Structure-activity relationship of triazafluorenone derivatives as potent and selective mGluR1 antagonists

Article abstract of DOI:10.1021/jm0504407

SAR (structure-activity relationship) studies of triazafluorenone derivatives as potent mGluR1 antagonists are described. The triazafluorenone derivatives are non-amino acid derivatives and noncompetitive mGluR1 antagonists that bind at a putative alloste

Full text of DOI:10.1021/jm0504407

7374
J. Med. Chem. 2005, 48, 7374-7388
Structure-Activity Relationship of Triazafluorenone Derivatives as Potent and
Selective mGluR1 Antagonists
Guo Zhu Zheng,* Pramila Bhatia, Jerome Daanen, Teodozyj Kolasa, Meena Patel, Steven Latshaw,
Odile F. El Kouhen, Renjie Chang, Marie E. Uchic, Loan Miller, Masaki Nakane, Sonya G. Lehto,
Marie P. Honore, Robert B. Moreland, Jorge D. Brioni, and Andrew O. Stewart
Neuroscience Research, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, Illinois 60064-6115
Received May 9, 2005
SAR (structure-activity relationship) studies of triazafluorenone derivatives as potent mGluR1
antagonists are described. The triazafluorenone derivatives are non-amino acid derivatives
and noncompetitive mGluR1 antagonists that bind at a putative allosteric recognition site
located within the seven-transmembrane domain of the receptor. These triazafluorenone
derivatives are potent, selective, and systemically active mGluR1 antagonists. Compound 1n,
for example, was a very potent mGluR1 antagonist (IC50 ) 3 nM) and demonstrated full efficacy
in various in vivo animal pain models.
Introduction
Glutamate is the most abundant excitatory neu-
rotransmitter in the central nervous system. Glutamate
modulates activity of many types of synapses by activat-
ing G-protein coupled receptors (GPCR) called metabo-
tropic glutamate receptors (mGluRs).1 There are three
different groups, groups I, II, and III, of mGluRs with
a total of eight distinct subtypes, mGluR1 to mGluR8,
based on their primary sequence similarity, signal
transduction linkages, and pharmacological profile.2
Group 1 mGluRs, mGluR1 and mGluR5, play key roles
in the central sensitization of pain, in addition to a
variety of functions with potential implications in
neurological and psychiatric disorders.3 Glutamate and
other excitatory amino acids are released from nerve
endings in the periphery under inflammatory condi-
tions. Glutamate or mGluR group 1 agonists induce
hyperalgesia when administrated peripherally.4 mGluRs
modulate pain transmission in the spinal cord, most
likely via sensitization of dorsal horn neurons to sustain
high-intensity C-fiber input.2c Normalization of gluta-
matergic neurotransmission in the spinal cord and
nociceptive afferents via inhibition of the group I
mGluRs is manifested in the attenuation of pain.5 There
is mounting evidence to support a role of peripheral
mGluRs in nociception and pain.6
We have investigated mGluR1 antagonists as a
therapy for the treatment of pain. The role of mGluR1
in the treatment of pain has not been validated, due to
lack of potent, selective, and systemically active mGluR1
antagonists. Earlier efforts to evaluate amino acid anta-
gonists, competing with the glutamate binding site, were
not successful due to poor selectivity, weak antagonism,
and lack of CNS availability.3 The first reported non-
competitive, non-amino acid-like mGluR1 antagonist,
CPCCOEt (7-(hydroxyimino)cyclopropa[b]chromen-1a-
carboxylate ethyl ester), elucidated an allosteric binding
site for the target.7 Other noncompetitive antagonists,
such as BAY 36-7620,8 R214127,9 dicarboxypyrroles,10
Figure 1. Triazafluorenone derivative identified from HTS.
and JNJ16259685,11 were also published. Instead of
competing at the glutamate binding N-terminal extra-
cellular domain (ECD), the reported noncompetitive
mGluR1 antagonists bind at the seven-transmembrane
domain (7-TMD).12 The allosteric modulation provides
additional possibilities for non-amino acid-like small
molecules of mGluR1 antagonists with superior physi-
cochemical properties which could facilitate the target
validation process for the treatment of pain.
Triazafluorenone derivatives (Figure 1) were identi-
fied as potent group I mGluRs antagonists in our high-
throughput screening (HTS) of this target. This type of
tricyclic heterocycle was first synthesized by Kadushkin
and co-workers.13 The synthetic approach described in
their paper was, however, lengthy, low yielding, and
difficult to apply to high-throughput synthesis. There-
fore, we needed to modify these procedures to suit high-
throughput synthesis for variations at several moieties
of the 5-thia-1,3,6-triazafluroenone pharmacophore.
These triazafluorenone derivatives consist of three
primary moieties: pyridine, thiophene, and pyrimidone
(Figure 1). We report here new triazafluorenone deriva-
tives 1a-ah and their analogues, as potent and selec-
tive mGluR1 antagonists. These new mGlu1 receptor
antagonists exhibited efficacy in several rat in vivo pain
models.
Chemistry
Variation at the P1 moiety (N3-substitutent) of the
triazafluorenone pharmacophore with both aryl and
* Corresponding author. Tel: 1-(847) 935 7513. Fax: 1-(847) 938
0072. E-mail: Guozhu.zheng@Abbott.com.
10.1021/jm0504407 CCC: $30.25 © 2005 American Chemical Society
Published on Web 10/21/2005

Triazafluorenone Derivatives as mGluR1 Antagonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23 7375
Scheme 1a
Scheme 2a
a
(a) HSCH2CONH2, NaOMe, MeOH, 96%. (b) t-BuOK, MeOH-
a
(a) Me2NCMe(OMe)2, EtOH, 23 °C, 2 days, 96%. (b)
pyridine, 98%. (c) NaNO2, 0 °C, 91%. (d) Cycloheptyl bromide,
NaH, DMF.
Me2NCH(OMe)2, 100 °C, 1.5 h, 93%. (c) HCl, MeOH, 0 °C, 9 h,
98%. (d) NaH, THF, HSCH2CO2Me (85%); HOCH2CO2Me (76%).
(e) Me2NCH(OMe)2, reflux, 5 h (X ) S 99%, X ) O 100%). (f)
R1NH2, toluene, p-TsOH.
Scheme 3a
alkyl groups required synthetic methods to accom-
modate the SAR studies. The Ullmann reaction could
be applied to aryl group attachment to the nitrogen
atom (N3) of the pyrimidone ring.14 Available aryl
reagents suitable for the Ullmann coupling reactions
only provided limited aryl groups using this approach.
Moreover, products generated as mixtures of O-aryla-
tion and N-arylation using this strategy also compli-
cated purification.
a
(a) (R2CO)2O, reflux, 2 h. (b) R1NH2, AcOH, reflux, 2 h.
thiophene analogue, 5-amino-4-dimethylaminothieno-
[8,9-b]pyridine-6-carboxylic acid methyl ester (6, Scheme
1).
Scheme 1 shows a viable synthetic approach for the
synthesis of triazafluorenone compounds with variation
of the R1 group for both aryl and alkyl substitutions (P1
moiety). A solution of malononitrile (2) in diethyl ether
was reacted with N,N-dimethylacetamide dimethyl ac-
etal in ethanol at 0 °C, and then warmed to room
temperature to give 2-(1-dimethylaminoethylidene)-
malononitrile (3) in quantitative chemical yield.15 Com-
pound 3 was then reacted with N,N-dimethylformamide
dimethyl acetal by heating at reflux to yield 2-(1,3-bis-
(dimethylamino)allylidene)malononitrile (4),16 which
could be purified through recrystallization in cold
methanol five times to give a combined 93% yield.
Intermediate 4 was then treated with anhydrous HCl
at 0 °C and cyclized to form the desired product,
4-dimethylamino-2-chloro-3-cyanopyridine (5, 98%).17
This product could be used for the next reaction without
further purification. In the presence of sodium hydride,
nucleophilic displacement of the 2-chloro atom of the
pyridine compound 5, followed by reaction with methyl
thioglycolate and cyclization in THF, gave a bicyclic
aminoester intermediate, 5-amino-4-dimethylamino-
thieno[8,9-b]pyridine-6-carboxylic acid methyl ester (6)
(85%).18 The formamidine derivative, 4-dimethylamino-
5-(dimethylaminomethyleneamino)thieno-[8,9-b]pyridine-
6-carboxylic acid methyl ester (8), was obtained in
quantitative yield after condensation with N,N-dimeth-
ylformamide dimethyl acetal. This formamidine deriva-
tive 8 was a key intermediate, which was used to make
a wide variety of R1 substituted derivatives, from aryl
to alkyl 5-thia-1,3,6-triazafluroenone derivatives 1a-
ah in parallel fashion, catalyzed by p-toluenesulfonic
acid in toluene (Scheme 1).
Tetraazafluorenone 14 was prepared as described in
Scheme 2. 2-Mercapto acetamide was reacted with
intermediate 5 in the presence of sodium methoxide in
methanol to generate the noncyclized intermediate, 2-(3-
cyano-4-dimethylaminopyridin-2-ylsulfanyl)acetamide
(11) in 96% yield. This intermediate was then treated
with potassium t-butoxide in a mixture of methanol and
pyridine to give the cyclized product 12 in quantitative
yield. The amino amide, 5-amino-4-dimethylaminothieno-
[8,9-b]pyridine-6-carboxylic acid amide (12), was oxi-
dized with sodium nitrate at 0 °C to form the tricyclic
heterocycle, 9-dimethylamino-3H-5-thia-1,2,3,6-tetra-
azafluoren-4-one (13) (91%).19 With this tetraazafluo-
renone intermediate 13 in hand, the 3-substituted
tetraazafluorenone 14 could be made by reaction of the
corresponding alkyl halide using sodium hydride in
dimethylformamide (DMF) with moderate chemical
yield (Scheme 2) accompanied by small amounts of
O-alkylation product.20
2-Substituted triazafluorenones 16a-c were prepared
as described in Scheme 3. The amino methylester 6
could be converted to another key intermediate oxazi-
nones 15a-c, in the presence of the desired carboxylic
acid anhydride with good yields (Scheme 3).21 The
oxazinones 15a-c were then treated with amine in
acetic acid to generate the 2-substituted triazafluo-
renone derivatives 16a-c with good yields.
The diazofluorenone analogue 22 was made using a
different route. 1,3-Dinitro-2-chlorobenzene (17) was
reacted with cuprous cyanide in DMF to give 1,3-dinitro-
2-cyanobenzene (18).22 One of the nitro groups of dini-
trocyanobenzene 17 was displaced with dimethylamine
to generate 1-dimethylamino-2-cyano-3-nitrobenzene
(19).23 The other nitro group of the nitrobenzene 19 was
reacted with methyl thioglycolate, followed by cycliza-
tion to form bicyclic aminoester, 5-amino-4-dimethy-
laminothieno[8,9-b]benzo-6-carboxylic acid methyl ester
(20), in the presence of sodium methoxide. The ben-
The furan analogues 10a,b (X ) O) were made from
the corresponding furan aminoester intermediate,
5-amino-4-dimethylaminofuro[8,9-b]pyridine-6-carboxy-
lic acid methyl ester (7). The latter was made using
methyl glycolate under the same conditions as the

