Substituted indole-2-carboxylates as in vivo potent antagonists acting as the strychnine-insensitive glycine binding site

Article abstract of DOI:10.1021/jm960644a

A series of indole-2-carboxylates bearing suitable chains at the C-3 position of the indole nucleus was synthesized and evaluated in terms of in vitro affinity using [³H]glycine binding assay and in vivo potency by inhibition of convulsions ind

Full text of DOI:10.1021/jm960644a

J . Med. Chem. 1997, 40, 841-850
841
Su bstitu ted In d ole-2-ca r boxyla tes a s in Vivo P oten t An ta gon ists Actin g a s th e
Str ych n in e-In sen sitive Glycin e Bin d in g Site
Romano Di Fabio,* Anna M. Capelli, Nadia Conti, Alfredo Cugola, Daniele Donati, Aldo Feriani, Paola Gastaldi,
Giovanni Gaviraghi, Cheryl T. Hewkin, Fabrizio Micheli, Andrea Missio, Manolo Mugnaini, Angelo Pecunioso,
Anna M. Quaglia, Emiliangelo Ratti, Luciana Rossi, Giovanna Tedesco, David G. Trist, and Angelo Reggiani
GlaxoWellcome S.p.A., Medicines Research Centre, Via A. Fleming 4, 37100 Verona, Italy
Received September 12, 1996X
A series of indole-2-carboxylates bearing suitable chains at the C-3 position of the indole nucleus
was synthesized and evaluated in terms of in vitro affinity using [3H]glycine binding assay
and in vivo potency by inhibition of convulsions induced by N-methyl-D-aspartate (NMDA) in
mice. 3-[2-[(Phenylamino)carbonyl]ethenyl]-4,6-dichloroindole-2-carboxylic acid (8) was an
antagonist at the strychnine-insensitive glycine binding site (noncompetitive inhibition of the
binding of [3H]TCP, pA2 ) 8.1) displaying nanomolar affinity for the glycine binding site (pKi
) 8.5), coupled with high glutamate receptor selectivity (>1000-fold relative to the affinity at
the NMDA, AMPA, and kainate binding sites). This indole derivative inhibited convulsions
induced by NMDA in mice, when administered by both iv and po routes (ED50 ) 0.06 and 6
mg/kg, respectively). The effect of the substituents on the terminal phenyl ring of the C-3 side
chain was investigated. QSAR analysis suggested that the pKi value decreases with lipophilicity
and steric bulk of substituents and increases with the electron donor resonance effect of the
groups present in the para position of the terminal phenyl ring. According to these results
the terminal phenyl ring of the C-3 side chain should lie in a nonhydrophobic pocket of limited
size, refining the proposed pharmacophore model of the glycine binding site associated with
the NMDA receptor.
In tr od u ction
In the last decade the role of excitatory amino acid
receptors in several disorders of the central nervous
system (CNS) has been extensively investigated. It is
now generally agreed that in ischaemic or hypoxic
conditions1 such as stroke, hypoglycemia, and traumatic
head and spinal cord injuries, the increased amount of
glutamate,2 released into the synaptic clefts, causes the
overstimulation of the ionotropic receptor complex
responding to the exogenous agonist N-methyl-D-aspar-
tate (NMDA). This key event results in a massive influx
of Ca2+ into the postsynaptic neurons leading ultimately
to cell death3 through the activation of several neuro-
toxic cascades. Intervention with competitive and
noncompetitive NMDA antagonists,4 to block the open-
ing of the ion channel associated with the NMDA
receptor,5 might be of therapeutic benefit. Among the
endogenous modulators of the NMDA receptor, glycine
has gained considerable interest in view of both its
functional role in the activation of the ionotropic recep-
tor complex as co-agonist of glutamate6 and the greater
therapeutic index seen for one of the first glycine
antagonists identified, the 7-chlorokynurenic acid (7-
Cl KA, 1) depicted in Figure 1, compared to different
series of NMDA antagonists.7 Therefore, in the last few
years the glycinergic site associated with the NMDA
receptor complex was perceived as a unique target for
F igu r e 1. Structure of various glycine antagonists.
medicinal chemistry, and considerable effort has been
devoted to find potent and selective ligands, resulting
in the identification of several classes of glycine antago-
nists.8 A selected number of these derivatives is shown
in Figure 1. These templates have been reported to be
endowed with nanomolar in vitro affinity at the strych-
nine-insensitive glycine binding site, but with the
exception of compounds 5 and 6, poor in vivo activity
was claimed in a variety of animal models, probably due
to their insufficient CNS bioavailability.9
* Corresponding author. Fax: 39-45-9218196. E-mail: rdf26781@
ggr.co.uk.
The indole-2-carboxylates of general structure 7,
depicted in Figure 2, represent a different class of
X Abstract published in Advance ACS Abstracts, February 1, 1997.
S0022-2623(96)00644-9 CCC: $14.00 © 1997 American Chemical Society

842 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6
Di Fabio et al.
for the glycine site and selectivity for glutamate recep-
tors; (b) “in vitro” functional antagonism studies to
evaluate potency and activity; (c) “in vivo” anticonvul-
sant activity.
Affinity for the glycine binding site associated with
the NMDA receptor channel complex was measured by
inhibition of the binding of [3H]glycine to crude synaptic
membranes, prepared from adult rat cerebral cortex as
described by Kishimoto et al.13 The incubation (20 min,
4 °C) was carried out in Tris/citrate (50 mM, pH 7.10)
using 20 nM [3H]glycine. Data of displacement experi-
ments, to determinate the inhibition constants (Ki) of
displacer ligands, were analyzed using a nonlinear
curve-fitting program LIGAND.14 Ki values were mea-
sured from at least six-point inhibition curves and are
the geometric mean of at least three different experi-
ments. The standard error associated with the mean
was less than 0.05.
F igu r e 2. Indole-2-carboxylates.
glycine antagonists widely investigated during the last
years by several research groups,10 as a consequence of
the preliminary evidence that the modulation at the C-3
position of the indole nucleus by different substituents
could considerably enhance the affinity of this template
at the glycine binding site.
The selectivity of NCE toward the glutamate binding
site of the NMDA receptor channel complex, AMPA, and
kainate ionotropic glutamate receptors was assessed by
inhibition of the binding to rat cortical membranes of
[3H]CPP, [3H]AMPA, and [3H]kainic acid, respectively.
In this case experiments were performed as reported
by Van Amsterdam et al.,15 Giberti et al.,16 and Honore´
et al.17
Functional antagonism at the glycine binding site was
demonstrated by the ability of NCE to antagonise the
glycine-induced enhancement of the binding of the
channel-blocking agent [3H]TCP18 in a glycine-sensitive
extensively washed rat cortical preparation.19 Binding
of [3H]TCP was carried out (2 h, 30 °C) in Tris/HCl (5
mM, pH 7.7) and in the presence of glutamic acid (1
µM). Nonspecific binding was determined by addition
of (+)-MK801 (30 µM). In the presence of increasing
concentrations of NCE, parallel rightward shifts of the
glycine concentration-response curves could be ob-
served, with no depression of the maximum response.
The antagonist potency (pA2) was calculated according
to the method of Arunlakshana and Schild.20
NCE were evaluated in vivo assessing their anticon-
vulsant effect in male CD-1 mice (18-29 g), both by iv
and po administration.7c Convulsions were induced by
icv injection of N-methyl-D-aspartate (1 nM/mouse) 1
min and 1 h after the iv and the po administration of
the NCE, respectively. Animals were observed for the
occurrence of generalized seizures during the first 30
min after NMDA treatment and considered protected
if convulsions did not occur within this period. The
percentage of animals showing convulsions in each
treatment group was recorded and ED50 values, i.e.
protecting dose of NCE providing 50% anticonvulsant
effect, with their 95% confidence limits were estimated.
The present paper deals with the synthesis and the
pharmacological characterization of a novel series of
indole-2-carboxylates substituted at the C-3 position
with suitable R,â-unsaturated side chains. This class
of glycine antagonists showed nanomolar affinity for the
glycine binding site coupled with high receptor selectiv-
ity and high in vivo potency in the NMDA-induced
convulsions model in mice, both by iv and po route. In
particular compound 8, GV150526A, shown in Figure
2, represents one of the most promising antagonists
acting at the strychnine-insensitive glycine binding site
identified to date.
Syn th esis
Compounds 14 have been prepared starting from the
known 3-unsubstituted indole derivative 910b following
both synthetic routes A and B shown in Scheme 1.
Formylation at position C-3 of the indole nucleus,
performed according to the Vilsmaier-Haack procedure,
afforded the intermediate 10 in high yield. This com-
pound was submitted to the Wittig-type olefination
reaction using a series of phosphorane derivatives,
widely substituted at the terminal phenyl ring, to give
compounds 11 with high regiocontrol in the formation
of the E double bond (route A). Alternatively, 10 was
transformed into the R,â-unsaturated acid intermediate
13, easily obtained from intermediate 12 by the chemose-
lective deprotection of the tert-butyl ester group (route
B). Amidation was then performed using different
synthetic methods.11 In particular, it is worth noting
that, in this case, the activation of the carboxyl group
through the formation of the corresponding 2-pyridyl
thioester, generated in situ by the mild “oxidation-
reduction” condensation reaction12 in the presence of
2,2′-dipyridyl disulfide and PPh3, was found to be
particularly efficient also in the case of poorly nucleo-
philic aromatic amines, affording the amide derivatives
11 in high yield (either from the isolated 2-pyridyl
thioester intermediate or from the acid 13 using a one-
pot procedure). Finally, the basic hydrolysis of the
2-ethyl ester protecting group gave quantitative yields
of the target compounds 14.
