Synthesis, 5-HT1A and 5-HT2A receptor affinity and QSAR study of 1-benzhydryl-piperazine derivatives with xanthine moiety at N4

Article abstract of DOI:10.1007/s00044-011-9555-y

Three novel 1-benzhydryl-piperazines with xanthine moiety at N4 were synthesized and tested for 5-HT_1A and 5-HT_2A receptor affinity. One of the compounds showed the highest affinity (58.6 nM) and selectivity (34 times) to 5-HT_2A receptor known for this class of compounds. A set of the three new and 31 previously synthesized 1-arylpiperazines with xanthine moiety at N4 was compiled and a QSAR study was performed in order to rationalize the further synthesis. It was found that the 5-HT_1A affinity depends on the shape of the molecules (ovality and number of circuits), the distribution of the electron density in the structures (partial charges at piperazine N1 and xanthine N1) and their charge transfer ability (HOMO energy). The 5-HT_2A affinity depends on the lipophilicity of the ligands and the distribution of the electron density in the structures (partial charges at piperazine N4 and xanthine O6). The QSAR results are in a good agreement with known pharmacophore models. Springer Science+Business Media, LLC 2011.

Full text of DOI:10.1007/s00044-011-9555-y

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2012) 21:477–486
DOI 10.1007/s00044-011-9555-y
ORIGINAL RESEARCH
Synthesis, 5-HT1A and 5-HT2A receptor affinity and QSAR study
of 1-benzhydryl-piperazine derivatives with xanthine moiety at N4
•
•
•
Iva Valkova Alexander Zlatkov Krystyna Ne˛dza
Irini Doytchinova
Received: 5 July 2010 / Accepted: 6 January 2011 / Published online: 19 January 2011
Ó Springer Science+Business Media, LLC 2011
Abstract Three novel 1-benzhydryl-piperazines with
xanthine moiety at N4 were synthesized and tested for
5-HT1A and 5-HT2A receptor affinity. One of the com-
pounds showed the highest affinity (58.6 nM) and selec-
tivity (34 times) to 5-HT2A receptor known for this class of
compounds. A set of the three new and 31 previously
synthesized 1-arylpiperazines with xanthine moiety at N4
was compiled and a QSAR study was performed in order to
rationalize the further synthesis. It was found that the
5-HT1A affinity depends on the shape of the molecules
(ovality and number of circuits), the distribution of the
electron density in the structures (partial charges at piper-
azine N1 and xanthine N1) and their charge transfer ability
(HOMO energy). The 5-HT2A affinity depends on the
lipophilicity of the ligands and the distribution of the
electron density in the structures (partial charges at piper-
azine N4 and xanthine O6). The QSAR results are in a
good agreement with known pharmacophore models.
nervous systems. It is important for a variety of physio-
logical functions, including platelet aggregation, smooth
muscle contraction, appetite, cognition, perception, mood,
and other CNS functions (Hoyer et al., 1994; Roth, 1994).
These diverse activities are mediated by large number of
5-HT receptor subtypes that are encoded by distinct genes.
It now appears that there are at least 15 receptor subtypes
that belong to seven families (Nichols and Nichols, 2008).
Apart from 5-HT3, all 5-HT receptors are coupled to
G-proteins (GPCR). The 5-HT1 subtypes decrease whereas
5-HT4, 5-HT5, 5-HT6 and 5-HT7 increase the cellular level
of cAMP. 5-HT2 increases the levels of inosine trisphos-
phate (IP3) and diglyceride (DAG). 5-HT3 is a ligand-gated
Na? and K? cation channel.
