Rigid analogues of buspirone and gepirone, 5-HT1A receptors partial agonists.

Article abstract of DOI:10.1016/S0014-827X(02)01279-X

Rigid analogues of buspirone and gepirone, 5-HT1A receptors partial agonists, were obtained. The compounds exhibited very low affinity to the receptors. Their structural features resembled to a large extent the arrangement of the respective structural elements found in the solid state of buspirone and in the theoretical structure of NAN-190 (5-HT1A postsynaptic antagonist) rigid analogue exhibiting high affinity to the receptor. The obtained results would thus suggest that the bioactive conformation of buspirone might not be the extended one. That would additionally suggest that either both groups of compounds could occupy different areas at the receptor binding sites (or bind to different receptor states) or the constrained structure of 2 does not represent well 5-HT1A receptor binding site requirements.

Full text of DOI:10.1016/S0014-827X(02)01279-X

Il Farmaco 57 (2002) 917ꢀ923
/
www.elsevier.com/locate/farmac
Rigid analogues of buspirone and gepirone, 5-HT1A receptors
partial agonists
Zdzis law Chilmonczyk a,b,*, Krzysztof J. Krajewski c, Jacek Cybulski c
a Drug Institute, 30/34 Che lmska Str., 00-725 Warsaw, Poland
b Institute of Chemistry, University of Bia lystok, Al. Pilsudskiego 11/4, 15-443 Bia lystok, Poland
c Pharmaceutical Research Institute, 8 Rydygiera St., 01-793 Warsaw, Poland
Received 21 February 2002; accepted 11 May 2002
Abstract
Rigid analogues of buspirone and gepirone, 5-HT1A receptors partial agonists, were obtained. The compounds exhibited very low
affinity to the receptors. Their structural features resembled to a large extent the arrangement of the respective structural elements
found in the solid state of buspirone and in the theoretical structure of NAN-190 (5-HT1A postsynaptic antagonist) rigid analogue
exhibiting high affinity to the receptor. The obtained results would thus suggest that the bioactive conformation of buspirone might
not be the extended one. That would additionally suggest that either both groups of compounds could occupy different areas at the
receptor binding sites (or bind to different receptor states) or the constrained structure of 2 does not represent well 5-HT1A receptor
binding site requirements.
´
# 2002 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
Keywords: 5-HT1A receptor ligands; Buspirone; Gepirone; Rigid analogues
1. Introduction
antagonistic properties at the postsynaptic 5-HT1A sites.
It was thus concluded that the constrained structure of 2
represents very well the 5-HT1A receptor binding [4].
In the present paper we report our results concerning
rigid analogues 6 and 7 of other well known 5-HT1A
Arylpiperazines have been largely investigated as a
group of compounds according to their ability to modify
serotonergic neurotransmission via 5-HT1A receptors
ligands*buspirone (3) and gepirone (4). Buspirone is as
/
interaction (for a review see Refs. [1ꢀ3]). And although
/
an approved anxiolytic drug and gepirone exhibited an
interesting anxiolytic and antidepressive profile in pre-
clinical studies. Both compounds are classified as 5-
HT1A partial agonists (for a review see Refs. [2,3]). It has
been found on the basis of crystallographic data and
quantum mechanical calculations that buspirone (and
some other related compounds) should appear in the
extended conformation [5]. On the other hand it was
also observed that the preferred solid state conformation
it has been suggested that this pharmacological class
failed to yield safer, non sedating and more effective
anxiolytic and antidepressant agents [3], considerable
efforts were exerted to investigate the receptors ligands.
The important problem of any receptor ligand is its
bioactive conformation, that is the conformation it
adopts at the molecular target site in a cell. It has
been recently postulated that NAN-190 (1), 5-HT1A
receptor postsynaptic antagonist, should adopt Z-con-
formation of the n-butyl spacer (Fig. 1). The assumption
was based on the fact that rigid NAN-190 analogues 2
(and some others) exhibited high affinity to the receptor
of buspirone analogue 5 (Kiꢁ40 nM) may depend on
/
the counterion in its salts. It was thus concluded that the
interaction with the 5-HT1A receptor might force that
class of ligands to adopt the bent conformation at the
receptor-binding site [6]. Thus we decided to obtain
buspirone and gepirone rigid analogues 6 and 7, as well
(Kiꢁ8 nM, as compared to 0.55 nM of NAN-190) and
/
as some related compounds 8ꢀ
could exhibit some affinity to 5-HT1A receptors (Fig. 2).
/
11 which in principle
* Corresponding author
E-mail address: chilmon@il.waw.pl (Z. Chilmonczyk).
´
0014-827X/02/$ - see front matter # 2002 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
PII: S 0 0 1 4 - 8 2 7 X ( 0 2 ) 0 1 2 7 9 - X

918
Z. Chilmonczyk et al. / Il Farmaco 57 (2002) 917ꢀ923
/
2. Experimental
2.1. Chemistry
aluminium oxide 90, neutral (70ꢀ230 mesh). Reagents
/
and solvents were purchased from common commercial
suppliers. The following reagents were synthesized by
published procedures: 4-hydroxycyclohexyl 4-methyl-
benzenesulfonate [7], cis- and trans-1,4-dibromo-cyclo-
hexane [8], 1-(2-pyrimidyl)piperazine [9].
