Positive and negative modulation of group I metabotropic glutamate receptors

Article abstract of DOI:10.1021/jm0611298

A discriminating pharmacophore model for noncompetitive metabotropic glutamate receptor antagonists of subtype 1 (mGluR1) was developed that facilitated the discovery of moderately active mGluR1 antagonists. One scaffold was selected for the design of sev

Full text of DOI:10.1021/jm0611298

634
J. Med. Chem. 2008, 51, 634–647
Positive and Negative Modulation of Group I Metabotropic Glutamate Receptors
Maksims Vanejevs,† Claudia Jatzke,‡ Steffen Renner,‡ Sibylle Müller,‡ Mirko Hechenberger,‡ Tanja Bauer,‡ Anna Klochkova,§
Ilya Pyatkin,§ Denis Kazyulkin,§ Elena Aksenova,§ Sergey Shulepin,§ Olga Timonina,§ Ariane Haasis,‡ Aleksandrs Gutcaits,†
Christopher G. Parsons,‡ Valerjans Kauss,† and Tanja Weil*,‡
Institute of Organic Synthesis, 21 Aizkraukles Street, Riga, LV1006, LatVia, Merz Pharmaceuticals GmbH, Altenhöferallee 3,
D-60438 Frankfurt am Main, Germany, and Asinex Ltd., 5 GabricheVskogo Street, Building 8, Moscow 125367, Russia
ReceiVed September 28, 2006
A discriminating pharmacophore model for noncompetitive metabotropic glutamate receptor antagonists of
subtype 1 (mGluR1) was developed that facilitated the discovery of moderately active mGluR1 antagonists.
One scaffold was selected for the design of several focused libraries where different substitution patterns
were introduced. This approach facilitated the discovery of potent mGluR1 antagonists, as well as positive
and negative mGluR5 modulators, because both receptor subtypes share similar binding pockets. For mGluR1
antagonists, a homology model of the mGlu1 receptor was established, and a putative binding mode within
the receptor’s transmembrane domain was visualized.
Introduction
studies, the mGluR1 receptor seems to be mainly involved in
therapeutic opportunities such as stroke,42 brain injury,17 and
pain.18 Excessive activation of mGluR5 receptors has been
implicated in a number of central nervous system disorders
including pain,19 anxiety,20 depression,20 other neurological
impairments such as drug addiction,21 and Parkinsons disease.22
G protein-coupled receptors (GPCRs) represent a large protein
superfamily that plays an important role in many physiological
and pathophysiological processes. In the past, the discovery of
molecules acting at GPCRs has been extremely successful with
about 50% of all recently launched drugs targeting GPCRs.1
Metabotropic glutamate receptors (mGluRs),2 together with
parathyroid calcium sensing receptors (CaSR),3 γ-aminobutyric
acid type B receptors (GABABR1 and GABABR2),4,5 some
vomeronasal receptors (V2Rs),6 taste receptors (T1R1–3),7 and
putative pheromone receptors,8 belong to family C GPCRs.
mGluRs mediate excitatory transmission on the cellular surface
through initial binding of glutamate, thus influencing the strength
of synaptic transmission.9,10 The mGluR family comprises eight
subtypes with several splice variants, which have been named
mGluR1–8 according to the succession of the molecular
cloning.11 These eight receptors are further subdivided into three
groups on the basis of their sequence homology, pharmacology,
and transduction mechanism: Group I (mGluR1 and mGluR5),a
group II (mGluR2 and mGluR3), and group III (mGluR4,
mGluR6, mGluR7, and mGluR8).11 A structural characteristic
of mGluRs is their large extracellular domain where the natural
ligand glutamate, as well as competitive agonists and antago-
nists, bind.11,12 The transmembrane domain (TM) bears binding
sites of allosteric modulators, which in general are considered
more selective than competitive ones.13 mGluR1 and mGluR5
are primarily localized postsynaptically.11 They are widely
distributed in many brain regions, including the hippocampus,
cerebellum, thalamic nuclei, and spinal cord.14 Stimulation of
mGluR1 and mGluR5 leads to phosphoinositide (PI) hydrolysis
and elevation of intracellular calcium levels via coupling of GR
q/11-protein to phospholipase C.15,16 According to the previous
Therefore, antagonists of mGluR5 could be of interest for
these indications. In contrast, the selective activation of mGluR5
via positive modulators could normalize hypofunctional activity
mediated Via N-methyl-D-aspartate (NMDA) receptors, and
could therefore be of interest for the development of novel
antipsychotic drug treatments.23,24
In the past, some potent antagonists and positive modulators
of mGluR1 and mGluR5 have been described.25 However, the
discovery of novel mGluR modulators via ligand-based or
structure-based virtual screening techniques is often limited
because of the lack of structurally diverse ligands, as well as
the low sequence identity of mGluRs and bovine rhodopsin,
which was at the time the studies were performed, the only
example of a successfully crystallized GPCR.26 To date, the
discovery of novel mGluR modulators is mainly based on high-
throughput screening (HTS) approaches, which bear inherent
drawbacks such as high cost and a high number of false-positive
or -negative hits.27 Therefore, we first focused on the develop-
ment of a pharmacophore hypothesis for allosteric mGluR1
antagonists because, here, some structurally different ligands
have been reported before.25 Since the TM binding sites of
mGluR1 and mGluR5 modulators reveal a high structural
similarity (“partial overlap”), the discovery of a new mGluR1
allosteric modulator could potentially serve as a reasonable
starting point for the development of novel mGluR5 modula-
tors.28 On the basis of the mGluR1 pharmacophore hypothesis,
a small library was bought and screened for both mGluR1 and
mGluR5 receptor subtypes. This approach allowed the discovery
of weakly active mGluR1 antagonists. Among those, one novel
scaffold was selected and chemically modified leading to highly
potent negative modulators of mGluR1 and both positive and
negative modulators of mGluR5. In addition, the putative
binding mode of such a novel mGluR1 antagonist was visual-
* To whom correspondence should be addressed. E-mail: Tanja.Weil@
merz.de. Fax: +49691503188. Phone: +49691503478.
†
Institute of Organic Synthesis.
Merz Pharmaceuticals GmbH.
Asinex Ltd.
‡
§
aAbbreviations: EL2, extracellular loop 2; EM-TBPC, 1-ethyl-2-methyl-
6-oxo-4-(1,2,4,5-tetrahydro-benzo[d]azepin-3-yl)-1,6-dihydro-pyrimidine-
5-carbonitrile; FLIPR, fluorescent imaging plate reader; HQ, hydroquino-
linone; [3H]MPEP, [3H]-2-methyl-6-(phenylethynyl)-pyridine; IP3, inositol-
1,4,5-trisphosphat or inositoltriphosphate; mGluR1, metabotropic glutamate
receptor 1; mGluR5, metabotropic glutamate receptor; TM, transmembrane.
10.1021/jm0611298 CCC: $40.75
2008 American Chemical Society
Published on Web 01/04/2008

Group I Metabotropic Glutamate Receptors
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 635
10a with respect to the three-dimensional alignment of 1, 2,
and 3. According to the alignment of 10a and the assignment
of all pharmacophore features substituents at position 1 and 2
of the HQ scaffold were considered crucial for achieving
mGluR1 potency. Therefore, a small library was designed where
first R1 was varied in a systematic way (Scheme 1). For R2, the
cyano substituent of 10a was kept, whereas for R3, either
hydrogen or methyl groups were introduced (Tables 1 and 2).
Compounds corresponding to the general formula Ia with a
thioether linker (Scheme 1) were obtained from the 5-oxo-2-
thioxo-1,2,5,6,7,8-hexahydro-quinoline-3-carbonitrile derivatives
8, which in turn were prepared from cyclohexane-1,3-diones 7
according to published procedures.35,36 Alkylation of 8 with
alkyl halides, for example 9, under basic conditions led to
2-alkylsulfanyl-5-oxo-5,6,7,8-tetrahydro-quinoline-3-carboni-
triles (10) of the general formula Ia bearing a thioether linker.
The synthesis of all main products is given in the Experimental
Section. Compound 12 represents a key building block for the
introduction of amino substituents. The reactive chloro group
at position 1 was gained after reaction of 1136,37 with phosphoryl
chloride in moderate yields;38 12a/b then underwent substitution
reactions with a library of appropriately functionalized amino
derivatives in moderate to excellent yields (14, general formula
Ib). All substituents R1 that were introduced via the above-
mentioned procedure are given in Table 1.
Figure 1. 2D-Structures and activity data of published mGluR1
antagonists.
ized, and the herein obtained structure–activity information
(SAR) was interpreted by application of this homology model.
Results and Discussion
Pharmacophore Hypothesis of mGluR1. The first goal was
directed toward the identification of novel mGluR1 antagonists
from commercially available compound libraries. For this
purpose, known mGluR1 antagonists were collected from public
sources, and three structurally different antagonists were chosen
for the development of an initial pharmacophore hypothesis.
Compounds 1,29 2,30 and 331 (Figure 1) represent highly potent
antagonists, and they were characterized in functional assays
by detection of intracellular IP3 changes32 (Experimental
Section). To improve our understanding of the active conforma-
tion of these known antagonists and to assess the number of
essential hydrogen bond acceptors, three derivatives of 3, model
compounds 4, 5, and 6 (Figure 1), were synthesized. These
compounds revealed less potency for mGluR1: 4 was found to
be about 10-fold less active than 3, and 5 was found to be about
10-fold less active than 4 and about 100-fold less active than
3. These first results already pointed toward a qualitatively
greater importance of the N-acceptor group of position 4 versus
position 1 of NPS 2390 (3, Figure 1). A reduction of the size
of the aromatic ring system, for example, from quinoxaline (3)
to pyrazine (6), gave a considerable decrease in activity. On
the basis of these results, a flexible alignment of mGluR1
antagonists 1, 2, and 3 was accomplished by application of the
software MOE (Chemical Computing Group, Figure 2a).
From this alignment, a pharmacophore hypothesis was created
(Figure 2b, protocol is given in the Experimental Section).
Essential groups within this pharmacophore query represent such
groups that are mandatory in the output structure, whereas the
presence of an optional group is not obligatory. Then, a data
set consisting of about 250 000 structurally diverse compounds
from the Asinex Gold Collection33 was virtually screened by
application of this pharmacophore model, and 39 compounds
were purchased and investigated in a functional mGluR1 assay
to detect changes in the IP3 level,32 in addition to being used
in a binding mGluR5 assay by displacement of the known
mGluR5 antagonist [3H]-2-methyl-6-(phenylethynyl)-pyridine34
([3H]MPEP, Experimental Section). Four compounds with an
mGluR1 IC50 below 10 µM were identified, corresponding to a
hit rate of about 10%. Among these hits, 10a represents a
moderately potent mGluR1 antagonist with a hydroquinolinone
(HQ) scaffold whose binding to mGluRs has not been reported
before (Figure 2a). Furthermore, this scaffold offers the ap-
plication of parallel synthesis methods for the straightforward
design of further HQ derivatives.
A synthetic procedure toward HQs carrying no substituents
at position 2 thus corresponding to formula IIa-c is given in
Scheme 2. Quinoline-2,5-diones 15a/b37,39 were reacted with
phosphoryl chloride to give the 2-chloro-substituted quinolin-
5-one derivatives 16a/b, which again offer the introduction of
different substituents. In this way, substitution of the chloro
group with either alcoholates 18 or amino derivatives 13 yielded
17 (formula IIa) and 19 (formula IIb) bearing ether or amino
linkers, respectively. Alternatively, compound 16 was coupled
to ethynyl derivatives 20 in the presence of a palladium(0)
catalyst to give additional derivatives of 21, corresponding to
the general formula IIc. All substituents R1 (where R1 denotes
a combination of substituent R1′, R1″ and a linker group) that
were introduced via the above-mentioned procedure are given
in Table 2. Introduction of a nitro substituent at R2 was achieved
via nitration of 15b with nitric acid at 40 °C to give 22 in
moderate yields. Thereafter, 22 reacted with phosphoryl chloride
leading to 23, which undergoes substitution reactions with
appropriate amino-derivatives 13 to yield a 24a/b in low to
moderate yields. A bromo group was introduced via the same
general procedure by bromination of 15b to give 25, and
subsequent reaction with phosphoryl bromide yielded 26 as
shown in Scheme 3.
The substitution of only one bromo group of 26 proceeded
in low to moderate yields by application of the appropriate
primary or secondary amino derivatives 13 (scheme 3) to give
27a/b. All substituents R1 and R3 introduced via the above-
mentioned procedure are shown in Table 3. A HQ with a
carboxylic acid group at R2, 28, was synthesized starting from
14d via acid hydrolysis in good yield (80%). The amide 29
was prepared via the coupling reaction of 28 by application of
O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium tetrafluo-
roborate (TBTU) as a condensation agent. Compounds 30 and
31 have been synthesized by consecutive substitution of the
bromo substituent with the corresponding amines first in position
R1 with 4-(4-fluoro-phenyl)-1,2,3,6-tetrahydro-pyridine and then
in position R2 by applying morpholine. It is worth noticing that
the bromo substituent of 30 only bears a low reactivity in
comparison with, for example, 25. Our attempts to carry out a
Synthesis of a HQ library. For the sake of clarity, Figure
2a depicts a sketch of the relative orientation of hit compound

