Bivalent molecular probes for dopamine D2-like receptors

Article abstract of DOI:10.1016/j.bmc.2011.10.063

Merging two arylamidoalkyl substituted phenylpiperazines as prototypical recognition elements for dopamine D₂-like receptors by oligoethylene glycol linkers led to a series of bivalent ligands. These dimers were investigated in comparison to th

Full text of DOI:10.1016/j.bmc.2011.10.063

Bioorganic & Medicinal Chemistry 20 (2012) 455–466
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Bivalent molecular probes for dopamine D2-like receptors
⇑
Daniela Huber, Stefan Löber, Harald Hübner, Peter Gmeiner
Department of Chemistry and Pharmacy, Emil Fischer Center, Friedrich Alexander University, Schuhstrasse 19, D-91052 Erlangen, Germany
a r t i c l e i n f o
a b s t r a c t
Article history:
Merging two arylamidoalkyl substituted phenylpiperazines as prototypical recognition elements for
Received 17 August 2011
Revised 14 October 2011
Accepted 19 October 2011
Available online 25 October 2011
dopamine D2-like receptors by oligoethylene glycol linkers led to a series of bivalent ligands. These
dimers were investigated in comparison to their monomeric analogues for their dopamine D2long, D2short
,
D3 and D4 receptor binding. Radioligand binding experiments revealed strong bivalent effects for some
para-substituted benzamide derivatives. For the D3 subtype, the target compounds 32, 34 and 36 showed
an up to 70-fold increase of affinity and a substantial enhancement of subtype selectivity when compared
to the monovalent analogue 24. Analysis of the binding curves displayed Hill slopes very close to one
indicating that the bivalent ligands displace 1 equiv of radioligand. Obviously, the two pharmacophores
occupy an orthosteric and an allosteric binding site rather than adopting a receptor-bridging binding
mode.
Keywords:
GPCR
Subtype selectivity
Dopamine
Bivalent ligand
D3 receptor
Ó 2011 Elsevier Ltd. All rights reserved.
1. Introduction
gain insights into the role of GPCR oligomerization on biosynthesis,
receptor function and internalization. On the other hand, bivalent
The dopamine D2 receptor family consists of three particular
subtypes named D2, D3 and D4.1 These aminergic GPCRs are associ-
ated with many central nervous system based diseases including
schizophrenia, Parkinson’s disease, drug addiction, and erectile
dysfunction.2 Numerous CNS-active drugs act through a stimula-
tion or blockade of the dopamine receptor controlled signal trans-
duction and a huge amount of investigations on structure–activity
relationships, rationally based tuning of subtype selectivity and
intrinsic activity as well as the propensity of the target proteins
to form homo- and heteromeric complexes has been accumu-
lated.3–16
Inspired by nature, hybrid molecules and bivalent ligands,
which incorporate two pharmacophores connected by an appropri-
ate linker, are currently evolving as a promising strategy in drug
discovery.17,18 Recently, design, synthesis and biological investiga-
tions of bivalent GPCR ligands have been described to target sero-
tonin,19 muscarinic,20,21 opioid22,23 and dopamine receptors.24 At
least two binding modes have been considered. Thus, the ligand
can simultaneously interact with two primary (orthosteric) bind-
ing sites of two neighboring protomers. Alternatively, the bivalent
ligand may address a primary binding site and a secondary (allo-
steric) binding site located in close proximity of an adjacent GPCR
or at the identical protomer leading to a bitopic or dualsteric bind-
ing mode.25 Bivalent ligands binding neighboring binding sites of
two physically interacting GPCRs may serve as pharmacological
tools to study the quaternary structure of receptor dimers and to
ligands with a dualsteric binding mode facilitate the generation
of subtype-selective agonists or antagonists because allosteric re-
gions are frequently less conserved than the orthosteric binding
pocket, which is usually very similar for all subtypes of a receptor
family.
Very recently, we reported on the synthesis and biological
investigations of bivalent ligands for the dopamine D2 receptor
N
N
O
N
O
N
N
O
N
N
O
N
N
N
O
linker
O
( )n
( )n
1
N
N
O
N
N
( )1,2
NH
O
( )1,2
HN
O
O
Ar
Ar
oligo ethylene glycol
⇑
Corresponding author.
E-mail address: peter.gmeiner@medchem.uni-erlangen.de (P. Gmeiner).
Chart 1. Bivalent dopamine D2 receptor ligand 1 and novel arylcarboxamide based
dimers.
0968-0896/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2011.10.063

456
D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
subtypes.26 The test compounds incorporating the privileged
structure of 1,4-disubstituted aromatic piperidines/piperazines
(1,4-DAPs) of type 1 (Chart 1) differ in length and structure of the
spacer unit that links two identical pharmacophores. Radioligand
binding assays revealed that the bivalent ligands exhibit a distinct
binding profile compared with monovalent analogues. Some of the
bivalent ligands revealed a steepening of the competition curve for
the D2 receptors with Hill slopes close to two indicating the libera-
tion of 2 equiv of radioligand. This behavior depended on the struc-
ture and the length of the linker arm. Whereas an octylene bridge
between the triazole units led to a bivalent binding mode occupying
two primary binding sites, Hill slopes close to one for oligoethylene
glycol linked analogues indicated the displacement of 1 equiv of
radioligand and thus, binding of a primary (orthosteric) and a second-
ary (allosteric) binding site. Because the oligoethylene glycol moiety
seemed to be associated with high dopamine receptor binding affin-
ity, we intended to explore oligoethylene glycol linked 1,4-DAPs
beyond the phenylpiperazinylmethyl based pharmacophore.
As an extension of this study, we herein present the synthesis of
a collection of bivalent amidopropyl- and amidobutylphenylpiper-
azine derivatives. The pharmacophore of this family of compounds
was described to show high affinity to the receptors of the dopa-
mine D2 family when the amidobutylphenylpiperazines revealed
substantial preference for the D3 subtype.2,27,28 We developed a fo-
cused library of bivalent target compounds and monovalent control
agents incorporating a 2-methoxyphenylpiperazine head group,
structurally different lipophilic appendages and oligoethylene
glycol linkers of varying length. Employing radioligand binding
studies including careful analysis of the competition curves, we
envisioned to learn if the target products were able to adopt a biva-
lent binding mode addressing two adjacent binding sites of a dopa-
mine receptor dimer or a dualsteric binding mode with subtype
selective binding properties.
HO
COOMe
a
O
COOMe
Ar
O
b
Ar
2: Ar = Ar1
3: Ar = Ar2
4a: Ar = Ar1; R = Me
5a: Ar = Ar2; R = Me
4b: Ar = Ar1; R = H
5b: Ar = Ar2; R = H
ROOC
O
O
COOR
a
(
)
3
O
Ar
Ar
6a: Ar = Ar1; R = Me
7a: Ar = Ar2; R = Me
b
6b: Ar = Ar1; R = H
7b: Ar = Ar2; R = H
COOMe
HO
O
COOR
Ar
c
O
Ar
8: Ar = Ar3
9: Ar = Ar2
10a: Ar = Ar1; R = Me
11a: Ar = Ar2; R = Me
b
10b: Ar = Ar1; R = H
11b: Ar = Ar2; R = H
d
COOMe
e
Br
Ar
O
ROOC
COOR
(
)
n
O
O
Ar
Ar
12: Ar = Ar3
13: Ar = Ar2
14a: Ar = Ar3; m = 3; R = Me
15a: Ar = Ar3; m = 4; R = Me
16a: Ar = Ar3; m = 5; R = Me
17a: Ar = Ar3; m = 7; R = Me
18a: Ar = Ar2; m = 3; R = Me
Ar2
Ar1
N
b
N
14b: Ar = Ar3; m = 3; R = H
15b: Ar = Ar3; m = 4; R = H
16b: Ar = Ar3; m = 5; R = H
17b: Ar = Ar3; m = 7; R = H
18b: Ar = Ar2; m = 3; R = H
Ar3
Scheme 1. Reagents and conditions: (a) BrCH2CH2OEt (for 4a, 5a) or tetra(ethylene
glycol)di-p-tosylate (for 6a, 7a), K2CO3, DMF, 70 °C, 16 h. (b) NaOH, H2O, MeOH,
reflux, 6 h. (c) BrCH2CH2OEt, KHMDS, toluene, DMF, 70 °C, 2 h. (d) for 9: PBr3,
CH2Cl2, rt, 16 h. (e) tetra(ethylene glycol), KHMDS, toluene, THF, 0 °C to rt, 2 h.
available from 931 by bromination with PBr3. The reaction se-
quence was conducted as described for the above mentioned ben-
zoic acid derivatives leading to the pyrazolopyridine-3-carboxylic
acid 5b,11b and the dimer 7b,18b.
2. Results and discussion
2.1. Synthesis
Finally, the newly prepared mono- and bis-carboxylic acids were
converted into the carboxamides 19–40 by TBTU-mediated cou-
pling with N-aminopropyl-N0-(2-methoxyphenyl)-piperazine and
N-aminobutyl-N0-(2-methoxyphenyl)-piperazine33 (Scheme 2).
Intending to evaluate the importance of the chosen attachment
position of the linker unit, we constructed an analogous derivative
featuring an alternative mode of linking when the para-position of
the phenylpiperazine moiety served as a point of attachment as well
as a monovalent control ligand to allow the investigation of a puta-
tive bivalency effect. The synthesis started from 2-bromo-5-hydrox-
ymethylanisole (41)34, which was O-alkylated with 2-ethoxyethyl
bromide to give the benzyl ether 42 (Scheme 3). Palladium assisted
amination with piperazine followed by N-alkylation of the resulting
intermediate 43 applying 4-bromobutyronitrile led to the N,N0-
disubstituted piperazine derivative 45. Finally, reduction of the cya-
no group gave the primary amine 44, which was benzoylated
yielding the desired carboxamide 46. The synthesis of the bivalent
ligand 51 started with cross-linking of the benzyl alcohol 41 with
bis-tosylated tetraethylene glycol. Subsequent Buchwald–Hartwig
reaction of the bis-benzylether 47 afforded the di-piperazine deriv-
ative 48. Finally, dialkylation with the N-tosyloxybutyl benzamide
50, which was derived from the respective butanol derivative 49,35
gave the final product 51.
Our first series of target compounds was based on a benzoic acid
amide moiety when the attachment point of the linker should be
varied. As a spacer element, oligoethylene glycol units of different
length were chosen. The mono- and bis-benzoic acid derivatives
that should serve as precursors for the respective carboxamide
based ligands were synthesized as outlined in Scheme 1.
Starting from methyl-3-hydroxybenzoate (2), reaction with
bromoethyl ethyl ether or tetraethylene glycol bis-tosylate in the
presence of a base (K2CO3) afforded the phenyl ethers 4a and 6a,
respectively. Subsequent saponification led to the corresponding
carboxylic acids 4b and 6b. An analogous procedure starting from
methyl-4-hydroxymethylbenzoat (8) led to the methyl ester 10a
and the corresponding carboxylic acid 10b. The respective
bis-carboxylates were prepared starting from the benzyl bromide
12. Reaction with tetra-, penta-, hexa- and octaethylene glycol
afforded the diester derivatives 14a, 15a, 16a and 17a, respectively,
which were converted into the corresponding carboxylic acids 14b,
15b, 16b and 17b.
It has been shown that bicyclic heteroaromatic moieties can act
as potent recognition elements for dopamine D2-like receptor sub-
types, when the pyrazolo[1,5-a]pyridine scaffold turned out to be
an excellent functionality to control ligand affinity and selectiv-
ity.29–31 Thus, we envisioned to incorporate a 3,5-disubstituted
7a-azaindole moiety as a heterocyclic bioisostere of the bis-substi-
tuted benzene. The syntheses of the mono- and bis-carboxylic
acids started from methyl-5-hydroxypyrazolo[1,5-a]pyridine-3-
carboxylate (3)32 and the 5-bromomethyl derivative 13 which is
2.2. Receptor-ligand binding experiments
In vitro binding affinities of the molecular probes 27–40 and 51,
along with their monovalent control agents 19–26 and 46 were