Melting points were determined on a Boetius appa-
ratus (Carl Zeiss Jena) and are uncorrected. IR spectra
were recorded on a PerkinꢀElmer 1725X spectrophot-
/
1
ometer. H NMR and 13C NMR spectra were obtained
using a Brucker AMX 500 (500 MHZ) or Varian
Gemini 2000 (200 MHz) with TMS as an internal
2.1.1. cis-2-[4-(4-Bromocyclohexyl)-piperazin-1-
yl]pyrimidine (16)
A mixture of trans-1,4-dibromocyclohexane (15) (20
g, 0.08 mol), 1-(2-pyrimidyl)piperazine (5.4 g, 0.03 mol),
anhydrous K2CO3 (6.8g, 0.05 mol), KOH (2.2 g, 0.04
mol) and 20 ml toluene was refluxed for 3 h, then cooled
to room temperature, washed with water and extracted
standard. 2D NMR (Hꢀ
/
H) COSY, (CꢀH) HETCOR
/
technique was used to support interpretation of 1D
spectra. Chemical shifts are given in d (ppm) and the
coupling constants J in hertz (Hz). GC/MS spectra were
recorded on a Hewlettꢀ/Packard GC model 5890 with
with aqueous 1 M HCl solution (3ꢂ10 ml). The
/
5970 mass detector by the electron impact (EI) method.
Elementary analyses were performed in the Institute of
Organic Chemistry, Polish Academy of Sciences (War-
saw, Poland) and were within 0.4% of theoretical value.
The radioligand binding studies were performed in the
Institute of Pharmacology, Polish Academy of Sciences
(Krako´w, Poland). Column chromatography separa-
tions were carried out on Merck Kieselgel 100 or
aqueous solution was separated, alkalized to pH 9 by
the addition of 5% aq. NaOH solution, extracted with
CHCl3 (3ꢂ
under reduced pressure. The residue was chromato-
graphed (CHCl3 as eluent, Kieselgel 100, 70ꢀ230 mesh)
/20ml), dried over MgSO4, and evaporated
/
1
to give 16 (5 g, 47%). M.p.ꢁ
(CDCl3) d (ppm): 1.70ꢀ1.73 (m, 2H, Ha7, Ha7?), 1.80ꢀ
1.94 (m, 4H, Ha8, Ha8?, He7, He7?), 2.16ꢀ2.20 (m, 2H,
/
119.5ꢀ
/
120.9 8C; H NMR
/
/
/
Fig. 1. Structures of NAN 190 (1), compound 2, buspirone (3), gepirone (4) and compound 5.
Fig. 2. Structures of compounds 6ꢀ11.
/

Z. Chilmonczyk et al. / Il Farmaco 57 (2002) 917ꢀ
/923
919
He8, He8?), 2.37 (tt, Jaaꢁ
(t, Jꢁ5.1, 4H, H5, H5?), 3.83 (t, Jꢁ
4.57 (m(t), Jꢁ3.3, 1H, He9), 6.47 (t, Jꢁ
8.30 (d, Jꢁ
4.7, 2H, H2); 13C NMR (CDCl3) d (ppm):
23.82(C7), 34.13(C8), 44.08(C4), 48.92(C5), 53.58(C9),
62.35(C6), 109.73(C3), 157.67(C2), 161.66(C1).
/
10.5, Jaeꢁ
/
3.3, 1H, Ha6), 2.65
5.1, 4H, H4, H4?),
4.7, 1H, H3),
with a 5% NaOH aq. solution and concentrated to
/
/
about 1/3 of initial volume. The precipitate was collected
by filtration and dried under reduced pressure (10
mmHg/50 8C) for 10 h to give 9.5 g 4-(4-pyrimidin-2-
yl-piperazin-1-yl)cyclohexan-1-ol (cis/trans mixture).
Yield 11.5%.
/
/
/
IR (CHCl3) n (cmꢃ1): 2963, 2937, 2808, 2762, 1584,
1544, 1486, 1440, 1363, 1307, 1260, 1231, 1134, 1021,
982, 952, 800, 692, 637; MS (70 eV); m/z (%) 326 (50
Mꢄ), 245(45), 218(60), 204(80), 163(17), 148(25),
2.1.4. trans-4-(4-Pyrimidin-2-yl-piperazin-1-yl)-
cyclohexan-1-ol (9)
121(73), 108(100), 80(48), 56(65); m.p.ꢁ
120.9 8C (hexane).
/
119.5ꢀ
/
The trans isomer was separated by crystallization.
The mixture of cis/trans 4-(4-pyrimidin-2-yl-piperazin-
1-yl)cyclohexan-1-ol (9 g) was dissolved in 20 ml hot
acetone and the precipitate was washed with 5 ml
2.1.2. 2-(4-Cyclohex-3-enyl-piperazinyl)pyrimidine (17)
A mixture of cis-1,4-dibromocyclohexane (14) (20 g,
0.08 mol), 1-(2-pyrimidyl)piperazine (5.4 g, 0.03 mol),
anhydrous K2CO3 (6.8 g, 0.05 mol), KOH (2.2 g, 0.04
mol) and 20 ml toluene was refluxed for 3 h, then cooled
to room temperature, washed with water and extracted
acetone and dried to give 3.7 g (4.2%) trans isomer 9.