636 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
VanejeVs et al.
Figure 2. (a) Flexible alignment of known mGluR1 antagonists 1, 2, and 3 and (b) allocation of their pharmacophore elements (Acceptor )
hydrogen bond acceptor, Hyd ) hydrophobic group, Aro ) aromatic group).
Scheme 1. Synthesis of 3-cyano-substituted 7,8-dihydro-6H-quinolin-5-ones (R3 ) H, methyl, Hal ) Cl, Br, I)
Table 1. Initial HQ Librarya
Discovery of mGluR1 Antagonists. An initial set of HQ
derivatives derived from hit compound 10a was synthesized to
prove that the mGluR1 potency found for 10a was not a
singleton (hit enrichment phase). The library contained ana-
logues of 10a where the linker group of R1 was varied, whereas
for R2, hydrogen and cyano substituents were chosen. Since
selectivity for mGluR1 versus mGluR5 is a key concern in the
reduction of potential side effects,42 all compounds were tested
in mGluR1 and mGluR5 assays (Experimental Section). Table
1 shows that the cyano substituent at R2 was crucial to inhibit
mGluR1 activity. This finding was in agreement with our
pharmacophore hypothesis of mGluR1, where the acceptor2
interaction point was present in most reference compounds. The
best inhibition of mGluR1 activity was obtained for derivatives
10a, 14a, and 14b. Furthermore, the combination of a cyano
group at R2 and a methyl group at R3 was favored. On the other
hand, the presence of hydrogen instead of the cyano substituent
at R2 enhances the potency for mGluR5, as shown for 17a and
a IC50 values were estimated via the simplified logistic equation, where
the hill slope is a constant value that equals unity, and the background and
the range were also constant values of 0 and 100%, respectively. Real DRC
measurements of at least 5 concentrations and 2 independent experiments
19a.
Obviously, the introduction of appropriate substituents at R2
favors selectivity for mGluR1 versus mGluR5. With consider-
were plotted in bold: mGluR1 (functional IP3 assay) and mGluR5 (binding).
ation of the initial results presented in Table 1, a set of
Some of the compounds could not displace [3H]MPEP up to 10 µM and
compounds based on an amino linker of R1 was developed. The
were therefore considered as not active (NA).
amino linker was favored over the thioether or ether linker
because it ensures a higher solubility and the possibility of
Suzuki coupling40,41 at R2 of 30 failed and resulted in debro-
mination. Therefore, the Suzuki coupling was performed via
introducing a broader variety of substituents. Changes in the
nature of substituents R1′ and R1″ considerably influenced the
the intermediate 25. A cyclopentylamine moiety was added after
application of triflouromethanesulfonate and resulted in 33a in
moderate yields. The tetrazole derivative 33b was achieved after
reacting 14d with sodium azide in DMF at elevated temperatures.
potency for mGluR1. Most potent examples include 14d, 14n,
14o, 14q, and 14r with IC50 values around and below 100 nM
(Table 2). In this way, the potency of 14d in comparison to the
initial hit 6 was improved by a factor of about 60.

Group I Metabotropic Glutamate Receptors
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 637
Table 3. Variation of R2 to achieve selective mGluR1 antagonistsa
Table 2. Variation of R1 to Achieve Selective mGluR1 Antagonistsa
a IC50 values were estimated via the simplified logistic equation, where
the hill slope is a constant value that equals unity, and the background and
the range were also constant values of 0 and 100%, respectively: mGluR1
(functional FLIPR assay except 27b) and mGluR5 (binding). Some of the
compounds could not displace [3H]MPEP up to 10 µM and were therefore
considered as not active (NA). b No DRC because of solubility problems.
example, an increase of the ring size from cyclopentyl (14d) to
cyclohexyl (14f), decreased the potency for mGluR1 by a factor
of 50. Apart from cyclopentyl (14d), the introduction of
tetrahydropiperidyl (14q, 14r) and piperazinyl (14n, 14o)
substituents at R1 resulted in potent mGluR1 antagonists (14n,
14o, 14q and 14r). According to Table 2, the presence of
substituents within the phenyl ring leads to ambiguous activity
results: The piperazinyl derivatives 14n, bearing no substituent
within the phenyl ring, and 14o, with a fluoro group, both
significantly antagonized the mGlu1 receptor within the same
order of magnitude, whereas the presence of a methoxyl
substituent in 14p was followed by a considerable decrease of
the potency. In contrast, tetrahydropiperidyl derivative 14r,
carrying a fluoro group, was found to be more potent than the
nonsubstituted analogue 14q. The same trend holds true for the
influence of the methyl substituent at R3 that was also less
obvious: In some cases an up to 5-fold increase of the
antagonistic effect was found (10a vs 10b), whereas in other
cases, no impact on the functional mGluR1 activity was
observed (14 e, 14f).
a IC50 values were estimated via the simplified logistic equation, where
the hill slope is a constant value that equals unity, and the background and
the range were also constant values of 0 and 100%, respectively. Real DRC
measurements of at least 5 concentrations were plotted in bold: mGluR1
(functional IP3 assay) and mGluR5 ([3H]-MPEP binding). Some of the
compounds could not displace [3H]MPEP up tp 10 µM and were therefore
considered as not active (NA).
Scheme 2. Synthesis of 3-Unsubstituted HQs (R3 ) H, methyl,
Hal ) Cl, Br, I)
A broader variation of R2 according to Scheme 3 is listed in
Table 3. The best results were obtained after the introduction
of a bromo (27b) or chloro (27c) substituent. However, here,
the mGluR1 activity was considerably influenced by the
combination of the substituent R1 and R2 (e.g., 27a, 27b).
Overall, among all substituents that were tested and listed in
Table 3, no alternative for the cyano group could be identified.
Unfortunately, the presence of a dimethylamido (29) or a
carboxylic acid group (28), which resulted in a considerable
improvement in solubility, gave a significant reduction of
potency. Heterocyclic groups such as pyrimidino or tetrazol
substituents also decreased or abolished potency for mGluR1.
Discovery of mGluR5 Antagonists and Positive
Allosteric Modulators. All compounds that were prepared
according to Scheme 2 were screened in a mGluR5 binding
assay as mentioned above, as well as in a functional assay
detecting changes in the calcium level (for details see Supporting
Information). Such compounds bearing an ethynyl linker (21,
Obviously, small hydrophobic or hydrophobic aromatic
substituents of a defined geometry at R1 were favored to inhibit
mGluR1 activity. However, slight structural changes, for

638 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
Scheme 3. Introduction of Substituents at Position R2
VanejeVs et al.
IIc) were found to interact with the mGluR5 receptor. This
observation was somewhat expected because derivatives of IIc
show a high structural similarity with the known mGluR5
antagonist MPEP43 (Table 4). In a binding assay, 21a moderately
displaced [3H]MPEP with an affinity of 800 nM. However, in
a functional assay, 21a did not behave like MPEP: instead of
acting as an inhibitor, 21a potentiated the calcium signals.
Functional activation of mGluR5 by orthosteric ligands (agonists
as L-quisqualate or L-glutamate) was measured in fluorescent
imaging plate reader (FLIPR) assays as transient increases of
intracellular calcium signals. In such a FLIPR assay with rat
astrocytes mGluR5 was selectively activated by L-quisqualate.
Here, a considerable and dose-dependent potentiation of a low
calcium signal, elicited by a 10 nM L-quisqualate (open triangle
in Figure 3a), was observed when 21a was preincubated prior
to agonist stimulation. The EC50 of this potentiation was 0.034
( 0.009 µM, and 21a gave no calcium signals (open circles).
When tested on agonist (L-quisqualate) concentration–re-
sponse curves, the addition of a fixed concentration (10 µM) of
21a lead to a 2.0-fold left shift, corresponding to an increase in
agonist potency (EC50) from 24 ( 1.9 to 12 ( 1.7 nM (Figure
3b). This behavior was even more pronounced for the human
mGluR5 receptor, where the native agonist L-glutamate was
applied (Figure 3c). Here, a 3.2-fold left shift was obtained
corresponding to a change of the L-glutamate EC50 from 0.77
( 0.07 to 0.24 ( 0.05 µM.
Because of the high structural similarity of 21a with the
known antagonist MPEP43 (Table 4), we were wondering
whether the positive modulation was achieved via the presence
of the carbonyl group of 21a, enabling 21a to form an additional

Group I Metabotropic Glutamate Receptors
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 639
Table 4. Potencies and Potentiation Properties of Positive Modulators
of mGluR5 Measured via Calcium Signala
a Antagonists of mGluR5 displacing [3H]MPEP in a binding assay.
hydrogen bond. Therefore, several derivatives of 21a were
synthesized according to Scheme 2: the most interesting
examples were listed in Table 4. A positive modulation was
achieved only for those compounds bearing a carbonyl or a
hydroxyl group at the unsaturated ring and two hydrogen
substituents at R3. However, even though the EC50 value of 34,
bearing a hydroxyl group, was considerably higher (0.49 µM),
its maximal change of quisqualate EC50 was found practically
unchanged in comparison to that of 21a. A separation of the
enantiomers of 34, 34a, and 34b was achieved by application
of chiral column chromatography. For both enantiomers, similar
activity and potentiation, compared to that of the racemic
compound 34, were observed.
In case where the carbonyl group was removed (35), a significant
negative modulation was achieved with an IC50 of 500 nM.
Furthermore, the orientation of the carbonyl group seems to be
crucial because the open analogue 36 again showed a moderate
antagonistic effect. Therefore, it seems that the presence of a
hydrogen-bond acceptor at a fixed position facilitates positive
modulation within the transmembrane domain. The etherification
of 34 yielded the methyl ether derivative 37, which was found to
be a potent mGluR5 antagonist with an IC50 value of about 80
nM. Interestingly, even small structural changes, such as the
introduction of two methyl-substituents at R3 (21c), abolished the
ability to induce a positive modulation of mGluR5, and only a
moderately active mGluR5 antagonist was obtained. The exchange
of the phenyl ring of 21a by a pyridyl ring in 21b gave only a
mGluR5 antagonist of weak potency. However, a slight variation
Figure 3. (a) Calcium signals of mGluR5 (rat) in dependence of
concentrations of 21a before (open circles) and after (filled circles)
stimulation with 10 nM L-quisqualate. Ten (open triangle) and 100 nM
(gray triangle) L-quisqualate signals were depicted as controls. (b)
Concentration–response curves of quisqualate without (circles) and with
(squares) after preincubation of 10 µM 21a at mGluR5 (rat). (c)
Concentration–response curves of L-glutamate with (squares) and
without (circles) 10 µM 21a at mGluR5 (human). Averaged from at
least 3 independent experiments that were performed in quadruplicate.
of the position of the dimethyl groups yielded the highly potent
mGluR5 antagonist 21d with an IC50 of 20 nM. These results show
that, in analogy to negative modulators of mGluR1 that have been
described before, minor structural changes could have a strong
impact in the functional profile of mGluR5 and that the interpreta-
tion of the structure–activity relationship is therefore rather chal-
lenginag.44

640 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
VanejeVs et al.
Figure 5. Putative binding mode of 14q inside the allosteric binding
site of mGluR1.
in the Supporting Information. Compound 14q was placed
manually in the transmembrane region in proximity to the pocket
where 11-cis-retinal was found in bovine rhodopsin26 and
cominimized with the receptor. The binding mode of 14q was
selected according to the observed SAR (Figure 5, see text
below). The calculated conformation of 14q in the receptor
reveals an internal energy near the global minimum obtained
from conformational analysis (80.281 vs 80.249 kcal/mol).
According to our homology model, the key interactions of the
pharmacophore for mGluR1 (Figure 2b) were hydrogen-bonding
interactions with residues R661 (3.29) for acceptor 1, T815
(7.39) for acceptor 2, and S668 (3.36) for acceptor 3. The
hydrophobic and aromatic features were placed close to W798
(6.48), V664 (3.32), and L640 (2.58). The carbonyl-oxygen of
the unsaturated ring of HQ potentially interacts as hydrogen-
bond acceptor 1 with a hydrogen-bonding network of R661
(3.29), N747 (45.51), and backbone atoms of EL2. This arginine
(which is conserved in mGluR5) has also been shown to affect
binding of the mGluR5 allosteric negative modulator MPEP
(Table 4).45 N750A (45.54) increased the effect of the mGluR1
negative allosteric modulator EM-TBPC (Figure 1)46 and might
thus be assumed to be in contact with the ligand, as found in
our proposed binding mode for 14q. The cyano group of 14q
(acceptor 2) could interact with T815 (7.39) that has been shown
previously to be involved in the binding of the mGluR1 negative
allosteric modulator EM-TBPC (Figure 1).46 The selectivity for
mGluR1 over mGluR5 for such molecules containing the cyano
group compared with molecules with hydrogen at R2 agrees
well with the model: At position 7.39, T815 was found in
mGluR1, whereas at position 7.39 in mGluR5, methionine was
present in mGluR5. This change from a small hydrophilic
(mGluR1) to a larger and more hydrophobic side chain in
mGluR5 might lead to a steric hindrance and thus repulsion of
the cyano group of these HQs in the mGluR5 binding site. The
third hydrogen-bonding interaction (acceptor 3) could be formed
via the aromatic acceptor nitrogen and S668 (3.36). Previously,
position 3.36 has been shown to be involved in the binding of
MPEP in mGluR5.45 We felt confident to assume a potential
interaction of mGluR1 modulators with residues that were
involved in the binding of mGluR5 modulators because the
binding site of mGluR1 is known to partially overlap with the
TM mGluR5 pocket.28 The two methyl groups of the R3 position
interact with a hydrophobic cluster of TM3 residues V753
(5.43), P756 (5.46), and V757 (5.47).
Figure 4. (a) Essential functionalities for mGluR1 antagonists, (b)
mGluR5 antagonists, and (c) mGluR5 positive modulators based on a
HQ scaffold.
Figure 4 summarizes the essential pharmacophore elements
based on the structure–activity relationship information from
Tables 1-4. At R1, a hydrophobic or hydrophobic/aromatic
substituent with a fixed geometry is one of the main elements
to achieve potency for mGluR1 (Figure 4a). To obtain potent
and subtype-selective mGluR1 antagonists, a small acceptor
group at R2, ideally a cyano group, seems to be essential. Within
the saturated ring close to the carbonyl group, the presence of
hydrophobic substituents is beneficial. To achieve mGluR5
antagonistic activity, an ethynyl spacer at R1, no substituents
at R2, and small, hydrophobic substituents close to the carbonyl
group are required.
In contrast, mGluR5 positive modulators based on a HQ
scaffold are characterized by the absence of functional groups
at the unsaturated HQ ring, in addition to the presence of an
acceptor group (e.g., carbonyl or hydroxyl group) in the HQ
ring, which could form hydrogen bonds with the TM helices.
Visualization of the Binding Cleft. To gain an improved
understanding of the SAR information, ligand 14q was selected
for a further analysis of the binding mode of the HQ scaffold
because of the combination of activity on mGluR1, selectivity
versus mGluR5, the presence of one of the most voluminous
R1 groups, and the structural high similarity with more complex
and potent derivatives (e.g., 19b, 14r). A detailed description
of the construction of a homology model of mGluR1 is given
The preference of nonaromatic over aromatic ring systems
at R1 in close proximity to the HQ scaffold agrees well with a