Resu lts a n d Discu ssion
Our interest in the indole-2-carboxylate derivatives
originated with the observation that the 3-unsubsti-
tuted-4,6-dichloro derivative 7 (R ) H) shown in Figure
2 was endowed with good glutamate receptor selec-
tivity.10b However, its in vitro affinity for the glycine
binding site was low (pKi ) 5.7) compared to the most
potent glycine antagonist hitherto identified, due to the
lack of the necessary hydrogen bond accepting group as
predicted by the proposed 3D-pharmacophore model of
the glycine binding site.21
Biology
The biological evaluation of the new chemical entities
(NCE) was performed according to the following screen-
ing sequence: (a) binding assay to evaluate the affinity

Indole-2-carboxylates as Potent Antagonists
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6 843
Sch em e 1a
a
Reagents and conditions: (a) PhN(Me)CHO, POCl3, ClCH2CH2Cl; (b) Ph3PdCHCOO-t-Bu, THF; (c) HCO2H; (d) 2-Aldrithiol, PPh3,
ArNH2, THF; (e) R3PdCHCONHAr; (f) NaOH, EtOH, H2O.
known derivatives 7 and 15,10b were synthesized and
Ta ble 1. Substitution at Position C-3
then characterized in terms of in vitro affinity at the
glycine binding site in rat cortical membrane prepara-
tions as previously described. On the basis of the
results summarized in Table 1, the following observa-
tions can be addressed: (a) all of the compounds tested
ED50 (mg/kg)b
iv
showed an affinity for the glycine binding site which
was found to be greater than the C-3 unsubstituted
analogue 7 (R ) H), confirming the validity of the
proposed 3D-pharmacophore model and in particular
the need of a suitable H-bond acceptor group on the C-3
side chain; (b) the affinity of ligands possessing an R,â-
unsaturated substituent directly conjugated with the
indole nucleus tends to be higher with respect to the
corresponding saturated system (compounds 8 vs 16 and
17 vs 15). This effect could be due to the greater
conformational entropy of compounds 8 and 17 vs 16
and 15, respectively, and both to the higher electron
density on the oxygen atom of the carbonyl group and
the improved H-bond donor character of the indole NH
group; (c) the observed 270-fold lower affinity at the
glycine binding site of the sulfonamide derivative 19
with respect to the amide analogue 8 confirms the
importance of a correct spatial orientation of the H-bond
accepting group and the reduced H-bond accepting
character of the sulfonamide group with respect to the
corresponding amide carbonyl group; (d) the homologa-
tion of the terminal amide (entry 20 vs entry 8) resulted
in a large reduction of affinity as a consequence of both
the increased steric bulk of the terminal substituent
and/or the presence of a phenyl ring not directly
conjugated with the nitrogen of the amide group; (e) the
no.
R
pKia
po
6
7
8
H
5.7
8.5
CHdCHCONHPh
0.06
(0.005 - 0.42) (4.40-7.94)
15 CH2CH2COOH
16 CH2CH2CONHPh
7.4
7.6
>10
3.70
(1.51 - 12.12)
17 CHdCHCOOH
7.7
17.21
>200
>200
(15.31 - 19.73)
18 CHdCHCOO-t-Bu
19 CHdCHSO2NHPh
20 CHdCHCONHCH2Ph 6.9
6.3
6.1
>30
3
5
6
8.2
9.0
8.1
0.21
(0.14 - 0.31)
0.78
(0.45 - 1.63)
0.21
(0.10 - 0.57)
a
b
Inhibition of binding of [3H]Gly (ref 13). NMDA-induced
convulsion model (ref 7c).
Suitable C-3 side chains, bearing different H-bond
accepting groups and terminal lypophilic substituents,
were designed with the aim of further increasing the
affinity of the indole-2-carboxylate template for the
glycinergic site, giving rise to a short series of saturated
and unsaturated indole derivatives, as reported in Table
1. All of the proposed compounds, in addition to the

844 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6
Di Fabio et al.
F igu r e 3. Superimposition of compound 8 with the pharmacophore model of the glycine binding site.
tert-butyl ester derivative 18 showed a significant
reduction in affinity with respect to compound 17 in
view of both the different H-bond accepting character
of the carboxyl group and the steric bulk associated with
the terminal lipophilic substituent.
the binding of [3H]glycine. Moreover, no evidence of
agonist activity was detected.
As reported in Table 1, the most potent in vitro indole
derivatives of this series were tested as antagonists of
NMDA-induced convulsions in mice both by iv and by
po routes. Compound 8, the glycine antagonist endowed
with the highest in vitro affinity for the glycine binding
site, was also found to be the most potent compound in
vivo, showing considerable anticonvulsant activity when
tested in the range of doses between 0.001-3 mg/kg iv
and 1-100 mg/kg po, according to the general procedure
described above. In both cases, dose-dependent inhibi-
tion of convulsions reached 80% at 3 mg/kg iv and 90%
at 100 mg/kg po, respectively. The estimated ED50 by
iv route was 0.06 mg/kg (0.005-0.42 mg/kg), whereas
the ED50 by oral route was 6 mg/kg (4.4-7.94 mg/kg).
As shown in Table 1, reference compounds 3, 5, and 6
were found to be slightly less active than 8.24
The excellent in vivo activity of 8 in mice was
confirmed by performing the same studies in rats by
the iv route. The compound was tested in the range of
doses between 0.03 and 3 mg/kg and a dose-dependent
inhibition of NMDA-induced convulsions (up to 90%
blockade) was observed, with an estimated ED50 of 0.38
mg/kg (0.03-1.34 mg/kg).
Su bstitu tion of th e Ter m in a l P h en yl Rin g. After
the identification of the indole derivative 8, the effect
of the substitution on the aromatic ring of the C-3 side
chain was investigated with the aim of further improv-
ing the affinity of this class of derivatives for the glycine
binding site associated with the NMDA receptor.
According to the Hansch approach, substituents are
described in terms of their electronic and steric effects
and lipophilicity. Compounds were chosen to ensure
maximum variance and minimum collinearity between
parameters characterizing the physicochemical proper-
ties of the substituents. The set of traditional descrip-
tors25-27 shown in Table 2 was considered, and MRA
In conclusion, the presence of an R,â-unsaturated side
chain in which both the carbonyl group and the terminal
phenyl ring play a key role in recognition seems to be
crucial to maximize the affinity of these indole templates
for the glycinergic site associated with the NMDA
receptor. In Figure 3 is shown the superimposition of
compound 8 with the known pharmacophore of the
glycine binding site,21 further confirming the validity
of the proposed model. This model was built up in house
using the active analog approach (AAA)22 implemented
within Sybyl 6.023a on the tetrahydroquinoline deriva-
tives 3 and 4 shown in Figure 1. As depicted in Figure
3 the indole derivative 8 as well as 5.7-dichlorokynurenic
acid 2, depicted in Figures 1 and 2, respectively, were
superimposed to this model satisfying the observed
pharmacophore arrangement.23b
Compound 8, the indole derivative endowed with the
highest affinity for the glycine binding site (pKi ) 8.5,
as reported in Table 1), showed more than 1000-fold
receptor selectivity for the strychnine-insensitive glycine
binding site with respect to other glutamate binding
sites (pKi ) 5.0, 5.1, and 5.4 for the NMDA, AMPA, and
kainate binding site, respectively). Moreover, this
compound was found to noncompetitively inhibit the
binding of [3H]TCP in extensively washed cortical
membranes preparation, according to the procedure
described above (this binding value can be considered
as an index of the functional activation of the NMDA-
gated ion channel). This study suggested that 8 is a
noncompetitive antagonist acting at the glycine binding
site with an estimated pA2 of 8.1 nM, in good agreement
with the affinity for the glycine recognition site (pKi )
8.5) assessed, as previously reported,18 by inhibition of

Indole-2-carboxylates as Potent Antagonists
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6 845
Ta ble 2. Physicochemical Parameters Utilized for MRAa
no.