7376 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23
Zheng et al.
Scheme 4a
Scheme 6a
a
(a) CuCN, DMF, 145 °C, 2 h, 70%. (b) Me2NH, DMF, 50 °C,
10 min, 95%. (c) HSCH2CO2Me, NaOMe, DMF, 23 °C, 16 h, 88%.
(d) Me2NCH(OMe)2, EtOH, reflux, 16 h, 90%. (e) p-Ethylaniline,
toluene, p-TsOH (cat.) reflux 16 h, 13%.
Scheme 5a
a
(a) Me2NCH(OMe)2, 100 °C, 1 h, 78%. (b) HCl, MeOH, 0 °C,
14.5 h, 98%. (c) HBr (30% in AcOH), 100 °C, 2 h, 100%. (d)
PMBnCl, NaH, DMF, 60 °C, 2.5 h, 37%. (e) HSCH2CO2Me,
NaOMe, DMF, 12 h, 86%. (f) Me2NCH(OMe)2, reflux 10 h, 100%.
(g) p-ethylaniline, toluene, p-TsOH (cat.), microwave 160 °C, 1 h,
39%. (h) TFA, 0 °C, 1.2 h, 100 %. (i) Tf2NPh, EtN(iPr)2, 80%. (j)
R3R4NH, CH2Cl2, 23 °C.
a
(a) NH2OH, CHO2H, H2SO4 (cat.), reflux 6 h, 83%. (b) Me2NH,
DMF, 23 °C, 4 h, 75%. (c) HSCH2CO2Me, NaOMe, DMF, 23 °C,
16 h, 88%. (d) Me2NCH(OMe)2, EtOH, reflux 16 h, 90%. (e) R1NH2,
toluene, p-TsOH (cat.), reflux 16 h.
2-chloro-3-cyano-4-ethoxypyridine (31a) and 2-chloro-
3-cyano-4-methoxypyridine (31b). The pyridine mixture
of 31a,b was deprotected with HBr (30%), giving the
desired product, 2-bromo-3-cyano-4-hydroxypyridine
(32a), with a small amount of 2-chloro-3-cyano-4-hy-
droxypyridine (32b).27 Protecting the 4-hydroxy group
of 32a and 32b with p-methoxybenzylchloride (PMBCl)
in the presence of NaH yielded protected pyridine
analogues 33a (37%) and 33b (8%). The protected
pyridines 33a and 33b were then condensed with
methyl thioglycolate, followed by cyclization to form
aminoester intermediate, 5-amino-4-(4′-methoxybenzy-
loxy)thieno[8,9-b]pyridine-6-carboxylic acid methyl ester
(34, 89%). This aminoester 34 was converted to forma-
midine derivative 35 in the presence of N,N-dimethyl-
formamide dimethyl acetal in quantitative yield. The
triazafluorenone analogue 36 was generated in the
presence of p-ethylaniline and catalyst, p-toluenesulfon-
ic acid, in toluene (39%). 9-(4′-Methoxybenzyloxy)-3-(4′-
ethylphenyl)-3H-5-thia-1,3,6-triazafluoren-4-one (36) was
then deprotected with trifluoroacetic acid (TFA) to give
9-hydroxy-3-(4′-ethylphenyl)-3H-5-thia-1,3,6-triazafluo-
ren-4-one (37, 100%).28 The hydroxy group of 37 was
converted to a triflate group in the presence of N,N-bis-
(trifluoromethanesulfonyl)phenylamide.29 The key tri-
flate intermediate 38 was used to prepare analogues
39a-f by addition of the appropriate amine in dichlo-
romethane at room temperature (Scheme 6).
zoaminoester 20 was converted to formamidine inter-
mediate 21 by heating with N,N-dimethylformamide
dimethyl acetal. Catalyzed by p-toluenesulfonic acid in
toluene, the intermediate 21 formed the pyrimidone ring
with an aryl or alkyl amine to produce 22 (Scheme 4).
Isomeric fused pyridine analogues 28a,b were pre-
pared as described in Scheme 5. Dichloropyridinecar-
boxaldehyde 23 was condensed with hydroxylamine,
which after dehydration formed 3,5-dichloro-4-cyanopy-
ridine (24).24 One of the chloro groups of 24 was easily
displaced by aqueous dimethylamine to generate 3-chloro-
4-cyano-5-dimethylaminopyridine (25). A nucleophilic
displacement of the 3-chloro group of 25 by thioglycolate,
followed by cyclization, afforded aminoester analogue,
3-amino-4-dimethylaminothieno[2,3-c]pyridine-2-car-
boxylic acid methyl ester (26).25 The aminoester 26 was
heated to reflux with N,N-dimethylformamide dimethyl
acetal yielding formamidine intermediate 27 quantita-
tively. The formamidine intermediate 27 was then
reacted with either 4-methyl- or 4-ethylanaline in the
presence of catalyst, p-toluenesulfonic acid, to form the
final triazafluorenone products 28a and 28b, respec-
tively (Scheme 5).
Variations of R3 and R4 at the 4-amino group of the
pyridine moiety (P3 moiety) were achieved via triflate
intermediate 38 (Scheme 6). 2-(1-Ethoxyethylidene)-
malononitrile (29) was reacted with N,N-dimethylfor-
mamide dimethyl acetal and generated a mixture of
The 9-methylamino triazafluorenone derivative 40
was obtained by demethylation of 1n using hydrogen
peroxide and formic acid in excellent chemical yield
(79%, Scheme 7).
The 9-dimethylamino substituent of triazafluorenone
derivatives 1n and 1ac could be converted to N-oxides
desired products 30 (30a, R5 ) ethyl; 30b, R5
)
methyl).26 The mixture of 30a,b was then treated with
anhydrous HCl and converted to a mixture of pyridines,

Triazafluorenone Derivatives as mGluR1 Antagonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23 7377
Scheme 7a
Table 1. N-Aryl Substituted Triazafluroenone Derivatives
Affect Both Potency and Selectivitya
Scheme 8a
41a and 41b respectively when treated with phthalic
anhydride and hydrogen peroxide-urea complex (Scheme
8).30
Results and Discussion
Both binding affinity (Ki) and functional antagonism
(IC50) were used to measure the biochemical potency of
mGluR1 antagonists for SAR studies.
Binding assays have been reported for both mGluR1
and 5 subtypes using radioligands that have been shown
to be noncompetitive to the extracellular N-terminal
glutamate binding site.9 The triazafluroenone deriva-
tives reported here compete with these allosteric binding
radioligands. Binding assays were used to generate
affinities for ligands at both mGluR1 and 5 subtypes.
In addition to affinity assays, functional antagonism and
potency (IC50) were measured using a calcium mobiliza-
tion assay (vide infra), measuring the ability of ligands
to block glutamate activation in engineered 1321 cells
expressing the mGlu1 or 5 receptor.
The first SAR table shown (Table 1) that similar SAR
trends were observed using both Ki values in the binding
assay and IC50 values in the calcium mobilization assay
within each subtype (mGluR1 and 5). Functional IC50
values in the calcium mobilization assays were therefore
used for additional SAR studies for measurement of
potency and subtype (mGluR1 vs mGluR5) selectivity
of antagonists (Table 1).
The triazafluorenone derivatives consisted of three
submoieties: aminopyridine ring (P3), thiophene ring
(P2), and pyrimidone ring (P1) (Figure 1). From Table
1, variation at the P1 moiety of the pyrimidone indicated
that the N3-aryl substituent of the pyrimidone affected
both in vitro potency and selectivity (mGluR1 vs
mGluR5). With a viable synthetic route to access
derivatives with different substituents at the N3 posi-
tion of the pyrimidone ring, we investigated SAR of this
key substituent. This substituent (P1) tolerated both
aryl and cyclic alkyl variations. Table 1 summarizes
typical aryl substitution. Various aryl groups, from
alkylaryl to haloaryl, hydroxyaryl, and heteroaryl, all
produced potent mGluR1 antagonists. Compound 1b,
4-methylphenyl triazafluorenone was a very potent
mGluR1 antagonist (IC50 ) 5 nM). This compound
exhibited weaker mGluR5 antagonism (IC50 ) 890 nM).
The 4-ethylphenyl triazafluorenone 1n maintained high
mGluR1 potency (IC50 ) 3 nM), and the selectivity of
this compound (mGluR1 vs mGluR5) was also high
(mGluR5 IC50 ) 442 nM) (entry 11). Similar results
were observed for the p-halide substituted phenyl ana-
logues. N3-4-Bromophenyl triazafluorenone 1f (mGluR1
a (a) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean. (b) Rat
mGluR1 binding (cerebellum) and rat mGluR5 binding (cortex),
mean of multiple results with standard error of mean. (c) Radio-
ligand [3H]R214127 was used (ref 9). (d) Radioliagnd [3H]MPEP
was used (ref 33).
IC50 ) 4 nM; mGluR5 IC50 ) 1250 nM) was the most
potent mGluR1 antagonist among 4-halophenyl deriva-
tives (entries 3-5). The strong electron withdrawing
p-CF3 group substituted phenyl analogue (1g) was also
a potent mGluR1 antagonist devoid of mGluR5 activity.
Disubstituted aryl triazafluorenone derivative, 1h, was
only slightly less potent (IC50 ) 17 nM) compared to the
monosubstituted aryl derivative 1b and now lacked
completely mGluR5 potency. Similar results were ob-
served for dichlorophenyl triazafluorenone derivative 1i,
which was more subtype selective than the monochlo-
rophenyl derivative 1e (entries 4 and 6). Heteroaryl
substituted triazafluorenone derivatives, 1j (IC50 ) 39
nM) and 1k (IC50 ) 122 nM), were quite potent mGluR1
antagonists with little activity at mGluR5 (entries 7 and
8). Benzyl substitution disfavored mGluR1 potency
slightly (1o, mGluR1 IC50 ) 281 nM, entry 12).
In general, para-substitution on the N3-aryl pyrimi-
done analogues boosted mGluR1 potency. This was
exemplified by the o-, m-, p-ethyl substituted aryl
analogues (1l, 1m, and 1n). This effect of the para-
substitution often improved mGluR5 potency of triaz-
afluorenone analogues as well.
Additional SAR studies examined the scope of para-
substituted phenyl analogues in more detail and are

7378 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23
Zheng et al.
Table 3. Alkyl Substituted Triazafluroenone Derivativesa
Table 2. Para Substitution Affects Both Potency and
Selectivitya
a (†) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean.
shown in Table 2. Replacing hydrogen (1a) with p-
methyl (1b) or p-ethyl (1n) at the para-position of the
N3-phenyl moiety, as stated, demonstrated a clear trend
in boosting mGluR1 potency (Table 2). There were limits
to size or lipophilicity of this para-substituent that were
tolerated. Substituted phenyl pyrimidone derivatives
containing larger alkyl groups, such as n-propyl or
isopropyl, , 1p and 1q, became less potent mGluR1
antagonists (IC50 ) 180 and 407 nM, respectively,
entries 4 and 5). When a bulky tert-butyl group was
incorporated at the para-position of the phenyl triaz-
afluorenone (1r), a dramatic loss of potency was ob-
served (IC50 ) 8.5 µM, entry 6). Cyclohexyl substituted
phenyl triazafluorenone 1t was also inactive in the
calcium influx assay (entry 8).
A lipophilic cycloalkyl substituent was an effective
replacement for the N3-aryl substituent. The size of the
cyclic alkyl group modulated in vitro potencies of these
cyclic alkyl triazafluorenone derivatives (Table 3). The
selectivity of these new mGlu1 receptor antagonists also
depended on the size of the cyclic alkyl ring. The trend
was that increasing the size of cyclic ring favored in vitro
potency for both mGluR1 and mGluR5. Once the ring
size reached seven-membered ring, the optimum in vitro
potency was obtained (1ab, IC50 ) 1 nM, entry 8).
Compound 1ab was also one of the most potent mGluR5
antagonists from this series (mGluR5 IC50 ) 147 nM).
The eight-membered cyclic alkyl substituted pyrimi-
done, 1ad (IC50 ) 13 nM), was not as potent as the
seven-membered alkyl pyrimidone derivative 1ab (en-
tries 8 and 10). Bulky adamantyl substituted pyrimi-
done, 1ae, was a mixed mGluR1 and mGluR5 antago-
nist (mGluR1 IC50 ) 76 nM, mGluR5 IC50 ) 154 nM,
entry 12) exhibiting some drop in potency. Incorporating
a nitrogen atom into the cyclic alkyl ring, such as
compound 1ac, was also well tolerated (IC50 ) 10 nM,
entry 9). A different substitution pattern on the cyclic
ring did not greatly affect in vitro potency or selectivity
a (†) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean.
to the degree that had been observed with the aryl
analogues (compounds 1y, 1z, and 1aa, entries 6-8).
Acyclic alkyl substituted triazafluorenone, 1af, was not
as potent as the corresponding cyclic compound 1w
(entries 3 and 13). Insertion of a methylene (CH2)
linkage between cyclic alkyl group and N3 atom of the
pyrimidone ring resulted in weaker mGluR1 antagonists
(entries 14 and 15). Compound 1ag (mGluR1 IC50 ) 26
nM) was 5 times weaker than its parent derivative 1w
(mGluR1 IC50 ) 5 nM). The elongated cyclohexyl tri-
azafluorenone, 1ah, was also less potent than its
truncated parent compound 1x, similar to the results
observed with the aryl triazafluorenone derivatives (1a
vs 1o). These methylene inserted derivatives, 1ag and
1ah, could however be argued to be more selective
mGluR1 antagonists (mGluR5 IC50 > 100 000 nM) than
analogues 1w and 1x since they showed no mGluR5
activity at the highest concentration tested (entires 14
to 15).
The pyrimidone ring could also be converted to a
triazinone ring. Analogue 14 was still a very potent
mGluR1 antagonist (IC50 ) 19 nM), comparable to its
parent compound, 1ab (entry 2, Table 4).
The C2 position of the triazafluorenone analogues
could be substituted with an alkyl group to produce
potent mGluR1 antagonists. Table 4 shows that there

Triazafluorenone Derivatives as mGluR1 Antagonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23 7379
Table 6. Diazafluorenone and Triazafluorenone Derivativesa
Table 4. Tetraazafluroenone and C2-Substituted
Triazafluroenone Derivativea
a (†) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean.
Table 7. 9-Alkylamino Triazafluorenone Derivativesa
a (†) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean.
Table 5. 5-Oxa-1,3,6-triazafluroenone Derivativesa
a (†) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean.
was also some size limitation for this alkyl substitution.
C2-methyl incorporated triazafluorenone derivative,
16a, was a 10-fold weaker mGluR1 antagonist (mGluR1
IC50 ) 44 nM, entry 4) than 1n. This compound was
also less selective (mGluR1 vs mGluR5) when compared
with its parent non-C2 substituted analogue, 1n. Larger
alkyl groups at C2 position of the triazafluorenone
derivatives further disfavored in vitro potency (entries
5 and 6).
The thieno moiety of the triazafluorenone analogues
could be replaced by a furan moiety (Table 5). The N3-
aryl substituted 5-oxa-1,3,6-triazafluorenone analogues
were weaker mGluR1 antagonists compared with the
5-thia-1,3,6-triazafluorenone analogues. Compound 10a
(IC50 ) 1460 nM), for example, was much weaker than
thieno analogue 1n (IC50 ) 3 nM). On the other hand,
the N3-cyclic alkyl 5-oxa-1,3,6-triazafluorenone ana-
logues were potent mGluR1 antagonists. Compound 10b
(IC50 ) 82 nM) retained the potency of the 5-thia-1,3,6-
triazafluorenone analogue 1aa, but lost selectivity (vs
mGluR5) (entry 4, Table 5).
a (†) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean.
C6-position of compound 1n to the C7-position (Figure
1) produced derivative 28a, which was still a potent
mGluR1 antagonist (IC50 ) 183 nM, entry 3). The IC50
value of the 1,3,7-triazafluorenone analogues 28a could
be improved by reoptimization of the P1 moiety. Com-
pound 28b, the p-methylphenyl analogue for example,
was a more potent and selective mGluR1 antagonist
(IC50 ) 43 nM, entry 4) than the p-ethylphenyl analogue
(28a).
The dimethylamino group of the pyridine ring could
also be replaced by various mono- or diamino groups
(Table 7). Small monoalkylamino group substituted
derivatives, such as cyclopropylamino derivative 39d
(IC50 ) 41 nM, entry 4), were potent mGluR1 antago-
nists. Similarly, small dialkylamino group substituted
triazafluorenone derivatives were also potent mGlu1
receptor antagonists. Ethylmethylaminopyridine de-
Replacement of the pyridine ring of the triazafluo-
renone derivatives by a phenyl ring resulted in analogue
22, a far less potent mGluR1 antagonist (IC50 ) 949 nM)
compared with the corresponding pyridine analogue 1n
(entry 2, Table 6). Moving the nitrogen group from the