1-Arylpiperazines are well known 5-HT receptor ligands
with 5-HT1A and/or 5-HT2A affinity (Oh et al., 2001;
Lopez-Rodriguez et al., 2002). The inclusion of a complex
heterocyclic fragment through a polymethylene spacer at
piperazine position N4 often leads to more potent ligands
in both 5-HT1A and 5-HT2A affinities (Nagamoto et al.,
2004). In the present study we describe three new
1-benzhydryl-piperazine derivatives with xanthine moiety
attached to N4 without any spacer or with a trimethylene
one. Xanthines are well known mild CNS agents acting as
antagonists of adenosine receptors (Daly, 2007). The new
compounds have been synthesized initially as vasodilators
and their 5-HT affinity was found by chance. They are
weak 5-HT1A binders but one of them is the best known
5-HT2A binder in the class of N4-xanthine derivatives of
1-arylpiperazine. It shows strong 5-HT2A selectivity being
34 times more affine to 5-HT2A than to 5-HT1A. Addi-
tionally, a set of similar 1-arylpiperazines with xanthine
moiety at N4, tested previously (Chojnacka-Wojcik et al.,
1995; Pawlowski et al., 1999; Pawlowski et al., 2000;
Chlon et al., 2001; Kolaczkowski et al., 2005), was
Keywords Piperazine ꢀ Xanthine ꢀ 5-HT1A and 5-HT2A
ꢀ
QSAR
Introduction
5-Hydroxytryptamine (5-HT, serotonin) is an indoleamine
neurotransmitter found in both the central and peripheral
I. Valkova ꢀ A. Zlatkov ꢀ I. Doytchinova (&)
Faculty of Pharmacy, Medical University of Sofia,
2 Dunav st, 1000 Sofia, Bulgaria
e-mail: idoytchinova@pharmfac.net
K. Ne˛dza
Institute of Pharmacology, Polish Academy of Sciences,
´
12 Smetna St, 31-343 Krakow, Poland
123

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Med Chem Res (2012) 21:477–486
compiled and a QSAR study was performed in order to
rationalize the further synthesis.
12 h. After the completion of reaction, the solvent was
removed and the waxy residue was treated with cold
chloroform to separate triethylamine hydrochloride. After
filtration the solvent was removed and the residue was
dissolved in acetone and was allowed to stand at 5°C
overnight. The solid crystals obtained were filtered, washed
with cold acetone, dried and recrystallized from, acetone.
Yield 62% (2.75 g). M.p. 222–224°C. IR: 3075 (CH aro-
matic), 1705 (CO xanthine), 1664 (CO xanthine), 1648
with shoulders at 1608 and 1637 (C=N, C=C aromatic),
1558 and 1539 (C=C xanthine, C=C aromatic), 755 and
746 (CH aromatic); 1H NMR: 2.53–2.49 m (4H, 2 9
–CH2–, piperazine), 2.36–3.29 m (4H, 2 9 –CH2–, piper-
azine), 3.61, 3.33 and 3.17 (s, 3 9 3H, CH3), 4.51s (1H,
–CH side chain), 7.49–7.19 (m, 10H, aromatic ring), 13C
NMR: 161.14 (C8xanth); 155.6 (C6xanth CO); 153.26
(C2xanth CO); 141.8, 128.67, 127.71, 127.18 (12 9 C
aromatic ring); 150.9 (C5xanth); 104.38 (C4xanth); 48.87
(2 9 N4piperazine, CH2), 50.94 (2 9 N1piperazine, CH2);
74.77 (CH–(Ph)2 side chain), 32.31.2 (N7xanth CH3),
29.35 (N1xant CH3), 27.34 (N3xanth CH3); MS m/z: 167
(100), 445 (M?1). Anal. C25H28N6O2 (C, H, N).
Materials and methods
Chemistry
Melting points were determined on an electrothermal
apparatus (Bu¨chi 535, Switzerland) in an open capillary
tube and are not corrected. The IR spectra 400–4000 cm-1
were recorded on a Shimadzu FTIR 8101 M spectropho-
tometer (Shimadzu, Japan) in Nujol. The 1H NMR and 13
C
NMR spectra were measured on a Bruker Avance DRX
250 (250 MHz) spectrometer (Germany). 1H NMR spectra
were measured for approximately 0.03 M while 13C NMR
spectra were measured for approximately 0.05 M solutions
using DMSO-d6 as solvent and chemical shifts were
expressed as d values in ppm against TMS as an inter-
nal standard. Mass spectra (MS) were recorded on an
Esquire-LC-00075 spectrometer.TLC was performed on
DC-Alufolien Kieselgel 60 F254, 0.20 mm (Merck, Ger-
many) sheets with the mobile phase: chloroform/acetone/
ethanol (3:3:4, V/V). The spots were detected at UV
254 nm. All names were generated by using structure-to-
name and name-to-structure algorithms included with
ChemBioDraw Ultra 11.0 (CambridgeSoft).
Synthesis of 1-benzhydryl-4-[1,3,7-trimethyl-2,6(3H,7H)-
dioxo-1H-purine-8-aminopropyl]-piperazine (6)
A mixture of 8-(3-bromopropylamino)-1,3,7-trimethyl-1H-
purine-2,6(3H,7H)-dione (2) (3.30 g, 0.01 mol), 1-ben-
zhydrylpiperazine (2.56 g, 0.01 mol) and triethylamine
(1.01 g, 0.01 mol) was refluxed in DMF (20 ml) for 13 h.
