Hit-to-lead optimization of disubstituted oxadiazoles and tetrazoles as mGluR5 NAMs

Article abstract of DOI:10.1016/j.bmcl.2010.04.075

Here we report the discovery and early SAR of a series of mGluR5 negative allosteric modulators (NAMs). Starting from a moderately active HTS hit we synthesized 3,5-disubstituted-oxadiazoles and tetrazoles as mGluR5 NAMs. Based on the analysis of ligand e

Full text of DOI:10.1016/j.bmcl.2010.04.075

Bioorganic & Medicinal Chemistry Letters 20 (2010) 3737–3741
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Hit-to-lead optimization of disubstituted oxadiazoles and tetrazoles
as mGluR5 NAMs
*
Gábor Wágner , Csaba Wéber, Olga Nyéki, Katalin Nógrádi, Attila Bielik, László Molnár, Amrita Bobok,
Attila Horváth, Béla Kiss, Sándor Kolok, József Nagy, Dalma Kurkó, Krisztina Gál, István Greiner,
}
Zsolt Szombathelyi, György M. Keseru, György Domány
Gedeon Richter Plc, Budapest 10, PO Box 27, H-1475, Hungary
a r t i c l e i n f o
a b s t r a c t
Article history:
Here we report the discovery and early SAR of a series of mGluR5 negative allosteric modulators (NAMs).
Starting from a moderately active HTS hit we synthesized 3,5-disubstituted-oxadiazoles and tetrazoles as
mGluR5 NAMs. Based on the analysis of ligand efficiency and lipophilic efficiency metrics we identified a
promising lead candidate as a starting point for further optimization.
Received 10 March 2010
Revised 16 April 2010
Accepted 16 April 2010
Available online 22 April 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
mGluR5
Negative allosteric modulator
Parallel synthesis
Glutamate is a major excitatory neurotransmitter in the mam-
malian central nervous system (CNS) and binds to neurons in the
CNS, thereby activating cell surface receptors. These receptors
can be divided into two major classes, ionotropic and metabotropic
glutamate receptors, based on the structural features of the recep-
tor proteins.
Metabotropic glutamate receptors are a family of eight G-pro-
tein coupled receptors which are classified into three groups
according to their sequence homology, effector coupling and phar-
macology. Group I mGlu receptors (mGluR1 and mGluR5) are pos-
itively coupled to phospholipase C; group II mGlu receptors
(mGluR2 and mGluR3) and group III mGlu receptors (mGluR4,
mGluR6, mGluR7 and mGluR8) are negatively coupled to adenylate
cyclase.1
Activation of group I mGlu receptors leads to a transient in-
crease in intracellular calcium via the production of inositol-tris-
phosphate. Generally, it has been shown that activation of group
I receptors enhances or facilitates the excitatory effects of gluta-
mate by modulation of ion channel activity.2 Although group I
mGlu receptors are related phylogenetically, both mGlu1 and
mGlu5 receptors have a distinct expression pattern in the brain,
which clearly suggests their different roles in nervous system func-
tion. mGlu5 receptors are found most abundantly throughout the
cerebral cortex, hippocampus, caudate-putamen, nucleus accum-
bens, and lamina I–III of the spinal cord.3
A large body of preclinical data were reported with mGluR5-
antagonists and mGluR5 knock-out mice emphasising the mGlu5
receptor as a potentially important therapeutic target for several
CNS disorders including anxiety,4 pain,5 depression,6 epilepsy,7
neurodegeneration,8 Parkinson’s disease9 and cocaine-depen-
dence.10 There is a large unmet medical need for new anti-anxiety
agents that relieve symptoms quickly and have no (benzodiaze-
pine-like) side effects. Recent findings have suggested an impor-
tant role for the mGlu5 receptor in anxiolysis.
According to the literature, the first selective non-competitive
mGluR5 antagonist compound, 2-methyl-6-(phenylethynyl)pyri-
dine (MPEP, 1)11 has a very broad and potent anxiolytic-like activ-
ity in rodent models of anxiety. It has a short onset of action and
lacks the potential to induce sedation or psychotomimetic effects,
in contrast to the benzodiazepines.12 (Fig. 1) Before cloning of
metabotropic glutamate receptor subtypes, the non-GABAergic
* Corresponding author.