1
m.p.ꢁ
(ppm): 1.18ꢀ
1.86ꢀ2.08 (m, 4H, H8, H8?), 2.24ꢀ
2.59 (t, Jꢁ5.1, 4H, H5, H5?), 3.50ꢀ
3.79 (t, Jꢁ5.1, 4H, H4, H4?), 6.45 (t, Jꢁ
8.29 (d, Jꢁ
4.7, 2H, H2); 13C NMR (CDCl3) d (ppm):
161.78(C1), 157.87(C2), 109.91(C3), 70.53(C9),
/
122.3ꢀ
/
123.6 8C (acetone); H NMR (CDCl3) d
/
1.45 (m, 4H, H7, H7?), 1.70 (s, 1H, Ã
2.40 (m, 1H, Ha6),
3.66 (m, 1H, Ha9),
4.7, 1H, H3),
/
OH),
/
/
/
/
with aqueous 1 M HCl solution (3ꢂ/10 ml). The
/
/
aqueous solution was separated, alkalized to pH 9 by
the addition of 5% aq. NaOH solution, extracted with
/
CHCl3 (3ꢂ
/
20 ml), dried over MgSO4, and evaporated
62.75(C6), 49.21(C5, C5?), 44.06(C4, C4?), 34.58(C8,
C8?), 26.46(C7, C7?); IR (KBr) n (cmꢃ1): 3391, 3186,
3021, 2937, 2806, 1670, 1582, 1542, 1493, 1455, 1359,
1306, 1264, 1125, 1070, 982, 963, 811, 797; MS (70 eV);
m/z (%) 262(41 Mꢄ), 203(100), 154(57), 148(14),
142(63), 134(7), 122(21),108(21), 80(4), 56(9); Anal.
under reduced pressure. The residue was chromato-
graphed (CHCl3 as eluent, Kieselgel 100, 70-230 mesh)
to give 17. m.p.ꢁ
(CDCl3) d (ppm): 1.47 (m, 1H, Ha11), 1.95ꢀ
5H, H7, H10, He11), 2.65 (m, 5H, H6, H5, H5?), 3.83 (t,
Jꢁ5.1, 4H, H4, H4?), 4.57 (m(t), 2H, H8, H9), 6.47 (t,
Jꢁ4.7, 1H, H3), 8.30 (d, Jꢁ
4.7, 2H, H2); 13C NMR
(CDCl3) d (ppm): 25.57(C10), 25.86(C11), 27.66(C7),
44.33(C4), 48.90(C5), 60.10(C6), 109.68(C3),
/
93.8ꢀ
/
94.8 8C (hexane); 1H NMR
2.24 (m,
/
/
(C14H22N4O×
m.p.ꢁ228.5ꢀ
1.4HCl×H2O) C, H, N, Cl.
/
H2O) C, H, N. 9×
/
HCl: white powder,
/
/
/
/
230.5 8C (EtOH); Anal. (C14H22N4O×
/
/
125.53(C8), 126.84(C9), 157.65(C2), 161.62(C1); IR
(CHCl3) n (cmꢃ1): 3016, 2943, 2815, 1585, 1542, 1500,
1436, 1363, 1307, 1262, 1217, 1132, 1014, 981, 795, 778,
725, 666, 637; MS (70 eV); m/z (%) 244(79 Mꢄ),
203(10), 190(100), 175(26),149(10), 121(66), 83(30),
56(16).
2.1.5. cis-4-(4-Pyrimidin-2-ylpiperazin-1-yl)cyclo-
hexan-1-ol (8)
The filtrate (after crystallization trans-isomer 9) was
concentrated under reduced pressure to give a crude
residue, which was three times chromatographed (THF/
hexane*
Yield 3.2 g of 8 (3.8%). m.p.ꢁ
tone).
1H NMR (CDCl3) d (ppm): 1.48ꢀ
H7?, H8, H8?), 2.24ꢀ2.42 (m(tt), Jaaꢁ
Ha6), 2.63 (t, Jꢁ5.1, 4H, H5, H5?), 3.81 (t, Jꢁ
H4, H4?), 3.94ꢀ4.03 (m(t), Jꢁ
4.7, 1H, H3), 8.30 (d, Jꢁ
4.7, 2H, H2); 13C NMR
(CDCl3) d (ppm): 160.10(C1), 157.87(C2), 109.85(C3),
66.24.7(C9), 62.49(C6), 49.01(C5, C5?), 44.07(C4, C4?),
31.97(C8, C8?), 22.95(C7, C7?); IR (KBr) n (cmꢃ1):
3386, 3257, 3155, 3039, 2933, 2811, 1690, 1582, 1542,
1497, 1445, 1359, 1306, 1265, 1126, 980, 963, 804; MS
(70 eV); m/z (%) 262(44 Mꢄ), 203(100), 154(79),
148(19), 142(84), 13411), 122(34),108(41), 96(11),
/
1:1 as eluent, Kieselgel 100, 70ꢀ
/
230 mesh).