Ro
Rm
Rp
MRomp MRo MRmp
πomp
πo
πmp
σmeta
σpara
Fmp
Fomp
Rmp
Romp
8
21
22
H
H
H
H
H
NH2
H
H
H
OH
H
H
0.00
1.24
0.37
0.37
0.37
0.15
0.15
0.15
0.72
1.23
1.08
0.74
0.74
0.03
0.65
0.88
1.29
0.72
0.01
1.08
1.39
1.21
2.22
0.51
2.88
0.00
0.00
0.00
0.37
0.00
0.00
0.00
0.15
0.73
0.00
0.46
0.73
0.02
0.02
0.00
0.15
0.00
0.00
0.00
0.00
1.39
0.00
0.00
0.00
0.00
0.00
1.24
0.37
0.00
0.37
0.15
0.15
0.00
0.00
1.23
0.61
0.01
0.73
0.01
0.65
0.73
1.29
0.72
0.01
1.08
0.00
0.74
2.22
0.51
2.88
0.00
-1.30
-1.23
0.00
0.00
0.00
0.00 -0.24
0.00
0.00
0.04
0.02
0.00
0.02
0.29
0.28
0.00
0.00
0.52
0.26
0.43
0.66
0.43
0.33
0.66
0.10
0.67
0.43
0.22
0.00
0.00
0.00
NHCONH2
H
H
NH2
OH
H
H
H
0.00 -1.30
0.00 -1.23 -0.16
0.00 0.00
0.06 -0.28 -0.28
0.00
0.00
0.02 -0.24 -0.24
23 NH2
-1.23 -1.23
0.03
0.00 -0.59
24
25
26
H
H
H
-1.23
-0.67
-0.67
-0.44 -0.44
-0.37 -0.37
-0.35
-0.19 -0.26
-0.15 -0.37
-0.09 -0.20
0.02 -0.20
0.04
0.11 -0.44
0.20
0.30
0.40
0.61
0.67
0.89
1.09
1.33
1.48
0.00 -1.23
0.00 -0.67
0.00 -0.67
0.00 -0.66
0.00 -0.37
0.12
0.00
0.00
0.12 -0.27
0.00 -0.27
0.00
0.71
0.00
0.00
0.71
0.00 -0.83
0.00
0.00
0.00 -0.24
0.00
0.71
0.00 -0.90
0.00 0.54
0.00 -0.40 -0.02 -0.02 -0.38 -0.38
0.02 -0.68 -0.68
0.29 -0.64 -0.64
0.28 -0.22 -0.22
0.36
0.84
0.00
0.00
0.00
27 OH
28 NO2
29
30 Me
0.00
0.00
0.00 -0.55
0.00 0.14
H
H
OMe OMe
H
H
NO2
H
H
NO2
H
H
H
H
H
NO2
H
H
0.00 -0.35
0.52 -0.69 -0.69
0.21 -0.51 -0.62
1.27 -0.34 -0.20
OMe
F
H
0.08
0.40
0.30
0.40
0.04
0.30
0.19
0.30
0.40
0.61
0.00
0.89
1.09
1.33
1.48
31 NO2
0.06
0.00
0.06
0.45
0.00
32
33
34
F
F
H
1.19
0.06 -0.24
F
0.97 -0.34 -0.63
0.33
1.02
0.10 -0.92 -0.92
0.67 0.16 0.16
0.43 -0.34 -0.34
0.22 -0.44 -0.44
COOH
H
N(CH3)2
NO2
F
OEt
H
Cl
0.00
0.15
0.06 -0.50
0.15
35 OH
36
37
38
39
H
H
H
H
0.00
0.00
0.00
0.00
0.67
0.00
0.00
0.00
0.00
0.78
0.06
40 i-Pr
0.00
0.23
0.00 -0.06
0.00 -0.09
41
42
43
44
H
H
H
H
1.07
0.01
0.38
1.07 -0.09 -0.09
0.01 -0.91 -0.91
0.38
N(Et2)
CF3
NHPh
0.19
0.19
H
a
MRomp ) sum of molar refractivities of ortho, meta, para substituents; *MRo ) molar refractivity of ortho substituent; MRmp ) sum
of molar refractivities of meta, para substituents; πomp ) sum of lipophilicity of ortho, meta, para substituents; *πo ) lipophilicity of ortho
substituent; πmp ) sum of lipophilicity of meta, para substituents;σmeta ) sigma meta of meta substituent;25 σpara ) sigma para of para
substituent;27 **Fomp ) global Swain and Lupton F25 for ortho, meta, para positions; Fmp ) global Swain and Lupton F25 for meta, para
positions; Romp ) global Swain and Lupton R25 for ortho, meta, para positions; Rmp ) global Swain and Lupton R25 for meta, para positions.
*Molar refractivities and lipophilicity were calculate with Daylight v4.4131 program. **Combinations of Swain and Lupton’s F and R
were calculated according to ref 26.
(multiple regression analysis)28 implemented with RS/
129 was employed to analyse the data. Data handling
and parametrization were performed with the programs
TSAR v2.230 and Daylight v4.41.31
The pKi values (observed affinity for the glycine
binding site) for compounds shown in Table 2 are
reported in Table 3. The following statistically signifi-
cant equation was derived from the analysis of these
results:
portion of a parabolic or bilinear34 trend; this aspect
could be verified by exploring a wider range of negative
values of lipophilicity).
The electronic donor term is more difficult to interpret
in terms of drug-receptor interactions. It can be
speculated that this effect improves the hydrogen bond-
ing acceptor ability of the carbonyl present within the
C-3 R,â-unsaturated side chain.
Com p ou n d 8 (GV150526A): F u r t h er Biologica l
Ch a r a cter iza tion . In view of its promising pharma-
cological profile, compound 8 (GV150526A) was selected
among the number of indole derivatives identified for
further evaluation in an experimental model of focal
cerebral ischaemia in rats (MCAo).35 Compound 8
showed an excellent neuroprotective activity (significant
reduction of the infarct volume with respect to control
groups) after iv and po administration, both pre- and
postischaemia.36 Moreover this indole derivative was
characterized in terms of safety and general pharma-
cology: neither ataxic effects (rotarod) nor impairment
of performance (passive avoidance) were observed in
mice up to a dose of 30 mg/kg administered iv (500-fold
the ED50 observed in inhibition of NMDA-induced
convulsions), confirming the wider therapeutic ratio
observed with glycine antagonists compared to competi-
tive NMDA antagonists and NMDA receptor ion chan-
nel blockers.7
pKi ) -0.53MRomp - 0.39πomp - 0.82σpara + 8.23
(1)
n ) 25
p < 0.0001
R2 ) 0.84
s ) 0.28
R2 ) 0.76 (ref 32)
F ) 37
cv
where MRomp represents the total bulk of ortho, meta,
and para25 substituents, πomp is the global lipophilicity
of ortho, meta, and para25 substituents, and finally σpara
27
corresponds to the Hammett’s σpara
.
The calculated
vs observed pKi values for eq 1 are shown in Figure 4,
whereas the correlation matrix33 for variables in eq 1
is reported in Table 4.
As shown in eq 1, the affinity for the glycine receptor
increases as total bulk (MRomp) and lipophilicity (πomp
of substituents decrease. Furthermore, pKi increases
as σpara decreases, i.e. as the electron donor resonance
effect of the para substituent increases.
According to these results the terminal aromatic ring
of the R,â-unsaturated C-3 side chain should lie in a
nonhydrophobic pocket of limited size (negative coef-
ficient of πomp and MRomp), refining the previous phar-
macophore model21 of the glycine binding site associated
with the NMDA receptor. The negative linear relation
between lipophilicity and pKi could be the descending
)
Con clu sion s
A novel class of indole-2-carboxylates was explored
with the aim of identifying potent in vivo glycine
antagonists. The substitution of the C-3 position of the
indole nucleus with a suitable R,â-unsaturated side
chain gave potent antagonists endowed with nanomolar
affinity at the glycine binding site. These compounds

846 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6
Di Fabio et al.
Ta ble 3. Substitution of the Terminal Phenyl Ring of the C-3
Side Chain: in Vitro Pharmacological Characterization
°C for 15 min under nitrogen atmosphere. Dry 1,2-dichloro-
ethane (60 mL) and ethyl 4,6-dichloroindole-2-carboxylate (6
g, 23.3 mmol) were added, and the suspension was stirred at
80 °C for 6 h. The reaction mixture was poured into a 50%
aqueous solution of NaOAc (300 mL) to give, after filtration,
4.1 g (62%) of the title compound as a yellow solid: mp 220-
222 °C; IR (Nujol) νmax 2725-2669, 1726, 1663, 1556 cm-1; 1H
NMR (DMSO-d6) δ 13.15 (s, 1H), 10.60 (s, 1H), 7.54 (d, 1H),
7.40 (d, 1H), 4.43 (q, 2H), 1.36 (t, 3H).
(E)-3-[2′-[(ter t-Bu tylca r bon yl)oxy]eth en yl]-4,6-d ich lor -
oin d ole-2-ca r boxylic Acid Eth yl Ester (12). A magneti-
cally stirred solution of [(tert-butoxycarbonyl)ethenyl]triphen-
ylphosphorane (5.2 g, 15 mmol) and ethyl 3-formyl-4,6-dichlor-
oindole-2-carboxylate (3.3 g, 11.6 mmol) in a 1:1 mixture of
CH3CN/dioxane (60 mL) was heated at 70 °C for 7 h under
nitrogen atmosphere. The solvent was evaporated under
reduced pressure and the crude residue purified by flash
chromatography (cyclohexane/ethyl acetate, 1:1) to give 3.3 g
of pure compound 12 (75%): mp 157-158 °C; IR (Nujol) νmax
no.
synthetic method
Ro
Rm
Rp
pKia
8
21
22
23
24
25
26
27
28
29b
30
31
32b
33
34
35
36
37
38
39
40
41
42
43
44
B
A
A
A
A
B
B
B
B
B
B
B
B
B
A
B
A
B
A
B
B
B
A
B
A
H
H
H
NH2
H
H
H
NH2
H
H
H
OH
H
H
OCH3
H
H
NO2
H
H
NO2
H
H
H
H
H
NO2
H
H
8.5
8.8
8.3
8.5
8.9
8.7
8.4
8.4
7.6
8.1
7.7
7.5
8.3
8.3
7.2
8.0
7.9
7.0
8.2
8.3
7.3
6.9
7.0
6.8
6.7
NHCONH2
H
H
NH2
OH
H
H
H
H
H
1
3302, 1703, 1674 cm-1; H NMR (DMSO-d6) δ 9.20 (bs, 1H),
8.32 (d, 1H, 16 Hz), 7.33 (d, 1H), 7.19 (d, 1H), 6.48 (d, 1H, 16
Hz), 4.43 (q, 2H), 1.56 (s, 9H), 1.42 (t, 3H).
