The identification of structurally novel, selective, orally bioavailable positive modulators of mGluR2

Article abstract of DOI:10.1016/j.bmcl.2009.11.032

The optimisation of an HTS hit series (1) leading to the identification of structurally novel, selective, orally bioavailable mGluR2 positive modulators GSK1331258 and GSK1331268 is described. Structure-activity relationships, attenuation of dopaminergic

Full text of DOI:10.1016/j.bmcl.2009.11.032

Bioorganic & Medicinal Chemistry Letters 20 (2010) 759–762
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
The identification of structurally novel, selective, orally bioavailable
positive modulators of mGluR2
Pier L. D’Alessandro a, Corrado Corti b, Adelheid Roth b, Annarosa Ugolini b, Anna Sava b, Dino Montanari b,
Federica Bianchi b, Stephen L. Garland c, Ben Powney c, Emma L. Koppe c, Magalie Rocheville c,
Greg Osborne c, Paloma Perez d, Jesús de la Fuente d, Maite De Los Frailes d, Paul W. Smith a,
Clive Branch a, David Nash a, Stephen P. Watson a,
*
a GlaxoSmithKline, Neurosciences Centre of Excellence for Drug Discovery, New Frontiers Science Park, Harlow, UK
b GlaxoSmithKline, Neurosciences Centre of Excellence for Drug Discovery, Medicines Research Centre, Verona, Italy
c GlaxoSmithKline, Molecular Discovery Research, New Frontiers Science Park, Harlow, UK
d GlaxoSmithKline, Molecular Discovery Research, Tres Cantos, Madrid, Spain
a r t i c l e i n f o
a b s t r a c t
Article history:
The optimisation of an HTS hit series (1) leading to the identification of structurally novel, selective, orally
bioavailable mGluR2 positive modulators GSK1331258 and GSK1331268 is described. Structure–activity
relationships, attenuation of dopaminergic activity, and potentiation of mGluR2 responses in rat hippo-
campal MPP-DG synapses are also reported.
Received 6 October 2009
Revised 6 November 2009
Accepted 9 November 2009
Available online 26 November 2009
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Metabotropic glutamate receptor ligand
Metabotropic glutamate receptor
Positive allosteric modulator
mGluR
mGlu
mGluR2
There is a strong body of evidence that modulation of metabo-
tropic glutamate receptors could afford novel approaches for the
treatment of neurodegenerative and neuropsychiatric diseases.1–4
The type 2 metabatropic glutamate receptor is of particular inter-
compounds such as 2 from Pfizer (Fig. 2), which prompted us to re-
port the work from our laboratories.
Herein we describe a novel series of 2-piperidine/piperazine-
methylene-benzimidazole mGluR2 positive modulators derived
from a high throughput screening (HTS) hit series exemplified by
compound 1 (Fig. 1). The synthesis, structure–activity relationships
(SAR) investigation, and the control of dopaminergic off-target
activities of this series is described: culminating in the identifica-
tion of the selective, orally bioavailable mGluR2 positive modula-
tors GSK1331258 and GSK1331268.
est, indeed,
a prodrug of the orthosteric mGluR2/3 agonist
LY404039 (Lilly) exhibited statistically significant improvements
in both positive and negative symptoms of schizophrenia in phase
II trials.5 Additionally a study using receptor deficient mice showed
that antipsychotic effects of the mGluR2/3 receptor agonist
LY404039 are dependent on mGluR2 and not mGluR3 receptors.6
Positive allosteric modulation could offer a superior mechanism
of potentiating mGluR2 function than classical orthosteric agon-
ism. The advantages positive modulation may afford, in terms of
enhanced selectivity and spatial and temporal control, have been
described extensively elsewhere7,8 suffice it to say that this mech-
anism has received a great deal of attention in recent patent and
scientific literature. Notable recent reports of mGluR2 positive
modulators include the pioneering work of Lilly, exemplified by
pyridylsulfonamides LY48737910,9–13 and, a promising report14 of
mGluR2 Pos Mod pEC50 6.9
mCluR2 Ag
mGluR1 Ag
mGluR1 Ant
mGluR5 Ag
mGluR5 Ant
mGluR5 Pos Mod pEC50 < 4.8
Dopamine D2 pIC50 7.4
pEC50 5.4
pEC50 < 4.8
pEC50 < 4.8
pEC50 < 4.8
pEC50 < 4.8
N
N
N
1
F
Mwt
PSA
323
21
cLogP 4.2
Solubility 31ug/ml
* Corresponding author. Tel.: +44 1279 875004.
