Synthesis of disubstituted 1,4-diazepines with affinity to GABA A-receptor subtypes

Article abstract of DOI:10.1691/ph.2010.0559

A series of tetrahydro-1H-1,4-diazepines 4a-c, dihydro-1H-1,4-diazepine 5 and pyrido diazepines 8 and 10 was prepared. Originated form dehydroacetic acid (DHA) and aromatic aldehydes cinnamoyl compounds 3a-c were obtained and converted with ethylenediamine to give tetrahydro-1H-1,4-diazepines 4a-c. For the synthesis of pyrido[1,2-d][1,4]diazepines 8 and 10 a new snythetic approach is described. Compounds 4b and 5 were investigated concerning their affinity to different benzodiazepine receptor subtypes. The determined IC₅₀ values for 5 are 1.5 μM and 1.1 μM at 10 μM respectively.

Full text of DOI:10.1691/ph.2010.0559

ORIGINAL ARTICLES
Institute of Pharmacy1, University of Leipzig; Biotie Therapies GmbH2, Radebeul, Germany
Synthesis of disubstituted 1,4-diazepines with affinity to GABAA-receptor
subtypes
D. Briel1, I. Rudolph1, K. Unverferth2, S. Mann1
Received March 2, 2010, accepted April 2, 2010
Prof. Dr. Detlef Briel, Bereich pharmazeutische Chemie, Institut für Pharmazie, Universität Leipzig, Brüderstrasse
34, D-04103 Leipzig, Germany
briel@rz.uni-leipzig.de
Pharmazie 65: 641–644 (2010)
doi: 10.1691/ph.2010.0559
A series of tetrahydro-1H-1,4-diazepines 4a–c, dihydro-1H-1,4-diazepine 5 and pyrido diazepines 8 and 10
was prepared. Originated form dehydroacetic acid (DHA) and aromatic aldehydes cinnamoyl compounds
3a–c were obtained and converted with ethylenediamine to give tetrahydro-1H-1,4-diazepines 4a–c. For
the synthesis of pyrido[1,2-d][1,4]diazepines 8 and 10 a new snythetic approach is described. Compounds
4b and 5 were investigated concerning their affinity to different benzodiazepine receptor subtypes. The
determined IC50 values for 5 are 1.5 ␮M and 1.1 ␮M at 10 ␮M respectively.
1. Introduction
Benzodiazepine derivatives are well-known for their anxi-
olytic, anticonvulsant and sedative effects. They affect the
γ-aminobutyric acid-A (GABAA) receptor by an allosteric
mechanism. The use of benzodiazepines is limited by the occur-
rence of side effects due to its unselective affinity to the α1, α2,
α3 and α5 subunits of the GABAA receptor. There is a neces-
sity for the development of subtype selective agents to improve
ditions (Hideg and Lloyd 1971; Lloyd et al. 1981). In several
cases only the olefinic carbon atom of the conjugated enone sys-
tem reacts with the amino group of ethylenediamine to form an
open-chain product and no cyclic diazepine (Hankovszky et al.
1979; Hideg and Lloyd 1971). The nucleophilic attack of the
second amino group of ethylenediamine at the carbonyl carbon
atom of the enone is dependent on the carbonyl activity and can
be favoured be electron withdrawing groups (Hankovszky et al.
1979;HidegandLloyd1971;RiedandStahlhofen1957). Conju-
pharmacological effects of benzodiazepines and simultaneously
avoid undesirable side effects (Atack et al. 2006; Collins et al.
gated enones with pyridyl-, (Samula and Jurkowska-Kowalczyk
1974) 4-hydroxy-coumarin-3-yl- (Tabakovic and Rapic 1987)
2002; Dias et al. 2005; Möhler and Rudolph 2004). The develop-
and 3-hydroxyphenyl-substituents (Fenton et al. 1985; Srivas-
ment of the sleep-inducing drugs zolpidem and zaleplon, which
tava et al. 1988) at the carbonyl group are known to give
affect the α1 subunit of the GABAA receptor, is only one exam-
the corresponding diazepines (Lloyd et al. 1981). Furthermore
ple of these efforts. Compounds selective for the subunits α2
the synthesis of 14-membered ring systems containing two
and α3 could lead to potent anxiolytic drugs with decreased hyp-
molecules of enone and two molecules of ethylenediamine was
noticeffectsandlessabusepotential(Atack2005, 2008;Dawson
reported.
et al. 2005; Dias et al. 2005; Dixon et al. 2008; Fradley et al.
Utilization of the electron withdrawing effect of the 4-hydroxy-
2007; Rowlett et al. 2005).