OH
NO2
H
CH3
NO2
F
F
H
OH
H
H
H
H
i-Pr
H
H
(E)-3-(2′-Ca r boxyeth en yl)-4,6-d ich lor oin d ole-2-ca r box-
ylic Acid (13). (E)-Ethyl 3-[2-(tert-butoxycarbonyl)ethenyl]-
4,6-dichloroindole-2-carboxylate (0.5 g, 1.3 mmol) was dissolved
in 100% HCOOH (60 mL), and the suspension was stirred at
23 °C for 2 h. The solvent was evaporated in vacuo to give
0.408 g of 3-(2-carboxyethenyl)-4,6-dichloroindole-2-carboxylate
13 as a white solid (96%): mp >250 °C; IR (Nujol) νmax 3246-
3128, 1699-1670, 1634 cm-1; 1H NMR (DMSO-d6) δ 12.6 (bs,
2H), 8.28 (d, 1H, 16.2 Hz), 7.51 (d, 1H), 7.32 (d, 1H), 6.44 (d,
1H, 16.2 Hz), 4.37 (q, 2H), 1.35 (t, 3H).
Gen er a l P r oced u r e for th e Syn th esis of Ar yl Am id es:
P r oced u r e A. To a solution of ethyl 3-(2′-carboxyethenyl)-
4,6-dichloroindole-2-carboxylate (13) (280 mg, 0.85 mmol)
dissolved in dry DMF (18 mL) were added portionwise 2,2′-
dipyridyl disulfide (267 mg, 1.2 mmol) and PPh3 (317 mg, 1.2
mmol) at room temperature under nitrogen atmosphere. The
solution was stirred for 3 h, and then the chosen arylamine
derivative (1.2 mmol) was added. The reaction mixture was
heated at 80 °C and stirred for additional 20 h. Resultant
precipitate was filtered to give pure title compounds in 65-
83% yields.
Gen er a l P r oced u r e for th e Syn th esis of Ar yl Am id es:
P r oced u r es B. A magnetically stirred solution of the chosen
triphenylphosphorane (15 mmol) and intermediate 10 (11.6
mmol) in a 1:1 mixture CH3CN/dioxane (60 mL) was stirred
at 70 °C for 7 h under nitrogen atmosphere. The solvent was
evaporated in vacuo, and the crude products were purified by
flash chromatography to give the final compounds (70-88%).
Gen er a l P r oced u r e for th e Ba sic Hyd r olysis of th e
Eth yl Ester s. To an aliquot of indole-2-carboxylic acid ethyl
ester (0.25 mmol) dissolved in isopropyl alcohol (20 mL) was
added NaOH (20 mg, 0.5 mmol). The solution was heated at
60 °C for 11 h and then diluted with water (30 mL). The
aqueous solution was concentrated in vacuo, and the precipi-
tate was filtered and washed with water to give pure sodium
salt derivative in quantitative yield.
OCH3
OCH3
F
H
F
COOH
H
N(CH3)2
NO2
F
OEt
H
Cl
N(C2H5)2
CF3
NHC6H5
H
H
H
H
a
b
Inhibition of binding of [3H]Gly (ref 13). Isolated and char-
acterized as acid derivative.
showed high receptor selectivity, coupled with an excel-
lent in vivo activity after systemic administration
inhibiting the convulsions induced by NMDA in mice
and in the MCAo model in rats.
In conclusion, the identification of this class of in vivo
potent and selective glycine antagonists will be useful
to further clarify the involvement of the glycine binding
site associated with the NMDA receptor antagonists in
different pathological conditions.
Exp er im en ta l Section
Infrared spectra were recorded on a Bruker IFS 48 spec-
trometer. 1H NMR spectra were recorded on a Varian Unity
400 (400 MHz); the data ar reported as follows: chemical shift
in ppm from the internal standard Me4Si on δ scale, multiplic-
ity (b ) broad, s ) singlet, d ) doublet, t ) triplet, q ) quartet,
m ) multiplet), and coupling constants (hertz).
3-[2′-[(P h en yla m in o)ca r bon yl]eth en yl]-4,6-d ich lor oin -
d ole-2-ca r boxylic Acid Sod iu m Sa lt (8). Prepared from
H11-
Chromatography was carried out with the use of Merck
silica gel 60 (230-400 mesh) as described by Still et al.37 Mass
spectra were recorded on a VG-4 triple quadrupole Fison
instrument in FAB mode. Elemental analyses were performed
by our own analytical group on Carlo Erba elemental analyzer.
Melting points were determined on a Bu¨chi 530 apparatus
(scale 0-250 °C) and are uncorrected. All reactions were
carried out under a controlled atmosphere in oven-dried
glassware. Anhydrous DMF was purchased from Aldrich; THF
was used after distillation over K/benzophenone; CH2Cl2 and
CH3CN were used after distillation from P2O5. Reactions were
monitored by analytical thin-layer chromatography (TLC)
using Merck silica gel 60 F-254 glass plates (0.25 mm).
Eth yl 3-F or m yl-4,6-d ich lor oin d ole-2-ca r boxyla te (10).
A solution of N-methylformanilide (5.19 g, 38.4 mmol) and
phosporous oxychloride (5.53 g, 36.1 mmol) was stirred at 23
compound 10 following the general procedure B: C18
Cl2N2O3Na; IR (Nujol) νmax 3404-3126, 1624, 1616, 1600 cm-1
;
MS m/e 397 (M + 1); 1H NMR (DMSO-d6) δ 11.8 (bs, 1H), 10.06
(s, 1H), 8.59 (d, 1H, 15.5 Hz), 7.75 (d, 2H), 7.44 (d, 1H), 7.27
(t, 2H), 7.21 (d, 1H, J ) 15.5 Hz), 7.10 (d, 1H), 6.98 (t, 1H).
3-[2′-[(P h en ylam in o)car bon yl]eth yl]-4,6-dich lor oin dole-
2-ca r boxylic Acid Sod iu m Sa lt (16). An aliquot of 3-(2′-
carboxyethyl)-4,6-dichloroindole-2-carboxylic acid ethyl ester
(Chem. Pharm. Bull. 1977, 25 (11), 3023) was treated with
equimolar amount of CDI and aniline in anhydrous DMF at
60 °C for 6 h. The reaction mixture was poured in H2O and
extracted with AcOEt. The organic layer was separated, dried
over Na2SO4, and evaporated in vacuo. The crude residue was
purified by flash chromatography (CH2Cl2/MeOH, 9:1) to give
the final compound (40%). The hydrolysis of the 2-carboxy-
ethyl ester group was performed according to the general

Indole-2-carboxylates as Potent Antagonists
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6 847
F igu r e 4. Calculated vs observed pKi values for compounds in eq 1.
Ta ble 4. Correlation Matrix for variables in Eq 1
pound 12 according to the general procedure above described
for basic hydrolysis: C16H14Cl2NO4Na; IR (Nujol) νmax 3339,
1700, 1653, 1612 cm-1; 1H NMR (DMSO-d6) 12.0 (bs, 1H), 8.61
(d, 1H), 7.41 (m, 1H), 7.11 (m, 1H), 7.07 (d, 1H), 1.45 (bs, 1H).
3-[2′-[(P h en yla m in o)su lfon yl]eth en yl]-4,6-d ich lor oin -
d ole-2-ca r boxylic Acid Sod iu m Sa lt (19). An aliquot of 4,6-
dichloroindole-2-carboxylic acid ethyl ester (J . Med. Chem.
1990, 33, 2244) was treated with 2 equiv of N-phenylethene-
sulfonamide (RN 3192-10-7), 1 equiv of PdCl2, and 4 equiv of
Cu(OAc)2 in CH3CN. The solution was refluxed for 16 h under
argon atmosphere. The reaction mixture was poured in H2O
and extracted with AcOEt. The organic layer was separated,
dried over Na2SO4, and evaporated in vacuo. The crude
residue was purified by flash chromatography (cyclohexane/
AcOEt, 7:3) to give the final compound (20%). The final
hydrolysis of the 2-carboxyethyl ester group was carried out
according to the general procedure described above, to give
title compound 19 in quantitative yield: C17H12Cl2N2O4SNa;
MRomp
1
πomp
σomp
MRomp
πomp
0.536
1
-0.415
0.106
1
σomp
Ta ble 5. Physical Data of New Compounds
mp, MS
no. Ro
Rm
Rp
formulaa
°Cb (m/z)
8
16
17
18
19
20
21
22
H
H
H
C
18H11Cl2N2O3Na
a
a
a
a
a
a
a
a
a
a
a
a
a
a
243
a
a
397
399
344
402
434
411
455
412
412
412
437
437
437
442
435
442
460
C18H13Cl2N2O3Na
C12H5Cl2NO4Na2
C16H14Cl2NO4Na
C17H12Cl2N2O4SNa
C19H13Cl2N2O3Na
H
H
H
NH2
H
NHCONH2 C19H13Cl2N4O4Na‚H2O
H
H
NH2
OH
H
C18H12Cl2N3O3Na‚H2O
C18H12Cl2N3O3Na‚5H2O
C18H12Cl2N3O3Na‚0.1H2O
C18H10Cl2N2O4Na2‚H2O
C18H10Cl2N2O4Na2‚3H2O
C18H10Cl2N2O4Na2‚H2O
C18H10Cl2N3O5Na‚H2O
C20H16Cl2N2O5‚0.5H2O
C20H15Cl2N2O4Na‚0.1H2O
C18H9Cl2FN3O5Na‚H2O
C18H10Cl2FO5N3
IR (Nujol) νmax 3288, 1691, 1603, 1315, 1138 cm-1; H NMR
1
23 NH2
24
25
26
H
H
H
H
H
OH
H
(DMSO-d6) 12.7 (s, 1H), 10.02 (s, 1H), 8.20 (d, 1H), 7.45 (d,
1H), 7.27 (d, 1H), 7.30-7.16 (m, 5H), 7.0 (m, 1H).
