A novel class of positive allosteric modulators of AMPA receptors: Design, synthesis, and structure-activity relationships of 3-biphenyl-4-yl-4-cyano-5-ethyl-1-methyl-1H-pyrrole-2-carboxylic acid, LY2059346

Article abstract of DOI:10.1016/j.bmcl.2006.07.012

The synthesis and structure-activity relationship (SAR) of novel and highly potent positive allosteric modulators of AMPA receptors, 3-biphenyl-4-yl-4-cyano-5-ethyl-1-methyl-1H-pyrrole-2-carboxylic acid, are described. These studies indicated that higher

Full text of DOI:10.1016/j.bmcl.2006.07.012

Bioorganic & Medicinal Chemistry Letters 16 (2006) 5203–5206
A novel class of positive allosteric modulators of AMPA
receptors: Design, synthesis, and structure–activity relationships of
3-biphenyl-4-yl-4-cyano-5-ethyl-1-methyl-1H-pyrrole-2-carboxylic
acid, LY2059346
Hamideh Zarrinmayeh,a,* Eric Tromiczak,a Dennis M. Zimmerman,a Nancy Rankl,a
Ken H. Ho,a Esteban Dominguez,b Ana Castano,b Ana Escribano,b Carmen Fernandez,b
˜
Alma Jimenez,b William J. Hornbacka and Eric S. Nisenbauma
aEli Lilly and Co, Lilly Research Laboratories, Indianapolis, IN 46285, USA
bEli Lilly and Co, Lilly S.A., Avenida de la Industria, 30, 28108 Alcobendas, Madrid, Spain
Received 26 May 2006; revised 23 June 2006; accepted 5 July 2006
Available online 25 July 2006
Abstract—The synthesis and structure–activity relationship (SAR) of novel and highly potent positive allosteric modulators of
AMPA receptors, 3-biphenyl-4-yl-4-cyano-5-ethyl-1-methyl-1H-pyrrole-2-carboxylic acid, are described. These studies indicated
that higher potency was achieved with ortho substitution of the distal (D) phenyl of the 3-biphenyl ring and resulted in the discovery
of a potent pyrrole LY2059346 (23q), that was selected for further evaluation in in vitro native tissue assays and in vivo experiments.
Ó 2006 Elsevier Ltd. All rights reserved.
L-Glutamate is the major excitatory neurotransmitter in
the central nervous system (CNS) acting on metabotro-
pic and ionotropic glutamate receptors. On the basis of
differences in their molecular sequences and sensitivity
to selective agonists, three subtypes of ionotropic recep-
tors have been identified, including AMPA (a-amino-3-
hydroxy-5-methyl-4-isoxazole-propionic acid), NMDA
(N-methyl-L-aspartate), and kainate receptors.1 Of these
subtypes, AMPA receptors mediated the majority of ra-
pid glutamatergic signaling in the CNS. Functional
AMPA receptors are proposed to be tetrameric struc-
tures, generated by the assembly of one or more distinct
subunits GluR1–4, yielding either homomeric or hetero-
meric configurations.2 Further diversity among native
AMPA receptors is conferred by alternative splicing in
an extracellular domain of each subunit, giving rise to
two isoforms named flip (i) and flop (o).3 Activation
of AMPA receptors is initiated by binding of glutamate
(or agonist) to each of the subunits which directs a con-
formational change in the ion channel gate, permitting
ion flux through the channel pore.4 Termination of cur-
rent flow can occur upon removal of glutamate and
unbinding of agonist (deactivation) or in the continued
presence of glutamate, uncoupling of agonist binding
from the channel gate, permitting channel closure
(desensitization).4
An accumulating body of evidence has indicated that
dysfunction of glutamatergic signaling in the CNS may
contribute to the deficits associated with a variety of
neurological and psychiatric disorders.5 Clinical and
experimental data have suggested that positive modula-
tion of AMPA receptors may be a novel therapeutic ap-
proach in the treatment of such disorders.6 Positive
allosteric modulators of AMPA receptors exert their ef-
fects only in the presence of agonist binding and act to
enhance ion flux through the channel either by attenuat-
ing the deactivation or desensitization process(es).4 Sev-
eral classes of positive allosteric modulators have been
identified and reported, including benzothiadiazides
(cyclothiazide, 1);5 pyrrolidinones (aniracetam, 2a, and
piracetam, 2b)6; and benzamides (CX516, 3)7 (Fig. 1).
However, the value of such tools for understanding
AMPA receptor pharmacology and physiology has been
limited by their relatively week potency. We previously
reported on a class of biarylpropylsulfonamide modula-
tors 4 that possess significantly higher potencies than
Keywords: Positive allosteric modulators; AMPA receptors.
*
Corresponding author. Tel.: +1 317 276 5964; e-mail:
Zarrinmayeh_Hamideh@lilly.com
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.07.012

