Phenylpiperazinylmethylindolecarboxylates and derivatives as selective D(4)-ligands.

Article abstract of DOI:10.1016/S0968-0896(02)00042-1

Novel phenylpiperazinylmethylindolecarboxylates were synthesized for evaluation as potential D(4)-ligands. Test compounds showed high affinity for the human dopamine D(4) receptor and great selectivity over the other receptor subtypes. Intrinsic effects of indole derivatives, which indicated most promising binding properties, were investigated in a mitogenesis assay.

Full text of DOI:10.1016/S0968-0896(02)00042-1

Bioorganic & Medicinal Chemistry 10 (2002) 1671–1679
Phenylpiperazinylmethylindolecarboxylates and Derivatives as
Selective D4-Ligands
Annette Moll, Harald Hubner, Peter Gmeiner and Reinhard Troschutz*
Department of Medicinal Chemistry, Emil Fischer Center, Friedrich-Alexander University, Schuhstrasse 19, D-91052 Erlangen, Germany
Received 17 December 2001; accepted 23 January 2002
Abstract—Novel phenylpiperazinylmethylindolecarboxylates were synthesized for evaluation as potential D4-ligands. Test com-
pounds showed high affinity for the human dopamine D4 receptor and great selectivity over the other receptor subtypes. Intrinsic
effects of indole derivatives, which indicated most promising binding properties, were investigated in a mitogenesis assay. # 2002
Elsevier Science Ltd. All rights reserved.
Introduction
negative carboxylate moiety. We report investigations
on the effect of the relative topicity of the basic side
chain and a carboxylate moiety (3a–e) on binding
properties, as well as the synthesis of target compounds
5 and 6 and the related structures 7 and 8. Finally the
carboxylate group is replaced by other electronegative
substituents and results of dopamine receptor binding
studies are presented.
Recently, there have been numerous investigations on
the role of the dopaminergic system in mechanisms of
neurological and psychiatric disorders such as Parkin-
son’s disease or schizophrenia. New insights were pos-
sible because of the advances in molecular biological
techniques, which led to the characterization of different
dopamine receptor subtypes. The dopamine receptors
are classified in two main families—the D1-like, con-
sisting of D1 and D5 subtype, and the D2-like, including
the D2, D3 and D4 subtypes. The findings of Seeman et
al.,1 who reported elevated density of dopamine D4
receptors in post-mortem brains of schizophrenic
patients, were followed by a controverse discussion.
Chemistry
In order to find out the optimum relative topicity of the
basic side chain and the carboxylate moiety, the 3-(4-
phenylpiperazin-1-ylmethyl)indolcarboxylates 3a–e were
prepared. Treatment of the Mannich bases 2a–d 4 with 4-
phenylpiperazine in boiling toluene gave 3a-d by elim-
ination-addition reaction. Compound 3e was obtained
by reductive amination of the methyl 3-formylindole-2-
carboxylate 4, which is known in the literature,5 with 4-
phenylpiperazine and sodium triacetoxyborohydride
(Scheme 1).
In spite of this, special interest is focused on selective D4
receptor ligands, because it is known that the atypical
neuroleptic drug clozapine binds preferentially to this
receptor subtype. The selectivity over the D2 receptor is
considered responsible for the lack of extrapyramidal
motoric symptoms. Following observations by Lober et
al.2 D4 selectivity of phenylpiperazinylmethylpyr-
azolo[1,5-a]pyridines is induced by a certain negative
molecular electrostatic potential (MEP).
Results from dopamine receptor binding studies (see
Table 1) showed best binding properties for the deriva-
In the course of synthesis of functionalised indoles, we
planned to modify the structure of 3-(4-phenylpiper-
azin-1-ylmethyl)indole (1) (Fig. 1), which shows only
moderate D4 selectivity,3 by introducing an electro-
*Corresponding author. Tel.: +49-9131-852-3923; fax: +49-9131-
852-2587; e-mail: troschuetz@pharmazie.uni-erlangen.de
Figure 1.
0968-0896/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(02)00042-1

1672
A. Moll et al. / Bioorg. Med. Chem. 10 (2002) 1671–1679
using sodium triacetoxyborohydride and the adequate
phenylpiperazine in 1,2-dichloroethane (DCE) at room
temperature (Scheme 2).9
Scheme 2. (a) HNR2, NaBH(OAc)3, DCE, rt.
Furthermore, we were interested in the binding proper-
ties of the derivatives bearing the piperazinylmethyl side
chain in position 2 of the indole. To yield compounds
7a–c and 8a,b the ethyl 2-formylindole-3-carboxylates
12a–c were required. Compound 12a was obtained in a
one pot procedure by treating 1,4-benzoquinone 11 with
the enamine 1010 in a Nenitzescu indole synthesis using
EtOAc/HOAc (10/1) as solvent.11 The diethyl acetal
moiety is cleaved by the acid conditions in this reaction.
The dimethylated derivative 12b could be obtained by
methylation of 12a with MeI and K2CO3 in acetone.
Further reductive amination of the resulting aldehydes
12a–b, as described above, gave 7a,b and 8a,b (Scheme
3).
Scheme 1. Reagents and conditions: (a) HNR2, toluene, reflux;
(b) HNR2, NaBH(OAc)3, DCE, rt.
tive bearing the carboxylate moiety in the indole-2-
position (3e). Thus indole derivatives 5a–e, 6a–e, as well
as their isomeric derivatives 7a–d and 8a,b, were syn-
thesised by reductive amination of the corresponding
aldehydes (Fig. 2).
The known aldehydes 9a–e6ꢀ8 were employed as starting
material leading to indole derivatives 5a–e and 6a–e
Table 1. Receptor binding data [Ki values (nM)] of different (4-phe-
nylpiperazin-1-ylmethyl)-indole-derivatives employing bovine D1 and
human D2, D3 and D4.4 receptorsa
Results from dopamine receptor binding studies (Table
1) showed loss of affinity with alkylation of the indole-
N. Similar effects have been reported by Thurkauf et
al.12 for the affinity of phenylpiperazinylmethyl sub-
Entry
[3H]SCH23390[
3H]spiperone
D2 long D2 short D3
3400
D1
D4.4
1
19
3100
570
900
28
660
820
2.9
0.51
680
Clozapine
3a
3b
3c
3d
42041
1700
450092705510660
46,000
4700
19660
600
870
3000
11
48,000 90,000 >100,000
49,000 53,000
800
4400
32,000
3500
3800
3e
5a
3000
3700
4200
5000
1300
5700
2.5
1.9
1.5
5.0
5.1
6.1
2.5
19
19
33
5b
5c
5d
5e
6a
6b
6c
6d
6e
7a
7b
7c
7d
8a
8b
16a
16b
16c
16d
20a
20b
20c
20d
20e
2400
22,000 25,000
43,000 80,000
36,000 32,000
45,000 50,000
15,000
16,000
17,000
56,000
2000
22,000
31,000
37,000
37,000
6700
10,000
12,000
3100
3100
4700
8900
18,000
21,000
18,000
16,000
970
16,000
2500
5200
570
4900
3500
4700
6400
5000
6000
17,000
24,000
5800
43,000 65,000
44,000 48,000
67,000 36,000
17,000 25,000
1900 1400
38,000 56,000
200
4.8
34,000 14,000
7600
2000
5100
7000
1500
6200
4100
2200
3900
22,000
3300
2400
2400
2500
4.6
1.1
6.9
27,000 30,000
3000
2100
4100
2600
4900
910
1600
5600
2900
3200
510
0.66
1.2
1.4
0.76
5.4
3.3
7.3
3500
56,000 73,000
13,000 14,000
1200 1300
8800 14,000
70,000
13,000
5000
4.7
0.50
14,000
3700
aKi values are the means of 2–4 independent experiments each carried
out in triplicate.