6-methyl-2H-pyranon structure in the present enones 3a–c and
In the course of our search for compounds with selective affin-
optimization of the reaction conditions gave the correspond-
ity to GABAA receptor subtypes (Grunwald et al. 2006) we
ing diazepines 4a–c in good yields up to 77% (Scheme 1).
were able to synthesize different partial hydrated 1,4-diazepine
We observed the best results using methylenechloride as
derivatives as potential agonists of the benzodiazepine receptor
solvent and two equivalents of ethylenediamine at room tem-
and to test them for their effect on benzodiazepine receptor.
perature. Changing the reaction settings according to already
reported studies, e.g., solvents to stabilize the diazepines (Lloyd
et al. 1981), using no solvent (Hankovszky et al. 1979; Hideg
and Lloyd 1971) or various work-up techniques (Lloyd et al.
1981), led to lower yields or no formation of the corresponding
diazepines. Although the formation of 14-membered ring sys-
2. Investigations and results
2.1. Synthesis of compounds
tems is known (Curtis 2001; Hankovszky et al. 1979; Schönherr
et al. 2006, 2004), we observed only the 7-membered diazepines
(HPLC-purity > 95%). The structure was confirmed by the eval-
uationofNOESY, COSYandHSQCspectraandNMR-diffusion
experiments in comparison to compound 2b.
Originated from dehydroacetic acid (DHA, 1) and aromatic
aldehydes 2a–c the 4-hydroxy-6-methyl-2H-pyran-2-ones 3a–c
were synthesized in a Claisen-Schmidt condensation according
to the method of Hassan et al. (1985) with small modifications
(Scheme 1). We observed the best yields (up to 46%) using
According to the method of Larsen and Jørgensen (1992) we
tolueneassolventinsteadofchloroform. Theconversionofthese
oxidized the 2,3,6,7-tetrahydro-1H-1,4-diazepine 4b to 2,3-
dihydro-1H-1,4-diazepine 5 using iodosobenzene in minor
4-hydroxy-6-methyl-2H-pyran-2-ones 3a–c with ethylenedi-
amine is known to be ambivalent. The formation of monocyclic
yields and several undefined oxidation products (Scheme 2). For
diazepines is dependent on the reactivity and on the reaction con-
Pharmazie 65 (2010)
641

ORIGINAL ARTICLES
OH
O
O
O
OH
O
O
O
OH
O
N
O
NH
a: R = H
H
a
b
b: R = 3-Cl
c: R = 4-F
O
DHA
1
R
R
R
2a-c
3a-c (31% – 46 %)
4a-c (74% – 77 %)
Scheme 1: Synthesis of cinnamoyl compounds 3a–c and 2,3,6,7-tetrahydro-1H-1,4-diazepines 4a–c a) toluene, piperidine, reflux 3 - 7 h b) CH2Cl2, H2N-(CH2)2-NH2, r.t., 2 h
Nachtwey (1924) and Wiley et al. (1955) originated from
DHA (1) yielding 2,6-dimethyl-4H-pyran-4-one or from
3-cinnamoyl-4-hydroxy-6-methyl-2H-pyranones delivering
2-methyl-6-styryl-4H-pyran-4-ones. Different trials to synthe-
size 8 according to literature methods mentioned above under
OH
O
N
OH
O
N
NH
NH
a
Cl
Cl
O
O
4b
acidic conditions failed. A further approach to synthesize 8 from
5
4b using a methanolic sodium hydroxide solution delivered
another new 1,2,3,4,5,9-hexahydropyrido[1,2-d][1,4]diazepine
c
b
10. The structure elucidation was performed by NMR. The
formation of 10 can be explained by a mechanism similar to that
described for 8. Under basic conditions an esterification of the
carboxylic acid takes place to give 9 and the decarboxylation
OH
N
OH N
NH
NH
step is no longer possible. We obtained 10 in 59% yield.
Cl
Cl
COOH
OH
COOCH
3
OH
2.2. Biological studies
6
9
The compounds 4b and 5 were tested for their affinity to different
GABAA receptor subtypes in a radioligand binding assay.
With 2,3,6,7-tetrahydro-1H-1,4-diazepine 4b (10 ␮M) we
observed an inhibition of [3H]-Ro15-1788 (flumazenil) binding
to the subtypes α2β3γ2 and α3β3γ2 of 83% and 89% respec-
tively. At a concentration of 1 ␮M 4b we observed 34% and
45% inhibition.
OH
N
N
NH
NH
O
Cl
COOCH
Cl
For the 2,3-dihydro-1H-1,4-diazepine 5 we measured similar
3
OH
affinities to both receptor subtypes. The determined IC50
values
10
7
are 1.5 ␮M to subtype α2β3γ2 and 1.1 ␮M to subtype α3β3γ2
respectively.
The results of the tests for 2,3-dihydro-1H-1,4-diazepine 5 and
pyrido[1,2-d][1,4]diazepine 8 show little affinity to the subtypes
α2β3γ2 and α3β3γ2 and no selectivity to the other GABAA
subtypes α1β2γ2 and α5β3γ2 (data not shown).