After the completion of reaction, the reaction content was
allowed to stand at 20°C overnight. The crude crystals
obtained were filtered, washed with water to separate tri-
ethylamine hydrochloride, dried and recrystallized from
isopropanol. Yield 55% (2.76 g). M.p. 187–188°C. IR:
3068 (CH aromatic), 1704 (CO xanthine), 1668 (CO xan-
thine), 1649 with shoulders at 1607 and 1635 (C=N, C=C
aromatic), 1555 and 1540 (C=C xanthine, C=C aromatic),
Synthetic grade chemicals procured from Merck, Ger-
many, were used for the synthesis of the target compounds,
as received. Non-commercially available intermediates
required for the synthesis of novel derivatives of 1-ben-
zhydrylpiperazine were prepared according to the literature
procedures without modifications as follows: 8-bromocaf-
feine (1) (Gagausov et al., 1987); 8-(3-bromopropylamino)-
1,3,7-trimethyl-1H-purine-2,6(3H,7H)-dione (2) (Peikov
et al., 1990), 1-(3-iodoropropyl)-3,7-dimethylxanthine (3)
(Peikov et al., 1988). The completion of reactions was
monitored through TLC.
1
Microanalyses were performed on a Perkin-Elmer
2400-II Element analyzer. Analyses indicated by the
symbols of the elements or functions were within 0.4% of
the theoretical values. All solutions were dried over
anhydrous sodium sulfate and evaporated on a Bu¨chi rotary
evaporator at reduced pressure.
754 and 744 (CH aromatic); H NMR: 1.73–1.67 t (2H,
–CH2–, J = 7 Hz, side chain), 2.51–2.31 m (10H, 4 9
–CH2–, piperazine, –CH2, N1 piperazine), 3.80–3.74 m
(2H, –CH2– side chain) 3.52, 3.30 and 3.16 (s, 3 9 3H,
CH3), 4.25 s (1H, –CH side chain), 4.34 s (1H, –NH side
chain), 7.42–6.96 (m, 10H, aromatic ring), 13C NMR:
154.2 (C6xanth CO); 152.9 (C2xanth CO); 150.9
(C8xanth); 142.9, 128.5, 127.6, 126.8 (12 9 C aromatic
ring); 148.4 (C5xanth); 101.8 (C4xanth); 52.9 (2 9
N4piperazine, CH2), 55.5 (2 9 N1piperazine, CH2); 75.16
(CH–(Ph)2 side chain), 51.6 (side chain CH2, N1pipera-
zine), 41.3 (CH2, –NH side chain), 29.18 (N7xanth CH3),
27.14 (N1xant CH3), 26.3 (side chain CH2), 25.49
(N3xanth CH3); MS m/z: 167 (100), 502 (M?1). Anal.
C28H35N7O2 (C, H, N).
The given yields are those of analytically pure product.
No efforts were made to optimize the yields.
Synthesis of 1-benzhydryl-4-[1,3,7-trimethyl-2,6(3H,7H)-
dioxo-1H-purine-8-yl]-piperazine (5)
A mixture of 8-bromocaffeine (1) (2.73 g, 0.01 mol),
1-benzhydrylpiperazine (2.56 g, 0.01 mol) and triethyl-
amine (1.01 g, 0.01 mol) was refluxed in DMF (20 ml) for
123

Med Chem Res (2012) 21:477–486
479
Synthesis of 1-benzhydryl-4-[3-(3,7-dimethyl-2,6(3H,7H)-
dioxo-1H-purine-1-yl)-propyl]-piperazine (7)
previously for 5-HT1A and 5-HT2A affinity (Chojnacka-
Wojcik et al., 1995; Pawlowski et al., 1999; Pawlowski
et al., 2000; Chlon et al., 2001; Kolaczkowski et al., 2005).
The pKi values range from 4.9 to 9.3 for 5-HT1A and from
4.4 to 7.2 for 5-HT2A (Table 1). Although the compounds
originate from different papers, their affinities are compa-
rable as they are tested by the same binding assay (Bojarski
et al., 1993) in the same lab (Institute of Pharmacology,
Polish Academy of Sciences).