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H. Zarrinmayeh et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5203–5206
O
S
enesulfonylacetonitrile. This intermediate was subse-
O
O
O
O
S
O
quently converted to pyrrole 9 when reacted with
ethylisocyanoacetate in the presence of DBU. N-methyl-
ation of 9 was followed by Suzuki cross coupling of 10
with phenylboronic acid using palladium tetrakis to
yield product 11. Lithiation of 11 at À78 °C in THF fol-
lowed by treatment with dialkyl disulfide yielded ana-
logs 13a–c. Pyrrole 11 was also converted to the
brominated analog 12 that was converted to the corre-
sponding products 13f,g upon coupling reaction with
tetramethyl tin or diethyl zinc, respectively.10 Hydrolysis
of the esters with lithium hydroxide yielded N-methyl-
pyrrole-2-carboxylic acids 6a–g.
H2N
Cl
NH
H2N
N
N
N
H
O
MeO
O
1 (Cyclothiazide)
2a (Aniracetam)
2b (Piracetam)
Me
Me
O
Me
N
N
N
S
O
O
N
3 (CX516)
4 (Sulfonamide)
D
P
CN
SMe
CN
As shown is Scheme 2, N-methylpyrrole-2-carboxylic
acid ethyl ester 21 was prepared in seven steps from
commercially available 4-benzyloxybenzaldehyde 14
which was converted to acrylonitrile 15 when treated
with p-toluenesulfonylacetonitrile under refluxing condi-
tion. Subsequent reaction of 15 with ethylisocyanoace-
tate resulted in the formation of pyrrole 16 which was
methylated to 17. Bromination of 17 to 18 followed by
reaction of 18 with diethyl zinc using Hartwig’s ligand
under a mild condition resulted in the formation of
5-ethylpyrrole 19. Deprotection of 19 and subsequent
triflation of the phenol 20 resulted in the synthesis of
intermediate pyrrole 21.
HO
HO
O
SMe
N
S
O
5
6a
Figure 1. Structures of positive allosteric modulators of AMPA
receptors.
those of 1–3.8,9 A subsequent high-throughput screen of
Lilly compound collections resulted in the discovery of
2-thiophene carboxylic acid 5 as the lead candidate. Pre-
liminary study of the core heterocyclic ring led to the
discovery of N-methylpyrrole 2-carboxylic acid 6a. In
this communication, we describe the SAR of these
pyrrole 2-carboxylic acid compounds as novel class of
allosteric modulators of AMPA receptors.
Intermediate 21 was converted to pyrrole 22a–q via
Suzuki cross coupling reaction with phenyl boronic
acids using palladium catalyst. Hydrolysis of the esters
by lithium hydroxide resulted in the formation of the
acids 23a–q as shown in Scheme 3.
Synthesis of pyrrole 6a–g is shown in Scheme 1.
Commercially available 4-bromo benzaldehyde 7 was
converted to acrylonitrile 8 upon refluxing with p-tolu-
The activity of allosteric modulators was evaluated
using HEK293 cells stably transfected with recombinant
human homomeric GluR2i, GluR2o, GluR4i, and
Br
Br
Br
CN
c
a
b
CN
BnO
BnO
EtO
O
BnO
H
Tos
N
H
8
9
7
H
b
O
CN
a
c
CN
Br
EtO
CHO
N
H
16
H
TOS
O
14
15
CN
BnO
BnO
e
d
BnO
CN
CN
Br
EtO
N
EtO
EtO
O
CN
CN
CN
N
N
f
d
e
O
O
10
11
12
EtO
EtO
EtO
Et
Br
f
N
N
N
g
h
O
O
O
18
OTf
17
19
HO
CN
CN
CN
h
h
CN
Et
CN
Et
EtO
HO
EtO
g
R
R
R
N
N
N
EtO
EtO
O
O
O
N
N
13a-c
6a-g
13e-g
O
O
21
20
Scheme 1. Synthesis of compounds 6a–g. Reagents and conditions: (a)
p-toluenesulfonylacetonitrile, piperidine, AcOH, toluene, reflux, 92%;
(b) ethylisocyanoacetate, DBU, THF, rt, 99%; (c) MeI, K2CO3, DMSO,
77%; (d) PhB(OH)2, Pd(PPh3)4, dioxane, 80 °C, 95%. (e), NBS, CCl4, rt,
82%. (f) i—LHMS, THF, À78 °C; ii—(R–S)2, À78 °C to rt, 72–80%; (g)
Me4Sn, Pd(PPh3)4, for 13f 62%, Et2Zn, Hartwig’s ligand, for 13g 95%;
(h) LiOH, THF/MeOH/H2O (3:2:1), 50 °C, 90–95%.
Scheme 2. Synthesis of compound 21. Reagents and conditions: (a) p-
toluenesulfonylacetonitrile, piperidine, AcOH, toluene, reflux, 95%; (b)
ethylisocyanoacetate, DBU, THF, rt, 91%; (c) MeI, K2CO3, DMSO,
77%; (d) NBS, THF, 60%; (e) Et2Zn, Hartwig’s ligand, THF, rt, 1 h,
95%; (f) H2, Pd(OH)2/C, EtOH/THF (1:2), 50 psi, rt, 97%; (g) Tf2O,
Et3N, CH2Cl2, rt, 93%.