N
NH
O
Cl
2.3. Conclusions
The synthetic route to 4-hydroxy-6-methyl-2H-pyranon substi-
tuted 1,4-diazepines 4a–c was optimized and shown to give good
yields. According to a known procedure we were able to oxidize
8
Scheme 2: Synthesis of 2,3-dihydro-1,4-diazepine 5 and pyrido diazepines 8 and 10
a) CH2Cl2, iodosobenzene, 0 ◦C, 3 h, 6% b) MeOH, SiO2, r.t., 7 d
c) MeOH, NaOH, reflux, 2 h, 59%
the 2,3,6,7-tetrahydro-1,4-diazepine 4b to 2,3-dihydro-1,4-
diazepine 5. For the synthesis of pyrido[1,2-d][1,4]diazepines 8
and 10 a new synthetic method is described. The 1,4-diazepines
4b and 5 and the pyrido[1,2-d][1,4]diazepine 8 were tested for
their inhibitory effect on benzodiazepine receptor subtypes. For
compound 5 we observed IC50 values of 1.5 ␮m for subtype
α2β3γ2 and 1.1 ␮M for subtype α3β3γ2 respectively.
the 2,3-dihydro-1H-1,4-diazepin 5 an equilibrium with the cor-
responding 3,6-dihydro-2H-1,4-diazepin is easily conceivable.
In our NMR experiments we observed only one compound, the
2,3-dihydro-1H-1,4-diazepine 5, distinctly marked by a signal at
5.85 ppm for the CH-proton in position 6 at the diazepin system.
In the course of optimization of the synthesis of diazepam
4b we also tried a procedure using ether, one equivalent
of ethylenediamine and silica gel at 80 ◦C. After leaving
the reaction mixture at room temperature for one week we
observed only traces of the initial product 3b and a new
compound. NMR-studies indicated that the structure was
3. Experimental
3.1. Chemistry
3.1.1. General considerations
Chemicals were purchased from Merck, Sigma-Aldrich, Fluka or Lancaster.
Melting points were determined on a Boetius melting point apparatus and are
uncorrected. Proton (1H), carbon (13C) and fluor (19F) NMR spectra were
a
2,3,4,5-tetrahydropyrido[1,2-d][1,4]diazepin-9(1H)-one 8
(Scheme 2). These observation can be explained by an rear-
recorded on a VARIAN MERCURY-300 for solutions in deuterio chloro-
rangement under DHA-ring cleavage to give the intermediate 6
forme unless otherwise indicated. In some cases APT, HMBC, HMQC and
followed by a decarboxlyation yielding 7 and an nucleophilic
attack of the nitrogen atom at the 2-carbon atom of 7 to give
compound 8. Similar reactions were described by Arndt and
COSY spectra were recorded. NOESY spectra were recorded on a BRUKER
DRX-400 for solutions in deuterio chloroform. Residue of solvent was used
as internal standard; chemical shifts (␦) are reported in parts per million
642
Pharmazie 65 (2010)

ORIGINAL ARTICLES
Table: HPLC settings
(E)-4-Hydroxy-6-methyl-3-(7-phenyl-2,3,6,7-tetrahydro-1H-1,4-
diazepin-5-yl)-2H-pyran-2-one (4a) Originated from 3a (0.80 g,
3.12 mmol). Yellow crystals: Yield 77%; m.p. 174–177 ◦C. 1H NMR: δ
2.10 (s, 3H, CH3); 2.85–2.96 (m, 2H); 3.26 (dd, J = 5.1, 13.2 Hz, 1H);
Column
Phenomenex Hyperclone 3u C8-BDS 130 Å
150 ×
4.6 mm
3.62–3.70 (m, 1H); 3.90–3.96 (m, 2H); 4.76 (d, J = 14.4 Hz, 1H); 5.96 (s,
Detection
Software
Flow rate
Time (min)
DAD UV detection at 300 nm with DIONEX
1H, CH); 7.24–7.55 (m, 5H, aromat); 14.24 (s, 1H, OH). 13C NMR: δ 19.9
CHROMELEON
1 ml/min
Eluent A (acetonitrile)
(CH3); 40.3 (CH2); 46.6 (CH2); 48.0 (CH2); 59.0 (CH); 96.5 (C-3); 107.5
(C-5); 126.6 (C-15, 19); 127.9 (C-17); 128.8 (C-16, 18); 143.5 (C-14);
163.0 (C-6); 163.9 (C=N); 179.6 (C-4); 184.9 (C-2). IR [KBr, cm−1]: 3313,
3034, 2997, 2887, 2831, 1694, 1661, 1591, 1574. MS (ESI) (m/z): 299
(M+ + 1, 100% rel.Int.), 597 (2 M+ + 1, 7.5% rel.int.).
Eluent B (phosphate
buffer pH 2.5) (%)
80
from 80 to 50
50
from 50 to 80
80
(%)
0–10
20
C17H18N2O3.