E-mail address: g.wagner@richter.hu (G. Wágner).
Figure 1. Proof of concept mGluR5 antagonists.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.04.075

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G. Wágner et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3737–3741
agent fenobam (2) was investigated in a double-blind placebo-con-
trolled clinical trial and showed efficacy and onset of action com-
parable with diazepam.13 In 2005 Roche reported that similar to
MPEP, fenobam is a negative allosteric modulator of mGluR514 that
provided clinical proof of principle for the mGluR5 approach in
anxiety. Consequently, several pharmaceutical companies have
initiated mGluR5 discovery programmes.15
The high throughput screening (HTS) of our corporate com-
pound library resulted in several hits. This Letter describes the
hit-to-lead optimization process of 3,5-disubstituted-oxadiazoles
represented by compound 3. (Fig. 2) Optimization of other clusters
were reported in several patent applications.16
Multiple objectives were set for our hit-to-lead optimization,
including the identification of the optimal central heterocycle,
the cyclic secondary amide, the substitution pattern of the aro-
matic ring etc. In order to achieve these goals we utilized a parallel
synthesis strategy, synthesizing both oxadiazoles and tetrazoles
as racemates. The hit-to-lead process was monitored calculating
size independent ligand efficiency (SILE)19 and a lipophilic ligand
efficiency metrics (LELP)20 recently introduced by us. LELP was de-
fined as the logP/LE ratio indicating the price of ligand efficiency
paid in logP. Consequently, the higher the absolute value of LELP
the less drug-like the lead compound.
Scheme 1 demonstrates the synthetic pathways leading to the
oxadiazole series.21 The synthesis of amines 8 was realized by pre-
paring amidoximes 6 from suitable nitriles 5 refluxed in methanol
with hydroxylamine, followed by acylation with Boc-protected
pipecolic acid, nipecotic acid, proline or thioproline under mild
conditions.22 Cyclocondensation of O-acylamidoximes using tetra-
butylammonium fluoride then provided the oxadiazoles ring23 (7).
Finally, amines 8 were obtained by deprotection of 7. In the last
step we used parallel synthesis to prepare final products (9). Acyl-
ation of the secondary amines 8 can be accomplished with carbox-
ylic acids (RCOOH) activated by EDC in the presence of a base (e.g.
TEA). Various commercially available substituted or unsubstituted
alkyl-, cycloalkyl-, aryl-, heteroaryl-carboxylic acids were used.
Final products (9) can be obtained in appropriate purity there-
fore after concentration of the solution. Biological experiments
were carried out without further purification. All compounds were
characterized by LC–MS.
Yields and purity data of the parallel synthetic step are summa-
rized in Table 1. The purity of most oxadiazoles was greater than
95%, and yields were also sufficient.
For the final acylation step, 15 amines and 61 acids were se-
lected, 915 reactions were performed and 701 endproducts were
isolated, 656 compounds fulfilled the purity criterion (>85%, LC–
MS) and were tested. The Ki values of the compounds,22 with only
a couple of exceptions, were above 200 nM, Affinities and func-
tional activities of the most active compounds are presented in
Table 2.
The 2-piperidinyl, 2-pyrrolidinyl, and 4-thiazolidinyl deriva-
tives were found to be the most active compounds, while 3-pipe-
ridinyl derivatives were generally inferior. The optimal acyl
groups depended on the nature of the saturated heterocycles.
The 2-piperidines with cycloalkylcarbonyl groups, (like cyclobutyl-
carbonyl (9c, 9e) and cyclopentylcarbonyl) gave active compounds
while 2-pyrrolidines acylated with 2-furoyl and 2-thiophenecar-
bonyl (9b, 9d) groups had higher affinity towards the target. The
Table 1
Purity and yield data of for the final products (9) obtained by parallel synthesis
Core Cyclic sec.