2.1.3. 4-(4-Pyrimidin-2-yl-piperazin-1-yl)cyclohexan-1-
/
138.5ꢀ140.4 8C (ace-
/
ol (8/9*mixture of cis and trans)
/
4-Hydroxycyclohexyl 4-methylbenzenesulfonate (18)
(114 g, 0.42 mol) was added dropwise during 2 h to a
warm (oil bath temperature ca. 120 8C) 1-(2-pyrimi-
dyl)piperazine (140 g, 0.85 mol). After the addition was
completed, the reaction mixture was cooled to 80 8C and
toluene was added. The resulting mixture was cooled to
0 8C and precipitate was removed by filtration. The
/1.89 (m, 8H, H7,
/
/
9.4, Jaeꢁ
/
4.1, 1H,
/
/
5.1, 4H,
/
/
3.0, 1H, He9), 6.47 (t, Jꢁ
/
/
filtrate was extracted with aqueous 1 M HCl aq. (3ꢂ
/
200 ml). The aqueous solution was alkalized to pH 9
with 25% ammonia, extracted with CHCl3, dried over
MgSO4, and evaporated under reduced pressure. The
resulting oil was dissolved in 500 ml of isopropanol,
filtered, 20 ml 48% aq. HBr was added, filtered again
and the filtrate evaporated to dryness. The solid
material was dissolved in 200 ml water, neutralized
80(13), 56(38); Anal. (C14H22N4O×
HCl; white powder, m.p.ꢁ240.0ꢀ
Anal. (C14H22N4O×1.3HCl×0.9H2O) C, H, N, Cl.
/
H2O) C, H, N. 8×
/
/
/
241.8 8C (EtOH);
/
/

920
Z. Chilmonczyk et al. / Il Farmaco 57 (2002) 917ꢀ923
/
2.1.6. cis-4-(4-pyrimidin-2-yl-piperazin-1-yl)-cyclohexyl
4-methylbenzenesulfonate (10)
white powder, m.p.ꢁ
/
167.8ꢀ169.7 8C (THF); Anal.
/
(C21H28N4O3S×2HBr×H2O) C, H, N, S, Br.
/
/
A solution of tosyl chloride (320 mg, 0.0016 mol) in 2
ml CHCl3 was added dropwise for 1 h to a cold (0 8C)
2.1.8. trans-8-Aza-8-[4-(4-pyrimidin-2-yl-piperazin-1-
yl)cyclohexyl]spiro[4.5]decane-7,9-dione (6)
solution
cis-4-(4-pyrimidin-2-yl-piperazin-1-yl)cyclo-
hexan-1-ol (8) (400 mg, 0.0015 mol) in 2 ml pyridine.
After the addition was completed, the reaction mixture
was stirred at room temperature for 24 h, diluted with 10
2.1.8.1. From cis-2-[4-(4-bromocyclohexyl)-piperazin-
1-yl]pyrimidine (16). A mixture of cis-2-[4-(4-bromocy-
clohexyl)-piperazin-1-yl]pyrimidine (16) (3.0 g, 0.009
mol), 8-azaspiro[4.5]decane-7,9-dione (4.6 g, 0.005
mol), anhydrous K2CO3 (0.8 g, 0.006 mol) and 7 ml
xylene was refluxed for 3 h, then cooled to room
temperature, washed with water and extracted with 2
ml CHCl3 and washed with water (3ꢂ20 ml). The
/
separated organic layer was dried over MgSO4 and
evaporated to dryness under reduced pressure. The
crude residue was chromatographed (CHCl3 as eluent,
Kieselgel 100, 70ꢀ
cyclohexane to give compound 10 as white needles.
Yield 370 mg (60%), m.p.ꢁ137.7ꢀ138.3 8C (cyclohex-
ane); 1H NMR (CDCl3) d (ppm): 1.42ꢀ
1.72 (m, 6H, H7,
H7?, Ha8, Ha8?), 1.94ꢀ2.00 (m, 2H, He8, He8?), 2.24ꢀ
2.42 (m, 1H, Ha6), 2.44 (s, 3H, H14), 2.59 (t, Jꢁ5.1, 4H,
H5, H5?), 3.80 (t, Jꢁ5.1, 4H, H4, H4?), 4.68ꢀ4.77 (m,
1H, He9), 6.47 (t, Jꢁ4.7, 1H, H3), 7.32 (d, Jꢁ8.1, 2H,
H12, H12?), 7.79 (d, Jꢁ8.5, 2H, H11, H11?), 8.30 (d,
Jꢁ
4.7, 2H, H2); 13C NMR (CDCl3) d (ppm):
/230 mesh) and crystallized from
M HCl aq. (3ꢂ
separated, basified to pH 9 by addition 5% NaOH aq.
solution, extracted with CHCl3 (3ꢂ20 ml), dried over
/10 ml). The aqueous solution was
/
/
/
/
/
/
MgSO4, and evaporated under reduced pressure. The
residue was chromatographed (CHCl3 as eluent, Kie-
/
/
/
selgel 100, 70ꢀ
acetone/hexane to give compound 6 (60 mg, 8%).
M.p.ꢁ183.8ꢀ185.8 8C (acetone/hexane).
/230 mesh) and then crystallized from
/
/
/
/
/
/
161.80(C1), 157.86(C2, C2?), 144.59(C10), 134.69(C13),
129.91(C11, C11?), 127.73(C12, C12?), 109.89(C3),
78.72(C9), 61.81(C6, C6?), 48.95 (C5, C5?), 44.07(C4,
C4?), 30.01(C8.C8?), 22.95(C7, C7?), 21.34(C14); IR
(KBr) n (cmꢃ1): 2957, 2835, 1592, 1543, 1510, 1353,
1167, 915, 872, 675, 555; MS (70 eV); m/z (%) 416(20
Mꢄ), 308(28), 296(35), 244(74), 203(62), 190(100),
175(31), 149(30), 136(50), 121(85), 108(95), 96(12),
2.1.8.2. From cis/trans mixture 4-(4-pyrimidin-2-yl-
piperazin-1-yl)cyclohexan-1-ol (8/9). A mixture of 4-
(4-pyrimidin-2-yl-piperazin-1-yl)cyclohexan-1-ol (0.4 g,
0.0015 mol), 1,3-dicyclohexylcarbodiimide (DCC) (0.3
g, 0.0016 mol), catalytic amount CuCl (1.5 mg) and 5 ml
CH2Cl2 was stirred for 72 h at room temperature, then
washed with water (2ꢂ
/
10ml), 25% ammonia aq. (3ꢂ10
/
ml), water (4ꢂ10 ml), dried over MgSO4, and evapo-
/
83(29), 56(30); Anal. (C21H28N4O3S) C, H, N, S. 10×
HBr, white powder, m.p.ꢁ149.1ꢀ151.1 8C (THF);
Anal. (C21H28N4O3S×1.1HBr×H2O) C, H, N, S, Br.