10–15
15–35
35–38
38–43
from 20 to 50
50
from 50 to 20
20
(E)-3-(7-(3-Chlorophenyl)-2,3,6,7-tetrahydro-1H-1,4-diazepin-5-yl)-4-
hydroxy-6-methyl-2H-pyran-2-one (4b) Originated from 3b (4.35 g, 15.0
mmol). Yellow crystals: Yield 75%; m.p. 133–135 ◦C. 1H NMR: δ 2.11 (s,
3H, CH3); 2.90–2.98 (m, 2H); 3.31 (dd, J = 5.1, 13.2 Hz, 1H); 3.64–3.74
(m, 1H); 3.90–4.01 (m, 2H); 4.74 (d, J = 13.8 Hz, 1H); 5.96 (s, 1H, CH);
7.24–7.56 (m, 4H, aromat); 14.23 (s, 1H, OH). 13C NMR: δ 19.8 (CH3);
40.4 (CH2); 46.7 (CH2); 48.0 (CH2); 58.4 (CH); 96.5 (C-3); 107.5 (C-5);
(ppm) and coupling constants (J) are given in Hertz (Hz). ESI mass spectra
were recorded on a BRUKER DALTONICS ESQUIRE 3000plus. EI mass
spectra were recorded on a VG ANALYTICS VG ZAB HSQ. High resolu-
tion ESI spectra were recorded on a BRUKER DALTONICS FTICR-APEX
II. IR spectra were determined from KBr discs with a PERKIN-ELMER 16
PC FT-IR. Thin layer chromatography was performed on DC foils of Merck
(DC-Alufolien Kieselgel F254) and detected by UV. HPLC was performed
on a DIONEX apparatus. Settings are listed in the Table.
125.1 (C-18); 126.5 (C-15); 128.0 (C-19); 130.1 (C-17); 134.5 (C-16);
145.7 (C-14); 163.0 (C-6); 163.9 (C=N); 179.2 (C-4); 184.9 (C-2). IR
[KBr, cm−1]: 3495, 3286, 3051, 2947, 2826, 1683, 1659, 1598, 1574. MS
(ESI) (m/z): 333 (M+ + 1, 100% rel.int.), 665 (2 M+ + 1, 62.5% rel.int.).
C17H17ClN2O3.
(E)-3-(7-(4-Fluorophenyl)-2,3,6,7-tetrahydro-1H-1,4-diazepin-5-yl)-4-
hydroxy-6-methyl-2H-pyran-2-one (4c) Originated from 3c (0.56 g, 2.04
For column chromatography silica gel 60 pure by Merck was used (0.063–
mmol). Orange crystals: Yield 74%; m.p. 182–186 ◦C. 1H NMR: δ 2.10 (s,
0.200 mm). It was dried at 150 ◦C for 3 h, cooled and deactivated with 5%
3H, CH3); 2.83–2.97 (m, 2H); 3.27 (dd, J = 4.8, 13.2 Hz, 1H); 3.63–3.71
water (w/w). It was set aside in a closed container before use for three hours.
(m, 1H); 3.88–3.96 (m, 2H); 4.71 (d, J = 13.8 Hz, 1H); 5.69 (s, 1H, CH);
7.03 (m, 2H, aromat); 7.49 (m, 2H, aromat); 14.22 (s, 1H; OH). 13C NMR:
3.1.2. Synthetic procedures
δ 19.8 (CH3); 40.5 (CH2); 46.5 (CH2); 48.1 (CH2); 58.3 (CH); 96.5 (C-3);
107.5 (C-5); 115.4 (d, JCF = 21.4 Hz, C-16, 18); 128.2 (d, JCF = 8.2 Hz,
C-15, 19); 139.2 (d, JCF = 2.8 Hz, C-14); 163.2 (C-6); 160.8; 164.1 (d,
JCF = 244.6 Hz, C-F); 164.0 (C = N); 179.4 (C-4); 184.9 (C-2). 19F NMR: δ
-114.8. IR [KBr, cm−1]: 3295, 2977, 2813, 2788, 1688, 1652, 1588, 1570.
MS (ESI) (m/z): 317 (M+ + 1, 100% rel.Int.), 633 (2 M+ + 1.2% rel.int.).
C17H17FN2O3.
3.1.2.1. General procedure for 4-hydroxy-6-methyl-2H-pyran-2-ones 3a–c
(Birch et al. 1960; Hassan et al. 1985; Wiley et al. 1955). Dehydroacetic
acid (DHA, 1) (1 eq.) and aldehydes 2a–c (1 eq.) were dissolved in toluene,
piperidine was added and the mixture was heated for several hours. Water
was removed by azeotropic distillation. Solvent was evaporated and the
residue recrystallized from 2-propanol.