Figure 2.

A. Moll et al. / Bioorg. Med. Chem. 10 (2002) 1671–1679
1673
from 2-cyanoindole (17) by aminomethylation (Scheme
5).
Scheme 3. Reagents and conditions: (a) 1,4-benzoquinone (11),
EtOAc/HOAc (10/1), 50 ꢁC; (b) HNR2, NaBH(OAc)3, DCE, rt; (c)
MeI, K2CO3, acetone, reflux; (d) (i) HC(OEt)3, 160 ꢁC; (ii) see (c);
(iii) EtOH, HCl (2 N), rt.
stituted pyrroles to the D2-receptor. To gain the mono-
methylated derivative 12c, in order to maintain the NH-
function and D4-affinity as well, the NH-group of 12a
had to be protected with the diethoxymethyl-(DEM)-
group13 before being methylated. Deprotection was
performed in EtOH/HCl (2 N) at room temperature and
led to 12c, which was used as a crude product in the
following reductive amination leading to 7c. The
5-unsubstituted indole derivative 7d could not be
achieved using the Nenitzescu method. Therefore the
formyl group was introduced by ortho-lithiation14,15 in
position 2 of the DEM-protected indole 13, which was
treated with DMF and subsequently hydrolysed to the
N-protected 2-formyl-indole-3-carboxylate 14. Related
structures have yet only been synthesised by oxidative
methods.16,17 Indole derivative 7d was obtained by
reductive amination and following deprotection with
EtOH/HCl (2 N) at room temperature (Scheme 4).
Scheme 5. Reagents and conditions: (a) LiAlH4, ether, 0 ꢁC; (b)
MnO2, CH2Cl2, rt; (c) H2C(CN)2, EtOH, NEt3, 0 ꢁC; (d)
H2C¼N(CH3)+2 Clꢀ, CH2Cl2, rt; (e) HNR2, toluene, reflux.
3-Z-Substituted 2-phenylpiperazinylmethylindoles 20a–
e were obtained from 2-(4-phenylpiperazin-1-ylmethyl)-
indole (19),19 a regioisomer of 1, as starting material.
Vilsmeier formylation of 19 gave the aldehyde 20b.
Following reduction of 20b with LiAlH4 in ether affor-
ded the alcohol 20a. 20b was also reacted with malo-
nonitrile in EtOH to yield the dicyanovinyl derivative
20c. Treatment of the aldehyde 20b with hydroxylamine
hydrochloride in EtOH provided the oxime 20e, which
was dehydrated to the nitrile 20d (Scheme 6).
Scheme 6. Reagents and conditions: (a) POCl3, DMF, 0 ꢁC; (b)
LiAlH4, THF, rt; (c) H2C(CN)2, EtOH, NEt3, 0 ꢁC; (d) NH2OH,
EtOH, NEt3, rt; (e) (CF3CO)2O, THF, NEt3, rt.
Scheme 4. Reagents and conditions: (a) n-BuLi, THF, ꢀ78 ꢁC; DMF,
rt; NaHCO3/H2O; (b) HNR2, NaBH(OAc)3, DCE, rt; (c) EtOH, HCl
(2 N), rt.
Results and Discussion
Dopamine D1 receptor binding was determined by
measuring the ability to displace [3H]SCH23390from
In order to get further information about the negative
MEP, that seems to induce D4 selectivity, indoles 16a–d
and 20a–e, containing an electronegative substituent Z,
were prepared. The indole-2-carboxylate 5a was reduced
with LiAlH4 in THF at room temperature to yield the
alcohol 16a, which could easily be transformed into the
aldehyde 16b by MnO2 oxidation. Following Knoeve-
nagel condensation with malononitrile in EtOH gave
rise to the dicyanovinyl derivative 16c. The indole-2-
carbonitrile 16d, an isomeric compound of previously
described 5-cyano -derivatives,18 could be achieved by
amine exchange with 4-phenylpiperazine in boiling tol-
uene from 2-cyanogramine (18), which was obtained
bovine D1 receptors.20 To assess D2 long, D2 short 21 D3
,
22
23
and D4.4 affinities cloned human dopamine receptor
subtypes stably expressed in Chinese hamster ovary cells
(CHO)20 and the radioligand [3H]spiperone were used
for competition experiments.
Binding data of the test compounds were depicted in
Table 1 and compared to those of lead structure 1 and
clozapine. An increased D4-selectivity could be observed
due to the introduction of the electronegative carboxy-
late moiety in target structures 5–8. Among the indole-
2-carboxylates bearing the basic side chain in the indole-

1674
A. Moll et al. / Bioorg. Med. Chem. 10 (2002) 1671–1679
3-position the 5-fluoro (5b) and the 5-H (5a) derivatives
showed highest affinity (Ki=1.5 and 1.9 nM, respec-
tively) to the dopamine D4 receptor and a 10- to 15-fold
selectivity over the D2-subtypes in comparison with 1. A
chloro substituent in the side chain neither increased
affinity nor selectivity for the D4-subtype in any of the
derivatives 6 or 8.
Methyl
3-(4-phenylpiperazin-1-ylmethyl)-1H-indole-6-
carboxylate (3b). Preparation and purification accord-
ing to 3a from 2b gave 3b (84.8 mg, 94%) as a colourless
powder. Mp 191–192 ꢁC; IR 3325, 2819, 1704, 1500, 1280,
1218 cmꢀ1; 1H NMR d 2.53–2.55 (m, 4H), 3.09–3.12 (m,
4H), 3.70(s, 2H), 3.85 (s, 3H), 6.73–6.76 (m, 1H), 6.88–
6.92 (m, 2H), 7.17–7.21 (m, 2H), 7.53 (d, J=2.5 Hz,
1H), 7.62 (dd, J=1.4, 8.5 Hz, 1H), 7.75 (d, J=8.5 Hz,
1H), 8.05 (d, J=1.4 Hz, 1H), 11.38 (s, br, 1H). MS (FD)
m/z: 350[M+1] +. Anal. C21H23N3O2 (349.41) C, H, N.
Variation of the electronegative group led to even more
selective dopamine D4-ligands. Best binding properties
were provided by the alcohol 16a (Ki=0.66 nM, about
3200-fold selectivity over D2 long), the nitrile 16d
Methyl
3-(4-phenylpiperazin-1-ylmethyl)-1H-indole-5-
(Ki=0.76 nM, about 6400-fold selectivity over D2 long
and the oxime 20e (Ki=0.50 nM, about 28,000-fold
selectivity over D2 short).
)
carboxylate (3c). Preparation and purification accord-
ing to 3a from 2c gave 3c (71.5 mg, 79%) as a colourless
powder. Mp 168–169 ꢁC; IR 3340, 2950, 1693, 1511,
1
1434, 1245 cmꢀ1; H NMR d 2.54–2.57 (m, 4H), 3.08–
Intrinsic effects of 5a, 7d, 16a, 16d and 20e, which
showed most promising binding profiles, were deter-
mined by a mitogenesis assay.18 All of them had partial
agonist effects between 32% (7d) and 53% (20e) com-
pared to quinpirole (100%).
