic data
and preclinical efficacy data of a series of 1,5-biaryl pyr-
role EP1 receptor antagonists, such as 1a (Fig. 1).7
Compound 1a7 showed good in vitro affinity at the re-
combinant human EP1 receptor with a pIC50 of 8.2 in a
[3H-]PGE2 binding assay in CHO cell membranes.7,8
Rat in vivo pharmacokinetic data showed 1a to have
moderate blood clearance (CLb = 42 mL/min/kg) whilst
data from the established complete Freund’s adjuvant
(CFA or FCA) model of inflammatory pain7 showed 1a
to have an ED50 of 9.2 mg/kg when dosed orally to rats.
O
EP1 binding pIC50 = 8.2
EP1 pA2 9.1
2
Br
N
OH
rat pharmacokinetics
CLb = 42 mL/min/kg
Vss = 0.7 L/kg
4
6
O
5
Keywords: EP1 antagonist; Pyrrole; Pain; Established FCA.
*
Corresponding
adrian.2.hall@gsk.com
author.
Tel.:
+44
1279
643464; e-mail:
established FCA ED50 9.2 mg/kg
1a
F
Present address: AstraZeneca, Mereside, Alderley Park, Macclesfield,
Cheshire SK11 4TG, UK.
Figure 1. Profile of lead compound 1a.
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.11.059

A. Hall et al. / Bioorg. Med. Chem. Lett. 17 (2007) 916–920
917
Table 1. In vitro EP1 binding data for 6-substituted derivatives 1a–p
Recently, we described the SAR of 4- and 5-substituted
benzoic acid derivatives.9
O
2
X
N
We now describe further SAR from this series where
substitution of the benzoic acid moiety at the 6-position
was investigated. Furthermore, we describe the SAR for
some 5,6-disubstituted and fused analogues. This pro-
gramme of work was directed at finding an analogue
of 1a which had improved pharmacokinetics and im-
proved in vivo efficacy. We were also interested in
assessing whether these changes would impact in vitro
affinity for the EP1 receptor.
OH
4
O
Z
6
5
Y
a
Compound
X
Y
Z
Binding pIC50
1a
Br
4-F
4-F
H
H
8.2 0.1
Compound affinities were determined using recombi-
nant human EP1 receptor stably expressed in CHO cell
membranes.8,10b
1b
1c
1d
1e
1f
Br
CF3
CF3
Me
F
F
F
F
F
F
8.9 0.2
8.7 0.1
8.1 0.2
7.8 0.0
7.9 0.1
8.6 0.1
8.3 0.1
8.6 0.1
8.9 0.1
8.7 0.1
9.4 0.0
8.3 0.2
8.7 0.2
9.2 0.1
9.4 0.1
Initial results demonstrated that installation of a fluo-
rine atom at the 6-position, 1b, led to a moderate
improvement in affinity, Table 1. Fixing the 6-fluoro-
benzoic acid on the right hand side and exploring alter-
native X-groups on the left hand side phenyl ring
revealed that Br and CF3 were generally better than
Me or F, 1b–f. Returning to the 6-position of the benzo-
ic acid, replacement of the fluorine atom by chlorine had
little effect on activity, 1g versus 1b, Table 1.
4-F
4-F
4-F
4-F
4-F
It was found that the electron-withdrawing halogen
group in the 6-position could be replaced by various
groups such as Me, OH, OMe, OCHF2 and NHAc,
1h–p. Addition of these groups led to the identification
of several analogues with exceptionally high affinity,
such as 1l, 1o and 1p. Comparison of 1i with 1j revealed
that Cl and Br (X-group on the left hand side) could be
interchanged with minimal effect on affinity.
1g
1h
1i
Br
Cl
Cl
Me
Cl
2,4-DiF
2,4-DiF
2,4-DiF
2,4-DiF
H
Me
The benzyl substitution (Y-group) effect is well illustrat-
ed by the stepwise increase in affinity moving from 1m to
1n to 1o.