A mixture of 1-(3-iodopropyl)-3,7-dimethyl-1H-purine-
2,6(3H,7H)-dione (3) (3.48 g, 0.01 mol), benzhydrylpip-
erazine (2.56 g, 0.01 mol) and triethylamine (1.01 g,
0.01 mol) was refluxed in DMF (20 ml) for 7 h. After the
completion of reaction, the solvent was removed and the
waxy residue was treated with isopropanol and the reaction
content was allowed to stand at 20°C overnight. The crude
crystals obtained were filtered, washed with water to sep-
arate triethylamine hydrochloride, dried, twofold recrys-
tallized from isopropanol and once from ethanol. Yield
74% (3.5 g). M.p. 225–226°C. IR: 3074 (CH aromatic),
1706 (CO xanthine), 1669 (CO xanthine), 1645 with
shoulders at 1606 and 1635 (C=N, C=C aromatic), 1555
and 1560 (C=C xanthine, C=C aromatic), 755 and 742 (CH
Molecular descriptors
The structures were built in HyperChem 7.5 (HyperCube
Ltd.) with extended polymethylene bridge and piperazine
ring in a chair conformation. Their geometries were
optimized by MM ? force field (Hocquet and Langgard,
1998). The electron density distribution was calculated by
semiempirical method AM1 (Dewar et al., 1985). The
charges at all common heteroatoms, HOMO and LUMO
energies form the set of electronic descriptors. Molecular
properties, like molecular weight, number of rings, num-
ber of H-bond donors and acceptors, etc. and 3D
descriptors, like volume, surface, ovality, dipole, etc. were
calculated by MDL QSAR software, version 2.2 (Symyx).
LogP values were calculated by ACD Labs software
(ACD Inc.). The initial set of descriptors included around
50 ones.
1
aromatic); H NMR: 2.05–1.85 t (2H, –CH2–, J = 7 Hz,
side chain), 2.81–2.21 m (10H, 4 9 –CH2–, piperazine,
–CH2, N1 piperazine), 3.95–3.90 t (2H, –CH2–, J = 7 Hz,
side chain), 3.88, 3.32 (s, 2 9 3H, CH3), 4.49 s (1H, –CH
side chain), 7.43–7.18 (m, 10H, aromatic ring), 8.05 s (1H,
–C8–H); 13C NMR: 154.6 (C6xanth CO); 151.02 (C2xanth
CO); 143.6 (C8xanth); 148.4, 128.7, 127.6, 127.2 (12 9 C
aromatic ring); 141.6 (C5xanth); 106.8 (C4xanth); 51.2
(2 9 N4piperazine, CH2), 53.2 (2 9 N1piperazine CH2);
73.4 (CH–(Ph)2 side chain), 48.1 (side chain CH2,
N4piperazine), 48.1 (CH2, N1piperazine), 33.2 (N7xanth
CH3), 37.7 (N1xant CH2), 29.5 (side chain CH2), 22.6
(N3xanth CH3); MS m/z: 167 (100), 473 (M?1). Anal.
C27H32N6O2 (C, H, N).
Variable selection
A genetic algorithm (GA) (Leardi et al., 1992) and step-
wise regression, as implemented in the MDL QSAR
package, were used as variable selection procedures. GA
allows one to select a subset of the most significant pre-
dictors using two evolutionary operations: random muta-
tion and genetic recombination (crossover). The parameters
of the GA used in this study were initial population with
size of 32, uniform crossover and one-point mutation.
Friedman’s lack-of-fit scoring function was used as a fit-
ness function. The GA regression equations were generated
on the basis of the selected variables by ordinary multiple
linear regression (MLR). The stepwise regression was used
in a forward mode with default value for F-to-enter (4.00)
and F-to-remove (3.99). The final descriptor set was
checked for intercorrelation.
5-HT1A and 5-HT2A receptor binding assay
Radioligand studies with native 5-HT1A and 5-HT2A recep-
tors were conducted according to the methods previously
described (Bojarski et al., 1993). Briefly, the following were
used: for 5-HT1A assays, rat hippocampal membranes, [3H]-
8-OH-DPAT (170 Ci/mmol, NEN Chemicals), and 5-HT
(10 lM) for non-specific binding; [3H]-ketanserin (88.0 Ci/
mmol, NEN Chemicals) and methysergide (1 lM) for non-
specific binding. Ki values were determined from at least
three competition-binding experiments, in which 7–9 drug
concentrations, run in triplicate, were used. The Cheng and
Prusoff equation (Cheng and Prusoff, 1973) was used for Ki
calculations.