Amine (num.
of subtypes)
Num. of
compound
Purity (%)
Yield (%)
>95
85–
95
<85 >60
30–
60
<30
8
Oxa
Oxa
Oxa
Oxa
Tet
Tet
Tet
Tet
3-Pip (8)
2-Pip (4)
2-Pyr (3)
Sum
3-Pip (2)
2-Pip (10)
2-Pyr (9)
Sum
274
244
183
701
57
344
228
629
1330
71.5 22.3 6.2
75 19.7 5.3
69.4 22.4 8.2
72.1 21.4 6.4
89.5 10.5
78.2 20
53.1 45.2 1.7
70.1 28.3 1.6
71.2 24.7 4.1
57.3 34.7
43.4 42.2 14.4
32.2 48.1 19.7
45.9 40.8 13.3
15.8 63.2 21
13.4 57.3 29.4
31.1 53.5 15.4
0
1.7
20
56.4 23.5
33.7 48.2 18.1
Total
Oxa, oxadiazoles; Tet, tetrazoles; pip, piperidines; pyr, pyrrolidines; num, number;
comp, compounds.
Figure 2. The HTS hit served as starting point of hit-to-lead optimization.
Scheme 1. Reagents and conditions: (a) Hydroxylamine, K2CO3, MeOH, rt, 3 h, 90%; (b) Boc-pipecolic acid or Boc-nipecotic acid or Boc-proline or Boc-thioproline, i-
butylchloroformate, N-Me-morpholine, DMF, 0 °C, 30 min, then tetrabutylammonium fluoride, 0 °C, 3 h, 60%; (c) HCl, EtOAc, rt, 3 h, 50–70%; (d) RCOOH, EDC, TEA, CH2Cl2, rt,
overnight, parallel synthesis.

G. Wágner et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3737–3741
3739
Table 2
Structures, activities and ligand efficiencies of the most potent oxadiazoles
rmGluR5
pKi17
pIC5018
Compound
Y
R
SILE19
LELP20
9a
9b
9c
9d
3-Cl
4-Thiazolidinyl
2-Pyrrolidinyl
2-Piperidinyl
2-Pyrrolidinyl
7.08
6.91
6.90
6.84
6.12
5.18
6.31
6.00
3.79
3.86
4.26
4.00
5.70
5.05
3.02
4.32
3-Cl
3-Me
3-Me
9e
9f
3-MeO
3-Me
3-Cl
2-Piperidinyl
2-Pyrrolidinyl
4-Thiazolidinyl
6.74
6.73
6.71
6.30
4.52
5.98
3.93
4.15
3.60
3.09
1.77
4.19
9g
9h
9i
3-Cl
2-Pyrrolidinyl
2-Piperidinyl
6.71
6.70
4.52
5.76
3.74
3.59
4.50
5.61
3-CN
most active compound of this series was, however, a thiazolidine
derivative 9a containing a thiophene-2-carbonyl group.
isomer (R-17c; optical purity: >90%, rmGluR5 pKi: 7.24; S-17c;
optical purity: >90%, rmGluR5 pKi: 6.13). After having prepared
several other pairs, we concluded that this observation could be
extended to the whole chemical series and therefore mostly race-
mic compounds were prepared during this stage of hit-to-lead
optimization.
After having established the SAR of the oxadiazole library, we
focused on the synthesis of the more promising tetrazole library
(Scheme 2). Aldehydes of formula 11 were prepared by the reduc-
tion of Weinreb amides and were not isolated. Weinreb amides
were prepared by reacting Boc-protected amino acids (10) with
N,O-dimethylhydroxylamine hydrochloride in the presence of DI-
PEA, DCC and DMAP25 and were subsequently reduced in anhy-
drous THF with LAH. Hydrazones 12 were prepared by reacting
aldehydes 11 with p-toluenesulfonhydrazide in ethanol.26 Diazo-
nium salts 14 were obtained from suitable substituted anilines
13, and converted to tetrazoles 15 by reaction with hydrazones
12 in aqueous ethanol with sodium hydroxide.27 Amines 16 were
This compound showed almost a 0.5 pKi unit improvement in
binding affinity while its ligand efficiency and lipophilic efficiency
were weaker than that of the HTS hit. Analysing SILE and LELP val-
ues we concluded that despite of their somewhat reduced affinity
9c and 9f could be considered as more promising leads. Higher SILE
and lower LELP of these compounds suggest those being better
starting points for further optimizations.