D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
457
O
O
Monovalent ligands
Bivalent ligands
OH
O
O
O
a
b
COOH
Br
Br
N
Ar
OMe
OMe
O
HN
OMe
c
L
i
n
k
e
r
R
COOH
41
42
43
Ar
COOH
4b, 5b, 10b, 11b
6b, 7b, 14b, 15b,
16b, 17b, 18b
O
Ar
O
(
)
n
H2N
N
OMe
a
N
N
a
d
N
N
OMe
NC
N
OMe
O
(
)
H2N
3
44
45
O
O
(
)
n
N
H
N
OMe
OMe
Ar
Ar
O
L
i
n
k
e
r
N
O
N
e
R
(
)
n
N
OMe
N
H
N
OMe
Ar
N
H
O
N
46
N
(
)
N
n
H
Ar
19: Ar1
20: Ar1
21: Ar2
22: Ar2
23: Ar3
24: Ar3
25: Ar2
26: Ar2
n
1
2
1
2
1
2
1
2
R
N
-OCH2CH2OCH2CH3
-OCH2CH2OCH2CH3
-OCH2CH2OCH2CH3
-OCH2CH2OCH2CH3
-CH2OCH2CH2OCH2CH3
-CH2OCH2CH2OCH2CH3
-CH2OCH2CH2OCH2CH3
-CH2OCH2CH2OCH2CH3
O
O
O
O
O
Ar
n
1
2
1
2
1
2
1
2
1
2
1
2
1
2
linker
-O(CH2CH2O)4-
-O(CH2CH2O)4-
Br
N
27: Ar1
28: Ar1
29: Ar3
30: Ar3
31: Ar3
32: Ar3
33: Ar3
34: Ar3
35: Ar3
36: Ar3
37: Ar2
38: Ar2
39: Ar2
40: Ar2
f
OMe
b
HN
OMe
41
O
O
-CH2O(CH2CH2O)4CH2-
-CH2O(CH2CH2O)4CH2-
-CH2O(CH2CH2O)5CH2-
-CH2O(CH2CH2O)5CH2-
-CH2O(CH2CH2O)6CH2-
-CH2O(CH2CH2O)6CH2-
-CH2O(CH2CH2O)8CH2-
-CH2O(CH2CH2O)8CH2-
-O(CH2CH2O)4-
O
O
O
Br
N
47
48
OMe
HN
OMe
Ar2
Ar1
Ar3
N
N
O
O
-O(CH2CH2O)4-
-CH2O(CH2CH2O)4CH2-
-CH2O(CH2CH2O)4CH2-
OH
OTos
g
N
N
H
H
h
50
49
Scheme 2. Reagents and conditions: (a) TBTU, DIPEA, CH2Cl2, DMF, 0 °C to rt, 4 h.
O
O
O
O
N
N
OMe
N
O
measured by displacement of the radioligand [3H]spiperone from
human D2long, D2short, D3, and D4.4 receptors stably expressed in Chi-
nese hamster ovary (CHO) cells (Table 1). Furthermore, binding data
for the porcine dopamine D1, serotonin 5-HT1A, 5-HT2 and the adren-
ergic a1 receptor were determined applying striatal and cortical
membranes and the radioligands [3H]SCH23390, [3H]WAY100635,
[3H]ketanserin and [3H]prazosin, respectively (Supplementary
data). The characterization of the test compounds involved receptor
affinity and steepness of the competition curve when the data of the
bivalent ligands was compared with that of the respective monova-
lent control agents.
H
O
O
N
N
OMe
N
H
51
Scheme 3. Reagents and conditions: (a) BrCH2CH2OEt, KHMDS, toluene, THF, 0 °C
to rt, 4 h; (b) piperazine, NaOtBu, Pd2(dba)3, 2-(di-t-butylphosphino)biphenyl,
toluene, 115 °C, 18 h; (c) Br(CH2)3CN, K2CO3, NaI, CH3CN, 80 °C, 24 h; (d) LiAlH4,
Et2O, ꢁ5 °C to rt, 3 h; (e) benzoyl chloride, Et3N, CHCl3, 0 °C to rt, 24 h; (f)
tetra(ethylene glycol)di-p-tosylate, NaH, THF, 0 °C to rt, 4 h; (g) p-TosCl, Et3N, THF,
40 °C, 16 h; (h) K2CO3, NaI, CH3CN, 80 °C, 16 h.
The monovalent amidopropylphenylpiperazines (19 and 23)
displayed moderate binding to the D2-like receptors. Formal dimer-
ization led to the bivalent analogues 27, 29, 31, 33, 35 indicating
significantly higher affinities for D2long, D2short and D3 whereas D4
affinity was more or less unchanged. The most substantial increase
of affinity was observed for the meta-disubstituted benzamide 27
with four oligoethylene glycol units displaying more than 20-fold
gain of affinity for D2long and D2short and a factor of approximately
15 for D3.
For the monovalent amidobutylphenylpiperazines 20 and 24, a
preference for D3 binding could be determined. This is in good
accordance to the unsubstituted N-[4-[4-(2-methoxyphenyl)pip-
erazin-1-yl]butyl]benzamide36 showing Ki values of 130, 90, 7.8
and 170 nM for D2long, D2short, D3 and D4, respectively. Again,
dimerization led to an improvement of D2long, D2short and D3 affin-
ity. For some representatives of this group, the improvement of
receptor recognition by formal dimerization was very strong. Thus,
the para-disubstituted test compounds 32, 34 and 36 incorporating
five, six and eight oligoethylene glycol units, respectively, revealed
an excellent gain of affinity for D3 reaching a factor of approxi-
mately 70-fold (for 32), 35-fold (for 34) and 55-fold (for 36),
respectively. In contrast, only a 2- to 15-fold increase of affinity
could be determined for D2long, D2short, D4 and also for the related
GPCRs D1 (ꢀfivefold), 5-HT1A (ꢀ0.5-fold), 5-HT2 (ꢀfivefold) and
a1 (2–4-fold), respectively, (Supplementary data). Moreover, these
dimers exhibited substantial D3 selectivity (>50-fold) over the
dopamine receptor subtypes D1, D2long, D2short, D4 and the serotonin
receptors 5-HT1A and 5-HT2. The behavior of the pyrazolo[1,5-a]
pyridine based ligands 21, 22, 25, 26 and 37–40 deviated from
the benzamide derivatives. Single-digit nanomolar or even subn-
anomolar Ki values were determined at the D2long and D2short recep-
tors for both mono- and bivalent azaindoles. Compared to the
benzamide analogues, the dimer-specific increase of affinity was
less pronounced. Interestingly, the 1,5-disubstituted bis-pyrazolo
[1,5-a]pyridine 38 displayed excellent binding affinity with Ki val-
ues of 1.8, 0.59 and 0.34 nM for D2long, D2short and D3 and strong
selectivity over D4 (Ki = 77 nM).
As expected, linkage of the pharmacophores via the phenylpip-
erazine head group resulted in dopamine receptor ligands with
poor binding affinity. According to receptor–ligand models that
are based on the recent D3 crystal structure,37 the arene system
of phenylpiperazines directs to the transmembrane helix 5 in the
lower part of the binding pocket. Large substituents or spacer
elements are, thus, expected to induce repulsive interactions lead-
ing to a decrease of binding energy.

458
D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
Table 1
Receptor binding data of compounds 19–40, 46 and 51 at the human dopamine D2long, D2short, D3 and D4.4 receptors
Compd
Valency/linkerc
Ar
Ki valuesa [nM] (Hill slopesb)
D2long
D2short
D3
D4.4
19
20
27
28
23
24
29
30
31
32
33
34
35
36
21
22
25
26
37
38
39
40
46
51
Mono
Mono
bi/4
Ar1
Ar1
Ar1
Ar1
Ar3
Ar3
Ar3
Ar3
Ar3
Ar3
Ar3
Ar3
Ar3
Ar3
Ar2
Ar2
Ar2
Ar2
Ar2
Ar2
Ar2
Ar2
270 (1.0)
95 (0.9)
16 (0.9)
6.6 (0.9)
780 (0.9)
290 (0.9)
42 (0.9)
25 (0.9)
47 (0.8)
20 (0.7)
51 (1.2)
40 (0.9)
150 (0.9)
38 (0.8)
29 (0.9)
18 (0.9)
67 (0.9)
25 (0.9)
8.4 (1.0)
1.8 (1.0)
4.7 (0.9)
8.3 (0.9)
19000 (0.9)
270 (1.1)
140 (1.1)
44 (0.9)
6.4 (1.1)
1.8 (0.8)
320 (1.0)
83 (0.9)
19 (0.9)
11 (0.9)
32 (0.8)
14 (0.8)
20 (1.0)
25 (0.9)
66 (1.0)
16 (1.0)
6.4 (1.0)
3.0 (0.9)
32 (0.9)
12 (0.9)
2.7 (1.1)
0.59 (0.8)
3.9 (0.8)
4.6 (0.8)
9700 (0.9)
280 (1.2)
170 (1.1)
4.9 (1.0)
11 (1.2)
0.61 (1.1)
330 (1.0)
16 (0.9)
140 (0.9)
220 (0.9)
38 (1.0)
bi/4
48 (0.8)
Mono
Mono
bi/4
bi/4
bi/5
bi/5
bi/6
bi/6
bi/8
240 (1.0)
250 (1.0)
82 (0.9)
68 (0.8)
190 (0.9)
78 (0.9)
160 (0.9)
130 (1.0)
360 (1.0)
140 (0.9)
70 (0.9)
64 (0.9)
45 (0.9)
160 (0.8)
51 (1.1)
77 (1.0)
140 (0.9)
100 (0.9)
2400 (0.9)
5400 (1.0)
83 (0.9)
0.88 (0.8)
100 (0.9)
0.23 (0.7)
11 (0.7)
0.44 (0.8)
350 (1.1)
0.30 (0.7)
78 (1.0)
0.66 (0.9)
160 (1.1)
1.2 (1.1)
3.5 (1.1)
0.34 (0.9)
10 (1.0)
0.50 (0.9)
930 (1.0)
110 (1.1)
bi/8
Mono
Mono
Mono
Mono
bi/4
bi/4
bi/4
bi/4
Mono
bi/4
a
Ki values in nM are based on the means of 2–9 experiments each done in triplicate with SD or SEM values <35%
Hill slopes are displayed as absolute values nH which are derived from the same binding curves recorded for the determination of Ki values; the original nH was negative
b
but is displayed as absolute value.
c
Number of ethylene glycol units.
Radioligand binding experiments normally lead to competitive
binding curves that follow the law of mass action and, thus, show
a Hill slope of one.38 For particular bivalent ligands of type 1, a
steepening of the competition curves was observed, compared to
the respective monomer, resulting in Hill slopes that reach the va-
lue of 2.0 for interactions with D2long and D2short. This is indicative of
a positive cooperative binding.9,39 Bivalent ligands addressing two
adjacent binding sites of receptor dimers will induce such coopera-
tivity because binding of the second pharamcophore is significantly
accelerated due to the vicinity of the ligand and the, thus, facilitated
enrichment of local concentration. In this case, bivalent binding
leads to the liberation of two equivalents of radioligand and a sub-
stantial steepening of the competition curve. Careful investigation
of the binding curves of our synthesized monomeric and dimeric
target compounds revealed Hill slopes not significantly greater than
1 (absolute value) in most of the cases. Following our above men-
tioned reasoning, the arylcarboxamide linked ligands do not feature
this bivalent binding mode with both recognition elements occupy-
ing the two binding pockets of the receptor dimer. SAR studies on
our bivalent linkers indicate that the behavior depended on the
structure and the length of the linker arm. Whereas an octylene
bridge between two triazole units led to a bivalent binding mode
occupying two primary binding sites, oligoethylene glycol linked
analogues indicated the displacement of 1 equiv of radioligand.
Obviously, the herein described bivalent ligands address a primary
binding site and a secondary (allosteric) binding site located in
close proximity of an adjacent GPCR or at the identical protomer.
incorporating 5–8 oligoethylene glycol units attached in para posi-
tion to a benzamide scaffold in combination with an amidobutyl-
phenylpiperazine derived pharmacophore. For the groups of
meta-disubstituted benzamides and pyrazolo[1,5-a]pyridine deriv-
atives, such strong effects could not be observed. Among the
amidopropylphenylpiperazine based dimers, the meta-disubsti-
tuted test compound 27 with four oligoethylene glycol units dis-
played more than 20-fold gain of affinity for D2long and D2short
.
The subtype specific gain of affinity by exploitation of secondary
binding elements (allosteric binding sites) shows that the bivalent
ligand approach is a fruitful concept in drug discovery.
4. Experimental section
Dry solvents and reagents were of commercial quality and were
used as purchased. MS were run on a JEOL JMS-GC Mate II spec-
trometer by EI (70 eV) with solid inlet or a Bruker Esquire 2000
by APC or ionization. HR-EIMS were run on a JEOL JMS-GC Mate
II using Peak-Matching (M/DM > 5000). NMR spectra were ob-
tained on a Bruker Avance 360 or a Bruker Avance 600 spectrom-
eter relative to TMS in the solvents indicated (J value in hertz).
Melting points were determined with a MEL-TEMP II melting point
apparatus (Laboratory Devices, USA) in open capillaries and are gi-
ven uncorrected. IR spectra were performed on a Jasco FT/IR 410
spectrometer. Purification by flash chromatography was per-
formed using Silica Gel 60; TLC analyses were performed using
Merck 60 F254 aluminum sheets and analyzed by UV light
(254 nm). Analytical HPLC was performed on Agilent 1100 HPLC
systems employing a VWL detector. As column, a ZORBAX ECLIPSE
3. Conclusion
XDB-C18 (4.6 ꢂ 150 mm, 5
lm) was used. HPLC purity was mea-
A series of amidoalkylphenylpiperazine based dimers along
with their monomeric analogues were synthesized. Radioligand
binding studies showed an up to 70-fold increase of D3 binding
affinity and substantial enhancement of subtype selectivity for
the target compounds 32, 34 and 36 when compared to the mono-
valent control agent 24. These dimeric ligands feature a linker
sured using following binary solvent systems: system A, eluent
CH3OH in 0.1% aqueous trifluoroacetic acid, 10% to 100% CH3OH
in 15 min, 100% for 3 min, flow rate 1.0 mL/min, k 254 nm; system
B, eluent CH3CN in 0.1% aqueous trifluoroacetic acid, 10% CH3CN
for 2 min, then 10–50% CH3CN in 13 min, then 50–100% CH3CN
in 9 min, then 100% for 1 min, flow rate 1.0 mL/min, k 254 nm.