/
rated under reduced pressure. The residue was dissolved
in DMF (4 ml), then was added 8-azaspiro[4.5]decane-
7,9-dione (4.6 g, 0.0018 mol) and the mixture was heated
for 10 h at 110 8C. After 10 h the precipitate was
removed by filtration and the filtrate was diluted CH2Cl2
/
/
/
/
2.1.7. trans-4-(4-Pyrimidin-2-yl-piperazin-1-yl)-
cyclohexyl 4-methylbenzenesulfonate (11)
Compound 11 was prepared in the same manner as 10
from trans-4-(4-pyrimidin-2-yl-piperazin-1-yl)cyclo-
hexan-1-ol (9). The crude product was chromato-
(10 ml), washed with water (2ꢂ
with 2 M HCl aq. (3ꢂ10 ml). The aqueous layer was
separated, basified to pH 9 by addition 5% NaOH aq.
solution, extracted with CHCl3 (3ꢂ20 ml), dried over
/10 ml) and extracted
/
/
graphed (CHCl3 as eluent, Kieselgel 100, 70ꢀ
mesh) to give compound 11 (170 mg, 30%). m.p.ꢁ
119.0ꢀ
120.0 8C (cyclohexane); 1H NMR (CDCl3) d
(ppm): 1.18ꢀ2.14 (m, 8H, H7, H7?, H8, H8?), 2.20ꢀ
2.38 (m, 1H, Ha6), 2.44 (s, 3H, H14), 2.55 (t, Jꢁ5.1,
4H, H5, H5?), 3.78 (t, Jꢁ5.1, 4H, H4, H4?), 4.38 (tt, J9a-
8a,8?aꢁ10.3, J9a-8e,8?e 4.2, 1H, Ha9), 6.46 (t, Jꢁ4.7,
1H, H3), 7.32 (d, Jꢁ8.1, 2H, H12, H12?), 7.79 (d, Jꢁ
8.5, 2H, H11, H11?), 8.30 (d, Jꢁ
4.7, 2H, H2); 13C
NMR (CDCl3) d (ppm): 161.75(C1), 157.86(C2, C2?),
144.65(C10), 134.13(C13), 129.91(C11, C11?),
/
230
MgSO4, and evaporated under reduced pressure. The
residue was chromatographed (CHCl3 as eluent, Kie-
/
/
selgel 100, 70ꢀ
mg of compound 6 (yield 4%). M.p.ꢁ
(acetone/hexane). 1H NMR (CDCl3) d (ppm): 1.25ꢀ
(m, 6H, H13, H13?Ha7, Ha7?), 1.56ꢀ176 (m, 6H, H14,
H14?, He8, He8?), 1.93ꢀ2.02 (m, 2H, He7, H 7?), 2.25ꢀ
2.52 (m, 3H, H6, H 8, Ha8?), 2.56 (s, 4H, H11, H11?),
/
230 mesh) to give (isomer trans only) 30
183.8ꢀ185.8 8C
1.51
/
/
/
/
/
/
/
/
/
ꢁ
/
/
/
/
e
/
/
a
/
2.63 (t, Jꢁ
H4?), 4.53 (tt, Jaaꢁ
Jꢁ4.7, 1H, H1), 8.3 (d, Jꢁ
(CDCl3) d (ppm): 172.87(C10, C10?), 161.78(C1),
157.89(C2), 109.84(C3), 62.58(C6), 52.35(C9),
/
5.1, 4H, H5, H5?), 3.82 (m, Jꢁ
12.3, Jaeꢁ3.8, 1H, Ha9), 6.46 (t,
4.7, 2H, H2); 13C NMR
/
5.1, 4H, H4,
/
/
/
/
127.73(C12, C12?), 109.95(C3), 81.24(C9), 61.64(C6,
C6?), 49.14(C5, C5?), 43.98(C4, C4?), 31.18(C8.C8?),
26.07(C7, C7?), 21.63(C14); IR (KBr) n (cmꢃ1): 2974,
2822, 1583, 1545, 1487, 1461, 1358, 1335, 1306, 1264,
1190, 1168, 924, 891, 853, 803, 690, 560; MS (EI, HR)
48.96(C5, C5?), 45.83(C4, C4?), 44.16(C11, C11?),
39.39(C12), 37.38(C13,C13?), 27.76(C7, C7?), 27.75(C8,
C8?), 24.17(C14, C14?); IR (KBr) n (cmꢃ1): 2941, 2860,
1722, 1662, 1583, 1545, 1480, 1446, 1357, 1257, 1226,
416(Mꢄ); Anal. (C21H28N4O3S) C, H, N, S. 11×
/
HBr,

Z. Chilmonczyk et al. / Il Farmaco 57 (2002) 917ꢀ
/923
921
1147, 1135, 983, 808, 637; MS (70 eV); m/z (%) 411(20
Mꢄ), 316(12), 303(67), 248(45), 244(25), 203(100),
168(41), 163(5), 148(14), 134(10), 122(23), 108(28),
bonds were rotated (scanned with the increment of 308
under the Conformational Search option). The rotamers
were rejected if their relative energy exceeded by 4 kcal
molꢃ1 the energy of the optimal structure.
