Synthesis, labelling and first evaluation of [18F]R91150 as a serotonin 5-HT2A receptor antagonist for PET

Article abstract of DOI:10.1002/jlcr.1565

In psychiatric disorders such as anxiety, depression and schizophrenia, 5-HT_2A receptors play an important role. In order to investigate them in vivo there is an increasing interest in selective and high-affinity radioligands for receptor binding studies using positron emission tomography (PET). Since available radioligands have disadvantages, R91150, which is a selective and high-affinity ligand for 5-HT_2A receptors, was labelled with fluorine-18. This was accomplished in six steps via 4-[ ¹⁸F]fluorophenol and 1-(3-bromopropoxy)-4-[¹⁸F] fluorobenzene within 190 min starting from no-carrier-added [ ¹⁸F]fluoride. The overall radiochemical yield was 3.8±2% and the specific activity was at least 335 GBq/μmol at the end of the synthesis. First ex vivo studies in mice proved the uptake of [¹⁸F]R91150 in the brain. Radiometabolite studies revealed no radiometabolites in the brain, whereas in the plasma at least two could be detected 30 min p.i. Further preclinical studies are encouraged to evaluate the potential of this new 5-HT_2A ligand as a radiotracer for PET. Copyright

Full text of DOI:10.1002/jlcr.1565

Research Article
Received 24 April 2008,
Revised 23 July 2008,
Accepted 15 October 2008
Published online 17 November 2008 in Wiley Interscience
(www.interscience.wiley.com) DOI: 10.1002/jlcr.1565
Synthesis, labelling and first evaluation
of [18F]R91150 as a serotonin 5-HT2A
receptor antagonist for PET
Ã
¨
Ute Muhlhausen, Johannes Ermert, and Heinz H. Coenen
In psychiatric disorders such as anxiety, depression and schizophrenia, 5-HT2A receptors play an important role. In order to
investigate them in vivo there is an increasing interest in selective and high-affinity radioligands for receptor binding
studies using positron emission tomography (PET). Since available radioligands have disadvantages, R91150, which is a
selective and high-affinity ligand for 5-HT2A receptors, was labelled with fluorine-18. This was accomplished in six steps via
4-[18F]fluorophenol and 1-(3-bromopropoxy)-4-[18F]fluorobenzene within 190 min starting from no-carrier-added [18F]fluor-
ide. The overall radiochemical yield was 3.872% and the specific activity was at least 335 GBq/mmol at the end of the
synthesis. First ex vivo studies in mice proved the uptake of [18F]R91150 in the brain. Radiometabolite studies revealed no
radiometabolites in the brain, whereas in the plasma at least two could be detected 30 min p.i. Further preclinical studies
are encouraged to evaluate the potential of this new 5-HT2A ligand as a radiotracer for PET.
Keywords: radiofluorination; 4-[18F]fluorophenol; [18F]R91150; 5-HT2A antagonist; PET
109.7 min. In summary, [11C]MDL 100907 is a very specific high-
affinity ligand for 5-HT2A receptors with the only disadvantage
Introduction
Serotonin (5-HT) receptors supposedly play a major role in
normal human functions such as sleep and appetite. Dysfunc-
tions in serotonergic transmission, mostly of the 5-HT2A
receptor, have been implicated in a great deal of human brain
disorders such as anxiety, depression, Alzheimer’s disease and
schizophrenia.1–3 For studying the role of those receptors in
physiology and pharmacology, there is an increasing interest in
high-affinity and selective radiolabelled ligands for in vivo
receptor binding studies using positron emission tomography
(PET) or single photon emission computer tomography (SPECT).
For this purpose several radiotracers are being used, among
others [11C]MDL 100907 and [18F]altanserin for PET and
5-[123I]I-R91150 for SPECT (cf. Figure 1).
of the short half-life of carbon-11, while the binding of
[18F]altanserin lacks specificity, but due to its availability it is
commonly used for 5-HT2A imaging.
For SPECT studies 5-[123I]I-R91150 has been developed. With a
Ki value of 0.2 nM and a KD value of 0.11 nM,11 it also shows high
affinity to 5-HT2A receptors. Its selectivity with regard to other
5-HT receptor subtypes and different receptors is at least a
factor of 50 higher.11–13 Only IC50 values and no Ki values are
reported13 for binding affinities to all receptors other than
5-HT2A, which makes a direct comparison with altanserin and
MDL 100907 difficult. However, MDL 100907 has at least a 300-
fold lower affinity for any other receptor type,8 while for
altanserin the selectivity is better or comparable to 5-[123I]I-
R91150.6,9 The radioiodinated compound exhibits rather high
non-specific binding and combined with the disadvantages of
SPECT vs PET it appears to be a less suitable ligand for 5-HT2A
imaging.
Both MDL 100907 and altanserin bind to the 5-HT2A receptor
with high affinity. MDL 100907 exhibits a Ki value of 0.2 nM4 and
a KD value of 0.56 nM (using [3H]MDL 100907),5 while altanserin
has a Ki of 0.13 nM6 and a KD of 0.3 nM.7 Binding to other 5-HT
receptor subtypes is very low for MDL 1009075 and moderate to
low for altanserin.7 While binding of MDL 100907 to receptors
outside the serotonergic system, such as dopaminergic or
a-adrenergic receptors, is negligible,8 this is not the case for
altanserin. It shows a relatively high affinity for D2 (62 nM) and
especially for adrenergic-a1 receptors (4.55 nM).6,9 Another
disadvantage of altanserin is the formation of at least four
different metabolites in humans, which may cross the blood–
The parent compound of 5-[123I]I-R91150 does not bear an
iodine atom but does contain a fluorine atom. In a comparative
study, the original R91150 exhibited an IC50 value of 0.18 vs
0.60 nM of the iodinated compound13 and has therefore a high
affinity for 5-HT2A receptors. A comparison of both compounds
Institute of Neurosciences and Biophysics (INB-4): Nuclear Chemistry, Research
brain-barrier.9 A shortcoming of [11C]MDL 100907, however, is Center Ju¨lich GmbH, D-52425 Ju¨lich, Germany
the short half-life of carbon-11 (20.4 min), which not only
requires the availability of a cyclotron on site of the PET scanner
but may also impair to achieve a state of reversible binding.7,10
*Correspondence to: Ute Mu¨hlhausen, Institute of Neurosciences and Biophysics
(INB-4): Nuclear Chemistry, Research Center Ju¨lich GmbH, D-52425 Ju¨lich,
Germany.
This should not be a problem with the half-life of fluorine-18 as E-mail: u.muehlhausen@fz-juelich.de
J. Label Compd. Radiopharm 2008, 52 13–22
Copyright r 2008 John Wiley & Sons, Ltd.