3.12 (m, 4H), 3.72 (s, 2H), 3.84 (s, 3H), 6.73–6.77 (m,
1H), 6.89–6.93 (m, 2H), 7.16–7.23 (m, 2H), 7.40(s, 1H),
7.45 (d, J=8.9 Hz, 1H), 7.74 (d, J=8.9 Hz, 1H), 8.39 (s,
+
1H), 11.36 (s, br, 1H). MS (FD) m/z: 350[M+1]
Anal. C21H23N3O2 (349.41) C, H, N.
.
Methyl
3-(4-phenylpiperazin-1-ylmethyl)-1H-indole-4-
carboxylate (3d). Preparation and purification accord-
ing to 3a from 2d gave 3d (61.5 mg, 68%) as a colourless
powder. Mp 155 ꢁC; IR 3345, 2822, 1715, 1598, 1496,
Experimental
All materials were employed as received from a com-
mercial supplier, unless noted otherwise. Solvents were
purified and dried by standard procedures. All anhy-
drous reactions were performed in oven-dried glassware.
Reactions were monitored by thin layer chromato-
graphy on analytical plates (Merck, silica gel 60F, alu-
minium back) and analysed with 254 nm UV light.
Chromatographic purification of the target compounds
was performed using silica gel 60(Merck). Melting
points: Buchi melting point apparatus, uncorrected. IR
Jasco FT/IR 410and Perkin-Elmer 1740. MS were run
on Finnigan MAT TSQ 70and 8200spectrometers with
EI (70eV) or on Jeol MS 700 with FD. 1H NMR spectra
were obtained on a Bruker AM 360(360MHz) spec-
trometer in DMSO-d6 relative to TMS. Microanalyses
were performed by the institute of inorganic chemistry
(Friedrich Alexander University, Erlangen) using Carlo
Erba Elemental Analyser 1108 and are withinÆ0.4% of
the theoretical values if not noted otherwise.
1
1349, 1269 cmꢀ1; H NMR d 2.35–2.42 (m, 4H), 2.98–
3.03 (m, 4H), 3.68 (s, 2H), 3.84 (s, 3H), 6.73–6.77 (m,
1H), 6.87–6.90(m, 2H), 7.12–7.20(m, 3H), 7.30–7.33
(m, 1H), 7.43 (s, br, 1H), 7.58 (d, J=7.5 Hz, 1H), 11.30
(s, br, 1H). MS (FD) m/z: 350[M+1] +. Anal.
C21H23N3O2 (349.41) C, H, N.
Methyl
3-(4-phenylpiperazin-1-ylmethyl)-1H-indole-2-
carboxylate (3e). To a mixture of 4 (140mg, .0691
mmol) and N-phenylpiperazine (112 mg, 0.691 mmol) in
5 mL of DCE sodium triacetoxyborohydride (205 mg,
0.967 mmol) was added under nitrogen atmosphere.
After 8 h of stirring at room temperature saturated
NaHCO3 solution was added and the mixture was
extracted with CH2Cl2. The dried and concentrated
organic layer was purified by flash chromatography
(cyclohexane/EtOAc=7:3) to afford 3e (135 mg, 56%)
as a colourless powder. Mp 170–172 ꢁC; IR 3316, 2950,
1
1685, 1492, 1454, 1234 cmꢀ1; H NMR d 2.56–2.61 (m,
Methyl
3-(4-phenylpiperazin-1-ylmethyl)-1H-indole-7-
4H), 3.07–3.11 (m, 4H), 3.89 (s, 2H), 4.07 (s, 3H), 6.73–
6.76 (m, 1H), 6.88–6.91 (m, 2H), 7.05–7.09 (dd, J=7.1,
8.2 Hz, 1H), 7.16–7.20(m, 2H), 7.25–7.29 (dd, J=7.1,
carboxylate (3a). Methyl 3-dimethylaminomethyl-
indole-7-carboxylate (2a) (60mg, 0.259 mmol) was dis-
solved in 5 mL of toluene and 4-phenylpiperazine (83.9
mg, 0.518 mmol) was added. The mixture was refluxed
for 4 h. After evaporation of the solvent, the residue was
purified by column chromatography (cyclohexane/
EtOAc=5:5) to give 3a (61.0mg, 68%) as a colourless
powder. Mp 133–135 ꢁC; IR 3448, 2915, 2819, 1697,
8.2 Hz, 1H), 7.43 (d, J=8.2 Hz, 1H), 7.89 (d, J=8.2 Hz,
+
1H), 11.71 (s, br, 1H). MS (FD) m/z: 350[M+1]
Anal. C21H23N3O2 (349.41) C, H, N.
.
Ethyl 3-(4-phenylpiperazin-1-ylmethyl)-1H-indole-2-car-
boxylate (5a). Preparation and purification according
to 3e from 9a gave 5a (195 mg, 78%) as a colourless
powder. Mp 175 ꢁC; IR 3320, 2935, 1681, 1496, 1454,
1500, 1284 cmꢀ1
;
1H NMR d 2.52–2.55 (m, 4H,
ArNCH2CH2), 3.09–3.11 (m, 4H, ArNCH2CH2), 3.72
(s, 2H, ArCH2), 3.95 (s, 3H, OCH3), 6.73–6.77 (m, 1H,
H-40), 6.88–6.92 (m, 2H, H-20/60), 7.11–7.20(m, 3H, H-
30/50/5), 7.34 (d, J=2.5 Hz, 1H, H-2), 7.78 (dd, J=0.7,
6.7 Hz, 1H, H-6), 8.00 (d, J=7.5 Hz, 1H, H-4), 11.05 (s,
br, 1H, NH, exchangeable with D2O). MS (FD) m/z:
350[M+1] +. Anal. C21H23N3O2 (349.41) calcd C 72.19;
H 6.63; N 12.03; found C 72.33; H; 7.10; N 11.91.
1
1226 cmꢀ1; H NMR d 1.37 (t, J=7.1 Hz, 3H), 2.56–
2.59 (m, 4H), 3.07–3.10 (m, 4H), 4.06 (s, 2H), 4.36 (q,
J=7.1 Hz, 2H), 6.73–6.77 (m, 1H), 6.87–6.92 (m, 2H),
7.05–7.09 (m, 1H), 7.15–7.20 (m, 2H), 7.22–7.28 (m,
1H), 7.44 (d, J=8.2 Hz, 1H), 7.88 (d, J=7.8 Hz, 1H),
11.66 (s, br, 1H). MS (EI) m/z: 363 [M]+. Anal.
C22H25N3O2 (363.44) C, H, N.