96(8), 81(17), 67(6), 56(12). 6×
m.p.ꢀ278 8C (dec.) (EtOH/THF);
(C23H33N5O2×HCl) C, H, N, Cl.
/HCl, white powder,
/
Anal.
/
2.3. Binding studies
2.1.9. trans-4,4-Dimethyl-1-[4-(4-pyrimidin-2-yl-
piperazin-1-yl)cyclohexyl]piperidine-2,6-dione (7)
The affinity of the compounds for central 5-HT1A and
5-HT2A receptors in vitro was assessed on the basis of
their ability to displace [3H]8-OH-DPAT and [3H]ke-
tanserin, respectively. Radioligand binding studies were
performed in the rat brain using the following struc-
A mixture of cis-2-[4-(4-bromocyclohexyl)-piperazin-
1-yl]pyrimidine (16) (4.0 g, 0.012 mol), 4,4-dimethylpi-
peridine-2,6-dione (5.2 g, 0.037 mol), anhydrous K2CO3
(1.7 g, 0.011 mol) and 8 ml xylene was refluxed for 3 h,
then cooled to room temperature, washed with water
tures: hippocampus (5-HT1A) and cortex (5-HT2A
)
according to the published procedures [10,11]. Buspir-
one and ritanserin were employed as reference com-
pounds. Radioligands used were [3H]8-OH-DPAT (190
Ci/mmol, Amersham) and [3H]ketanserin (60 Ci/mmol,
NEN Chemicals) for 5-HT1A and 5-HT2A, respectively.
Ki values were determined from three competition
binding experiments in which several drug concentra-
tions run in triplicates were used.
and extracted with 2 M HCl aq. (3ꢂ10 ml). The
/
aqueous solution was separated, basified to pH 9 by
addition 5% NaOH aq. solution, extracted with CHCl3
(3ꢂ
reduced pressure. The residue was chromatographed
(CHCl3 as eluent, Kieselgel 100, 70ꢀ230 mesh) and then
crystallized from acetone/hexane to give compound 7
100 mg (2.1%). M.p.ꢁ192.9ꢀ194.3 8C (AcOEt/hexane);
1H NMR (CDCl3) d (ppm): 1.05 (s, 6H, H13, H13?),
1.39 (dq, Ja7-e8 3.6, Ja7-a8 12.6, Ja7-a6 12.4, Ja7-e7
12.4, 2H, Ha7, Ha7?), 1.60 (m, 2H, He8, He8?), 1.95 (m,
2H, He7, H 7?), 2.36 (dq, Ja8-e7 3.5, Ja8-a7 12.7, Ja8-
12.7, 2H, H 8, Ha8?), 2.43ꢀ
/20 ml), dried over MgSO4, and evaporated under
/
/
/
ꢁ
/
ꢁ
/
ꢁ
/
ꢁ
/
3. Results and discussion
ꢁ
/
ꢁ
/
Compounds 6 and 7 were obtained in a reaction
sequence as shown in Fig. 3. In brief, 1,4-cyclohexane-
diol (12) was converted to 1,4-dibromocyclohexane (13)
followed by the separation of cis and trans isomers 14
and 15. trans derivative 15 was subjected to the reaction
with 1-(2-pyrimidyl)piperazine to give compound 16
which upon the reaction with 8-azaspiro[4,5]decane-7,9-
dione gave compound 6. Similarly compound 7 was
obtained by the reaction of derivative 16 with 4,4-
dimethylpiperidine-2,6-dione. In another reaction se-
quence compound 6 was obtained from the mixture of
cis and trans 4-(4-pyrimidin-2-yl-piperazin-1-yl)cyclo-
hexan-1-ol (8/9, Fig. 4). cis- (8) and trans- (9) 4-(4-
pyrimidin-2-yl-piperazin-1-yl)cyclohexan-1-ol were ob-
tained from isomeric mixture and compounds 10 and 11
by tosylation of the respective isomers 8 and 9.
The compounds’ affinities to 5-HT1A and 5-HT2A
receptors were measured in radioligand displacement
experiments. Unexpectedly the affinities of rigid analo-
gues 6 and 7 to 5-HT1A (as well as to 5-HT2A) receptors
were very low. The affinities of the buspirone (com-
pound 6) and gepirone (compound 7) analogues to 5-
HT1A receptors were 1607 and 472 nM, respectively.