7380 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23
Zheng et al.
Table 8. Triazafluorenone Compound 1n in Animal Pain
indication that mGluR1 antagonism may be a valid
therapeutic target for treatment of pain.
Modelsa
Biology
Rat mGluR1 and Rat mGluR5 binding assays were
carried out as described by El-Kouhen et al.31 Briefly,
mGluR1 binding was performed using rat cerebellum
membrane preparation using [3H]R214127 (9 Ci/mmol)
as radioligand, and nonspecific binding was determined
in the presence of 1 µM LY-456066.32 Rat mGluR5
binding was performed using rat cortex preparation and
the selective mGluR5 radioligand. Nonspecific binding
was determined using 10 µM MTEP.33 Specific binding
was obtained by calculating the difference between total
binding and nonspecific. Radioligand saturation binding
data were analyzed using Prism GraphPad software
(San Diego, CA). Competition binding data were ana-
lyzed by nonlinear regression curve fitting. Ki values
were determined by the method of Cheng and Prusoff.34
The Ca2+ mobilization assay was performed using
Fluorometric Imaging Plate Reader (FLIPR) assays.
Calcium mobilization induced by L-glutamate was modu-
lated by an mGlu1 receptor antagonist. Cloning of
human mGluR1 and human mGluR5 is described by El-
Kouhen et al.,31 as well as FLIPR assays. Human
mGluR1 and human mGluR5 receptor-transfected
1321N1 cells (coexpressing rat GLAST) were plated into
96-well biocoat black-wall/clear bottom microplates
(Becton Dickinson, Boston, MA) at a density of 30 000
cells per well, in DMEM glutamate-free media (contain-
ing 10% FBS, 1% GluMax, 500 µg/mL G418, 200 µg/mL
hygromycin B). After 2 days of culture, the culture
medium was removed and replaced by 100 µL per well
of D-PBS containing 0.04% Pluronic F127 and 4 µM
Fluo-4 AM (fluorescent calcium indicator dye). After
incubation for 1 h at room temperature, the cells were
washed four times with D-PBS in a plate washer
(Molecular Devices, Sunnyvale, CA), and 150 µL of
D-PBS was left in each well. To assess the effect of
antagonists on 10 µM L-glutamate-induced calcium
released, antagonists were preincubated for 3 min before
the addition of the agonist. FLIPR instrument made
fluorescent reading for 5 min (every second for the first
minute of compound addition (antagonist and agonist),
then every 5 s for the remaining time). The instrument
software normalizes the fluorescent reading to give
equivalent initial reading at time zero, and all data were
normalized with the response of 10 µM L-glutamate.
Detailed experimental procedures for animal pain
models are described by Lehto et al.35 Male Sprague-
Dawley rats (Charles River, Wilmington, MA) weighing
200-300 g were utilized in all experiments.
a (a) 1321N1 cells expressing either human mGluR1 or mGluR5,
mean of multiple results with standard error of mean. (b) Test
performed 30 min after intraperitoneal administration of com-
pound in rats (6 rats per group). Vehicle was 10% DMSO/PEG (5
mL/kg) (ref 35).
rivative, 39a, was a potent example (mGluR1 IC50
)
37 nM, entry 1). In general, the monoalkylamino
substituted pyrimidone derivatives were slightly less
potent than dialkylamino substituted derivatives.
Monomethylaminopyridine derivative 40 (IC50 ) 98 nM,
entry 7), for example, was weaker than the dimethy-
laminopyridine derivative 1n (IC50 ) 3 nM). Various
alkyl, alkoxyalkyl, and halide alkyl amino groups could
be used to replace the original dimethylamino group and
maintained good in vitro potency. It was interesting to
note that even a polar N-oxide group was tolerated.
Converting the dimethylamino group of the triazafluo-
renone derivative 1n to the corresponding N-oxide
moiety gave a new polar mGluR1 antagonist 41a with
reasonable in vitro potency (IC50 ) 306 nM, entry 8).
Better in vitro potency of the N-oxide pyrimidone
derivatives was achieved by modifying N-substituents
at triazafluorenone ring (P1 moiety). Compound 41b,
for example, was a potent polar mGluR1 antagonist
(IC50 )167 nM, entry 9).
To test the hypothesis that mGluR1 antagonists could
be useful analgesics, we conducted initial in vivo
pharmacology studies with selected triazafluorenone
antagonists. Compound 1n (PK profile in rat: t1/2 ) 1.9
h, Vd ) 4.8 L/kg, Cmax(ip) ) 137.5 ng/mL, F(ip) ) 45%,
F(oral) ) 12%), for example, was fully efficacious in CFA
(complete Freund’s adjuvant-induced thermal hyperal-
gesia, ED50 ) 15 µmol/kg, ip), carrageenan-induced
hyperalgesia (carrageenan ED50 ) 11 µmol/kg, ip), and
formalin (phase II, ED50 ) 19 µmol/kg, ip) rat models
of pain. No rotorod side effect was observed at doses up
to 300 µmol/kg (rotorod ED50 > 300 µmol/kg), and no
sedative effect was observed at effective doses indicated
above (Table 8).
In summary, we have identified a new triazafluo-
renone series of mGluR1 antagonists. These novel
triazafluorenone derivatives are non-amino acid-like,
potent, and subtype selective mGluR1 antagonists that
interact with a well-defined allosteric recognition site
of the metabotropic glutamate receptor. Therefore these
novel triazafluorenone mGluR1 antagonists are non-
competitive with glutamate. SAR studies have identified
the N3-phenyl substituent as a key moiety contributing
to potency and selectivity. A specific site demonstrating
this substituent effect was the para-position of this
phenyl ring, and this site was optimized by introduction
of small alkyl groups. One of these optimized antago-
nists, 1n, is a single digit nanomolar mGluR1 antagonist
with at least 40-fold selectivity over the mGluR5
subtype. Analogue 1n also exhibits efficacy in preclinical
rodent pain models without showing any rotorod side
effect up to 300 µmol/kg. These data are a preliminary
In the formalin-induced spontaneous pain model
(Formalin), following a 30-min acclimation period to
individual observation cages, 50 µL of a 5% formalin
solution was injected (sc) into the dorsal aspect of the
right hind paw and the rats were then returned to the
clear observation cages. Rats were observed for periods
of time corresponding to phase 1 (0-10 min) and phase
2 (30-50 min) of the formalin test. Nociceptive behav-
iors were recorded from animals during the session by
observing each animal for one 60-s observation period
during each 5-min interval. Nociceptive behaviors re-
corded included flinching, licking, or biting the injected
paw.

Triazafluorenone Derivatives as mGluR1 Antagonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23 7381
(113.7 g, 81%). The final residue was purified by a short
column chromatography (SiO2, ethyl acetate) to give an
additional 17.2 g of product (12%).
Carrageenan- and Complete Freund’s Adjuvant-
Induced Thermal Hyperalgesia. Unilateral inflam-
mation was induced by injecting 100 µL of a 1% solution
of λ-carrageenan or 150 µL of a 50% solution of complete
Freund’s adjuvant (CFA) (Sigma Chemical Co., St.
Louis, MO) in physiological saline into the plantar
surface of the right hind paw of the rat. The hyperal-
gesia to thermal stimulation was determined 2 or 48 h
following carrageenan or CFA injection.
Rotorod performance was measured using an ac-
celerating rotorod apparatus (Omnitech Electronics,
Inc., Columbus, OH). For the rotorod assay, rats were
allowed a 30 min acclimation period in the testing room
and then placed on a 9 cm diameter rod that increased
in speed from 0 to 20 rpm over a 60 s period. The time
required for the rat to fall from the rod was recorded,
with a maximum score of 60 s. Each rat was given 3
training sessions. Latencies to fall from the rotorod were
determined 30 (ip) or 60 (po) min following compound
injection.
1H NMR (CDCl3, 300 MHz): δ ) 7.42 ppm (d, J ) 12.2 Hz,
1H); 4.36 (d, J ) 12.2 Hz, 1H); 3.15 (s, 6H), 3.05 (s, br, 6H).
MS (ESI/NH3): m/z ) 191.1 (M + H+).
4-Dimethylamino-2-chloro-3-cyanopyridine (5). To a
slurry of 2-(1,3-bisdimethylaminoallylidene)malononitrile (4,
120.0 g, 632 mmol) in methanol (1500 mL), HCl gas was
introduced gently at 0 °C. The reaction mixture became
homogeneous in about 1 h, and was allowed to stir under
constant HCl flow for additional 9 h at 0 °C. N2 was bubbled
though the reaction mixture for 2 h, and all the solvent was
removed. The residue solid was redissolved in CH2Cl2, and
washed with water/K2CO3/water. The organic layer was sepa-
rated and dried over Na2SO4. Removal of salt and solvent gave
a pure product (113.0 g, 98%).
1H NMR (CDCl3, 300 MHz): δ ) 7.96 ppm (d, J ) 6.4 Hz,
1H); 6.59 (d, J ) 6.4 Hz, 1H); 3.30 (s, 6H). MS (ESI/NH3): m/z
) 181.9 (M + H+).
5-Amino-4-dimethylaminothieno[8,9-b]pyridine-6-car-
boxylic Acid Methyl Ester (6). To a solution of methyl
thioglycolate (40 mL, 442 mmol) in anhydrous THF (500 mL),
sodium hydride (60%, 20.0 g, 500 mmol) was added in small
portions at 0 °C. A solution of 4-dimethylamino-2-chloro-3-
cyanopyridine (5, 80.0 g, 442 mmol) in THF (1000 mL) was
added. The reaction mixture was allowed to stir for 1 day at
room temperature. Additional NaH (60%, 11.0 g, 276 mmol)
was added, and the reaction mixture was then heated to reflux
for 1 h 50 min. After being cooled down to room temperature,
the reaction mixture was quenched with saturated NH4Cl.
Organic solvent was removed under vacuum, and the aqueous
layer was extracted with dichloromethane (500 mL). The
organic layer was separated and washed with water and brine,
then dried over Na2SO4. The solvent was concentrated, and a
pure product was precipitated. Solid was collected and washed
with cold methanol (3 × 5 mL), then ether, and dried under
vacuum (82.3 g, 74%). Additional solid was collected via the
same, repeated procedures (12.1 g, 11%).
All tests were performed 30 min after intraperitoneal
administration of compound 1n, using 10%DMSO/PEG
as vehicle (5 mL/kg).
Experimental Section
General Information. Unless otherwise noted, all sol-
vents, including anhydrous solvents, and chemicals were
purchased from Aldrich Co. and/or Acros Organics, and used
without further purification. NMR spectra were obtained at
either 300 or 500 MHz using deuterated solvent and TMS
(tetramethylsilane) as the internal standard. The mass spectra
were recorded on a Finnigin-4000 instrument using EI or CI.
Elemental analyses were performed by Robertson Microlit
Laboratories with (0.4% of theoretical value. Column chro-
matography purifications used silica gel 60 (230-400 mesh)
from E. Merck. Thin-layer chromatography analyses were
performed on silica gel plate 60 F254 with a thickness of 0.25
mm.
1H NMR (CDCl3, 300 MHz): δ ) 8.43 ppm (d, J ) 5.2 Hz,
1H), 6.83 (d, J ) 5.2 Hz, 1H), 6.74, s, br, 2H), 3.86 (s, 3H),
2.84 (s, 6H). MS (ESI/NH3): m/z ) 251.9 (M + H+).
4-Dimethylamino-5-(dimethylaminomethyleneamino)-
thieno[8,9-b]pyridine-6-carboxylic Acid Methyl Ester (8).
5-Amino-4-dimethylaminothieno[8,9-b]pyridine-6-carboxylic acid
methyl ester (6, 32.0 g, 127 mmol) was dissolved in ethanol
(150 mL) and N,N-dimethylformamide dimethyl acetal (100
mL), and heated to reflux for 4.5 h. Excess of solvent and
reagent were removed to give a yellow solid product (38.5 g,
99%).
LY-456066 and [3H]R214127 were synthesized at Abbott
Laboratories (Abbott Park, IL). [3H]MPEP was from Tocris
(Bristol, U.K.). Rat cerebellum and rat cortex were purchased
from Pel-Freez Biologicals (Rogers, AR). Pluronic F127 and 4
µM Fluo-4 AM were bought from Molecular Probes (Eugene,
OR). D-PBS, neomycin (G418), hygromycin B, GluMax, and
tissue culture reagents were from Invitrogen/Life Technology.
All other chemicals were purchased from Sigma unless oth-
erwise noted. Male Sprague-Dawley rats (Charles River,
Wilmington, MA) weighing 200-300 g were utilized in all
experiments.
2-(1-Dimethylaminoethylidene)malononitrile (3). To a
solution of N,N-dimethylacetamide dimethyl acetal (90%, 125.0
g, 845 mmol) in ethanol (75 mL), a solution of malononitrile
(2, 56.0 g, 847.2 mmol) in diethyl ether (500 mL) was added
slowly at 0 °C. The reaction mixture was then gradually
warmed to room temperature and stirred for 2 days. Solvent
was removed, and the solid was redissolved in ethyl acetate,
and was purified via a short column chromatography (SiO2,
ethyl acetate) to give a pale yellow solid product (109.2 g, 96%).
1H NMR (CDCl3, 300 MHz): δ ) 3.32 ppm (s, br, 6 H); 2.28
(s, 3H). MS (ESI/NH3): m/z ) 135.9 (M + H+).
2-[1,3-Bis(dimethylaminoallylidene)]malononitrile (4).
2-(1-Dimethylaminoethylidene)malononitrile (3, 100.0 g, 740
mmol) was dissolved in N,N-dimethylformamide dimethyl
acetal (94%, 210 mL, 1.48 mol), and the reaction mixture was
heated to reflux at 100 °C for 1 h 20 min and then cooled down
to room temperature. Solid was collected, and washed with
cold methanol (10 × 3 mL) to give a yellow solid product. The
mother liquor was concentrated, and the solid was collected
again, and washed with cold methanol. This procedure was
repeated 5 times, and all the solid products were combined
1H NMR (CDCl3, 300 MHz): δ ) 8.30 ppm (d, J ) 5.4 Hz,
1H), 7.41 (s, 1H), 6.60 (d, J ) 5.4 Hz, 1H), 3.82 (s, 3H), 3.17
(s, br, 3H), 3.07 (s, br, 3H), 2.99 (s, 6H). MS (ESI/NH
3): m/z )
307.0 (M + H+).
General Procedure for the Synthesis of Tricyclic
Pyrimidone Derivatives 1a-ah. 9-Dimethylamino-3-(p-
tolyl)-3H-5-thia-1,3,6-triazafluoren-4-one (1b). 4-Dimethy-
lamino-5-(dimethylaminomethyleneamino)thieno[8,9-b]pyridine-
6-carboxylic acid methyl ester (8, 1.0 g, 3.2 mmol), p-toluene-
sulfonic acid (25 mg, 0.13 mmol), and p-toluidine (512 µL, 4.8
mmol) were placed in a flask with toluene (25 mL) and then
heated to 130 °C overnight. The mixture was cooled down to
room temperature. Solvent was removed under vacuum, and
the residue was treated with cold methanol and sonicated.
White precipitate was formed. The product was then collected,
washed with cold methanol (2 × 2 mL), and dried under
vacuum to give a pure product (451 mg, 42%).
1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.43 ppm (d, J )
5.9 Hz, 1H), 8.29 (s, 1H), 7.37 (d, J ) 8.4 Hz, 2H), 7.34 (d, J
) 8.4 Hz, 2H), 6.84 (d, J ) 5.9 Hz, 1H), 3.26 (s, 6H), 2.46 (s,
3H). MS (ESI/NH3): m/z ) 337.0 (M + H+). Anal. (C18H16N4-
OS‚0.2H2O) C, H, N.
9-Dimethylamino-3-(o-hydroxyphenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1c): prepared using the same procedure