The modification of the central heterocycle of the original HTS
hit (3) to tetrazole, a known oxadiazole bioisostere, resulted in a
second starting point for lead discovery 17c. Since all the com-
pounds synthesized in the oxadiazole library were racemates, the
first question was whether enantiomers have different biological
activities. Since the synthetic protocol to 17c (Scheme 2)24 was
not affected by stereochemical inversion we were able to prepare
its pure enantiomers with known absolute configuration. Testing
the enantiomers of 17c revealed that the R enantiomer is the active
Scheme 2. Reagents and conditions: (a) i—N,O-dimethylhydroxylamine hydrochloride, DIPEA, DCC, DMAP, 0 °C, 30 min, then rt, 3 h, 85%; ii—LAH, THF, 0 °C, 10 min, 90%;
(b) p-toluenesulfonhydrazide, ethanol, 25 °C, 30 min, 90%; (c) NaNO2, hydrochloric acid, water, 0 °C; (d) NaOH, aqueous ethanol, 0 °C, 30 min, then 25 °C, 1 h, 60%, two steps
(c and d); (e) HCl, EtOAc, rt, 3 h, 80–90%; (f) RCOOH, EDC, TEA, CH2Cl2, rt, overnight, parallel synthesis.

3740
G. Wágner et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3737–3741
Table 3
Structures, activities and ligand efficiencies of the most potent tetrazoles.
rmGluR5
pKi17
Compound
Y
R
SILE19
LELP20
pIC5018
17a
17b
17c
17d
17e
17f
3-Cl
3-Me
3-Cl
3-Cl
3-Cl
3-Me
3-Cl
2-Piperidinyl
2-Piperidinyl
2-Piperidinyl
2-Piperidinyl
2-Pyrrolidinyl
2-Piperidinyl
2-Pyrrolidinyl
7.31
7.14
7.11
7.06
7.05
6.99
6.98
6.67
6.65
6.47
6.53
6.60
3.78
4.17
3.96
3.94
3.78
4.08
3.74
4.31
2.87
3.42
3.84
4.72
3.28
3.07
17g
17h
3-Cl
2-piperidinyl
6.90
3.70
3.55
17i
17j
17k
17l
3-Cl
3-CN
3-Cl
3-F
2-Piperidinyl
2-Piperidinyl
3-Piperidinyl
2-Piperidinyl
6.86
6.64
6.50
6.34
6.42
3.83
3.70
3.48
3.28
3.65
2.92
4.53
4.43
obtained by deprotecting the basic nitrogen of 15. Acylation of the
secondary amines 16 was accomplished with acids (RCOOH) acti-
vated by EDC in the presence of a base (e.g. TEA) in a parallel syn-
thesis setup. The final tetrazole products 17 were characterized by
LC–MS (Table 1). Purities of the tetrazoles were similar to those of
the oxadiazoles, but yields were somewhat lower.
Before the preparation of the tetrazole library orthogonal com-
binations of 21 amines and 85 acids were synthesized and tested.
Binding affinity data were used selecting the best 24 acids used
for the acylation of the amines during library preparation.
Altogether 629 tetrazoles were obtained; those 619 that ful-
filled the purity criterion were tested. Binding data of the most ac-
tive subset are shown in the Table 3. Similarly to the oxadiazole
series 2-piperidyl derivatives were more active than the 3-piperi-
dyl analogues. With suitable acyl parts, the 2-pyrrolidinyl deriva-
tives could also have acceptable affinity.
In the piperidinyl series the cyclobutylcarbonyl 17b, 17c and
the cyclopentylcarbonyl 17d groups were the best ones, but in
one case 5-bromofurane-2-carbonyl substitution (17a) resulted
in a compound with remarkable affinity. In the pyrrolidinyl series
the most active compound was obtained by the acylation with 5-
methylthiophene-2-carboxylic acid (17e). Substitution at position
3 of the benzene ring was generally tolerated; the best compounds
contained Cl or Me at this position.