Group I Metabotropic Glutamate Receptors
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 641
and bis-(ditertbutylphosphine)-palladium(0) was purchased from
Strem Chemicals, Inc. The purity of all compounds was determined
via HPLC column chromatography (Shimadzu LC-20Aprominence;
compound dissolved in acetonitrile/water 50/50 with 0.5% DMSO;
column Phemomenex Gemini 50 × 4.6 mm, 3 µM; mobile phase
acetonitrile/water with 0.1% formic acid; flow 2 mL/min; detection
UV at 310 nm, ambient temperature). The synthesis of several HQ
derivatives has been described in a patent application.47,48
close interaction of this ring with V664 (3.32), as predicted by
the model. V664 might also be involved in the observed
selectivity of 14q for mGluR1. In GluR5, isoleucine was found
at position 3.32, which results in a decrease in the accessible
volume. The presence of the relatively large isoleucine group
might also explain the preference for mGluR5 antagonists
bearing an ethynyl linker at R1, which was found in positive
and negative HQ modulators of mGluR5. The distant aromatic
ring at R1 could interact with a hydrophobic cluster of L640
(2.58), P645 (2.63), V819 (7.43), and potentially, F599 (1.42).
The predicted binding pocket of 14q seems to overlap with
the binding pocket of the mGluR1 negative allosteric inhibitor
EM-TBPC (Figure 1), for which residues influencing the binding
of the ligand have been determined experimentally by mutational
analysis.46 V757 (5.47), W798 (6.48), F801 (6.51), Y805 (6.55),
and T815 (7.39) were found to affect EM-TBPC binding when
mutated to the respective mGluR5 residues. All of these residues
were found close to the putative binding site of 14q. However
14q exceeds the binding pocket of EM-TBPC, and it might form
additional interactions with, for example, TM2 and TM1. The
putative binding mode for 14q is in agreement with our SAR
and published mutational data from other ligands and might
thus provide a further starting point for the design and
optimization of other mGluR1 modulators. In addition, these
investigations might pave the way toward a better understanding
of the ligand–receptor interactions in class C GPCRs.
2-Benzylsulfanyl-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydroquino-
line-3-carbonitrile (10a). To a solution of 7,7-dimethyl-5-oxo-2-
thioxo-1,2,5,6,7,8-hexahydroquinoline-3-carbonitrile (0.5 g, 2.2
mmol) in DMF (4 mL) was added 10% aqueous KOH (1.23 mL),
followed by dropwise addition of benzyl bromide (0.25 mL, 2.2
mmol). The mixture was stirred at room temperature for 12 h; then
water (12 mL) was added. The precipitate formed within 2 h was
filtered off, washed with water, and recrystallized from hexane/
ethyl acetate mixture. A yellow solid was obtained (0.61 g, 86%)
1
with mp ) 129–130 °C. H NMR (CDCl3, TMS): δ 1.13 (s, 6H),
2.54 (s, 2H), 3.05 (s, 2H), 4.55 (s, 2H), 7.3–7.4 (m, 5H), 8.32
ppm (s, 1H). MS: m/z (APCI+) 323 (M + H+). Purity (HPLC):
97.89%.
2-Benzylsulfanyl-5-oxo-5,6,7,8-tetrahydroquinoline-3-carbo-
nitrile (10b). To a solution of 5-oxo-2-thioxo-1,2,5,6,7,8-hexahy-
droquinoline-3-carbonitrile (0.45 g, 2.2 mmol) in DMF (4 mL) was
added 10% aqueous potassium hydroxide (1.23 mL), followed by
dropwise addition of benzyl bromide (0.25 mL, 2.2 mmol). The
mixture was stirred at room temperature for 12 h; then water (12
mL) was added. The product was extracted with diethyl ether. The
extract was washed with water and dried over sodium sulfate.
Filtration and concentration under reduced pressure afforded a
residue that was purified by flash chromatography on silica gel
(petroleum ether/ethyl acetate, 10:1) to give the title compound in
a 91% yield as a colorless solid with mp ) 137–138 °C. 1H NMR
(CDCl3, TMS): δ 2.20 (m, 2H), 2.69 (t, 6 Hz, 2H), 3.16 (t, 6 Hz,
2H), 4.54 (s, 2H), 7.3-7.4 (m, 5H); 8.33 (s, 1H). MS: m/z (APCI+)
295 (M + H+). Purity (HPLC): 97.89%.
Conclusion
We have presented the development of a pharmacophore model
for allosteric mGluR1 antagonists based on published mGluR1
reference ligands. This model was applied successfully to screen
a diverse compound library, and it facilitated the identification of
mGluR1 antagonists with low micromolar affinity with a hit rate
of about 10%. Compound 10a was chosen for a hit optimization
program because of its novelty, as well as the suitability of its
scaffold for a parallel synthesis approach. A focused library of HQ
derivatives led to the discovery of highly potent mGluR1 and
mGluR5 antagonists, as well as potent mGluR5 positive modula-
tors. The presence of both positive and negative modulators of
mGluR1 and mGluR5 based on the same HQ scaffold was mainly
attributed to the fact that these receptors have similar “overlapping”
allosteric binding sites. Furthermore, it turned out that minor
structural changes at distinct positions of the HQ scaffold could
have a considerable impact on their potency and selectivity profile.
Our results show that a novel scaffold interacting within this binding
side could be fine-tuned toward positive and negative modulation
of mGluR1 and mGluR5. The approach described herein could,
in principle, be applied for the identification of other mGluR
modulators where only little or no ligand information is available.
2-Benzylamino-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydroquino-
line-3-carbonitrile (14a). A solution of 7,7-dimethyl-2-methylsul-
fanyl-5-oxo-5,6,7,8-tetrahydroquinoline-3-carbonitrile (0.25 g, 1
mmol), benzylamine (0.13 g, 1.2 mmol), and sodium acetate (0.41
g, 3 mmol) in dry ethanol (3 mL) was stirred while boiling at reflux
for 60 h. The reaction mixture was then evaporated under reduced
pressure. The residue was purified by flash chromatography on silica
gel (chloroform/methanol, 30:1) and treated with dry HCl solution
in diethyl ether to give the title compound (0.072 g, 21%) as a
1
colorless solid with mp ) 163–164 °C. H NMR (CDCl3, TMS):
δ 1.11 (s, 6H), 2.48 (s, 2H), 2.95 (s, 2H), 4.88 (d, 4.2 Hz, 2H),
7.38 (m, 5H), 8.38 (s, 1H). Purity (HPLC): 100%.
2-(Benzyl-methyl-amino)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahy-
droquinoline-3-carbonitrile (14b). Compound 12b (0.220 g, 0.94
mmol), N-methyl-benzylamine (1.70 g, 1.40 mmol), and triethy-
lamine (0.10 g, 1.0 mmol) in ethanol (3 mL) were refluxed for 5 h
(TLC control, chloroform/hexane, 2:1). Then, the mixture was
diluted with water (20 mL) and extracted with dichloromethane (2
× 10 mL). The combined extracts were concentrated, and the
residue was separated by column chromatography on silica gel
(dichloromethane/hexane, 1:1). The subsequent recrystallization
from ethanol resulted in 0.145 g (48%) of pure compound 14b (mp
Experimental Section
(A) Chemistry. (The synthesis of all intermediate compounds
is given in the Supporting Information.)
1
) 97–99 °C). H NMR (DMSO-d6, 400 MHz): δH 1.01 (s, 6H),
2.43 (s, 2H), 2.80 (s, 2H), 3.33 (s, 3H), 5.05 (s, 2H), 7.24–7.38
(m, 5H), 8.21 (s, 1H). MS: m/z (APCI+) 320 (M + H+). Purity
(HPLC): 99.55%.
General Information. The solvents used were of commercial
grade; tetrahydrofuran (THF) was dried over potassium, and toluene
was distilled from sodium. 1H and 13C NMR spectra were recorded
on a Bruker DRX 500 (500 and 125 MHz, respectively), Bruker
AMX 300 (300 and 75 MHz, respectively), or Varian MERCURY
(400 MHz) spectrometer. Melting points were taken on a Sanyo
Gallenkamp melting point apparatus (MPD350.BM3.5) and were
uncorrected. The LC/MS analyses for the compounds were done
at Surveyor MSQ (Thermo Finnigan, U.S.A.) with APCI ionization.
Tetrakis(triphenylphosphine)-palladium(0) (Pd(PPh3)4) and dichlo-
ro[1,1′-bis(diphenylphosphino)ferrocene]palladium(II) dichlorom-
ethane adduct (Pd(dppf)2Cl2 catalysts were purchased from ABCR,
2-(Benzyl-methyl-amino)-5-oxo-5,6,7,8-tetrahydroquinoline-
3-carbonitrile (14c). Compound 12a (0.154 g, 0.75 mmol),
N-methyl-benzylamine (1.82 g, 1.50 mmol), and triethylamine
(0.083 g, 0.82 mmol) in ethanol (3 mL) were refluxed for 5 h (TLC
control, chloroform/hexane, 2:1). Then, the mixture was diluted
with water (10 mL). The precipitate was filtered off and recrystal-
lized from ethanol to give 0.085 g (39%) of compound 14c (mp )
1
103–105 °C). H NMR (DMSO-d6, 400 MHz): δH 1.95–2.08 (s,
2H), 2.48–2.58 (m, 2H), 2.87 (t, J ) 7 Hz, 2H), 3.33 (s, 3H), 5.05