H. Zarrinmayeh et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5203–5206
5205
Table 2. EC50 values of selected novel AMPA receptor positive
alosteric modulators
OTf
EtO
R
R
CN
CN
Et
CN
Et
a
b
R
EtO
HO
Et
N
N
N
O
O
O
CN
23a-q
21
22a-q
HO
Et
N
Scheme 3. Synthesis of compounds 23a–q. Reagents and conditions:
(a) ArB(OH)2, Pd catalyst, 30–90%; (b) LiOH, THF/MeOH/H2O
(3:2:1), 50 °C, 80–95%.
O
Compound
R
EC50 (nM)
GluR4o receptors. Changes in intracellular calcium ion
(Ca2+) were measured in the presence of glutamate alone
and in response to co-application with the modulators at
concentrations of 0.1–3 M using a FLIPR technology.
Responses were normalized to those produced by co-ap-
plication of 5 and 100 lM glutamate (maximum re-
sponse) and 100 lM glutamate alone (minimum
response). The in vitro potency of the compounds was
reported as EC50 values in nanomolars as shown in
Tables 1 and 2.
iGluR4i iGluR4o iGluR2i iGluR2o
6ga
H
213
1143
2085
790
1766
>3000
>3000
1181
>3000
>3000
386
103
412
965
270
423
391
280
83
398
1647
2133
990
370
924
902
36
23ab
23bb
23cb
23db
23eb
23fb
p-Me
m-Me
o-Me
p-OMe
738
m-OMe 1051
o-OMe
o-OEt
o-SMe
p-F
673
158
105
242
223
80
23gb
23hb
23ib
23jb
23kb
23lb
23mb
23nb
23ob
23pb
23qc
LY2059346
146
77
57
32
2947
>3000
185
345
426
111
142
186
42
1035
1490
143
984
900
55
m-F
o-F
In this study, the 2-carboxylic acid, 3-biaryl, and 4-cya-
no groups substitutions on the pyrrole ring were kept
constant since preliminary studies revealed the necessity
of such functionalities for minimum activity. We first fo-
cused on the evaluation of the 5-thiomethyl functional-
ity, its significance and possible replacement with more
neutral and metabolically stable groups.
p-Cl
910
1082
225
373
631
56
>3000
>3000
157
m-Cl
o-Cl
p-CN
m-CN
o-CN
>3000
>3000
53
423
395
82
370
974
73
a EC50 value (n P 10).
b EC50 value (n P 3).
c EC50 value (n P 20).
As shown in Table 1, parent pyrrole 6a exhibited reason-
able activity with EC50 values of 394, 1301 nM at
GluR4i/o and 174 nM, 473 nM at GluR2i/o, respective-
ly. Flip splice variant selectivity was observed for both
subunits with higher potency at GluR2i. Larger thio-
alkyl groups were not tolerated as shown in 6b,c. Lack
of activity of 5-proto pyrrole 6d demonstrated the need
for substitution of 5 position with groups larger than
proton. Alkyl-substituted analogs 6f,g were prepared
and tested. Even though the activity of 5-methyl analog
6f was modest, the 5-ethyl analog 6g showed a desired
improvement in activity compared to that of the parent
6a, especially at the flip isoform with EC50 values of 213
and 103 nM at GluR4i and GluR2i receptors, respec-
tively. In addition, 6g was the first pyrrole analog to
possess similar activity to that of the potent lead thio-
phene 5. Ethyl functionality was also superior to other
groups that were studied and not presented in Table 1.
As a result, ethyl was selected as the desired functional-
ity to substitute 5 position of the pyrrole for subsequent
SAR studies.
Table 1. EC50 values of selected novel AMPA receptor’s positive
allosteric modulators
CN
HO
R
N
O
Compound
R
EC50 (nM)
iGluR4i iGluR4o iGluR2i iGluR2o
5a
—
214
1559
1301
NT
97
174
309
473
6ab
6bb
6cb
6db
6ec
6fb
6gc
S-Me 394
S-Et 619
S-i-Pr >3 lM
To optimize substituents and the pattern of substitution
on the distal phenyl ring D of the 3-biaryl, we prepared
analogs 23a–q. As shown in Table 2, methyl substitution
of the phenyl ring provided pyrrole 23a–c with only
modest activities. Methoxy analogs 23d–f also demon-
strated similar potencies. However, some pattern of sub-
stitution was observed in that higher activity resided in
ortho-substituted analogs. Therefore, analogs 23g,h were
prepared and displayed an increase in activity at all
receptors. In addition, thiomethyl anolog 23h possessed
NT
NT
>3 lM
>3 lM
1689
2388
1766
>3 lM
>3 lM
>3 lM
308
>3 lM
>3 lM
>3 lM
391
H
>3 lM
679
532
Br
Me
Et
213
103
398
NT, not tested.
a EC50 value (n P 100).
b EC50 value (n P 3).
c EC50 value (n P 10).