Despite of its improved binding affinity ligand efficiency and
lipophilic efficiency of the most active compound (17a) were infe-
rior to that of the HTS hit. Comparative analysis of these metrics re-
vealed that 17b would be a more promising lead having improved
binding affinity and efficiency profile.
tive non-competitive mGluR5 antagonist. Structural modification
of the hit led to new 1,3 disubstituted heterocycles with reason-
able mGluR5 affinities. Two central heterocycles were investi-
gated in detail. During the hit-to-lead process we improved the
binding affinity of the original HTS hit by about 0.4 and 0.6 pKi
unit (2.5-fold and 5-fold) in both the oxadiazole (compound 9a)
and tetrazole (compound 17a) series, respectively. Our results
confirmed that tetrazoles are generally more active than their
oxadiazole analogues.28 Analysing SILE and LELP data we identi-
fied the tetrazole 17b as having improved affinity, ligand effi-
ciency and lipophilic efficiency relative to the original HTS hit.
This lead was considered as a promising starting point for further
optimization.
References and notes
1. (a) Schoepp, D. D.; Conn, P. J. Trends Pharmacol. Sci. 1993, 14, 13; (b) Schoepp, D.
D. Neurochem. Int. 1994, 24, 439; (c) Pin, J. P.; Duvoisin, R. Neuropharmacology
1995, 34, 1; (d) Bordi, F.; Ugolini, A. Prog. Neurobiol. 1999, 59, 55.
2. Alagarsamy, S.; Sorensen, S. D.; Conn, P. J. Curr. Opin. Neurobiol. 2001, 11, 357.
3. Spooren, W. P.; Gasparini, F.; Salt, T. E.; Kuhn, R. Trends Pharmacol. Sci. 2001, 22,
331.
4. Spooren, W. P.; Vassout, A.; Neijt, H. C.; Kuhn, R.; Gasparini, F.; Roux, S.; Porsolt,
R. D.; Gentsch, C. J. Pharmacol. Exp. Ther. 2000, 295, 1267.
5. Walker, K.; Reeve, A.; Bowes, M.; Winter, J.; Wotherspoon, G.; Davis, A.;
Schmid, P.; Gasparini, F.; Kuhn, R.; Urban, L. Neuropharmacology 2001, 40, 10.
6. Tatarczyn´ ska, E.; Klodzin´ ska, A.; Chojnacka-Wójcik, E.; Palucha, A.; Gasparini,
F.; Kuhn, R.; Pilc, A. Br. J. Pharmacol. 2001, 132, 1423.
7. Chapman, A. G.; Nanan, K.; Williams, M.; Meldrum, B. S. Neuropharmacology
2000, 39, 1567.
8. (a) Bruno, V.; Ksiazek, I.; Battaglia, G.; Lukic, S.; Leonhardt, T.; Sauer, D.;
Gasparini, F.; Kuhn, R.; Nicoletti, F.; Flor, P. J. Neuropharmacology 2000, 39,
2223; (b) Movsesyan, V. A.; O’Leary, D. M.; Fan, L.; Bao, W.; Mullins, P. G.;
Knoblach, S. M.; Faden, A. I. J. Pharmacol. Exp. Ther. 2001, 296, 41.
9. Ossowska, K.; Konieczny, J.; Wolfarth, S.; Wieron´ ska, J.; Pilc, A.
Neuropharmacology 2001, 41, 413.
In summary, HTS of our corporate compound library identified
a 3,5-disubstituted-oxadiazole (compound 3) as moderately ac-

G. Wágner et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3737–3741
3741
10. Chiamulera, C.; Epping-Jordan, M. P.; Zocchi, A.; Marcon, C.; Cottiny, C.;
Tacconi, S.; Corsi, M.; Orzi, F.; Conquet, F. Nat. Neurosci. 2001, 4, 873.
11. Allgeier, H.; Auberson, Y.; Biollaz, M.; Cosford, N. D.; Gasparini, F.; Heckendorn,
R.; Johnson, E. C.; Kuhn, R.; Varney, M. A.; Velicelebi, G. WO 9902497, 2007.
