Structure-activity relationships of alkyl- and alkoxy-substituted 1,4- dihydroquinoxaline-2,3-diones: Potent and systemically active antagonists for the glycine site of the NMDA receptor

Article abstract of DOI:10.1021/jm960654b

We report on a series of alkyl- and alkoxy-substituted 1,4- dihydroquinoxaline-2,3-diones (QXs), prepared as a continuation of our structure-activity relationship (SAR) study of QXs as antagonists for the glycine site of the N-methyl-D-aspartate (NMDA) re

Full text of DOI:10.1021/jm960654b

730
J . Med. Chem. 1997, 40, 730-738
Str u ctu r e-Activity Rela tion sh ip s of Alk yl- a n d Alk oxy-Su bstitu ted
1,4-Dih yd r oqu in oxa lin e-2,3-d ion es: P oten t a n d System ica lly Active An ta gon ists
for th e Glycin e Site of th e NMDA Recep tor
Sui Xiong Cai,*,† Sunil M. Kher,‡ Zhang-Lin Zhou,† Victor Ilyin,† Stephen A. Espitia,† Minhtam Tran,†
J on E. Hawkinson,† Richard M. Woodward,† Eckard Weber,† and J ohn F. W. Keana‡
CoCensys Inc., 213 Technology Drive, Irvine, California 92618, and Department of Chemistry, University of Oregon,
Eugene, Oregon 97403
Received September 18, 1996X
We report on a series of alkyl- and alkoxy-substituted 1,4-dihydroquinoxaline-2,3-diones (QXs),
prepared as a continuation of our structure-activity relationship (SAR) study of QXs as
antagonists for the glycine site of the N-methyl-D-aspartate (NMDA) receptor. The in vitro
potency of these antagonists was determined by displacement of the glycine site radioligand
[3H]-5,7-dichlorokynurenic acid ([3H]DCKA) in rat brain cortical membranes. In general, methyl
is a good replacement for chloro or bromo in the 6-position, and alkoxy-substituted QXs have
lower potencies than alkyl- or halogen-substituted QXs. Ethyl-substituted QXs are generally
less potent than methyl-substituted QXs, especially in the 6-position of 5,6,7-trisubstituted
QXs. Fusion of a ring system at the 6,7-positions results in QXs with low potency. Several
methyl-substituted QXs are potent glycine site antagonists that have surprisingly high in vivo
activity in the maximal electroshock (MES) test in mice. Among these, 7-chloro-6-methyl-5-
nitro QX (14g) (IC50 ) 5 nM) and 7-bromo-6-methyl-5-nitro QX (14f) (IC50 ) 9 nM) are
comparable in potency to 6,7-dichloro-5-nitro QX (2) (ACEA 1021) as glycine site antagonists.
QX 14g has an ED50 value of 1.2 mg/kg iv in the mouse MES assay. Interestingly, alkyl QXs
with log P values of 0.5 or less tend to be more bioavailable than QXs with higher log P values.
QX 14g has 440-fold selectivity for NMDA vs R-amino-3-hydroxy-5-methyl-4-isoxazolepropionic
acid (AMPA) receptors, as determined electrophysiologically under steady-state conditions in
oocytes expressing rat cerebral cortex poly(A)+ RNA. Overall, 14g was found to have the best
combination of in vitro and in vivo potency of all the compounds tested in this and previous
studies on the QX series.
Ch a r t 1
In tr od u ction
Glutamate is the primary excitatory neurotransmitter
in the central nervous system. Glutamate receptors are
classified into two main groups: ionotropic and metabo-
tropic.1 The N-methyl-D-aspartate (NMDA) type of
ionotropic glutamate receptors plays a major role in
glutamate excitotoxicity, a process implicated in a
number of pathophysiological conditions that lead to
neuronal death and degeneration including cerebral
ischemia and brain trauma.2 The discovery that the
amino acid glycine is a necessary coagonist for NMDA
receptor activation3 has stimulated research into the
development of antagonists for the NMDA receptor
glycine site.4 By inhibiting the glycine site these drugs
prevent NMDA receptor activation and hence could
potentially be used to reduce damage to the nervous
system incurred by cerebral stroke and head injury.
Several structural classes of highly potent glycine site
antagonists have been discovered.5 Many of these
compounds, however, have weak in vivo activity, prob-
ably due to poor penetration of the blood-brain barrier.6
4-Hydroxy-3-arylquinolin-2(1H)-ones such as 1,7 1,4-
dihydroquinoxaline-2,3-diones (QXs) such as 2,8 and
pyridazino[4,5-b]quinolinediones such as 39 are among
the few glycine site antagonists shown to be centrally
active following systemic administration (Chart 1).
We have recently reported the structure-activity
relationship (SAR) of QXs substituted with electron-
withdrawing groups such as halogen and nitro.10 This
study led to the discovery of 6,7-dichloro-5-nitro QX
(ACEA 1021) (2), a potent and systemically active
glycine site antagonist11 currently undergoing clinical
evaluation for stroke. As a continuation of SAR studies
in the QX series, we elected to explore substitution in
the 6- and 7-positions with electron-donating groups
such as alkyl, resulting in the discovery of 6,7-dimethyl-
5-nitro QX (9a ) as a potent glycine site antagonist with
high in vivo activity.12 Although QX 9a is about one-
fifth as potent as QX 2 at the glycine site, it is slightly
more potent in vivo as an anticonvulsant in the maximal
electroshock (MES) test in mice.13 On the basis of this
observation, we speculated that the combination of
alkyl, alkoxy, and halogen substituents in the 6- and
7-positions with a nitro group in the 5-position might
† CoCensys Inc.
‡ University of Oregon.
X Abstract published in Advance ACS Abstracts, J anuary 15, 1997.
S0022-2623(96)00654-1 CCC: $14.00 © 1997 American Chemical Society

SAR of Substituted 1,4-Dihydroquinoxaline-2,3-diones
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5 731
Sch em e 1
Sch em e 3
Sch em e 2a
Sch em e 4a
a
(a) (1) (CH3CO)2O; (2) KNO3/H2SO4; (b) (CF3CO)2O/CF3CO2H,
then KNO3; (c) 2 N HCl; (d) K2CO3 or NaOH; (e) SnCl2; (f) (NH4)2S;
(g) H2/Pd/C.
produce QXs with a favorable combination of in vitro
and in vivo potencies. Herein we report the synthesis
and potency of a series of alkyl- and alkoxy-substituted
QXs designed to test this hypothesis and the discovery
of 7-chloro-6-methyl-5-nitro QX (14g), a glycine site
antagonist that has the best combination of in vitro and
in vivo activities of all the QXs we have yet tested.14
a
(a) KNO3/H2SO4; (b) NH2CH2CO2Na; (c) BrCH2CO2Et; (d)
SnCl2/EtOH or Na2S2O4; (e) HNO3/CF3CO2H.
Ch em istr y
using KNO3/H2SO4 gave a complicated mixture, while
mild nitration using KNO3/CF3CO2H gave nitro 9g as
the sole product. Similarly, nitration of 6,7-dimethyl
QX (5a ) by KNO3/CF3CO2H gave a purer product 9a
than that using KNO3/H2SO4. However, when 6-chloro-
7-ethyl QX (5b) was nitrated, a 1:1 mixture of 5- and
8-nitro isomers was obtained.
QXs 5a -i were prepared by condensation of oxalic
acid with the corresponding 1,2-diaminobenzenes 4a -i
(Scheme 1). 6,7-Dihydroxy QX (5j) was obtained by
demethylation of 6,7-dimethoxy QX (5d ).
Those 1,2-diaminobenzenes that were not commer-
cially available were prepared from the corresponding
anilines using known procedures.10 Briefly, the amino
group of the anilines was first protected by reaction with
acetic anhydride to give the amides, which were then
nitrated in H2SO4 with KNO3. Alternatively, the aniline
was treated with trifluoroacetic anhydride, and then
KNO3 was added to the reaction mixture to give the
nitro product. The amides were then hydrolyzed to give
the o-nitroanilines which were then reduced to give the
1,2-diaminobenzenes (Scheme 2).
3-Chloro-4-ethylaniline (6b) was prepared from 2-eth-
ylaniline according to Lambooy and Lambooy.15 1,2-
Diamino-4,5-diethylbenzene (4c) was prepared from 1,2-
diethylbenzene using a modified procedure of Lambooy.16
1,2-Diamino-5-methyl-3-nitrobenzene (4h ) was obtained
by selectively reducing one of the nitro groups in
4-methyl-2,6-dinitroaniline using (NH4)2S. 1,2-Diamino-
4,5-(methylenedioxy)benzene (4e) was prepared by hy-
drogenation of 4,5-(methylenedioxy)-1,2-dinitroben-
zene.17
5-Nitro-6,7-disubstituted QXs 14a -h were prepared
by a regiospecific oxidative nitration procedure and the
structures were assigned as shown based on our earlier
observations (Scheme 4).18 The intermediate N-phen-
ylglycinates (12) were prepared by nucleophilic aromatic
substitution of 4,5-disubstituted-2-halonitrobenzenes
(11) using sodium glycinate. Alternatively, nucleophilic
substitution on ethyl bromoacetate by 4,5-disubstituted
2-nitroanilines (8a ,b) gave the N-phenylglycinates. The
nitro group was reduced by SnCl2 or Na2S2O4 and the
product spontaneously cyclized to give the 3,4-dihydro-
quinoxalin-2(1H)-ones (13). Nitration of the quinoxalin-
2-ones with fuming HNO3 in trifluoroacetic acid resulted
in both nitration and oxidation to give 5-nitro-6,7-
disubstituted QXs (14a -h ) regiospecifically. Similarly,
4-chloro-3-nitrobenzonitrile was reacted with sodium
glycinate followed by reductive cyclization to give 7-cy-
anoquinoxalin-2-one. Oxidative nitration of the qui-
noxalin-2-one gave 7-cyano-5-nitro QX (14i) regiospe-
cifically.
6,7-Disubstituted QXs (5a -c,g) were treated in con-
centrated H2SO4 or trifluoroacetic acid with KNO3 to
give 5-nitro-6,7-disubstituted QXs (Scheme 3). It was
found that KNO3/CF3CO2H is the better nitration
system for alkyl QXs. For instance, nitration of 5g
The 4,5-disubstituted-2-halonitrobenzenes (11) were
prepared by nitration of 3,4-disubstituted halobenzenes
(10). 2,5-Dichloro-1-ethylbenzene (10c) was prepared

732 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5
Cai et al.