642 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
VanejeVs et al.
(s, 2H), 7.23–7.38 (m, 5H), 8.21 (s, 1H). MS: m/z (APCI+) 292
(M + H+). Purity (HPLC): 98.64%.
formed sediment was filtered off, washed with water, and crystal-
lized with ethanol. As a result, 0.04 g (11%) of compound 14j was
obtained as a white solid (mp ) 266–268 °C). 1H NMR (DMSO-
d6, 400 MHz): δH 1.02 (s, 6H), 1.68 (s, 6H), 2.09 (s, 3H), 2.20 (s,
6H), 2.43 (s, 2H), 2.84 (s, 2H), 6.20 (br. s, 1H), 8.16 (s, 1H).MS:
m/z (APCI+) 350 (M + H+). Purity (HPLC): 97.59%.
7,7-Dimethyl-5-oxo-2-propylamino-5,6,7,8-tetrahydroquino-
line-3-carbonitrile (14k). Compound 14k was synthesized from
12b and propylamine, using the same procedure as described for
14b, as a white solid (mp ) 122–125 °C, yield ) 43%). 1H NMR
(DMSO-d6, 400 MHz): δH 0.88 (t, J ) 7 Hz, 3H), 1.00 (s, 6H),
1.53–1.67 (m, 2H), 2.41 (s, 2H), 2.80 (s, 2H), 3.42 (m, 2H), 7.83
(br. s, 1H), 8.15 (s, 1H).MS: m/z (APCI+) 258 (M + H+). Purity
(HPLC): 98.10%.
2-Butylamino-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydroquinoline-
3-carbonitrile (14l). Compound 14l was synthesized from 12b and
butylamine, using the same procedure described for 14b, as a white
solid (mp ) 107–110 °C, yield ) 41%). 1H NMR (DMSO-d6, 400
MHz): δH 0.92 (t, J ) 7 Hz, 3H), 1.02 (s, 6H), 1.25–1.43 (m, 2H),
1.44–1.65 (m, 2H), 2.41 (s, 2H), 2.80 (s, 2H), 3.46 (m, 2H), 7.88
(t, J ) 7 Hz, 1H), 8.16 (s, 1H). MS: m/z (APCI+) 272 (M + H+).
Purity (HPLC): 98.52%.
2-Hexylamino-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydroquinoline-
3-carbonitrile (14m). Compound 14m was synthesized from 12b
and hexylamine, using the same procedure described for 14b, as a
white solid (mp ) 88–89 °C, yield ) 30%). 1H NMR (DMSO-d6,
400 MHz): δH 0.80–0.93 (m, 3H), 1.00 (s, 6H), 1.20–1.35 (m, 6H),
1.49–1.65 (m, 2H), 2.40 (s, 2H), 2.78 (s, 2H), 3.38–3.52 (m, 2H),
7.78 (br. s, 1H), 8.14 (s, 1H). MS: m/z (APCI+) 300 (M+H+).
Purity (HPLC): 99.97%.
2-Cyclopentylamino-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-
quinoline-3-carbonitrile (14d). Compound 14d was synthesized
from 12b and cyclopentylamine, using the procedure described for
14b, as a white solid (mp ) 154–156 °C, yield ) 80%). 1H NMR
(DMSO-d6, 400 MHz): δH 1.00 (s, 6H), 1.47–1.82 (m, 6H),
1.85–2.05 (m, 2H), 2.43 (s, 2H), 2.80 (s, 2H), 4.43–4.56 (m, 1H),
7.54 (br. s, 1H), 8.14 (s, 1H). MS: m/z (APCI+) 284 (M + H+).
Purity (HPLC): 98.39%.
2-Cyclohexylamino-5-oxo-5,6,7,8-tetrahydroquinoline-3-car-
bonitrile (14e). Compound 14e was synthesized from 12a and
cyclohexylamine, using the procedure described for 14c, as a white
solid (mp ) 143–145 °C, yield ) 59%). 1H NMR (DMSO-d6, 400
MHz): δH 1.00 (s, 6H), 1.00–1.90 (m, 10H), 2.40 (s, 2H), 2.80 (s,
2H), 4.01–4.15 (m, 1H), 7.41 (br. s, 1H), 8.14 (s, 1H). MS: m/z
(APCI+) 270 (M + H+). Purity (HPLC): 99.80%.
2-Cyclohexylamino-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-
quinoline-3-carbonitrile (14f). Compound 12b(0.236 g, 1 mmol),
cyclohexylamine (0.120 g, 1.2 mmol), and triethylamine (0.165 mL,
1.2 mmol) were dissolved in acetonitrile (5 mL). The mixture was
refluxed for 6 h, and the reaction was monitored by TLC (ethyl
acetate/hexane, 1:4). When the reaction was over, the mixture was
diluted with 10 mL of water. The formed sediment was filtered off
and crystallized from ethanol. As a result, 0.170 g (57%) of
compound 14f was obtained (mp ) 164–167 °C). 1H NMR
(DMSO-d6, 400 MHz): δH 1.03–1.20 (m, 2H), 1.20–1.50 (m, 5H),
1.55–1.66 (m, 1H), 1.68–1.78 (m, 2H), 1.78–1.88 (m, 2H),
1.95–2.05 (m, 2H), 2.78–2.95 (m, 2H), 4.02–4.18 (m, 1H), 7.36
(br. s, 1H), 8.16 (s, 1H). MS: m/z (APCI+) 298 (M + H+). Purity
(HPLC): 98.34%.
2-Azepanyl-5-oxo-5,6,7,8-tetrahydroquinoline-3-carboni-
trile (14g). Compound 14g was synthesized from 12a and azepane,
using the procedure described for 14c, as a white solid (mp ) 76–78
°C, yield ) 38%). 1H NMR (DMSO-d6, 400 MHz): δH 1.46–1.58
(m, 4H), 1.74–1.87 (m, 4H), 1.95–2.07 (m, 2H), 2.48–2.58 (m, 2H),
2.82–2.92 (m, 2H), 3.85–3.97 (m, 4H), 8.17 (s, 1H). MS: m/z
(APCI+) 270 (M + H+). Purity (HPLC): 99.60%.
7,7-Dimethyl-5-oxo-2-(4-phenyl-piperazin-1-yl)-5,6,7,8-tet-
rahydroquinoline-3-carbonitrile (14n). Compound 14n was syn-
thesized from 12b and 4-phenylpiperazine, using the procedure
described for 14b, as a white solid (mp ) 167–170 °C, yield )
1
62%). H NMR (DMSO-d6, 400 MHz): δH 1.02 (s, 6H), 2.47 (s,
2H), 2.84 (s, 2H), 3.32 (br. s, 4H), 4.05 (br. s, 4H), 6.80 (t, J ) 7
Hz, 1H), 6.97 (d, J ) 7 Hz, 2H), 7.23 (t, J ) 7 Hz, 2H), 8.28 (s,
1H). MS: m/z (APCI+) 361 (M + H+). Purity (HPLC): 99.18%.
2-[4-(4-Fluoro-phenyl)-piperazin-1-yl]-7,7-dimethyl-5-oxo-
5,6,7,8-tetrahydroquinoline-3-carbonitrile (14o). Compound 12b
(0.236 g, 1 mmol), 4-fluoro-phenyl-piperazine (0.216 g, 1.2 mmol),
and triethylamine (0.165 mL, 1.2 mmol) were dissolved in 5 mL
of acetonitrile. The mixture was refluxed for 6 h, and the reaction
was monitored by TLC (ethyl acetate/hexane, 1:4). When the
reaction was over, the mixture was diluted with water (10 mL).
The formed sediment was filtered off, and recrystallized from
ethanol. As a result, 0.162 g (43%) of compound 14o was obtained
(mp ) 123–125 °C). 1H NMR (DMSO-d6, 400 MHz): δH 1.02 (s,
6H), 2.47 (s, 2H), 2.83 (s, 2H), 3.28 (br. s, 4H), 4.03 (br. s, 4H),
6.94–7.13 (m, 4H), 8.27 (s, 1H).MS: m/z (APCI+) 379 (M + H+).
Purity (HPLC): 99,62%. Anal. Calcd (C22H23FN4O): C, 69.82; H,
6.13; N, 14.8. Found: C, 69.74; H, 6.10; N, 14.51.
2-(Indan-2-ylamino)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-
quinoline-3-carbonitrile (14h). Compound 12b (0.236 g, 1 mmol),
indane-2-yl-amine (0.160 g, 1.2 mmol), and triethylamine (0.165
mL, 1.2 mmol) were dissolved in 5 mL of ethanol. The mixture
was refluxed for 6 h, and the reaction was monitored by TLC (ethyl
acetate/hexane, 1:4). When the reaction was over, the mixture was
diluted with 10 mL of water. The formed sediment was filtered
off, washed with water, and crystallized from ethanol. As a result,
0.175 g (53%) of compound 14h was obtained as a white solid
(mp ) 184–186 °C). 1H NMR (DMSO-d6, 400 MHz): δH 1.03 (s,
6H), 2.43 (s, 2H), 2.82 (s, 2H), 2.96–3.15 (m, 2H), 3.17–3.35 (m,
2H), 4.90–5.05 (m, 1H), 7.14 (br. s, 2H), 7.24 (br. s, 2H), 7.93 (br.
s, 1H), 8.19 (s, 1H).MS: m/z (APCI+) 332 (M + H+). Purity
(HPLC): 99.00%.
2-(3,4-Dihydro-1H-isoquinolin-2-yl)-7,7-dimethyl-5-oxo-5,6,7,8-
tetrahydroquinoline-3-carbonitrile (14i). Compound 12b (0.236
g, 1 mmol), 1,2,3,4-tetrahydroisoquinoline (0.16 g, 1.2 mmol), and
triethylamine (0.165 mL, 1.2 mmol) were dissolved in 5 mL of
acetonitrile. The mixture was refluxed for 6 h, and the reaction
was monitored by TLC (ethyl acetate/hexane, 1:4). When the
reaction was over, the mixture was diluted with water (10 mL).
The formed sediment was filtered off and crystallized from ethanol.
As a result, 0.200 g (60%) of compound 14i was obtained (mp )
2-[4-(4-Methoxy-phenyl)-piperazin-1-yl]-7,7-dimethyl-5-oxo-
5,6,7,8-tetrahydroquinoline-3-carbonitrile (14p). Compound 12b
(0.236 g, 1 mmol), 4-methoxy-phenyl-piperazine (0.23 g, 1.2
mmol), and triethylamine (0.165 mL, 1.2 mmol) were dissolved in
acetonitrile (5 mL). The mixture was refluxed for 6 h, and the
reaction was monitored by TLC (ethyl acetate/hexane, 1:4). When
the reaction was over, the mixture was diluted with water (10 mL).
The formed sediment was filtered off and recrystallized from
ethanol. As a result, 0.300 g (76%) of compound 14p was obtained
(mp ) 158–160 °C). 1H NMR (DMSO-d6, 400 MHz): δH 1.02 (s,
6H), 2.47 (s, 2H), 2.84 (s, 2H), 3.28 (br. s, 4H), 3.70 (s, 3H), 4.03
(br. s, 4H), 6.85 (d, J ) 7 Hz, 2H), 6.95 (d, J ) 7 Hz, 2H), 8.27
(s, 1H).MS: m/z (APCI+) 391 (M+H+). Purity (HPLC): 98.30%.
7,7-Dimethyl-5-oxo-2-(4-phenyl-3,6-dihydro-2H-pyridin-1-yl)-
5,6,7,8-tetrahydroquinoline-3-carbonitrile (14q). Compound 12b
(0.236 g, 1 mmol), 4-phenyl-1,2,3,6-tetrahydropyridine hydrochlo-
ride (0.235 g, 1.2 mmol), and triethylamine (0.165 mL, 1.2 mmol)
were dissolved in acetonitrile (5 mL). The mixture was refluxed
for 6 h, and the reaction was monitored by TLC (ethyl acetate -
1
132–134 °C). H NMR (DMSO-d6, 400 MHz): δH 1.02 (s, 6H),
2.46 (s, 2H), 2.87 (s, 2H), 3.03 (t, J ) 7 Hz, 2H), 4.08 (t, J ) 7
Hz, 2H), 4.7 (s, 2H), 7.17–7.29 (br. s, 4H), 8.28 (s, 1H). MS: m/z
(APCI+) 332 (M + H+).
2-(Adamantan-1-ylamino)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahy-
droquinoline-3-carbonitrile (14j). Compound 12b (0.236 g, 1
mmol), adamantanylamine (0.450 g, 2 mmol), and triethylamine
(0.28 mL, 2 mmol) were dissolved in 5 mL of ethanol, and the
mixture was refluxed for 24 h. The reaction was monitored by TLC
(ethyl acetate/hexane, 1:4). Then, the mixture was cooled. The