3-[2′-[(Ben zyla m in o)ca r bon yl]eth en yl]-4,6-d ich lor oin -
d ole-2-ca r boxylic Acid Sod iu m Sa lt (20). Prepared ac-
cording to the general procedure A: C19H13Cl2N2O3Na; IR
(Nujol) νmax 3400, 1649, 1609 cm-1; 1H NMR (DMSO-d6) 11.70
(bs, 1H), 8.45 (d, 1H), 8.44 (t, 1H), 7.38 (d, 1H), 7.35-7.25 (m,
4H), 7.20 (t, 1H), 7.07 (d, 1H), 6.99 (d, 1H), 4.34 (d, 2H).
3-[2′-[[(4-Ur eid op h en yl)a m in o]ca r bon yl]eth en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (21). Pre-
pared according to the general procedure A: C19H13Cl2-
N4O4Na‚H2O; IR (Nujol) νmax 3268-3190, 1684, 1650 cm-1; 1H-
NMR (DMSO-d6) 11.86 (bs, 1H), 9.96 (bs, 1H), 9.77 (s, 1H),
8.76 (d, 1H), 7.63 (d, 2H), 7.51 (d, 2H), 7.41 (d, 1H), 7.13 (d,
1H), 7.00 (d, 1H), 6.36 (bs, 2H).
27 OH
H
H
28 NO2
H
29
H
OCH3 OCH3
30 CH3
31 NO2
H
H
OCH3
F
H
F
32
33
34
F
F
H
NO2
H
H
>250 438
C18H9Cl2F2N2O3Na‚H2O
C19H10Cl2N2O5Na2‚2H2O
C18H9Cl2N3O6Na2‚H2O
C20H16Cl2N3O3Na
C18H10Cl2N3O5Na‚H2O
C18H10Cl2N2O3FNa
a
a
a
a
a
a
a
a
a
a
434
463
480
439
442
415
441
439
476
468
466
488
COOH
H
N(CH3)2
NO2
F
35 OH NO2
36
37
38
39
H
H
H
H
H
H
H
H
H
OEt
H
C20H15Cl2N2O4Na‚0.5H2O
C21H17Cl2N2O3Na
40 i-Pr
41
42
43
44
H
H
H
H
NO2 Cl
C18H9Cl3N3O5Na‚H2O
3-[2′-[[(2-Am in op h en yl)a m in o]ca r b on yl]et h en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (22). Pre-
pared according to the general procedure A: C18H12Cl2N3O3-
Na‚H2O; IR (Nujol) νmax 3500-3100, 1647, 1607 cm-1; 1H NMR
(DMSO-d6) δ 11.85 (bs, 1H), 9.32 (bs, 1H), 8.56 (d, 1H, 15.6
Hz), 7.43 (d, 1H), 7.32 (d, 1H), 7.22 (d, 1H, 15.6 Hz), 7.09 (d,
1H), 6.86 (m, 1H), 6.71 (dd, 1H), 6.54 (m, 1H), 4.94 (bs, 2H).
3-[2′-[[(3-Am in op h en yl)a m in o]ca r b on yl]et h en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (23). Pre-
pared according to the general procedure A: C18H12Cl2N3O3-
H
H
H
N(C2H5)2
C22H20Cl2N3O3Na‚0.5H2O
CF3
C19H11Cl2F3N2O3Na‚0.5H2O a
C24H16Cl2N3O3Na‚H2O
NHC6H5
a
a
C, H, N analyses were within 0.4% of the theoretical values
for the formulae given unless otherwise noted. a: sodium salt
derivatives melting over 250 °C.
b
procedure described above to give title compound 16 in
quantitative yield: C18H13Cl2N2O3Na; IR (Nujol) νmax 1661
cm-1; 1H NMR (DMSO-d6) 13.4 (bs, 1H), 11.9 (s, 1H), 9.93 (bs,
1H), 7.57 (d, 2H), 7.38 (s, 1H), 7.26 (t, 2H), 7.15 (s, 1H), 7.0 (t,
1H), 3.62 (t, 2H), 3.57 (t, 2H).
Na‚5H2O; IR (Nujol) νmax 3188, 1601 cm-1; H NMR (DMSO-
1
d6) δ 11.8 (bs), 10.05 (s), 9.72 (s), 8.62 (d), 8.55 (d), 7.43 (d),
7.12 (m), 7.11 (d), 6.91 (t), 6.84 (d), 6.23 (d), 5.00 (bs).
3-[2′-[[(4-Am in op h en yl)a m in o]ca r b on yl]et h en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (24). Pre-
pared according to the general procedure A: C18H12Cl2N3O3-
3-(2′-Ca r b oxye t h e n yl)-4,6-d ich lor oin d ole -2-ca r b ox-
ylic Acid Disod iu m Sa lt (17). Prepared from compound 13
according to the general procedure described above for basic
hydrolysis: C12H5Cl2NO4Na2; IR (Nujol) νmax 1684, 1636, 1610
Na‚0.1H2O; IR (Nujol) νmax 3389-3196, 1607 cm-1; H NMR
1
cm-1; H NMR (DMSO-d6) 12.26 (bs, 1H), 8.34 (d, 1H), 7.49
1
(DMSO-d6) δ 11.80 (bs, 1H), 9.61 (bs, 1H), 8.47 (d, 1H), 7.40
(d), 7.38 (d), 7.08 (d), 7.05 (d), 6.48 (d, 2H), 4.79 (bs, 2H).
3-[2′-[[(4-Hyd r oxyp h en yl)a m in o]ca r bon yl]eth en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (25). Pre-
(d, 1H), 7.32 (d, 1H), 6.50 (d, 1H).
3-[2′-(ter t-Bu toxyca r bon yl)eth en yl]-4,6-d ich lor oin dole-
2-ca r boxylic Acid Sod iu m Sa lt (18). Prepared from com-

848 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6
Di Fabio et al.
pared according to the general procedure B: C18H10Cl2N2O4-
Sa lt (36). Prepared according to the general procedure A:
Na2‚H2O; IR (Nujol) νmax 3437-2670, 1684 cm-1
;
1H NMR
C20H16Cl2N3O3Na; IR (Nujol) νmax 3184, 2670, 1700-1620 cm-1
;
(DMSO-d6) δ 13.7 (bs, 1H), 12.47 (bs, 1H), 9.94 (bs, 1H),
9.20 (bs, 1H), 8.21 (d, 1H), 7.49 (d), 7.46 (d), 7.28 (6.73, 1H),
6.69 (d).
1H NMR (DMSO-d6) δ 13.70 (bs, 1H), 12.42 (bs, 1H), 9.82 (bs,
1H), 8.21 (d, 1H), 7.53 (d), 7.46 (d), 7.26 (d, 1H), 6.75 (d), 7.69
(d), 2.84 (s, 6H).
3-[2′-[[(3-Hyd r oxyp h en yl)a m in o]ca r bon yl]eth en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (26). Pre-
pared according to the general procedure B; C18H10Cl2N2O4-
3-[2′-[[(4-Nit r op h en yl)a m in o]ca r b on yl]et h en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (37). Pre-
pared according to the general procedure B: C18H10Cl2N3O5-
Na2‚3H2O; IR (Nujol) νmax 3098, 1705, 1676, 1607 cm-1 1H
;
Na‚H2O; IR (Nujol) νmax 3346 cm-1 1H NMR (DMSO-d6) δ
;
NMR (DMSO-d6) δ 13.70 (bs, 1H), 12.53 (s, 1H), 10.05 (s, 1H),
9.38 (s, 1H), 8.23 (d, 1H, 15.9 Hz), 7.47 (d, 1H), 7.30 (d, 2H),
7.06 (m, 2H), 6.74 (d, 1H, 15.9 Hz), 6.44 (dt, 1H).
3-[2′-[[(2-Hyd r oxyp h en yl)a m in o]ca r bon yl]eth en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (27). Pre-
pared according to the general procedure B: C18H10Cl2N2O4-
Na2‚H2O; IR (Nujol) νmax 3150, 1736, 1656, 1630 cm-1; 1H NMR
(DMSO-d6) δ 14.2-13.2 (bs, 1H), 12.56 (s, 1H), 9.97 (bs, 1H),
9.7 (s, 1H), 8.24 (s, 1H), 7.8 (d, 1H), 7.49 (d, 1H), 7.30 (d, 1H),
6.96 (d, 1H), 6.96 (dt), 6.88 (dd), 6.79 (td).