D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
459
The purity of all test compounds and key intermediates was deter-
mined to be >95%. CHN elementary analyses were performed at the
chair of Organic Chemistry of the Friedrich–Alexander University
Erlangen–Nuernberg.
4.5. Tetraethylene glycole di(3-methoxycarbonyl phenyl) ether
(6a)
Tetraethylene glycol di(p-toluenesulfonate) (0.1 mL, 0.25 mmol)
was added dropwise to a solution of methyl 3-hydroxybenzoate
(229 mg, 1.5 mmol) and K2CO3 (415 mg, 3.0 mmol) in DMF
(4.6 mL). After being stirred at 70 °C for 16 h the mixture was trea-
ted with a saturated solution of NaHCO3 and subsequently ex-
tracted with EtOAc. The organic layer was washed with brine and
dried over Na2SO4. The solvent was removed under reduced pres-
sure and the residue was purified by flash chromatography (hex-
ane/EtOAc 1:1) to give 6a in 90% yield (104 mg). EI-MS: m/z 463
(M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 3.67–3.75 (m, 8H),
3.84–3.89 (m, 4H), 3.90 (s, 6H), 4.14–4.19 (m, 4H,), 7.09–7.13 (m,
2H), 7.32 (t, J = 8.0 Hz, 2H,); 7.55–7.58 (m, 2H), 7.60–7.64 (m, 2H);
13C NMR: (CDCl3, 90 MHz) d (ppm): 52.1, 67.7, 69.7, 70.7, 70.9,
4.1. Methyl 3-(2-ethoxyethoxy)benzoate (4a)
A
suspension of methyl 3-hydroxybenzoate (104 mg,
0.68 mmol), 2-bromoethyl ethyl ether (0.22 mL, 2.0 mmol) and
K2CO3 (550 mg, 4.0 mmol) in DMF (13 mL) was stirred for 16 h at
70 °C. After addition of a saturated solution of NaHCO3 the mixture
was extracted with EtOAc. The organic layer was washed with brine
and dried over Na2SO4. The solvent was removed under reduced
pressure and the residue was purified by flash chromatography
(hexane/EtOAc 5:1) to give 4a in 89% yield (136 mg). EI-MS: m/z
224 (M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.25 (t, J = 7.0 Hz,
3H), 3.61 (q, J = 7.0 Hz, 2H), 3.78–3.83 (m, 2H), 3.91 (s, 3H),
4.14–4.19 (m, 2H), 7.11–7.15 (m, 1H), 7.33 (t, J = 8.0 Hz, 1H),
7.58–7.60 (m, 1H), 7.61–7.65 (m, 1H); 13C NMR: (CDCl3, 90 MHz) d
(ppm): 15.1, 52.1, 66.9, 67.7, 68.9, 114.8, 120.2, 122.2, 129.4,
114.8, 120.1, 122.2, 129.4, 131.4, 158.8, 167.0; IR: (NaCl)
m
(cmꢁ1): 2875, 1723, 1445, 1290, 1230, 1106, 756.
4.6. Tetraethylene glycole di(3-hydroxycarbonyl phenyl) ether
(6b)
131.4, 158.9, 167.0; IR: (NaCl)
1230, 1109, 756.
m
(cmꢁ1): 2872, 1724, 1445, 1289,
Compound 6b was synthesized from 6a (101 mg) according to
the procedure described for 4b yielding 88 mg (92%) of 6b. 1H
NMR: (CD3OD, 600 MHz) d (ppm): 3.64–3.72 (m, 8H), 3.82–3.87
(m, 4H), 4.12–4.18 (m, 4H), 7.12–7.17 (m, 2H), 7.35 (t, J = 8.0 Hz,
2H), 7.53–7.56 (m, 2H,); 7.58–7.61 (m, 2H); 13C NMR: (CD3OD,
90 MHz) d (ppm): 68.9, 70.8, 71.7, 71.8, 116.3, 120.7, 123.3,
4.2. 3-(2-Ethoxyethoxy)benzoic acid (4b)
A mixture of methyl 3-(2-ethoxyethoxy)benzoate (4a) (104 mg,
0.47 mmol), aq. NaOH (1 mol/L, 8.7 mL) and MeOH (8.9 mL) was
heated at reflux temperature for 6 h. After cooling to ambient tem-
perature the solution was washed with Et2O and subsequently
acidified to pH 3 with aq. HCl. After extraction with CH2Cl2 the or-
ganic layer was dried over Na2SO4 evaporated under reduced pres-
sure. The resulting solid (98 mg, 99%) was used without further
purification. EI-MS: m/z 210 (M+); 1H NMR: (CD3OD, 360 MHz) d
(ppm): 1.22 (t, J = 7.0 Hz, 3H), 3.60 (q, J = 7.0 Hz, 2H), 3.77–3.83
(m, 2H), 4.12–4.19 (m, 2H), 7.14–7.20 (m, 1H), 7.37 (t, J = 8.0 Hz,
130.6, 133.4, 160.4, 169.7; IR: (NaCl)
1289 1243, 1102, 759.
m
(cmꢁ1): 2888, 1690, 1449,
4.7. Tetraethylene glycole di(3-methoxycarbonylpyrazolo[1,5-
a]pyridine-5-yl) ether (7a)
Compound 7a was synthesized from methyl 5-hydroxypyrazol-
o[1,5-a]pyridine-3-carboxylate32 (70 mg) according to the proce-
dure described for 6a yielding 28 mg (86%) of 7a. EI-MS: m/z 543
(M+); 1H NMR: (CDCl3, 600 MHz) d (ppm): 3.69–3.76 (m, 8H),3.88
(s, 6H), 3.90–3.93 (m, 4H), 4.22–4.26 (m, 4H), 6.64 (dd,
J1 = 7.5 Hz, J2 = 3.0 Hz, 2H), 7.40 (d, J = 3.0 Hz, 2H), 8.26 (s, 2H),
8.31 (d, J = 7.5 Hz, 2H); 13C NMR: (CDCl3, 90 MHz) d (ppm): 51.0,
68.1, 69.2, 70.7, 70.9, 97.0, 102.5, 108.3, 130.0, 142.7, 145.3,
1H), 7.56–7.58 (m, 1H), 7.59–7.63 (m, 1H); IR: (NaCl)
1684, 1454, 1307, 1247, 1119, 759.
m
(cmꢁ1):
4.3. Methyl 5-(2-ethoxyethoxy)pyrazolo[1,5-a]pyridine-3-carbo-
xylate (5a)
m
(cmꢁ1): 2876, 1698, 1649, 1540, 1277,
Compound 5a was synthesized from methyl 5-hydroxypyrazol-
o[1,5-a]pyridine-3-carboxylate32 (50 mg) according to the proce-
dure described for 4a yielding 60 mg (88%) of 5a. EI-MS: m/z 265
(M+); 1H NMR: (CDCl3, 600 MHz) d (ppm): 1.26 (t, J = 7.0 Hz, 3H),
3.62 (q, J = 7.0 Hz, 2H), 3.83–3.86 (m, 2H), 3.89 (s, 3H), 4.23–4.26
(m, 2H); 6.67 (dd, J1 = 7.5 Hz, J2 = 2.5 Hz, 1H), 7.42 (d, J = 3.0 Hz,
1H), 8.28 (s, 1H, H-2), 8.32 (d, J = 7.5 Hz, 1H); 13C NMR: (CDCl3,
90 MHz) d (ppm): 15.1, 51.0, 66.9, 68.2, 68.4, 97.0, 102.5, 108.5,
158.7, 164.0; IR: (NaCl)
1248, 1215, 1109, 1056.
4.8. Tetraethylene glycole di(3-hydroxycarbonylpyrazolo[1,5-
a]pyridine-5-yl) ether (7b)
Compound 7b was synthesized from 7a (34 mg) according to the
procedure described for 4b yielding 25 mg (78%) of 7b. EI-MS: m/z
130.0, 142.7, 145.3, 158.9, 164.1; IR: (NaCl)
1648, 1539, 1275, 1248, 1213, 1119, 1056.
m
(cmꢁ1): 2870, 1697,
427 (M+-2ꢂ COO); 1H NMR: (DMSO-d6, 600 MHz)
d
(ppm):
3.54–3.63 (m, 8H); 3.77–3.83 (m, 4H), 4.19–4.25 (m, 4H), 6.79
(dd, J1 = 7.5 Hz, J2 = 3.0 Hz, 2H), 7.33 (d, J = 3.0 Hz, 2H), 8.26 (s,
2H), 8.67 (d, J = 7.5 Hz, 2H); 12.25 (s, 2H); 13C NMR: (DMSO-d6,
90 MHz) d (ppm): 68.0, 68.5, 69.8, 69.9, 96.4, 102.5, 107.9, 130.9,
4.4. 5-(2-Ethoxyethoxy)pyrazolo[1,5-a]pyridine-3-carboxylic
acid (5b)
141.9, 145.1, 158.3, 164.2; IR: (NaCl)
1537, 1284, 1227, 1110, 1057.
m
(cmꢁ1): 3433, 2892, 1663,
Compound 5b was synthesized from methyl 5-(2-ethoxyeth-
oxy)pyrazolo[1,5-a]pyridine-3-carboxylate (5a, 50 mg)31 according
to the procedure described for 4b yielding 43 mg (91%) of 5b. EI-
MS: m/z 250 (M+); 1H NMR: (DMSO-d6, 600 MHz) d (ppm): 1.34
(t, J = 7.0 Hz, 3H), 3.52 (q, J = 7.0 Hz, 2H), 3.73–3.78 (m, 2H),
4.21–4.25 (m, 2H), 6.81 (dd, J = 7.5 Hz, 3.0 Hz, 1H), 7.34 (d,
J = 3.0 Hz, 1H), 8.27 (s, 1H), 8.69 (d, J = 7.5 Hz, 1H), 12.26 (s, 1H);
13C NMR: (DMSO-d6, 90 MHz) d (ppm): 15.0, 65.7, 67.8, 68.0,
4.9. Methyl 4-(2-ethoxyethoxymethyl)benzoate (10a)
To a solution of methyl 4-(hydroxymethyl)benzoate (500 mg,
3.0 mmol) and 2-bromoethyl ethyl ether (2.0 mL, 18 mmol) in dry
DMF (25 mL) was added dropwise KHMDS (18 mL, 0.5 M in
toluene, 9.0 mmol). After being stirred at 70 °C was 2 h the mixture
was treated with waterand extracted with CH2Cl2. The combined or-
ganic layers were washed with brine and dried over MgSO4. After
96.4, 102.4, 108.0, 130.9, 141.9, 145.1, 158.3, 164.1. IR: (NaCl)
m
(cmꢁ1): 3435, 2877, 1662, 1531, 1281, 1218, 1121, 1056.