Group I Metabotropic Glutamate Receptors
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 643
hexane, 1:4). When the reaction was over, the mixture was diluted
with 10 mL of water. The formed sediment was filtered off and
recrystallized from ethanol. As a result, 0.106 g (30%) of compound
2H), 6.97 (d, J ) 8 Hz, 2H), 7.23 (t, J ) 8 Hz, 2H), 7.90 (d, J )
8 Hz, 1H). MS: m/z (APCI+) 308 (M + H+). Purity (HPLC):
99.68%.
1
14q was obtained (mp ) 131–133 °C). H NMR (DMSO-d6, 400
2-Phenylethynyl-7,8-dihydro-6H-quinolin-5-one (21a). Com-
pound 16a (0.2 g, 1.1 mmol) was dissolved in triethylamine (5 g,
49.5 mmol), and then ethynylbenzene (0.17 g, 1.6 mmol), Pd[PPh3]4
(0.02 g, 0.062 mmol), and CuI (21 mg, 0.11 mmol) were added
under an argon atmosphere. The mixture was stirred for 3 h at 100
°C under an argon atmosphere. Compound 21a was isolated by
column chromatography on silica gel (hexane/ethyl acetate, 50:1)
and was obtained as colorless crystals (0.04 g, 15%) with mp )
121.1–121.7 °C. 1H NMR (DMSO-d6, 400 MHz): δH 2.13 (t, J )
7 Hz, 2H), 2.60–2.76 (m, 2H), 3.10 (t, J ) 7 Hz, 2H), 7.43–7.55
(m, 3H), 7.59–7.68 (m, 3H), 8.19 (d, J ) 8 Hz, 1H). MS: m/z
(APCI+) 248 (M + H+). Purity (HPLC): 99.79%. Anal. Calcd
(C17H13NO): C, 82.57; H, 5.30; N, 5.66. Found: C, 82.34; H, 5.24;
N, 5.53.
MHz): δH 1.02 (s, 6H), 2.47 (s, 2H), 2.63–2.77 (m, 2H), 2.85 (s,
2H), 4.12 (t, J ) 7 Hz, 2H), 4.51 (br. s, 2H), 6.38 (br. s, 1H),
7.22–7.42 (m, 3H), 7.44–7.54 (m, 2H), 8.27 (s, 1H).MS: m/z
(APCI+) 358 (M + H+). Purity (HPLC): 96.17%.
2-[4-(4-Fluoro-phenyl)-3,6-dihydro-2H-pyridin-1-yl]-7,7-di-
methyl-5-oxo-5,6,7,8-tetrahydroquinoline-3-carbonitrile (14r).
Compound 12b (0.236 g, 1 mmol), 4-(4-fluorophenyl)-1,2,3,6-
tetrahydropyridine (0.212 g, 1.2 mmol), and triethylamine (0.165
mL, 1.2 mmol) were dissolved in 5 mL of ethanol. The mixture
was refluxed for 6 h, and the reaction was monitored by TLC (ethyl
acetate/hexane, 1:4). When the reaction was over, the mixture was
diluted with 20 mL of water and extracted with dichloromethane
(3 × 10 mL). The resulting solution was filtered through a small
layer of silica gel and concentrated. The residue was crystallized
from ethanol. As a result, 0.155 g (41%) of compound 14r was
2-Pyridin-2-ylethynyl-7,8-dihydro-6H-quinolin-5-one (21b).
Compound 16a (0.3 g, 1.7 mmol) was dissolved in triethylamine
(10 mL). Then, 2-ethynylpyridine (0.26 g, 2.5 mmol), Pd[PPh3]4
(0.095 g, 0.082 mmol), and CuI (32 mg, 0.17 mmol) were added
under an argon atmosphere, and the mixture was stirred for 5 h at
100 °C under an argon atmosphere. The residue was purified by
column chromatography on silica gel (hexane/ethyl acetate, 1:2).
As a result, 0.1 g (22%) of crystalline white compound 21b was
obtained with mp ) 150–151 °C. 1H NMR (DMSO-d6, 400 MHz):
δH 2.13 (t, J ) 7 Hz, 2H), 2.60–2.76 (m, 2H), 3.10 (t, J ) 7 Hz,
2H), 7.48–7.55 (m, 1H), 7.67 (d, J ) 8 Hz, 1H), 8.08 (d, J ) 8
Hz, 1H), 8.22 (d, J ) 8 Hz, 1H), 8.66 (d, J ) 4 Hz, 1H), 8.84 (s,
1H). MS: m/z (APCI+) 249 (M + H+). Purity (HPLC): 99.58%.
7,7-Dimethyl-2-pyridin-2-ylethynyl-7,8-dihydro-6H-quinolin-
5-one (21c). Compound 16b (0.3 g, 1.4 mmol) was dissolved in
triethylamine (10 mL). Then, 3- ethynylpyridine (0.22 g, 2.1 mmol),
Pd[PPh3]4 (0.023 g, 0.071 mmol), and CuI (26 mg, 0.14 mmol)
were added under an argon atmosphere. The mixture was stirred
for 3 h at 100 °C under an argon atmosphere. Then, the solvent
was evaporated, and the residue was purified by column chroma-
tography on silica gel (hexane/ethyl acetate, 1:1). As a result, 0.05 g
(13%) of crystalline 21c was obtained with mp ) 108–109 °C. 1H
NMR (DMSO-d6, 400 MHz): δH 1.08 (s, 6H), 2.62 (s, 2H), 3.06
(s, 2H), 7.54 (m, 1H), 7.71 (d, J ) 10 Hz, 1H), 8.10 (d, J ) 10
Hz, 1H), 8.24 (d, J ) 10 Hz, 1H), 8.68 (d, J ) 6 Hz, 1H), 8.87 (s,
1H). MS: m/z (APCI+) 277 (M + H+). Purity (HPLC): 99.20%.
6,6-Dimethyl-2-phenylethynyl-7,8-dihydro-6H-quinolin-5-
one (21d). Sodium hydride (0.048 g, 1.2 mmol, 1.5 equiv) was
added in an argon atmosphere to a solution of 2-phenylethynyl-
7,8-dihydro-6H-quinolin-5-one (21a) (0.2 g, 0.8 mmol, 1 equiv)
in dry THF (5 mL), and the mixture was stirred for 30 min at room
temperature. Then iodomethane (0.16 g, 1.2 mmol, 1.5 equiv) was
added to the reaction mixture, and stirring was continued for 3 h
at room temperature. Then the reaction mixture was evaporated,
and the residue was subjected to column chromatography on silica
gel (60–100 µ, l ) 20 cm, d ) 2 cm, hexane/ethyl acetate 3:1). As
a result, 0.033 g (13%) of 21d was obtained. 1H NMR (400 MHz,
CDCl3): δ 1.25 (6H, s), 2.06 (2H, t), 3.20 (2H, t), 7.33–7.43 (3H,
m), 7.49 (1H, d), 7.63 (2H, d), 8.29 (1H, d). MS: m/z (CI MS) 276
(M + H+). Purity (HPLC): 95.83%.
1
obtained (mp 108–110 °C). H NMR (DMSO-d6, 400 MHz): δH
1.02 (s, 6H), 2.45 (s, 2H), 2.63–2.75 (m, 2H), 2.84 (s, 2H), 4.12 (t,
J ) 7 Hz, 2H), 4.51 (br. s, 2H), 6.25 (br. s, 1H), 7.17 (t, J ) 6 Hz,
2H), 7.52 (t, J ) 6 Hz, 2H), 8.26 (s, 1H).MS: m/z (APCI+) 376
(M + H+). Purity (HPLC): 95.46%. Anal. Calcd (C23H22FN3O):
C, 73.58; H, 5.91; N, 11.19. Found: C, 73.06; H, 5.83; N, 11.11.
2-Benzyloxy-7,8-dihydro-6H-quinolin-5-one (17a). The com-
pound was prepared according to ref 49. Colorless solid. mp: 80–81
°C. MS: m/z (APCI+) 254 (M + H+).1H NMR (CDCl3, TMS): δ
2.16 (m, 2H), 2.63 (t, 6.4 Hz, 2H), 3.03 (t, 6.4 Hz, 2H), 5.44 (s,
2H), 6.70 (d, 8.9 Hz, 1H), 7.3 – 7.5 (m, 5H), 8.17 (d, 8.9 Hz, 1H).
2-Benzyloxy-7,7-dimethyl-7,8-dihydro-6H-quinolin-5-one hy-
drochloride (17b). To a solution of benzyl alcohol (0.162 g, 1.5
mmol) in diethyl ether (10 mL) was added sodium (0.035 g, 1.5
mmol), and it was stirred at room temperature for 2.5 h. Then
2-chloro-7,7-dimethyl-7,8-dihydro-6H-quinolin-5-one (0.21 g, 1.0
mmol) was added, and the resulting mixture was stirred at 30 °C
for 24 h. Water (12 mL) was added and the mixture was extracted
with ethyl acetate (2 × 10 mL). The organic phase was washed
with water (10 mL) and dried over magnesium sulfate; then it was
filtered and evaporated under reduced pressure. The residue was
purified by flash chromatography on silica gel (petroleum ether/
ethyl acetate, 10:1), followed by treatment with 0.5 M dry HCl
solution in diethyl ether to give the title compound in a 33% yield.
MS: m/z (APCI+) 282 (M + H+). mpL 87–88 °C. 1H NMR(CDCl3,
TMS): δ 1.15 (s, 6H), 2.55 (s, 2H), 3.50 (s, 2H), 5.70 (s, 2H), 7.07
(d, 8.8 Hz, 1H), 7.30–7.60 (m, 5H), 8.58 (d, 8.8 Hz, 1H).
2-Benzylamino-7,7-dimethyl-7,8-dihydro-6H-quinolin-5-one
(19a). 2-Chloro-7,7-dimethyl-7,8-dihydro-6H-quinolin-5-one (0.315
g, 1.5 mmol) and potassium carbonate (0.83 g, 6 mmol) were added
to a solution of benzylamine (0.2 g, 1.8 mmol) in dry acetonitrile
(4 mL). The mixture was stirred while boiling under reflux for 48 h.
Then DMSO (3 mL) and sodium hydride (0.05 g) were added, and
heating was continued for an additional 5 h. Water (10 mL) was
added, and the mixture was extracted with chloroform (2 × 10
mL). The extract was dried over Na2SO4, filtered, and evaporated
under reduced pressure. The residue was purified by flash chro-
matography on silica gel (chloroform/methyl alcohol, 40:1); then
it was treated with dry HCl in diethyl ether to give the title
compound (0.1 g, 21%) as a colorless solid with mp ) 215–216
°C. MS: m/z (APCI+) 281 (M + H+). 1H NMR (CDCl3, TMS): δ
1.14 (s, 6H), 2.48 (s, 2H), 3.08 (s, 2H), 4.60 (d, 6.8 Hz, 2H), 6.64
(d, 9.4 Hz, 1H), 7.3–7.4 (m, 5H), 8.23 (d, 9.4 Hz, 1H), 9.67 (br s,
1H).
7,7-Dimethyl-3-nitro-2-(4-phenyl-3,6-dihydro-2H-pyridin-1-
yl)-7,8-dihydro-6H-quinolin-5-one (24b). To the solution of the
compound 23 (0.12 g, 0.47 mmol) in acetonitrile (10 mL),
diisopropylethylamine (0.15 g, 1.1 mmol) and 4-phenyl-1,2,3,6-
tetrahydro-pyridine hydrochloride (0.138 g, 0.7 mmol) were added.
The mixture was refluxed for 3 h. The solvent was evaporated.
The residue was purified by column chromatography on silica gel
(hexane/ethyl acetate, 4:1). As a result, compound 24 was obtained
2-(4-Phenyl-piperazin-1-yl)-7,8-dihydro-6H-quinolin-5-one
(19b). A mixture of compound 16a (0.31 g, 1.7 mmol) and
N-phenylpiperazine (5.5 g, 34 mmol) was stirred at 100 °C for 1 h
(TLC control). The reaction mixture was worked out by column
chromatography on silica gel (hexane/ethyl acetate, 3:1) to give
0.220 g (41%) of compound 19b as colorless crystals (mp )
157.1–158.0 °C). 1H NMR (DMSO-d6, 400 MHz): δH 2.01 (t, J )
7 Hz, 2H), 2.86 (t, J ) 7 Hz, 2H), 3.83 (s, 4H), 6.75–6.85 (m,
1
(0.06 g, 34%) as an oil. H NMR (CDCl3, TMS, 400 MHz): δH
1.25 (s, 3H), 1.28 (s, 3H), 2.70 (s, 2H), 2.90 (d, J ) 20 Hz, 1H),
3.10 (d, J ) 20 Hz, 1H), 4.08 (s, 2H), 4.35 (s, 2H), 5.19 (s, 1H),
6.19 (br. s, 1H), 6.62 (d, J ) 11 Hz, 1H), 7.20–7.55 (m, 5H), 8.09
(d, J ) 11 Hz, 1H). MS: m/z (APCI+) 378 (M + H+).