1j
Br
Br
Br
Br
Br
Br
Br
Me
Having identified several groups in the 6-position that
increased in vitro affinity, we were eager to combine
one of these substituents with the NH2 group in the 5-
position which we had previously found to increase in vi-
tro affinity.9 Previously we had found that when an NH2
group was introduced to the 5-position of the benzoic
acid (X = Br, Y = H), the pIC50 value increased from
8.17,8 to 8.5.9 Hence, derivatives 2a–f were prepared,
where the 5-amino group was incorporated with the
6-methyl group, Table 2. The selection of the methyl
group in the 6-position was based mainly on the avail-
ability of starting material, see later (Scheme 2).
Although the in vitro affinity was good, it was not as
high as might be anticipated and the addition of a sec-
ond substituent did not give an additive rise in affinity,
although no direct comparisons of 1h–j were made.
Again, affinity was lowest when X was Me, compound
2f, Table 2.
1k
1l
OH
OMe
1m
1n
1o
1p
OCHF2
OCHF2
OCHF2
NHAc
4-F
2,4-DiF
2,4-DiF
a Mean of at least three experiments standard deviation.
ring was well tolerated. As previously observed the
2,4-difluorobenzyl group yielded the most active com-
pounds, whilst bromine was the best X-group and meth-
yl the least active.
As an alternative approach to combine substitution at
the 5- and 6-position we decided to investigate the activ-
ity of fused systems such as the naphthyl derivatives, 3a–
f, Table 3. As the results show, the additional phenyl

918
A. Hall et al. / Bioorg. Med. Chem. Lett. 17 (2007) 916–920
Table 2. In vitro EP1 binding data for 5,6-disubstituted derivatives
2a–f
Table 3. In vitro EP1 binding data for naphthoic acid derivatives 3a–f
O
O
2
X
N
OH
X
N
OH
O
6
4
O
5
NH2
Y
Y
a
Compound
X
Y
H
Binding pIC50
a
Compound
X
Y
Binding pIC50
3a
Cl
8.2 0.1
2a
Cl
H
8.7 0.2
3b
3c
3d
3e
3f
Br
Br
Br
Me
F
H
7.9 0.3
8.3 0.2
8.6 0.2
7.1 0.1
7.6 0.2
2b
2c
2d
2e
2f
Br
H
8.5 0.1
8.4 0.1
9.1 0.1
8.4 0.1
7.8 0.1
4-F
CF3
Br
H
2,4-DiF
4-F
4-F
4-F
4-F
CF3
4-F
Me
a Mean of at least three experiments standard deviation.
a Mean of at least three experiments standard deviation.
Due to the activity of the naphthyl derivatives, we inves-
tigated other fused ring systems, especially those that
might mimic the hydrogen bond donating (HBD) ability
of the acetamide group in the 5-position which we had
previously discovered to impart high affinity.9 Hence,
the indole-4-carboxylic acid 4 was prepared in an at-
tempt to increase in vitro affinity. Pleasingly, this com-
pound showed excellent activity with a pIC50 value of
8.9, Figure 2. It is also worth noting that this level of
activity was attained without the preferred substitution
pattern on the left hand side of the molecule.
O
Cl
N
OH
4: binding pIC50 8.9 (0.1)a
O
N
H
Figure 2. In vitro EP1 binding data for compound 4. aMean of at least
three experiments, standard deviation in parentheses.
Based on a balance of in vitro affinity at the EP1 recep-
tor, minimal CYP450 interactions and in vitro metabolic
stability, several compounds were profiled in a rat phar-
macokinetic assay to investigate their oral exposure,
Table 4.
Table 4. In vivo rat pharmacokinetic data for selected compoundsa
Compound AUC (0–t)/dose (min kg/L) Tmax (h) Cmax (lM)
1c
1h
1l
27
18
4
8
9
1
9
3
0.6
0.5
0.5
0.5
0.5
1.1
4.0
0.5
0.87 0.03
0.59 0.18
0.34 0.17
2.94 1.16
0.91 0.10
1.35 0.32
0.22 0.11
0.52 0.25
Substitution of the 6-position generally resulted in good
exposure, 1c and 1h, defined as a dose-normalized area
under the curve (AUC) P10 min kg/L, with the excep-
tion of the methoxy derivative 1l. The 5,6-disubstituted
analogues 2a, 2c and 2d displayed the highest exposure,
whereas the naphthoic acids had considerably lower
exposure.
2a
2c
2d
3a
3c
39
31
52 14
9
9
3
3
a Mean of three experiments. Compounds administered orally in 1%
aqueous methylcellulose at 3 mg/kg.