460
D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
evaporation the crude product was purified by flash chromatogra-
phy (hexane/EtOAc 5:1) to give 10a in 49% yield (350 mg). EI-MS:
m/z 238 (M+); 1H NMR: (CDCl3, 600 MHz) d (ppm): 1.23 (t,
J = 7.0 Hz, 3H), 3.55 (q, J = 7.0 Hz, 2H), 3.61–3.67 (m, 4H), 3.91 (s,
3H), 4.64 (s, 2H), 7.42 (m, 2H), 8.01 (m, 2H); 13C NMR: (CDCl3,
90 MHz) d (ppm): 15.2, 52.0, 66.7, 69.9, 69.9, 72.6, 127.2, 129.3,
layers were washed with brine, dried (MgSO4) and evaporated.
Purification by flash chromatography (hexane/EtOAc 3:1) yielded
pure 13 (118 mg, 90%). EI-MS: m/z 269 (M+); 1H NMR: (CDCl3,
600 MHz) d (ppm): 3.96 (s, 3H), 4.56 (s, 2H), 7.02 (dd, J1 = 7.0 Hz,
J2 = 2.0 Hz, 1H); 8.18 (dd, J1 = 2.0 Hz, J2 = 1.0 Hz, 1H), 8.43 (s, 1H),
8.52 (dd, J1 = 7.0 Hz, J2 = 1.0 Hz, 1H); 13C NMR: (CDCl3, 150 MHz)
d (ppm): 31.4, 51.3, 104.3, 114.7, 118.3, 129.5, 137.6, 140.4,
129.7, 143.7, 167.0; IR: (NaCl)
1119, 764.
m
(cmꢁ1): 2868, 1692, 1426, 1292,
145.4, 163.6; IR: (NaCl)
1274, 1243, 1049, 778.
m
(cmꢁ1): 2359, 1691, 1643, 1536, 1367,
4.10. 4-(2-Ethoxyethoxymethyl)benzoic acid (10b)
4.14. Dimethyl 4,40-(2,5,8,11,14-pentaoxapentadecan-1,15-
diyl)dibenzoate (14a)
2 N NaOH (15.4 ml) was added to a solution of 10a (367 mg,
1.54 mmol) in MeOH (29.5 mL). After being stirred for 4 h at reflux
temperature the solution was allowed to cool to room temperature.
After washing with CH2Cl2, the mixture was acidified with 2 N HCl
and extracted several times with CH2Cl2. The combined organic lay-
ers were dried over Na2SO4 and evaporated under reduced pressure.
Purification by flash chromatography (hexane/EtOAc 1:3 + 1%
HCOOH) afforded 10b in 70% yield (245 mg).). EI-MS: m/z 224
(M+); 1H NMR: (DMSO-d6, 600 MHz) d (ppm): 1.11 (t, J = 7.0 Hz,
3H), 3.44 (q, J = 7.0 Hz, 2H), 3.52–3.60 (m, 4H), 4.57 (s, 2H), 7.44
(m, 2H), 7.92 (m, 2H), 12.89 (s, 1H); 13C NMR: (DMSO-d6, 90 MHz)
d (ppm): 15.1, 65.5, 69.1, 69.4, 71.4, 127.1, 129.2, 129.7, 143.7, 167.1.
Compound 14a was synthesized from commercially available
methyl 4-bromomethylbenzoate (2.1 g, 9.2 mmol) according to
the procedure described for 18a yielding 0.91 g (50%) of 14a.
EI-MS: m/z 491 (M+); 1H NMR: (CDCl3, 600 MHz) d (ppm):
3.62–3.71 (m, 16H), 3.91 (s, 6H), 4.61 (s, 4H), 7.41 (m, 4H), 8.00
(m, 4H); 13C NMR: (CDCl3, 150 MHz) d (ppm): 52.0, 69.9, 70.6,
70.6, 70.7, 72.6, 127.2, 129.3, 129.7, 144.7, 166.9; IR: (NaCl)
m
(cmꢁ1): 2867, 1723, 1613, 1436, 1280, 1107, 756.
4.15. 4,40-(2,5,8,11,14-Pentaoxapentadecan-1,15-diyl)dibenzoic
acid (14b)
4.11. Methyl 5-(2-ethoxyethoxymethyl)pyrazolo[1,5-a]pyridin-
3-carboxylate (11a)
Compound 14b was synthesized from 14a (0.49 g) according to
the procedure described for 18b yielding 0.38 g (98%) of 14b. 1H
NMR: (CD3OD, 360 MHz) d (ppm): 3.60–3.70 (m, 16H), 4.60 (s,
4H), 7.44 (m, 4H), 7.99 (m, 4H). 13C NMR: (CD3OD, 150 MHz) d
(ppm): 71.0, 71.6, 71.6, 71.6, 73.4, 128.4, 130.8, 131.2, 145.3,
To a solution of 9 (62 mg, 0.3 mmol) and 2-bromoethyl ethyl
ether (0.2 mL, 1.8 mmol) in dry THF (2.5 mL) was added dropwise
KHMDS (1.8 mL, 0.5 M in toluene, 0.9 mmol) at 0 °C. After being stir-
red at ambient temperature for 2 h, the mixture was heated to 70 °C
over night. After being cooled to room temperature the mixture was
treated with water and extracted with CH2Cl2. The combined organ-
ic layers were washed with brine and dried over Na2SO4. After evap-
oration the crude product was purified by flash chromatography
(hexane/EtOAc 3:2) to give 11a in 18% yield (15 mg). APCI-MS: m/z
279 (M++1); 1H NMR: (CDCl3, 600 MHz) d (ppm): 1.24 (t, J = 7.0 Hz,
3H), 3.56 (q, J = 7.0 Hz, 2H), 3.64–3.67 (m, 2H), 3.68–3.71 (m, 2H),
3.91 (s, 3H), 4.67 (s, 2H), 7.01 (dd, J1 = 7.0 Hz, J2 = 2.0 Hz, 1H),
8.08–8.11 (m, 1H), 8.38 (s, 1H), 8.46–8.50 (m, 1H); 13C NMR: (CDCl3,
90 MHz) d (ppm): 15.2, 51.2, 66.8, 69.9, 70.2, 72.0, 103.5, 113.4,
116.6, 129.2, 139.1, 140.7, 145.1, 163.9.
169.7; IR: (NaCl) m
(cmꢁ1): 2874, 1695, 1613, 1427, 1295, 1100, 760.
4.16. Dimethyl 4,40-(2,5,8,11,14,17-hexaoxaoctadecan-1,18-
diyl)dibenzoate (15a)
Compound 15a was synthesized from commercially available
methyl 4-bromomethylbenzoate (2.1 g, 9.2 mmol) and pentaethyl-
ene glycol according to the procedure described for 18a yielding
0.92 g (56%) of 15a. APCI-MS: m/z 535 (M++1); 1H NMR: (CDCl3,
360 MHz) d (ppm): 3.62–3.72 (m, 20H), 3.91 (s, 6H, 4.61 (s, 4H),
7.38–7.43 (m, 4H), 7.98–8.03 (m, 4H); 13C NMR: (CDCl3, 90 MHz)
d (ppm): 52.1, 69.9, 70.7, 70.7, 72.6, 127.2, 129.4, 129.7, 143.7,
167.0; IR: (NaCl)
m
(cmꢁ1): 2867, 1721, 1280, 1107, 757.
4.12. 5-(2-Ethoxyethoxymethyl)pyrazolo[1,5-a]pyridin-3-
carboxylic acid (11b)
4.17. 4,40-(2,5,8,11,14,17-hexaoxaoctadecan-1,18-diyl)dibenzoic
acid (15b)
1 N NaOH (0.5 mL) was added to a solution of 11a (15 mg,
54
l
mol) in MeOH (1 mL). After being stirred for 4 h at reflux tem-
Compound 15b was synthesized from 15a (775 mg) according
to the procedure described for 18b yielding 670 mg (91%) of 15b.
APCI-MS: m/z 507 (M++1); 1H NMR: (CDCl3, 360 MHz) d (ppm):
3.63–3.73 (m, 20H), 4.63 (s, 4H), 7.41–7.47 (m, 4H), 8.02–8.08
(m, 4H); 13C NMR: (CDCl3, 90 MHz) d (ppm): 69.9, 70.7, 70.7,
perature the solution was allowed to cool to room temperature.
After washing with CH2Cl2, the mixture was acidified with 2 N HCl
and extracted several times with CH2Cl2. the combined organic lay-
ers were dried over MgSO4 and evaporated under reduced pressure
to give crude 11b (16 mg, 112% yield), which was used for the next
reaction without further purification. EI-MS: m/z 264 (M+); 1H
NMR: (CDCl3, 600 MHz) d (ppm): 1.25 (t, J = 7.0 Hz, 3H), 3.58 (q,
J = 7.0 Hz, 2H), 3.65–3.72 (m, 4H), 4.69 (s, 2H), 7.06 (dd, J1 = 7.0 Hz,
J2 = 1.5 Hz, 1H), 8.13–8.16 (m, 1H), 8.46 (s, 1H), 8.51–8.55 (m, 1H).
72.6, 127.3, 128.5, 130.3, 144.6, 170.9; IR: (NaCl)
1715, 1248, 1103, 753.
m
(cmꢁ1): 2870,
4.18. Dimethyl 4,40-(2,5,8,11,14,17,20-heptaoxahenicosan-1,21-
diyl)dibenzoate (16a)
4.13. Methyl 5-bromomethylpyrazolo[1,5-a]pyridin-3-carboxy
late (13)
Compound 16a was synthesized from commercially available
methyl 4-bromomethylbenzoate (1.8 g, 7.8 mmol) and hexaethyl-
ene glycol according to the procedure described for 18a yielding
0.59 g (38%) of 16a. APCI-MS: m/z 579 (M++1); 1H NMR: (CDCl3,
600 MHz) d (ppm): 3.59–3.72 (m, 24H), 3.91 (s, 6H), 4.62 (s, 4H),
7.39–7.44 (m, 4H), 7.98–8.04 (m, 4H); 13C NMR: (CDCl3, 150 MHz)
d (ppm): 52.1, 69.9, 70.6, 70.6, 70.6, 70.7, 72.6, 127.2, 129.3, 129.7,
A solution of 9 (124 mg, 0.6 mmol) in CH2Cl2 (2 mL) was cooled
to 0 °C. PBr3 (0.11 mL, 1.2 mmol) was added dropwise and the mix-
ture was stirred at room temperature over night. After evaporation
of the solvent the crude product was treated with a saturated solu-
tion of NaHCO3 and extracted with EtOAc. The combined organic
143.7, 167.0; IR: (NaCl)
m
(cmꢁ1): 2868, 1721, 1280, 1107, 757.