644 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
VanejeVs et al.
3-Bromo-2-cyclopentylamino-7,7-dimethyl-7,8-dihydro-6H-
quinolin-5-one (27a). Compound 26 (0.15 g, 0.45 mmol) and
cyclopentylamine (0.38 g, 4.5 mmol) were fused at 80 °C for 2 h.
Purification of the cake by column chromatography on silica gel
(hexane/ethyl acetate, 2:1) gave 0.053 g (34%) of light yellow
crystalline compound 27a (mp ) 176–177 °C). 1H NMR (DMSO-
d6, 400 MHz): δH 1.20 (s, 3H), 1.27 (s, 3H), 1.47–1.85 (m, 6H),
2.03–2.15 (m, 2H), 2.61 (d, J ) 18 Hz, 1H), 3.12 (d, J ) 18 Hz,
1H), 4.04–4.16 (m, 1H), 4.29 (s, 1H), 5.10–5.23 (m, 1H), 6.36 (d,
J ) 10 Hz, 1H), 8.07 (d, J ) 10 Hz, 1H). MS: m/z (APCI+) 337
(M + H+). Purity (HPLC): 96.19%.
3-Bromo-2-[4-(4-fluoro-phenyl)-piperazin-1-yl]-7,7-dimethyl-
7,8-dihydro-6H-quinolin-5-one (27b). Compound 26 (0.1 g, 0.3
mmol) and 1-(4-fluoro-phenyl)-piperazine (0.5 g, 3 mmol) were
fused at 80 °C for 1 h. Purification of the residue by column
chromatography on silica gel (hexane/ethyl acetate, 3:2 v/v) gave
0.02 g (16%) of compound 27b as a colorless oil. 1H NMR (DMSO-
d6, 400 MHz): δH 1.11 (s, 6H), 2.45 (s, 2H), 2.82 (s, 2H), 3.36 (br.
s, 4H), 3.74 (br. s, 4H), 6.86–7.05 (m, 4H), 8.29 (s, 1H). MS: m/z
(APCI+) 432 (M + H+).
3-Chloro-2-cyclopentylamino-7,7-dimethyl-7,8-dihydro-6H-
quinolin-5-one (27c). (a) Synthesis of 3-Chloro-7,7-dimethyl-
7,8-dihydro-1H,6H-quinoline-2,5-dione. N-chlorosuccinimide (0.8
g, 5.5 mmol) was added to a solution of 7,7-dimethyl-7,8-dihydro-
1H,6H-quinoline-2,5-dione (1.0 g, 5.0 mmol) in 1,2-dichloroethane
(50 mL). The reaction mixture was refluxed for 8 h. Then the
solvent was removed on a rotary evaporator. The residue was
crystallized with ethanol (5 mL). The colorless crystals formed were
filtered off and dried on air to give 0.3 g (25%) of the title
compound. MS: m/z (APCI+) 226 (M + H+).
(b) Synthesis of 2-Bromo-3-chloro-7,7-dimethyl-7,8-dihydro-
6H-quinolin-5-one. POBr3 (0.46 g, 1.6 mmol) was added to a
solution of 3-chloro-7,7-dimethyl-7,8-dihydro-1H,6H-quinoline-2,5-
dione (0.3 g, 1.3 mmol) in acetonitrile (25 mL). The reaction
mixture was refluxed for 5 h. Then the solvent was evaporated.
The residue was dissolved in dichloromethane (20 mL) and washed
with 10% aqueous potassium carbonate and water. The organic layer
was separated, dried over sodium sulfate, and concentrated. The
residue was purified by column chromatography on silica gel
(hexane/ethyl acetate, 1:1) to give 0.06 g (16%) of the title
compound as a light yellow oil. MS: m/z (APCI+) 289 (M+H+).
(c) Synthesis of 3-Chloro-2-cyclopentylamino-7,7-dimethyl-
7,8-dihydro-6H-quinolin-5-one (27c). A mixture of 2-bromo-3-
chloro-7,7-dimethyl-7,8-dihydro-6H-quinolin-5-one(0.06g,0.21mmol)
and cyclopentylamine (0.18 g, 2.1 mmol) was fused at 80 °C for
2 h. Purification of the cake by column chromatography on silica
gel (hexane/ethyl acetate, 2:1) resulted in 0.032 g (45%) of light
yellow crystalline 27c (mp ) 80–81 °C). 1H NMR (CDCl3, TMS,
400 MHz): δH 1.08 (s, 6H), 2.40 (s, 2H), 2.78 (s, 2H), 4.48 (m,
1H), 5.42 (br. s, 1H), 7.98 (s, 1H). MS: m/z (APCI+) 293 (M +
H+). Purity (HPLC): 95.91%.
2-Cyclopentylamino-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-
quinoline-3-carboxylic acid (28). A solution of compound 14d
(0.283 g, 1.0 mmol) in a mixture of HCl/AcOH/H2SO4/H2O (1/1/
1/1 v/v; 20 mL) was refluxed for 12 h (TLC control; ethyl acetate/
hexane, 1:2). Then, mixture was cooled and carefully basified with
aqueous NH3 to pH 7–8. The formed precipitate was filtered off,
washed with water, and dried in air to give 0.21 g (70%) of
compound 28 as white solid (mp ) 196–197 °C). 1H NMR (DMSO-
d6, 400 MHz): δH 1.00 (s, 6H), 1.35–1.76 (m, 6H), 1.92–2.06 (m,
2H), 2.32 (s, 2H), 2.73 (s, 2H), 4.35–4.51 (m, 1H), 8.39 (s, 1H),
9.64 (br. s, 1H). MS: m/z (APCI+) 303 (M + H+). Purity (HPLC):
99.61%.
2-Cyclopentylamino-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-
quinoline-3-carboxylic acid dimethylamide (29). TBTU (0.13 g,
0.4 mmol), dimethylamine hydrochloride (0.03 g, 0.35 mmol), and
triethylamine (0.2 mL) were added to a solution of compound 28
(0.1 g, 0.3 mmol) in acetonitrile (4 mL). The mixture was stirred
at room temperature for 2 h (TLC control; ethyl acetate/hexane,
1:2). Then, the mixture was diluted with water (15 mL) and
extracted with dichloromethane (3 × 10 mL). The extract was dried
over Na2SO4 and evaporated. The residue was purified by column
chromatography on silica gel (ethyl acetate/hexane, 1:5) to give
0.052 g (50%) of compound 29 as a white solid (mp ) 133–134
1
°C). H NMR (DMSO-d6, 400 MHz): δH 1.02 (s, 6H), 1.41–1.76
(m, 6H), 1.89–2.06 (m, 2H), 2.38 (s, 2H), 2.77 (s, 2H), 2.94 (s,
6H), 4.37–4.51 (m, 1H), 6.87 (d, J ) 11 Hz, 1H), 7.71 (s, 1H).
MS: m/z (APCI+) 330 (M + H+). Purity (HPLC): 99.60%.
2-[4-(4-Fluoro-phenyl)-3,6-dihydro-2H-pyridin-1-yl]-7,7-di-
methyl-3-morpholin-4-yl-7,8-dihydro-6H-quinolin-5-one (31). A
mixture of compound 30 (0.15 g, 0.35 mmol), sodium tert-pentoxide
(0.093 g, 0.85 mmol), morpholine (0.061 g, 0.7 mmol), bis(diben-
zylideneacetone)palladium (0.03 g, 0.052 mmol), and 2-(di-t-
butylphosphino)biphenyl (0.016 g, 0.053 mmol) in tetrahydrofuran
(5 mL) was stirred under an argon atmosphere at 66 °C for 8 h.
Then, the reaction mixture was concentrated under reduced pressure.
The residue was purified by column chromatography on silica gel
(hexane/ethyl acetate, 5:1). As a result, 0.026 g (17%) of compound
31 was isolated as light yellow oil. 1H NMR (DMSO-d6, 400 MHz):
δH 1.10 (s, 6H), 2.42 (s, 2H), 2.65 (s, 2H), 2.81 (s, 2H), 3.03 (s,
4H), 3.86 (s, 4H), 4.02 (t, J ) 7 Hz, 1H), 4.40 (s, 1H), 6.15 (s,
1H), 7.03 (t, J ) 12 Hz, 2H), 7.38 (t, J ) 12 Hz, 2H), 7.67
(s, 1H). MS: m/z (APCI+) 436 (M + H+).
2-Cyclopentylamino-7,7-dimethyl-3-pyrimidin-5-yl-7,8-dihy-
dro-6H-quinolin-5-one (33a). To a solution of compound 32 (0.22
g, 0.8 mmol), pyridine (0.135 g, 1.7 mmol), and diisopropylethy-
lamine (1.18 g, 1.4 mmol) in dichloromethane (10 mL) trifluo-
romethanesulfonic anhydride (0.38 g, 1.4 mmol) was added. The
reaction mixture was stirred at room temperature for 3 h; then, the
solvent was evaporated. Purification of the residue by column
chromatography on silica gel (hexane/ethyl acetate, 3:1) gave 0.25 g
(80%) of the product as a dark oil. A mixture of this oil (0.25 g,
0.64 mmol), cyclopentylamine (0.38 g, 4.5 mmol) and diisopro-
pylethylamine (0.12 g, 0.9 mmol) in acetonitrile (10 mL) was
refluxed for 4 h. Then the solvent was removed in vacuo, and the
residue was purified by HPLC to give colorless crystalline
compound 33 (0.32 g, 17%, mp ) 191–193 °C). 1H NMR (DMSO-
d6, 400 MHz): δH 1.12 (s, 6H), 1.27–1.41 (m, 2H), 1.57–1.73 (m,
4H), 2.03–2.15 (m, 2H), 2.42 (s, 2H), 2.87 (s, 2H), 4.45–4.55 (m,
1H), 4.71 (br. s, 1H), 7.26 (s, 1H), 7.83 (s, 1H), 8.78 (s, 2H), 9.23
(s, 1H). MS: m/z (APCI+) 337 (M + H+). Purity (HPLC): 100%.
2-Cyclopentylamino-7,7-dimethyl-3-(2H-tetrazol-5-yl)-7,8-di-
hydro-6H-quinolin-5-one (33b). To a solution of 14d (0.566 g, 2
mmol) in dry DMF (5 mL), sodium azide (0.260 g, 4 mmol) and
ammonium chloride (0.214 g, 4 mmol) were added. The mixture
was refluxed for 3 h (TLC control, ethyl acetate/hexane, 1:2). Then,
the mixture was diluted with water (20 mL) and acidified with acetic
acid to pH 5. The formed precipitate was filtered off and crystallized
with ethanol to give 0.45 g (69%) of 34 (mp 269–271 °C). MS:
m/z (APCI+) 327 (M + H+). Purity (HPLC): 99.47%.
2-Phenylethynyl-5,6,7,8-tetrahydro-quinolin-5-ol (34). To a
suspension of the compound 27a (0.12 g, 0.48 mmol) in isopropyl
alcohol (25 mL), NaBH4 (0.2 g, 5.3 mmol) was added, and the
mixture was stirred for 5 h at RT (room temperature). Then, the
solvent was evaporated under reduced pressure. The residue was
dissolved in chloroform (50 mL), washed with water, and dried
over Na2SO4. The chloroform was evaporated. As a result of HPLC
separation, 0.12 g (99%) of compound 34 was obtained as an oil.
1H NMR (DMSO-d6, 400 MHz): δH 1.65–1.87 (m, 2H), 1.95–2.07
(m, 2H), 2.80–2.95 (m, 2H), 4.69 (br. s, 1H), 7.45–7.53 (m, 3H),
7.56 (d, J ) 8 Hz, 1H), 7.60–7.68 (m, 3H), 7.93 (d, J ) 8 Hz,
1H). MS: m/z (APCI+) 250 (M + H+). Anal. Calcd (C17H15NO)
C, 81.90; H, 6.06; N, 5.62. Found: C, 81.60; H, 5.93; N, 5.41.
(S)-2-Phenylethynyl-5,6,7,8-tetrahydro-quinolin-5-ol (34a) and
(R)-2-Phenylethynyl-5,6,7,8-tetrahydro-quinolin-5-ol (34b). Ra-
cemic mixture of isomers of 2-phenylethynyl-5,6,7,8-tetrahydro-
quinolin-5-ol (34) (0.15 g, 0.6 mmol) was separated by HPLC
(column Chiralpak AD-H 4.6 × 250 mm 5 mkm; mobile phase
hexane/IPA 90/10; flow1.0 mL/min; detection UV at 240 nm;
ambient temperature) to give 0.064 g (43%) of (S)-2-phenylethynyl-
5,6,7,8-tetrahydro-quinolin-5-ol (34a) (e.e. > 99.5%) and 0.069 g
(46%) of (R)-2-phenylethynyl-5,6,7,8-tetrahydro-quinolin-5-ol (34b)