E-mail address: steve.p.watson@gsk.com (S.P. Watson).
Figure 1. Structure and profile of HTS hit compound 1.
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.11.032

760
P. L. D’Alessandro et al. / Bioorg. Med. Chem. Lett. 20 (2010) 759–762
N
N
N
N
O
O
N
N
N
N
Cl
N
O
H
S
O
H
O
N
LY487379
GSK1331258
mGluR2 pEC50 = 7.1
2
CF3
Figure 2. Exemplar mGluR2 positive modulators and GSK1331258.
Me
Me
O
H
N
N
R2 NH R1
N
R1
N
N
a,b
R2
3
7-26
R3
H
H
N
O
H
N
R2 NH R1
N
c,d
N
N
R1
N
N
R1
N
a,b
R2
R2
4
27-34
Scheme 1. Reagents and conditions: (a) DCM/AcOH = 98:2 (V/V), rt, 15 min; (b) MP–BH(OAc)3 (5.0 equiv), microwave irradiation, 110 °C, 10 min; 50–90%; (c) 60% NaH in
mineral oil, DMF/THF = 30:70 (V/V), 10 min at rt; (d) R3X, microwave irradiation, 100 °C, 10 min (X = Cl, Br), 40–75%.
From a HTS in a chimeric CHO cell line using a FLIPR readout15
several promising mGluR2 modulator chemotypes were identified.
Compound 1 was selected for further progression based on its dis-
playing the best balance of pharmacology, selectivity, developabil-
ity and properties suitable to afford good CNS penetration (Mol. wt
<400, c Log P <4.5 and PSA <80). It was anticipated that dopaminer-
gic activity associated with this template could be readily
attenuated.
The desired mechanism of action of this compound/chemotype
was confirmed by blockade of the positive modulation by the
orthosteric mGluR2 antagonist16 LY341495. A medicinal chemistry
programme was initiated with the aim of identifying selective,
orally bioavailable and brain penetrant mGluR2 positive modula-
tors suitable for progression to studies in in vivo models.
Compounds were typically prepared according to the chemistry
shown in Schemes 1 and 2.17
preparation cannot be discounted). Compounds typically potenti-
ated an EC20 of glutamate to 70–100% of the glutamate EC100 re-
sponse, but there is no apparent correlation between this efficacy
of potentiation and pEC50
.
Close analogues of the chemotype exemplified by compound 1
are known to display activities at dopaminergic receptors,18–20
hence it was important to investigate and attenuate any dopami-
nergic activity21 such that dopaminergic pharmacology could not
confound readout in any downstream in vivo models. Data in Ta-
ble 1 illustrates that piperidine based analogues of compound 1 in-
deed typically demonstrated significant D2 and D3 antagonism.
However this activity could be successfully attenuated in the
NH
Simple reductive amination of 1-methyl-3-formyl benzimid-
azole (3) with appropriate amines allowed investigation of the
aryl-piperazine/piperidine region of the template (Table 1). Varia-
tion of the benzimidazole N-substituent was achieved via a final
step alkylation of intermediates generated by reductive amination
of 2-formylbenzimidazole (4) (Table 2). Variation of the benzimid-
azole benzosubstituents (Table 2) was effected via reaction of N-
methyl bis-anilines (5) and the corresponding piperazine/piperi-
dine derived glycine (6).