Ta ble 1. SAR of 1,4-Dihydroquinoxaline-2,3-diones at the
NMDA Receptor Glycine Site
Sch em e 5
no.
R5
NO2
H
H
H
H
H
H
H
NO2
H
H
H
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
NH2
NO2
NO2
NO2
NO2
NO2
NO2
NO2
OCOMe
OH
H
R6
R7
Cl
Me
Et
Et
OMe
[3H]DCKA IC50 (µM)a
2
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
9a
9c
9g
9i
14a
14b
14c
14d
14e
14f
14g
14h
14i
15
18a
18b
18c
18d
18e
18f
18g
19a
19b
20
Cl
Me
Cl
Et
0.0059 ( 0.0010
3.3 ( 0.7
1.4 ( 0.2
1.8 ( 0.2
OMe
OCH2O
CHdCHCHdCH
17 ( 1
>100
16 ( 1
CH2CH2CH2
15 ( 4
by reduction of 2′,5′-dichloroacetophenone. 3-Bromo-4-
ethyl-6-nitroaniline (8a ) was prepared from 2-ethyl-5-
nitroaniline in a manner similar to that of 3-chloro-4-
ethyl-6-nitroaniline (8b).15
6-Chloro-7-methyl QX (5k ) was prepared by oxidation
(H2O2/AcOH) of 6-chloro-7-methyl-3,4-dihydroquinoxa-
lin-2(1H)-one (13g). Reduction of 7-chloro-6-methyl-5-
nitro QX (14g) with SnCl2 gave 5-amino-7-chloro-6-
methyl QX (15).
H
Me
Br
OH
Me
Me
Et
9.3 ( 1.9
OMe
OH
Cl
1.0 ( 0.2
>100
0.78 ( 0.06
0.029 ( 0.003
0.16 ( 0.03
3.1 ( 0.5
Me
Et
CH2CH2CH2
OMe
Br
Br
Et
Et
Cl
CN
F
Br
Cl
Me
CN
Cl
F
F
F
F
F
Cl
Cl
OMe
OMe
Cl
0.064 ( 0.003
0.082 ( 0.019
0.029 ( 0.002
0.13 ( 0.01
0.073 ( 0.010
0.095 ( 0.024
0.0087 ( 0.0004
0.0047 ( 0.0006
0.045 ( 0.008
2.8 ( 0.4
Cl
Et
Me
Me
Me
Me
Cl
H
During our formulation and stability experiments
with 6-halo-5-nitro-7-substituted QXs, we found that
some of these compounds are unstable toward nucleo-
philes such as amines. In particular, replacement of
the fluoro ortho to the nitro group by an amino group
was observed. We recognized that the instability of
these compounds provides access to a group of 5-nitro-
6,7-disubstituted QXs. Thus, nucleophilic substitution
of the fluoro ortho to the nitro group in 6,7-difluoro-5-
nitro QX (16)10 and 7-chloro-6-fluoro-5-nitro QX (17)10
with different nucleophiles gave QXs 18a -g (Scheme
5). The reaction could be followed by 1H NMR spec-
troscopy. For QX 16, replacement of the fluoro ortho
to the nitro group by a nucleophile converts the aromatic
proton from a doublet of doublets to a doublet. The F-H
coupling constants for QXs 18a -e are all around 11.5
Hz, which confirms that the remaining fluoro is ortho
to the ring proton.
Me
0.15 ( 0.03
11 ( 1
NH2
OMe
OEt
OBu-n
O(CH2)3Ph
OMe
SEt
OMe
OMe
Cl
1.9 ( 0.1
3.6 ( 0.7
4.3 ( 0.5
4.0 ( 0.9
0.15 ( 0.02
0.64 ( 0.13
53 ( 16
17 ( 2
0.13 ( 0.03
a
Potency is expressed as the concentration producing 50%
inhibition (IC50) of [3H]DCKA binding. Values are means ( SEMs
of at least three independent experiments.
Resu lts a n d Discu ssion
5-Acetoxy-6,7-dimethoxy QX (19a ) was obtained by
acetoxylation (HNO3/HOAc) of 6,7-dimethoxy QX (5d ).26
Hydrolysis of 19a gave 5-hydroxy QX (19b).
The SAR of QXs as antagonists for the NMDA
receptor glycine site is summarized in Table 1. All the
6,7-disubstituted QXs (5a -g,i-k ) are less active than
6,7-dichloro QX10 20, implying that electron-withdraw-
ing groups are important for high potency of 6,7-
disubstituted QXs. QXs 5b, 5i, and 5k , substituted with
one chlorine or bromine atom, are among the most
potent inhibitors in this group. 6,7-Dimethyl QX 5a and
6,7-diethyl QX 5c have moderate potency, whereas
dimethoxy QX 5d , substituted with two strong electron-
donating groups, has low affinity. Dihydroxy QX 5j is
inactive, indicating that hydrophilic groups are not
tolerated in the 6- and 7-positions. QX 5e, with a
methylenedioxy ring in the 6,7-position, is inactive, and
QXs 5f and 5g have low affinity, all suggesting that
incorporation of the 6,7-positions into ring systems is
not tolerated.
As expected from our earlier SAR of QXs,10 introduc-
tion of a nitro group at the 5-position consistently results
in higher affinity. Thus, 6,7-dimethyl-5-nitro QX 9a is
about 100-fold more potent than 6,7-dimethyl QX 5a ,
and 6,7-diethyl-5-nitro QX 9c is about 10-fold more
potent than 6,7-diethyl QX 5c. Likewise, 7-chloro-6-
methyl-5-nitro QX 14g and 6-chloro-7-methyl-5-nitro
P h a r m a cology
The potency of QXs for the NMDA receptor glycine
site was measured by displacement of [3H]-5,7-dichlo-
rokynurenic acid ([3H]DCKA) binding in rat brain
cortical membranes as described previously.19 For
selected QXs, potencies at NMDA receptor glycine sites
and at R-amino-3-hydroxy-5-methyl-4-isoxazole propi-
onic acid (AMPA) receptors were determined electro-
physiologically in Xenopus oocytes expressing rat brain
poly(A)+ RNA or, in certain cases for NMDA receptor,
using the cloned rat NMDA receptor (NR) 1a/2C subunit
combinations.20 Apparent antagonist dissociation con-
stants (Kb values) were estimated by assuming competi-
tive inhibition and assaying suppression of membrane
current responses elicited by fixed concentrations of
agonist: 1 µM glycine and 100 µM glutamate for NMDA
receptors; 10 µM AMPA for AMPA receptors.19 Levels
of receptor expression were similar to those reported
previously.10,19 Anticonvulsant activity of selected QXs
was measured in a mouse MES model11,14 which was
used as a rough estimate of systemic bioavailability.

SAR of Substituted 1,4-Dihydroquinoxaline-2,3-diones
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5 733
Ta ble 2. Functional Antagonism of NMDA and AMPA Receptors Expressed in Xenopus Oocytes by QXs
Kb (µM)
selectivity
no.
glycine
AMPA
for NMDAa
nb
9a c
9g
0.039 (0.034-0.043)d
0.87e (0.72-1.0)
3.1 (2.7-3.6)
17f (15-20)
79
20
20
4, 4
3, 3
3, 4
4, 3
14d
0.029e (0.024-0.035)
0.0079 (0.0072-0.0087)
0.59f (0.51-0.68)
3.5 (3.1-4.0)
14gg
440
a
b
The steady-state selectivity index for inhibition of NMDA receptors was estimated by dividing Kb AMPA by Kb NMDA. Indicates
the number of independent experiments (cells examined); numbers refer to NMDA and AMPA, respectively. c Data from ref 12b. Numbers
in parentheses are 95% confidence intervals adjusted to the linear scale. e Apparent antagonist dissociation constants (Kb) for NMDA
receptor glycine sites were determined from inhibition of currents activated by 1 µM glycine and 100 µM glutamate in oocytes expressing
the cloned rat NMDA receptor subunit combination NR1a/2C. f Kb values for AMPA receptors were determined from inhibition of currents
d
activated by 10 µM AMPA in oocytes expressing rat cerebral cortex poly(A)+ RNA. Data from ref 14.
g
QX 14h are 160- and 17-fold more potent than 6-chloro-
7-methyl QX 5k , respectively. QX 9g is the least potent
inhibitor among 5-nitro-6,7-disubstituted QXs, again
suggesting that incorporation of the 6,7-positions into
a ring system considerably reduces potency.
6,7-Dimethyl-5-nitro QX 9a is >5-fold more potent
than 6,7-diethyl-5-nitro QX 9c, implying the existence
of a size-limited pocket in the region surrounding the
6- and 7-positions for 5-nitro-6,7-disubstituted QXs.
Since 6-chloro-7-methyl-5-nitro QX 14h and 6-chloro-
7-ethyl-5-nitro QX 14b have comparable activity while
7-chloro-6-methyl-5-nitro QX 14g is 26-fold more potent
than 7-chloro-6-ethyl-5-nitro QX 14c, the size-limited
effect might be more pronounced at the 6-position
compared to the 7-position.
7-Chloro-6-methoxy-5-nitro QX 18f has affinity com-
parable to that of 7-chloro-6-ethyl-5-nitro QX 14c,
indicating the similarity of methoxy and ethyl at the
6-position. QX 18f and QX 14c is 30 times and 26 times
less potent than 7-chloro-6-methyl-5-nitro QX 14g,
respectively, and 7-fluoro-6-methoxy-5-nitro QX 18b is
about 20 times less potent than 7-fluoro-6-methyl-5-
nitro QX 14e. This sharp drop in affinity from methyl
to ethyl or methoxy is another indication that there is
a size-limited pocket adjacent to the 6-position. Inter-
estingly, QXs 18b-e, with groups ranging in size from
methoxy to 3-phenylpropoxy at the 6-position, have
comparable affinity, suggesting that after the sharp
reduction in potency from methyl to methoxy a further
increase in the size of substituents at the 6-position has
little effect on potency. Similarly, QX 18g, with a
thioethoxy group at the 6-position, is only 4 times less
active than QX 18f. 6-Amino-7-fluoro-5-nitro QX 18a
is the least potent ligand among QXs 18a -e, again
suggesting that hydrophilic groups are not tolerated at
the 6-position.
optimal at the 7-position.21,22 Interestingly, 6-chloro-
7-ethyl-5-nitro QX 14b is about 5-fold more potent than
7-chloro-6-ethyl-5-nitro QX 14c, while 7-chloro-6-meth-
yl-5-nitro QX 14g is about 9-fold more potent than
6-chloro-7-methyl-5-nitro 14h , indicating that ethyl is
better tolerated at the 7-position than at the 6-position,
while methyl is better tolerated at the 6-position than
at the 7-position.