Compounds with acceptable oral exposure were assessed
in the established FCA model of inflammatory pain.7
Compounds were either assayed at a single dose of
5 mg/kg or in dose–response format, Table 5. The
6-methyl derivative 1h showed considerably better effica-
cy than the 6-F derivative 1c. Disappointingly, the
disubstituted derivatives 2a, 2c and 2d showed consider-
ably less efficacy despite the excellent exposure shown.
On the basis of these results, 1h (GW855454X) was pro-
filed in a dose–response assay, and showed improved
efficacy relative to 1a, with an ED50 of 2.5–4.9 mg/
kg,11 Table 5.

A. Hall et al. / Bioorg. Med. Chem. Lett. 17 (2007) 916–920
919
Table 5. In vivo rat pain data from FCA model of inflammatory paina
Table 7. Rat blood and brain levels of compound 1h (GW855454)
from FCA studya
a,b
ED50 (mg/kg)
Compound
% reversal of hypersensitivity
at 5 mg/kga (%)
Time (h)
Blood concnb
Brain concnc
Br:Bl ratiod
(lM)
(lM)
1c
1h
2a
2c
2d
56
75
35
40
50
n/t
2.5–4.9c
n/t
n/t
0.25
7.358 1.842
10.089 2.598
5.930 2.304
3.561 1.285
2.309 0.682
1.600 0.322
0.037 0.020
2.598 0.441
3.564 0.669
2.484 0.516
1.221 0.618
0.913 0.282
0.753 0.080
0.031e
0.4
0.4
0.4
0.4
0.4
0.4
0.5e
1
2
n/t
4
6
a Compounds administered orally in 1% aqueous methyl cellulose at
doses of 1, 3 and 10 mg/kg or 3, 10 and 30 mg/kg or at a single dose
of 5 mg/kg. Pain readout taken 1 h post-dose of test compound.
b n/t = not tested.
8
24
a Compound dosed orally in 1% aqueous methylcellulose at 30 mg/kg.
b Mean of seven data points.
c Mean of three data points.
c See Ref. 11.
d Br:Bl ratios calculated by dividing individual brain concentrations by
the corresponding blood concentration for the same animal. Mean of
three data points.
Due to the excellent efficacy of compound 1h its phar-
macokinetic profile was investigated in a rat iv-po cross-
over study, detailed in Table 6. In vitro, 1h showed good
metabolic stability with intrinsic clearance (CLi) values
63.6 mL/min/g liver in rat, dog, monkey and human liv-
er microsomes. Compound 1h showed improved in vivo
metabolic stability relative to 1a with a large volume of
distribution which is unusual for a carboxylic acid. The
combination of the large volume of distribution and low
clearance results in a half-life of 4.6 h.
e Single data point.
time point. Furthermore, the analgesia is maintained
for approximately 8 h, at which point there are still con-
siderable drug concentrations in both tissues, Figure 3
and Table 7. From these data the estimated blood
EC50 is 3.5 lM at the 4 h time point.
Based on these data, the effects of 1h were studied in a
time-course (PK-PD) experiment over a 24 h time period
using a dose of 30 mg/kg in the established FCA model
of inflammatory pain. Results from this study are shown
in Figure 3 and Table 7. As the results show, 1h showed
good reversal of the mechanical hyperalgesia induced by
FCA at the 1 and 2 h time points. This effect correlates
with peak blood and brain concentrations at the 1 h
The in vitro selectivity profile of 1h was ascertained at
related prostanoid receptors (EP2, EP3, EP4, IP, FP
and TP). Functional antagonism at the EP1, EP3 and
TP receptors was ascertained using a calcium mobilisa-
tion assay (FLIPR, recombinant human receptor ex-
pressed in CHO cells). In these assays 1h demonstrated
a pKi of 9.0 1.1 (n = 2) at the EP1 receptor and a
pKi of 6.3 0.0 (n = 4) at the EP3 receptor. At the TP
receptor 1h showed a pIC50 of 7.4 and a pKi of 7.8.
No agonist activity was observed at the TP receptor.