D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
461
4.19. 4,40-(2,5,8,11,14,17,20-heptaoxahenicosan-1,21-diyl)
dibenzoic acid (16b)
6.96 (dd, J1 = 7.0 Hz, J2 = 1.5 Hz, 2H), 8.14–8.17 (m, 2H), 8.44 (s,
2H), 8.50–8.54 (m, 2H); 13C NMR: (CDCl3, + TFA, 150 MHz) d
(ppm): 70.1, 70.8, 70.9, 71.0, 71.6, 103.4, 113.3, 116.5, 129.1,
139.7, 140.2, 146.0, 168.9; IR: (NaCl)
1703, 1649, 1533, 1371, 1238, 1049, 779.
Compound 16b was synthesized from 16a (0.57 g) according to
the procedure described for 18b yielding 0.39 g (71%) of 16b. APCI-
m
(cmꢁ1): 3510, 2924, 2858,
MS: m/z 551 (M++1); 1H NMR: (CDCl3, 360 MHz)
d (ppm):
3.60–3.75 (m, 24H), 4.63 (s, 4H), 7.39–7.46 (m, 4H), 8.00–8.08
4.24. 3-(2-Ethoxyethoxy)-N-[3-[4-(2-methoxyphenyl)piperazin-
1-yl]propyl]benzamide (19)
(m, 4H); 13C NMR: (CDCl3, 150 MHz) d (ppm): 69.9, 70.6, 70.6,
70.6, 70.7, 72.6, 127.3, 128.7, 130.3, 144.5, 171.2; IR: (NaCl)
m
(cmꢁ1): 2871, 1717, 1692, 1247, 1105, 755.
A solution of 4b (10 mg, 48
in CH2Cl2 was cooled to 0 °C. After consecutive addition of solutions
of TBTU (19 mg, 60 mol) in DMF (1 mL) and 3-[4-(2-methoxy-
phenyl)piperazin-1-yl]propylamine (22 mg, 90 mol) in CH2Cl2
lmol) and DIPEA (30 lL, 0.18 mmol)
4.20. Dimethyl 4,40-(2,5,8,11,14,17,20,23,26-nonoxa heptacosan-
1,27-diyl)dibenzoate (17a)
l
l
(5 mL), the mixture was stirred for 4 h at room temperature. Addi-
tion of a saturated solution of NaHCO3 was followed by extraction
with CH2Cl2. The combined organic layers were dried over MgSO4
and evaporated under reduced pressure. Purification by flash chro-
matography (CH2Cl2/MeOH 95:5) gave 19 in 94% yield (20 mg). EI-
MS: m/z 442 (M+); 1H NMR: (CDCl3, 600 MHz) d (ppm): 1.21 (t,
J = 7.5 Hz, 3H), 1.82 (br quint, J = 5.5 Hz, 2H), 2.65 (br t, J = 5.5 Hz,
2H), 2.69–2.75 (m, 4H), 3.07–3.13 (m, 4H), 3.51 (q, J = 7.0 Hz, 2H),
3.59 (br q, J = 5.5 Hz, 2H), 3.70 (t, J = 5.0 Hz, 2H), 3.86 (s, 3H), 4.13
(t, J = 5.0 Hz, 2H), 6.85–6.95 (m, 3H), 6.99–7.04 (m, 2H), 7.27 (t,
J = 8.0 Hz, 1H), 7.35–7.39 (m, 1H), 7.43–7.46 (m, 1H), 8.30 (br s,
1H); 13C NMR: (CDCl3, 90 MHz) d (ppm): 15.1, 24.3, 40.9, 50.6,
53.6, 55.4, 58.5, 66.8, 67.6, 68.8, 111.3, 113.5, 117.8, 118.1, 119.1,
Compound 17a was synthesized from commercially available
methyl 4-bromomethylbenzoate (1.0 g, 4.5 mmol) and octaethyl-
ene glycol according to the procedure described for 18a yielding
290 mg (28%) of 17a. APCI-MS: m/z 668 (M++1); 1H NMR: (CDCl3,
600 MHz) d (ppm): 3.57–3.72 (m, 32H), 3.91 (s, 6H), 4.62 (s, 4H),
7.36–7.44 (m, 4H), 7.95–8.04 (m, 4H); 13C NMR: (CDCl3, 90 MHz)
d (ppm): 52.1, 69.9, 70.6, 70.6, 70.7, 72.6, 127.2, 129.4, 129.7,
143.7, 167.0; IR: (NaCl)
m
(cmꢁ1): 2869, 1720, 1281, 1108, 758.
4.21. 4,40-(2,5,8,11,14,17,20,23,26-nonoxaheptacosan-1,27-
diyl)dibenzoic acid (17b)
Compound 17b was synthesized from 17a (185 mg) according
to the procedure described for 18b yielding 170 mg (96%) of 17b.
APCI-MS: m/z 640 (M++1); 1H NMR: (CD3OD, 600 MHz) d (ppm):
3.55–3.71 (m, 32H), 4.62 (s, 4H), 7.42–7.49 (m, 4H), 7.97–8.02
(m, 4H); 13C NMR: (CD3OD, 90 MHz) d (ppm): 71.1, 71.6, 71.6,
71.6, 71.6, 71.7, 73.5, 128.5, 130.9, 131.3, 145.3, 169.7; IR: (NaCl)
121.0, 123.1, 129.3, 136.4, 141.1, 152.3, 159.1, 167.1; IR: (NaCl) m
(cmꢁ1): 3325, 2875, 1643, 1581, 1538, 1500, 1303, 1241, 1120,
750. HRMS (C25H35N3O4): calcd: 441.2628, found: 441.2627. HPLC
(method A): tR = 13.4 min; purity: 98.1%.
4.25. 3-(2-Ethoxyethoxy)-N-[4-[4-(2-methoxyphenyl)piperazin-
1-yl]butyl]benzamide (20)
m
(cmꢁ1): 2871, 1715, 1249, 1105, 756.
4.22. Dimethyl 5,50-(2,5,8,11,14-pentaoxapentadecan-1,15-
diyl)bispyrazolo[1,5-a]pyridin-3-carboxylate (18a)
Compound 20 was synthesized from 4b (10 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 19 yielding 20 mg (90%) of 20. EI-MS: m/z
A solution of tetraethylen glycol (400
l
L, 2.31 mmol) in THF
456 (M+); 1H NMR: (CDCl3, 360 MHz)
d (ppm): 1.24 (t,
(30 mL) was cooled to 0 °C. After dropwise addition of KHMDS
(0.5 M in toluene, 13.9 mL, 6.93 mmol) the mixture was stirred
for 15 min. Subsequently, a solution of 13 (1.86 g, 6.93 mmol) in
THF (8 mL) was added. After being stirred for 2 h at room temper-
ature the mixture was treated with water and extracted with
EtOAc. The combined organic layers were washed with brine and
dried over Na2SO4. After evaporation the crude product was puri-
fied by flash chromatography (hexane/EtOAc 1:1) to give 18a in
46% yield (620 mg). APCI-MS: m/z 571 (M++1); 1H NMR: (CDCl3,
360 MHz) d (ppm): 3.64–3.74 (m, 16H), 3.91 (s, 6H), 4.65 (s, 4H),
7.00 (dd, J1 = 7.0 Hz, J2 = 2.0 Hz, 2H), 8.04–8.08 (m, 2H), 8.36 (s,
2H), 8.47 (dd, J1 = 7.0 Hz, J2 = 1.0 Hz, 2H); 13C NMR:(CDCl3,
90 MHz) d (ppm): 51.2, 70.1, 70.6, 70.7, 70.8, 71.9, 103.5, 113.4,
J = 7.0 Hz, 3H), 1.61–1.75 (m, 4H), 2.47 (br t, J = 6.5 Hz, 2H),
2.63–2.67 (m, 4H), 3.05–3.08 (m, 4H), 3.48 (br q, J = 6.5 Hz, 2H),
3.59 (q, J = 7.0 Hz, 2H), 3.76–3.81 (m, 2H), 3.85 (s, 3H), 4.13–
4.18 (m, 2H), 6.83–6.95 (m, 3H), 6.96–7.07 (m, 2H), 7.27–7.33
(m, 2H), 7.34–7.38 (m, 1H); 13C NMR: (CDCl3, 150 MHz)
d
(ppm): 15.2, 24.4, 27.4, 40.0, 50.5, 53.4, 55.3, 58.0, 66.9, 67.6,
68.8, 111.2, 113.2, 118.0, 118.2, 119.0, 121.0, 122.9, 129.5,
136.4, 141.2, 152.3, 159.0, 167.5; IR: (NaCl)
m
(cmꢁ1): 3332,
2932, 1642, 1583, 1540, 1500, 1301, 1241, 1129, 751. HRMS
(C26H37N3O4): calcd: 455.2784, found: 455.2784. HPLC (method
B): tR = 14.6 min; purity: 99.2.
4.26. 5-(2-Ethoxyethoxy)-N-[3-[4-(2-methoxyphenyl)piperazin-
1-yl]propyl] pyrazolo[1,5-a]pyridine-3-carboxamide (21)
116.5, 129.1, 139.1, 140.7, 145.1, 163.8; IR: (NaCl)
m
(cmꢁ1):
3510, 2924, 2858, 1703, 1649, 1533, 1371, 1238, 1049, 779.
Compound 21 was synthesized from 5b (10 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the pro-
cedure described for 19 yielding 17 mg (89%) of 21. EI-MS: m/z 482
(M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.25 (t, J = 7.0 Hz, 3H),
1.84 (br quint, J = 6.0 Hz, 2H), 2.63 (br t, J = 6.0 Hz, 2H), 2.72–2.75
(m, 4H), 3.14–3.17 (m, 4H), 3.56–3.67 (m, 4H), 3.80–3.84 (m,
2H), 3.86 (s, 3H), 4.21–4.26 (m, 2H), 6.63 (dd, J1 = 7.5 Hz,
J2 = 3.0 Hz, 1H), 6.80–6.89 (m, 1H), 6.90–7.04 (m, 3H), 7.64 (d,
J = 3.0 Hz, 1H), 8.10 (s, 1H), 8.26 (d, J = 7.5 Hz, 1H); 13C NMR:
(CDCl3, 90 MHz) d (ppm): 15.1, 24.4, 46.2, 49.1, 53.3, 55.4, 63.9,
66.8, 68.2, 68.4, 97.2, 105.4, 108.5, 111.3, 118.7, 121.2, 123.8,
4.23. 5,50-(2,5,8,11,14-pentaoxapentadecan-1,15-
diyl)bispyrazolo[1,5-a]pyridin-3-carboxylic acid (18b)
1 N NaOH (21 mL) was added to a solution of 18a (0.66 g, 1.2 mol)
in MeOH (22 mL). After being stirred at reflux temperature for 4 h,
the solutionwas allowedto cool to room temperature. After washing
with CH2Cl2, the mixture was acidified with 2 N HCl and extracted
several times with CH2Cl2. The combined organic layers were dried
over MgSO4 and evaporated under reduced pressure. Purification by
flash chromatography (CH2Cl2/MeOH 98:2 + 1% HCOOH) gave 18b
in 99% yield (630 mg). APCI-MS: m/z 499 (M++1–CO2); 1H NMR:
(CDCl3 + TFA, 360 MHz) d (ppm): 3.71–3.79 (m, 16H), 4.69 (s, 4H),
129.6, 138.8, 141.9, 142.1, 152.2, 158.1, 164.3; IR: (NaCl)
m (cm
ꢁ1): 3309, 2928, 1650, 1540, 1502, 1453, 1280, 1240, 1119, 752;

462
D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
HRMS (C26H35N5O4): calcd: 481.2689, found: 481.2689. HPLC
496 (M++1); 1H NMR: (CDCl3, 600 MHz)
d (ppm): 1.22 (t,
(method A): tR = 17.4 min; purity: 97.0.
J = 7.0 Hz, 3H), 2.02–2.13 (m, 2H), 2.87–3.14 (m, 6H), 3.23–3.37
(m, 4H), 3.54 (q, 2H), 3.60–3.69 (m, 6H), 3.86 (s, 3H), 4.64 (s, 2H),
6.86–6.89 (m, 1H), 6.91–6.96 (m, 2H), 6.99 (dd, J1 = 7.0 Hz,
J2 = 1.5 Hz, 1H), 7.02–7.07 (m, 1H), 7.73–7.81 (m, 1H), 8.23–8.26
(m, 2H), 8.43–8.46 (m, 1H); 13C NMR: (CDCl3, 150 MHz) d (ppm):
15.2, 24.3, 37.9, 49.2, 53.3, 55.4, 56.7, 66.7, 69.8, 69.9, 72.1, 106.6,
111.3, 113.2, 117.3, 118.7, 121.2, 123.9, 128.8, 138.0, 139.9, 140.3,
4.27. 5-(2-Ethoxyethoxy)-N-[4-[4-(2-methoxyphenyl)piperazin-
1-yl]butyl] pyrazolo[1,5-a]pyridine-3-carboxamide (22)
Compound 22 was synthesized from 5b (10 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 19 yielding 16 mg (82%) of 22. EI-MS: m/z
496 (M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.25 (t, J = 7.0 Hz,
3H), 1.76 (br quint, J = 6.50 Hz, 2H), 1.98 (br quint, J = 7.5 Hz, 2H),
2.93–346 (m, 10H), 3.51 (br q, J = 6.0 Hz, 2H), 3.61 (q, J = 7.0 Hz,
2H). 3.81–3.84 (m, 2H), 3.86 (s, 3H), 4.21–4.26 (m, 2H), 6.64 (dd,
J1 = 7.5 Hz, J2 = 2.5 Hz, 1H); 6.88 (d, J = 8.5 Hz, 1H), 6.93 (d,
J = 4.0 Hz, 2H), 7.04–7.09 (m, 1H); 7.61 (d, J = 3.0 Hz, 1H), 8.24 (s,
1H), 8.32 (d, J = 7.5 Hz, 1H), 8.40 (s, 1H); 13C NMR: (CDCl3,
150 MHz) d (ppm): 15.1, 21.1, 26.3, 37.9, 47.5, 52.3, 55.4, 56.7,
66.8, 68.1, 68.4, 97.3, 105.3, 108.6, 111.4, 118.8, 121.2, 124.5,
141.5, 152.2, 164.0; IR: (NaCl)
m
(cmꢁ1): 3413, 2925, 1647, 1554,
1502, 1454, 1246, 1117, 793, 752; HRMS (C27H37N5O4): calcd:
495.2846, found: 495.2844. HPLC (method B): tR = 13.2 min; purity:
>99.5.
4.31. 5-(2-Ethoxyethoxymethyl)-N-[4-[4-(2-
methoxyphenyl)piperazin-1-yl]butyl] pyrazolo[1,5-a]pyridine-
3-carboxamide (26)
Compound 26 was synthesized from 11b (7.4 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 26 yielding 11 mg (79%) of 26. APCI-MS: m/z
129.5, 138.9, 141.9, 142.3, 152.1, 158.2, 164.4; IR: (NaCl)
m (cm
ꢁ1): 3303, 2917, 1650, 1542, 1501, 1451, 1280, 1242, 1119, 753;
HRMS (C27H37N5O4): calcd: 495.2846, found: 495.2846. HPLC
(method A): tR = 17.3 min; purity: 98.3.
510 (M++1); 1H NMR: (CDCl3, 600 MHz)
d (ppm): 1.23 (t,
J = 7.0 Hz, 3H), 1.66-1.79 (m, 4H), 2.64 (br t, J = 7.0 Hz, 2H),
2.77–2.91 (m, 4H), 3.10–3.24 (m, 4H), 3.48–3.56 (m, 4H),
3.61–3.67 (m, 4H), 3.86 (s, 3H), 4.63 (s, 2H), 6.45–6.51 (m, 1H),
6.85–6.88 (m, 1H), 6.89–6.94 (m, 2H), 6.97–7.04 (m, 2H), 8.20 (s,
1H), 8.21–8.23 (m, 1H), 8.42–8.45 (m, 1H); 13C NMR: (CDCl3,
150 MHz) d (ppm): 15.2, 23.6, 27.4, 38.9, 49.9, 53.4, 55.4, 57.9,
66.7, 69.8, 69.9, 72.0, 106.8, 111.2, 113.3, 117.3, 118.4, 121.1,
4.28. 4-(2-Ethoxyethoxymethyl)-N-[3-[4-(2-methoxyphenyl)-
piperazin-1-yl]propyl] benzamide (23)
Compound 23 was synthesized from 10b (6.1 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the pro-
cedure described for 19 yielding 9.1 mg (74%) of 23. EI-MS: m/z 496
(M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.21 (t, J = 7.0 Hz, 3H),
1.83 (br quint, J = 5.5 Hz, 2H), 2.65 (br t, J = 6.0 Hz, 2H), 2.70–2.74
(m, 4H), 3.07–3.11 (m, 4H), 3.52 (q, J = 7.0 Hz, 2H), 3.57–3.64
(m, 6H), 3.86 (s, 3H), 4.59 (s, 2H), 6.85–6.97 (m, 3H), 6.99–7.05
(m, 1H), 7.37 (d, J = 8.5 Hz, 2H), 7.82 (d, J = 8.5 Hz, 2H), 8.24
(s, 1H). 13C NMR: (CDCl3, 150 MHz) d (ppm): 15.2, 24.3, 40.9, 50.6,
53.6, 55.4, 58.5, 66.7, 69.7, 69.8, 72.6, 111.3, 118.1, 121.0, 123.1,
123.3, 128.7, 137.9, 140.3, 140.7, 140.8, 152.2, 163.5; IR: (NaCl)
m
(cmꢁ1): 3323, 2933, 1647, 1554, 1502, 1454, 1242, 1117, 750;
HRMS (C28H39N5O4): calcd: 509.3002, found: 509.3002. HPLC
(method A): tR = 14.4 min; purity: 95.2.
4.32. Tetraethylene glycol di[[3-N-[3-[4-(2-methoxyphenyl)
piperazin-1-yl]propyl]aminocarbonyl]phenyl] ether (27)
127.2, 127.4, 134.1, 141.1, 141.7, 152.3, 167.1; IR: (NaCl)
m (cm
ꢁ1): 3333, 2940, 2871, 1641, 1543, 1501, 1304, 1241, 1114, 750;
HRMS (C26H37N3O4): calcd: 455.2784, found: 455.2785. HPLC
(method A): tR = 17.6 min; purity: 95.7.
A solution of 6b (10 mg, 0.023 mmol) and DIPEA (0.03 mL,
0.18 mmol) in CH2Cl2 (2 ml) was cooled to 0 °C. TBTU (19 mg,
0.06 mmol) in DMF (1 ml) and 3-[4-(2-methoxyphenyl)-pipera-
zin-1-yl]propylamine (34 mg, 0.14 mmol) in CH2Cl2 (4.6 ml)
were added and the solution was stirred for 4 h at room tem-
perature. The reaction was treated with a saturated aqueous
solution of NaHCO3 followed by extraction with CH2Cl2. The
combined organic layers were washed with brine, dried
(MgSO4) and evaporated. The crude product was purified by
flash chromatography (CH2Cl2/MeOH 9:1) yielding 14 mg
(68%) of 27. EI-MS: m/z 898 (M+); 1H NMR: (CDCl3, 600 MHz)
d (ppm): 1.84 (br quint, J = 6.0 Hz, 4H), 2.66 (br t, J = 5.0 Hz,
4.29. 4-(2-Ethoxyethoxymethyl)-N-[4-[4-(2-methoxyphenyl)-
piperazin-1-yl]butyl] benzamide (24)
Compound 24 was synthesized from 10b (5.0 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 19 yielding 6.6 mg (64%) of 24. EI-MS: m/z
496 (M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.22 (t, J = 7.0 Hz,
3H), 1.62–1.76 (m, 4H), 2.49 (br t, J = 7.0 Hz, 2H), 2.65–2.69
(m, 4H), 3.06–3.10 (m, 4H), 3.49 (br q, J = 6.5 Hz, 2H), 3.54
(q, J = 7.0 Hz, 2H), 3.59–3.64 (m, 4H), 3.85 (s, 3H), 4.61 (s, 2H),
6.83–6.94 (m, 3H), 6.96–7.03 (m, 1H), 7.40 (d, J = 8.5 Hz, 2H), 7.75
(d, J = 8.5 Hz, 2H); 13C NMR: (CDCl3, 150 MHz) d (ppm): 15.2, 24.4,
27.4, 39.9, 50.4, 53.4, 55.4, 58.0, 66.7, 69.8, 69.9, 72.6, 111.3, 118.2,
121.0, 123.0, 127.0, 127.5, 134.2, 141.2, 141.9, 152.3, 167.5; IR:
4H), 2.71–2.77 (m,
8 H), 3.08–3.15 (m, 8H), 3.58 (br q,
J = 5.5 Hz, 4H), 3.63–3.68 (m, 8H), 3.74–3.78 (m, 4H), 3.86 (s,
6H), 4.10–4.13 (m, 4H), 6.84–6.95 (m, 6H), 6.99–7.03 (m, 4H),
7.26 (t, J = 8.0 Hz, 2H), 7.36–7.40 (m, 2H), 7.42–7.45 (m, 2H),
8.30 (s, 2H); 13C NMR: (CDCl3, 90 MHz) d (ppm): 24.3, 40.6,
50.4, 53.6, 55.4, 58.2, 67.6, 69.6, 70.6, 70.8, 111.3, 113.4,
117.8, 118.2, 119.2, 121.0, 123.1, 129.3, 136.3, 141.0, 152.3,
(NaCl)
m
(cmꢁ1): 3335, 2937, 2867, 1637, 1543, 1501, 1303, 1241,
m
(cmꢁ1): 3306, 2819, 1644, 1582,
1116, 750; HRMS (C27H39N3O4): calcd: 469.2941, found: 469.2941.
HPLC (method A): tR = 17.4 min; purity: 97.1.
159.00, 167.1; IR: (NaCl)
1537, 1500, 1303, 1241, 1117, 751; HRMS (C50H68N6O9): calcd:
896.5048, found: 896.5046.
4.30. 5-(2-Ethoxyethoxymethyl)-N-[3-[4-(2-
methoxyphenyl)piperazin-1-yl]propyl] pyrazolo[1,5-
a]pyridine-3-carboxamide (25)
4.33. Tetraethylene glycol di[[3-N-[4-[4-(2-methoxyphenyl)
piperazin-1-yl]butyl]aminocarbonyl]phenyl] ether (28)
Compound 25 was synthesized from 11b (4.2 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the pro-
cedure described for 19 yielding 5.9 mg (75%) of 25. APCI-MS: m/z
Compound 28 was synthesized from 6b (10 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 27 yielding 24 mg (95%) of 28. EI-MS: m/z 926