5-Amino-7-chloro-6-methyl QX 15 is 30 times less
potent than the corresponding 5-nitro QX 14g, indicat-
ing the importance of the 5-nitro group. Similarly,
5-acetoxy-6,7-dimethoxy QX 19a and 5-hydroxy-6,7-
dimethoxy QX 19b26 have very low potency, again
confirming the importance of electron-withdrawing
groups at the 5-position. Furthermore, 6,7-dimethyl-
5-nitro QX 9a and 6-chloro-7-methyl-5-nitro QX 14h are
>100-fold more potent than 6,7-dimethyl QX 5a and
7-methyl-5-nitro QX 5h , indicating that a substitution
pattern encompassing all three positions is critical for
high potency of 5-nitro-6,7-disubstituted QXs.10
Functional antagonism of NMDA receptor by four
selected QXs was tested using electrophysiological
recording techniques in Xenopus oocytes expressing the
cloned rat NMDA receptor subunit combination NR1a/
2C, or native rat brain poly(A)+ RNA8 (Table 2). For
QXs 9a and 14g, which were measured on rat brain
NMDA receptor, the inhibitory potency was similar to
that measured in [3H]DCKA binding assay. For QXs
9g and 14d , which were measured on NR1a/2C receptor,
potency in the electrophysiological assay was ∼3-fold
higher than that in the [3H]DCKA binding. This small
discrepancy between assays may be due to a slightly
higher potency of QXs at NR1a/2C receptors as com-
pared to the mixture of receptors expressed in oocytes
by rat whole brain poly(A)+ RNA.8
Electrophysiological recordings were also used to test
the four selected QXs for functional antagonism of
AMPA receptors in oocytes expressing rat cerebral
cortex poly(A)+ RNA. Similar to halogen- and nitro-
substituted QXs,10 the alkyl-substituted QXs were all
AMPA receptor antagonists (Table 2). 7-Chloro-6-
methyl-5-nitro QX 14g and 6,7-dimethyl-5-nitro QX 9a
have comparable, low micromolar potency for AMPA
receptors, but still show >400- and ∼80-fold selectivity
for the NMDA glycine site, respectively. These results
suggest that, for the AMPA receptors, methyl and chloro
are equivalent substituents at the 7-position of QXs. Of
the QXs tested, 7-cyano-6-methyl-5-nitro QX 14d is the
most potent AMPA antagonist with a Kb of 590 nM,
indicating that for the AMPA receptor, cyano is pre-
ferred over methyl and chloro at the 7-position, and this
substituent pattern results in a QX which has relatively
low selectivity for the glycine site. QX 9g, with the 6,7-
7-Chloro-6-methyl-5-nitro QX 14g and 7-bromo-6-
methyl-5-nitro QX 14f are the most potent antagonists
in this series with IC50 values of 5 and 9 nM, respec-
tively. The potency of 14g is similar to 6,7-dichloro-5-
nitro QX 2, showing that methyl and chloro are com-
parable at the 6-position. In contrast, 6-chloro-7-
methyl-5-nitro QX 14h is about 7 times less active than
QX 2, indicating that methyl is not such a good replace-
ment for chloro at the 7-position. 7-Fluoro-6-methyl-
5-nitro QX 14e and 7-cyano-6-methyl-5-nitro QX 14d
are about 19 times and 15 times less active than 14g,
respectively, again suggesting that chloro is preferred
at the 7-position. Similarly, 7-fluoro-6-methoxy-5-nitro
18b is about 12 times less potent than 7-chloro-6-
methoxy-5-nitro QX 18f. These observations are in
agreement with SAR developed in several other series
of glycine antagonists in which a chlorine atom is

734 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5
Ta ble 3. Anticonvulsant Activity (MES) of QXs in Mouse
Cai et al.
Con clu sion s
In conclusion, a series of alkyl- and alkoxy-substituted
QXs were synthesized and evaluated as antagonists of
the NMDA receptor glycine site. Methyl is a good
replacement for chloro in the 6-position, and alkoxy-
substituted QXs generally have potency lower than
alkyl- or halogen-substituted QXs. QXs substituted at
the 6-position by groups larger than methyl or chloro
show a reduction in potency. 7-Chloro-6-methyl-5-nitro
QX (14g) and 7-bromo-6-methyl-5-nitro QX (14f) are the
most active inhibitors with IC50 values of 5 and 9 nM
in the [3H]DCKA assay, respectively. These two QXs
are among the most potent glycine antagonists reported
to date. Under steady-state conditions, QX 14g is >400-
fold selective for NMDA receptors vs AMPA receptors
as measured electrophysiologically. Most importantly,
a number of the alkyl-substituted QXs are potent,
systemically active anticonvulsants in the MES assay
in mice. QXs 9a and 14e-g are also neuroprotective
in a rat model of focal cerebral ischemia.25 These
compounds therefore have potential as neuroprotectants
for the clinical treatment of ischemic stroke.
[3H]DCKA
IC50 (nM)
MES ED50
mg/kg
no.
R6
R7
log Pa
14g
14e
14h
14f
9a
2
14b
21
Me
Me
Cl
Me
Me
Cl
Cl
Br
Br
Cl
F
4.7
95
45
8.7
29
5.9
29
1.2 (0.9-1.6)
1.8 (1.3-2.6)
1.9 (1.2-2.8)
2.0 (1.4-2.9)
2.6 (1.8-3.8)
4.0 (3.2-5.0)
4.5 (3.1-6.4)
7.3 (5.7-9.4)
11 (8.5-14)
0.49
0.08
0.44b
0.61b
0.58
0.21
0.92c
0.74b
1.04
Me
Br
Me
Cl
Et
Br
Et
20
82
14a
a
Measured by shake-flask method in octanol vs pH 7.4 buffer.
Estimated by a HPLC method. c Calculated value.
b
positions cyclized into a ring system, has lower activity
at both NMDA and AMPA receptors compared to that
of QX 9a .
To estimate systemic bioavailability, anticonvulsant
activity of seven alkyl substituted QXs was measured
in a mouse MES model. Table 3 summarizes the
anticonvulsant potency and the log P values of the seven
QXs, together with QX 2 and 6,7-dibromo-5-nitro QX
Exp er im en ta l Section
Ch em istr y. Melting points were determined in open
capillary tubes on a Mel-Temp apparatus and are uncorrected.
The 1H NMR spectra were recorded at 300 MHz. Chemical
shifts are reported in ppm (δ), and J coupling constants are
reported in hertz. 19F NMR spectra were recorded with C6F6
(-162.9 ppm) as internal standard. Elemental analyses were
performed by Desert Analytics, Tucson, AZ. Mass spectra
(MS) were obtained using a VG 12-250 or a VG ZAB-2FHF
mass spectrometer. Substituted anilines were obtained from
Aldrich or Lancaster and used as received. Reagent grade
solvents were used without further purification unless other-
wise specified. Reverse phase HPLC were obtained at 254 nM
on a 4.6 × 250 mM microsorb-MV C18 column, using as
solvents 0.1% trifluoroacetic acid in water (A) and 0.1%
trifluoroacetic acid in acetonitrile (B). The linear gradient was
20% B in A to 95% B in A with a flow rate of 1 mL/min. The
preparation of QXs 5d , 5i, 9i, and 19a ,b26 and QXs 14e-h 18
have been reported previously.
3-Br om o-4-et h yln it r ob en zen e. To a mixture of 6.61 g
(39.8 mmol) of 2-ethyl-5-nitroaniline15 in 15 mL of 48% HBr
stirred in ice bath was added a solution of 2.84 g (41.1 mmol)
of NaNO2 in 5 mL of H2O. The resulting mixture was stirred
in ice bath for 30 min, and then was added in portions into a
boiling solution of 3.62 g (25.2 mmol) of CuBr in 8 mL of 48%
HBr. The resulting solution was boiled for 1 h, cooled to room
temperature, and extracted with CHCl3 (3 × 20 mL). The
extract was washed with aqueous 1 N NaOH (10 mL), aqueous
1 N HCl (10 mL), and water (2 × 10 mL), dried (MgSO4), and
evaporated to leave 8.5 g (93%) of yellow liquid: 1H NMR
(CDCl3) 1.28 (t, J ) 7.5, 3H), 2.88 (q, J ) 7.5, 2H), 7.40 (d, J
) 8.5, 1H), 8.12 (dd, J ) 2.3, 8.4, 1H), 8.42 (d, J ) 2.2, 1H).
3-Br om o-4-eth yla n ilin e (6a ). A solution of 7.8 g (33.9
mmol) of 3-bromo-4-ethylnitrobenzene and 30.6 g (135 mmol)
of SnCl2‚2H2O in 75 mL of absolute alcohol was refluxed for 1
h. It was evaporated to remove most of the solvent, and the
residue was treated with aqueous 2 N NaOH to pH ) 5. The
mixture was filtered, and the solid was washed with ethanol
(20 mL) and ethyl acetate (40 mL). The filtrate was separated,
and the aqueous phase was extracted with ethyl acetate (15
mL). The combined organic phase was dried (MgSO4) and
evaporated to leave 6.0 g (88%) of pale-yellow liquid: 1H NMR
(CDCl3) 1.17 (t, J ) 7.5, 3H), 2.64 (q, J ) 7.7, 2H), 3.53 (sb,
2H), 6.58 (dd, J ) 2.3, 8.1, 1H), 6.90 (d, J ) 2.3, 1H), 7.00 (d,
J ) 8.1, 1H).
21, two of the most potent QXs reported earlier.10
A
surprising number of the alkyl-substituted QXs were
potent anticonvulsants following iv administration.