Group I Metabotropic Glutamate Receptors
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 645
1
(e.e. > 99.5%) as oils. H NMR (400 MHz, CDCl3): δ 1.78–1.98
in a stable CHO cell line under control of an inducible promoter
(LacSwitchII, Stratagene). In this case, the Ca-Kit contained
glutamic-pyruvate transferase (GPT), Na-pyruvate, and pyridoxal
phosphate to avoid accumulation of glutamate during Ca-dye
loading. Positive allosteric modulators were characterized in two
ways. First, the EC50 of the potentiation was tested by preincubation
of concentration range of the modulator and subsequent stimulation
with a fixed, low concentration of agonist (∼EC20). Second, the
increase in agonist potency by a positive modulator was determined
by running two agonist curves in parallel: one without and one
with a preincubated modulator. This potency increase was expressed
as a left shift and determined as the ratio of the EC50 values.
mGluR1 Assay: Accumulation of [3H]-Inositol Phosphates.
After cerebellar granule cells were cultured for 7 days, BEM was
removed, and inositol-free DMEM (ICN) supplemented with
[3H]myo-inositol (0.5 µCi/well; Perkin-Elmer) was added. After
48 h, the medium was replaced with 100 µL of Locke’s buffer
(plus 20 mM Li+, pH 7.4) and incubated for 15 min at 37 °C before
replacement with agonists/antagonists in Locke’s buffer. The
incubation (45 min at 37 °C) was terminated by replacement of
the Locke’s solutions with 100 µL of 0.1 M HCl (10 min on ice).
The 96-well plates can be frozen at -20 °C at this stage until further
analysis. Home-made resin exchange columns (AG1-X8 Biorad,
140–14 444) were used to separate labeled inositol phosphates by
elution with 1 mL of 1 M ammonium formate/0.1 M formic acid
into 24-well visiplates (Perkin-Elmer). Scintillation liquid (Ulti-
maFlow AF, Perkin-Elmer) was added, and the plate was sealed
and vortexed before the radioactivity was determined by conven-
tional liquid scintillation counting (Microbeta,Perkin-Elmer) as
disintegrations per minute (DPM). At least three independent
experiments were performed.
(3H, m), 2.00–2.17 (2H, m), 2.86–3.08 (2H, m), 4.77–4.87 (1H,
m), 7.30–7.42 (4H, m), 7.55–7.63 (2H, m), 7.77 (1H, d). CI MS:
m/z 250 (M + H+).
2-Phenylethynyl-5,6,7,8-tetrahydro-quinoline (35). A solution
of the compound 34 (0.12 g, 0.48 mmol), DMAP (0.06 g, 0.049
mmol), Et3N (0.1 mL, 0.1 mmol), and tosyl chloride (0.2 g, 0.1
mmol) in dichloroethane (20 mL) was heated at 70 °C for 3 days.
Then, the solvent was evaporated. The residue was purified by
column chromatography on silica gel (60–100 µm, hexane/ethyl
acetate, 2:1). As a result, the Tos-derivative of 34 (0.1 g, 51%)
was obtained as an oil. To a solution of the obtained compound
(0.1 g, 0.25 mmol) in DMSO (1 mL), NaBH4 (0.05 g, 1.2 mmol)
was added. The reaction mixture was stirred at 60 °C for 6 h,
dissolved in chloroform (30 mL), and washed with water (3 × 25
mL). The organic layer was dried over Na2SO4, and the solvent
was evaporated. The product was isolated by column chromatog-
raphy on silica gel (60–100 µm, hexane/ethyl acetate, 2:1). As a
result, compound 34 (0.03 g, 52%) was obtained as a colorless oil.
1H NMR (DMSO-d6, 400 MHz): δH 1.65–2.07 (m, 4H), 2.75–2.85
(m, 2H), 2.92–3.02 (m, 2H), 7.23–7.39 (m, 5H), 7.55–7.63 (m, 2H).
MS: m/z (APCI+) 234 (M + H+).
6-Phenylethynyl-nicotinic acid methyl ester (36). Ethynylben-
zene (0.265 g (2.6 mmol), Pd(Ph3P)2Cl2 (0.07 g, 0.1 mmol), and
CuI (0.04 g, 0.2 mmol) were added to 6-chloro-nicotinic acid methyl
ester (0.342 g, 2 mmol) in 5 mL of triethylamine under argon
atmosphere. Then, the mixture was refluxed for 8 h under an argon
atmosphere and was monitored by TLC (ethyl acetate/hexane, 1:3).
Thereafter, the mixture was cooled, and the sediment that formed
was filtered off. The crude material was dissolved in dichlo-
romethane and was purified by column chromatography on silica
gel (dichloromethane). As a result, 0.165 g (35%) of compound
mGluR5 Binding Assay: [3H]MPEP] Assay. On the day of
assay, the membranes were thawed and washed four times by
resuspension in 50 mM Tris-HCl, pH 8.0, and centrifugation at
48 000 × g for 20 min, and finally, they were resuspended in 50
mM Tris-HCl, pH 7.5. The amount of protein in the final membrane
preparation (250–500 µg/ml) was determined according to the
method of Lowry.50 Incubations were started by the addition of
[3H]-MPEP (50.2 Ci/mmol, 5 nM, Tocris) to vials with 125–250
µg of protein (total volume 0.5 mL) and various concentrations of
the agents. The incubations were continued at room temperature
for 60 min (equilibrium was achieved under the conditions used).
Nonspecific binding was defined by the addition of unlabeled MPEP
(10 µM). Incubations were terminated using a Millipore filter
system. The samples were rinsed twice with 4 mL of ice cold assay
buffer over glass fiber filters (Schleicher and Schuell) under a
constant vacuum. Following the separation and rinse, the filters were
placed into scintillation liquid (5 mL of Ultima Gold), and the
radioactivity retained on the filters was determined with a conven-
tional liquid scintillation counter (Hewlett-Packard, Liquid Scintil-
lation Analyzer). The solubility of most of the compounds was
increased by using up to 5% DMSO in the final assay solution.
Specific binding was extremely high, that is, normally >85% and
essentially independent of buffer (Tris or HEPES oth 50 mM) and
pH (6.8–8.9). There was a clear saturable protein dependence,
and the chosen protein concentration used for subsequent assays
(250–500 µg/mL) was within the linear portion of this dependence.
Cold MPEP displaced hot ligand with an IC50 of 18.8 ( 4.1 nM.
The Kd of [3H]MPEP of 13.6 nM was determined by Scatchard
analysis and used according to the Cheng Prussoff relationship to
calculate the affinity of displacers as Kd values (IC50 of cold MPEP
equates to a Ki of 13.7 nM). Bmax was 0.56 pm/mg protein. Since
this assay is very stable, only two independent experiments were
performed.
1
36 was obtained as a white solid (mp ) 138–140 °C). H NMR
(DMSO-d6, 400 MHz): δH 3.92 (s, 3H), 7.44–7.54 (m, 3H), 7.65
(d, J ) 7 Hz, 2H), 7.78 (d, J ) 7 Hz, 1H), 8.32 (d, J ) 7 Hz, 1H),
9.09 (s, 1H). MS: m/z (APCI+) 238 (M + H+). Purity (HPLC):
100%.
5-Methoxy-2-phenylethynyl-5,6,7,8-tetrahydro-quinoline (37).
Sodium hydride (0.048 g, 1.2 mmol, 1.5 equiv) was added in an
argon atmosphere to a solution of 2-phenylethynyl-5,6,7,8-tetrahy-
dro-quinolin-5-ol (21a) (0.2 g, 0.8 mmol, 1 equiv) in dry dioxane
(5 mL), and the mixture was stirred for 30 min at room temperature.
Then iodomethane (0.16 g, 1.2 mmol, 1.5 equiv) was added to the
reaction mixture, and stirring was continued for 3 h at room
temperature. Then the reaction mixture was evaporated, and the
residue was subjected to column chromatography on silica gel
(60–100 µ, l ) 20 cm, d ) 2 cm, hexane/ethyl acetate 2:1). As a
result, 0.065 g (31%) of crystalline 37 was obtained with mp ) 94
°C. 1H NMR (400 MHz, CDCl3): δ 1.75–2.17 (4H, m), 2.86–3.09
(2H, m), 3.47 (3H, s), 4.30–4.40 (1H, m), 7.30–7.42 (4H, m),
7.56–7.65 (2H, m), 7.70 (1H, d). MS: m/z (CI MS) 264 (M + H+).
Purity: (HPLC): 100%.
(B) Pharmacology Assays. (Cell culture, cell preparation,
membrane preparation, and compound preparation are reported in
the Supporting Information.)
mGluR5 Assay: Calcium FLIPR Studies. The increase of
intracellular calcium after stimulation with the mGluR5 agonist
DHPG or L-quisqualate was measured using a fluorometric imaging
plate reader (FLIPR tetra) and the Ca-Kit (both Molecular Devices,
CA). Prior to the addition of agonist or antagonist, the medium
was aspirated, and the cells were loaded for 2 h at RT with 150 µL
of loading buffer consisting of Ca-sensitive dye (MD R8033)
reconstituted in sodium chloride (123 mM), potassium chloride (5.4
mM), magnesium chloride (0.8 mM), calcium chloride (1.8 mM),
D-glucose (15 mM), and HEPES (20 mM), pH 7.3. Subsequently,
the plates were transferred to FLIPR to detect calcium increase
with the addition of DHPG (300 µM) or L-quisqualate (100 nM)
measured as relative fluorescence units (RFU). Compounds were
preincubated for 10 min prior to the addition of the agonist. Effects
on the human mGluR5 were tested with L-glutamate as agonist and
a preincubation time of 5 min. The human mGluR5 was expressed
(C) Computational Chemistry: Generation of a Pharma-
cophore Model for mGluR1. (The generation of the mGluR1
homology model and the alignment of TM helices and EL2 of rat
mGluR1 to the bovine rhodopsin structural template is given in
the Supporting Information.)

646 Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3
VanejeVs et al.
(3) Brown, E. M.; Gamba, G.; Riccardi, D.; Lombardi, M.; Butters, R.;
Kifor, O.; Sun, A.; Hediger, J.; Lytton, M. A.; Hebert, S. C. Cloning
and characterization of an extracellular Ca21-sensing receptor from
bovine parathyroid. Nature 1993, 366, 575–580.
Flexible Alignment of 1, 2, and 3 by Application of MOE
(Chemical Computing Group). After generation of 3D structures
for all molecules (1, 2, and 3), using a favored force field for
small compounds (MMFF94), molecules 1 and 2 were super-
imposed in a flexible manner using the Flexible Alignment tool
included in the MOE software package (version 2003.02,
Chemical Computing Group) according to the following settings:
iteration limit, 200 attempts; failure limit, 20 configurations in
a row; energy cutoff, 10.0 kcal/mol greater than the minimum
generated value; configuration limit, 1000 alignment configura-
tions; R, 2.5; gradient test, 0.01; rmsd tolerance, 0.5Å; maximum
steps, 500 energy minimization steps; similarity terms, H-bond
donor/acceptor (1/1), aromaticity (3), acid/base (1), hydrophobe
(1), polar hydrogens (1), and volume (3)). Several orientations
have been calculated including their corresponding energy values.
One orientation with a low energy value and a reasonable overlay
(i.e., the most complete overlay with respect to the core
structures) was manually selected and both molecules were kept
in a fixed position relative to each other for further alignments.
In the next step, molecule 2 was superimposed onto this
orientation, and a sensible overlay was saved and fixed for the
next alignment procedure. Eventually, the remaining structure
was aligned on this orientation, resulting in a final alignment
for allosteric mGluR1 antagonists (Figure 2). Alignment settings
remained unchanged throughout the whole process.
Assignment of the Pharmacophore Elements. A pharmaco-
phore hypothesis was established based upon this flexible overlay
using the Pharmacophore Query Editor of the MOE software. This
tool displays certain chemical properties (H-bond donor/acceptor,
etc.) of nonhydrogen atoms of each molecule to be edited, which
are here referred to as annotation points. For this model, the default
PCH-type was employed. Here, annotation points of the same label,
which occur in all molecules at nearly the same spatial position
were presumed to be important. Query features were then assigned
to these important annotation points. A query feature denotes a point
in space with a radiuslike tolerance on spatial proximity and an
associated expression. The expression and the radius of each query
feature were manually edited. The diameter of each tolerance radius
was chosen to be sufficiently large to cover identical annotation
points of every molecule in that region.
(4) Jones, K. A.; Borowsky, B.; Tamm, J. A.; Craig, D. A.; Durkin, M. M.;
Dai, M.; Yao, W. J.; Johnson, M.; Grunwaldsen, C.; Huang, L. Y.;
Tang, C.; Shen, Q.; Salon, J. A.; Morse, K.; Laz, T.; Smith, K. E.;
Nagarathnam, D.; Noble, S. A.; Branchek, T. A.; Gerald, C. GABA(B)
receptors function as a heteromeric assembly of the subunits
GABA(B)R1 and GABA(B)R2. Nature 1998, 396 (6712), 674–679.
(5) Kaupmann, K.; Malitschek, B.; Schuler, V.; Heid, J.; Froestl, W.; Beck,
P.; Mosbacher, J.; Bischoff, S.; Kulik, A.; Shigemoto, R.; Karschin,
A.; Bettler, B. GABA(B)-receptor subtypes assemble into functional
heteromeric complexes. Nature 1998, 396 (6712), 683–687.
(6) Charest, P. G.; Oligny-Longpré, G.; Bonin, H.; Azzi, M.; Buvier, M.
The V2 vasopressin receptor stimulates ERK1/2 activity independently
of heterotrimeric G protein signalling. Cell. Signal. 2007, 19 (1), 32–
41.
(7) Palmer, R. K. The pharmacology and signaling of bitter, sweet, and
umami taste sensing. Mol InterV. 2007, 7 (2), 87–98.
(8) Hoon, M. A.; Adler, E.; Lindemeier, J.; Battey, J. F.; Ryba, N. J.;
Zuker, C. S. Putative mammalian taste receptors: a class of taste-
specific GPCRs with distinct topographic selectivity. Cell 1999, 96
(4), 541–551.
(9) Nakanishi, S.; Masu, M. Molecular diversity and functions of glutamate
receptors. Annu. ReV. Biophys. Biomol. Struct. 1994, 23, 319–348.
(10) Hollmann, M.; Heinemann, S. Cloned glutamate receptors. Annu. ReV.
Neurosci. 1994, 17, 31–108.
(11) Pin, J. P.; Galvez, T.; Prezeau, L. Evolution, structure, and activation
mechanism of family 3/C G-protein-coupled receptors. Pharmacol.
Ther. 2003, 98 (3), 325–354.
(12) O’Hara, P. J.; Sheppard, P. O.; Thogersen, H.; Venezia, D.; Haldeman,
B. A.; McGrane, V.; Houamed, K. M.; Thomsen, C.; Gilbert, T. L.;
Mulvihill, E. R. The ligand-binding domain in metabotropic glutamate
receptors is related to bacterial periplasmic binding proteins. Neuron
1993, 11 (1), 41–52.
(13) Kew, J. N. Positive and negative allosteric modulation of metabotropic
glutamate receptors: emerging therapeutic potential. Pharmacol Ther.
2004, 104 (3), 233–244.
(14) Lavreysen, H.; Pereira, S. N.; Leysen, J. E.; Langlois, X.; Lesage,
A. S. Metabotropic glutamate 1 receptor distribution and occupancy
in the rat brain: a quantitative autoradiographic study using
[3H]R214127. Neuropharmacology 2004, 46 (5), 609–619.
(15) Pin, J.-P.; Acher, F. The metabotropic glutamate receptors: Structure,
activation mechanism and pharmacology. Curr. Drug Targets: CNS
Neurol. Disord. 2002, 1 (3), 297–317.
Virtual Screening of a Compound Database. In the search for
new ligands, we virtually screened the Asinex Gold compound
collection, version of April 2003 (Asinex Ltd., Moscow, Russia,
http://www.asinex.com), which contained 194 563 molecules. All
molecules which did not fulfill the criteria of Lipinski’s Rule of
Five were removed from the database.51
(16) Conn, P. J.; Pin, J.-P. Pharmacology and functions of metabotropic
glutamate receptors. Annu. ReV. Pharmacol. Toxicol 1997, 37, 205–
237.
(17) Nicoletti, F.; Bruno, V.; Copani, A.; Casabona, G.; Knöpfel, T.
Metabotropic glutamate receptors: a new target for the therapy of
neurodegenerative disorders. Trends Neurosci. 1996, 19 (7), 267–271.
(18) Young, M. R.; Fleetwood-Walker, S. M.; Mitchell, R.; Munro, F. E.
Evidence for a role of metabotropic glutamate receptors in sustained
nociceptive inputs to rat dorsal horn neurons. Neuropharmacology
1994, 33 (1), 141–144.
(19) Varney, M. A.; Gereau, R. W. I. Metabotropic glutamate receptor
involvement in models of acute and persistent pain: Prospects for the
development of novel analgesics. Curr. Drug Targets: CNS Neurol.
Disord. 2002, 1 (3), 283–296.
(20) Spooren, W. P. J. M.; Vassout, A.; Neijt, H. C.; Kuhn, R.; Gasparini,
F. Anxiolytic-like effects of the prototypical metabotropic glutamate
receptor 5 antagonist 2-methyl-6-(phenylethynyl)pyridine in rodents.
J. Pharmacol. Exp. Ther. 2000, 295 (3), 1267–1275.
(21) Chiamulera, C.; Epping-Jordan, M. P.; Zocchi, A.; Marcon, C.; Cottiny,
C. Reinforcing and locomotor stimulant effects of cocaine are absent
in mGluR5 null mutant mice. Nat. Neurosci. 2001, 4 (9), 873–874.
(22) Spooren, W. P. J. M.; Gasparini, F.; Bergmann, R.; Kuhn, R. Effects
of the prototypical mGlu(5) receptor antagonist 2-methyl-6-(phenyl-
ethynyl)-pyridine on rotarod, locomotor activity and rotational re-
sponses in unilateral 6-OHDA-lesioned rats. Eur. J. Pharmacol. 2000,
406 (3), 403–410.
(23) Chavez-Noriega, L. E.; Schaffhauser, H. J. L.; Campbell, U. C.
Metabotropic glutamate receptors: potential drug targets for the
treatment of schizophrenia. Curr. Drug Targets CNS Neurol. Disord.
2002, 1 (3), 261–281.
Acknowledgment. The authors would like to thank the Merz
Compound Management Group for their great support and for
numerous HPLC measurements to assess the purity of the
compounds. Furthermore, we gratefully acknowledge the help
of Sylvia Willy, Andrea Baude, Sabine Denk, and Christina
Wollenburg for performing functional mGluR1/5 experiments
and mGluR5 binding assays.
Supporting Information Available: Table listing the analytical
techniques used to determine degree of purity for all target
compounds, the synthesis procedures of all intermediate products,
a detailed description of the cell culture, membrane preparation
and compound preparation, and a detailed description of the
generation of the mGluR1 homology model and the alignment of
TM helices and EL2 of rat mGluR1 to the bovine rhodopsin
structural template. This material is available free of charge via
the Internet at http://pubs.acs.org.
References
(24) Moghaddam, B. Targeting metabotropic glutamate receptors for
treatment of the cognitive symptoms of schizophrenia. Psychophar-
macol. 2004, 174 (1), 39–44.
(1) Klabunde, T.; Hessler, G. Drug design strategies for targeting
G-protein-coupled receptors. ChemBioChem 2002, 3 (10), 928–944.
(2) Wess, J. Molecular basis of receptor/G-protein-coupling selectivity.
(25) Noeske, T.; Gutcaits, A.; Parsons, C. G.; Weil, T. Allosteric Modulation
of Family 3 GPCRs. QSAR Comb. Sci. 2006, 25 (2), 134–146.
Pharmacol. Ther. 1998, 80 (3), 231–264.