D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
463
(M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.61–1.77 (m, 8H), 2.54
(br t, J = 7.0 Hz, 4H), 2.71–2.75 (m, 8H), 3.09–3.12 (m, 8H), 3.47
(br q, J = 6.5 Hz, 4H), 3.64–3.72 (m, 8H), 3.80–3.84 (m, 4H), 3.85
(s, 6H), 4.10–4.14 (m, 4H), 6.83–6.94 (m, 6H), 6.97–7.04 (m, 4H),
7.25–7.30 (t, J = 7.5 Hz, 2H), 7.31–7.34 (m, 2H), 7.34–7.37 (m, 2H);
13C NMR: (CDCl3, 90 MHz) d (ppm): 24.0, 27.3, 39.8, 50.1, 53.4,
55.4, 57.9, 67.6, 69.6, 70.7, 70.8, 111.3, 113.1, 118.1, 118.3, 119.2,
procedure described for 27 yielding 36 mg (97%) of 32. APCI-MS:
m/z 998 (M++1); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.61–1.78
(m, 8H), 2.49 (br t, J = 6.5 Hz, 4H), 2.61–2.77 (m, 8H),
3.00–3.16 (m, 8H), 3.48 (dt, J1 = 6.5 Hz, J2 = 6.0 Hz, 4H), 3.57–3.74 (m,
20H), 3.85 (s, 6H), 4.58 (s, 4H), 6.74–6.82 (m, 2H), 6.83–7.03 (m, 8H),
7.35–7.41 (m, 4H), 7.71–7.78 (m, 4H); 13C NMR: (CDCl3, 90 MHz) d
(ppm): 24.3, 27.4, 39.9, 50.4, 53.4, 55.4, 58.1, 69.7, 70.6, 70.7, 72.6,
111.3, 118.2, 121.0, 123.0, 127.1, 127.5, 134.2, 141.2, 141.8, 152.3,
121.0, 123.1, 129.5, 136.3, 140.9, 152.3, 159.0, 167.5; IR: (NaCl)
m
(cmꢁ1): 3320, 2939, 1643, 1582, 1537, 1501, 1301, 1242, 1118,
753; HRMS (C52H72N6O9): calcd: 924.5361, found: 924.5361. HPLC
(method A): tR = 17.0 min; purity: 95.6.
167.5; IR: (NaCl) m
(cmꢁ1): 3334, 2936, 2817, 1642, 1542, 1501, 1452,
1303, 1241, 1115, 751; HPLC (method B): tR = 15.4 min; purity: 96.5.
4.38. Hexaethylene glycol di[[4-N-[3-[4-(2-methoxyphenyl)
piperazin-1-yl]propyl]aminocarbonyl]benzyl] ether (33)
4.34. Tetraethylene glycol di[[4-N-[3-[4-(2-methoxyphenyl)
piperazin-1-yl]propyl]aminocarbonyl]benzyl] ether (29)
Compound 33 was synthesized from 16b (20 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the pro-
cedure described for 27 yielding 31 mg (84%) of 33. APCI-MS: m/z
1014 (M++1); 1H NMR: (CDCl3, 600 MHz) d (ppm): 1.86–2.05 (m,
4H), 2.80 (t, J = 5.5 Hz, 4H), 2.85–3.01 (m, 8H), 3.14–3.30 (m, 8H),
3.57–3.70 (m, 28H), 3.86 (s, 6H), 4.58 (s, 4H), 6.85–6.96 (m, 6H),
7.01–7.06 (m, 2H), 7.35–7.40 (m, 4H), 7.83–7.90 (m, 4H), 8.22–
8.27 (m, 2H); 13C NMR: (CDCl3, 90 MHz) d (ppm): 24.1, 39.6,
49.7, 53.3, 55.4, 57.3, 69.8, 70.6, 70.7, 72.7, 111.4, 118.4, 121.1,
Compound 29 was synthesized from 14b (9 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the pro-
cedure described for 27 yielding 15 mg (85%) of 29. EI-MS: m/z 926
(M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.83 (br quint, J = 6.0 Hz,
4H), 2.64 (br t, J = 6.0 Hz, 4H), 2.70–2.74 (m, 8H), 3.07–3.11 (m,
8H), 3.56–3.68 (m, 20H), 3.86 (s, 6H), 4.56 (s, 4H), 6.84–6.96 (m,
6H), 6.98–7.04 (m, 2H), 7.36 (d, J = 8.5 Hz, 4H), 7.8 (d, J = 8.5 Hz,
4H), 8.22 (s, 2H); 13C NMR: (CDCl3, 90 MHz) d (ppm): 24.4, 40.8,
50.6, 53.6, 55.4, 58.4, 69.7, 70.6, 70.6, 70.7, 72.6, 111.3, 118.1,
121.1, 123.1, 127.2, 127.4, 134.1, 141.1, 141.7, 152.3, 167.1; IR:
123.6, 127.3, 127.5, 133.7, 140.4, 141.9, 152.3, 167.3; IR: (NaCl)
m
(cmꢁ1): 3584, 2925, 2871, 1652, 1541, 1501, 1453, 1303, 1242,
(NaCl)
m
(cmꢁ1): 3333, 2940, 2819, 1643, 1542, 1501, 1304, 1241,
1114, 1026, 752; HPLC (method B): tR = 15.5 min; purity: 99.4.
1113, 1028, 751; HPLC (method A): tR = 16.0 min; purity: 97.4%.
HPLC (method A): tR = 17.0 min; purity: 97.4.
4.39. Hexaethylene glycol di[[4-N-[4-[4-(2-methoxyphenyl)
piperazin-1-yl]butyl]aminocarbonyl]benzyl] ether (34)
4.35. Tetraethylene glycol di[[4-N-[4-[4-(2-methoxyphenyl)
piperazin-1-yl]butyl]aminocarbonyl]benzyl] ether (30)
Compound 34 was synthesized from 16b (20 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 27 yielding 31 mg (82%) of 34. APCI-MS: m/z
1042 (M++1); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.58–1.77 (m,
8H), 2.50 (br t, J = 6.0 Hz, 4H), 2.61–2.82 (m, 8H), 2.95–3.19 (m,
8H), 3.48 (dt, J1 = 6.0 Hz, J2 = 6.0 Hz, 4H), 3.57–3.76 (m, 24H), 3.85
(s, 6H), 4.58 (s, 4H), 6.71–6.79 (m, 2H), 6.83–6.94 (m, 6H),
6.95–7.03 (m, 2H), 7.35–7.42 (m, 4H), 7.71–7.78 (m, 4H); 13C
NMR: (CDCl3, 90 MHz) d (ppm): 24.3, 27.5, 39.9, 50.4, 53.4, 55.4,
58.1, 69.7, 70.6, 70.7, 72.6, 111.3, 118.3, 121.0, 123.0, 127.1,
Compound 30 was synthesized from 14b (8 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 27 yielding 16 mg (95%) of 30. EI-MS: m/z
954 (M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.63–1.76 (m, 8H),
2.53 (br t, J = 7.0 Hz, 4H), 2.70–2.73 (m, 8H), 3.08–3.11 (m, 8H),
3.48 (br q; J = 6.5 Hz, 4H), 3.59–3.70 (m, 16H), 3.85 (s, 6H), 4.58 (s,
4H), 6.83–6.94 (m, 6H), 6.97–7.03 (m, 2H), 7.37 (d, J = 8.5 Hz, 4H),
7.74 (d, J = 8.5 Hz, 4H). 13C NMR: (CDCl3, 90 MHz) d (ppm): 24.1,
27.3, 39.8, 50.2, 53.4, 55.4, 58.0, 69.7, 70.6, 70.6, 70.7, 72.6, 111.3,
118.3, 121.0, 123.1, 127.1, 127.5, 134.1, 141.0, 141.8, 152.3, 167.5;
127.5, 134.2, 141.2, 141.9, 152.3, 167.5; IR: (NaCl)
3334, 2934, 2817, 1642, 1542, 1501, 1452, 1302, 1241, 1116,
771; HPLC (method B): tR = 15.4 min; purity: 98.7.
m
(cmꢁ1):
IR: (NaCl)
m
(cmꢁ1): 3345, 2938, 1643, 1543, 1500, 1302, 1241,
1096, 1028, 751; HPLC (method A): tR = 16.9 min; purity: 98.0.
4.40. Octaethylene glycol di[[4-N-[3-[4-(2-methoxyphenyl)
piperazin-1-yl]propyl]aminocarbonyl]benzyl] ether (35)
4.36. Pentaethylene glycol di[[4-N-[3-[4-(2-methoxyphenyl)
piperazin-1-yl]propyl]aminocarbonyl]benzyl] ether (31)
Compound 35 was synthesized from 17b (20.5 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the
procedure described for 27 yielding 33 mg (94%) of 35. APCI-MS:
m/z 1102 (M++1); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.78–1.87
(m, 4H), 2.63 (br t, J = 6.0 Hz, 4H), 2.67–2.78 (m, 8H),
3.00–3.15 (m, 8H), 3.56–3.69 (m, 36H), 3.86 (s, 6H), 4.57 (s, 4H),
6.85–6.96 (m, 6H), 6.99–7.05 (m, 2H), 7.33–7.40 (m, 4H),
7.79–7.85 (m, 4H), 8.18–8.25 (m, 2H); 13C NMR: (CDCl3,
150 MHz) d (ppm): 24.4, 40.9, 50.7, 53.6, 55.4, 58.5, 69.7, 70.6,
70.6, 70.6, 72.6, 111.3, 118.1, 121.1, 123.2, 127.2, 127.4, 134.1,
Compound 31 was synthesized from 15b (20 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the
procedure described for 27 yielding 35 mg (95%) of 31. APCI-MS:
m/z 970 (M++1); 1H NMR: (CDCl3, 600 MHz) d (ppm): 1.79–1.89
(m, 4H), 2.66 (br t, J = 5.5 Hz, 4H), 2.69–2.84 (m, 8H), 2.87–3.27 (m,
8H), 3.56–3.68 (m, 24H), 3.86 (s, 6H), 4.57 (s, 4H), 6.85–6.91 (m,
4H), 6.92–6.96 (m, 2H), 6.99–7.05 (m, 2H), 7.33–7.40 (m, 4H),
7.79–7.85 (m, 4H), 8.22–8.28 (m, 2H); 13C NMR: (CDCl3, 150 MHz)
d (ppm): 24.4, 40.7, 50.6, 53.4, 55.4, 58.3, 69.7, 70.6, 70.7, 72.6,
111.4, 118.1, 121.1, 123.2, 127.2, 127.5, 134.1, 141.0, 141.8, 152.3,
m
(cmꢁ1): 3341, 2874, 2820,
167.1; IR: (NaCl)
1451, 1304, 1241, 1114, 733; HPLC (method B): tR = 15.5 min; pur-
ity: 99%. HPLC (method B): tR = 15.5 min; purity: 95.6.
m
(cmꢁ1): 3326, 2937, 2818, 1639, 1542, 1500,
141.1, 141.7, 152.4, 167.1; IR: (NaCl)
1643, 1542, 1501, 1453, 1304, 1241, 1113, 791, 753; HPLC (method
B): tR = 15.1 min; purity: 97.3.
4.37. Pentaethylene glycol di[[4-N-[4-[4-(2-methoxyphenyl)
4.41. Octaethylene glycol di[[4-N-[4-[4-(2-methoxyphenyl)
piperazin-1-yl]butyl]aminocarbonyl]benzyl] ether (32)
piperazin-1-yl]butyl]aminocarbonyl]benzyl] ether (36)
Compound 32 was synthesized from 15b (20 mg) and 4-[4-(2-
Compound 36 was synthesized from 17b (20 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine
according
to
the
methoxyphenyl)-piperazin-1-yl]butylamine according to the