12. O’Brien, J. A.; Lemaire, W.; Wittmann, M.; Jacobson, M. A.; Ha, S. N.; Wisnoski,
D. D.; Lindsley, C. W.; Schaffhauser, H. J.; Rowe, B.; Sur, C.; Duggan, M. E.;
Pettibone, D. J.; Conn, P. J.; Williams, D. L., Jr. J. Pharmacol. Exp. Ther. 2004, 309,
568.
in this brain area (Romano, C.; Sesma, M. A.; McDonald, C. T.; O’Malley, K.;
Van den Pol, A. N.; Olney, J.W. J. Comp. Neurol. 1995, 355, 455.). The
neocortical cell cultures were prepared as described in Nagy et al. (Nagy, J.;
Horváth, C.; Farkas, S.; Kolok, S.; Szombathelyi, Z. Neurochem. Int. 2004, 44,
17.) Functional activity at mGluR5 was measured by Ca2+-fluorometry
according to Nagy et al. with modifications. Briefly, cells were isolated from
E17 rat embryos, seeded in 96-well plates and cultured at least for 5 days
before being subjected to Ca2+-measurements. For the Ca2+-measurements
cells were loaded with the Ca2+-sensitive dye, fluo-4/AM. Baseline and agonist
evoked signals were recorded with a plate reader fluorometer. Agonist was
(S)-3,5-dihydroxyphenylglycine. For IC50 determination sigmoidal (4-
parameter) concentration–inhibition curves were fitted to the percent
inhibition data derived from at least three independent experiments using
GraphPad Prism software.
13. Pecknold, J. C.; McClure, D. J.; Appeltauer, L.; Wrzesinski, L.; Allan, T. J. Clin.
Psychopharmacol. 1982, 2, 129.
14. Porter, R. H.; Jaeschke, G.; Spooren, W.; Ballard, T. M.; Büttelmann, B.;
Kolczewski, S.; Peters, J. U.; Prinssen, E.; Wichmann, J.; Vieira, E.; Mühlemann,
A.; Gatti, S.; Mutel, V.; Malherbe, P. J. Pharmacol. Exp. Ther. 2005, 315, 711.
15. Jaeschke, G.; Wettstein, J. G.; Nordquist, R. E.; Spooren, W. Expert Opin. Ther.
Patents 2008, 18, 123.
19. Nissink, J. W. M. J. Chem. Inf. Model. 2009, 49, 1617.
}
}
}
16. (a) Galambos, J.; Vastag, M.; Bobok, A. Á.; Keseru, G. M.; Gál, K.; Benko, B.; Rill,
20. Keseru, G. M.; Makara, G. M. Nat. Rev. Drug Disc. 2009, 8, 203.
21. Gál, K.; Wéber, C.; Wágner, G. A.; Horváth, A.; Nyéki, G.; Vastag, M.; Keseru, G.
}
}
A.; Demeter, Á. WO 2007072089, 2007.; (b) Nógrádi, K.; Wágner, G. A.; Keseru,
G. M.; Bielik, A.; Gáti, T.; Háda, V.; Kóti, J.; Gál, K.; Vastag, M.; Bobok, A. Á. WO
M. WO 2007039781, 2007.
}
2007072090, 2007.; (c) Nógrádi, K.; Keseru, G. M.; Bielik, A.; Gáti, T.; Gál, K.;
22. Poulain, R. F.; Tartar, A. L.; Déprez, D. P. Tetrahedron Lett. 2001, 42, 1495.
23. Gangloff, A. R.; Litvak, J.; Shelton, E. J.; Sperandio, D.; Wang, V. R.; Rice, K. D.
Tetrahedron Lett. 2001, 42, 1441.
}
Vastag, M.; Bobok, A. Á. WO 2007072091, 2007.; (d) Keseru, G. M.; Wéber, C.;
Bielik, A.; Bobok, A. Á.; Gál, K.; Meszlényiné S. M.; Molnár, L.; Vastag, M. WO
}
}
2007072093, 2007.; (e) Nógrádi, K.; Wágner, G. A.; Keseru, G. M.; Bielik, A.;
24. Gál, K.; Wéber, C.; Wágner, G. A.; Bobok, A. Á.; Nyéki, G.; Vastag, M.; Keseru, G.
Gáti, T.; Háda, V.; Kóti, J.; Gál, K.; Vastag, M.; Bobok, A. Á. WO 2007072094,
2007.; (f) Nógrádi, K.; Wágner, G. A.; Keseru, G. M.; Bielik, A.; Gáti, T.; Háda, V.;
Kóti, J.; Gál, K.; Vastag, M.; Bobok, A. Á. WO 2007072095, 2007.