Indeed, five of the seven alkyl-substituted QXs tested
were more potent anticonvulsants than QX 2, the most
active QX identified in previous studies.10,11
The reason for the high in vivo activity of these alkyl-
substituted QXs remains unclear. As seen in other
series of NMDA receptor glycine site antagonists,6 there
is no direct correlation between in vitro and in vivo
activities (Table 3). There is also no direct correlation
between in vivo activity and log P value. Interesting
observations can be made, however, by considering the
in vitro, in vivo, and log P data together. For example,
QXs 14e and 14a have similar in vitro potencies, but
14e has a log P value lower than that of 14a and is >5-
fold more active in vivo. QXs 9a , 14b, and 2110 also
have similar in vitro potencies, but 9a has a log P value
lower than that of 14b and 21, and 9a is about 2-fold
and 3-fold more active in vivo than 14b and 21,
respectively. In addition, although QX 14e is 19 times
less active than 14g in vitro, it is only <2 times weaker
in vivo. QXs 14h and 14f have log P values around 0.5
and are highly active in vivo. These data imply that
QXs with log P values of ∼0.5 or less might be more
bioavailable than QXs with higher log P values, which
is a deviation from the conventional rule of thumb that
compounds with a log P value around 2 are optimal for
CNS drugs.23 Since QX 2 is highly bound to plasma
proteins (>99%),24 we hypothesize that part of the
reason might be that QXs with log P values ∼0.5 or less
achieve an optimum for limiting binding to plasma
proteins and still being sufficiently hydrophobic to
penetrate the blood-brain barrier. We speculate that
QXs with higher log P values might be too tightly bound
to plasma proteins, resulting in low in vivo activity.
Other physiochemical and pharmacokinetic parameters,
such as solubility of the QXs in plasma, and clearance
rates, are also likely to contribute to the observed
difference in in vivo potency.
4-Br om o-5-eth yl-2-(tr iflu or oa ceta m id o)n itr oben zen e
(7a ). To a solution of 30 mL of (CF3CO)2O and 30 mL of CF3-
CO2H stirred in ice bath was added dropwise 5.80 g (29.0
mmol) of 6a , and the resulting mixture was stirred at room
temperature for 1 h. To the mixture stirred in ice bath was

SAR of Substituted 1,4-Dihydroquinoxaline-2,3-diones
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5 735
added in portion 3.15 g (31.2 mmol) of KNO3, and it was stirred
in an ice bath for 1 h and at room temperature overnight. The
solution was added into 150 mL of ice-water, and the
precipitate was filtered, washed with water, and dried to give
9.8 g (98%) of pale-yellow solid: 1H NMR (CDCl3) 1.29 (t, J )
7.5, 3H), 2.83 (q, J ) 7.4, 2H), 8.16 (s, 1H), 8.99 (s, 1H), 11.30
(s, 1H). The crude product was used for the next reaction
without purification.
The mixture was diluted with 20 mL of H2O, filtered, washed
with water, and dried to give pale-brown solid 3.57 g (94%):
mp >250 °C; H NMR (DMSO-d6) 2.16 (s, 6H), 6.87 (s, 2H),
11.79 (s, 2H). Anal. (C10H10N2O2) C, H, N.
1
QXs 5b,c,e-h were prepared from the corresponding 1,2-
diaminobenzenes in a manner similar to that for 5a .
7-Ch lor o-6-eth yl-1,4-dih ydr oqu in oxalin e-2,3-dion e (5b):
1
mp >400 °C; H NMR (DMSO-d6) 1.14 (t, J ) 7.3, 3H), 2.64
3-Br om o-4-eth yl-6-n itr oa n ilin e (8a ). A mixture of 5.10
g (14.7 mmol) of 7a in 35 mL of 7 wt % K2CO3 methanol/H2O
(3:2) was stirred at room temperature for 2 h. It was diluted
with 20 mL of water, filtered, washed with water and dried to
give 1.81 g (49%) of yellow solid: mp 75-76 °C; 1H NMR
(CDCl3) 1.22 (t, J ) 7.4, 3H), 2.68 (q, J ) 7.6, 2H), 5.93 (br s,
2H), 7.08 (s, 1H), 7.96 (s, 1H).
(q, J ) 7.4, 2H), 7.31 (s, 1H), 7.11 (s, 1H), 11.91 (s, 1H), 11.91
(s, 1H); HRMS calcd for C10H9ClN2O2 224.0348, found 224.0359.
Anal. Calcd (C10H9ClN2O2) C, H, N.
6,7-Dieth yl-1,4-d ih yd r oqu in oxa lin e-2,3-d ion e (5c): mp
1
>360 °C; H NMR (DMSO-d6) 1.09 (t, J ) 7.5 Hz, 6H), 2.51
(q, J ) 7.5 Hz, 4H), 6.87 (s, 2H), 11.74 (s, 2H). Anal.
(C12H14N2O2) C, H, N.
5-Aceta m id o-6-n itr oin d a n (7g). To a stirred solution of
5.2 g (39 mmol) of 5-aminoindan (6g) in 15 mL of dioxane in
an ice bath was added dropwise 8 mL (8.6 g, 84 mmol) of acetic
anhydride. The solution was stirred at room temperature for
16 h. It was diluted with 70 mL of water, and the mixture
was filtered, washed by water, and dried to give a gray solid:
6.21 g (91%); 1H NMR (CDCl3) 2.07 (m, 2H), 2.17 (s, 3H), 2.88
(m, 4H), 7.20 (mb, 1H), 7.15 (s, 2H), 7.44 (s, 1H).
1,4-Dih yd r o-6,7-(m et h ylen ed ioxy)q u in oxa lin e-2,3-d i-
on e (5e): mp >360 °C; 1H NMR (DMSO-d6) 6.00 (s, 2H), 6.78
(s, 2H), 11.70 (s, 2H); HRMS calcd for C9H6N2O4 206.2304,
found 206.2306. Anal. (C9H6N2O4) C, H, N.
1,4-Dih ydr oben zo[g]qu in oxalin e-2,3-dion e (5f): mp >250
°C; 1H NMR (DMSO-d6) 7.38 (dd, J ) 3.2, 6.2, 2H), 7.53 (s,
2H), 7.82 (dd, J ) 3.2, 6.2, 2H), 12.09 (s, 2H). Anal.
(C12H8N2O2‚H2O) C, H, N.
To a solution of 5.59 g (31.9 mmol) of the amide in 55 mL of
H2SO4 kept in an ice bath was added portionwise 3.62 g of
KNO3 (35.8 mmol). The solution was stirred in ice bath for 2
h and at room temperature overnight. It was added into 500
mL of ice-water, and the mixture was stirred for 1 h. The
precipitate was filtered, washed with water, and dried to leave
black solid which was purified by chromatography (silica gel,
eluted with hexane/ethyl acetate ) 10:1) to give 1.06 g (15%)
of a yellow solid: 1H NMR (CDCl3) 2.14 (m, 2H), 2.28 (s, 3H),
3.00 (m, 4H), 8.44 (s, 1H), 8.58 (s, 1H), 10.39 (s, 1H).
5-Am in o-6-n itr oin d a n (8g). A mixture of 259 mg (1.16
mmol) of 7g in 4 mL of 2 N HCl was heated at 85 °C for 9 h
and cooled to room temperature. The solid precipitate was
filtered, washed with water, and dried to leave 201 mg (97%)
of a crystalline yellow solid: 1H NMR (CDCl3) 2.07 (m, 2H),
2.84 (m, 4H), 6.00 (sb, 2H), 6.65 (s, 1H), 7.94 (s, 1H).
5,6-Dia m in oin d a n (4g). A solution of 200 mg (1.12 mmol)
of 8g and 1.14 g (6.01 mmol) of SnCl2 in 8 mL of ethanol was
heated at 70 °C for 2 h. It was evaporated to remove the
ethanol, and the residue was treated with 40% aqueous NaOH
to pH ) 12. The mixture was diluted by 4 mL of water and
extracted with CHCl3 (3 × 10 mL). The extract was dried
(MgSO4) and evaporated to leave 162 mg (97%) of a yellow
crystalline solid: 1H NMR (CDCl3) 2.01 (m, 2H), 2.77 (t, J )
7.3, 4H), 3.30 (sb, 4H), 6.61 (s, 2H).
1,4-Dih yd r ocyclop en to[g]qu in oxa lin e-2,3-d ion e (5g):
1
mp >360 °C; H NMR (DMSO-d6) 1.99 (m, 2H), 2.81 (t, J )
7.3, 4H), 6.957 (s, 2H), 11.83 (s, 2H); HRMS calcd for
C11H10N2O2 202.0738, found 202.0747. Anal. (C11H10N2O2) C,
H, N.
1,4-Dih ydr o-7-m eth yl-5-n itr oqu in oxalin e-2,3-dion e (5h ):
mp 319-323 °C dec; 1H NMR (DMSO-d6) 2.35 (s, 3H), 7.27(s,
1H), 7.75 (s, 1H), 11.04 (s, 1H), 12.30 (s, 1H). Anal.
(C9H7N3O4‚0.35H2O) C, H, N.
1,4-Dih yd r o-6,7-d ih yd r oxyqu in oxa lin e-2,3-d ion e (5j).
To a suspension of 5d (222 mg, 1.0 mmol) in 2 mL of methylene
chloride was added 5 mL of a solution of boron tribromide in
methylene chloride (1 M, Aldrich). The resulting mixture was
allowed to stir at room temperature for 24 h. The mixture
was poured into ice-water (10 g) to form a suspension.
Aqueous sodium hydroxide (20%, 10 mL) was added to the
suspension to form a red solution. Then the solution was
acidified with 6 N HCl (10 mL) to pH ) 1. The suspension
was centrifuged, washed with methanol, and dried in vacuo
1
to give 170 mg (88%) of 5j as a brown solid: mp >350 °C; H
NMR (DMSO-d6) 6.54 (s, 2H), 8.99 (s, 2H), 11.54 (s, 2H);
HRMS calcd for C8H6N2O4 194.1480, found 194.1475. Anal.
(C8H6N2O4) C, H, N.
6-Ch lor o-1,4-d ih yd r o-7-m et h ylq u in oxa lin e-2,3-d ion e
(5k ). A suspension of 7-chloro-3,4-dihydro-6-methylquinoxa-
lin-2(1H)-one18 (38 mg, 0.19 mmol), 30% H2O2 (0.48 mL), and
AcOH (0.8 mL) was stirred at room temperature for 3 days.