A. Moll et al. / Bioorg. Med. Chem. 10 (2002) 1671–1679
1675
Ethyl
5-fluoro-3-(4-phenylpiperazin-1-ylmethyl)-1H-
(m, 2H), 7.07 (dd, J=8.2 Hz, 1H), 7.19–7.21 (m, 2H),
7.26 (dd, J=8.2 Hz, 1H), 7.44 (d, J=8.2 Hz, 1H), 7.87
(d, J=8.2 Hz, 1H), 11.65 (br s, 1H). MS (EI) m/z: 397
[M]+. Anal. C22H24N3O2Cl (397.89) (ꢂ0.25H2O) C, H,
N.
indole-2-carboxylate (5b). Preparation and purification
(cyclohexane/EtOAc=95:5) according to 3e from 9b
gave 5b (192 mg, 73%) as a colourless powder. Mp
175 ꢁC; IR 3320, 2935, 2850, 1681, 1600, 1500, 1461,
1238 cmꢀ1; H NMR d 1.37 (t, J=7.1 Hz, 3H), 2.56–
1
2.58 (m, 4H), 3.08–3.10 (m, 4H), 4.03 (s, 2H), 4.36 (q,
J=7.1 Hz, 2H), 6.73–6.77 (m, 1H), 6.88–6.92 (m, 2H),
7.11–7.20(m, 3H), 7.45 (dd, J=4.6, 8.9 Hz, 1H), 7.61
(dd, J=2.5, 9.9 Hz, 1H), 11.78 (br s, 1H). MS (FD) m/z:
416 [M+1]+. Anal. C22H24N3O2F (381.43) C, H, N.
Ethyl 3-[4-(4-chlorophenyl)piperazin-1-ylmethyl]-5-fluoro-
1H-indole-2-carboxylate (6b). Preparation and purifica-
tion (cyclohexane/EtOAc=95:5) according to 6a from
9b gave 6b (124 mg, 65%) as a colourless powder. Mp
178–179 ꢁC; IR 3320, 1681, 1496, 1458, 1346, 1241
cmꢀ1; 1H NMR d 1.36 (t, J=7.1 Hz, 3H), 2.54–2.57 (m,
4H), 3.08–3.10 (m, 4H), 4.02 (s, 2H), 4.36 (q, J=7.1 Hz,
2H), 6.88–6.92 (m, 2H), 7.14 (ddd, J=2.5, 8.9, 9.2 Hz,
1H), 7.18–7.22 (m, 2H), 7.45 (dd, J=4.6, 9.2 Hz, 1H),
7.61 (dd, J=2.5, 8.9 Hz, 1H), 11.78 (br s, 1H). MS (FD)
m/z: 416 [M+1]+. Anal. C22H23N3O2ClF (415.88) C,
H, N.
Ethyl
5-chloro-3-(4-phenylpiperazin-1-ylmethyl)-1H-
indole-2-carboxylate (5c). Preparation and purification
(cyclohexane/EtOAc=95:5) according to 3e from 9c
gave 5c (150mg, 55%) as a colourless powder. Mp 205–
206 ꢁC; IR 3297, 2766, 1682, 1455, 1397, 1260cm ꢀ1; 1H
NMR d 1.37 (t, J=7.1 Hz, 3H), 2.56–2.59 (m, 4H),
3.08–3.11 (m, 4H), 4.04 (s, 2H), 4.37 (q, J=7.1 Hz, 2H),
6.73–6.77 (m, 1H), 6.88–6.92 (m, 2H), 7.16–7.21 (m,
2H), 7.27 (dd, J=8.8, 1.8 Hz, 1H), 7.46 (d, J=8.5 Hz,
1H), 7.93 (d, J=1.8 Hz, 1H), 11.88 (br s, 1H). MS (EI)
m/z: 397 [M]+. Anal. C22H24N3O2Cl (397.89) C, H, N.
Ethyl 5-chloro-3-[4-(4-chlorophenyl)piperazin-1-ylmethyl]-
1H-indole-2-carboxylate (6c). Preparation and purifica-
tion (cyclohexane/EtOAc=95:5) according to 6a from
9c gave 6c (114 mg, 57%) as a colourless powder. Mp
170–171 ꢁC; IR 3320, 2923, 1681, 1496, 1454, 1234
cmꢀ1; 1H NMR d 1.37 (t, J=7.1 Hz, 3H), 2.55–2.59 (m,
4H), 3.08–3.12 (m, 4H), 4.03 (s, 2H), 4.36 (q, J=7.1 Hz,
2H), 6.88–6.93 (m, 2H), 7.18–7.22 (m, 2H), 7.26 (dd,
J=2.0, 8.9 Hz, 1H), 7.45 (d, J=8.9 Hz, 1H), 7.92 (d,
J=2.0Hz, 1H), 11.87 (br s, 1H). MS (EI) m/z: 432
[M]+. Anal. C22H23N3O2Cl2 (432.33) (ꢂ2H2O) C, H, N.
Ethyl 5-methoxy-3-(4-phenylpiperazin-1-ylmethyl)-1H-
indole-2-carboxylate (5d). Preparation and purification
(cyclohexane/EtOAc=95:5) according to 3e from 9d
gave 5d (171 mg, 63%) as a colourless powder. Mp
184 ꢁC; IR 3333, 2769, 1681, 1466, 1385, 1259 cmꢀ1; 1H
NMR d 1.36 (t, J=7.1 Hz, 3H), 2.57–2.59 (m, 4H),
3.09–3.12 (m, 4H), 3.76 (s, 3H), 4.04 (s, 2H), 4.34 (q,
J=7.1 Hz, 2H), 6.73–6.77 (m, 1H), 6.88–6.91 (m, 2H),
6.93 (dd, J=8.9, 2.5 Hz, 1H), 7.17–7.21 (m, 2H), 7.34
(d, J=8.9 Hz, 1H), 7.34 (d, J=2.5 Hz, 1H), 11.53 (br s,
1H). MS (FD) m/z: 394 [M+1]+. Anal. C23H27N3O3
(393.43) C, H, N.
Ethyl 3-[4-(4-chlorophenyl)piperazin-1-ylmethyl]-5-meth-
oxy-1H-indole-2-carboxylate (6d). Preparation and
purification (CHCl3/acetone=9:1) according to 6a from
9d gave 6d (124 mg, 63%) as a colourless powder. Mp
ꢁ
1
173–174 C; IR 3305, 2827, 1677, 1496, 1241 cmꢀ1; H
NMR d 1.36 (t, J=7.1 Hz, 3H), 2.55–2.59 (m, 4H),
3.09–3.11 (m, 4H), 3.76 (s, 3H), 4.03 (s, 2H), 4.34 (q,
J=7.1 Hz, 2H), 6.88–6.93 (m, 2H), 6.93 (dd, J=2.3, 8.7
Hz, 1H), 7.17–7.21 (m, 2H), 7.31–7.35 (m, 2H), 11.52
(br s, 1H). MS (EI) m/z: 428 [M]+. Anal.
C23H26N3O3Cl (427.95) C, H, N.
Ethyl 5,6-dimethoxy-3-(4-phenylpiperazin-1-ylmethyl)-
1H-indole-2-carboxylate (5e). Preparation and purifi-
cation (cyclohexane/EtOAc=95:5) according to 3e from
9e gave 5e (180mg, 62%) as a colourless powder. Mp
170 ꢁC; IR 3328, 2942, 2842, 1673, 1496, 1234 cmꢀ1; 1H
NMR d 1.35 (t, J=7.1 Hz, 3H), 2.55–2.58 (m, 4H),
3.07–3.11 (m, 4H), 3.76 (s, 3H), 3.79 (s, 3H), 4.03 (s,
2H), 4.31 (q, J=7.1 Hz, 2H), 6.71–6.74 (m, 1H), 6.87 (s,
1H), 6.87–6.93 (m, 2H), 7.18–7.22 (m, 2H), 7.35 (s, 1H),
11.36 (br s, 1H). MS (FD) m/z: 424 [M+1]+. Anal.