3-Cinnamoyl-4-hydroxy-6-methyl-2H-pyran-2-one (3a) Originated from
dehydroacetic acid (1) (3.36 g, 20 mmol), benzaldehyde (2a) (2.05 ml,
20 mmol) and piperidine (0.5 ml) in toluene (100 ml). Orange crystals: Yield
46%; m.p. 129–133 ◦C (Lit.: 130–132 ◦C) (Ait-Baziz et al. 2008). 1H NMR:
δ 2.27 (d, J = 0.9 Hz, 3H, CH3); 5.95 (d, J = 0.6 Hz, 1H, CH); 7.40–7.70 (m,
5H, aromat); 7.96 (d, J = 15.6 Hz, 1H, CH = CH); 8.31 (d, J = 15.9 Hz, 1H,
CH = CH). 13C NMR: δ 20.8 (CH3); 99.7 (C-3); 102.6 (C-5); 123.2 (C-9);
129.1 (C-11, 15); 129.4 (C-12, 14); 131.3 (C-13); 134.9 (C-10); 146.5 (C-8);
161.4 (C-6); 168.9 (C-4); 183.4 (C-2); 193.0 (C-7). IR [KBr, cm−1]: 3081,
1722, 1624, 1577. MS: (EI) (m/z) 256 (M+). C15H12O4.
3.1.2.3. 3-((4E,6Z)-7-(3-Chlorophenyl)-2,3-dihydro-1H-1,4-diazepin-
5-yl)-4-hydroxy-6-methyl-2H-pyran-2-one (5). Compound 4b (1.50 g,
4.50 mmol) was dissolved in dichlormethane (100 ml), freshly prepared
iodosobenzene (Saltzman and Sharefkin 1973) (0.30 g, 1.50 mmol) was
added and the suspension was stirred at 0 − 5 ◦C for 3 h. The addition of
iodosobenzene was repeated. Solvent was evaporated, keeping the bath
temperatur below 25 ◦C. The residue was purified by column chromatog-
raphy (eluent chloroform:methanol 100:3) and slowly recrystallized from
n-hexane. Slight brown powder: Yield 6%; m.p. 105–108 ◦C. 1H NMR: δ
(E)-3-[3-(3-Chlorophenyl)acryloyl]-4-hydroxy-6-methyl-2H-pyran-2-
one (3b) Originated from dehydroacetic acid (1) (16.8 g, 100 mmol),
3-chlorobenzaldeyde (2b) (12 ml, 100 mmol) and piperidine (2.0 ml) in
toluene (250 ml). Yellow needles: Yield 39%; m.p. 147–150 ◦C (Lit.:
154–156 ◦C) (Ait-Baziz et al. 2008). 1H NMR: δ 2.28 (s, 3H, CH3); 5.96 (s,
1H, CH); 7.33–7.63 (m, 4H, aromat); 7.84 (d, J = 16.2 Hz, 1H, CH = CH);
8.28 (d, J = 15.9 Hz, 1H, CH=CH); 17.68 (s, 1H, OH). 13C NMR: δ 20.9
(CH3); 99.7 (C-3); 102.4 (C-5); 124.6 (C-9);127.3 (C-14); 128.9 (C-11);
130.3 (C-13); 131.0 (C-15); 135.2 (C-10); 136.7 (C-12); 144.5 (C-8); 161.3
(C-6); 169.2 (C-4); 183.2 (C-2); 192.8 (C-7). IR [KBr, cm−1]: 3098, 1719,
1654, 1634, 1540. MS (EI) (m/z): 290 (M+).
2.10 (s, 3H, CH3); 3.67–3.71 (m, 4H, 2 CH2); 5.84 (s, 1H, CH); 6.83 (s,
1H, CH); 6.93 (s, 1H, NH); 7.29–7.56 (m, 4H, aromat); 12.80 (s, 1H; OH).
13C NMR: δ 18.6 (CH3); 46.8 (CH2); 48.1 (CH2); 91.4 (C-12); 94.1 (C-3);
106.0 (C-5); 124.7 (C-18); 126.3 (C-15); 129.2 (C-19); 129.6 (C-17);
133.7 (C-16); 139.2 (C-14); 157.4 (C-13); 160.5 (C-6); 164.0 (C=N); 166.0
(C-4); 180.5 (C-2). IR [KBr, cm−1]: 3295, 2977, 2813, 2788, 2717, 1688,
1652, 1588, 1570. MS (ESI) (m/z): 331 (M+ + 1, 100% rel.int.).
C17H15ClN2O3.
3.1.2.4. 2-(3-Chlorophenyl)-7-methyl-2,3,4,5-tetrahydropyrido[1,2-
d][1,4]diazepin-9(1H)-one (8). Compound 3b (1.16 g, 4.00 mmol) was
grinded with silica gel (4 g) for 30 min while ethylene diamine (2.8 ml,
40 mmol) was added slowly. The mixture was kept at RT. After 12 h
C15H11ClO4.