QSAR study
Model assessment
Data set
Final models were assessed by explained variance r2,
standard error of estimate SEE, F-ratio, leave-one-out
cross-validated (LOO-CV) correlation coefficient q2LOO and
correlation coefficient qC2 V5 after cross validation in groups
The three novel compounds were added to a set of 31
1-arylpiperazines with xanthine moiety at N4, tested
123

480
Med Chem Res (2012) 21:477–486
Table 1 Structures and their affinities to 5-HT1A and 5-HT2A receptors, used in the QSAR study
R2
R1
(CH2)n
N
N
No
R1
n
R2
pKi (5-HT1A
)
pKi (5-HT2A
)
Reference
This study
O
H3
C
5
0
4.965
4.647
CH3
N7
N1
N9
N3
O
CH3
O
H3
C
6
7
3
3
5.804
5.702
6.512
7.232
This study
This study
CH3
O
N7
N1
NH
N9
N3
CH3
O
CH3
N7
N1
N9
O
N3
CH3
O
N
8
3
3
4
7.796
7.509
8.678
6.375
6.588
6.614
Chojnacka-Wojcik et al., (1995)
Chojnacka-Wojcik et al., (1995)
Chojnacka-Wojcik et al., (1995)
CH3
N
N
N
N
O
CH3
O
N
9
CH3
N
N
N
N
O
CH3
O
10
CH3
N
N
N
N
N
O
CH3
O
11
12
3
3
7.602
7.444
5.330
5.089
Chojnacka-Wojcik et al., (1995)
Chojnacka-Wojcik et al., (1995)
O
CH3
N
N
N
N
N
O
CH3
O
N
N
N
CH3
N
N
N
N
O
CH3
O
N
N
13
14
3
3
7.377
6.324
4.553
4.411
Chojnacka-Wojcik et al., (1995)
Chojnacka-Wojcik et al., (1995)
O
CH3
N
N
N
N
N
C
O
H3
O
N
N
CH3
N
N
O
N
N
N
O
CH3
O
N
15
16
3
3
7.590
8.553
6.900
6.313
Pawlowski et al., (1999)
Pawlowski et al., (1999)
CH3
N
N
N
N
O
Cl
CH3
O
CH3
N
N
N
O
N
N
O
H3C
CH3
123

Med Chem Res (2012) 21:477–486
481
Table 1 continued
No
R1
n
R2
pKi (5-HT1A
)
pKi (5-HT2A
)
Reference
O
N
17
3
7.573
6.481
Pawlowski et al., (1999)
Pawlowski et al., (1999)
Pawlowski et al., (1999)
Pawlowski et al., (1999)
Pawlowski et al., (1999)
Pawlowski et al., (1999)
Pawlowski et al., (1999)
Pawlowski et al., (1999)
Pawlowski et al., (2000)
Pawlowski et al., (2000)
CH3
N
N
N
Cl
N
O
CH3
O
18
19
20
21
22
23
24
25
26
3
3
3
3
3
3
3
3
3
7.764
7.368
7.666
7.484
7.983
7.220
6.775
6.415
7.000
6.250
6.740
5.375
5.967
5.411
6.445
5.300
6.706
6.212
CH3
N
O
O
O
O
N
N
H3
C
C
C
C
N
N
O
CH3
O
O
O
CH3
N
N
N
N
Cl
Cl
Cl
N
C
O
H3
O
CH3
N
N
N
N
H3
H3
H3
N
C
O
H3
O
O
O
CH3
N
N
N
N
N
O
CH3
O
CH3
N
N
N
N
N
O
CH3
O
CH3
N
O
N
N
N
N
O
CH3
O
CH3
N
N
O
N
N
C
N
O
CH3
O
N
H3
NH
CH3
N
N
N
O
CH3
CH3
O
CH3
N
N
NH
N
N
O
CH3
CH3
O
27
28
3
3
6.228
6.340
6.747
5.996
Pawlowski et al., (2000)
Pawlowski et al., (2000)
O
O
N
CH3
N
N
N
NH
O
CH3
CH3
O
O
O
CH3
N
N
N
NH
N
O
CH3
CH3
O
29
3
6.658
6.348
Chlon et al., (2001)
O
O
CH3
N
N
N
NH
N
O
CH3
123

482
Med Chem Res (2012) 21:477–486
Table 1 continued
No
R1
n
R2
pKi (5-HT1A
)
pKi (5-HT2A
)
Reference
30
3
5.997
6.939
Chlon et al., (2001)
CH3
O
N
CH3
N
N
N
NH
O
CH3
31
32
33
34
35
36
3
3
3
3
3
4
7.301
7.319
8.097
6.678
8.000
7.668
6.352
6.260
6.517
6.924
6.461
5.701
Chlon et al., (2001)
Chlon et al., (2001)
Chlon et al., (2001)
Chlon et al., (2001)
Chlon et al., (2001)
Kolaczkowski et al., (2005)
O
N
CH3
N
N
N
NH
O
CH3
O
N
O
CH3
H3
C
N
N
N
NH
O
CH3
O
H3C
O
N
CH3
N
N
N
NH
O
CH3
Cl
O
CH3
N
N
N
NH
N
O
CH3
Cl
O
N
CH3
N
N
N
NH
O
CH3
O
O
O
O
CH3
N
N
N
N
N
O
CH3
O
37
38
4
4
9.301
8.056
5.971
6.815
Kolaczkowski et al., (2005)
Kolaczkowski et al., (2005)
CH3
N
N
N
N
O
N
O
H3C
CH3
O
CH3
N
N
N
Cl
N
N
O
CH3
groups. To check the validity of the selected descriptor set,
100 randomizations of the dependent variable values
among the compounds were carried out (y scrambling).