7382 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23
Zheng et al.
as for 1b, but employing o-hydroxyaniline as the amine source
(15% yield). 1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.41 ppm
(d, J ) 5.9 Hz, 1H), 8.27 (s, 1H), 7.39 (m, 1H), 7.28 (m, 1H),
7.11 (m, 1H), 7.05 (m, 1H), 6.83 (d, J ) 5.9 Hz, 1H), 3.26 (s,
6H). MS (ESI, M+1): 338.9 (M + H+). Anal. (C17H14N4O2S‚
0.3H2O) C, H, N.
9-Dimethylamino-3-(p-fluorophenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1d): prepared using the same procedure
as for 1b, but employing p-fluoroaniline as the amine source
(81% yield). 1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.43 ppm
(d, J ) 5.9 Hz, 1H), 8.28 (s, 1H), 7.46 (m, 2H), 7.27 (m, 2H),
7.36 (m, 2H), 6.84 (d, J ) 5.9 Hz, 1H), 3.25 (s, 6H). MS (ESI/
NH3): 340.9 (M + H+). Anal. (C17H13FN4OS) C, H, N.
9-Dimethylamino-3-(p-chlorophenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1e): prepared using the same procedure
as for 1b, but employing p-chloroaniline as the amine source
(72% yield). 1H NMR (CDCl3, 300 MHz): δ ) 8.43 ppm (d, J )
5.9 Hz, 1H), 8.27 (s, 1H), 7.55 (m, 2H), 7.42 (m, 2H), 6.84 (d,
J ) 5.9 Hz, 1H), 3.24 (s, 6H). MS (ESI/NH3): 356.9 (M + H+).
Anal. (C17H13ClN4OS) C, H, N.
7.40 (m, 1H), 7.26 (m, 1H), 6.87 (d, J ) 6.6 Hz, 1H), 2.52 (m,
2H), 2.48 (s, 6H), 1.17 (t, J ) 7.5 Hz, 3H). MS (ESI/NH3): m/z
) 351.0 (M + H+). Anal. (C19H18N4OS) C, H, N.
9-Dimethylamino-3-(3-ethylphenyl)-3H-5-thia-1,3,6-tri-
azafluoren-4-one (1m): prepared using the same procedure
as for 1b, but employing m-ethylaniline as the amine source
(41% yield). 1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.44 ppm
(d, J ) 6.2 Hz, 1H), 8.30 (s, 1H), 7.48 (t, J ) 7.8 Hz, 1H), 7.37
(d, J ) 7.8 Hz, 1H), 7.26 (m, 2H), 6.84 (d, J ) 6.2 Hz, 1H),
2.76 (q, J ) 7.8 Hz, 2H), 2.28 (s, 3H), 1.30 (t, J ) 7.8 Hz, 3H),
1.17 (t, J ) 7.5 Hz, 3H). MS (ESI/NH3): m/z ) 351.0 (M +
H+). Anal. (C19H18N4OS) C, H, N.
9-Dimethylamino-3-(4-ethylphenyl)-3H-5-thia-1,3,6-tri-
azafluoren-4-one (1n): prepared using the same procedure
as for 1b, but employing p-ethylaniline as the amine source
(73% yield). 1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.43 ppm
(d, J ) 5.9 Hz, 1H), 8.29 (s, 1H), 7.40 (d, J ) 8.4 Hz, 2H), 7.36
(d, J ) 8.4 Hz, 2H), 6.84 (d, J ) 5.9 Hz, 1H), 3.27 (s, 6H), 2.76
(q, J ) 7.8 Hz, 2H), 1.30 (t, 7.8 Hz, 3H). MS (ESI/NH3): m/z
) 351.0 (M + H+). Anal. (C19H18N4OS) C, H, N.
9-Dimethylamino-3-(p-bromophenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1f): prepared using the same procedure
as for 1b, but employing p-bromoaniline as the amine source
(62% yield). 1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.43 ppm
(d, J ) 5.9 Hz, 1H), 8.26 (s, 1H), 7.72 (d, J ) 8.7 Hz, 2H), 7.36
(d, J ) 8.7 Hz, 2H), 6.84 (d, J ) 5.9 Hz, 1H), 3.26 (s, 6H). MS
(ESI/NH3): 400.0, 402.8 (M + H+). Anal. (C17H13BrN4OS) C,
H, N.
9-Dimethylamino-3-(p-trifluoromethylphenyl)-3H-5-
thia-1,3,6-triazafluoren-4-one (1g): prepared using the same
procedure as for 1b, but employing p-trifluoromethylaniline
as the amine source (21% yield). 1H NMR (CDCl3/MeOD, 300
MHz): δ ) 8.45 ppm (s, br, 1H), 8.29 (s, 1H), 7.87 (d, J ) 8.4
Hz, 2H), 7.64 (d, J ) 8.4 Hz, 2H), 6.85 (d, J ) 5.3 Hz, 1H),
3.23 (s, 6H). MS (ESI/NH3): 390.9 (M + H+). Anal. (C18H13F3N4-
OS) C, H, N.
9-Dimethylamino-3-(2,4-dimethylphenyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (1h): prepared using the same
procedure as for 1b, but employing 2,4-dimethylaniline as the
amine source (85% yield). 1H NMR (CDCl3/MeOD, 300 MHz):
δ ) 8.44 ppm (d, J ) 5.9 Hz, 1H), 8.17 (s, 1H), 7.23 (s, 1H),
7.18 (d, J ) 8.1 Hz, 1H), 7.15 (d, J ) 8.1 Hz, 1H), 6.85 (d, J )
5.9 Hz, 1H), 3.28 (s, 6H), 2.42 (s, 3H), 2.17 (s, 3H). MS (ESI/
NH3): 351.0 (M + H+). Anal. (C19H18N4OS) C, H, N.
9-Dimethylamino-3-(2,4-dichlorophenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1i): prepared using the same procedure
as for 1b, but employing 2,4-dichloroaniline as the amine
source (77% yield). 1H NMR (CDCl3/MeOD, 300 MHz): δ )
8.42 ppm (d, J ) 6.2 Hz, 1H), 8.12 (s, 1H), 7.67 (d, J ) 2.2 Hz,
1H), 7.48 (dd, J ) 8.7, 2.2 Hz, 1H), 7.44 (d, J ) 8.7 Hz, 1H),
6.87 (d, J ) 6.2 Hz, 1H), 3.33 (s, 6H). MS (ESI/NH3): 390.9
(M + H+). Anal. (C17H12Cl2N4OS‚0.1H2O) C, H, N.
9-Dimethylamino-3-benzyl-3H-5-thia-1,3,6-triazafluo-
ren-4-one (1o): prepared using the same procedure as for 1b,
but employing benzylamine as the amine source (65% yield).
1H NMR (CDCl3, 300 MHz): δ ) 8.34 ppm (d, J ) 6.6 Hz,
1H), 8.33 (s, 1H), 7.38 (m, 3H), 7.36 (m, 2H), 6.86 (d, J ) 6.6
Hz, 1H), 5.28 (s, 2H), 3.34 (s, 6H). MS (ESI/NH3): m/z ) 337.0
(M + H+). Anal. (C18H16N4OS‚0.1H2O) C, H, N.
9-Dimethylamino-3-(4-n-propylphenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1p): prepared using the same procedure
as for 1b, but employing a mixture of 4-n-propylaniline as the
1
amine source (63% yield). H NMR (DMSO-d6, 300 MHz): δ
) 8.58 ppm (s, 1H), 8.41 (d, J ) 6.1 Hz, 1H), 7.48 (m, 2H),
7.40 (m, 2H), 6.97 (d, J ) 6.1 Hz, 1H), 3.16 (s, 6H), 2.66 (t, J
) 7.5 Hz, 2H), 1.65 (m, 2H), 0.95 (t, J ) 7.1 Hz, 3H). MS (ESI/
NH3): m/z ) 365.0 (M + H+). Anal. (C20H20N4OS) C, H, N.
9-Dimethylamino-3-(4-isopropylphenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1q): prepared using the same procedure
as for 1b, but employing a mixture of 4-isopropylaniline as
the amine source (58% yield). 1H NMR (DMSO-d6, 300 MHz):
δ ) 8.59 ppm (s, 1H), 8.40 (d, J ) 5.8 Hz, 1H), 7.49 (m, 2H),
7.46 (m, 2H), 6.97 (d, J ) 5.8 Hz, 1H), 3.16 (s, 6H), 3.01 (m,
1H), 1.27 (d, J ) 7.1 Hz, 6H). MS (ESI/NH3): m/z ) 365.1 (M
+ H+). Anal. (C20H20N4OS‚0.1H2O) C, H, N.
9-Dimethylamino-3-(4-tert-butylphenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1r): prepared using the same procedure
as for 1b, but employing a mixture of 4-tert-butylaniline as
the amine source (66% yield). 1H NMR (DMSO-d6, 300 MHz):
δ ) 8.59 ppm (s, 1H), 8.40 (d, J ) 5.8 Hz, 1H), 7.62 (m, 2H),
7.50 (m, 2H), 6.97 (d, J ) 5.8 Hz, 1H), 3.15 (s, 6H), 1.35 (s,
9H). MS (ESI/NH3): m/z ) 379.1 (M + H+). Anal. (C21H22N4-
OS) C, H, N.
9-Dimethylamino-3-(4-cyclopropylphenyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (1s): prepared using the same
procedure as for 1b, but employing a mixture of 4-cyclopro-
pylaniline as the amine source (55% yield). 1H NMR (DMSO-
d6, 300 MHz): δ ) 8.55 ppm (s, 1H), 8.38 (d, J ) 5.8 Hz, 1H),
7.44 (m, 2H), 7.26 (m, 2H), 6.94 (d, J ) 5.8 Hz, 1H), 3.13 (s,
6H), 2.02 (m, 1H), 1.03 (m, 2H), 0.76 (m, 2H). MS (ESI/NH3):
m/z ) 363.1 (M + H+). Anal. (C20H18N4OS) C, H, N.
9-Dimethylamino-3-(4-cyclohexylphenyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (1t): prepared using the same
procedure as for 1b, but employing a mixture of 4-cyclohexy-
laniline as the amine source (64% yield). 1H NMR (DMSO-d6,
300 MHz): δ ) 8.59 ppm (s, 1H), 8.41 (d, J ) 6.1 Hz, 1H),
7.46 (m, 2H), 7.42 (m, 2H), 6.98 (d, J ) 6.1 Hz, 1H), 3.17 (s,
6H), 2.62 (m, 1H), 1.20-1.90 (m, 10H), 0.76 (m, 2H). MS (ESI/
NH3): m/z ) 405.1 (M + H+). Anal. (C23H24N4OS‚0.1H2O) C,
H, N.
9-Dimethylamino-3-(2′-methoxy-5′-pyridyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (1j): prepared using the same
procedure as for 1b, but employing 5-amino-2-methoxypyridine
1
as the amine source (57% yield). H NMR (CDCl3, 300 MHz):
δ ) 8.44 ppm (d, J ) 5.8 Hz, 1H), 8.25 (s, 1H), 8.23 (d, J ) 2.7
Hz, 1H), 7.75 (dd, J ) 8.8, 2.7 Hz, 1H), 6.92 (d, J ) 8.8 Hz,
1H), 6.81 (d, J ) 5.8 Hz, 1H), 4.01 (s, 3H), 3.18 (s, 6H). MS
(ESI/NH3): m/z ) 354.0 (M + H+). Anal. (C17H15N5O2S) C, H,
N.
9-Dimethylamino-3-(1H-indazol-6-yl)-3H-5-thia-1,3,6-
triazafluoren-4-one (1k): prepared using the same procedure
as for 1b, but employing 6-amino-1H-indazole as the amine
source (65% yield). 1H NMR (DMSO-d6, 300 MHz): δ ) 13.42
ppm (s, 1H), 8.65 (s, 1H), 8.40 (d, J ) 5.8 Hz, 1H), 8.22 (s,
1H), 7.95 (d, J ) 8.5 Hz, 1H), 7.82 (s, 1H), 7.28 (dd, J ) 8.5,
1.7 Hz, 1H), 6.95 (d, J ) 5.8 Hz, 1H), 3.13 (s, 6H). MS (ESI/
NH3): m/z ) 363.0 (M + H+). Anal. (C18H14N6OS) C, H, N.
9-Dimethylamino-3-(2-ethylphenyl)-3H-5-thia-1,3,6-tri-
azafluoren-4-one (1l): prepared using the same procedure
as for 1b, but employing o-ethylaniline as the amine source
(51% yield). 1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.45 ppm
(d, J ) 6.6 Hz, 1H), 8.19 (s, 1H), 7.52 (m, 1H), 7.48 (m, 1H),
9-Dimethylamino-3-cyclopropyl-3H-5-thia-1,3,6-triaz-
afluoren-4-one (1u): prepared using the same procedure as
for 1b, but employing cyclopropylamine as the amine source
1
(21% yield). H NMR (CDCl3, 300 MHz): δ ) 8.40 ppm (d, J
) 6.2 Hz, 1H), 8.23 (s, 1H), 6.84 (d, J ) 6.2 Hz, 1H), 3.35 (m,