Compound 1h showed no activity up to a concentration
of 1 lM at the EP2 or IP receptors and no activity at the
FP receptor (pIC50 < 5.5). At the EP4 receptor, 1h
showed weak activity with a pIC50 of 4.8 and pKi of
5.1 in a binding assay and a pKi of 5.2 in a cAMP mobi-
lisation assay. No significant activity (<25%) was seen at
a battery of 50 targets (Cerep) at a concentration of
1 lM.12 Thus, 1h shows excellent selectivity over a range
of related and unrelated targets with the exception of the
TP receptor.
Table 6. In vivo rat pharmacokinetic data for compound 1ha
Compound CLb (mL/min/kg) Vss (L/kg) t1/2 (h)
Fpo
1h 22 4 2.6 0.9 4.6 3.2 45%
a Compound administered intravenously in 5% (w/v) glucose contain-
ing 2% DMSO (v/v) at 1 mg/kg and orally in 1% aqueous methyl-
cellulose at 3 mg/kg. Data are mean values of three experiments.
time course in established FCA for compound 1h (30mg/kg p.o.)
100
75
50
25
0
The data for 1h (GW855454X) thus support its use as a
tool compound to investigate the in vivo effects of EP1
receptor antagonism. In addition, despite being a car-
boxylic acid, 1h shows considerable penetration into
the central nervous system (CNS) as evidenced by the
brain concentrations from the time-course study. Thus,
1h may prove useful in elucidating the CNS effects of
EP1 receptor antagonism.
-25
-50
Compounds were prepared by Paal-Knorr condensation
of the requisite 1,4-diketone with an appropriately func-
tionalized aniline as described previously.7 Full experi-
mental procedures and characterizing data have been
disclosed.10 The anilines were generally commercially
available or were prepared by literature procedures.
Conveniently, if the aniline was not available it could
0
2
4
6
8
10 12 14 16 18 20 22 24 26
Time (hours post dose)
Figure 3. PK-PD data for 1h in established FCA at 30 mg/kg (po).

920
A. Hall et al. / Bioorg. Med. Chem. Lett. 17 (2007) 916–920
CO2Me
OCHF2
H2N
H2N
CO2Me
OMe
O2N
CO2Me
OH
a, b
c, b
6
5
7
Scheme 1. Reagents and conditions: (a) ClCF2CO2Na, Na2CO3, DMF, 100 °C; (b) H2, Pd/C, EtOH; (c) NaH, MeI, DMF.
O
O
O
H2N
H2N
O2N
b, a
a
OMe
OMe
OMe
N
H
NH2
NO2
9
8
10
Scheme 2. Reagents and conditions: (a) H2, Pd/C, MeOH; (b) DMF–DMA, DMF, 60 °C, lwave, 15 min.
Toxicol. 2001, 41, 661; (c) Narumiya, S.; Sugimoto, Y.;
Ushikubi, F. Physiol. Rev. 1999, 79, 1193; (d) Coleman, R.
A., Kennedy, I., Humphrey, P. P. A., Bunce, K., Lumley,
P., Eds.; Comprehensive Medicinal Chemistry; 1990, Vol. 3,
Pergamon: Oxford, UK, p 643.
be prepared by reduction of the analogous nitro
derivative.
Methyl 5-amino-2-(methyloxy)benzoate 6 and methyl 5-
amino-2-[(difluoromethyl)oxy]benzoate 7 were prepared
from 5 as detailed in Scheme 1. Paal-Knorr condensa-
tion7 with the appropriate 1,4-diketone followed by ester
hydrolysis yielded derivatives 1m–o. The condensation
reaction was conducted under thermal (PhMe, pTSA,
reflux) or microwave (NMP, pTSA, 150 °C, 10 min)
conditions.
2. Huntjens, D. R. H.; Danhof, M.; Della Pasqua, O. E.
Rheumatology 2005, 44, 846.
3. Stock, L. S.; Shinjo, K.; Burkhardt, J.; Roach, M.;
Taniguchi, K.; Ishikawa, T.; Kim, H.-S.; Flannery, P. J.;
Coffman, T. M.; McNeish, J. D.; Audoly, L. P. J. Clin.
Invest. 2001, 107, 325.
4. (a) Omote, K.; Kawamata, T.; Nakayama, Y.; Kawa-
mata, M.; Hazama, K.; Namiki, A. Anesth. Analg. 2001,
92, 233; (b) Omote, K.; Yamamoto, H.; Kawamata, T.;
Nakayama, Y.; Namiki, A. Anesth. Analg. 2002, 95,
1708.