Cream-colored solid was collected by vacuum filtration, washed
with water (5 mL), and dried in vicuo to give 18 mg (44%) of
5k as cream-colored powder: mp > 365 °C; 1H NMR (DMSO-
d6) 2.27 (s, 3H), 7.02 (s, 1H), 7.11 (s, 1H), 11.89 (s, 1H), 11.95
(s, 1H); Anal. (C9H7ClN2O2‚0.72H2O) C, H: calcd, 3.81; found,
3.24; N.
1,4-Dih yd r o-6,7-d im et h yl-5-n it r oq u in oxa lin e-2,3-d i-
on e (9a ). To a suspension of 1.90 g (10.0 mmol) of 5a in 50
mL of TFA was added 1.104 g (11.0 mmol) of KNO3 at room
temperature, and the mixture was stirred at room temperature
for 24 h. The resulting solution was evaporated to dryness,
and the residue was treated with 30 mL of MeOH and
evaporated again to dryness. The solid was treated with 20
mL of water, filtered, washed with water, and dried to give
1.86 g (79%) of 9a as a yellow solid: mp 326-328 °C; 1H NMR
(DMSO-d6) 2.08 (s, 3H), 2.25 (s, 3H), 7.05 (s, 1H), 11.77 (s,
1H), 12.03 (s, 1H). Anal. (C10H9N3O4) C, H, N.
1,2-Dia m in o-4-ch lor o-5-eth ylben zen e (4b). Diamine 4b
was prepared from 8b15 in a manner similar to that for 4g: 1H
NMR (CDCl3) 1.17 (t, J ) 7.5, 3H), 2.59 (q, J ) 7.4, 2H), 6.56
(s, 1H), 6.70 (s, 1H).
1,2-Dia m in o-4,5-(m eth ylen ed ioxy)ben zen e (4e). 4,5-
(Methylenedioxy)-1,2-dinitrobenzene17 (1.37 g, 6.47 mmol) was
dissolved into ethyl acetate (20 mL). To this solution was
added 10% Pd/C (343 mg, 20%). The mixture was then stirred
at room temperature under the pressure of 40 psi (H2) for 14
h. The catalyst was removed through a column of Celite (5 g)
and washed with ethyl acetate (3 × 15 mL) under nitrogen.
The filtrates were combined, and the solvent was removed to
give the diamine 4e as a nearly colorless solid: 1H NMR
(CDCl3) 5.80 (s, 2H), 6.34 (s, 2H).
1,2-Diam in o-4-m eth yl-6-n itr oben zen e (4h ). A dark black
solution of 4-methyl-2,6-dinitroaniline (0.100 g, 0.561 mmol)
in 6.6% aqueous (NH4)2S (3.3 mL) and ethanol (3.5 mL) was
refluxed for 45 min. It was then cooled to room temperature,
and the volatiles were removed in vacuo (inside the hood to
avoid the stench of ammonium sulfide). The slurry so obtained
was diluted with water (10 mL), and the resulting red solid
was filtered and dried under vacuum to give 0.072 g (85%) of
4h as a red powder which was used as such for the next
reaction: 1H NMR (acetone-d6) 2.21 (s, 3H), 6.34 (br s, 2H),
6.62 (s, 1H), 7.57 (s, 1H).
QXs 9c,g,b (mixture of 14b and 14c) were obtained from
nitration of the corresponding 6,7-disubstituted QX in
a
manner similar to QX 9a .
6,7-Diet h yl-1,4-d ih yd r o-5-n it r oq u in oxa lin e-2,3-d ion e
1
(9c): mp 270-272 °C dec; H NMR (DMSO-d6) 1.14 (m, 6H),
2.62 (m, 4H), 7.09 (s, 1H), 11.76 (s, 1H), 12.03 (s, 1H). Anal.
(C12H13N3O4) C, H, N.
1,4-Dih yd r o-6,7-d im eth ylqu in oxa lin e-2,3-d ion e (5a ). A
mixture of 2.72 g (20.0 mmol) of 1,2-diamino-4,5-dimethylben-
zene (4a ) and 1.92 g (21.3 mmol) of oxalic acid in 30 mL of 2
N HCl was refluxed for 2.5 h and cooled to room temperature.
1,4-Dih yd r o-5-n it r ocyclop en t o[g]q u in oxa lin e-2,3-d i-
1
on e (9g): mp 310 °C dec; H NMR (DMSO-d6) 2.04 (m, 2H),
2.97 (t, J ) 7.5, 2H), 3.09 (t, J ) 7.5, 2H), 7.26 (s, 1H), 11.18

736 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5
Cai et al.
(s, 1H), 12.23 (s, 1H); HRMS calcd for C11H9N3O4 247.0588,
found 247.0593. Anal. (C10H6N4O4‚0.5H2O) C, H; N: calcd,
16.44; found, 15.77.
6-Ch lor o-7-et h yl-1,4-d ih yd r o-5-n it r oq u in oxa lin e-2,3-
d ion e (14b) a n d 7-Ch lor o-6-eth yl-1,4-d ih yd r o-5-n itr oqu i-
n oxa lin e-2,3-d ion e (14c): 1H NMR (DMSO-d6) 1.14 (m, 3H),
2.58 (q, J ) 7.4, 1H), 2.71 (q, J ) 7.4, 1H), 7.19 (s, 0.5H), 7.30
(s, 0.5H), 12.00 (mb, 0.5H), 12.17 (mb, 1.5H).
Na2SO4 and evaporated in vacuo, and the resulting oil was
dried further in vacuo to give 0.943 g (92%) of 11c as an oil:
1H NMR (CDCl3) 1.27 (t, J ) 7.5, 3H), 2.80 (q, J ) 7.5, 2H),
7.42 (s, 1H), 7.94 (s, 1H).
N-(4-Ch lor o-5-eth yl-2-n itr op h en yl)glycin e (12c). To a
solution of 11c (0.585 g, 2.66 mmol) in ethanol (10.0 mL) at
room temperature was added a solution of sodium glycinate
(0.260 g, 2.68 mmol) in water (2.5 mL), and the resulting
suspension was refluxed for 2 days. The solution was then
cooled to room temperature, and the precipitated red solid was
collected by filtration, washed with ethanol (4 mL), and dried
in vacuo to give 0.086 g (13%) of 12c as a red powder: 1H NMR
(DMSO-d6) 1.15 (t, J ) 7.2, 3H), 2.63 (q, J ) 7.2, 2H), 3.47 (d,
J ) 3.0, 2H), 6.77 (s, 1H), 7.97 (s, 1H), 8.77 (s, 1H). The
unreacted 2,5-dichloro-4-ethylnitrobenzene was recovered quan-
titatively from the filtrate.
7-Ch lor o-6-eth yl-3,4-dih ydr oqu in oxalin -2(1H)-on e (13c).
A suspension of 12c (0.082 g, 0.31 mmol) and SnCl2‚2H2O
(0.215 g, 0.953 mmol) in ethanol (1.0 mL) was refluxed for 45
min. It was then cooled to room temperature, and the
precipitated solid was collected by filtration, washed with
ethanol (1.0 mL), and dried in vacuo to yield 0.048 g (72%) of
13c as a light yellow powder: 1H NMR (DMSO-d6) 1.05 (t, J
) 7.2, 3H), 2.50-2.57 (m, 2H), 3.67 (s, 2H), 6.04 (s, 1H), 6.54
(s, 1H), 6.67 (s, 1H), 10.25 (s, 1H).
Eth yl N-(3-Br om o-4-eth yl-6-n itr oph en yl)glycin ate (12a).
A mixture of 452 mg (1.81 mmol) of 8a , 396 mg (2.87 mmol) of
K2CO3, and 2.0 mL (3.0 g, 18 mmol) of ethyl bromoacetate was
heated at 137 °C for 3 days. The mixture was cooled to room
temperature, 12 mL of aqueous 1 N NaOH was added
dropwise, and the mixture was stirred at room temperature
for 1 h. The mixture was filtered, washed with water, and
dried to leave 620 mg (100%) of a black solid: 1H NMR (CDCl3)
1.22 (t, J ) 7.6, 3H), 1.33 (t, J ) 7.2, 3H), 2.68 (q, J ) 7.4,
2H), 4.05 (d, J ) 5.4, 2H), 4.29 (m, 2H), 6.92 (s, 1H), 8.05 (s,
1H), 8.24 (sb, 1H). It was used for the next reaction without
purification.
6-Br om o-7-eth yl-3,4-dih ydr oqu in oxalin -2(1H)-on e (13a).
A solution of 620 mg (1.84 mmol) of 12a , 1.62 g (7.18 mmol) of
SnCl2‚2H2O, and 6 mL of absolute alcohol was refluxed for 4
h, and the mixture was cooled to room temperature. It was
evaporated, and the residue was treated with aqueous 1 N
NaOH to pH ) 10. The mixture was filtered, washed with
water, and dried to leave a solid. The solid was stirred with
20 mL of ethyl acetate for 1 h and filtered. The filtrate was
evaporated to leave 342 mg (72%) of a yellow solid: 1H NMR
(CDCl3) 1.17 (t, J ) 7.4, 3H), 2.63 (q, J ) 7.4, 2H), 3.80 (s,
1H), 3.95 (s, 2H), 6.55 (s, 1H), 6.86 (s, 1H), 7.83 (s, 1H). It
was used for the next reaction without purification.
7-Ch lor o-6-et h yl-1,4-d ih yd r o-5-n it r oq u in oxa lin e-2,3-
d ion e (14c). QX 14c was prepared from 13c in a manner
similar to that for 14a : mp 302-306 °C dec; 1H NMR (DMSO-
d6) 1.09 (t, J ) 7.2, 3H), 2.52 (q, J ) 7.2, 2H), 7.27 (s, 1H),
11.99 (s, 1H), 12.15 (s, 1H); HRMS calcd for C10H8ClN3O4
269.0203, found 269.0190. Anal. (C10H8ClN3O4) C, H; N:
calcd, 15.58; found, 14.40.
N-(4-Cya n o-5-m eth yl-2-n itr op h en yl)glycin e (12d ). Gly-
cine 12d was prepared from 4-chloro-2-methylbenzonitrile
(10d ) in two steps similar to the preparation of 12c: 1H NMR
(DMSO-d6) 2.37 (s, 3H), 3.51 (d, J ) 3.9, 1H), 6.85 (s, 1H),
8.38 (s, 1H), 9.13 (s, 1H).