Their affinity to 5-HT2A receptors was negligible.
e
a9ꢁ
5H, H6, H11, H11?), 2.62 (t, Jꢁ
(m, Jꢁ
1H, Ha9), 6.45 (t, Jꢁ
H2); 13C NMR (CDCl3) d (ppm): 172.28(C10, C10?),
161.62(C1), 157.64(C2), 109.63(C3), 62.47(C6),
/12.7, Ja8-e8
ꢁ
/
/
2.49 (m,
a
/
5.1, 4H, H5, H5?), 3.81
/
5.1, 4H, H4, H4?), 4.54 (tt, Jaaꢁ
/
12.3, Jaeꢁ
/
3.9,
/
4.7, 1H, H1), 8.3 (d, Jꢁ
/
4.7, 2H,
52.22(C9), 48.88(C5, C5?), 47.30(C11, C11), 44.11(C4,
C4?), 28.89(C12), 27.93(C7, C7?), 27.68(C8, C8?),
24.44(C13, C13?); IR (KBr) n (cmꢃ1): 2983, 2871,
1722, 1673, 1581, 1547, 1479, 1451, 1360, 1328, 1260,
1230, 1147, 1131, 978, 805, 637, 601; MS (70 eV); m/z
(%) 385(10 Mꢄ), 277(66), 265(99), 244(22), 222(46),
203(100), 175(7), 149(12), 122(24), 108(31), 96(8),
83(18), 81(11), 67(6), 56(17); Anal. (C21H31N5O2) C,
H, N. 7×
/
HCl white powder, m.p.ꢀ
/
270 8C (dec.)
(MeOH); Anal. (C21H31N5O2×
/HBr) C, H, N, Br.
2.2. Molecular modelling
The calculations were carried out with the aid of
molecular mechanics implemented in the Chem Plus 2.0
package (HyperChem 5.1., 1998). The internal MMꢄ
force field was used molecular geometry optimization
and for the conformational analysis the conjugate
/
Compounds 8ꢀ
/
11 did not exhibit an affinity (Kiꢀ
/
gradient (Polakꢀ
/
Ribier) method was used. The four
10 000) to any of the receptors (Table 1) although n-
hexyl and n-heptyl N-4-derivatives of 1-(2-pyrimidi-
nyl)piperazine were shown to be relatively good 5-HT1A
isomeric structures were constructed, corresponding to
different conformations of the piperazine ring, with
substituents located at the axial or equatorial positions.
Each of the structures was optimized in the course of the
minimal internal energy search. In the next step the three
receptor ligands (Kiꢁ84 and 60 nM, respectively) [12].
/
1H NMR spectra showed that compounds 6 and 7,
similarly to compound 2, existed in a diequatorial 1e,4e

922
Z. Chilmonczyk et al. / Il Farmaco 57 (2002) 917ꢀ923
/
Fig. 3. Synthesis of compounds 6 and 7. Reagents and conditions: (a) HBr, reflux; (b) separation; (b1) chromatography, (b2) crystallization; (c) 1-(2-
pyrimidyl)piperazine, K2CO3, KOH, xylene, reflux; (d) 8-azaspiro[4,5]decane-7,9-dione, K2CO3, xylene, reflux; (e) 4,4-dimethylpiperazine-2,6-dione,
K2CO3, xylene, reflux.
Fig. 4. Synthesis of compounds 8ꢀ
/
11. Reagents and conditions: (a) TsCl, pyridine, r.t., 24 h; (b) 1-(2-pyrimidyl)piperazine, 115 8C; (c) separation;
(c1) chromatography, (c2) crystallization; (d) DCC, CuCl; (e) 8-zaspiro[4.5]decane-7,9-dione, DMF.
˚
much more near the imide carbonyl group (7.93 A) than
chair form with respect to the cyclohexane ring. In order
˚
11.96 A).
to recognize the flexibility of the investigated structures
found in the extended structures (11.59ꢀ
/
conformational analysis with the aid of MMꢄ force
/
field implemented in the Chem Plus 2.0 package
(HyperChem 5.1., 1998) was performed. It was found
that the energetically most stable conformations of
compounds 6 and 7, differing in the orientation of
substituents at the piperazine ring, were very similar to
those found in buspirone and compound 5 hydrochlor-
ides solid states (Table 2). Those conformations of
buspirone and gepirone analogues 6 and 7 also re-
sembled the most stable theoretical conformations of
NAN-190 rigid analogue 2, the respective distances
Table 1
Affinities of compounds 6ꢀ
/
11 to 5-HT1A and 5-HT2A receptors
Comp.
Ki (nM)
5-HT1A
5-HT2A
a
c
b
Buspirone (3)
Gepirone (4)
14
32
794
3630
d
6
16079501
49299
44 36793628
117 000927 000
74 18696075
32 02095016
27 76298251
196 566931 026
7
between the aromatic and imide ring centres being
˚
8
68 903912 056
75 827912 855
13 45093526
55 693913 293
9
12.52ꢀ
/
12.67 A for different conformers of 2 [4] and
10
11
˚
12.73 A for 6. They, however, differed from the
bent conformation found in the solid state of compound
11.08ꢀ
/
a,b,c,d
5 perchlorate, where the heteroaromatic ring centre was
Data taken from Refs. [6,13ꢀ15], respectively.
/

Z. Chilmonczyk et al. / Il Farmaco 57 (2002) 917ꢀ
/923
923
Table 2
˚
Intramolecular distances (A) between an aromatic ring centroid, N4-nitrogen atom and imide oxygen atom in buspirone hydrochloride (3),
compound 5 hydrochloride and perchlorate solid state structures and theoretical low energy conformations of buspirone rigid analogue 6*
*For gepirone analogue 7 very similar results were obtained.
#Data taken from Ref. [3].
$Conformations of substituents at the piperazine ring.
[6] Z. Chilmonczyk, A. Szelejewska-Woz´niakowska, J. Cybulski, M.
Cybulski, A.E. Kozio l, M. Gdaniec, Conformational flexibility of
4. Conclusions
serotonin1A receptor ligands from crystallographic data. Updated
model of receptor pharmacophore, Arch. Pharm. Pharm. Med.