U. Mu¨hlhausen et al.
H
N
OH
OCH3
S
O11CH3
N
18F
N
N
O
F
11C]MDL 100907
[
18F]altanserin
[
O
F
O
F
O
O
O
OMe
OMe
N
N
N
N
H
H
5-[123I]I-R91150
R91150
NH2
NH2
123I
Figure 1. 5-HT2A receptor antagonists.
with regard to binding to other receptors is very favorable for N-alkylated to N-[1-(3-(4-fluorophenoxy)propyl)-4-methyl-4-pi-
the parent compound.13 Further, due to its lower lipophilicity peridinyl]acetamide (5) with 1-(3-bromopropoxy)-4-fluoroben-
less non-selective binding can be expected. Thus, in order to use zene (4) without prior purification. The latter was previously
the advantages of fluorine-18 for PET studies R91150 was prepared by alkylation of 4-fluorophenol with 1,3-dibromopro-
radiofluorinated expecting a highly selective ligand with high pane in a 4 M sodium hydroxide solution similar to the literature
affinity for 5-HT2A receptors.
procedure.18 The bromo-derivative 4 was preferred over the
The non-labelled compound R91150 as well as the labelling earlier used 1-(3-chloropropoxy)-4-fluorobenzene13 because of
precursors were synthesized. While part of this study was its greater reactivity in nucleophilic substitutions. For the
published as an abstract,14 we report here on the synthetic alkylation at the piperidine NH-group, potassium carbonate
strategy for labelling of [18F]R91150 via n.c.a. 4-[18F]fluorophenol and potassium iodide were added and 5 was obtained in a
and on preliminary preclinical data, such as lipophilicity, moderate yield of 26%. Lithium hydride for deprotonation of the
blood–brain-barrier penetration and metabolism.
piperidine NH-group showed no improvement.
For the formation of the benzamide part of R91150, a free
amino-group at the 4-position of the piperidine ring was
necessary. Similar to the literature procedure,16 the acetamide 5
was cleaved with concentrated hydrochloric acid at 1001C for
72 h instead of 24 h13 to get 1-(3-(4-fluorophenoxy)propyl)-4-
methylpiperidin-4-amine (6). Earlier its condensation to the
benzamide was achieved by activating the respective 4-amino-
benzoic acid with ethyl chloroformate as a mixed acid anhydride
and its subsequent reaction with 6.13 However, applying this
procedure using 4-amino-2-methoxybenzoic acid, 4-(ethoxycar-
bonylamino)-2-methoxybenzoic acid was obtained. The mixed
acid anhydride was not formed and therefore the subsequent
amidation to 6 could not take place. This result is under-
standable because the amino function is more reactive than the
acid function.
Results and discussion
Chemistry
For evaluation studies and the determination of appropriate
high-performance liquid chromatography (HPLC) conditions for
the identification of radiolabelled compounds, the standard
compound R91150 had to be synthesized as well as the
corresponding labelling precursors and standard compounds of
intermediates occurring in the labelling process. The general
synthesis of R91150 as well as of different derivatives such as, for
example, 5-I-R91150 is described in a patent by Leysen and Van
Daele.13 However, some reaction steps leading to R91150 could
not be reproduced and modifications or even different reactions
had to be employed. The total synthesis of R91150 was
performed here as depicted in Scheme 1.
In a first step commercially available N-benzyl-piperid-4-one
was reacted with methyllithium to get 1-benzyl-4-methyl-
piperidin-4-ol (1) similar to the literature procedure.15 As
described,16,17 1 was converted with acetonitrile and concen-
trated sulfuric acid to the acetamide derivative N-(1-benzyl-4-
methylpiperidin-4-yl)acetamide (2) in a Ritter reaction. The next
step was deprotection at the nitrogen of the piperidine ring.
Because of easier handling debenzylation was conducted using
ammonium formate and palladium black instead of hydrogen
gas and palladium-on-charcoal (10%) as described earlier.13 The
In order to avoid an unprotected amino-group on the benzoic
acid, a benzamide formation to N-[1-(3-(4-fluorophenoxy)pro-
pyl)-4-methyl-piperidin-4-yl]-2-methoxy-4-nitrobenzamide
(7)
was carried out with 2-methoxy-4-nitrobenzoic acid instead,
which is commercially available. Furthermore, rather than the
formation of the mixed acid anhydride for activation, (benzo-
triazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate
(PyBOP) was used as the coupling reagent. PyBOP is utilized in
peptide chemistry and does not form a toxic by-product like
(benzotriazol-1-yloxy)tri(dimethylamino)phosphonium
hexa-
fluorophosphate (BOP).19 Using this reaction pathway, 7 was
free base (4-methylpiperidin-4-yl)acetamide (3) was directly almost quantitatively achieved. However, it could not be
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J. Label Compd. Radiopharm 2008, 52 13–22