7-Cya n o-3,4-d ih yd r o-6-m et h ylq u in oxa lin -2(1H )-on e
(13d ). To a stirred solution of 12d (0.100 g, 0.389 mmol) in
water (5.0 mL) at 100 °C was added sodium dithionite (0.540
g, 3.10 mmol) in two equal portions over 10 min. The resulting
white suspension was stirred at 100 °C for 1 h and then cooled
to room temperature. The solid was collected by filtration in
vacuo, washed with water (4 mL), and dried in vacuo to give
0.040 g (55%) of 13d as an off white powder: 1H NMR (DMSO-
d6) 2.22 (s, 3H), 3.83 (s, 2H), 6.51 (s, 1H), 6.81 (br s, 1H), 6.83
(s, 1H), 10.38 (s, 1H).
6-Br om o-7-et h yl-1,4-d ih yd r o-5-n it r oq u in oxa lin e-2,3-
d ion e (14a ). To a solution of 321 mg (1.24 mmol) of 13a in 6
mL of CF3CO2H stirred in an ice bath was added dropwise
1.5 mL of fuming HNO3, and the solution was stirred in an
ice bath for 1 h and at room temperature overnight. It was
added into 30 mL of ice-water, and the precipitate was
filtered, washed with water, and dried to leave 211 mg of a
yellow solid. The solid was stirred with 20 mL of 1 N NaOH
for 1 h and filtered. The filtrate was acidified by 6 N HCl to
pH ) 1. The precipitate was filtered, washed with water, and
1
dried to leave 174 mg (44%) of yellow solid: mp >290 °C; H
NMR (DMSO-d6) 1.16 (t, J ) 7.4, 3H), 2.72 (q, J ) 7.4, 2H),
7.19 (s, 1H), 12.20 (sb, 2H); HRMS calcd for C10H8BrN3O4
312.9695, found 312.9711. Anal. (C10H8BrN3O4‚0.1H2O) C, H,
N.
7-Cya n o-1,4-d ih yd r o-6-m eth yl-5-n itr oqu in oxa lin e-2,3-
d ion e (14d ). A solution of 13d (0.030 g, 0.16 mmol), fuming
HNO3 (0.070 mL), and TFA (0.700 mL) was stirred overnight
at room temperature. The resulting solution was poured into
ice water (7 mL), and the solid was collected by filtration in
vacuo, washed with water (7 mL), and dried in vacuo to give
0.027 g of a 2:1 mixture of 7-cyano-1,4-dihydro-6-methyl-7-
nitroquinoxaline-2,3-dione and 7-cyano-6-methyl-5-nitroqui-
noxalin-2(1H)-one as a yellow powder: 1H NMR (DMSO-d6)
2.34 (s, 3H), 2.41 (s, 3H), 7.47 (s, 1H), 7.75 (s, 1H), 8.35 (s,
1H), 12.21 (br s, 1H), 12.28 (s, 1H), 12.93 (s, 1H). This mixture
was again stirred in fuming HNO3 (0.10 mL) and TFA (1.0
mL) at room temperature for 72 h. The solution was poured
into ice water (10 mL), and the solid was collected by filtration
in vacuo, washed with water (10 mL), and dried in vacuo to
give 0.018 g (36%) of 14d as yellow powder: mp 341 °C dec;
1H NMR (DMSO-d6) 2.39 (s, 3H), 7.47 (s, 1H), 12.21 (br s, 1H),
12.27 (s, 1H); HRMS calcd for C9H6N4O4 246.0388, found
246.0375.
6-Ch lor o-7-et h yl-1,4-d ih yd r o-5-n it r oq u in oxa lin e-2,3-
d ion e (14b). QX 14b was prepared from 8b in a manner
similar to that of 14a : mp >330 °C; 1H NMR (DMSO-d6) 1.16
(t, J ) 7.4, 3H), 2.71 (q, J ) 7.6, 2H), 7.19 (s, 1H), 12.21 (mb,
2H); HRMS calcd for C10H8ClN3O4 269.0198, found 269.0196.
Anal. (C10H8ClN3O4) C, H, N.
2,5-Dich lor oeth ylben zen e (10c). A suspension of mossy
zinc (10 g) and mercuric chloride (0.5 g) in concentrated HCl
(0.5 mL) and water (5 mL) was shaken for ∼5 min. The
aqueous layer was then decanted. To the residue was added
concentrated HCl (7.5 mL) and water (7.5 mL), followed by
2′,5′-dichloroacetophenone (3.106 g, 16.43 mmol) and the
suspension was refluxed for 4 h during which time, hourly
addition of concentrated HCl (90.5 mL) was carried out. The
resulting suspension was cooled to room temperature, and the
aqueous layer was decanted. The residual solid was washed
with ether (3 × 30 mL). The aqueous layer was extracted with
ether (3 × 50 mL). The combined ether layer was washed with
brine, dried over anhydrous Na2SO4, and removed in vacuo.
The residue was dried further in vacuo to give 2.664 g of crude
product which was purified on silica gel (hexane) to give 0.813
g (28%) of 10c as a colorless liquid: 1H NMR (DMSO-d6) 1.23
(t, J ) 7.5, 3H), 2.73 (q, J ) 7.5, 2H), 7.09-7.27 (m, 3H).
2,5-Dich lor o-4-eth yln itr oben zen e (11c). To a stirred
solution of 10c (0.795 g, 4.68 mmol) in concentrated H2SO4
(4.5 mL) at 0 °C was added KNO3 (0.473 g, 4.68 mmol) in one
portion. The resulting pale yellow solution was allowed to
warm to room temperature and was stirred overnight at room
temperature. It was then poured into ice (80 g) and extracted
with ether (3 × 30 mL). The extract was dried over anhydrous
N-(4-Cya n o-2-n it r op h en yl)glycin e (12i). Glycine 12i
was prepared from 4-chloro-3-nitrobenzonitrile in a manner
similar to that of 12c: 1H NMR (DMSO-d6) 3.510 (d, J ) 3.9,
2H), 6.92 (d, J ) 9.0, 1H), 7.72 (dd, J ) 1.2, 9.0, 1H), 8.45 (d,
J ) 1.5, 1H), 9.19 (s, 1H).
7-Cya n o-3,4-d ih yd r oqu in oxa lin -2(1H)-on e (13i). QX-2-
one 13i was prepared from 12i in a manner similar to that
1
for 13d : H NMR (DMSO-d6) 3.86 (s, 2H), 0.641 (d, J ) 8.4,
1H), 6.90 (s, 2H), 7.12 (dd, J ) 1.2, 8.4, 1H), 10.47 (s, 1H).
7-Cyan o-1,4-dih ydr o-5-n itr oqu in oxalin e-2,3-dion e (14i).
QX 14i was prepared from 13i in a manner similar to that for

SAR of Substituted 1,4-Dihydroquinoxaline-2,3-diones
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5 737
14a : mp 339-40 °C dec; 1H NMR (DMSO-d6) 7.64 (d, J ) 1.2,
1H), 8.34 (d, J ) 1.5, 1H), 11.47 (s, 1H), 12.47 (s, 1H). Anal.
(C9H6N4O4‚0.3H2O) C, H, N.
solid was collected by filtration in vacuo, washed with water
(10 mL), and dried in vacuo to obtain 0.098 g (84%) of 18g as
a yellow powder: mp >323 °C; 1H NMR (DMSO-d6) 1.04 (t, J
) 7.2, 3H), 2.82 (q, J ) 7.5, 2H), 7.34 (s, 1H), 12.20 (s, 1H),
12.25 (s, 1H); HRMS calcd for C10H8ClN3O4S 300.9924, found
300.9919.
5-Am in o-7-ch lor o-1,4-d ih yd r o-6-m et h ylq u in oxa lin e-
2,3-d ion e (15). A solution of QX 14g18 (75 mg, 0.29 mmol)
and SnCl2‚2H2O (200 mg, 0.88 mmol) in ethanol (1.0 mL) was
refluxed for 6.5 h. The reaction mixture was cooled to room
temperature and allowed to stand overnight at room temper-
ature. The yellow solid was collected by filtration in vacuo,
washed with ethanol (1.5 mL), and dried in vacuo to give 54
P h a r m a cology
DCKA Bin d in g Assa y. The potency of the QXs as
inhibitors of [3H]DCKA binding was determined in rat
brain cortical membranes as previously described.19
Briefly, well-washed membranes (400 µg) were incu-
bated in 50 mM HEPES-KOH (pH 7.5) with 15 nM [3H]-
DCKA for 30 min at 0 °C in the presence of nine
concentrations of test compound added in 5 µL DMSO
(1% final). Nonspecific binding was defined using ACEA
1021 (10 µM). The assays were terminated by rapid
filtration, and filter-bound radioactivity was determined
by liquid scintillation spectrometry. IC50 values were
determined using the sigmoidal equation in Prism
(GraphPad).
Electr op h ysiology. QXs were made up in DMSO.
Ringer solutions were made by 300-1000-fold dilution
of DMSO stocks. Agonist concentration-response curves
were analyzed as described previously.8 For NR1a/2C
receptors the EC50 value for glycine was 0.17 µM and
the slope was 1.5 (n ) 6). For AMPA receptors
expressed by rat cerebral cortex poly(A+) RNA, the EC50
value was 5.9 µM and the slope was 2.0 (n ) 5).
Previously unreported Kb values in Table 2 were cal-
culated from three- or four-point concentration-inhibi-
tion curves using the equation
1
mg (81%) of 15 as a yellow powder: mp 323 °C dec; H NMR
(DMSO-d6) 2.09 (s, 3H), 5.47 (s, 2H), 6.46 (s, 1H), 11.15 (s,
1H), 11.72 (s, 1H). Anal. (C9H6N4O4‚0.88 H2O) C, H; N: calcd,
17.40; found, 16.32.