Multiple r2 were computed for each of corresponding
regressions and the mean value is given as r2scr. Addition-
ally, the newly synthesized compounds were separated into
an external test set and their binding affinities were pre-
dicted by the derived models.
123

Med Chem Res (2012) 21:477–486
Results and discussion
Chemistry
483
Pharmacology
The affinity constants of the three novel 1-arylpiperazines
are given in Table 2. Compound 5 is very weak binder to
both 5-HT1A and 5-HT2A receptors (10.8 and 22.5 lM,
respectively). Compound 6 is weak 5-HT1A binder
(1.6 lM) and moderate 5-HT2A binder (307.5 nM) Thus, it
The synthetic route for the preparation of the piperazine
derivatives 5–7 are summarized in Scheme 1.
The readily available materials 8-bromocaffeine (1);
8-(3-bromopropylamino)-1,3,7-trimethyl-1H-purine-2,6
(3H,7H)-dione (2) and 1-(3-iodoropropyl)-3,7-dimethyl-
xanthine (3) were aminated with benzhydrylpiperazine in
the presence of triethylamine to give the target compounds
5–7 in high yield.
is 5 times more selective on 5-HT2A than on 5-HT1A
.
Compound 7 shows weak 5-HT1A affinity (2 lM) and high
5-HT2A affinity (58.6 nM). It is 34 times more selective on
5-HT2A than on 5-HT1A receptors.
All the synthesized compounds were freely soluble in
chloroform, dichloromethane, DMF and DMSO but insol-
uble in non-polar solvents like n-hexane. The structures of
the synthesized compounds were confirmed by IR, 1H
NMR, 13C NMR, FABMS spectra and elemental analysis.
The results were consistent with the assigned structures.
The 1H NMR spectra of the compounds 5–7 showed the
signals of the respective protons of the synthesized com-
pounds, which were verified on the basis of their chemicals
shifts, multiplicities and coupling constants.
QSAR study
5-HT1A affinity model
Both variable selection procedures, GA and stepwise regres-
sion, led toanidenticalmodel for affinityto5-HT1A receptors:
pKi ¼ ꢁ6:202ðꢂ2:384ÞqN1pip þ 0:470ðꢂ0:088Þncirc
þ 5:659ðꢂ1:629Þovality þ 2:107ðꢂ0:491ÞHOMO
þ 222:1ðꢂ66:83ÞqN1xant þ 80:597
The values of the chemical shifts of the protons and
carbon atoms registered by 1H NMR and 13C-NMR spectra
were compared with simulated values. A very good corre-
lation between the registered and computed values was
observed with 13C-NMR spectra. We observed only small
deviations of computed from experimental values. The
deviations of registered in comparison with calculated val-
ues at 1H NMR spectra are larger, due to an impossibility to
render an account of influence of the solvent. Regardless, the
Table 2 5-HT1A/5-HT2A affinities of the novel 1-benzhydryl-piper-
azine derivatives
Compound
Ki SEM (nM)
5-HT2A selectivity
5-HT1A/5-HT2A
5-HT1A
5-HT2A
5
6
7
10830 710
1569 90
22540 1860
307.5 20
0.5
5.1
1
simulated H NMR spectra are in good correlation with
1988 116
58.6
4
33.9
experimental ones.