464
D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
procedure described for 27 yielding 32 mg (89%) of 36. APCI-MS:
m/z 1130 (M++1); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.60–1.75
(m, 8H), 2.47 (br t, J = 7.0 Hz, 4H), 2.58–2.71 (m, 8H), 2.98–3.13
(m, 8H), 3.48 (dt, J1 = 6.0 Hz, J2 = 6.0 Hz, 4H), 3.56–3.72 (m, 32H),
3.85 (s, 6H), 4.59 (s, 4H), 6.67–6.77 (m, 2H), 6.83–6.94 (m, 6H),
6.95–7.02 (m, 2H), 7.35–7.42 (m, 4H), 7.71–7.78 (m, 4H); 13C
NMR: (CDCl3, 90 MHz) d (ppm): 24.3, 27.5, 39.9, 50.4, 53.4, 55.4,
58.1, 69.8, 70.6, 70.6, 70.6, 70.7, 72.6, 111.3, 118.3, 121.0, 123.0,
2819, 1648, 1552, 1500, 1453, 1241, 1115, 792, 752; HPLC (method
A): tR = 9.4 min; purity: 98.1.
4.45. 5,50-(2,5,8,11,14-Pentaoxapentadecan-1,15-diyl)bis[N-[4-
[4-(2-methoxyphenyl)piperazin-1-yl]butyl]}pyrazolo[1,5-
a]pyridin-3-carboxamide] (40)
Compound 40 was synthesized from 18b (7.3 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 27 yielding 13 mg (93%) of 40. APCI-MS: m/z
1034 (M++1); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.61–1.75 (m,
8H), 2.50 (br t, J = 6.5 Hz, 4H), 2.65–2.77 (m, 8H), 3.03–3.17 (m,
8H), 3.49 (dt, J1 = 6.0 Hz, J2 = 6.0 Hz, 4H), 3.58–3.75 (m, 16H), 3.85
(s, 6H), 4.59 (s, 4H), 6.40–6.54 (m, 2H), 6.83–7.04 (m, 10H), 8.17
(s, 2H), 8.19–8.23 (m, 2H), 8.41 (dd, J1 = 7.0 Hz, J2 = 0.5 Hz, 2H).
13C NMR: (CDCl3, 90 MHz) d (ppm): 24.3, 27.8, 39.3, 50.4, 53.4,
55.4, 58.1, 69.9, 70.6, 70.6, 70.7, 72.0, 106.9, 111.3, 113.3, 117.2,
118.3, 121.0, 123.0, 128.7, 137.9, 140.3, 140.7, 141.2, 152.3,
127.1, 127.5, 134.2, 141.2, 141.9, 152.3, 167.5; IR: (NaCl)
m (cm
ꢁ1): 3565, 2872, 1642, 1548, 1501, 1454, 1303, 1242, 1115, 752;
HPLC (method B): tR = 15.4 min; purity: 97.6.
4.42. Tetraethylene glycol di[[3-N-[3-[4-(2-methoxyphenyl)
piperazin-1-yl]propyl]aminocarbonyl]pyrazolo[1,5-a]pyridine-
5-yl] ether (37)
Compound 37 was synthesized from 7b (10 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the
procedure described for 27 yielding 16 mg (86%) of 37. EI-MS:
m/z 978 (M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.95 (br quint,
J = 6.0 Hz, 4H), 2.79 (br t, J = 6.0 Hz, 4H), 2.87–2.92 (m, 8H), 3.22–
3.25 (m, 8H), 3.59 (br q, J = 6.0 Hz, 4H), 3.65–3.74 (m, 8H), 3.86 (s,
6H), 3.85–3.89 (m, 4H), 4.17–4.23 (m, 4H), 6.60 (dd, J1 = 7.5 Hz,
J2 = 2.77 Hz, 2H), 6.84–6.97 (m, 6H), 6.99–7.05 (m, 2H), 7.59 (d,
J = 2.5 Hz, 2H), 7.64 (s, 2H), 8.23 (s, 2H), 8.25 (dd, J1 = 7.5 Hz,
J2 = 0.5 Hz, 2H); 13C NMR: (CDCl3, 90 MHz) d (ppm): 24.7, 38.6,
49.8, 53.4, 55.4, 57.3, 68.0, 69.3, 70.7, 70.8, 97.2, 105.6, 108.4,
111.3, 118.3, 121.2, 123.5, 129.5, 140.4, 141.5, 142.1, 152.2,
163.4; IR: (NaCl)
m
(cmꢁ1): 3313, 2935, 1647, 1554, 1502, 1452,
1242, 1115, 793, 750; HPLC (method A): tR = 9.5 min; purity: 94.7.
4.46. 1-Bromo-4-(2-ethoxyethoxy)methyl-2-methoxybenzene
(42)
Compound 42 was synthesized from 41 (200 mg) according to
the procedure described for 11a yielding 97 mg (36%) of 42. EI-MS:
m/z 288 (M+), 288 (M++2); 1H NMR: (CDCl3, 360 MHz) d (ppm):
1.23 (t, J = 7.0 Hz, 3H), 3.56 (q, J = 7.0 Hz, 2H), 3.63–3.68 (m, 2H),
3.69–3.74 (m, 2H), 3.80 (s, 3H), 4.60 (s, 2H), 6.69 (dd, J1 = 9.0 Hz,
J2 = 3.0 Hz, 1H), 7.11 (d, J = 3.0 Hz, 1H), 7.39 (d, J = 8.5 Hz, 1H). 13C
NMR: (CDCl3, 90 MHz) d (ppm): 15.2, 55.5, 66.7, 69.9, 70.2, 72.3,
157.9, 164.0; IR: (NaCl)
m
(cmꢁ1): 3308, 2941, 1650, 1541, 1501,
1453, 1281, 1240, 1118, 769. HPLC (method A): tR = 16.9 min;
purity: 96.2.
112.6, 114.2, 114.8, 133.0, 138.8, 159.2; IR: (NaCl)
m
(cmꢁ1): 2974,
4.43. Tetraethylene glycol di[[3-N-[4-[4-(2-methoxyphenyl)
piperazin-1-yl]butyl]aminocarbonyl]pyrazolo[1,5-a]pyridine-5-
yl] ether (38)
2866, 1595, 1574, 1471, 1296, 1273, 1240, 1119, 1055, 1018, 806.
4.47. 1-[4-(2-Ethoxyethoxymethyl)-2-methoxyphenyl]
piperazine (43)
Compound 38 was synthesized from 7b (10 mg) and 4-[4-(2-
methoxyphenyl)-piperazin-1-yl]butylamine according to the pro-
cedure described for 27 yielding 18 mg (97%) of 38. EI-MS: m/z
1006 (M+); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.66–1.82 (m,
8H), 2.61 (br t, J = 6.5 Hz, 4H), 2.79–2.83 (m, 8H), 3.17–3.20 (m,
8H), 3.49 (br q, J = 6.0 Hz, 4H), 3.65–3.72 (m, 8H), 3.85 (s, 6H),
3.83–3.89 (m, 4H), 4.15–4.20 (m, 4H), 6.60 (dd, J1 = 7.5 Hz,
J2 = 2.5 Hz, 2H), 6.84–7.05 (m, 8H), 7.58 (d, J = 2.5 Hz, 2H), 8.19
(s, 2H), 8.25 (dd, J1 = 7.5 Hz, J2 = 0.5 Hz, 2H). 13C NMR: (CDCl3,
90 MHz) d (ppm): 23.6, 27.5, 38.8, 49.9, 53.2, 55.4, 57.8, 68.0,
69.3, 70.7, 70.8, 97.2, 105.6, 108.5, 111.3, 118.5, 121.1, 123.3,
A solution of 42 (391 mg, 1,35 mmol), piperazine (256 mg,
3.0 mmol), NaOtBu (156 mg, 1.62 mmol), Pd2(dba)3 (12.4 mg,
0.01 mol %) and 2-(di-tert-butyl-phosphino)biphenyl (16 mg
(4 mol %) in toluene (4.6 mL) was heated for 18 h at 115 °C. After
being cooled to room temperature the mixture was filtered through
a pad of CeliteÓ and the filtrate was evaporated. The residue was
purified by flash-chromatography (CH2Cl2/MeOH/Me2EtN 9:1:0.1)
to give 43 in 56% yield (225 mg). APCI-MS: m/z 295 (M++1); 1H
NMR: (CDCl3, 360 MHz) d (ppm): 1.22 (t, J = 7.0 Hz, 3H), 2.18–2.25
(m, 1H), 2.76–2.90 (m, 4H), 2.97–3.11 (m, 4H), 3.54 (q, J = 7.0 Hz,
2H), 3.60–3.64 (m, 2H), 3.65–3.69 (m, 2H), 3.79 (s, 3H), 4.65 (s,
2H), 6.80 (dd, J1 = 8.5 Hz, J2 = 3.0 Hz, 1H), 7.01–7.08 (m, 2H). 13C
NMR: (CDCl3, 90 MHz) d (ppm): 15.2, 46.6, 54.3, 55.5, 66.7, 68.8,
129.5, 140.7, 141.3, 142.2, 152.2, 157.9, 163.8; IR: (NaCl)
m (cm
ꢁ1): 3305, 2938, 1650, 1541, 1500, 1450, 1280, 1240, 1117, 751.
HPLC (method A): tR = 17.1 min; purity: 96.3.
69.9, 70.0, 113.6, 114.3, 121.0, 134.9, 145.1, 156.2; IR: (NaCl)
m (cm
4.44. 5,50-(2,5,8,11,14-Pentaoxapentadecan-1,15-diyl)bis[N-[3-
[4-(2-methoxyphenyl)piperazin-1-yl]propyl]pyrazolo[1,5-
a]pyridin-3-carboxamide] (39)
ꢁ1): 3431, 2929, 1610, 1500, 1458, 1298, 1219, 1119, 1049, 910, 818.
4.48. 4-[4-[4-(2-Ethoxyethoxymethyl)-2-methoxyphenyl]
piperazin-1-yl]butyronitrile (45)
Compound 39 was synthesized from 18b (23.5 mg) and 3-[4-(2-
methoxyphenyl)-piperazin-1-yl]propylamine according to the pro-
cedure described for 27 yielding 38.5 mg (88%) of 39. APCI-MS: m/z
1006 (M++1); 1H NMR: (CDCl3, 360 MHz) d (ppm): 1.82–1.91 (m,
4H), 2.66 (br t, J = 6.1, 4H), 2.70–2.82 (m, 8H), 3.07–3.20 (m, 8H),
3.55–3.70 (m, 20H), 3.86 (s, 6H), 4.59 (s, 4H), 6.82–6.88 (m, 2H),
6.91–7.05 (m, 8H), 7.58–7.65 (m, 2H), 8.21–8.25 (m, 4H), 8.41
(dd, J1 = 7.0 Hz, J2 = 0.5 Hz, 2H); 13C NMR: (CDCl3, 90 MHz) d
(ppm): 25.0, 39.6, 50.6, 53.7, 55.4, 58.1, 69.9, 70.6, 70.7, 70.7,
72.0, 107.1, 111.3, 113.2, 117.4, 118.4, 121.2, 123.2, 128.7, 137.7,
A mixture of 43 (86 mg, 0.29 mmol), K2CO3 (89 mg, 0.65 mmol),
4-bromo butyronitrile (59 lL, 0.59 mmol), NaI (23 mg, 0.16 mmol)
and CH3CN (5 mL) was stirred for 24 h at reflux temperature. After
being cooled to room temperature the mixture was filtered
through a pad of CeliteÓ followed by washing with acetone. After
evaporation the residue was purified by flash-chromatography
(EtOAc) to give 44 in 68% yield (73 mg). APCI-MS: m/z 362
(M++1); 1H NMR: (CDCl3, 600 MHz) d (ppm): 1.22 (t, J = 7.0 Hz,
3H), 1.83–1.89 (m, 2H), 2.46 (t, J = 7.0 Hz, 2H), 2.52 (t, J = 7.0 Hz,
2H), 2.54–2.64 (m, 4H), 3.87 (br t, J = 4.5 Hz, 4H), 3.54 (q, J = 7.0
140.3, 140.8, 140.9, 152.3, 163; IR: (NaCl)
m
(cmꢁ1): 3324, 2938,