R6
O
NH
a
N
NO2
N
N
OH
Ar
Ar
g
b
O
NH
N
R6
+
Cl
N
Ar
NH2
Table 1 illustrates the profiles of a representative set of ana-
logues of lead compound 1, in which the piperazine, and corre-
sponding piperidine, aryl ring is modified. There are apparent
differences in SAR between the piperazines and piperidines. Piper-
idines are typically more potent mGluR2 positive modulators than
the corresponding piperazines (e.g., contrast the potency of the
two simple phenyl analogues 7 and 17) but in some instances
potencies are comparable (e.g., 4-trifluoromethyl analogues 11
and 22). Most compounds also demonstrated weak agonism of
the receptor, this could be due to low level allosteric agonism
(but effects due to contaminating traces of glutamate in the assay
6
5
c,d
N
N
R6
N
N
Ar =
N
Cl
CF3
37-41
Ar
Scheme 2. Reagents and conditions: (a) BrCH2COOH, MeCN, rt, overnight, 62%; (b)
1-chloro-N,N,2-trimethyl-1-propen-1-amine MeCN, rt, 40 min; (c) MeCN, 20 min at
rt; (d) MeCN, microwave irradiation, 140 °C, 20 min; (b–d in one pot-11–25%
overall); (g) Fe, NH4Cl, 50% MeOH/H2O (V/V), 2 h at reflux, 40–60%.

P. L. D’Alessandro et al. / Bioorg. Med. Chem. Lett. 20 (2010) 759–762
761
Table 1
Positive modulation of EC20 glutamate15 at mGluR2 and dopaminergic antagonism20 by piperazine and piperidine analogues
Me
N
N
X
N
Ar
mGluR2 pos. mod.
Compound
Ar
X
mGluR2 pos. mod.
pEC50
mGluR2 agonist.
pEC50
D2
pKi
D3
pKi
D4 pKi
max.
7
1
8
9
10
11
12
13
14
15
16
17
18
19
20
Phenyl
2-F-Phenyl
4-Chloro-phenyl
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
CH
N
N
N
N
N
N
N
N
N
6.7
6.9
7.1
6.5
7
89
91
91
94
94
110
92
76
84
70
5.2
5.4
5.3
5.2
5.6
5.7
5.4
5.5
5.6
6.2
<4.8
<4.8
5.2
<4.8
<4.8
5.7
5.9
5.6
5.2
5.7
5.2
7.6
7.4
7.8
<5.5
6.9
6.6
7.4
6.4
7.1
6.1
6.9
7.0
6.5
6.3
<5
<5
5.8
<5
7.0
7.2
2-Trifluoromethyl-phenyl
3-Trifluoromethyl-phenyl
4-Trifluoromethyl-phenyl
3-Chloro-phenyl
1,3-Benzothiazole-2-yl
1,2-Benzisothiazole-3-yl
3-Chloro-5-(trifluoromethyl)-2-pyridinyl
2-Pyridyl
6.6
6.9
6.6
8.1
6.7
5.6
7
7
6.8
6.7
7.4
<5.5
5.3
6.1
6.1
6.5
7.1
6.9
6.9
6.7
6.5
6.3
4.8
<4.9
5.4
5.8
n/a
100
64
Phenyl
2-F-Phenyl
4-Chloro-phenyl
<5
<5
6.9
6.2
6.2
5.6
6.6
6.4
6.1
6.0
2-Trifluoromethyl-phenyl
3-Chloro-5-(trifluoromethyl)-2-pyridinyl
5-Chloro-2-pyrimidinyl
4-Chloro-2-fluoro-phenyl
4-Trifluoromethyl-phenyl
1,3-Benzothiazole-2-yl
5-(Trifluoromethyl)-2-pyridinyl
2-Pyridyl
84
71
69
69
84
88
81
100
n/a
GSK1331258
GSK1331268
5.3
<5
5.3
<5
21
22
23
24
25
26
<5
<5
<5
<5
<5
<5
N
N
N
4-Cyanophenyl
majority of the piperazines. D4 antagonism was more persistent
within the analogues, but was reduced to a very low level in com-
pounds such as GSK1331268.