(E)-3-[3-(4-Fluorophenyl)acryloyl]-4-hydroxy-6-methyl-2H-pyran-2-
one (3c) Originated from dehydroacetic acid (1) (3.36 g, 20 mmol),
4-fluorobenzaldeyde (2c) (2.1 ml, 20 mmol) and piperidine (0.5 ml) in
toluene (60 ml). Reddish crystalline powder: Yield 31%; m.p. 140–145 ◦C
(Lit.: 147–149 ◦C) (Ait-Baziz et al. 2008). 1H NMR: δ 2.27 (d, J = 0.6 Hz,
3H, CH3); 5.95 (d, J = 0.9 Hz, 1H, CH); 7.09 (m, 2H, aromat); 7.67 (m,
2H, aromat); 7.89 (d, J = 15.9 Hz, 1H, CH=CH); 8.22 (d, J = 15.9 Hz, 1H,
CH = CH). 13C NMR: δ 20.8 (CH3); 99.6 (C-3); 102.5 (C-5); 116.5 (d,
JCF = 21.9 Hz, C-12, 14); 122.9 (d, JCF = 2.1 Hz, C-10); 131.2 (C-9); 131.4
(d, JCF = 8.7 Hz, C-11, 15); 145.1 (C-8); 161.4 (C-6); 162.9, 166.3 (d,
methanol (70 ml) was added and the suspension was stirred at RT
Silica gel was removed by filtration. The filtrate was kept in a closed
container at RT for 14 d. Methanol was evaporated and the oily residue was
for 3 h.
purified by column chromatography (eluent chloroform:methanol 100:8)
and recrystallized from acetone. Slight brown powder: Yield 30%; m.p.
168–172 ◦C. 1H NMR: δ 2.31 (s, 3H, CH3); 2.76 (d, J = 15.3 Hz, 1H of
CH2); 2.91 (dd, J = 8.7, 13.5 Hz, 1H of CH2); 3.27 (dd, J = 9.6, 15.0 Hz,
1H of CH2); 3.44 (dd, J = 6.3, 13.5 Hz, 1H of CH2); 3.83 (d, J = 9.0 Hz,
1H, CH); 4.10 (m, 1H of CH2); 4.34 (m, 1H of CH2); 6.16 (m, 2H, CH);
7.23–7.40 (m, 4H, aromat). 13C NMR: δ 21.3 (CH3); 44.7 (CH2); 48.4
(CH2); 50.6 (CH2); 62.2 (CH); 118.6, 118.8 (C-8, 10); 125.0 (C-15); 126.9
(C-12); 128.5 (C-14); 130.5 (C-16); 134,9 (C-13); 145.4; 148.6 (C-7);
152.1 (=C-N); 179.2 (C-9). IR [KBr, cm−1]): 3426, 1632, 1548. MS (ESI)
(m/z): 289 (M+ + 1, 29% rel.Int.), 577 (2 M+ + 1, 100% rel.int.).
J
CF = 251.7 Hz, C-F); 168.9 (C-4); 183.3 (C-2); 192.8 (C-7). IR [KBr,
cm−1]: 3099, 1722, 1625, 1597, 1523. MS (ESI) (m/z): 275 (M+ + 1).
C15 11FO4.
H
3.1.2.2. General procedure for 2,3,6,7-tetrahydro-1H-1,4-diazepin-5-
yl)-2H-pyran-2-ones 4a–c. 3-Cinnamoyl-4-hydroxy-6-methyl-2H-pyran-
2-ones 3a–c were dissolved in dichlormethane and ethylene diamine (2 eq.)
C16H17ClN2O.
was added. The mixtures were stirred at room temperature. After reaction
was finished, solvent was evaporated and the residue was recrystallized from
ethanol.
3.1.2.5. Methyl 2-(3-chlorophenyl)-7-methyl-9-oxo-1,2,3,4,5,9-
hexahydropyrido[1,2-d][1,4]diazepine-10-carboxylate (10). Compound
Pharmazie 65 (2010)
643

ORIGINAL ARTICLES
4b (1.08 g, 3.25 mmol) was dissolved in methanol (50 ml) and aq. 0.1 M
NaOH solution (25 ml) was added. The mixture was refluxed for 2 h, the
Dawson GR, Collinson N, Atack JR (2005) Development of subtype selec-
tive GABAA modulators. CNS Spectr 10: 21–27.
solvent was evaporated and the residue was recrystallized from acetone
(5 ml). Slightly yellow crystalline powder: Yield 59%; m.p. 222–227 ◦C. 1H
NMR: δ 2.30 (s, 3H, CH3); 2.83 (dd, J = 1.5, 15.0 Hz, 1H of CH2); 2.89 (dd,
J = 8.4, 13.2 Hz, 1H of CH2); 3.04 (dd, J = 9.0, 15.9 Hz, 1H of CH2); 3.40
(dd, J = 6.0, 13.2 Hz, 1H of CH2); 3.76 (s, 3H, OCH3); 3.89 (d, J = 8.1 Hz,
1H, CH); 4.09 (dd, J = 8.7, 15.6 Hz, 1H of CH2); 4.34 (dd, J = 6.0, 15.6 Hz,
1H of CH2); 6.02 (s, 1H, CH); 7.23–7.41 (m, 4H, aromat). 13C NMR: δ
21.7 (CH3); 42.5 (CH2); 48.3 (CH2); 50.9 (CH2); 52.6 (OCH3); 62.2 (CH);
119.3 (C-8); 124.7 (C-15); 125.1 (C-11); 126.5 (C-12); 128.2 (C-14); 130.3
(C-16); 134.8 (C-13); 146.2; 148.2; 150.0; 167.8; 175.2. IR [KBr, cm−1]:
3307, 2950, 2805, 1726, 1634, 1572. MS (HR-ESI) (m/z): 369 (M+ + Na,
100% rel.int.), 715 (2 M+ + Na, 74.7% rel.int.).