12.20 (bs, 1H), 10.89 (bs, 1H), 8.74 (d, 1H), 8.19 (m, 2H), 8.05
(m, 2H), 7.52 (d, 2H), 7.45 (d, 1H), 7.13 (d, 1H).
3-[2′-[[(4-F lu or op h en yl)a m in o]ca r b on yl]et h en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (38). Pre-
pared according to the general procedure A: C18H10Cl2N2O3-
FNa; IR (Nujol) νmax 3206, 1628-1609 cm-1; 1H NMR (DMSO-
d6) δ 11.08 (bs, 1H), 10.14 (bs, 1H), 8.62 (d, 1H, 15.6 Hz), 7.79
(m, 1H), 7.45 (d, 1H), 7.25 (d, 1H, 15.6 Hz), 7.11 (m), 7.11 (d).
3-[2′-[[(4-Eth oxyp h en yl)a m in o]ca r bon yl]eth en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (39). Pre-
pared according to the general procedure B: C20H15Cl2N2O4-
3-[2′-[[(2-Nit r op h en yl)a m in o]ca r b on yl]et h en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (28). Pre-
pared according to the general procedure B: C18H10Cl2N3O5Na‚
1
Na‚0.5H2O; IR (Nujol) νmax 3248, 1663, 1632, 1610 cm-1; H
NMR (DMSO-d6) δ 13.70 (bs, 1H), 12.50 (bs, 1H), 10.04 (bs,
1H), 8.22 (d, 1H, 15.6 Hz), 7.61 (d, 2H), 7.47 (d, 1H), 7.29 (d,
1H), 6.86 (d, 2H), 6.74 (d, 1H, 15.6 Hz), 3.97 (q, 2H), 1.29 (t,
3H).
H2O; IR (Nujol) νmax 3234, 1684, 1636, 1609 cm-1 1H NMR
;
(DMSO-d6) δ 12.20 (bs, 1H), 10.51 (bs, 1H), 8.59 (d, 1H), 7.95
(dd, 1H), 7.81 (dd, 1H), 7.69 (m, 1H), 7.48 (d, 1H), 7.38-7.28
(m, 2H), 7.20 (d, 1H).
3-[2′-[[(2-Isop r op ylp h en yl)a m in o]ca r b on yl]et h en yl]-
4,6-d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (40).
Prepared according to the general procedure B: C21H17Cl2N2O3-
3-[2′-[[(3,4-Dim eth oxyph en yl)am in o]car bon yl]eth en yl]-
4,6-d ich lor oin d ole-2-ca r boxylic Acid (29). Prepared ac-
cording to the general procedure B: C20H16Cl2N2O5‚0.5H2O;
Na; IR (Nujol) νmax 3261, 1610 cm-1 1H NMR (DMSO-d6) δ
;
1
IR (Nujol) νmax 3420, 3281, 1690, 1680, 1620, 1607 cm-1; H
12.10 (bs, 1H), 9.39 (bs, 1H), 8.57 (d, 1H), 7.57 (bs, 1H), 7.38-
7.28 (m), 7.28-7.10 (m), 3.25 (m, 1H), 1.15 (d, 6H).
NMR (DMSO-d6) δ 13.8-13.6 (bs, 1H), 12.53 (bs, 1H), 10.08
(bs, 1H), 8.23 (d, 1H, 16 Hz), 7.47 (m, 2H), 7.29 (d, 1H), 7.2
(dd, 1H), 6.89 (d, 1H), 6.74 (d, 1H, 16 Hz), 3.73 (s, 3H), 3.7 (s,
3H).
3-[2′-[[(4-Ch lor o-3-n itr op h en yl)a m in o]ca r bon yl]eth e-
n yl]-4,6-d ich lor oin d ole-2-ca r b oxylic Acid Sod iu m Sa lt
(41). Prepared according to the general procedure B: C18H9-
Cl3N3O5Na‚H2O; IR (Nujol) νmax 3383-3182, 1610, 1587 cm-1
;
3-[2′-[[(2-Met h yl-4-m et h oxyp h en yl)a m in o]ca r b on yl]-
eth en yl]-4,6-d ich lor oin d ole-2-ca r boxylic Acid Sod iu m
Sa lt (30). Prepared according to the general procedure B:
C20H15Cl2N2O4Na‚0.1H2O; IR (Nujol) νmax 3279, 1703, 1661,
1630 cm-1; 1H NMR (DMSO-d6) δ 12.41 (bs, 1H), 9.39 (bs, 1H),
8.26 (d, 1H), 7.48 (d), 7.36 (d), 7.27 (d), 6.90 (d), 6.80 (d), 6.75
(dd), 3.73 (s, 3H), 2.19 (s, 3H).
1H NMR (DMSO-d6) δ 12.18 (bs, 1H), 10.87 (bs, 1H), 8.71 (d,
1H, 15.6 Hz), 7.67 (d, 1H), 8.00 (dd, 1H), 7.69 (d, 1H), 7.53 (d,
1H), 7.48 (d, 1H, 15.6 Hz), 7.13 (d, 1H).
3-[2′-[[[4-(d ie t h y la m in o )p h e n y l]a m in o ]c a r b o n y l]-
eth en yl]-4,6-d ich lor oin d ole-2-ca r boxylic Acid Sod iu m
Sa lt (42). Prepared according to the general procedure A:
C22H20Cl2N3O3Na‚0.5H2O; IR (Nujol) νmax 3248, 3192, 1649,
1610 cm-1; 1H NMR (DMSO-d6) δ 11.80 (bs, 1H), 9.67 (s, 1H),
8.47 (d, 1H, 15.6 Hz), 7.51 (d, 2H), 7.39 (d, 1H), 7.08 (d, 1H),
7.08 (d, 1H, 15.6 Hz), 6.59 (d, 2H), 3.4 (q, 4H), 1.05 (t, 6H).
3-[2′-[[[4-(Tr iflu or om et h yl)p h en yl]a m in o]ca r b on yl]-
eth en yl]-4,6-d ich lor oin d ole-2-ca r boxylic Acid Sod iu m
Sa lt (43). Prepared according to the general procedure A:
C19H11Cl2F3N2O3Na‚0.5H2O; IR (Nujol) νmax 3430, 3000, 1700,
1678, 1636, 1614 cm-1; 1H NMR (DMSO-d6) δ 14.00-13.50 (bs,
1H), 12.55 (bs, 1H), 10.54 (bs, 1H), 8.37 (d, 1H, 15.8 Hz), 7.91
(d, 2H), 7.67 (d, 2H), 7.48 (d, 1H), 7.30 (d, 1H), 6.86 (d, 1H,
15.8 Hz).
3-[2′-[[(4-F lu or o-2-n itr op h en yl)a m in o]ca r bon yl]eth e-
n yl]-4,6-d ich lor oin d ole-2-ca r b oxylic Acid Sod iu m Sa lt
(31). Prepared according to the general procedure B: C18H9-
1
Cl2FN3O5Na‚H2O; IR (Nujol) νmax 3312, 1653, 1634 cm-1; H
NMR (DMSO-d6) δ 12.00 (bs, 1H), 10.44 (bs, 1H), 8.65 (d, 1H),
7.88 (dd), 7.79 (dd), 7.62 (m, 1H), 7.45 (d), 7.45 (d), 7.15 (d,
1H).
3-[2′-[[(2-flu or o-3-n itr oph en yl)am in o]car bon yl]eth en yl]-
4,6-d ich lor oin d ole-2-ca r boxylic Acid (32). Prepared ac-
cording to the general procedure B: C18H10Cl2FO5N3; IR (Nujol)
νmax 3414-3100, 1684-1614, 1539-1348 cm-1 1H NMR
;
(DMSO-d6) δ 13.82 (bs), 12.60 (bs), 10.46 (bs), 9.21 (dd), 8.41
(d), 8.05 (m), 7.59 (t), 7.50 (d), 7.33 (d), 7.06 (d).
3-[2′-[[[4-(P h e n y la m in o )p h e n y l]a m in o ]c a r b o n y l]-
eth en yl]-4,6-d ich lor oin d ole-2-ca r boxylic Acid Sod iu m
Sa lt (44). Prepared according to the general procedure A:
3-[2′-[[(2,4-Diflu or op h en yl)a m in o]ca r b on yl]et h en yl]-
4,6-d ich lor oin d ole-2-ca r boxylic Acid Sod iu m n Sa lt (33).
Prepared according to the general procedure B: C18H9Cl2F2N2O3-
C24H16Cl2N3O3Na‚H2O; IR (Nujol) νmax 3389, 1595, 1650 cm-1
;
Na‚H2O; IR (Nujol) νmax 3431-3233, 1707, 1678, 1612 cm-1
;
1H NMR (DMSO-d6) δ 9.88 (bs, 1H), 8.55 (d, 1H, 15.7 Hz), 8.00
(s, 1H), 7.63 (d, 2H), 7.40 (d, 1H), 7.17 (t, 2H), 7.16 (d, 1H,
15.7 Hz), 7.10 (d, 1H), 7.02 (d, 2H), 6.98 (d, 2H), 6.73 (t, 1H).
1H NMR (DMSO-d6) δ 14.0-13.6 (bs, 1H), 12.54 (bs, 1H), 9.99
(bs, 1H), 8.29 (d, 1H, 15.6 Hz), 7.97 (m, 1H), 7.48 (d, 1H), 7.30
(m, 1H), 7.29 (d, 1H), 7.07 (m, 1H), 6.90 (d, 1H, 15.6 Hz).