Triazafluorenone Derivatives as mGluR1 Antagonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23 7383
1H), 3.27 (s, 6H), 1.20-1.40 (m, 4H). MS (ESI/NH3): m/z )
287.0 (M + H+). Anal. (C14H14N4OS‚0.1H2O) C, H, N.
(d, J ) 6.6 Hz, 1H), 8.41 (s, 1H), 6.82 (d, J ) 6.6 Hz, 1H), 3.30
(s, 6H), 2.49 (m, 6H), 2.30 (m, 3H), 1.81 (m, 6H). MS (ESI/
NH3): m/z ) 381.0 (M + H+). Anal. (C21H24N4OS‚0.1H2O) C,
H, N.
9-Dimethylamino-3-cyclobutyl-3H-5-thia-1,3,6-triaz-
afluoren-4-one (1v): prepared using the same procedure as
for 1b, but employing cyclobutylamine the amine source (51%
9-Dimethylamino-3-(3-n-pentyl)-3H-5-thia-1,3,6-triaz-
afluoren-4-one (1af): prepared using the same procedure as
for 1b, but employing 3-n-pentylamine as the amine source
1
yield). H NMR (CDCl3, 300 MHz): δ ) 8.42 ppm (d, J ) 6.2
Hz, 1H), 8.34 (s, 1H), 6.82 (d, J ) 6.2 Hz, 1H), 5.12 (m, 1H),
3.31 (s, 6H), 2.64 (m, 2H), 2.42 (m, 2H), 1.98 (m, 2H). MS (ESI/
NH3): m/z ) 300.7. Anal. (C15H16N4OS) C, H, N.
1
(35% yield). H NMR (CDCl3, 300 MHz): δ ) 8.42 ppm (d, J
) 6.9 Hz, 1H), 8.16 (s, 1H), 6.87 (d, J ) 6.9 Hz, 1H), 4.82 (m,
1H), 3.36 (s, 6H), 1.95 (m, 2H), 1.85 (m, 2H), 0.92 (t, J ) 7.5
Hz, 6H). MS (ESI/NH3): m/z ) 317.0 (M + H+). Anal.
(C16H20N4OS‚0.1H2O) C, H, N.
9-Dimethylamino-3-cyclopentyl-3H-5-thia-1,3,6-triaz-
afluoren-4-one (1w): prepared using the same procedure as
for 1b, but employing cyclopentylamine as the amine source
1
(24% yield). H NMR (CDCl3, 300 MHz): δ ) 8.39 ppm (d, J
9-Dimethylamino-3-cyclopentylmethyl-3H-5-thia-1,3,6-
triazafluoren-4-one (1ag): prepared using the same proce-
dure as for 1b, but employing cyclopentylmethylamine as the
amine source (54% yield). 1H NMR (CDCl3, 300 MHz): δ )
8.40 ppm (d, J ) 6.9 Hz, 1H), 8.23 (s, 1H), 6.86 (d, J ) 6.9 Hz,
1H), 4.04 (d, J ) 7.5 Hz, 2H), 3.35 (s, 6H), 2.45 (m, 1H), 1.30-
1.8 (m, 8H). MS (ESI/NH3): m/z ) 329.0 (M + H+). Anal.
(C17H20N4OS‚0.2H2O) C, H, N.
9-Dimethylamino-3-cyclohexylmethyl-3H-5-thia-1,3,6-
triazafluoren-4-one (1ah): prepared using the same proce-
dure as for 1b, but employing cyclohexylmethylamine as the
amine source (15% yield). 1H NMR (CDCl3, 300 MHz): δ )
8.40 ppm (d, J ) 6.2 Hz, 1H), 8.16 (s, 1H), 6.85 (d, J ) 6.2 Hz,
1H), 3.92 (d, J ) 7.5 Hz, 2H), 3.30 (s, 6H), 1.00-2.00 (m, 11H).
MS (ESI/NH3): m/z ) 343.1 (M + H+). Anal. (C18H22N4OS‚
0.1H2O) C, H, N.
5-Amino-4-dimethylaminofuro[8,9-b]pyridine-6-car-
boxylic Acid Methyl Ester (7). To a solution of methyl
glycolate (2.7 mL, 35 mmol) in anhydrous THF (100 mL),
sodium hydride (60%, 2.8 g, 70 mmol) was added in small
portions at 0 °C. A solution of 4-dimethylamino-2-chloro-3-
cyanopyridine (5, 5.0 g, 27.6 mmol) in THF (25 mL) was added.
The reaction mixture was allowed to stir for 3 days at room
temperature. The reaction mixture was then heated to reflux
for 2 h. After being cooled down to room temperature, the
reaction mixture was quenched with saturated NH4Cl. Organic
solvent was removed under vacuum, and the aqueous layer
was extracted with dichloromethane (500 mL). The organic
layer was separated and washed with water and brine, then
dried over Na2SO4. The solvent was concentrated, and purified
via column chromatography (SiO2, ethyl acetate:hexanes ) 1:9
then 1:4) to give 4.9 g product (76%).
) 6.9 Hz, 1H), 8.30 (s, 1H), 6.86 (d, J ) 6.9 Hz, 1H), 5.22 (m,
1H), 3.37 (s, 6H), 2.30 (m, 2H), 1.95 (m, 2H), 1.84 (m, 4H). MS
(ESI/NH3): m/z ) 315.0 (M + H+). Anal. (C16H18N4OS) C, H,
N.
9-Dimethylamino-3-cyclohexyl-3H-5-thia-1,3,6-triaz-
afluoren-4-one (1x): prepared using the same procedure as
for 1b, but employing cyclohexylamine as the amine source
1
(16% yield). H NMR (CDCl3, 300 MHz): δ ) 8.39 ppm (d, J
) 6.6 Hz, 1H), 8.30 (s, 1H), 6.85 (d, J ) 6.6 Hz, 1H), 4.84 (m,
1H), 3.34 (s, 6H), 1.30-2.10 (m, 10H). MS (ESI/NH3): m/z )
329.0 (M + H+). Anal. (C17H20N4OS‚0.3H2O) C, H, N.
9-Dimethylamino-3-(2-methylcyclohexyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (1y): prepared using the same
procedure as for 1b, but employing a mixture of cis and trans
2-methylcyclohexylamine as the amine source (27% yield). 1H
NMR (CDCl3/MeOD, 300 MHz): δ ) 8.42 ppm (m, 1H), 8.18
(m, 1H), 6.80 (m, 1H), 3.50 (m, 1H), 2.92 (s, 6H), 2.80 (m, 1H),
1.00-2.00 (m, 8H), 0.80 (m, 3H). MS (ESI/NH3): m/z ) 343.1
(M + H+). Anal. (C18H22N4OS‚0.1H2O) C, H, N.
9-Dimethylamino-3-(2-methylcyclohexyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (1z): prepared using the same
procedure as for 1b, but employing a mixture of cis and trans
3-methylcyclohexylamine as the amine source (33% yield). 1H
NMR (CDCl3/MeOD, 300 MHz): δ ) 8.22 ppm (m, 1H), 8.20
(m, 1H), 6.78 (m, 1H), 5.00 (m, 1H), 3.22 (m, 6H), 3.02 (m,
1H), 1.00-2.00 (m, 8H), 0.90 (m, 3H). MS (ESI/NH3): m/z )
343.1 (M + H+). Anal. (C18H22N4OS) C, H, N.
9-Dimethylamino-3-(4-methylcyclohexyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (1aa): prepared using the same
procedure as for 1b, but employing a mixture of cis and trans
4-methylcyclohexylamine as the amine source (27% yield). 1H
NMR (CDCl3/MeOD, 300 MHz): δ ) 8.25 ppm (m, 1H), 8.20
(m, 1H), 6.75 (m, 1H), 4.42 (m, 1H), 3.30 (m, 6H), 1.00-2.00
(m, 9H), 0.90 (m, 3H). MS (ESI/NH3): m/z ) 343.1 (M + H+).
Anal. (C18H22N4OS‚0.2H2O) C, H, N.
1H NMR (CDCl3, 300 MHz): δ ) 8.22 ppm (d, J ) 5.8 Hz,
1H), 7.27 (d, J ) 5.8 Hz, 1H), 5.27 (s, br, 2H), 3.94 (s, 3H),
3.01 (s, 6H). MS (ESI/NH3): m/z ) 236.0 (M + H+).
4-Dimethylamino-5-(dimethylaminomethyleneamino)-
furo[8,9-b]pyridine-6-carboxylic Acid Methyl Ester (9).
5-Amino-4-dimethylaminofuro[8,9-b]pyridine-6-carboxylic acid
methyl ester (7, 2.0 g, 8.5 mmol) was dissolved in ethanol (12
mL) and N,N-dimethylformamide dimethyl acetal (6 mL), and
heated to reflux for 4.5 h. Excess of solvent and reagent were
removed under vacuum to give a yellow solid product (2.5 g,
100%).
9-Dimethylamino-3-cycoheptyl-3H-5-thia-1,3,6-triaz-
afluoren-4-one (1ab): prepared using the same procedure as
for 1b, but employing cycloheptylamine as the amine source
1
(54% yield). H NMR (CDCl3, 300 MHz): δ ) 8.40 ppm (d, J
) 6.6 Hz, 1H), 8.29 (s, 1H), 6.85 (d, J ) 6.6 Hz, 1H), 4.96 (m,
1H), 3.35 (s, 6H), 1.50-2.10 (m, 12H). MS (ESI/NH3): m/z )
343.0 (M + H+). Anal. (C18H22N4OS‚0.2H2O) C, H, N.
9-Dimethylamino-3-(N-hexamethyleneiminyl)-3H-5-
thia-1,3,6-triazafluoren-4-one (1ac): prepared using the
same procedure as for 1b, but employing 1-aminohomopiperi-
dine as the amine source (45% yield). 1H NMR (CDCl3/MeOD,
300 MHz): δ ) 8.40 ppm (d, J ) 5.8 Hz, 1H), 8.37 (s, 1H),
6.78 (d, J ) 5.8 Hz, 1H), 3.88 (m, 4H), 3.15 (s, 6H), 1.78 (m,
8H). MS (ESI/NH3): m/z ) 344.1 (M + H+). Anal. (C17H21N5-
OS‚0.1H2O) C, H, N. 9-Dimethylamino-3-cyclooctyl-3H-5-
thia-1,3,6-triazafluoren-4-one (1ad): prepared using the
same procedure as for 1b, but employing cyclooctylamine as
the amine source (48% yield). 1H NMR (DMSO-d6, 300 MHz):
δ ) 8.69 ppm (s, 1H), 8.39 (d, J ) 6.1 Hz, 1H), 6.97 (d, J ) 6.1
Hz, 1H), 4.90 (m, 1H), 3.18 (s, 6H), 1.50-2.15 (m, 14H). MS
(ESI/NH3): m/z ) 357.1 (M + H+). Anal. (C19H24N4OS‚0.1H2O)
C, H, N.
1H NMR (CDCl3, 300 MHz): δ ) 8.12 ppm (d, J ) 5.8 Hz,
1H), 7.68 (s, 1H), 6.44 (d, J ) 5.8 Hz, 1H), 3.87 (s, 3H), 3.16
(s, 6H), 3.14 (s, br, 3H), 3.09 (s, br, 3H). MS (ESI/NH3): m/z )
291.1 (M + H+).
9-Dimethylamino-3-(4′-methyl-1′-cyclohexyl)-3H-1,3,6-
triazafluoren-4-one (10b). Dimethylamino-5-(dimethylami-
nomethyleneamino)furo[8,9-b]pyridine-6-carboxylic acid meth-
yl ester (9, 150 mg, 0.52 mmol), p-toluenesulfonic acid (10 mg,
0.05 mmol), and 4-methylcyclohexylamine (88 mg, 0.78 mmol)
were placed in a flask with toluene (10 mL) and then heated
to 130 °C for overnight. The mixture was cooled down to room
temperature. Solvent was removed under vacuum, and the
residue was purified via column chromatography (SiO2, ethyl
acetate:hexanes ) 1:9 then 1:4) to give a pure product (75 mg,
44%).
9-Dimethylamino-3-(1-adamantyl)-3H-5-thia-1,3,6-tri-
azafluoren-4-one (1ae): prepared using the same procedure
as for 1b, but employing 1-adamantylamine as the amine
source (55% yield). 1H NMR (CDCl3, 300 MHz): δ ) 8.41 ppm
1H NMR (CDCl3/MeOD, 300 MHz): δ ) 8.19 ppm (m, 1H),
8.16 (m, 1H), 6.50 (m, 5.8 Hz, 1H), 4.90 (m, 1H), 3.42 (m, 6H),
1.50-2.10 (m, 8H), 1.30 (m, 1H), 1.00 (m, 3H). MS (ESI/NH3):
m/z ) 327.1 (M + H+). Anal. (C18H22N4O2) C, H, N.