C24H29N3O4 (423.47) C, H, N.
Ethyl 3-[4-(4-chlorophenyl)piperazin-1-ylmethyl]-5,6-di-
methoxy-1H-indole-2-carboxylate (6e). Preparation and
purification (cyclohexane/EtOAc=95:5) according to
6a from 9e gave 6e (180mg, 67%) as a colourless pow-
der. Mp 195 ꢁC; IR 3336, 2823, 1673, 1496, 1245 cmꢀ1
;
1H NMR d 1.35 (t, J=7.1 Hz, 3H), 2.54–2.57 (m, 4H),
3.08–3.12 (m, 4H), 3.76 (s, 3H), 3.79 (s, 3H), 4.02 (s,
2H), 4.31 (q, J=7.1 Hz, 2H), 6.88 (s, 1H), 6.90–6.94 (m,
2H), 7.18–7.21 (m, 2H), 7.33 (s, 1H), 11.36 (br s, 1H);
MS (EI) m/z: 458 [M]+. Anal. C24H28N3O4Cl (457.92)
(ꢂ0.25H2O) C, H, N.
Ethyl
3-[4-(4-chlorophenyl)piperazin-1-ylmethyl]-1H-
indole-2-carboxylate (6a). To a mixture of 9a (100 mg,
0.461 mmol) and N-(4-chlorphenyl)piperazine (90 mg,
0.461 mmol) in 5 mL of DCE sodium triacetoxybor-
ohydride (137 mg, 0.645 mmol) was added under nitro-
gen atmosphere. After 8 h of stirring at room
temperature saturated NaHCO3 solution was added and
the mixture was extracted with CH2Cl2. The dried and
concentrated organic layer was purified by flash chro-
matography (cyclohexane/EtOAc=9:1) to afford 6a
(98.6 mg, 58%) as a colourless powder. Mp 153–155 ꢁC;
IR 3342, 2772, 1772, 1594, 1498, 1235 cmꢀ1; 1H NMR d
1.37 (t, J=7.1 Hz, 3H), 2.55–2.58 (m, 4H), 3.07–3.10
(m, 4H), 4.05 (s, 2H), 4.36 (q, J=7.1 Hz, 2H), 6.88–6.92
Ethyl
5-hydroxy-2-(4-phenylpiperazin-1-ylmethyl)-1H-
indole-3-carboxylate (7a). Preparation and purification
(cyclohexane/EtOAc=95:5) according to 3e from 12a
gave 7a (167 mg, 64%) as a colourless powder. Mp 192–
194 ꢁC; IR 3289, 2932, 1655, 1466, 1232 cmꢀ1; 1H NMR
d 1.35 (t, J=7.1 Hz, 3H), 2.60–2.63 (m, 4H), 3.17–3.19
(m, 4H), 4.05 (s, 2H), 4.26 (q, J=7.1 Hz, 2H), 6.65 (dd,
J=8.5, 2.5 Hz, 1H), 6.74–6.78 (m, 1H), 6.91–6.95 (m,

1676
A. Moll et al. / Bioorg. Med. Chem. 10 (2002) 1671–1679
2H), 7.17–7.25 (m, 3H), 7.35 (d, J=2.5 Hz, 1H), 8.87
(s,1H), 11.78 (br s, 1H); MS (EI) m/z: 379 [M]+. Anal.
C22H25N3O3 (379.43) C, H, N.
MS (EI) m/z: 413 [M]+. Anal. C22H24N3O3Cl (413.90)
C, H, N.
Ethyl 2-[4-(4-chlorophenyl)piperazin-1-ylmethyl]-5-meth-
oxy-1-methyl-1H-indole-3-carboxylate (8b). Preparation
and purification (cyclohexane/EtOAc=95:5) according
to 6a from 12b gave 8b (152 mg, 75%) as a colourless
powder. Mp 146–148 ꢁC; IR 2905, 2813, 1678, 1499,
Ethyl
5-methoxy-1-methyl-2-(4-phenylpiperazin-1-yl-
methyl)-1H-indole-3-carboxylate (7b). Preparation and
purification (cyclohexane/EtOAc=95:5) according to 3e
from 12b gave 7b (213 mg, 76%) as a colourless powder.
Mp 166–167 ꢁC; IR 2923, 2820, 2383, 1677, 1531, 1407
cmꢀ1; 1H NMR d 1.38 (t, J=7.1 Hz, 3H), 2.60–2.63 (m,
4H), 3.08–3.11 (m, 4H), 3.79 (s, 3H), 3.84 (s, 3H), 4.15
(s, 2H), 4.30(q, J=7.1 Hz, 2H), 6.74–6.78 (m, 1H),
6.88–6.92 (m, 3H), 7.16–7.21 (m, 2H), 7.46 (d, J=8.9
Hz, 1H), 7.55 (d, J=2.5 Hz, 1H). MS (EI) m/z: 40 7
[M]+. Anal. C24H29N3O3 (407.48) C, H, N.
1
1382 cmꢀ1; H NMR d 1.38 (t, J=7.1 Hz, 3H), 2.59–
2.62 (m, 4H), 3.08–3.11 (m, 4H), 3.79 (s, 3H), 3.83 (s,
3H), 4.14 (s, 2H), 4.29 (q, J=7.1 Hz, 2H), 6.89 (dd,
J=8.9, 2.5 Hz, 1H), 6.91–6.93 (m, 2H), 7.20–7.23 (m,
2H), 7.46 (d, J=8.9 Hz, 1H), 7.54 (d, J=2.5 Hz, 1H);
MS (EI) m/z:441 [M]+. Anal. C24H28N3O3Cl (441.95)
C, H, N.
Ethyl 5-methoxy-2-(4-phenylpiperazin-1-ylmethyl)-1H-
indole-3-carboxylate (7c). 12a (100 mg, 0.429 mmol)
was refluxed in ethyl orthoformiate for 8 h. Evaporation
of the reagent and purification by column chromato-
graphy (cyclohexane/ EtOAc=94:6) gave a crude pro-
duct (112 mg, 78%), which was methylated according to
12b. Chromatographic purification (cyclohexane/
EtOAc=95:5) afforded (93.5 mg, 80%) of the methy-
lated raw product. Hydrolysis in EtOH/HCl (2 N)
according to 7d gave 12c (66.2 mg, 100%). Following
reductive amination according to 3e from 12c (60mg,
0.25 mmol), N-phenylpiperazine (41 mg, 0.25 mmol)
and sodium triacetoxyborohydride (74 mg, 0.35 mmol)
gave 7c (65.1 mg, 68%) as colourless crystals. Mp 185–
186 ꢁC; IR 3023, 2823, 1596, 1496, 1234 cmꢀ1; 1H NMR
d 1.37 (t, J=7.1 Hz, 3H), 2.61–2.63 (m, 4H), 3.17–3.20
(m, 4H), 3.77 (s, 3H), 4.07 (s, 2H), 4.28 (q, J=7.1 Hz,
2H), 6.74–6.77 (m, 1H), 6.80(dd, J=2.5, 8.5 Hz, 1H),
6.91–6.94 (m, 2H), 7.18–7.23 (m, 2H), 7.35 (d, J=8.5
Hz, 1H), 7.48 (d, J=2.5 Hz, 1H), 11.66 (br s, 1H). MS
(FD) m/z: 394 [M+1]+. Anal. C23H27N3O3 (393.43)
(ꢂ0.25H2O) C, H, N.