3-[2′-[[(4-Ca r boxyp h en yl)a m in o]ca r bon yl]eth en yl]-4,6-
d ich lor oin d ole-2-ca r boxylic Acid Sod iu m Sa lt (34). Pre-
pared according to the general procedure A: C19H10Cl2N2-
O5Na2‚2H2O; IR (Nujol) νmax 3406, 1734 cm-1; 1H NMR (DMSO-
d6) δ 9.94 (bs, 1H), 8.57 (d, 1H, 15.6 Hz), 7.72 (d, 2H), 7.59 (d,
2H), 7.37 (d, 1H), 7.17 (d, 1H, 15.6 Hz), 7.09 (d, 1H).
Ack n ow led gm en t. The authors thank Dr. C. Mar-
1
chioro and her co-workers for H-NMR spectra, Dr. M.
Handam and his co-workers for mass spectra data, and
Dr. A. Garofalo for elemental analysis. Thanks are due
to Dr. P. Albertini, Mr. S. Costa, and Mr. E. Valerio for
the in vivo characterization in the NMDA-induced
convulsions model in mice.
3-[2′-[[(2-Hyd r oxy-3-n itr op h en yl)a m in o]ca r bon yl]eth e-
n yl]-4,6-d ich lor oin d ole-2-ca r b oxylic Acid Sod iu m Sa lt
(35). Prepared according to the general procedure B: C18H9-
Cl2N3O6Na2‚H2O; IR (Nujol) νmax 3383, 3298, 1700, 1530, 1377
Refer en ces
1
cm-1; H NMR (DMSO-d6) δ 12.53 (s, 1H), 9.74 (s, 1H), 9.10
(1) (a) Meldrum, B.; Garthwaite, J . Excitatory Amino Acid Neuro-
toxicity and Neurodegenerative Diseases. Trends Pharmacol.
1990, 11, 379-387. (b) Choi, D. W. Cerebral Hypoxia: Some
New Approaches and Unanswered Questions. J . Neurosci. 1990,
10, 2493-2501. (c) Meldum, B. Protection Against Ischaemic
(d, 1H), 8.30 (d, 1H), 7.90 (dd, 1H), 7.48 (d, 1H), 7.29 (d, 1H),
7.06 (d, 1H), 7.02 (d, 1H).
3-[2′-[[[4-(Dim et h yla m in o)p h en yl]a m in o]ca r b on yl]-
eth en yl]-4,6-d ich lor oin d ole-2-ca r boxylic Acid Sod iu m

Indole-2-carboxylates as Potent Antagonists
J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6 849
Neuronal Damage by Drugs Acting on Excitatory Neurotrans-
mission. Cerebrovasc. Brain Metab. Rev. 1990, 2, 27-57. (d)
McCullogh, J . Excitatory Amino Acid Antagonists and their
Potential for the Treatment of Ischemic Brain Damage in Man.
Br. J . Clin. Pharmacol. 1992, 34, 106-114.
membered Ring Conformation of Affinity. Bioorg. Med. Chem.
Lett. 1992, 2 (12), 1627-1630. (g) Rao, T. S.; Gray, N. M.;
Dappen, M. S.; Cler, J . A.; Mick, S. J .; Emmett, M. R.; Iyergan,
S.; Monahan, J . B.; Cordi, A. A.; Wood, P. L. Indole-2-Carboxy-
lates, Novel Antagonists of the N-Methyl-D-Aspartate (NMDA)-
Associated Glycine Recognition Sites: in vivo Characterization.
Neuropharmacology 1993, 32 (2), 139-147. (h) Smith, E. C. R.;
McQuaid, L. A.; Calligaro, D. O.; OåopMalley, P. J . Structure-
Activity Relationship of a Series of Glycine Antagonists Related
to 5,7-Dichlorokynurenic acid and 3-(2-Carboxy-6-Chloroindol-
3-yl)Acetic Acid. Bioorg. Med. Chem. Lett. 1993, 3 (1), 81-84.
(11) Larock, R. C. Comprehensive Organic Transformations 972-976;
VCH Publishers, Inc.: New York, 1989.
(12) Di Fabio, R.; Summa, V.; Rossi, T. Synthesis of Amides: An
Efficient and Chemoselective Method for the Preparation of
â-lactam Derivatives Related to HLE Inhibitors. Tetrahedron
1993, 49 (11), 2239-2306 and references therein.
(13) Kishimoto, H.; Simon, J . R.; Aprison, M. H. Determination of
the Equilibrium Dissociation Constants at a Number of Glycine
Binding Sites in Several Areas of the Rat Central Nervous
System, Using a Na-Indipendent System. J . Neurochem. 1981,
37, 1015-1024.
(14) Munson, P. D.; Rodbard, D. LIGAND: A Versatile Computerized
Approach for Characterization of Ligand-binding Data. Anal.
Biochem. 1980, 107, 220-239.
(15) Van Amsterdam, F. Th. M.; Giberti, A.; Mugnaini, M.; Ratti, E.
3-[(()-2-Carboxypiperazin-4-yl]propyl-1-Phosphonic Acid Rec-
ognizes Two N-Methyl-D-Aspartate Binding Sites in Rat Cere-
bral Cortex Membranes. J . Neurochem. 1992, 59 (5), 1850-
1855.
(16) Giberti, A.; Ratti, E.; Gaviraghi, G.; van Amsterdam, F. Th. M.
Binding of DL-[3H]-R-Amino-3-Hydroxy-5-Methyl-Isoxazole-4-
Propionic Acid (AMPA) to Rat Cortex Membranes Reveals
Two Sites or Affinity States. J . Recept. Res. 1991, 11 (5), 727-
741.
(17) Honore´, T.; Drejer, J .; Nielsen, M. Calcium Discriminates Two
[3H]Kainate Binding Sites with Different Molecular Target Sizes
in Rat Cortex. Neurosci. Lett. 1986, 65, 47-52.
(18) Kloog, Y.; Haring, R.; Sokolovsky, M. Kinetic Characterization
of the Phenylcyclidine-N-Methyl-D-Aspartate Receptor Interac-
tion: Evidence for Steric Blockade of the Channel. Biochemistry
1988, 27, 843-848.
(2) (a) Cotman, C. W.; Foster, A. C.; Lanthorn, T. H. Adv. Biochem.
Biopharmacol. 1981, 27, 1-27. (b) Watchins, J . C.; Evans, R.
H. Excitatory Amino Acids Transmitters. Annu. Rev. Pharma-
col. Toxicol. 1981, 21, 165-204.
(3) Choi, D. W.; Rothman, S. M. The Role of Glutamate Neurotox-
icity in Hypoxic-Ischemic Neuronal Death. Annu. Rev. Phar-
macol. Toxicol. 1991, 31, 171-182.
(4) (a) Wong, E. H. F.; Kemp, J . A. Site for Antagonism on the
N-methyl-D-aspartate Receptor Channel Complex. Annu. Rev.
Pharmacol. Toxicol. 1991, 31, 401-425. (b) Meldrum, B., Eds.
Excitatory Amino Acid Antagonists; Blackwall: Oxford, England,
1991.
(5) Collingridge, G. L., Watkins, J . C., Eds. The NMDA Receptor,
2nd ed.; IRL Press: Oxford, England, 1994.
(6) (a) J ohnson, J . W.; Ascher, P. Glycine Potentiates the NMDA
Receptor Response in Cultured Brain Mouse Neurones. Nature
1987, 325, 529-531. (b) Kleckner, N. W.; Dingledine, R.
Requirements for Glycine in Activation of NMDA Receptors Ex-
pressed in Xenopus Oocytes. Science 1988, 241, 835-837. Re-
views: (c) Thomson, A. M. Glycine is a Coagonist at the NMDA
Receptor/Ion Channel Complex. Prog. Neurobiol. 1990, 35, 53-
74. (d) Huettner, J . E. Competitive Antagonism of Glycine at
the N-Methyl-D-Aspartate (NMDA) Receptor. Biochem. Phar-
macol. 1991, 41, 9-16.
(7) (a) Morris, R. G. M.; Anderson, E.; Lynch, G. S.; Baudry, M.
Selective Impairment of Learning and Blockage of Long-term
Potentiation by an N-Methyl-D-Aspartate antagonist, AP5.
Nature 1986, 319, 774-776. (b) Koek, W.; Colpaert, F. C.
Selective Blockade of N-Methyl-D-aspartate (NMDA)-induced
Convulsions by NMDA Antagonists and Putative Glycine Antag-
onists: Relationship with Phenylcyclidine-like Behavioral Ef-
fects. J . Pharmacol. Exp. Ther. 1990, 252, 349-357. (c)
Chiamulera, C.; Costa, S.; Reggiani, A. Effect of NMDA and
Strychnine-Insensitive Glycine Site Antagonists on NMDA-
Mediated Convulsions and Learning. Psycopharmacology 1990,
112, 551-552.