6-Am in o-7-flu or o-1,4-d ih yd r o-5-n itr oqu in oxa lin e-2,3-
d ion e (18a ). To a solution of 20 mg (0.082 mmol) of 1610 in
0.5 mL of DMSO-d6 was added 2 drops of 30% ammonium
hydroxide, and the solution was heated at 80 °C for 24 h. To
the mixture was added one more drop of 30% ammonium
hydroxide, and it was heated at 80 °C for 10 h. The mixture
was added into 4 mL of water and acidified with 2 N HCl to
pH ) 1. The precipitate was filtered, washed with water, and
dried to leave 15 mg (76%) of a yellow solid: mp 250 °C dec;
1H NMR (DMSO-d6) 7.20 (d, J ) 11.5, 1H), 7.23 (sb, 2H), 11.15
(mb, 1H), 12.0 (mb, 1H); 19F NMR -133.84 (d, J ) 11.6); HRMS
calcd for C8H5FN4O4 240.0290, found 240.0294. Anal. (C8H5-
FN4O4‚0.6H2O) C, H; N: calcd, 22.32; found, 21.00.
7-F lu or o-1,4-d ih yd r o-6-m et h oxy-5-n it r oq u in oxa lin e-
2,3-d ion e (18b). To a mixture of 162 mg (0.667 mmol) of 16
and 162 mg (2.85 mmol) of sodium methoxide was added 4
mL of DMSO, and the solution was stirred at room tempera-
ture overnight. The solution was diluted with 15 mL of water
and acidified with 2 N HCl to pH ) 4. The precipitate was
filtered, washed with water, and dried to leave 157 mg (92%)
1
of a yellow solid: mp >300 °C; H NMR (DMSO-d6) 3.90 (s,
3H), 7.16 (d, J ) 11.6, 1H), 11.97 (mb, 1H), 12.14 (s, 1H); 19F
NMR -134.48 (mb). Anal. (C9H6FN3O5) C, H, N.
IC50
6-Eth oxy-7-flu or o-1,4-d ih yd r o-5-n itr oqu in oxa lin e-2,3-
d ion e (18c). QX 18c was prepared from 16 and sodium
ethoxide in a manner similar to the preparation of 18b: mp
Kb )
{2 + ([agonist]f/EC50)n}1/n - 1
1
293-5 °C; H NMR (DMSO-d6) 1.24 (t, J ) 7.0, 3H), 4.14 (q,
J ) 7.2, 2H), 7.15 (d, J ) 11.7, 1H), 11.98 (s, 1H), 12.14 (s, 1);
19F NMR -134.06 (mb); HRMS calcd for C10H8FN3O5 269.0443,
found 269.0454. Anal. (C10H8FN3O5‚0.2H2O) C, H, N.
where IC50 is the concentration of QX that reduces the
control response by 50%, [agonist]f is the fixed dose of
agonist used to construct the inhibition curve, EC50 is
the concentration of agonist evoking a half-maximal
response, and n is the slope of the agonist concentra-
tion-response relation.27
Mou se Ma xim u m Electr osh ock -In d u ced Seizu r e
Tests. Procedures for the mouse MES assay were as
reported previously.11,14 QXs 2, 9a , 14a ,b,e-h , and 21
were dissolved in 0.05 M Tris and were tested for
anticonvulsant effects at the peak of activity which
occurred 2 min after iv administration. For QX 14a the
onset of activity was slower and the compound was
tested at a peak of activity occurring 15 min after
injection. ED50 values were determined by Litchfield
and Wilcoxon analysis.
6-n -Bu t oxy-7-flu or o-1,4-d ih yd r o-5-n it r oq u in oxa lin e-
2,3-d ion e (18d ). To a mixture of 25 mg (1.0 mmol) of NaH
and 50 mg (0.67 mmol) of 1-butanol was added 1.5 mL of
DMSO, and the mixture was stirred for 1 h. To the resulting
solution was added 27 mg (0.11 mmol) of 16, and the solution
was stirred at room temperature for 5 days. The solution was
diluted with 3 mL of water and acidified with 2 N HCl to pH
) 1. The precipitate was filtered, washed with water, and
dried to leave 11 mg (33%) of a yellow solid: mp 290-292 °C;
1H NMR (DMSO-d6) 0.88 (t, J ) 7.4, 3H), 1.36 (m, 2H), 1.60
(m, 2H), 4,08 (t, J ) 6.3, 2H), 7.15 (d, J ) 11.54, 1H), 11.98
(m, 1H), 12.12 (m, 1H); HRMS calcd for C12H12FN3O5 297.0755,
found 297.0757. Anal. (C12H12FN3O5‚0.2H2O) C, H, N.
7-F lu or o-1,4-d ih yd r o-5-n itr o-6-(3-p h en ylp r op oxy)qu i-
n oxa lin e-2,3-d ion e (18e). QX 18e was prepared in a manner
1
similar to that for 18d : mp 280-2 °C; H NMR (DMSO-d6)
log P Mea su r em en t. log P values for QXs 2, 9a ,
and 14a,e,g were obtained by classic shake-flask method
in octanol vs pH 7.4 buffer. Eight QXs with log P values
between -0.97 and 1.04 obtained from classic shake-
flask method were chromatographed at three concentra-
tions, and the log K′0 for each compound was deter-
mined. A linear plot of log K′0 vs log P was made from
these eight QXs and used for the estimation of log P
values from log K′0 values. log P values for QXs 14f,h
and 21 were obtained by the HPLC method and esti-
mated from their log K′0 values. The log P value for
QX 14b was calculated on the basis of the log P of 14a
(1.04) and the log P difference between 14f and 14g
(0.12).
1.92 (m, 2H), 2.65 (t, J ) 7.8, 2H), 4.10 (t, J ) 6.1, 2H), 7.14-
7.31 (m, 6H), 11.98 (m, 1H), 12.131 (s, 1H); 19F NMR -133.87
(mb); HRMS calcd for C17H14FN3O5 359.0911, found 359.0927.
Anal. (C17H14FN3O5‚0.2H2O) C, H, N.
7-Ch lor o-1,4-d ih yd r o-6-m et h oxy-5-n it r oq u in oxa lin e-
2,3-d ion e (18f). QX 18f was prepared from 1710 in a manner
similar to that for 18b: mp > 316 °C; 1H NMR (DMSO-d6) 3.82
(s, 3H), 7.28 (s, 1H), 12.08 (s, 1H), 12.12 (s, 1H). Anal. (C9H6-
ClN3O5‚0.1DMSO) C, H, N.
7-Ch lor o-1,4-d ih yd r o-5-n itr o-6-th ioeth oxyqu in oxa lin e-
2,3-d ion e (18g). A solution of 17 (0.100 g, 0.385 mmol) and
ethanethiol sodium salt (0.130 g, 1.55 mmol) in DMSO (1.0
mL) was stirred at room temperature for 24 h. The resulting
solution was diluted with water (5 mL) and acidified with
concentrated HCl (4 or 5 drops) to pH ∼5. The precipitated

738 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 5
Cai et al.
ones: Novel Potent Glycine Site NMDA Antagonists Showing
Neuroprotectant Activity in Stroke Models. Presented at the
211th ACS National Meeting, New Orleans, LA, 1996. Division
of Medicinal Chemistry Abstract 105.
Ack n ow led gm en t. We thank Dr. Yan Ni and Dr.
Ricardo Miledi (University of California, Irvine) for the
generous gift of rat cerebral cortex poly(A)+ RNA used
in this study, Dr. P. H. Seeburg (Heidelburg University,
Heidelburg, Germany) for the cDNAs encoding rat NR1
and NR2 subunits, Dr. J ohn Guastella (CoCensys Inc.)
for preparation of cRNAs encoding NMDA receptor
subunits, and Mr. J oe Richman (CoCensys, Inc.) for
measurement of log P. Financial support was provided
in part by Acea Pharmaceuticals, Inc., a wholly owned
subsidiary of CoCensys, Inc., and the National Institute
of Drug Abuse (DA-06726).
(10) Keana, J . F. W.; Kher, S. M.; Cai, S. X.; Dinsmore, C. M.; Glenn,
A. G.; Guastella, J .; Huang, J . C.; Lu, Y.; Ilyin, V.; Lu, Y.;
Mouser, P. L.; Woodward, R. M.; Weber, E. Synthesis and
Structure-Activity Relationships of Substituted 1,4-Dihydroqui-
noxaline-2,3-diones (QXs): Antagonists of the Glycine Site on
the NMDA Receptor and of Non-NMDA Glutamate Receptors.
J . Med. Chem. 1995, 38, 4367-4379.
(11) Woodward, R. M.; Huettner, J . E.; Tran, M.; Guastella, J .; Keana,
J . F. W.; Weber, E. Pharmacology of 5-Chloro-7-trifluoromethyl-
1,4-dihydro-2,3-quinoxalinedione: A Novel Systemically Active
Ionotropic Glutamate Receptor Antagonist. J . Pharmacol. Exp.
Ther. 1995, 275, 1209-1218.
(12) (a) Lufty, K.; Shen, K.-Z.; Kwon, I.-S.; Cai, S. X.; Woodward, R.
M.; Keana, J . F. W.; Weber, E. Blockade of Morphine Tolerance
by 5-Nitro-6,7-dimethyl-2,3-quinoxalinedione (Acea-1328),
A
Refer en ces
Novel NMDA Receptor/Glycine Site Antagonist. Eur. J . Pharm.
1995, 273, 187-189. (b) Lufty, K.; Shen, K.-Z.; Woodward, R.
M.; Weber, E. Inhibition of Morphine Tolerance by NMDA
Receptor Antagonists in the Formalin Test. Brain Res. 1996,
731, 171-181.
(1) Knopfel, T.; Kuhn, R.; Allgeier, H. Metabotropic Glutamate
Receptors: Novel Targets for Drug Development. J . Med. Chem.
1995, 38, 1417-1426.
(2) Doble A. Excitatory Amino Acid Receptors and Neurodegenera-
tion. Therapie 1995, 50, 319-337.
(13) Tran, M.; Lutfy, K.; Xu, Z.; Cai, S. X.; Keana, J . F. W.; Weber,
E. The Anticonvulsant Structure-Activity Relationship of a
Series of Novel NMDA Receptor/Glycine Site Antagonists in the
Maximal Electroshock Seizure (MES) Model in Mice. Presented
at 25th Annual Meeting of Society for Neuroscience, San Diego,
CA. November, 1995. Abstract 84.18.
(14) Ilyin, V. I.; Whittemore, E. R.; Tran, M.; Shen, K.-Z.; Cai, S. X.;
Kher, S. M.; Keana, J . F. W.; Weber, E.; Woodward, R. M.