The obtained results suggest that the bioactive con-
formation of buspirone might not be the extended one.
NAN-190 (1) and its rigid analogue 2 differ from
buspirone (3), gepirone (4) and their constrained analo-
gues 6 and 7 both in the composition of the imide and
aromatic parts of their molecules. Thus the results could
suggest that either both groups of compounds occupy
different areas at the receptor binding sites, bind to
different receptor states [16] or the constrained structure
of 2 does not represent well 5-HT1A receptor binding site
requirements. Although all three rigid analogues 2, 6
and 7 exhibit smaller affinity to 5-HT1A receptors than
compounds 1, 3, and 4 (15, 94 and 15 times, respec-
tively), none of the possibilities can be excluded on the
basis of the available data.
Chem. 330 (1997) 146ꢀ160.
[7] N.A. Nelson, G.A. Mortimer, Bicyclo[3.1.0]hexane derivatives. I.
Synthesis of bicyclo[3.1.0]-2-hexanone and methyl bicy-
/
clo[3.1.0]hexane-1-carboxylate, J. Org. Chem. 22 (1957) 1146ꢀ
1153.
[8] B. Franzus, B.E. Hudson, Jr., Structural determination of cis-
/
and trans-1,3-dibromocyclohexane, J. Org. Chem. (1963) 2238ꢀ
/
2244.
[9] K.L. Howard, H.W. Steward, E.A. Conroy, J.J. Denton, Piper-
azines. II. 1-Heterocyclicpiperazines and 1-heterocyclic-4-car-
bethoxypiperazines, J. Org. Chem. 18 (1953) 1484ꢀ1488.
[10] D.N. Middlemis, J.R. Fozard, 8-Hydroxy-2-(di-n-propylamino)-
/
tetralin discriminates between subtypes of the 5-HT1 recognition
site, Eur. J. Pharmacol. 9 (1983) 151ꢀ
[11] A.J. Bojarski, M.T. Ceg la, S. Charakchieva-Minol, M.J. Mok-
rosz, M. Mac´kowiak, S. Misztal, I.L. Mokrosz, Structureꢀ
/153.
/
activity relationship studies of CNS agents. Part 9. 5-HT1A and
5-HT2 receptor affinity of some 2- and 3-substituted 1,2,3,4-
tetrahydro-carbolines, Pharmazie 48 (1993) 289ꢀ294.
/
[12] J.L. Mokrosz, M.J. Mokrosz, S. Charakchieva-Minol, M.H.
Paluchowska, A.J. Bojarski, B. Duszyn´ska, Quantitative analysis
of alkyl chain effects on the 5-HT1A and 5-HT2 receptor affinities
of 4-alkyl-1-arylpiperazines and their analogs, Arch. Pharm.
References
[1] Z. Chilmonczyk, H.Y. Aboul-Enein, 5-HT1A receptors and their
(Weinheim) 328 (1995) 143ꢀ
[13] D. Hoyer, P. Schoeffter, 5-HT receptors: Subtypes and second
messengers, J. Receptor Res. 11 (1991) 197ꢀ214.
/
148.
role in anxiety and depression, Saudi Pharm. J. 9 (2001) 123ꢀ135.
/
[2] L.E. Schechter, P. McGonigle, J.E. Barett, Serotonergic anti-
/
depressants: Current and future perspectives, Curr. Opin. CPNS
[14] J.L. Mokrosz, M. Pietrasiewicz, B. Duszyn´ska, M.T. Ceg la, Part
5. Effect of the hydrocarbon chain on the affinity of 4-substituted
1-(3-chlorophenyl)piperazines for 5-HT1A receptor site, J. Med.
Invest. Drugs 1 (1999) 432ꢀ447.
/
[3] L.R. Levine, W. Potter, 5-HT1A agonist, partial agonists and
antagonists in anxiety and depression: A lost case?, Curr. Opin.
Chem. 35 (1992) 2369ꢀ
[15] J.L. Mokrosz, L. Strekowski, B. Duszyn´ska, D.B. Harden, M.J.
Mokrosz, A.J. Bojarski, Struktureꢀactivity relationship studies of
/
2374.
CPNS Invest. Drugs 1 (1999) 448ꢀ452.
/
[4] M.H. Paluchowska, M.J. Mokrosz, A. Bojarski, A. Weso lowska,
J. Borycz, S. Charakchieva-Minol, E. Chojnacka Wo´jcik, On the
bioactive conformation of NAN-190 (1) and MP3022 (2), 5-
/
CNS agents. Part 14: Structural requirements for the 5-HT1A and
5-HT2A receptor selectivity 1-(2-pyrimidyl)piperazine derivatives,
HT1A receptor antagonists, J. Med. Chem. 42 (1999) 4952ꢀ
/4960.
Pharmazie 49 (1994) 801ꢀ806.
/
[5] Z. Chilmonczyk, A. Les´, A. Woz´niakowska, J. Cybulski, A.E.
Kozio l, M. Gdaniec, Buspirone analogues as ligands of the 5-
HT1A receptor. 1. The molecular structure of buspirone and its
[16] L. Pardo, M. Campillo, J. Giraldo, The effect of the molecular
mechanism of G protein-coupled receptor activation on the
process of signal transduction, Eur. J. Pharmacol. 335 (1997)
two analogues, J. Med. Chem. 38 (1995) 1701ꢀ
/
1710.
73ꢀ87.
/