Dias R, Sheppard WFA, Fradley RL, Garrett EM, Stanley JL, Tye SJ,
Goodacre S, Lincoln RJ, Cook SM, Conley R, Hallett D, Humphries
AC, Thompson SA, Wafford KA, Street LJ, Castro JL, Whiting PJ,
Rosahl TW, Atack JR, McKernan RM, Dawson GR, Reynolds DS (2005)
Evidence for a significant role of ␣3-containing GABAA receptors in
mediating the anxiolytic effects of benzodiazepines. J Neurosci 25:
10682–10688.
Dixon CI, Rosahl TW, Stephens DN (2008) Targeted deletion of the
GABRA2 gene encoding ␣2-subunits of GABAA receptors facilitates
performance of a conditioned emotional response, and abolishes anxi-
olytic effects of benzodiazepines and barbiturates. Pharmacol Biochem
Behav 90: 1–8.
Fenton DE, Gonzalez L, S. M (1985) The reaction of 2,3,6,7-tetrahydro-
(2-hydroxyphenyl)-7-phenyl-1-H-1,4-diazepine with copper(II) and
C18H19ClN2O3.
3.2. Biology
nickel(II) salts. Trans Metal Chem 10: 437–438.
The radioligand binding assay was performed on membranes of transiently
Fradley RL, Guscott MR, Bull S, Hallett DJ, Goodacre SC, Wafford KA,
transfected HEK293 tsa cells. These cells are transfected with GABAA
Garrett EM, Newman RJ, O’Meara GF, Whiting PJ, Rosahl TW, Dawson
GR, Reynolds DS, Atack JR (2007) Differential contribution of GABAA
receptor subtypes to the anticonvulsant efficacy of benzodiazepine site
subunits α2β3γ2 and α3β3γ2 respectively by calcium phosphate DNA pre-
cipitation. 24 h after transfection the cells were scraped from the plates in
PBS and the membrane fractions prepared after different steps of separation.
3
ligands. J Psychopharmacol 21: 384–391.
The radioactive binding assay with [ H]-Ro15–1788 was performed in Mul-
Grunwald C, Rundfeldt C, Lankau H-J, Arnold T, Hofgen N, Dost R,
tiScreen glass fiber filterplates. 10–100 ␮g protein per well was incubated
3
Egerland U, Hofmann H-J, Unverferth K (2006) Synthesis, pharma-
with 1 nM [ H]-Ro15–1788 in 50 mM Tris, 200 mM NaCl, pH = 7.1 for
60 min at 4 ◦C. Nonspecific binding was estimated in the presence of 10 ␮M
cology, and structure-activity relationships of novel imidazolones and
diazepam. The assay was terminated by aspiration using the MultiScreen
pyrrolones as modulators of GABAA receptors. J Med Chem 49:
1855–1866.
Hankovszky OH, Hideg K, Lloyd D, McNab H (1979) The formation and
decomposition of 1,4,8,11-tetra-azacyclotetradeca-4,11-dienes. J Chem
Soc, Perkin Trans 15: 1345–1350.
Hassan MA, El-Kady M, El-Borai M, Abd El-Moaty AA (1985) Base cat-
vacuum manifold and washing with 200 ␮l assay buffer. After removing
residues of buffer from the plate, the plate was dryed at 50 ◦C for 2 h. 50 ␮l
liquid scintillator was applied to each well and radioactivity was counted in
a Wallac Microplate Beta counter in order to determine the specific binding
of [3H]-Ro15–1788.
alyzed additions of dehydroacetic acid an its 3-cinnamoyl derivatives.
Acknowledgement: We thank Biotie Therapies GmbH Radebeul for the per-
Rev Roum Chim 30: 245–248.
formanceoftheradioligandbindingassay. Thepresentedinvestigationswere
Hideg K, Lloyd D (1971) Reaction products from α,β -unsaturated ketones
funded by EFRE grants from the EU and by grants from the Free State of
and aliphatic diamines or dithiols. J Chem Soc C 20: 3441–3445.
Saxony (project no. 12525).
Larsen J, Jørgensen KA (1992) A facile oxidation of secondary amines to
imines by iodosobenzene or by a terminal oxidant and manganese or iron
References
porphyrins and manganese salen as the catalysts. J Chem Soc, Perkin
Trans 28: 1213–1217.