7384 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23
Zheng et al.
9-Dimethylamino-3-(4′-ethylphenyl)-3H-1,3,6-triazaflu-
oren-4-one (10a): prepared using the same procedure as for
10b, but employing p-ethylaniline as the amine source (18%
9-Dimethylamino-2-ethyl-3-oxa-5-thia-1,6-diazafluoren-
4-one (15b): prepared using the same procedure as for 15a,
but employing propanoic acid anhydride (64% yield). 1H NMR
(300 MHz, CDCl3): δ ) 2.67 ppm (s, 3H), 2.94-3.05 (m, 2H),
3.19 (s, 6H), 6.78 (d, J ) 3.0 Hz, 1H), 8.40 (d, J ) 3.0 Hz, 1H).
MS (DCI/NH3): m/z ) 276.0 (M + H+).
9-Dimethylamino-2-n-butyl-3-oxa-5-thia-1,6-diazafluo-
ren-4-one (15c): prepared using the same procedure as for
15a, but employing n-pentanoic acid anhydride (62% yield).
1H NMR (300 MHz, CDCl3): δ ) 1.00 (t, J ) 11.0 Hz, 2H),
1.28-1.42 (m, 3H), 1.75-2.05 (m, 4H), 3.21 (s, 6H), 6.95 (d, J
) 4.0 Hz, 1H), 8.43 (d, J ) 4.0 Hz, 1H). MS (DCI/NH3): m/z )
316.0 (M + H+).
1
yield). H NMR (CDCl3, 300 MHz): δ ) 8.21 ppm (d, J ) 6.1
Hz, 1H), 8.15 (s, 1H), 7.37 (m, 4H), 6.54 (d, J ) 6.1 Hz, 1H),
3.43 (s, 6H), 2.75 (q, J ) 7.8 Hz, 2H), 1.30 (t, J ) 7.8 Hz, 3H).
MS (ESI/NH3): m/z ) 335.0 (M + H+). Anal. (C19H18N4O2) C,
H, N.
2-(3-Cyano-4-dimethylaminopyridin-2-ylsulfanyl)ace-
tamide (11). 2-Mercapto acetamide (in methanolic ammonia)
(1.0 g, 10.0 mmol) in methanol (15 mL) was added sodium
methoxide (2.28 g, 25.0 mmol) and then 2-chloro-4-dimethy-
laminonicotinonitrile (5, 1.6 g, 9.0 mmol). The reaction mixture
was refluxed for 20 h. After cooling to room temperature, the
mixture was evaporated to dryness, water was added to the
residue, and solid was collected and dried in the drying oven
to give the desired product (2.1 g, 96%) as a white solid.
1H NMR (300 MHz, DMSO-d6) δ ) 3.32 ppm (s, 6H), 3.85
(s, 2H), 6.60 (d, J ) 6.4 Hz, 1H), 7.10 (s, 1H), 7.50 (s, 1H),
8.06 (d, J ) 6.4 Hz, 1H). MS (DCI/NH3): m/z ) 237.0 (M +
H+).
5-Amino-4-dimethylaminothieno[8,9-b]pyridine-6-car-
boxylic Acid Amide (12). 2-(3-Cyano-4-dimethylaminopyri-
din-2-ylsulfanyl)acetamide (11, 2.1 g, 9.0 mmol) in THF (20
mL)/methanol (5 mL)/pyridine (0.5 mL) was treated with
potassium t-butoxide (2.3 g, 18.5 mmol), and the reaction
mixture was refluxed for 20 h. After cooling to room temper-
ature, the mixture was evaporated to dryness, water was
added to the residue, and yellow solid was collected and dried
in the drying oven to give the desired product (1.9 g, 98%).
1H NMR (300 MHz, DMSO-d6): δ ) 2.81 ppm (s, 6H), 6.82-
7.21 (m, 5H), 8.38 (d, J ) 5.4 Hz, 1H). MS (DCI/NH3): m/z )
237.0 (M + H+).
General Procedure To Make 9-Dimethylamino-2-
methyl-3H-5-thia-1,3,6-triazafluoren-4-one (16). 9-Di-
methylamino-2-methyl-3H-5-thia-1,3,6-triazafluoren-4-
one (16a). To a round-bottom flask containing AcOH (10 mL)
was added the oxazinone intermediate 15a (1.0 g, 4.0 mmol),
and the reaction mixture was heated at reflux for 2 h. The
reaction mixture was cooled and extracted with CH2Cl2. The
combined organic layers were dried over Na2SO4 and concen-
trated. The mixture was purified via column chromatography
(SiO2, CH2Cl2 to 10%MeOH/CH2Cl2) to give 0.68 g of the
desired product in 49% yield as a white solid/foam.
1H NMR (300 MHz, CDCl3): δ ) 1.30 ppm (t, J ) 9.0 Hz,
3H), 2.37 (s, 3H), 2.75 (dd, J ) 6.0, 12.0 Hz, 2H), 3.20 (s, 6H),
6.87 (d, J ) 6.0 Hz, 1H), 7.18 (d, J ) 9.0 Hz, 2H), 7.39 (d, J )
9.0 Hz, 2H), 8.40 (d, J ) 6.0 Hz, 1H). MS (DCI/NH3): m/z )
365.0 (M + H+). Anal. (C20H20N4OS) C, H, N.
9-Dimethylamino-2-ethyl-3H-5-thia-1,3,6-triazafluoren-
4-one (16b): prepared using the same procedure as for 16a,
but employing intermediate 15b and propanoic acid anhydride
1
(52% yield). H NMR (300 MHz, CDCl3) δ ) 1.35-1.39 ppm
(m, 6H), 2.60 (dd, J ) 10.9, 22.0 Hz, 2H), 2.78-2.84 (m, 2H),
3.24 (s, 6H), 6.78 (d, J ) 5.8 Hz, 1H), 7.18 (d, J ) 8.5 Hz, 2H),
7.37 (d, J ) 8.5 Hz, 2H), 8.40 (d, J ) 5.76 Hz, 1H). MS (DCI/
NH3): m/z ) 378.0 (M + H+). Anal. (C21H22N4OS) C, H, N.
9-Dimethylamino-2-n-butyl-3H-5-thia-1,3,6-triazafluo-
ren-4-one (16c): prepared using the same procedure as for
16a, but employing intermediate 15c and n-pentanoic acid
9-Dimethylamino-3H-5-thia-1,2,3,6-tetraazafluoren-4-
one (13). 5-Amino-4-dimethylaminothieno[8,9-b]pyridine-6-
carboxylic acid amide (12, 1.9 g, 8.0 mmol), in 5N HCl (20 mL)
was cooled to 0 °C and treated dropwise with a solution of
NaNO2 (1.1 g, 16.0 mmol) in water (15 mL). The reaction
mixture was vigorously stirred for 30 min. Then a solution of
5% aq NaHCO3 was added until pH ) 7 was reached,
whereupon a solid precipitated. The solid was collected by
filtration and washed with cold water to give the desired
product (1.8 g, 91% yield) as an off-white solid.
1
anhydride (42% yield). H NMR (300 MHz, CDCl3): δ ) 0.85
ppm (t, J ) 11.2 Hz, 2H), 1.22-1.38 (m, 6H), 1.70-1.82 (m,
2H), 2.55 (t, J ) 11.2 Hz, 2H), 2.75 (dd, J ) 15.3 Hz, 2H), 3.21
(s, 6H), 6.77 (d, J ) 5.8 Hz, 1H), 7.18 (d, J ) 8.5 Hz, 2H), 7.39
(d, J ) 8.5 Hz, 2H), 8.40 (d, J ) 5.8 Hz, 1H). MS (DCI/NH3):
m/z ) 407.0 (M + H+). Anal. (C23H26N4OS) C, H, N.
1-Cyano-2,6-dinitrobenzene (18). A mixture of 2,6-d-
dinitro-1-chlorobenzene (17, 14.7 g, 73 mmol), copper cyanide
(8.0 g, 90.0 mmol), and DMF (75 mL) was heated at 145 °C
under N2 for 2 h. The mixture was cooled to room temperature
and poured into ice water. The solid was filtered off, and then
the solid was extracted with hot EtOAc (2 × 200 mL).
Combined extracts were evaporated. The residue was sus-
pended in 25 mL of hot EtOH, filtered off, and dried to give
the desired 1-cyano-2,6-dinitrobenzene (18, 9.8 g, 70%).
1H NMR (300 MHz, DMSO-d6): δ ) 8.22 ppm (t, J ) 9.0
Hz, 1H), 8.67 (d, J ) 9.0 Hz, 2H). MS (DCI/NH3): m/z 194 (M
+ H)+.
1H NMR (300 MHz, DMSO-d6): δ ) 3.01-3.25 ppm (m, 6H),
7.03 (d, J ) 5.76 Hz, 1H), 8.44 (d, J ) 5.76 Hz, 1H), 15.54 (s,
1H). MS (DCI/NH3): m/z ) 248.0 (M + H+).
3-Cycloheptyl -9-dimethylamino-3H-5-thia-1,2,3,6-tet-
raazafluoren-4-one (14). 9-Dimethylamino-3H-5-thia-1,2,3,6-
tetraazafluoren-4-one (13, 70 mg, 0.28 mmol) in DMF (4 mL)
was treated with NaH 60% disp. in oil (18 mg, 0.4 mmol) and
then with cycloheptyl bromide (0.04 mL, 0.29 mmol). The
reaction mixture was heated at 80 °C for 18 h. After cooling
to room temperature, the mixture was poured into water (25
mL) and extracted with ethyl acetate (30 mL). The organic
layer was washed with water (3 × 25 mL), dried over MgSO4,
and concentrated in vacuo. The residue was purified by
chromatography (silica gel, 1:1 hexane:EtOAc) to provide the
desired product (55 mg, 57% yield).
1H NMR (300 MHz, CDCl3): δ ) 1.47-1.80 ppm (m, 4H),
1.83-2.02 (m, 4H), 2.03-2.46 (m, 4H), 3.23 (s, 6H), 4.99-5.59
(m, 1H), 6.85 (d, J ) 5.8 Hz, 1H) 8.43 (d, J ) 5.8 Hz, 1 H). MS
(DCI/NH3): m/z ) 344.0 (M + H+). Anal. (C17H21N5OS) C, H, N.
General Procedure To Make Oxazinone 15. 9-Di-
methylamino-2-methyl-3-oxa-5-thia-1,6-diazafluoren-4-
one (15a). To a flask containing the aminomethyl ester
intermediate 6 (1.0 g, 4.0 mmol) was added acetic anhydride
(15 mL), and the reaction mixture was refluxed for 2 h. After
cooling the reaction to room temperature and standing for 24
h, the desired material crystallized and was filtered off. The
product was recrystallized from toluene to give white needles
(0.80 g) in 76% yield.
2-Dimethylamino-1-cyano-6-nitrobenzene (19). To the
nitrile intermediate 18 from above (1.0 g, 5.2 mmol) dissolved
in DMF (6 mL) was added 40% aqueous dimethylamine (1.5
mL, 14.0 mmol). The mixture was heated to 50 °C for 10 min
and then cooled to room temperature. The mixture was poured
into ice water. The precipitate formed was filtered off and
vacuum-dried to obtain 0.93 g (95%) of desired product as a
brown color solid.
1H NMR (300 MHz, CDCl3): δ ) 3.15 ppm (s, 6H), 7.22 (m,
2H), 7.55 (m, 2H). MS (DCI,/NH3): m/z ) 191.0 (M + H+).
5-Amino-4-dimethylaminothieno[8,9-b]benzo-6-carbox-
ylic Acid Methyl Ester (20). A mixture of nitrile intermedi-
ate 19 (1.9 g, 10.4 mmol) and methyl thioglycolate (1.2 g, 10.4
mmol) was dissolved in dry DMF (20 mL). The mixture was
cooled to 0 °C. Sodium methoxide (1.2 g, 22.0 mmol) was then
added under N2. The reaction mixture was allowed to warm
1H NMR (300 MHz, CDCl3): δ ) 2.58 ppm (s, 3H), 3.17 (s,
6H), 6.75 (d, J ) 3.0 Hz, 1H), 8.39 (d, J ) 3.0 Hz, 1H). MS
(DCI/NH3): m/z ) 262.0 (M + H+).