Based on the data in Table 1, the 5-chloro-2-pyrimidinyl and 3-
chloro-5-(trifluoromethyl)-2-pyridinyl substituted piperazine
fragments of compounds GSK1331258 and GSK1331268 were se-
lected for use in further studies (see Table 2), as they represented
the best balance of high mGluR2 activity and low dopaminergic
activity.
Compounds 27–34 (Table 2) illustrate that increasing the size of
the benzimidazole N-substituent affords little enhancement of
mGluR2 pharmacology. Interestingly the two N-unsubstituted ana-
logues 35 and 36 have effectively no mGluR2 activity.
Table 2 also reports data for modifications to benzimidazole
ring substitution of 3-chloro-5-(trifluoromethyl)-2-pyridinyl
piperazines (37–41). Similarly to further Benzimidazole N-substi-
tution, these ring substitutions conferred no significant improve-
ment on mGluR2 pharmacology.
In contrast to the majority of analogues GSK1331258 and
GSK1331268 demonstrated good in vitro metabolic stability (data
not shown) and were selected for progression for in vivo pharma-
cokinetic investigation. In rat, in a 1 mg/kg oral cassette study both
compounds showed a promising profile, viz. GSK1331258 exhib-
ited low clearance (CLb 18.0 ml/min/kg), a half life of 4.1 h, a
brain:blood ratio of 3.2 with brain Cmax of 147 ng/g, and bioavail-
ability of 58%; GSK1331268 exhibited moderate clearance (CLb
32.0 ml/min/kg), a half life of 1.0 h, a brain–blood ratio of 1.3 with
brain Cmax of 32 ng/g and bioavailability of 22%.
Table 2
FLIPR pEC50 positive modulator activity as potentiation of EC20 glutamate15
R3
N
N
X
N
N
R6
N
R5
R4
Furthermore GSK1331258 and GSK1331268 preserved the
mGluR selectivity observed in initial hit compound 1, with neither
compound displaying significant activity at mGluRs1, 4 or 5 in ago-
nist, antagonist or positive modulator modes (data not shown).
The pharmacology of GSK1331268 was characterised further on
Medial Perforant-Path Dentate Gyrus (MPP-DG) Synapses in rat
hippocampal slices,22 in which synaptic transmission is inhibited
by activation of group II mGluR receptors.23
X
R3
R4
R5
R6
mGluR2 pos.
mod. pEC50
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
C
C
N
C
N
N
N
C
C
N
C
C
C
C
C
–Et
2-Cl
2-Cl
5-Cl
2-Cl
5-Cl
5-Cl
5-Cl
2-Cl
2-Cl
5-Cl
2-Cl
2-Cl
2-Cl
2-Cl
2-Cl
5-CF3
5-CF3
H
5-CF3
H
H
H
H
H
H
H
H
H
H
H
7.2
6.9
6.9
6.8
6.8
6.3
6.1
5.6
4.9
4.9
6.6
7.3
5.7
6.9
6.7
–CH2Ph
–CH2Ph
–(CH2)2OCH3
–Et
–(CH2)2OCH3
–nProp
–nProp
–H
–H
Me
Me
Me
H
H
Orthosteric agonist LY35474024,25 inhibits MPP–DG fEPSP with
an EC50 of 134 nM. Co-application of LY354740 with GSK1331268
5-CF3
5-CF3
H
5-CF3
5-CF3
5-CF3
5-CF3
5-CF3
(10 lM) shifts the concentration–response curve for LY354740 to
the left by threefold, to 43 nM (see Fig. 3).