O
O
N
CH3
CH3
N
H3C
N
N
N
H3C
N
Br
N
N CH
N
O
N
O
CH3
1
CH3
5
O
O
CH3
CH3
N
N
N
DMF / (C2H5)3N
- (C2H5)3N.HX
H3C
N
H3C
N
NH (CH2)3
N
N CH
HN
N CH
NH (CH2)3 Br
+
N
O
N
O
N
6
2
CH3
CH3
4
O
CH3
O
N
CH3
N
I
(H2C)3
N
N
N
CH
N
N
(H2C)3
N
N
N
O
O
CH3
7
CH3
3
Scheme 1
123

484
Med Chem Res (2012) 21:477–486
n = 34, r2 = 0.806, SEE = 0.437, F = 23.3, q2LOO
0.713, q2CV5 = 0.681, rs2cr = 0.146
=
show any significant difference in the charge of N1 in
comparison with the 7- and 8-substituted xanthines.
However, the removing of the descriptor qN1xant from the
model decreases both r2 and qL2OO by 0.1. This subtle
influence of the partial charge of xanthine N1 atom on
5-HT1A receptor affinity will be explored in our further
synthesis.
No intercorrelation exists between the descriptors inclu-
ded in the model. The descriptor qN1pip shows the partial
atomic charge of piperazine N1 atom. Its values range from
-0.266 (6) to -0.382 (17). The negative regression coeffi-
cient means that as more negative is the piperazine N1 atom
as more affine is the compound. The charge of N1 depends on
the aryl substituent. The inductive effect of the N1 substit-
uents decrease in the order: phenyl [ pyrimidinyl [ benz-
hydryl. In the same order increases the 5-HT1A binding
affinity of the investigated piperazines.
The correlation plot between predicted and experi-
mental pKi values for 5-HT1A affinity is given in Fig. 1.
The highest residual is 0.848 for compound 13. Removing
it led to a slightly improvement of r2 and q2 (data not
shown).
The descriptor ncirc accounts for the number of graph
circuits, the total number of all cycles in the molecular
graph. For example, ncirc equals 2 for biphenyl, but 3 for
naphthalene, that is two cycles of 6 edges and 1 cycle of 10
edges. The compounds in this study are divided into three
groups according to the value of ncirc. Compounds 25–28
have 5 circuits in their structures. Compounds 5–7 and
29–35 have 6 circuits, while compounds 8–24 and 36–38
have 8 circuits. The positive regression coefficient of ncirc
indicates that the more circuits in the structure of the ligand
the higher is the affinity.
The predictive ability of the model was tested on the
newly synthesized compounds. They were excluded from
the training set, the model was re-built and used to predict
the pKi (5-HT1A) values of the three new compounds
(Table 3). Compounds 6 and 7 were predicted well with
residuals below 0.5 log unit, while compound 5 was
overestimated by 0.7 log unit. The r2pred value is 0.834.
5-HT2A affinity model
The initial model for 5-HT2A affinity derived by GA and
stepwise regression had low r2 and q2. The removing of
two outliers with residuals[0.9 (compounds 25 and 35) led
to the following model:
Ovality describes the overall size and shape of the
molecules (Bodor et al., 1989). It is defined as the ratio
between the area of the surface of the molecule and that of
the minimum surface corresponding to the volume of the
molecule. For a sphere, ovality equals 1. As higher is the
ovality value as far from a sphere is the shape of a mole-
cule. Ovality of the investigated structures ranges from
1.76 for compound 5 to 1.99 for compound 35. The posi-
tive correlation between ovality and affinity suggests that
bulky substituents at piperazine N4 are favour for the
5-HT1A affinity. Elongation of the spacer also increases
ovality and should increase affinity.
pKi ¼ 0:464ðꢂ0:073ÞlogP þ 175:2ðꢂ49:72:ÞMaxNeg
þ 14:72ðꢂ4:59ÞqN4pip þ 81:576
n = 32, r2 = 0.652, SEE = 0.458, F = 17.47, q2LOO
0.569, q2CV5 = 0.582, rs2cr = 0.099
=
No intercorrelation exists between the descriptors
included in the model. The lipophilicity of compounds,
expressed by the descriptor logP, correlates positively with
the affinity, e.g. more lipophilic ligands bind more tightly.