M.; Háda, V.; Kóti, J. WO 2007039782, 2007.
25. Balboni, G.; Marastoni, M.; Merighi, M.; Borea, P. A.; Tomatis, R. Eur. J. Med.
Chem. 2000, 35, 979.
}
17. The mGluR5 receptor binding was determined according to Gasparini et al.
(Gasparini, F.; Andres, H.; Flor, P. J.; Heinrich, M.; Inderbitzin, W.; Lingenhöhl,
K.; Müller, H.; Munk, V. C.; Omilusik, K.; Stierlin, C.; Stoehr, N.; Vranesic, I.;
Kuhn, R. Bioorg. Med. Chem. Lett. 2002, 2, 407.) with modifications. Rat cerebro-
cortical membrane preparation was used to determine the binding
characteristics of reference compounds and novel compounds to the rat
mGluR5. As radioligand [3H]-M-MPEP (2 nM) was used. The nonspecific
26. Chandrasekhar, S.; Reddy, M. V.; Rajaiahet, G. Tetrahedron Lett. 2000, 41, 10131.
27. (a) Ito, S.; Tanaka, Y.; Kakehi, A.; Kondo, K. Bull. Chem. Soc. Jpn. 1976, 49, 1920;
(b) Shawali, A. S.; Fahmi, A. A.; Eweiss, N. F. J. Heterocyclic Chem. 1979, 16, 123.
28. (a) Merck&Co. Inc.: Cosford, N. D.; Roppe, J.; Chen, C.; Smith, N.; Reger, T. WO
2003029210, 2003.; (b) Smith, N. D.; Cosford, N. D. P.; Reger, T. R.; Roppe, J.
R.; Poon, S. F.; Huang, D.; Chen, C.; Eastman, B. W. WO 2003077918;
AstraZeneca AB; (c) Wensbo, D.; Xin, T.; Stefanac, T.; Arora, J.; Edwards, L.;
Isaac, M.; Slassi, A.; Stefanac, T. WO 2004014902, 2004.; (d) Wensbo, D.;
Edwards, L.; Isaac, M.; McLeod, D. A.; Slassi, A.; Xin, T.; Stormann, T. M. WO
2005066155, 2005.; (e) Johansson, M.; Minidis, A.; Staaf, K.; Wensbo, D.;
McLeod, D. A.; Edwards, L.; Isaac, M.; O’Brien, A.; Slassi, A.; Xin, T. WO
2005080356, 2005.; (f) Arora, J.; Edwards, L.; Isaac, M.; Kers, A.; Staaf, K.;
Slassi, A.; Stefanac, T.; Wensbo, D.; Xin, T.; Holm, B. WO 2005080386. 2005.;
(g) Isaac, M.; Slassi, A.; Edwards, L.; Xin, T.; Stefanac, T. WO 2007130821,
2007.; (h) Isaac, M.; Slassi, A.; Edwards, L.; Xin, T.; Stefanac, T. WO
2007130822, 2007; Addex Pharm. SA: (i) Bessis, A.; Bonnet, B.; Le Poul, E.;
Rocher, J.; Epping-Jordan, M. WO 2005044797, 2007.
binding was determined in the presence of 10 lM M-MPEP. The ligand
displacement by the compounds was determined in duplicates or triplicates.
For IC50 (Ki) determinations concentration–displacement curves were
generated consisting of minimum of six concentrations. Ki values (i.e.
inhibition constants) were calculated using the Cheng-Prusoff equation:
Ki = IC50/[1 + (L/Kd)], where [L] is the radioligand concentration and Kd the
affinity of the labelled ligand for receptor. Kd was determined from the
Scatchard plot.
18. Functional activity at mGluR5 was measured in primary rat neuronal cultures
of neocortical origin, taking advantage of the high expression level of mGluR5