Triazafluorenone Derivatives as mGluR1 Antagonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23 7385
up to room temperature and stirred for 16 h, it was poured
into ice water (250 mL), and a yellow precipitate that formed
was filtered off and vacuum-dried to obtain 2.3 g (88%) of the
desired 5-amino-4-dimethylaminothieno[8,9-b]benzo-6-carbox-
ylic acid methyl ester (20) as a yellow amorphous solid.
1H NMR (300 MHz, DMSO-d6): δ ) 2.71 ppm (s, 6H), 3.78
(s, 3H), 7.19 (d, J ) 9.0 Hz, 1H), 7.38 (bs, 2H), 7.44 (t, J ) 9.0
Hz, 1H), 7.54 (d, J ) 9 Hz, 1H). MS (DCI/NH3): m/z 251 (M +
H)+.
9-Dimethylamino-3-(4-ethylphenyl)-3H-5-thia-1,3-diaz-
afluoren-4-one (22). A mixture of intermediate 20 (1.9 g, 7.6
mmol), dimethylformamide dimethylacetal (6.0 mL), and EtOH
(6 mL) was refluxed under N2 for 16 h, cooled to room
temperature, and concentrated under vacuum. The solid
obtained was recrystallized from EtOH-water to obtain 2.1 g
(90%) of the desired 4-dimethylamino-5-(dimethylaminometh-
yleneamino)thieno[8,9-b]benzene-6-carboxylic acid methyl es-
ter (21).
temperature, and concentrated under vacuum. The solid
obtained was recrystallized from EtOH-water to obtain 2.1 g
(90%) of the desired 4-dimethylamino-5-(dimethylaminometh-
yleneamino)-2-aza-thieno[8,9-b]pyridine-6-carboxylic acid meth-
yl ester (27).
1H NMR (300 MHz, DMSO-d6): δ ) 2.81 ppm (s, 6H), 3.04
(s, 6H), 3.76 (s, 3H), 7.52 s, 1H), 8.04 (s, 1H), 8.65 (s, 1H). MS
(DCI/NH3): m/z 307 (M + H)+.
9-Dimethylamino-3-(4-ethylphenyl)-3H-5-thia-1,3,7-tri-
azafluoren-4-one (28a). A mixture of 27 (0.306 g, 1.0 mmol),
4-ethylaniline (0.181 g, 1.5 mmol), p-toluenesulfonic acid (0.02
g, 0.1 mmol), and toluene (7 mL) was refluxed under N2 for
16 h. The reaction mixture was cooled to room temperature
and concentrated under vacuum, and the residue was purified
by flash column chromatography (silica gel, 1:1 hexane:EtOAc)
to give 0.11 g (38%) of the desired pyrimidone as a yellow color
solid.
1H NMR (300 MHz, DMSO-d6): δ )1.24 ppm (t, J ) 9.0
Hz, 3H), 2.71 (q, J ) 9.0 Hz, 2H), 3.01 (s, 6H), 7.42 (d, J ) 9.0
Hz, 2H), 7.50 (d, J ) 9 Hz, 2H), 8.27 (s, 1H), 8.63 (s, 1H), 8.96
(s, 1H). MS (DCI/NH3): m/z ) 351.0 (M + H+). Anal. (C19H18N4-
OS) C, H, N.
9-Dimethylamino-3-(4-methylphenyl)-3H-5-thia-1,3,7-
triazafluoren-4-one (28b): prepared using the same proce-
dure as for 28a, but employing p-toluidine as the amine source
(58% yield). 1H NMR (300 MHz, DMSO-d6): δ ) 2.42 ppm (s,
3H), 3.02 (s, 6H), 7.40 (d, J ) 9.0 Hz, 2H), 7.46 (d, J ) 9.0 Hz,
2H), 8.27 (s, 1H), 8.63 (s, 1H), 8.97 (s, 1H). MS (DCI/NH3):
m/z ) 337.0 (M + H+). Anal. (C18H16N4OS) C, H, N.
2-(3-Dimethylamino-1-ethoxyallylidene)malononi-
trile (30). 2-(1-Ethoxyethylidene)malononitrile (29, 110.0 g,
808 mmol) was dissolved in N,N-dimethylformamide dimethyl
acetal (94%, 230 mL, 1.62 mol), and the reaction mixture was
heated to reflux at 100 °C for 1 h. The mixture was cooled
down to room temperature. Solid was collected and washed
with cold methanol to give an orange solid product. The mother
liquor was concentrated, and the solid was collected again, and
washed with cold methanol. This procedure was repeated a
couple of times, and all the solid products were combined. The
final residue was purified by a short column chromatography
(SiO2, ethyl acetate). The combined yield of the reaction is 78%
(120.0 g).
A mixture of 4-dimethylamino-5-(dimethylaminomethylene-
amino)thieno[8,9-b]benzene-6-carboxylic acid methyl ester (21,
0.306 g, 1.0 mmol), 4-ethylaniline (0.181 g, 1.5 mmol), p-
toluenesulfonic acid (0.02 g, 0.1 mmol), and toluene (7 mL)
was refluxed under N2 for 16 h. The reaction mixture was
cooled to room temperature and concentrated under vacuum,
and the residue was purified by flash column chromatography
(SiO2, 1:1 hexane:EtOAc) to give 0.11 g (13%) of the desired
diazafluorenone as yellow color solid.
1H NMR (300 MHz, DMSO-d6): δ ) 1.25 ppm (t, J ) 9 Hz,
3H), 2.72 (q, J ) 9.0 Hz, 2H), 2.95 (s, 6H), 7.07 (d, J ) 9.0 Hz,
1H), 7.51 (m, 5H), 7.65 (d, J ) 9 Hz, 1H), 8.58 (s, 1H). MS
(DCI/NH3): m/z ) 350.0 (M + H+). Anal. (C20H19N3OS‚0.1H2O)
C, H, N.
3,5-Dichloroisonicotinonitrile (24). A mixture of 3,5-
dichloro-4-pyridinecarboxaldehyde (23, 10.0 g, 57.1 mmol),
hydroxylamine hydrochloride (5.25 g 76 mmol), formic acid (50
mL) and concentrated H2SO4 (5 drops) was refluxed under N2
for 6 h. The mixture was concentrated under vacuum. The
solids were taken in Et2O (250 mL), washed with NaHCO3 and
brine, and the organic layer was dried over MgSO4, filtered,
concentrated, and recrystallized from hexane to give 8.2 g
(83%) of the desired nitrile 24 (mp 114 °C).
1H NMR (300 MHz, CDCl3): δ ) 8.70 ppm (s, 2H). MS (DCI/
NH3): m/z 173 (M + H)+.
1H NMR (CDCl3, 300 MHz): δ ) 7.47 ppm (d, J ) 12.6 Hz,
1H); 5.16 (d, J ) 12.6 Hz, 1H); 4.43 (q, J ) 7.1 Hz, 2H), 3.19
(s, br. 3H), 2.94 (s, br 3H), 1.41 (t, J ) 7.1 Hz, 3H). MS (ESI/
NH3): m/z ) 192.1 (M + H+).
3-Chloro-5-dimethylaminoisonicotinonitrile (25). The
nitrile intermediate 24 from above (3.0 g, 17.2 mmol) was
dissolved in DMF (25 mL). The mixture was cooled to 0 °C.
To this mixture was added 40% aqueous dimethylamine (7.0
mL, 35 mmol). The mixture was allowed to warm up room
temperature, stirred at that temperature for 4 h, and then
poured into ice water. The precipitate that formed was filtered
off and vacuum-dried to obtain 2.3 g (75%) of the desired
3-chloro-4-cyano-5-dimethylaminopyridine (25) as a beige color
solid (mp114 °C).
2-Chloro-3-cyano-4-ethoxypyridine (31). To a slurry of
2-(3-dimethylamino-1-ethoxyallylidene)malononitrile (30, 85.0
g, 445.0 mmol) in methanol (1200 mL), HCl gas was introduced
gently at 0 °C. The reaction mixture became homogeneous in
about 1 h, and was allowed to stir under constant HCl flow
for additional 14.5 h at 0 °C. N2 was bubbled though the
reaction mixture overnight, and all the solvent was removed.
The residue solid was redissolved in CH2Cl2 and washed with
water/K2CO3/water. The organic layer was separated and dried
over Na2SO4. Removal of salt and solvent gave a pure product
(113.0 g, 98%).
1H NMR (300 MHz, CDCl3): δ ) 3.11 ppm (s, 6H), 8.05 (s,
1H), 8.10 (s, 1H); MS (DCI/NH3) m/z 182 (M + H)+.
3-Amino-4-dimethylaminothieno[2,3-c]pyridine-2-car-
boxylic Acid Methyl Ester (26). A mixture of 25 (1.9 g, 10.4
mmol) and methyl thioglycolate (1.2 g, 10.4 mmol) was
dissolved in dry DMF (20 mL). The mixture was cooled to 0
°C. Sodium methoxide (1.2 g, 22 mmol) was then added under
N2. The reaction mixture was allowed to warm up to room
temperature and allowed to stir for 16 h, it was poured into
ice water (250 mL), and a yellow precipitate that formed was
filtered off and vacuum-dried to obtain 2.3 g (88%) of 5-amino-
4-dimethylamino-2-aza-thieno[8,9-b]pyridine-6-carboxylic acid
methyl ester (26) as a yellow amorphous solid.
1H NMR (CDCl3, 300 MHz): δ ) 7.96 ppm (d, J ) 6.4 Hz,
1H); 6.59 (d, J ) 6.4 Hz, 1H); 3.30 (s, 6H). MS (ESI/NH3): m/z
) 181.9 (M + H+).
2-Bromo-3-cyano-4-hydroxypyridine (32). A mixture of
2-chloro-3-cyano-4-ethoxypyridine (31, 8.5 g, 46.7 mmol) and
HBr in acetic acid (30%, 85 mL) was heated to 100 °C for 2 h.
The mixture was cooled down to room temperature, and solid
was collected, washed with cold water, and dried under
vacuum to give a white solid with 91.6% bromonation product,
and 8.4% chlorination product.
1H NMR (300 MHz, DMSO-d6): δ ) 2.80 ppm (s, 6H), 3.81
(s, 3H), 7.18 (bs, 2H), 8.35 (s, 1H), 8.83 (s, 1H). MS (DCI/
NH3): m/z 252 (M + H)+.
4-Dimethylamino-3-(dimethylaminomethyleneamino)-
thieno[2,3-c]pyridine-2-carboxylic Acid Methyl Ester
(27). A mixture of aminocarboxylate intermediate 26 (1.9 g,
7.6 mmol), dimethyl-formamide dimethylacetal (6.0 mL), and
EtOH (6 mL) was refluxed under N2 for 16 h, cooled to room
2-Bromo-3-cyano-4-hydroxypyridine (32a). 1H NMR
(DMSO-d6, 300 MHz): δ ) 8.20 ppm (d, J ) 5.9 Hz, 1H); 6.95
(d, J ) 5.9 Hz, 1H). MS (ESI/NH3): m/z ) 200.9 (M + H+).
2-Chloro-3-cyano-4-hydroxypyridine (32b). 1H NMR
(DMSO-d6, 300 MHz): δ ) 8.27 ppm (d, J ) 6.3 Hz, 1H); 6.98
(d, J ) 6.3 Hz, 1H). MS (ESI/NH3): m/z ) 172.0 (M + H+).