In summary a successful SAR investigation of a HTS hit series (1)
was conducted. Understanding of factors controlling mGluR2 activ-
ity and dopaminergic activity allowed the identification of struc-
turally novel, selective,26 and orally bioavailable mGluR2 positive
5-OMe
5-CN
5-CF3
6-F
Me
Me
5-F

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P. L. D’Alessandro et al. / Bioorg. Med. Chem. Lett. 20 (2010) 759–762
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15. mGluR2 pharmacology characterised using a FLIPR (Fluorometric Imaging Plate
Reader) assay utilising CHO cells stably expressing both the human mGluR2
receptor and Ga16 G-protein. 11 Point dose–response curves were generated,
and for positive modulation in presence of an EC20 of glutamate. Values are
typically average of four or more experiments. Positive Modulation Max
Response is calculated relative to response of EC100 of glutamate.
16. Johnson, B. G.; Wright, R. A.; Arnold, M. B.; Wheeler, W. J.; Ornstein, P. L.;
Schoepp, D. D. Neuropharmacology 1999, 38, 1519.
17. Structure of final compounds confirmed via H NMR and LC–MS analysis.
18. Stewart, A. O.; Cowart, M. D.; Moreland, R. B.; Latshaw, S. P.; Matulenko, M. A.;
Bhatia, P. A.; Wang, X.; Daanen, J. F.; Nelson, S. L.; Terranova, M. A.; Namovic,
M. T.; Donnelly-Roberts, D. L.; Miller, L. N.; Nakane, M.; Sullivan, J. P.; Brioni, J.
D. J. Med. Chem. 2004, 47, 2348.
Figure 3. The effect of GSK1331268 on excitatory post-synaptic potentials (fEPSP)
at MPP–DG synapses in rat hippocampal slices (data points indicate the mean
-
SEM of fEPSP peak amplitude expressed as a percentage of predrug control baseline;
n = 9–11 slices per data point).
19. Nakane, M.; Cowart, M. D.; Hsieh, G. C.; Miller, L.; Uchic, M. E.; Chang, R.;
Terranova, M. A.; Donnelly-Roberts, D. L.; Namovic, M. T.; Miller, T. R.; Wetter,
J. M.; Marsh, K.; Stewart, A. O.; Brioni, J. D.; Moreland, R. B. Neuropharmacology
2005, 49, 112.
20. Cowart, M.; Latshaw, S. P.; Bhatia, P. A.; Daanen, J. F.; Rohde, J. J.; Nelson, S. L.;
Patel, M. V.; Kolasa, T.; Nakane, M.; Uchic, M. E.; Miller, L. N.; Terranova, M. A.;
Chang, R.; Donnelly-Roberts, D. L.; Namovic, M. T.; Hollingsworth, P. R.;
Martino, B. R.; Lynch, J. J.; Sullivan, J. P.; Hsieh, G. C.; Moreland, R. B.; Brioni, J.
D.; Stewart, A. O. J. Med. Chem. 2004, 47, 3853.
21. D2 and D3, [35S]-GTPgS assay CHO membranes expressing G0 with hD3 or hD2
transfected; D4 FLIPR assay in HEK293 cells stably expressing hD4 and
transiently expressing the chimeric Gqo5 G-protein, Values are typically an
average of four or more experiments generated from 11 point dose–response
curves.
22. Schaffhauser, H.; Rowe, B. A.; Morales, S.; Chavez-Noriega, L. E.; Yin, R.; Jachec,
C.; Rao, S. P.; Bain, G.; Pinkerton, A. B.; Vernier, J. M.; Bristow, L. J.; Varney, M.
A.; Daggett, L. P. Mol. Pharmacol. 2003, 64, 798.
modulators GSK1331258 and GSK1331268. Indeed these com-
pounds have suitable profiles for investigation of the effects
mGluR2 positive modulation in in vivo disease models.
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