Synthesis of potent and selective dopamine D4 antagonists as candidate radioligands

Article abstract of DOI:10.1016/S0960-894X(01)00241-4

A series of dopamine D₄ antagonists was synthesized and evaluated as potential candidates for development as positron emission tomography (PET) radioligands. All new compounds display high affinity and selectivity for the D₄ receptors and compounds 5b, 5d, and 5e were identified as candidates for radioligand development.

Full text of DOI:10.1016/S0960-894X(01)00241-4

Bioorganic & Medicinal Chemistry Letters 11 (2001) 1375–1377
Synthesis of Potent and Selective Dopamine D4
Antagonists as Candidate Radioligands
Yiyun Huang,a,* Lawrence S. Kegeles,a Sung-A Bae,a Dah-ren Hwang,a
Bryan L. Roth,b Jason E. Savageb and Marc Laruellea
aDepartment of Psychiatry, Columbia University College of Physicians and Surgeons, New York, NY 10032, USA
bNIMH Psychoactive Drug Screening Program, Departments of Biochemistry and Psychiatry, Case Western Reserve University
Medical School, Cleveland, OH 44106, USA
Received 30 January 2001; accepted 23 March 2001
Abstract—A series of dopamine D4 antagonists was synthesized and evaluated as potential candidates for development as positron
emission tomography (PET) radioligands. All new compounds display high affinity and selectivity for the D4 receptors and com-
pounds 5b, 5d, and 5e were identified as candidates for radioligand development. # 2001 Elsevier Science Ltd. All rights reserved.
The dopamine D4 receptor is a member of the dopamine
D2-like family of receptors, which include the D2, D3,
and D4 receptor subtypes.1,2 The D4 receptor differs
from the D2 and D3 receptors in that it is mostly loca-
lized in the extrastriatal areas of the brain such as cor-
tex, hippocampus and thalamus and thus does not
follow the dopamine concentration gradient in the CNS,
which has the highest DA concentrations in the stria-
tum.1,3 The initial interest in the D4 receptor stemmed
from the findings that the atypical antipsychotic cloza-
pine had a higher affinity for the dopamine D4 receptor
compared to the D2 and that postmortem studies
reported results consistent with an up-regulation of D4
receptors in the striatum of patients with schizo-
phrenia.4,5 Although this initial enthusiasm was somewhat
dampened due to the failure to replicate the postmortem
findings, more recently published results have in general
supported the importance of the D4 receptor in brain
functions and its involvement in neuropsychiatric dis-
orders.6,7 For example, a recent autoradiographic study
reported a selective increase of D4 receptors in the
entorhinal cortex of patients with schizophrenia.8
Jentch et al.9 reported the reversal of PCP-induced cog-
nitive deficits in monkeys by the selective D4 antagonist
NGD 94-1 (Ki 3.0 nM at hD4), suggesting a possible role
for the D4 receptors in the cognitive and memory
impairments seen in schizophrenia and Alzheimer’s
disease. Preclinically, several D4-selective antagonists,
notably NRA 0160 (Ki 0.5 nM at hD4) and CI 1030 (Ki
4.3 at hD4), have been shown to be active in animal
models predictive of antipsychotic efficacy and thus,
may be useful atypical antipsychotics.10,11 In addition,
the investigation of the D4 receptor’s involvement in
attention deficit hyperactivity disorder (ADHD), drug
and alcohol abuse, Parkinsonism, and depression
remain active areas of research.12
Despite the unique localization of the D4 receptor and its
possible involvement in a number of neuropsychiatric
disorders, the study of this receptor subtype in vivo has
so far been hindered by the lack of selective radio-
ligands. To date only two selective D4 radioligands,
[3H]NGD 94-1 and [3H]PNU-101958, have been repor-
ted and utilized in binding and autoradiography studies
in vitro.12,13 However, there are no positron emission
tomography (PET) radioligands available for in vivo
study of the D4 receptor, despite some recent develop-
ment efforts in this direction.14,15 The successful devel-
opment of a selective D4 PET radioligand will allow for
the study of this receptor in vivo to probe its functions
and possible involvement in psychiatric diseases. Herein
we report our effort to synthesize selective D4 receptor
antagonists as candidate compounds for possible devel-
opment as PET radioligands.
The synthetic program was initiated based on the lead
compound L745,870 (Fig. 1), the first potent and selec-
tive D4 receptor antagonist reported in the literature.16
*Corresponding author. Tel.:+1-212-543-6629; fax: +1-212-568-
6171; e-mail: hh285@columbia.edu
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00241-4