This result is in a good agreement with different 5-HT2A
The highest occupied molecular orbital (HOMO) ener-
gies of the investigated compounds correlates positively
with their 5-HT1A affinities, e.g. high HOMO energies
correspond to high affinities. The HOMO energy reflects
the compounds’ ability to donate electrons in a charge-
transfer type of interaction. The charge-transfer mechanism
of action of hallucinogenic drugs, like LSD, is well known
(Kang and Green, 1970; Kolb, 1987). Also there is
empirical evidence that the hallucinogenic effects of LSD
are mediated by a direct agonist effect at 5-HT2A receptors
(Ouagazzal et al., 2001). Our result shows that a charge
transfer is a possible mechanism also in the interaction
between ligand and 5-HT1A receptor.
The participation of the partial charge of xanthine N1
atom, expressed by the descriptor qN1xant, in the 5-HT1A
model was surprising, because the substitutions in the set
of investigated compounds are at positions far from N1.
Even the 1-substituted xanthine derivative 7 does not
Fig. 1 Plot of predicted versus experimental pKi values for 5-HT-1A
affinity after LOO-CV. The new 1-benzhydryl-piperazine derivatives
are given in pink (gray) diamonds (Color figure online)
123

Med Chem Res (2012) 21:477–486
485
Table 3 Predicted and experimental pKi (5-HT1A) and pKi (5-HT2A) values of the novel 1-benzhydryl-piperazine derivatives
Compound
pKi (5-HT1A
)
pKi (5-HT2A)
Predicted
Experimental
Residual
Predicted
Experimental
Residual
5
6
7
5.698
6.268
5.979
4.965
5.804
5.702
-0.733
-0.464
-0.277
4.842
6.009
7.383
4.647
6.512
7.232
-0.195
0.503
-0.151
pharmacophore models, reviewed extensively by Bojarski
(2006), which include one or two hydrophobic areas,
necessary for receptor binding of agonists and antagonists.
Descriptor MaxNeg reflects the largest negative charge
over the atoms in a molecule. In the studied molecules the
largest negative charge is associated with xanthine O6
atom. The positive regression coefficient for MaxNeg
means that less negative O6 atoms are more favorable for
5-HT2A affinity. The partial charge of O6 depends on the
substituent at N1 in the xanthine moiety. Compound 7 has
the lowest negative charge of O6 due to the stronger
positive inductive effect of the three-methylene bridge than
that of the methyl group.
Fig. 2 Plot of predicted versus experimental pKi values for 5-HT-2A
affinity after LOO-CV. Outliers are given in blank diamonds.
New 1-benzhydryl-piperazine derivatives are given in pink (gray)
diamonds (Color figure online)
The descriptor qN4pip shows the partial atomic charge of
piperazine N4 atom. The xanthine moiety is bound to N4
directly in 5 or through a three- or four-methylene spacer in
the rest of ligands. The positive correlation between this
charge and the affinity means that less negative N4 atoms
are more favorable for 5-HT2A affinity. The charge of N4
depends on the length of methylene bridge, as longer is the
chain as lower is the charge.
The correlation plot between predicted and experimental
pKi values for 5-HT2A affinity is given in Fig. 2. Com-
pounds 25 and 35 are outliers with residuals above |0.9|.
Compound 25 has the highest positive residual, while
compound 35 has the highest negative one. Both structures
do not differ significantly from the rest of molecules in the
set. Compound 35 is the most lipophilic (logP = 5.82) not
being the most affine (pKi on 5-HT2A = 6.461) as it is
expected according to the 5-HT2A model. Regarding
compound 25 we do not have any reasonable explanation
why it acts as an outlier.
The predictive ability of the model was tested on the
newly synthesized compounds. They were excluded from
the training set, the model was re-built and used to predict
the pKi (5-HT2A) values of the three new compounds
(Table 3). Compounds were predicted very well with
residuals up to 0.5 log unit and r2pred = 0.914.
Fig. 3 5-HT2A pharmacophore model according to Mokrosz et al.
(1994), shown on compounds 6 and 7
The structures of the new selective 5-HT2A ligands are
in a good agreement with the pharmacophore models,
available in the literature (Bojarski, 2006). According to
Glennon’s model of a 5-HT2A binding site (Glennon et al.,
1991) the pharmacophore consists of region of bulk toler-
ance at a distance from a basic nitrogen atom, area of
hydrogen bond acceptor (HBA), electron acceptor site and
hydrophobic area (Fig. 3). Most of these elements, like
region of bulk tolerance, basic nitrogen, HBA and
123

486
Med Chem Res (2012) 21:477–486
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study reveals several structural features responsible for the
affinity and selectivity of 5-HT ligands with benzhydryl
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