Ethyl 2-formyl-5-hydroxy-1H-indole-3-carboxylate (12a).
1,4-Benzoquinone 11 (648 mg, 6 mmol) was dissolved
in EtOAc/HOAc (10:1) and the enamine 10 (434 mg, 2
mmol) was added dropwise at 50 ꢁC; the reaction mix-
ture was stirred at 50 ꢁC for 16 h; after cooling a solu-
tion of 1.03g NaOAc in H2O was added dropwise under
stirring at room temperature for 2 h. Then pH was
adjusted to 7.5 with 2 N NaOH and a yellow precipitate
was collected. Recrystallisation from ethylacetate gave
12a (242 mg, 52%) of pale yellow crystals. Mp 159–
160 ꢁC; IR 3284, 2989, 1676, 1651, 1589, 1235 cmꢀ1; H
1
NMR d 1.39 (t, J=7.1 Hz, 3H), 4.37 (q, J=7.1 Hz, 2H),
6.94 (dd, J=8.9, 2.5 Hz, 1H), 7.36 (d, J=8.9 Hz, 1H),
7.46 (d, J=2.5 Hz, 1H), 9.41 (br s, 1H), 10.45 (s, 1H),
12.50(br s, 1H). MS (EI) m/z: 233 [M]+. Anal.
C12H11NO4 (233.22) (ꢂ0.25H2O) C, H, N.
Ethyl 2-formyl-5-methoxy-1-methyl-1H-indole-3-carboxyl-
ate (12b). 12a (233 mg, 1.0mmol) was dissolved in ace-
tone. K2CO3 (276 mg, 2.0mmol) and MeI (2.5 mL, 40
mmol) were added and the mixture was refluxed for 10
h. After evaporation of the solvent the residue was sus-
pended in H2O. Extraction with ether and crystal-
lisation from EtOH gave 12b (154 mg, 59%) as pale
yellow crystals. Mp 115–116 ꢁC; IR 22987, 1700, 1651,
Methyl
2-(4-phenylpiperazin-1-ylmethyl)-1H-indole-3-
carboxylate (7d). 15 (30 mg, 0.067 mmol) was dissolved
in EtOH. After addition of one drop of aqueous HCl
(2 N) and 2 h of stirring at room temperature, the mix-
ture was neutralised by addition of saturated NaHCO3
solution (aq.). Extraction with ether gave 7d (23.4 mg,
100%) as colourless crystals. Mp 177–178 ꢁC; IR 3401,
1
1493, 1275 cmꢀ1; H NMR d 1.41 (t, J=7.1 Hz, 3H),
3.83 (s, 3H), 4.04 (s, 3H), 4.39 (q, J=7.1 Hz, 2H), 7.15
(dd, J=9.2, 2.5 Hz, 1H), 7.56 (d, J=2.5 Hz, 1H), 7.65
(d, J=9.2 Hz, 1H), 10.60 (s, 1H). MS (EI) m/z: 261
[M]+. Anal. C14H15NO4 (261.28) C, H, N.
1
2927, 1681, 1454, 1226 cmꢀ1; H NMR d 2.62–2.66 (m,
4H), 3.18–3.22 (m, 4H), 3.83 (s, 3H), 4.11 (s, 2H), 6.75–
6.79 (m, 1H), 6.92–6.95 (m, 2H), 7.13–7.27 (m, 2H),
Methyl 1-diethoxymethyl-2-formyl-1H-indole-3-carboxyl-
ate (14). 13 (145.5 mg, 0.5 mmol) was dissolved in
THF under nitrogen atmosphere and cooled to-78 ꢁC; n-
BuLi (1.6 M in hexane) (0.34 mL, 0.55 mmol) was
added. The mixture was stirred at ꢀ78 ꢁC for 15 min
and at room temperature for another 30min. After
cooling again to ꢀ78 ꢁC DMF (40 mL, 0.52 mmol) was
added. The mixture was allowed to warm and the reac-
7.18–7.23 (m, 2H), 7.45–7.48 (m, 1H), 7.94–7.97 (m,
+
1H), 11.80(br s, 1H). MS (FD) m/z: 350[M+1]
Anal. C21H23N3O2 (349.41) C, H, N.
.
Ethyl
2-[4-(4-chlorophenyl)piperazin-1-ylmethyl]-5-
hydroxy-1H-indole-3-carboxylate (8a). Preparation and
purification (cyclohexane/EtOAc=95:5) according to
6a from 12a gave 8a (139 mg, 73%) as a colourless
powder. Mp 203 ꢁC; IR 3286, 2815, 1654, 1461, 1338,
tion was stopped after 10min by addition of NaHCO
solution. Extraction with ether gave 14 (154 mg, 96%)
3
as a colourless powder. Mp 141–143 ꢁC; IR 2977, 1712,
1
1
1230cm ꢀ1; H NMR d 1.35 (t, J=7.1 Hz, 3H), 2.59–
1673, 1454, 1261 cmꢀ1; H NMR d 1.09–1.13 (m, 6H),
2.62 (m, 4H), 3.16–3.19 (m, 4H), 4.04 (s, 2H), 4.26 (q,
J=7.1 Hz, 2H), 6.64 (dd, J=8.5, 2.1 Hz, 1H), 6.93–6.96
(m, 2H), 7.20–7.23 (m, 2H), 7.23 (d, J=8.5 Hz, 1H),
7.35 (d, J=2.1 Hz, 1H), 8.87 (s,1H), 11.51 (br s, 1H).
3.41–3.50(m, 2H), 3.67–3.76 (m, 2H), 3.96 (s, 3H), 7.26
(s, 1H), 7.33–7.39 (m, 1H), 7.43–7.49 (m, 1H), 7.95–8.00
(m, 1H), 8.13–8.20(m, 1H), 10.62 (s, 1H). MS (EI) m/z:
305 [M]+. Anal. C16H19NO5 (305.28) C, H, N.

A. Moll et al. / Bioorg. Med. Chem. 10 (2002) 1671–1679
1677
Methyl
1-diethoxymethyl-2-(4-phenylpiperazin-1-yl-
3 h. After evaporation of the solvent and column chro-
matography (cyclohexane/ethylacetate 9:1) 16d (44.5
mg, 80%) was yielded as a colourless powder. Mp 184–
methyl)-1H-indole-3-carboxylate (15). Preparation and
purification (cyclohexane/EtOAc=9:1) according to 3e
from 14 (42 mg, 0.138 mmol), N-phenylpiperazine (21.7
mg, 0.138 mmol) and sodium triacetoxyborohydride
(40.5 mg, 0.195 mmol) gave 15 (34.2 mg, 55%) as col-
ourless crystals. Mp 159–160 ꢁC; IR 2977, 1700, 1500,
186 ꢁC; IR 3444, 2946, 2221, 1558, 1538, 1234 cmꢀ1; H
1
NMR d 2.56–2.58 (m, 4H), 3.10–3.12 (m, 4H), 3.84 (s,
2H), 6.72–6.76 (m, 1H), 6.88–6.91 (m, 2H), 7.13–7.20
(m, 3H), 7.31–7.36 (m, 1H), 7.43 (d, J=8.2 Hz, 1H),
7.83 (d, J=8.2 Hz, 1H), 12.23 (s, br, 1H). MS (FD) m/z:
317 [M+1]+. Anal. C20H20N4 (316.21) C, H, N.