Pharmacology of ACEA-1416: A Potent Systemically Active
N-Methyl-D-asparatate Receptor Glycine Site Antagonist. Eur.
J . Pharmacol. 1996, 310, 107-114.
(15) Lambooy, P.; Lambooy, J . P. Synthesis and Biological Activities
of 7-Ethyl-8-chloro-10-(1′-D-ribityl)isoalloxazine and 7-Chloro-
8-ethyl-10-(1′-D-ribityl)isoalloxazine, Analogs of Riboflavin. J .
Med. Chem. 1973, 16, 765-770.
(16) Lambooy, J . P. The Synthesis of 4,5-Diethyl-o-phenylenediamine
through the Nitration of o-Diethylbenzene. J . Am. Chem. Soc.
1949, 71, 3756.
(3) J ohnson, J . W.; Ascher, P. Glycine Potentiates the NMDA
Response in Cultured Mouse Brain Neurons. Nature (London)
1987, 325, 529-531.
(4) (a) Kemp, J . A.; Leeson, P. D. The Glycine Site of the NMDA
Receptor-Five Years on. Trends Pharmacol. Sci. 1993, 14, 20-
25. (b) Leeson, P. D. Glycine-site N-Methyl-D-aspartate Receptor
Antagonists. In Drug Design for Neuroscience; Kozikowski, A.
P., Ed.; Raven Press: New York, 1993; Chapter 13.
(5) (a) McQuaid, L. A.; Smith, E. C. R.; Lodge, D.; Pralong, E.; Wikel,
J . H.; Calligaro, D. O.; O’Malley, P. J . 3-Phenyl-4-hydroxyquino-
line-2(1H)-ones: Potent and Selective Antagonists at the Strych-
nine-Insensitive Glycine Site on the N-Methyl-D-aspartate Re-
ceptor Complex. J . Med. Chem. 1992, 35, 3423-3425. (b) Gray,
N. M.; Dappen, M. S.; Cheng, B. K.; Cordi, A. A.; Biesterfeldt,
J . P.; Hood, W. F.; Monahan, J . B. Novel Indole-2-carboxylates
as Ligands for the Strychnine-Insensitive N-Methyl-D-aspartate-
Linked Glycine Receptor. J . Med. Chem. 1991, 34, 1283-1292.
(c) Swartz, K. J .; Koroshetz, W. J .; Rees, A. H.; Huettner, J . E.
Competitive Antagonism of Glutamate Receptor Channels by
Substituted Benzazepines in Cultured Cortical Neurons. Mol.
Pharmacol. 1992, 41, 1130-1141. (d) Nagata R.; Tanno, N.;
Kodo, T.; Ae, N.; Yamaguchi, H.; Nishimura, T.; Antoku, F.;
Tatsuno, T.; Kato, T.; Tanaka, Y.; Nakamura, M. Tricyclic
Quinoxalinediones: 5,6-Dihydro-1H-pyrrolo[1,2,3-de]quinoxa-
line-2,3-diones and 6,7-Dihydro-1H,5H-pyrido[1,2,3-de]quinoxa-
line-2,3-diones as Potent Antagonists for the Glycine Binding
Site of the NMDA Receptor. J . Med. Chem. 1994, 37, 3956-
3968. (e) Rowley, M.; Leeson, P. D.; Stevenson, G. I.; Moseley,
A. M.; Stansfield, I.; Sanderson, I.; Robinson, L.; Baker, R.;
Kemp. J . A.; Marshall, G. R.; Foster, A. C.; Grimwood, S.;
Tricklebank, M. D.; Saywell, K. L. 3-Acyl-4-hydroxyquinolin-
2(1H)-ones. Systemically Active Anticonvulsants Acting by
Antagonism at the Glycine Site of the N-Methyl-D-aspartate
Receptor Complex. J . Med. Chem. 1993, 36, 3386-3396. (f)
Carling, R. W.; Leeson, P. D.; Moseley, A. M.; Baker, R.; Foster,
A. C.; Grimwood, S.; Kemp, J . A.; Marshall, G. R. 2-Carboxytet-
rahydroquinolines. Conformational and Stereochemical Require-
ments for Antagonism of the Glycine Site on the NMDA
Receptor. J . Med. Chem. 1992, 35, 1942-1953. (g) Leeson, P.
D.; Carling, R. W.; Moore, K. W.; Moseley, A. M.; Smith, J . D.;
Stevenson, G.; Chan, T.; Baker, R.; Foster, A. C.; Grimwood, S.;
Kemp, J . A.; Marshall, G. R.; Hoogsteen, K. 4-Amido-2-car-
boxytetrahydroquinolines. Structure-Activity Relationships for
Antagonism at the Glycine Site of the NMDA Receptor. J . Med.
Chem. 1992, 35, 1954-1968.
(17) Wulfman, D. S.; Cooper, C. F. Monoreduction of Dinitroarenes
with Iron/Acetic Acid. Synthesis 1978, 924.
(18) Kher, S. M.; Cai, S. X.; Weber, E.; Keana, J . F. W. Regiospecific
Oxidative Nitration of 3,4-Dihydro-6,7-disubstituted Quinoxalin-
2(1H)-ones Gives 1,4-Dihydro-5-nitro-6,7-disubstituted Quinoxa-
line-2,3-diones, Potent Antagonists at the NMDA/Glycine Site.
J . Org. Chem. 1995, 60, 5838-5842.
(19) Cai, S. X.; Zhou, Z.-L.; Huang, J .-C.; Whittemore, E. R.; Egbu-
woku, Z. O.; Lu, Y.; Hawkinson, J . E.; Woodward, R. M.; Weber,
E.; Keana, J . F. W. Synthesis and Structure-Activity Relation-
ships of 1,2,3,4-Tetrahydroquinoline-2,3,4-trione 3-Oximes: Novel
and Highly Potent Antagonists for the NMDA Receptor Glycine
Site. J . Med. Chem. 1996, 39, 3248-3255.
(20) Monyer, H. R.; Sprengel, R.; Schoepfer, A.; Herb, M.; Higuchi,
H.; Lomeli, N.; Burnashev, B.; Sakmann B.; Seeburg, P. H.
Heteromeric NMDA Receptors: Molecular and Functional Dis-
tinction of Subtypes. Science (Washington D.C.) 1992, 256,
1217-1221.
(21) Leeson, P. D.; Baker, R.; Carling, R. W.; Curtis, N. R.; Moore,
K. W.; Williams, B. J .; Foster, A. C.; Donald, A. E.; Kemp. J . A.;
Marshall, G. R. Kynurenic Acid Derivatives. Structure-Activity
Relationships for Excitatory Amino Acid Antagonism and Iden-
tification of Potent and Selective Antagonists at the Glycine Site
on the N-Methyl-D-aspartate Receptor. J . Med. Chem. 1991, 34,
1243-1252.
(22) Bigge, C. F.; Malone, T. C.; Boxer, P. A.; Nelson, C. B.; Ortwine,
D. F.; Schelkun, R. M.; Retz, D. M.; Lescosky, L. J .; Borosky, S.
A.; Vartanian, M. G.; Schwarz, R. D.; Campbell, G. W.; Ro-
bichaud, L. J .; Watjen, F. Synthesis of 1,4,7,8,9,10-Hexahydro-
9-methyl-6-nitropyrido[3,4-f]-quinoxaline-2,3-dione and Related
Quinoxalinediones: Characterization of R-Amino-3-hydroxy-5-
methyl-4-isoxazolepropionic Acid (and N-Methyl-D-aspartate)
Receptor and Anticonvulsant Activity. J . Med. Chem. 1995, 38,
3720-3740.
(23) Hansch, C.; Bjorkroth, J . P.; Leo, A. Hydrophobicity and Central
Nervous System Agents: On the Principle of Minimal Hydro-
phobicity in Drug Design. J . Pharm. Sci. 1987, 76, 663-687.
(24) Hawkinson, J . E. Unpublished results.
(6) Leeson, P. D.; Iversen, L. L. The Glycine Site on the NMDA
Receptor: Structure-Activity Relationships and Therapeutic
Potential. J . Med. Chem. 1994, 37, 4053-4067.
(7) Kulagowski, J . J .; Baker, R.; Curtis, N. R.; Leeson, P. D.; Mawer,
I. M.; Moseley, A. M.; Ridgill, M. P.; Rowley, M.; Stansfield, I.;
Foster, A. G.; Grimwood, S.; Hill, R. G.; Kemp, J . A.; Marshall,
G. R.; Saywell, K. L.; Tricklebank, M. D. 3′-(Arylmethyl)- and
3′-(Aryloxy)-3-phenyl-4-hydroxyquinoline-2(1H)-ones: Orally Ac-
tive Antagonists of the Glycine Site on the NMDA Receptor. J .
Med. Chem. 1994, 37, 1402-1405.
(8) Woodward, R. M.; Huettner, J . E.; Guastella, J .; Keana, J . F.
W.; Weber, E. In Vitro pharmacology of ACEA-1021 and ACEA-
1031: Systemically Active quinoxalinediones with High Affinity
and Selectivity for N-Methyl-D-aspartate Receptor Glycine Sites.
Mol. Pharmacol. 1995, 47, 568-581.
(9) Bare, T. M.; Sparks, R. B.; Smith, R. W.; Schreffler-Smith, C.
M.; Draper, C. W.; Rhile, I. J .; Empfield, J . R.; Forst, J . M.;
Herzog, K. J .; Pastore, W. M.; J ackson, P. F.; Garcia-Davenport,
L.; Davenport, T. W.; Chapdelaine, M. J .; McLaren, C. D.; Pullan,
L. M.; Goldstein, J . M.; Patel, J . B. Pyridazino[4,5-b]quinolinedi-
(25) Daniell, G.; Marek, P.; Weber, E. Unpublished results.
(26) Zhou, Z.-L.; Weber, E.; Keana, J . F. W. Acetoxylation of 6,7-
Dialkoxy-Substituted 1,4-Dihydroquinoxaline-2,3-diones (QXs)
Using Fuming Nitric Acid in Acetic Acid: A Facile Synthesis of
5-Acyloxy-6,7-dialkoxy QXs. Tetrahedron Lett. 1995, 36, 7583-
7586.
(27) Leff, P.; Dougall, I. G. Further Concerns over Cheng-Prusoff
Analysis. Trend. Pharmacol. Sci. 1993, 14, 110-112.
J M960654B