1376
Y. Huang et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1375–1377
In binding studies L745,870 displays a high affinity (Ki
0.43 nM at hD4, 1.53 nM at rD4) and excellent selectiv-
ity for D4 over other receptors.17,18 Based on this lead
compound, a number of derivatives bearing functional
groups amenable to labeling with positron-emitting iso-
topes were prepared and screened in a comprehensive in
vitro binding study. Other molecules with alkyl sub-
stituent of various length and type were also synthesized
and screened to probe the structure–activity relation of
this class of compounds.
formed by displacement of the radioligand [3H]spiper-
one at the cloned rat D2 and D4 receptors with the test
compounds. For binding to cloned rat D3 receptor the
radioligand [3H]sulpiride was used and for D1 and D5
[3H]SCH23982 was used. In addition to dopamine
receptors, binding affinities of these compounds at var-
ious cloned human serotonin (5-HT) receptors, includ-
ing 5-HT1A, 5-HT1B, 5-HT1D, 5-HT1E, 5-HT2A, 5-HT2C
,
5-HT5A, 5-HT6, and 5-HT7, as well as a large number of
other biogenic amine receptors were also determined as
previously described.22 Selected biological data are pre-
sented in Table 1.
As is evident from Table 1 the test compounds demon-
strated high affinity for the dopamine D4 receptors and
excellent selectivity over other DA and 5-HT receptors.
All compounds display negligible affinity (Ki >10 mM)
for the following cloned receptors: D1, D5, 5-HT1B, 5-
HT1D, 5-HT1E, 5-HT2C, 5-HT5A, k, d, m H1, NMDA,
M1–M5, nicotinic acetylcholine receptors and the
NMDA PCP site.
Figure 1. Structural representation of L-745,870.
Preparation of these compounds is described in Scheme
1 and was achieved by the condensation of substituted
phenylpiperazines 3 and azagramine 4.14 Synthesis of
azagramine followed the literature procedures,19 while
the preparation of substituted phenylpiperazine, if not
commercially available, was accomplished by the palla-
dium catalyzed aromatic amination of substituted phe-
nylbromides 2 with piperazine 1.20 Monosubstituted
phenylpiperazines 3e–j were obtained in 32–75% yields.
Desired compounds 5a–j were prepared in 45–82%
yields following purification by recrystallization.21 All
From the biological data it is clear that substituents as
diverse as fluorine, methoxy, thiomethyl, vinyl and tri-
fluoromethyl, placed at either the ortho or para position
of the phenylpiperazine moiety in L745,870, are well
tolerated by the D4 binding site and the resulting com-
pounds all retain their high binding affinity and selec-
tivity for the D4 receptor. Replacement of the chlorine
in L745,870 with an alkyl group such as ethyl, propyl,
or vinyl group also resulted in compounds with high
affinity and selectivity for the D4 receptor (5h–i). How-
ever, with a bulky group such as the phenyl in the para
position (5j) the binding affinity at D4 is diminished,
although the selectivity remains.
1
new compounds were characterized by H NMR, MS,
HRMS and/or elemental analysis.
All compounds were tested in receptor binding studies.
Binding to dopamine D2 and D4 receptors was per-
Scheme 1. Synthesis of compounds 5a–j.
Table 1. In vitro binding affinities (Ki, nM) of compounds 5a–j at selected DA and 5-HT receptorsa
Compound
rD4
rD2
rD3
5-HT1A
5-HT2A
5-HT6
5-HT7
L745,870
3.7Æ0.6
2.3Æ0.7
3.6Æ1.1
3.3Æ0.5
1.4Æ0.2
4.7Æ0.5
34.1Æ6.8
2.8Æ0.5
5.0Æ1.0
6.7Æ1.1
148.0Æ22.2
>10,000
>10,000
>10,000
>10,000
438.0
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
668.0
6620.9
2759.7
>10,000
990.7
405.6
>10,000
>10,000
3967.7
>10,000
>10,000
>10,000
453.0
232.2
>10,000
242.0
2311.6
>10,000
2791.0
509.0
627.0
>10,000
1932.0
1656.2
>10,000
>10,000
>10,000
7385.0
>10,000
7109.5
>10,000
4460.7
6486.5
226.3
412.3
>10,000
212.9
102.8
335.9
318.2
>10,000
411.0
713.0
5a
5b
5c
5d
5e
5f
5g
5h
5i
4507.0
>10,000
>10,000
1692.0
921.0
>10,000
5j
>10,000
>10,000
aAll assays were conducted in duplicate. For each Ki value at rD4 the data are the meanÆSD of computer-derived estimates for n=4 separate
determinations. Ki’s for other receptors are the means of n=4 separate determinations.