1457, 1222 cmꢀ1; H NMR d 1.28–1.33 (m, 6H) 2.62–
1
2.68 (m, 4H), 3.08–3.12 (m, 4H), 3.44–3.52 (m, 2H),
3.76–3.84 (m, 2H), 3.87 (s, 3H), 4.20(s, 2H), 6.66 (s,
1H), 6.74–6.79 (m, 1H), 6.90–6.93 (m, 2H), 7.17–7.23
(m, 4H), 7.82–7.85 (m, 1H), 8.00–8.03 (m, 1H). MS (EI)
m/z: 452 [M+1]+. Anal. C26H33N3O4 (451.53) C, H, N.
3-Dimethylaminomethyl-1H-indole-2-carbonitrile
(18).
Indole-2-carbonitrile (17) (70mg, 0.50mmol) and N,N-
dimethylmethyleneimmonium chloride (56.1 mg, 0.60
mmol) were dissolved in 8 mL of dry CH2Cl2. After 18 h
of stirring at room temperature and neutralisation with
NaHCO3 (aq) the mixture was extracted with CH2Cl2.
Purification was performed by column chromatography
(cyclohexane/ethylacetate 8:2) to give 18 (86.6 mg, 87%)
as a colourless powder. Mp 173–174 ꢁC; IR 3316, 2944,
[3-(4-Phenylpiperazin-1-ylmethyl)-1H-indol-2-yl]-metha-
nol (16a). 5a (180mg, 0.496 mmol) was suspended in 5
mL of dry ether (0 ꢁC, nitrogen atmosphere). Dry THF
was added until dissolution of 5a. 0.50 mL of a LiAlH4
solution (1 M in THF) were added. The reaction mix-
ture was stirred for 10min Saturated NaHCO 3 soln was
added. Neutralisation with 2 N HCl and extraction with
ether gave 16a (132 mg, 83%) as a colourless powder.
1
2825, 2220, 1718, 1455, 1348 cmꢀ1; H NMR d 2.17 (s,
6H), 3.68 (s, 2H), 7.13–7.16 (m, 1H), 7.31–7.36 (m, 1H),
7.42–7.45 (m, 1H), 7.76–7.79 (m, 1H), 12.20(s, br, 1H).
MS (FD) m/z: 200 [M+1]+. Anal. C12H13N3 (199.26)
(ꢂ0.25H2O) C, H, N.
ꢁ
ꢀ1
Mp 175–177 C; IR 3231, 2825, 1598, 1455, 1230cm
;
1H NMR d 2.52–2.55 (m, 4H), 3.06–3.09 (m, 4H), 3.67
(s, 2H), 4.66 (s, 2H), 5.75 (s, br, 1H), 6.73–6.77 (m, 1H),
6.88–6.91 (m, 2H), 6.95 (ddd, J=1.4, 7.5, 8.5 Hz, 1H),
7.03 (ddd, J=1.4, 7.5, 8.5 Hz, 1H), 7.16–7.21 (m, 2H),
7.32 (d, J=8.5 Hz, 1H), 7.59 (d, J=7.5 Hz, 1H), 10.97
(s, br, 1H). MS (FD) m/z: 322 [M+1]+. Anal.
C20H23N3O (321.41) (ꢂ0.8H2O) C, H, N.
[2-(4-Phenylpiperazin-1-ylmethyl)-1H-indol-3-yl]-methanol
(20a). 20b (180mg, 0.564 mmol) was suspended in 5 mL
of dry ether (0 ꢁC, nitrogen atmosphere). Dry THF was
added until dissolution of 20b. 0.54 mL of a LiAlH4
solution (1 M in THF) were added. The reaction mix-
ture was stirred for 10min and quenched with saturated
NaHCO3 soln. Neutralisation with 2 N HCl and
extraction with ether gave 20a (166 mg, 92%) as a col-
ourless powder. Mp 189–191 ꢁC; IR 3178, 2823, 1596,
3-(4-Phenylpiperazin-1-ylmethyl)-1H-indole-2-carbalde-
hyde (16b). 16a (100 mg, 0.31 mmol) was dissolved in
CH2Cl2. MnO2 was added and the mixture stirred for 4
h at room temperature. After filtering off the MnO2 and
evaporation of the solvent, the residue was purified by
column chromatography (cyclohexane/ethylacetate 8:2)
to give 1ꢁ6b (88.0mg, 89%) as a colourless powder. Mp
1
1446, 1245 cmꢀ1; H NMR d 2.55–2.58 (m, 4H), 3.12–
3.14 (m, 4H), 3.70(s, 2H), 4.65 (s, 3H), 6.74–6.78 (m,
1H), 6.90–6.93 (m, 2H), 6.96 (ddd, J=1.0, 7.0, 8.1 Hz,
1H), 7.04 (ddd, J=1.0, 7.0, 8.5 Hz, 1H), 7.17–7.22 (m,
2H), 7.31 (d, J=8.5 Hz, 1H), 7.57 (d, J=8.1 Hz, 1H),
10.92 (s, br, 1H). MS (FD) m/z: 322 [M+1]+. Anal.
C20H23N3O (321.41) (ꢂ0.85H2O) C, H, N.
176–177 C; IR 3286, 2814, 1643, 1454, 1242 cmꢀ1; H
1
NMR d 2.57–2.61 (m, 4H), 3.08–3.13 (m, 4H), 4.06 (s,
2H), 6.73–6.77 (m, 1H), 6.88–6.92 (m, 2H), 7.09–7.13
(m, 1H), 7.17–7.21 (m, 2H), 7.31–7.35 (m, 1H), 7.43 (d,
J=8.2 Hz, 1H), 7.89 (d, J=8.2 Hz, 1H), 10.18 (s, 1H),
11.80(s, br, 1H). MS (FD) m/z: 320[M+1] +. Anal.
C20H21N3O (319.40) ꢂ0.75H2O) C, H, N.
2-(4-Phenylpiperazin-1-ylmethyl)-1H-indole-3-carbalde-
hyde (20b). To a solution of POCl3 (79.0mg, .0515
mmol) in 5 mL of DCE, DMF (37.6 mg, 0.515 mmol)
was added at 0 ꢁC. After stirring the mixture at room
temperature for 30min, 2-(4-phenylpiperazin-1-yl-
methyl)indole (19) (100 mg 0.344 mmol) dissolved in 2
mL of DCE was added followed by 2 more hours of
stirring at room temperature. After addition of 0.5 g
NaOAc in 1 mL H2O and 1 h of stirring, the pH was
adjusted to 8 (NaHCO3). Extraction with CH2Cl2 and
column chromatography (cyclohexane/ethylacetate 5:5)
gave 20b (84.4 mg, 77%) as a colourless powder. Mp
2-[3-(4-Phenylpiperazin-1-ylmethyl)-1H-indol-2-yl-methyl-
ene]malononitrile (16c). 16b (40mg, 0.126 mmol) and
malononitrile (16.6 mg, 0.126 mmol) were dissolved in 3
mL of EtOH at 0 ꢁC. One drop of triethylamine was
added and the mixture stirred at room temperature.