Ait-Baziz N, Rachedi Y, Chemat F, Hamdi M (2008) Solvent free
microwave-assistedknoevenagelcondensationofdehydroaceticacidwith
benzaldehyde derivatives. Asian J Chem 20: 2610–2622.
Arndt F, Nachtwey P (1924) Über Darstellung von Dehydracets Üure aus
Acetessigester und Über den Mechanismus dieser Reaktion. Ber Dtsch
Chem Ges 57: 1489–1491.
Lloyd D, Scheibelein W, Hideg K (1981) Further studies of the mixture
obtained from reactions between conjugated enones and ethylenediamine,
and from conjugated enones and 1-aminopropane. J Chem Res Synopses
3: 62–3.
Möhler H, Rudolph U (2004) Selective GABAA circuits for novel CNS
drugs. Drug Discov. Today: Therap Strat 1: 117–123.
Ried W, Stahlhofen P (1957) Über heterocyclische Siebenringsysteme,
V. Umsetzung von o-Phenylendiamin mit alpha,beta-Ungesättigten Car-
Atack JR (2005) The benzodiazepine binding site of GABAA
receptors as
a target for the development of novel anxiolytics. Expert Opin Investig
Drugs 14: 601–618.
bonylverbindungen. Chem Ber 90: 815–824.
Atack JR (2008) GABAA Receptor Subtype-Selective Efficacy: TPA023,
Rowlett JK, Platt DM, Lelas S, Atack JR, Dawson GR (2005) Differ-
ent GABA receptor subtypes mediate the anxiolytic, abuse-related,
an ␣/␣3 Selective Non-sedating Anxiolytic and ␣5IA, an ␣5 Selective
Cognition Enhancer. CNS Neurosci Ther 14: 25–35.
A
and motor effects of benzodiazepine-like drugs in primates. PNAS 102:
Atack JR, Wafford KA, Tye SJ, Cook SM, Sohal B, Pike A, Sur
915–920.
C, Melillo D, Bristow L, Bromidge F, Ragan I, Kerby J, Street
L, Carling R, Castro JL, Whiting P, Dawson GR, McKernan RM
Saltzman H, Sharefkin JG (1973) Iodosobenzene. Organic Syntheses, Coll.
Vol. 5: 658–659.
(2006) TPA023
[7-(1,1-Dimethylethyl)-6-(2-ethyl-2H-1,2,4-triazol-3-
ylmethoxy)-3-(2-fluorophenyl)-1,2,4-triazolo[4,3-b]pyridazine], an ago-
nist selective for ␣2- and ␣3-containing GABAA receptors is a
Samula K, Jurkowska-Kowalczyk E (1974) Condensation of azachalcones
with diamines. Roczniki Chemii 48: 2287–2292.
Schönherr T, Seichter W, Weber E (2006) Complexes of pendant-armed
nonsedating anxiolytic in rodents and primates. J Pharmacol Exp Ther
cyclam derivatives with copper(II) perchlorate: Synthesis and crystal
structures. Polyhedron 25: 3463–3473.
Schönherr T, Weber E, Seichter W (2004) A new tetraaza macrotricycle and
its inclusion compound with chloroform. Synthesis and crystal structures.
J Incl Phenom Macro 50: 187–191.
316: 410–422.
Birch AJ, Cameron DW, Rickards RW (1960) Studies in relation to biosyn-
thesis. XXIII. Formation of aromatic compounds from β-polyketones. J
Chem Soc 4395–4000.
Collins I, Moyes C, Davey WB, Rowley M, Bromidge FA, Quirk K,
Srivastava YK, Singh S, Verma BL (1988) Synthesis and spectral studies of
Atack JR, McKernan RM, Thompson S-A, Wafford K, Dawson GR,
some 2,3,6,7-tetrahydro-5,7-diphenyl-1H-1,4-diazepines. Nat Acad Sci
Pike A, Sohal B, Tsou NN, Ball RG, Castro JL (2002) 3-Heteroaryl-
Lett 11: 387–389.
2-pyridones: benzodiazepine site ligands with functional selectivity for
␣2/␣3-subtypes of human GABAA receptor-ion channels. J Med Chem
45: 1887–1900.
Tabakovic I, Rapic V (1987) Chemistry of coumarins. Syntheses of some
coumarin heterocycles. J Serb Chem Soc 52: 3–9.
Wiley RH, Jarboe CH, Ellert HG (1955) 2-Pyrones. XV. Substituted
3-cinnamoyl-4-hydroxy-6-methyl-2-pyrones from dehydroacetic acid. J
Am Chem Soc 77: 5102–5105.
Curtis NF (2001) Compounds of nickel(II) with rac- and meso-5,12-
dimethyl-7,14-diphenyl-1,4,8,11-tetraazacyclotetradeca-4,11-diene.
Inorg Chim Acta 317: 27–32.
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