7386 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23
Zheng et al.
2-Bromo-3-cyano-4-(4′-methoxybenzyloxy)pyridine (33).
A mixture of 2-bromo-3-cyano-4-hydroxypyridine (32a) and
2-chloro-3-cyano-4-hydroxypyridine (32b) (10.0 g, 35.7 mmol)
was dissolved in DMF (50 mL), followed by NaH in portions
(60%, 2.86 g, 71.5 mmol). 4-Methoxybenxyl chloride (6.85 mL,
50.3 mmol) was added. The reaction mixture was then heated
to 60 °C for 2.5 h and quenched with NH4Cl (saturated). The
mixture was then extracted with ethyl acetate (5 × 20 mL).
The combined organic layers were washed with water, and
dried over Na2SO4. Column chromatographic purification
(SiO2, ethyl acetate:hexanes ) 1:9) gave an off-white solid
product (4.21 g, 37%) with 92% the title product and 8% of
2-chloro-3-cyano-4-(4′-methoxybenzyloxy)pyridine.
mixture was sonicated. Solid was then collected to give the
desired product (1.61 g, 100%).
1H NMR (DMSO-d6, 300 MHz): δ ) 8.54 ppm (s, 1H), 8.42
(d, J ) 5.9 Hz, 1H), 7.48 (d, J ) 6.6 Hz, 2H), 7.43 (d, J ) 6,6
Hz, 2H), 6.92 (d, J ) 5.9 Hz, 2H), 2.72 (q, J ) 7.5 Hz, 2H),
1.25 (t, J ) 7.5 Hz, 3H). MS (ESI/NH3): m/z ) 324.0 (M +
H+).
9-(Trifluromethanesulfonyl)-3-(4′-ethylphenyl)-3H-5-
thia-1,3,6-triazafluoren-4-one (38). 9-Hydroxy-3-(4′-eth-
ylphenyl)-3H-5-thia-1,3,6-triazafluoren-4-one (37, 1.66 g, 3.82
mmol), N-phenyltrifluoromethanesulfonimide (3.42 g, 9.57
mmol), and N-ethyl-diisopropylamine (1.7 mL, 9.76 mmol)
were dissolved in 1,4-dioxane (80 mL) at room temperature.
The reaction mixture was allowed to stir for 3 days. Solvent
was removed, and the residue was purified via column chro-
matography (SiO2, ethyl acetate:hexanes ) 1:9) to give a white
solid product (1.42 g, 82%).
2-Bromo-3-cyano-4-(4′-methoxybenxyloxy) pyridine
1
(33a). H NMR (CDCl3, 300 MHz): δ ) 8.33 ppm (d, J ) 5.7
Hz, 1H); 7.34 (d, J ) 8.8 Hz, 2H), 6.94 (d, J ) 8.8 Hz, 2H),
5.23 (s, 2H), 3.83 (s, 3H). MS (ESI/NH3): m/z ) 320.0 (M +
H+).
1H NMR (CDCl3, 300 MHz): δ ) 8.88 ppm (d, J ) 5.1 Hz,
1H), 8.35 (s, 1H), 7.39 (m, 5H), 2.74 (q, J ) 7.5 Hz, 2H), 1.31
(t, J ) 7.5 Hz, 3H). MS (ESI/NH3): m/z ) 455.9 (M + H+).
General Procedure for Making Mono and Disubsti-
tuted Triazafluorenone Analogues 39. 9-N-Azetidinyl-3-
(4′-ethylphenyl)-3H-5-thia-1,3,6-triazafluoren-4-one (39c).
9-(Trifluromethanesulfonyl)-3-(4′-ethylphenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (38, 60 mg, 0.13 mmol) was dissolved in
dichloromethane (2 mL). Azetidine hydrochloride (50 mg, 0.52
mmol) was neutralized with sodium hydroxide (1 N, 1 mL, 1.0
mmol) in dichloromethane. The organic layer was separated
and dried over sodium sulfate. The solution of azetidine in
dichloromethane was then filtrated and added to the reaction
mixture. The reaction mixture was then allowed to stir at room
temperature for 1 day. Solvent was removed, and the crude
mixture was purified via a short column chromatography
(SiO2, ethyl acetate:hexanes ) 1:4) to give a white solid product
(42 mg, 89%).
2-Chloro-3-cyano-4-(4′-methoxybenzyloxy)pyridine(33b).
1H NMR (CDCl3, 300 MHz): δ ) 8.36 ppm (d, J ) 5.8 Hz,
1H); 7.31 (d, J ) 8.8 Hz, 2H), 6.89 (d, J ) 8.8 Hz, 2H), 4.63 (s,
2H), 3.82 (s, 3H). MS (ESI/NH3): m/z ) 275.0 (M + H+).
5-Amino-4-(4′-methoxybenzyloxy)thieno[8,9-b]pyridine-
6-carboxylic Acid Methyl Ester (34). 2-Bromo-3-cyano-4-
(4′-methoxybenzyloxy)pyridine (33a, 4.0 g, 12.5 mmol) was
dissolved in DMF (20 mL), followed by methyl thioglycolate
(1.2 mL, 13.2 mmol), and then sodium methoxide (95%, 1.56
g, 27.4 mmol) in portions at room temperature. The reaction
mixture was allowed to stir overnight, and then quenched with
water. Solid was collected and washed with water several
times, then dried under vacuum to give a pure product (3.7 g,
86%).
1H NMR (CDCl3, 300 MHz): δ ) 8.48 ppm (d, J ) 5.8 Hz,
1H), 7.39 (m, 2H), 6.96 (m, 2H), 6.76 (d, J ) 5.8 Hz, 1H), 6.53
(s, br. 1H), 5.19 (s, 2H), 3.85 (s, 3H), 3.84 (s, 3H). MS (ESI/
NH3): m/z ) 344.9 (M + H+).
4-(4′-Methoxybenzyloxy)-5-(dimethylaminomethylene-
amino)thieno[8,9-b]pyridine-6-carboxylic Acid Methyl
Ester (35). 5-Amino-4-(4′-methoxybenzyloxy)thieno[8,9-b]py-
ridine-6-carboxylic acid methyl ester (34, 3.7 g, 10.8 mmol) was
dissolved in N,N-dimethylformamide dimethyl acetal (30 mL),
and heated to reflux for 10 h. Excess of reagent was removed
to give a yellow solid product (4.3 g, 100%).
1H NMR (CDCl3, 300 MHz): δ ) 8.26 ppm (d, J ) 6.1 Hz,
1H), 8.17 (s, 1H), 7.37 (m, 4H), 6.25 (d, J ) 6.1 Hz, 2H), 4.54
(m, 4H), 2.75 (q, J ) 7.8 Hz, 2H), 2.47 (m, 2H), 1.30 (t, J ) 7.8
Hz, 3H). MS (ESI/NH3): m/z ) 363.0 (M + H+). Anal.
(C20H18N4OS) C, H, N.
9-(N,N-Ethylmethylamino)-3-(4′-ethylphenyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (39a): prepared using the same
procedure as for 39c, but employing ethylmethylamine hy-
drochloride as the amine source (92% yield). 1H NMR (CDCl3,
300 MHz): δ ) 8.43 ppm (d, J ) 6.6 Hz, 1H), 8.27 (s, 1H),
7.40 (d, J ) 8.4 Hz, 2H), 7.35 (d, J ) 8.4 Hz, 2H), 6.84 (d, J )
6.6 Hz, 1H), 3.82 (q, J ) 7.2 Hz, 2H), 3.33 (s, 3H), 2.75 (q, J )
7.8 Hz, 2H), 1.38 (t, 7.2 Hz, 3H), 1.30 (t, J ) 7.8 Hz, 3H). MS
(ESI/NH3): m/z ) 365.0 (M + H+). Anal. (C20H20N4OS‚0.1H2O)
C, H, N.
9-(2′-Methoxyethylmethylamino)-3-(4′-ethylphenyl)-
3H-5-thia-1,3,6-triazafluoren-4-one (39b): prepared using
the same procedure as for 39c, but employing 2-methoxyeth-
ylmethylamine as the amine source (78% yield). 1H NMR
(CDCl3, 300 MHz): δ ) 8.42 ppm (d, J ) 5.8 Hz, 1H), 8.26 (s,
1H), 7.37 (m, 4H), 6.85 (d, J ) 5.8 Hz, 1H), 3.91 (t, J ) 5.4
Hz, 2H), 3.68 (t, J ) 5.4 Hz, 2H), 3.27 (s, 3H), 3.21 (s, 3H),
2.75 (q, J ) 7.8 Hz, 2H), 1.30 (t, J ) 7.8 Hz, 3H). MS (ESI/
NH3): m/z ) 395.0 (M + H+). Anal. (C21H22N4O2S‚0.3H2O) C,
H, N.
9-Cyclopropylamino-3-(4′-ethylphenyl)-3H-5-thia-1,3,6-
triazafluoren-4-one (39d): prepared using the same proce-
dure as for 39c, but employing cyclopropylamine as the amine
source (74% yield). 1H NMR (CDCl3, 300 MHz): δ ) 8.67 ppm
(s, 1H), 8.49 (d, J ) 6.6 Hz, 1H), 8.27 (s, 1H), 7.40 (d, J ) 8.0
Hz, 2H), 7.34 (d, J ) 8.0 Hz, 2H), 7.05 (d, J ) 6.6 Hz, 1H),
2.80 (m. 1H), 2.76 (q, J ) 7.8 Hz, 2H), 1.31 (t, J ) 7.8 Hz,
3H), 1.09 (m, 2H), 0.83 (m, 2H). MS (ESI/NH3): m/z ) 363.1
(M + H+). Anal. (C20H18N4OS‚0.1H2O) C, H, N.
1H NMR (CDCl3, 300 MHz): δ ) 8.47 ppm (d, J ) 5.8 Hz,
1H), 7.44 (s, 1H), 7.40 (m, 2H), 6.91 (m, 2H), 6.75 (d, J ) 5.8
Hz, 1H), 5.09 (s, 2H), 3.84 (s, 3H), 3.82 (s, 3H), 2.83 (s, br,
6H). MS (ESI/NH3): m/z ) 400.0 (M + H+).
9-(4′-Methoxybenzyloxy)-3-(4′-ethylphenyl)-3H-5-thia-
1,3,6-triazafluoren-4-one (36). 4-(4′-Methoxybenzyloxy)-5-
(dimethylaminomethyleneamino)thieno[8,9-b]pyridine-6-car-
boxylic acid methyl ester (35, 4.0 g, 10.0 mmol), 4-ethylanaline
(2.42 g, 20.0 mmol), and a catalytic amount of p-toluenesulfonic
acid (200 mg, 1.1 mmol) in toluene (50 mL) were heated under
microwave to 160 °C for 1 h. The reaction mixture was then
cooled, and solvent was removed. The residue was purified via
column chromatography (SiO2, ethyl acetate:hexanes ) 1:4,
then 1:1) to give a solid product, which was recrystallized from
cold methanol (1.7 g, 39%).
1H NMR (DMSO-d6, 300 MHz): δ ) 8.64 ppm (d, J ) 5.5
Hz, 1H), 8.56 (s, 1H), 7.53 (d, J ) 8.6 Hz, 2H), 7.48 (d, J ) 8.3
Hz, 2H), 7.40 (d, J ) 8.3 Hz, 2H), 7.33 (d, J ) 5.5 Hz, 1H),
6.97 (d, J ) 8.6 Hz, 2H), 5.45 (s, 2H), 3.75 (s, 3H), 3.71 (q, J
) 7.4 Hz, 2H), 1.25 (t, J ) 7.4 Hz, 3H). MS (ESI/NH3): m/z )
444.0 (M + H+).
9-Hydroxy-3-(4′-ethylphenyl)-3H-5-thia-1,3,6-triazaflu-
oren-4-one (37). 9-(4′-Methoxybenzyloxy)-3-(4′-ethylphenyl)-
3H-5-thia-1,3,6-triazafluoren-4-one (36, 1.6 g, 3.6 mmol) was
added to cold (0 °C) trifluoroactic acid (10 mL) in portions, and
then allowed to stir for 1.2 h. Excess of TFA was removed
under vacuum. The residue was allowed to sit overnight, then
treated with ethyl acetate (20 mL). Ethyl acetate was removed
and the procedure was repeated five times until a yellow solid
was obtained. Hexanes were added to the solid, and the
9-(1′-Cyanomethylamino)-3-(4′-ethylphenyl)-3H-5-thia-
1,3,6-triaza-fluoren-4-one (39e): prepared using the same
procedure as for 39c, but employing cyanomethylamine as the
amine source (82% yield). 1H NMR (CDCl3, 300 MHz): δ )
8.71 ppm (t, J ) 5.9 Hz, 1H), 8.62 (d, J ) 6.5 Hz, 1H), 8.29 (s,

Triazafluorenone Derivatives as mGluR1 Antagonists
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 23 7387
1H), 7.41 (d, J ) 8.4 Hz, 2H), 7.35 (d, J ) 8.4 Hz, 2H), 6.79 (d,
J ) 6.5 Hz, 1H), 4.47 (d, J ) 5.9 Hz, 2H), 2.76 (q, J ) 7.5 Hz,
2H), 1.31 (t, J ) 7.5 Hz, 3H). MS (ESI/NH3): m/z ) 361.9 (M
+ H+). Anal. (C19H15N5OS‚0.2H2O) C, H, N.
9-(2′,2′,2′-Trifluoroethylamino)-3-(4′-ethylphenyl)-3H-
5-thia-1,3,6-triazafluoren-4-one (39f): prepared using the
same procedure as for 39c, but employing 2,2,2-trifluorethy-
lamine as the amine source (67% yield). 1H NMR (CDCl3, 300
MHz): δ ) 8.94 ppm (t, J ) 6.9 Hz, 1H), 8.55 (d, J ) 6.5 Hz,
1H), 8.32 (s, 1H), 7.97 (s, br. 1H), 7.40 (d, J ) 8.4 Hz, 2H),
7.34 (d, J ) 8.4 Hz, 2H), 6.82 (d, J ) 6.5 Hz, 1H), 4.16 (m,
2H), 2.76 (q, J ) 7.5 Hz, 2H), 1.31 (t, J ) 7.5 Hz, 3H). MS
(ESI/NH3): m/z ) 404.9 (M + H+). Anal. (C19H15F3N4OS) C,
H, N.
9-(Methylamino)-3-(4′-ethylphenyl)-3H-5-thia-1,3,6-tri-
azafluoren-4-one (40). To a solution of 1n (53 mg, 0.15 mmol)
in formic acid (5 mL) at 0 °C was added 30% H2O2 (1 mL),
and the mixture was allowed to warm to room temperature
for 24 h. Solid NaHCO3 was added to pH ) 8, and the mixture
was extracted with ethyl acetate, washed with brine, and dried
with anhydrous MgSO4. The ethyl acetate was removed under
reduced pressure and the residue was chromatographed (SiO2,
hexanes:EtOAc 1:1) to provide 35 mg (70%) of the desired
product.
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1H NMR (300 MHz, DMSO-d6): δ ) 1.24 ppm (t, J ) 7.0
Hz, 3H), 2.70 (q, J ) 7.0 Hz, 2H), 3.03 (d, J ) 4.5 Hz, 3H),
6.66 (d, J ) 6.0 Hz, 1H), 7.41 (d, J ) 9.0 Hz, 2H), 7.48 (d, J )
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8.59 (s, 1H). MS (DCI/NH3): m/z ) 337.0 (M + H+). Anal.
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was continued at room temperature until disappearance of
starting material (∼30 min). A 2 N Solution of Na2CO3 (10
mL) was added, and the layers were separated. The water
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1H NMR (300 MHz, DMSO-d6) δ ) 1.23 ppm (t, J ) 7.0 Hz,
3H), 2.71 (q, J ) 7.0 Hz, 2H), 2.91 (s, 6H), 7.42 (d, J ) 9.0 Hz,
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complex (141 mg, 1.5 mmol) and phthalic anhydride (75 mg,
0.5 mmol) in dichloromethane (4 mL). The resulting reaction
mixture was stirred at room temperature for 4 h. Saturated
K2CO3 (20 mL) and dichloromethane (20 mL) were added to
the reaction mixture. The organic layer was separated and
washed with brine, dried over MgSO4, and then concentrated
in vacuo. The residue was purified by flash chromatography
using 10% methanol in dichloromethane to give the desired
product (105 mg, 58%): 1H NMR (300 MHz, DMSO-d6) δ )
1.32-2.02 ppm (m, 12H) 2.88 (s, 6H) 7.51 (d, J ) 5.8 Hz, 1H),
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359.0 (M + H+). Anal. (C17H21N5O2S‚0.15H2O) C, H, N.
Acknowledgment. We would like to thank Dr.
Bruce W. Surber, Shirish Raja, and Jia Du of Abbott
Laboratories for synthesis and purification of the ra-
diolabeled ligand [3H]R214127.
Supporting Information Available: Elemental analysis.
This material is available free of charge via the Internet at
http://pubs.acs.org.

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