(8) Reviews: (a) Palfreyman, M. G.; Baron, M. B. Non-competitive
NMDA Antagonists Acting at the Glycine Site. In Excitatory
Amino Acid Antagonists; Meldrum, B., Ed.; Blackwell: Oxford,
England, 1991; Chapter 6, pp 101-129. (b) Carter, A. J . Glycine
Antagonists: Regulation of the NMDA Receptor-Complex by the
Strychnine-Insensitive Glycine Site. Drugs Future 1992, 17 (7),
595-613. (c) Kemp, J . A.; Leeson, P. D. The Glycine Site of the
NMDA Receptor - Five Years On. Trends Pharmacol. Sci. 1993,
14, 20-25. (d) Leeson, P. D.; Iversen, L. L. The Glycine Site on
the NMDA receptor: Structure-Activity Relationships and Thera-
peutic Potential. J . Med. Chem. 1994, 37, 4053-4067. (e)
Iversen, L. L.; Kemp, J . A. Non competitive NMDA Antagonists
as Drug. In The NMDA Receptor, 2nd ed.; Collingridge, G. L.,
Watkins, J . C., Eds.; IRL Press: Oxford, England, 1994; Chapter
20, pp 469-486. (f) Di Fabio, R.; Gaviraghi, G.; Reggiani, A.
Strychnine-Insensitive Glycine Binding Site and the NMDA
Receptor. Chim. Ind. 1996, 78, 283-289.
(9) Moore, K. W.; Leeson, P. D.; Carling, R. W.; Tricklebank, M. D.;
Singh, L. Anticonvulsant Activity of Glycine-Site NMDA An-
tagonists. 1. 2-Carboxyl Prodrugs of 5,7-Dichlorokynurenic
Acid. Bioorg. Med. Chem. Lett. 1993, 3 (1), 61-64. Carling, R.
W.; Leeson, P. D.; Moseley, A. M.; Smith, J . D.; Saywell, K.;
Tricklebank, M. D.; Kemp, J . A.; Marshall, J . R.; Foster, A. C.;
Grimwood, S. Anticonvulsant Activity of Glycine-Site NMDA
Antagonists. 2. Trans 2-Carboxy-4-Substituted Tetrahydro-
quinolines. Bioorg. Med. Chem. Lett. 1993, 3 (1), 65-70.
(10) (a) Huettner, J . E. Indole-2-Carboxylic Acid: A Competitive
Antagonist of Potentiation By Glycine at the NMDA Receptor.
Science 1989, 243, 1611-1613. (b) Salituro, F. G.; Harrison, B.
L.; Baron, B. M.; Nyce, P. L.; Stewart, K. T.; McDonald, I. A.
Indole Derivatives: Antagonists of the Strychnine-Insensitive
Glycine Receptor Associated with the N-Methyl-D-Aspartate
Receptor Complex. J . Med. Chem. 1990, 33, 2944-2946. (c)
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. (d) Salituro, F. G.; Tomlinson, R. C.;
Baron, B. M.; Demeter, D. A.; Weintraub, H. J . R.; McDonald,
I. A. Design, Synthesis and Molecular Modeling of 3-Acylamino-
2-carboxyindole NMDA Receptor Glycine Site Antagonists.
Bioorg. Med. Chem. Lett. 1991, 1 (9), 455-460. (e) Salituro, F.
G.; Harrison, B. L.; Baron, B. M.; Nyce, P. L.; Stewart, K. T.;
Kehne, J . H.; White, H. S.; McDonald, I. A. 3-(2-Carboxylindol-
3-yl)propionic Acid-Based Antagonists of the N-Methyl-D-as-
partic Acid Receptor Associated Glycine Binding Site. J . Med.
Chem. 1992, 35, 1791-1799. (f) Rowley, M.; Leeson, P. D.;
Grimwood, S.; Foster, A.; Saywell, K. 2-Carboxy Indolines and
Indoles as Potential Glycine/NMDA Antagonists: Effect of Five-
(19) Ratti, E.; Tacconi, S.; Graziani, F.; Gaviraghi, G. Requirement
of the Glycine for the Glutamate Activity at the NMDA Receptor
Site. Eur. J . Pharmacol. 1990, 183, 1665.
(20) Arunlakshana, O.; Schild, H. O. Some Quantitative Use of Drug
Antagonists Br. J . Pharmacol. 1959, 14, 48-58.
(21) (a) Leeson, P. D.; Baker, R.; Carling, R. W.; Curtis, N. R.; Moore,
K. W.; Williams, B. J .; Foster, A. C.; Donald, A. D.; Kemp, J . A.;
Marshall, G. R. Kynurenic Acid Derivatives: Structure-Activity
Relationship 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. (b) 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-Amino-2-Carboxytetrahydroquinolines. Struc-
ture-Activity Relationship for Antagonism at the Glycine Site
of the NMDA Receptor. J . Med. Chem. 1992, 35, 1954-1968.
(22) Mayer, D.; Naylor, C. B.; Motoc, I.; Marshall, G. R. A Unique
Geometry of the Active Site of the Angiotensin-Converting
Enzyme Consist with Structure-Activity Studies. J . Comput-
Aided Mol. Des. 1987, 1, 3-16.
(23) (a) Sybyl 6.0, TRIPOS Inc., St. Louis, MO. (b) In this case,
distance maps were built up considering the NH of the indole
nucleus, the Cl substituent present at position C-6, the lone pairs
of the carboxylic group at position C-2 of the indole ring, and
those present on the oxygen atom of carbonyl group belonging
to the C-3 side chain. In addition, several parameters affecting
the AAA results were tuned with a “trial and error” procedure
in order to increase the accuracy of the model. Two solutions
(tuples) were obtained using superconstrained distance maps
with a grid size of 0.5 Å and 10 degrees as rotatable bond
increment. Conformations satisfying the distance map con-
straints were minimized and superimposed according to the
previously defined pharmacophore points.
(24) Although moderately, compound 16 was found to be active (ED50
) 3.7 mg/kg) in our study in contrast to what published by
Rowley et al. (ref 10f). This discrepancy could be due to the
different model used (DBA/2 model in mice).
(25) Hansch, C.; Leo, A. Substituent constants for Correlation Analy-
sis in Chemistry and Biology; J ohn Wiley & Sons: New York,
1979.
(26) Williams, S. G.; Norrington, F. E. Determination of Positional
Weighting Factors for the Swain and Lupton Substituent
Constants F and R. J . Am. Chem. Soc. 1976, 98, 508-516.
(27) Hammett, L. P. Physical Organic Chemistry; McGraw-Hill: New
York, 1940 and 1970.
(28) Draper, N. R.; Smith, H. Applied Regression Analysis; Wiley:
New York, 1981.

850 J ournal of Medicinal Chemistry, 1997, Vol. 40, No. 6
Di Fabio et al.
(29) RS/1 v 4.4.1, Bolt Beranek and Newman Inc., 1992.
(30) TSAR v.2.2, Oxford Molecular Ltd., 1993.
(36) Preischaemia, a dose dependent reduction of the infarct volume
was observed; the maximal effect (78%) was seen at 3 mg/kg iv
(ED50 ) 0.76 mg/kg). Postischaemia, the administration of 3
mg/kg iv dose, 1, 3, and 6 h after the MCA occlusion resulted in
a significant neuroprotective effect giving 83%, 65%, and 53%
reduction of the infarct volume, respectively. (a) Reggiani, A.;
Costa, S.; Pietra, C.; Ratti, E.; Trist, D.; Ziviani, L.; Gaviraghi,
G. Neuroprotective Activity of GV150526A a Novel, Potent and
Selective Glycine Antagonist. Eur. J . Neurol. 1995, 2 (2), 6. (b)
Neuroprotection of GV150526A. Comparison with Reference
Compounds. Pietra, C.; Ziviani, L.; Valerio, E.; Tarter, G.;
Reggiani, A. Stroke 1996, 27 (1), 26. (c) Di Fabio, R.; Cugola,
A.; Donati, D.; Feriani, A.; Gaviraghi, G.; Ratti, E.; Reggiani,
A.; Trist, D. Novel Substituted Indole-2-Carboxylates as Potent
Glycine Antagonists. ACS Book of Abstracts, 211th ACS
National Meeting, New Orleans, LA, 24-28 March 1996,
Abstract 107. (c) Cugola, A.; Gaviraghi, G. Giacobbe, S. EP
568136, 3 Nov 1993.
(31) Daylight v4.41, Daylight Chemical Information Systems, 1995.
(32) (a) R2 ) cross-validated R2, LOO (leave one out),32b calcu-
cv
lated according to ref 32c. (b) Wold, S. Cross-Validatory Estima-
tion of a Number of Components in Factor and Principal
Components. Techometrics 1978, 20, 397-405. (c) Wold, S.
Validation of QSARs. Quant. Struct. Act. Relat. 1991, 10, 191-
193.
(33) The correlation matrix is the default choice for measuring the
degree of association of the parameters in the MRA module of
RS1, the program used for the statistical calculations. For this
reason, we use it here instead of the parameters’ covariance
table.
(34) Kubinyi, H. Quantitative Structure Activity Relationships. Non
Linear Dependence of Biological Activity on Hydrophobic Char-
acter: a New Model. Arzneim.-Forsch. 1976, 26, 1991-1997.
(35) Tamura, A.; Graham, D. I.; McCullogh, J .; Teasdale, G. M. Focal
Cerebral Ischemia in Rat: 1. Description of Technique and
Early Neuropathological Consequences. J . Cereb. Blood. Flow
Metab. 1981, 1, 53-60.
(37) Still, C. W.; Kahan, M.; Mitra, A. Rapid Chromatographic
Technique for Preparative Separations with Moderate Resolu-
tion. J . Org. Chem. 1978, 43, 2923-2925.
J M960644A