5. Kawahara, H.; Sakamoto, T.; Takeda, S.; Onodera, H.;
Imaki, J.; Ogawa, R. Anesth. Analg. 2001, 93, 1012.
6. Maruyama, T.; Koketsu, M.; Yaamamoto, H.; Yamam-
oto, K.; Yamamoto, L. T.; Hayashida, K.-i.; Ohchida, S.;
Kondo, K. Prostaglandins and Other Lipid Mediators
1999, 59, 217.
Methyl 3,5-diamino-2-methylbenzoate 9 was prepared
from 8 as detailed in Scheme 2. Paal-Knorr condensation
took place selectively with the less hindered 5-amino
moiety, to give derivatives 2a–f, upon basic hydrolysis
of the ester group. Methyl 6-amino-1H-indole-4-carbox-
ylate 10 was prepared as described in Scheme 2 and used
without purification to prepare compound 4.
7. Hall, A.; Atkinson, S.; Brown, S. H.; Chessell, I. P.;
Chowdhury, A.; Clayton, N. M.; Coleman, T.; Giblin, G.
M. P.; Gleave, R. J.; Hammond, B.; Healy, M. P.;
Johnson, M. J.; Michel, A. D.; Naylor, A.; Novelli, R.;
Spalding, D. J.; Tang, S. P. Bioorg. Med. Chem. Lett.
2006, 16, 3657.
8. Hall, A.; Bit, R. A.; Brown, S. H.; Chaignot, H. M.;
Chessell, I. P.; Coleman, T.; Giblin, G. M. P.; Hurst, D.
N.; Kilford, I. R.; Lewell, X. Q.; Michel, A. D.; Moham-
ed, S.; Naylor, A.; Novelli, R.; Skinner, L.; Spalding, D.
J.; Tang, S. P.; Wilson, R. J. Bioorg. Med. Chem. Lett.
2006, 16, 2666, and references therein.
In conclusion, substitution of the benzoic acid moiety of
the 1,5-biaryl pyrrole template led to the identification
of compounds with sub-nanomolar affinity for the EP1
receptor. Several analogues showed good oral exposure
and efficacy in a preclinical model of inflammatory pain.
Compound 1h (GW855454X) demonstrated good meta-
bolic stability, good bioavailability and excellent efficacy
in established FCA model of inflammatory pain with an
ED50 of 2.5–4.9 mg/kg. These data support the use of 1h
as a tool compound to elucidate the in vivo effects of
EP1 receptor antagonism.
9. Hall, A.; Brown, S. H.; Chessell, I. P.; Chowdhury, A.;
Clayton, N. M.; Coleman, T.; Giblin, G. M. P.; Ham-
mond, D.; Healy, M. P.; Johnson, M. R.; Metcalf, A.;
Michel, A. D.; Naylor, A.; Novelli, R.; Spalding, D. J.;
Sweeting, J. Bioorg. Med. Chem. Lett. 2006, 16.
doi:10.1016/j.bmcl.2006.10.078.
10. (a) Giblin, G. M. P.; Hall, A.; Healy, M. P.; Lewell, X. Q.;
Miller, N. D.; Novelli, R. WO2003/101959A1.; (b) Giblin,
G. M. P.; Hall, A.; Healy, M. P.; Lewell, X. Q.; Miller, N.
D.; Novelli, R.; King, F. D.; Naylor, A. WO2005/
054191A1.
Acknowledgments
The authors thank Emma Ward for generating the EP1
and EP3 FLIPR data and Davina Mitchell for generat-
ing the FP data.
References and notes
11. ED50 = 2.5 mg/kg when dose–response assay carried out
at doses of 1, 3 and 10 mg/kg, ED50 = 4.9 mg/kg when
dose–response assay carried out at 3, 10 and 30 mg/kg.
12. www.cerep.fr.
1. (a) Coleman, R. A.; Smith, W. L.; Narumiya, S. Pharma-
col. Rev. 1994, 46, 205; (b) Breyer, R. M.; Bagdassarian, C.
K.; Myers, S. A.; Breyer, M. D. Annu. Rev. Pharmacol.