Y. Huang et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1375–1377
1377
Among the analogues of L745,870, the 4-methoxy-
phenyl derivative (5b) has an affinity for D4 receptor
that is comparable to the lead (Ki 3.6 at rD4 vs 3.7 nM
for L745,870), yet with a slightly better selectivity, while
the 2-methoxyphenyl analogue (5d) has a slightly higher
affinity (Ki 1.4 nM) for the D4 receptor than the lead and
somewhat diminished selectivity. Another compound, 5e,
shows affinity and selectivity very similar to the lead (Ki
4.7 nM). These affinities are determined using the cloned
rat dopamine receptors. It has been demonstrated that
L745,870 displays lower affinity at the rat D4 receptor
than at the human D4 receptor.18 Therefore, the affinity
of 5b, 5d, and 5e at the human D4 receptor might also
be in the subnanomolar range, similar to that of
L745,870 (Ki 0.43 at hD4). Compounds with nanomolar
or subnanomolar affinity at the target receptor are in
general good candidates for development as in vivo
radioligands for PET imaging. As all these three com-
pounds have a group (methoxy or methylthio) that is
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amenable to labeling with
a
positron-emitting
14. Staley, J. K.; Tamagnan, G.; Baldwin, R. M.; Fujita, M.;
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[11C]methyl group, they are suitable candidates for
radiolabeling and development as potential PET radio-
ligands.
In summary, compounds with high affinity and selec-
tivity for the dopamine D4 receptors are synthesized and
assayed in receptor binding studies. Three compounds
(5b, 5d, and 5e) were identified as potential candidates
for development as PET radioligands to investigate the
dopamine D4 receptors in vivo.
17. Bristow, L. J.; Kramer, M. S.; Kulagowski, J.; Patel, S.;
Ragan, C. I.; Seabrook, G. R. Trends Pharmacol. Sci. 1997,
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Fletcher, A. E.; Knowles, M.; Marwood, R.; McAllister, G.;
Myers, J.; Patel, S.; Curtis, N.; Kulagowski, J. J.; Leeson,
P. D.; Ridgill, M.; Graham, M.; Matheson, S.; Rathbone, D.;
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19. Williamson, W. R. N. J. Chem. Soc. 1962, 2833.
20. Zhao, S.-H.; Miller, A. K.; Berger, J.; Flippin, L. A. Tet-
rahedron Lett. 1996, 37, 4463.
Acknowledgements
This study was supported by the National Alliance for
Research in Schizophrenia and Depression (NARSAD),
the Lieber Center for Schizophrenia Research, and the
Public Health Service (NIMH MH59342-01 and K02-
MH01603-01). In vitro binding studies were supported
by NO1-MH80005 and KO2-MH01366.
21. The preparation of compound 5b is typical: Azagramine 4
(175 mg, 1.0 mmol) and 4-(4-methoxyphenyl)piperazine 3b
(1.2 mmol) were dissolved in xylene, stirred at 150 ꢀC for 36 h,
and cooled to room temperature. The solid was collected,
rinsed twice with ether, and dried to give the product 5b
(245 mg, 75%). Recrystallization from acetone afforded an
analytical sample as a colorless solid, mp 209–212 ꢀC. 1H
NMR (400 MHz, CDCl3): d 9.60 (s, 1H), 8.34 (dd, 1H,
J=1.38, 4.75 Hz), 8.14 (dd, 1H, J=1.38, 7.86 Hz), 7.32 (s,
1H), 7.12 (dd, 1H, J=4.75, 7.86 Hz), 6.92 (d, 2H, J=9.2 Hz),
6.84 (d, 2H, J=9.2 Hz), 3.83 (s, 5H), 3.13 (m, 4H), 2.70 (m,
4H). Anal. calcd: C, 70.78; H, 6.88; N, 17.38; found: C, 70.91;
H, 6.87; N, 17.35.
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