Group I Metabotropic Glutamate Receptors
Journal of Medicinal Chemistry, 2008, Vol. 51, No. 3 647
(26) Palczewski, K.; Kumasaka, T.; Hori, T.; Behnke, C. A.; Motoshima,
H.; Fox, B. A.; Trong, I. L.; Teller, D. C.; Okada, T.; Stenkamp, R. E.;
Yamamoto, M.; Miyano, M. Crystal structure of rhodopsin: A G
protein-coupled receptor. Science 2000, 289 (5480), 739–745.
(38) Ioannis, N.; Molina, A.; Lynch, J.; Reamer, R.; Volante, R.; Reider,
P. Condensation of 2-methylbenzoxazole with aromatic aldehydes
bearing acidic protons. A convenient coupling in the synthesis of the
HIV-reverse transcriptase inhibitor L-696,229. J. Org. Chem. 1993,
58 (11), 3176–3178.
(39) Pettit, G. R.; Fleming, W. C.; Paull, K. D. Synthesis of the 6- and
7-hydroxy-5,8-dioxocarbostyrils. J. Org. Chem. 1968, 33 (3), 1089–
1092.
(40) Miyaura, N.; Yanagi, T.; Suzuki, A. The palladium-catalyzed cross-
coupling reaction of phenylboronic acid with haloarenes in the presence
of bases. Synth. Commun. 1981, 11 (7), 513–519.
(41) Suzuki, A.; Saveda, S.; Saegusa, T. New ring-opening polymerization
via a π-allyl complex. 1. Polymerization of diethyl 2-vinylcyclopro-
pane-1,1-dicarboxylate catalyzed by a palladium(0) complex. Mac-
romolecules 1989, 22 (3), 1505–1507.
(42) Gravius, A.; Pietraszek, M.; Schäfer, D.; Schmidt, W. J.; Danysz, W.
Effects of mGlu1 and mGlu5 receptor antagonists on negatively
reinforced learning. BehaV. Pharmacol. 2005, 16 (2), 113–121.
(43) Gasparini, F.; Lingenhohl, K.; Stoehr, N.; Flor, P. J.; Heinrich, M.;
Vranesic, I.; Biollaz, M.; Allgeier, H.; Heckendorn, R.; Urwyler, S.;
Varney, M. A.; Johnson, E. C.; Hess, S. D.; Rao, S. P.; Sacaan, A. I.;
Santori, E. M.; Velicelebi, G.; Kuhn, R. 2-Methyl-6-(phenylethynyl)-
pyridine (MPEP), A potent, selective and systemically active mGlu5
receptor antagonist. Neuropharmacology 1999, 38 (10), 1493–1503.
(44) O’Brien, J. A.; Lemaire, W.; Chen, T. B.; Chang, R. S.; Jacobson,
M. A.; Ha, S. N.; Lindsley, C. W.; Schaffhauser, H. J.; Sur, C.;
Pettibone, D. J.; Conn, P. J., Jr. A family of highly selective allosteric
modulators of the metabotropic glutamate receptor subtype 5. Mol.
Pharmacol. 2003, 64 (3), 731–740.
(45) Malherbe, P.; Kratochwil, N.; Zenner, M. T.; Piussi, J.; Diener, C.;
Kratzeisen, C.; Fischer, C.; Porter, R. H. Mutational analysis and
molecular modeling of the binding pocket of the metabotropic
glutamate 5 receptor negative modulator 2-methyl-6-(phenylethynyl)-
pyridine. Mol. Pharmacol. 2003, 64, 823–832.
(46) Malherbe, P.; Kratochwil, N.; Knoflach, F.; Zenner, M. T.; Kew, J. N.;
Kratzeisen, C.; Maerki, H. P.; Adam, G.; Mutel, V. Mutational analysis
and molecular modeling of the allosteric binding site of a novel,
selective, noncompetitive antagonist of the metabotropic glutamate 1
receptor. J. Biol. Chem. 2003, 278 (10), 8340–8347.
(27) Gribbon, P.; Sewing, A. Fluorescence readouts in HTS: no gain without
pain. Drug DiscoV. Today 2003, 8 (22), 1035–1043.
(28) Pagano, A.; Ruegg, D.; Litschig, S.; Stoehr, N.; Stierlin, C.; Heinrich,
M.; Floersheim, P.; Prezeau, L.; Carroll, F.; Pin, J. P.; Cambria, A.;
Vranesic, I.; Flor, P. J.; Gasparini, F.; Kuhn, R. The non-competitive
antagonists 2-methyl-6- (phenylethynyl)pyridine and 7-hydroxyimi-
nocyclopropan [b]chromen-1a-carboxylic acid ethyl ester interact with
overlapping binding pockets in the transmembrane region of group I
metabotropic glutamate receptors. J. Biol. Chem. 2000, 275, 33750–
33758.
(29) Mabire, D.; Coupa, S.; Adelinet, C.; Poncelet, A.; Simonnet, Y.; Venet,
M.; Wouters, R.; Lesage, A. S.; Beijsterveldt, L. V.; Bischoff, F.
Synthesis, structure-activity relationship, and receptor pharma-
cology of a new series of quinoline derivatives acting as selective,
noncompetitive mGlu1 antagonists. J. Med. Chem. 2005, 48 (6), 2134–
2153.
(30) Knoflach, F.; Mutel, V.; Jolidon, S.; Kew, J. N.; Malherbe, P.; Vieira,
E.; Wichmann, J.; Kemp, J. A. Positive allosteric modulators of
metabotropic glutamate 1 receptor: Characterization, mechanism of
action, and binding site. Proc. Natl. Acad. Sci. U.S.A. 2001, 98 (26),
13402–13407.
(31) Lavreysen, H.; Janssen, C.; Bischoff, F.; Langlois, X.; Leysen, J. E.;
Lesage, A. S. [3H]R214127: A novel high-affinity radioligand for the
mGlu1 receptor reveals a common binding site shared by multiple
allosteric antagonists. Mol. Pharmacol. 2003, 63 (5), 1082–1093.
(32) Noeske, T.; Jirgensons, A.; Stachenkovs, I.; Renner, S.; Jaunzeme, I.;
Trifanova, D.; Hechenberger, M.; Bauer, T.; Schneider, G.; Parsons,
C. G.; Weil, T. Virtual screening for selective allosteric mglur1
antagonists and structure-activity relationship investigations for
coumarine derivatives ChemMedChem 2007, in press.
(33) Asinex_Gold_Collection, http://www.asinex.com, 2003.
(34) Gasparini, F.; Andres, H.; Flor, P. J.; Heinrich, M.; Inderbitzin, W.;
Lingenhohl, K.; Muller, H.; Munk, V. C.; Omilusik, K.; Stierlin, C.;
Stoehr, N.; Vranesic, I.; Kuhn, R. [(3)H]-M-MPEP, A potent, subtype-
selective radioligand for the metabotropic glutamate receptor subtype
5. Bioorg. Med. Chem. Lett. 2002, 12 (3), 407–409.
(35) Shanazarov, A. K.; Kuzovkin, V. A.; Chistyakov, V. V.; Granik, V. G.
Acetals of lactams and amides. 60. N- and O-alkylation of 5-oxo-
5,6,7,8-tetrahydrocarbostyryils. Synthesis of isoxazolo[5,4-f]- and
pyrazolo[3,4-f]quinolines. Chem. Heterocycl. Compd. (Engl.Transl.)
1991, 27 (1), 71–76.
(36) Abu-Shanab, F. A.; Redhouse, A. D.; Thompson, J. R.; Wakefield,
B. J. Synthesis of 2,3,5,6-tetrasubstituted pyridines from enamines
derived from N,N-dimethylformamide dimethyl acetal. Synthesis 1995,
557–560.
(37) Baraldi, P. G.; Simoni, D.; Manfredini, S. An improved preparation
of enaminones from 1,3-diketones and ammonium acetate or amine
acetates. Synthesis 1983, 11, 902–903.
(47) Jirgensons, A.; Parsons, C. G.; Jaunzeme, I.; Kalvinsh, I.; Henrich,
M.; Vanejevs, M.; Kauss, V.; Danysz, W.; Weil, T. Tetrahydroqui-
nolones and their use as modulators of metabotropic glutamate
receptors. U.S. Patent 2006004 001, 2006.
(48) Parsons, C. G.; Jirgensons, A.; Jaunzeme, I.; Kalvinsh, I.; Henrich,
M.; Vanejevs, M.; Kauss, V.; Danysz, W.; Jatzke, C.; Weil, T.,
Tetrahydroquinolones and their use as modulators of metabotropic
glutamate receptors. Int. Patent WO2007023245, 2007.
(49) Carlier, P. R.; Du, D.-M.; Han, Y.-F.; Liu, J.; Perola, E.; Williams,
I. D.; Pang, Y.-P. Angew. Chem. Int. Ed. 2000, 39 (10), 1775–1777;
Angew. Chem. 2000, 112, 1845–1847.
(50) Lowry, O. H. et al. J. Biol. Chem. 1951, 193, 256–275.
(51) Lipinski, C. A.; Lombardo, F.; Dominy, B. W.; Feeney, P. J. AdV.
Drug Del. ReV. 2001, 46, 3–26.
JM0611298