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H. Zarrinmayeh et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5203–5206
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significant potency and selectivity for the flop isoform.
More ortho preference for substitution of the phenyl D
ring was observed with fluorinated analogs 23i–k where
ortho fluoro anaolog 23k was the most potent of the
three regioisomers. The same pattern of substitution
was also observed with chlorinated analogs 23l–n, where
ortho chloro analog 23n was significantly more potent
than the other regioisomers 23l,m. Within the cyano
analog series 23o–q, of the three regioisomers, ortho cya-
no analog 23q showed the highest activities at GluR4i/o
and desired activities at GluR2i/o. A unique property of
23q was its equally high potency at all receptor subtypes.
This broad range of activity has prompted further eval-
uation of 23q, LY2059346,11 in in vivo assays.
In summary, we have designed, synthesized, and eval-
uated a novel class of positive allosteric modulators of
AMPA receptors. Initial SAR studies on the N-methyl-
pyrrole 6a resulted in replacement of thiomethyl with
ethyl functionality at position 5. Further SAR on the
distal phenyl ring D of the biphenyl ring system at 3
position resulted in analogs where their higher poten-
cies came from not only the substituents, but also from
the pattern of substitution. ortho Substitution of ring
D with ethoxy, thiomethoxy, fluoro, and cyano func-
tionalities, independent of the electronic features of
the substituents, resulted in significant increase of
potency. Isoform selectivity was not predominant in
this SAR and only a mode preference for GluR2 ver-
sus GluR4 receptors was observed. The most potent
pyrrole 23q, LY2059346, exhibited equally high poten-
cy at all four homomeric receptors and has been select-
ed for further in vitro and in vivo biological studies.
10. Uno, H.; Tanaka, M.; Inoue, T.; Ono, N. Synthesis 1999, 3,
471.
11. 1H NMR (DMSO-d6, 500.0 MHz): d 1.26 (t,3H), 2.87 (q,
2H), 3.88 (s, 3H), 7.52 (d, 2H), 7.62 (t, 1H), 7.65 (d, , 2H),
7.71 (d, 1H), 7.83 (t,1H), 7.99 (d,1H), 12.85 (s, 1H); IR
(KBr): 2938, 2225, 1654, 1476, 1440, 1356, 1280, 1252,
1166, 763cmÀ1. PCT Int. Appl. WO 2005040110.
References and notes
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