After 3 h, 16c (38.6 mg, 84%) was collected as a yellow
precipitate. Mp 175–176 ꢁC; IR 3367, 2933, 2825, 2221,
1587, 1492 cmꢀ1; H NMR d 2.53–2.57 (m, 4H), 3.07–
1
206–208 ꢁC; IR 3234, 2822, 1627, 1460cm ꢀ1; H NMR
1
3.11 (m, 4H), 3.95 (s, 2H), 6.73–6.77 (m, 1H), 6.88–6.92
(m, 2H), 7.13–7.21 (m, 3H), 7.38–7.42 (m, 1H), 7.63 (d,
J=8.2 Hz, 1H), 7.88 (d, J=8.2 Hz, 1H), 8.62 (s, 1H),
11.24 (s, br, 1H);. MS (FD) m/z: 368 [M+1]+. Anal.
C23H21N5 (367.45) C, H, N.
d 2.61–2.64 (m, 4H), 3.16–3.18 (m, 4H), 4.06 (s, 2H),
6.75–6.79 (m, 1H), 6.90–6.94 (m, 2H), 7.16–7.24 (m,
4H), 7.46–7.48 (m, 1H), 8.10(m, 1H), 10.21 (s, 1H),
12.09 (s, br, 1H). MS (FD) m/z: 320[M+1] +. Anal.
C20H21N3O (319.40) (ꢂ0.75H2O) C, H, N.
3-(4-Phenylpiperazin-1-ylmethyl)-1H-indole-2-carbonitrile
(16d). 18 (35 mg, 0.176 mmol) and 4-phenylpiperazine
(65 mg, 0.40 mmol) were refluxed in 5 mL of toluene for
2-[2-(4-Phenylpiperazin-1-ylmethyl)-1H-indol-3-ylmethyl-
ene]malononitrile (20c). 20b (40mg, 0.126 mmol) and

1678
A. Moll et al. / Bioorg. Med. Chem. 10 (2002) 1671–1679
malononitrile (16.6 mg, 0.126 mmol) were dissolved in 3
mL of EtOH at 0 ꢁC. One drop of triethylamine was
added and the mixture stirred at room temperature.
After 3 h 20c (38.9 mg, 84%) was collected as a yellow
precipitate. Mp 211–212 ꢁC; IR 3270, 2923, 2211, 1573,
In brief, 10,000 cells/well were seeded in a 96-well plate
and grown for 75 h. Medium was removed and the test
compounds were incubated in medium without serum
for 20 h with a final concentration of 0.001–10000 nM.
After addition of 0.25 mCi of [3H]thymidine and further
incubation for 2 h, cells were trypsinised, harvested onto
GF/C filters and the incorporated radioactivity was
counted in a 96-well scintillation counter.
1
1502, 1454 cmꢀ1; H NMR d 2.59–2.62 (m, 4H), 3.16–
3.18 (m, 4H), 4.00 (s, 2H), 6.75–6.79 (m, 1H), 6.91–6.95
(m, 2H), 7.18–7.22 (m, 2H), 7.24–7.33 (m, 2H), 7.51–
7.55 (m, 1H), 8.08 (dd, J=1.4, 7.1 Hz, 1H), 8.56 (s, 1H),
12.76 (s, br, 1H). MS (FD) m/z: 368 [M+1]+. Anal.
C23H21N5 (367.45) C, H, N.
Protein concentration was established by the method of
Lowry using bovine serum albumin as standard.25
2-(4-Phenylpiperazin-1-ylmethyl)-1H-indole-3-carbonitrile
(20d). 20e (50mg, 0.150mmol) was dissolved in 5 mL of
THF (0 ꢁC, N2-atmosphere). After addition of triethyl-
amine (15.2 mg, 0.30 mmol) and trifluoroacetic acid
anhydride (15.8 mg, 0.150 mmol) the mixture was stir-
red for 4 h at room temperature. After extraction with
CH2Cl2 the combined organic layers were washed with
water, dried and concentrated in vacuo. Purification
was performed by column chromatography (cyclohex-
ane/ethylacetate 7:3) to give 20d (31.8 mg, 67%) as a
colourless powder. Mp 149–150 ꢁC; IR 3259, 2931,
Data analysis
The resulting competition curves of the receptor binding
experiments were analysed by nonlinear regression
using the algorithms in PRISM (GraphPad Software,
San Diego, CA, USA). The data were fit in accordance
to a sigmoid model to provide an IC50 value, represent-
ing the concentration corresponding to 50% of maximal
inhibition, and then transformed to Ki values applying
the equation of Cheng and Prusoff.26 Data resulting
from experiments investigating the stimulation of mito-
genesis were each normalised and then combined to get
a mean curve. Analysing this curve as described above
yielded an EC50 value expressing the concentration,
which caused half of the maximal rate of incorporation
of the radioactive marker to derive the rate of agonistic
effect for each test compound in correlation to the
reference agonist quinpirole.
1
2827, 2213, 1504, 1226 cmꢀ1; H NMR d 2.60–2.63 (m,
4H), 3.16–3.19 (m, 4H), 3.60(s, 2H), 6.75–6.80(m, 1H),
6.92–6.95 (m, 2H), 7.18–7.25 (m, 4H), 7.50(d, J=7.8
Hz, 1H), 7.58 (d, J=7.1 Hz, 1H), 12.24 (s, br, 1H). MS
(FD) m/z: 317 [M+1]+. Anal. C20H20N4 (316.41) C, H,
N.
2-(4-Phenylpiperazin-1-ylmethyl)-1H-indole-3-carbalde-
hyde oxime (20e). 20b (50mg, .0157 mmol) and
hydroxylamine hydrochloride (12 mg, 0.174 mmol) were
dissolved in EtOH. After addition of one drop of tri-
ethylamine, the mixture was refluxed for 3 h. After
cooling 20e (33 mg, 63%) was collected as colourless
crystals. Mp 224 ꢁC; IR 3320, 2819, 1596, 1442, 1226,
1133 cmꢀ1; 1H NMR d 2.55–2.59 (m, 4H), 3.13–3.17 (m,
4H), 3.80(s, 2H), 6.74–6.78 (m, 1H), 6.90–6.93 (m, 2H),
7.03–7.07 (m, 1H), 7.11–7.15 (m, 1H), 7.18–7.22 (m,
2H), 7.37 (d, J=8.2 Hz, 1H), 7.97 (d, J=7.8 Hz, 1H),
8.44 (s, 1H), 10.45 (s, br, 1H), 11.41 (s, br, 1H). MS
(FD) m/z: 335 [M+1]+. Anal. C20H22N4O (334.41)
(ꢂ0.25H2O) C, H, N.
Acknowledgements
The authors thank Dr. H. H. M. Van Tol (Clarke
Institute of Psychiatry, Toronto), Dr. J.-C. Schwartz
and Dr. P. Sokoloff (INSERM, Paris) as well as Dr. J.
Shine (The Garvan Institute of Medical Research, Syd-
ney) for providing dopamine D4, D3 and D2 receptor
expressing cell lines, respectively. Dr. R. Huff (Pharma-
cia & Upjohn, Inc., Kalamazoo, MI, USA) is acknowl-
edged for providing a D4 expressing cell line employed
for mitogenesis. Thanks are also due to Mrs. H. Szcze-
panek, Mrs. P. Schmitt and Mrs. P. Huebner for skilful
technical assistance.
Receptor binding studies
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