D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
465
Hz, 2H), 3.60–3.68 (m, 4H), 3.79 (s, 3H), 4.64 (s, 2H), 6.79 (dd,
J1 = 8.5 Hz, J2 = 3.0 Hz, 1H), 7.02–7.07 (m, 2H). 13C NMR: (CDCl3,
90 MHz) d (ppm): 14.9, 15.2, 22.8, 53.1, 53.8, 55.5, 56.4, 66.7,
68.8, 69.9, 70.0, 113.5, 114.3, 119.8, 120.9, 134.8, 144.7, 156.2;
150 MHz) d (ppm): 55.5, 70.2, 70.6, 70.7, 70.8, 72.3, 112.6, 114.3,
114.7, 133.0, 138.7, 159.1; IR: (NaCl)
1354, 1296, 1111, 876, 810, 754.
m
(cmꢁ1): 2868, 1595, 1469,
IR: (NaCl)
m
(cmꢁ1): 3514, 2871, 2247, 1500, 1458, 1300, 1215,
4.52. 1,10-[2,5,8,11,14-Pentaoxapentadecan-1,15-diylbis(2-
methoxy-4,1-phenylene)]dipiperazine (48)
1132, 1028, 816, 752, 715.
4.49. 4-[4-[4-(2-Ethoxyethoxymethyl)-2-methoxyphenyl]
piperazin-1-yl]butylamine (44)
Compound 48 was synthesized from 47 (1.24 g, 2.1 mmol)) and
piperazine (0.79 g, 9.2 mmol) according to the procedure described
for 43 yielding 0.50 g (40%) of 48. APCI-MS: m/z 603 (M++1); 1H
NMR: (CDCl3, 360 MHz) d (ppm): 2.11–2.28 (m, 2H), 2.77–2.89
(m, 8H), 2.96–3.06 (m, 8H), 3.62–3.73 (m, 16H), 3.78 (s, 6H), 4.63
(s, 4H), 6.79 (dd, J1 = 9.0 Hz, J2 = 3.0 Hz, 2H), 7.00–7.07 (m, 4H).
13C NMR: (CDCl3, 150 MHz) d (ppm): 46.6, 54.3, 55.5, 68.8, 69.8,
A solution of 45 (39 mg, 0.11 mmol) in Et2O (2 mL) was cooled
to ꢁ5 °C. Addition of LiAlH4 (270
lL, 1 M in Et2O, 0.27 mmol) was
followed by stirring for 3 h at room temperature. After addition
of one drop of NaHCO3 (sat. aqueous solution), the mixture was fil-
tered through a layered pad of CeliteÓ/MgSO4/ CeliteÓ. Washing
with Et2O was followed by evaporation and purification by flash-
chromatography (CH2Cl2/MeOH/Me2EtN 95:5:0.1) to give 45 in
36% yield (14.4 mg). APCI-MS: m/z 366 (M++1); 1H NMR: (CDCl3,
360 MHz) d (ppm): 1.22 (t, J = 7.0 Hz, 3H), 1.49–1.64 (m, 4H),
2.42 (t, J = 7.0 Hz, 2H), 2.48–2.67 (m, 6H), 2.76 (t, J = 6.5 Hz, 2H),
2.88 (br t, J = 5.0 Hz, 4H), 3.54 (q, J = 7.0 Hz, 2H), 3.60–3.69 (m,
4H), 3.79 (s, 3H), 4.63 (s, 2H), 6.78 (dd, J1 = 8.5 Hz, J2 = 3.0 Hz,
1H), 7.02–7.09 (m, 2H). 13C NMR: (CDCl3, 90 MHz) d (ppm): 15.2,
24.5, 31.2, 41.8, 53.0, 53.9, 55.5, 58.5, 66.7, 68.8, 69.8, 70.0,
70.6, 70.6, 113.5, 114.4, 121.0, 134.8, 145.2, 156.2; IR: (NaCl)
m
(cmꢁ1): 2924, 2854, 1500, 1462, 1265, 1211, 1095, 814, 756.
4.53. 4-(Benzamido)butyl p-toluene sulfonate (50)
To a cooled solution (0 °C) of 49 (382 mg, 1.98 mmol) in THF
(3.2 mL) was added Et3N (385 L, 2.77 mmol). After stirring for
l
15 min a solution of TosCl (566 mg, 2.97 mmol) in THF (5 mL) was
added dropwise and the mixture was stirred at 40 °C for 16 h. Addi-
tion of ice-water was followed by extraction with CH2Cl2, drying
(Na2SO4) and evaporation of the solvent. The residue was purified
by flash-chromatography (hexane/EtOAc 4:1 to 1:1) to give 50 in
53% yield (370 mg). APCI-MS: m/z 348 (M++1); 1H NMR: (CDCl3,
360 MHz) d (ppm): 1.62–1.80 (m, 4H), 2.43 (s, 3H), 3.43 (dt,
J1 = 6.5 Hz, J2 = 6.5 Hz, 2H), 4.07 (t, J = 6.0 Hz, 2H), 6.23–6.39 (m,
1H), 7.30–7.36 (m, 2H), 7.37–7.45 (m, 2H), 7.46–7.52 (m, 1H),
7.71–7.81 (m, 4H). 13C NMR: (CDCl3, 150 MHz) d (ppm): 21.6, 25.8,
26.4, 39.2, 70.2, 126.9, 127.9, 128.5, 129.9, 131.5, 132.9, 134.5,
113.5, 114.3, 121.0, 134.9, 144.7, 156.2; IR: (NaCl)
3361, 2868, 1500, 1462, 1298, 1215, 1124, 816, 717.
m
(cmꢁ1):
4.50. N-[4-[4-[4-(2-Ethoxyethoxymethyl)-2-methoxyphenyl]
piperazin-1-yl]butyl] benzamide (46)
A solution of 44 (8.5 mg, 23
CHCl3 (1 mL) was cooled to 0 °C. After 15 min benzoyl chloride
(4 L, 35 mol) was added. After being stirred for 24 h at room
lmol) and Et3N (10 lL, 72 lmol) in
l
l
144.9, 167.7; IR: (NaCl) m
(cmꢁ1): 3321, 2927, 1643, 1539, 1358,
temperature, the reaction was treated with a saturated aqueous
solution of NaHCO3 followed by extraction with CHCl3. The com-
bined organic layers were washed with brine, dried (MgSO4) and
evaporated. The crude product was purified by preparative HPLC
(ZORBAX ECLIPSE XDB-C8, 0.1% aqueous TFA/MeOH, gradient)
yielding 6.8 mg (60%) of 46. APCI-MS: m/z 470 (M++1); 1H NMR:
(CDCl3, 360 MHz) d (ppm): 1.21 (t, J = 6.9 Hz, 3H), 1.60–1.77 (m,
4H), 2.49 (br t, J = 6.0 Hz, 2H), 2.54–2.71 (m, 4H), 2.86 (br t,
J = 4.5 Hz, 4H), 3.46–3.57 (m, 4H), 3.59–3.68 (m, 4H), 3.78 (s, 3H),
4.62 (s, 2H), 6.68–6.74 (m, 1H), 6.78 (dd, J1 = 8.5 Hz, J2 = 3.0 Hz,
1H), 6.98 (d, J = 8.5 Hz, 1H), 7.04 (d, J = 3.0 Hz, 1H), 7.38–7.57 (m,
3H), 7.74–7.85 (m, 2H). 13C NMR: (CDCl3, 150 MHz) d (ppm):
15.2, 23.8, 27.4, 39.8, 52.7, 53.7, 55.5, 57.9, 66.7, 68.9, 69.8, 70.0,
113.6, 114.5, 121.0, 127.0, 128.5, 131.3, 134.8, 135.0, 144.5,
1176, 941, 818, 710, 663. HPLC (method A): tR = 13.4 min; purity: 96.
4.54. N,N0-[2,5,8,11,14-Pentaoxapentadecan-1,15-diylbis[(2-
methoxy-4,1-phenylene)piperazin-4,1-diylbutan-4,1-diyl]]
dibenzamide (51)
A suspension of 48 (147 mg, 0.24 mmol), 50 (73 mg, 0.48 mg),
K2CO3 (339 mg, 0.98 mmol) and NaI (73 mg, 0.49 mmol) ind CH3CN
(15 mL) was stirred for 16 h at reflux temperature. After being
cooled to room temperature, the solvent was removed under re-
duced pressure and the residue was treated with a saturated solu-
tion of NaHCO3. Extraction with CH2Cl2, drying (Na2SO4),
evaporation of the solvent and subsequent purification by prepara-
tive HPLC (ZORBAX ECLIPSE XDB-C8, 0.1% aqueous TFA/MeOH, gra-
dient) gave 51 in 12% yield (28 mg). APCI-MS: m/z 954 (M++1); 1H
NMR: (CDCl3, 600 MHz) d (ppm): 1.62–1.73 (m, 8H), 2.47 (br t,
J = 7.0 Hz, 4H), 2.52–2.74 (m, 8H), 2.84 (br t, J = 4.5 Hz, 8H), 3.44–
3.51 (m, 4H), 3.60–3.69 (m, 16H), 3.77 (s, 6H), 4.60 (s, 4H), 6.77
(dd, J1 = 8.5 Hz, J2 = 3.5 Hz, 2H), 6.80–6.84 (m, 2H), 6.97 (d,
J = 8.5 Hz, 2H), 7.02 (d, J = 3.0 Hz, 2H), 7.36–7.51 (m, 6H), 7.75–
7.81 (m, 4H). 13C NMR: (CDCl3, 90 MHz) d (ppm): 24.4, 27.5, 39.9,
52.9, 53.8, 55.5, 58.1, 68.8, 69.8, 70.6, 70.6, 113.4, 114.4, 120.9,
156.3, 167.8; IR: (NaCl)
1296, 1093, 802, 710; HRMS (C27H39N3O4): calcd: 469.2941, found:
469.2941; HPLC (method B): tR = 14.1 min; purity: 95.3.
m
(cmꢁ1): 3313, 2927, 1645, 1539, 1498,
4.51. 1,15-Bis(4-bromo-3-methoxyphenyl)-2,5,8,11,14-
pentaoxapentadecane (47)
A suspension of NaH (1.8 g, 60% in paraffin, 45 mmol) in THF
(50 mL) was cooled to 0 °C. Addition of a cooled solution of 41
(3.66 g, 17 mmol) in THF (30 mL) was followed by stirring for
15 min. After adding tetraethylene glycol di-p-tosylate (1.12 mL,
2.8 mmol) the mixture was warmed to room temperature and stir-
red for 4 h. Addition of water was followed by extraction with
EtOAc, drying (Na2SO4) and evaporation of the solvent. The residue
was purified by flash-chromatography (hexane/EtOAc 6:1 to 1:1)
to give 47 in 74% yield (1.24 mg). EI-MS: m/z 592 (M+); 1H NMR:
(CDCl3, 360 MHz) d (ppm): 3.67–3.69 (m, 8H), 3.70–3.73 (m, 8H),
3.79 (s, 6H), 4.58 (s, 4H), 6.69 (dd, J1 = 8.5 Hz, J2 = 3.0 Hz, 1H),
7.08 (d, J = 3.0 Hz, 1H), 7.38 (d, J = 9.0 Hz, 1H). 13C NMR: (CDCl3,
127.0, 128.5, 131.3, 134.8, 135.1, 144.5, 156.2, 167.8; IR: (NaCl)
m
(cmꢁ1): 3321, 2935, 1643, 1539, 1496, 1296, 1126, 802, 714. HPLC
(method B): tR = 14.6 min; purity: 95.7.
4.55. Radioligand binding studies
Receptor binding studies were carried out as described previ-
ously.40 In brief, competition binding experiments with the human
D2long, D2short, D3 and D4.4 receptors were run on membrane prepara-
tions from CHO cells stably expressing the corresponding receptor.
Assays were run with membranes at protein concentrations per well

466
D. Huber et al. / Bioorg. Med. Chem. 20 (2012) 455–466
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Acknowledgment
This research was funded by a Grant of the Deutsche Fors-
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matograms and 13C NMR spectra of the target compounds and
receptor binding data for the porcine dopamine D1, serotonin
5-HT1A, 5-HT2 and the drenergic alpha1 receptor) associated with
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