U. Mu¨hlhausen et al.
OH
O
O
H2SO4
MeLi
Bn
N
N
N
N
Diethyl ether
-78 °C / RT
64 %
MeCN
93 %
Bn
Bn
H
2
1
F
HCOONH4
Pd black
O
4
Br
O
F
O
HN
K2CO3, KI
MeOH
65 °C
100%
N
H
O
DMF, 80 °C
N
26 %
5
N
3
H
COOH
OCH3
F
O
conc. HCl
NO2
100 °C
N
50 %
NH2
PyBOP, TEA
CH2Cl2, RT
100 %
6
F
O
O
OMe
HCOONH4
N
Pd black
N
H
MeOH
65 °C
55 %
7
NO2
F
O
O
OMe
N
N
H
8
R91150
NH2
Scheme 1. Synthesis of R91150.
obtained without small impurities from pyrrolidine derivatives
after purification by column chromatography, but those could
be removed after the next reaction step. Finally the nitro-group
in 4-position of the benzamide part was reduced with
ammonium formate and palladium black in methanol. Purifica-
tion gave 4-amino-N-[1-(3-(4-fluorophenoxy)propyl)-4-methylpi-
peridin-4-yl]-2-methoxy-benzamide (8), also known as R91150.
In order to get the labelling precursor and non-radioactive
standard compounds of intermediates of the labelling process, a
slightly different and more universally applicable concept was
chosen to avoid possible side reactions. Instead of primary
alkylation at the nitrogen of the piperidine ring, first the
formation of the benzamide part was conducted (cf. Scheme 2).
For labelling with fluorine-18 it is essential that all functional
groups containing acidic protons are derivatized by a suitable
protection group. For protection of the amino-group at the
benzamide part, the tert-butoxycarbonyl (Boc) group was
chosen because it is easily cleaved under acidic conditions.20
Starting with compound 2 cleavage of the acetamide bond
with concentrated hydrochloric acid gave 1-benzyl-4-methyl-
piperidin-amine (9) similar to the literature procedure.17 For the
benzamide formation, 4-(tert-butoxycarbonylamino)-2-methoxy-
benzoic acid (10) was synthesized by reducing the nitro-group
of commercially available 2-methoxy-4-nitrobenzoic acid with
ammonium formate and palladium-on-charcoal (10%).21 Then
the amino-group was protected with di-tert-butyl dicarbonate
providing the intermediate 10. The formation of tert-butyl 4-(1-
benzyl-4-methylpiperidin-4-ylcarbamoyl)-3-methoxyphenylcar-
bamate (11) was achieved similar to the synthesis of 7. As
described above, this compound was also contaminated with
pyrrolidine derivatives, but used without further purification in
the next reaction step. For getting the free NH-function at the
piperidine ring, debenzylation of 11 was performed as described
for 3 using ammonium formate and palladium black. Thus, with
tert-butyl 3-methoxy-4-(4-methylpiperidin-4-ylcarbamoyl)phe-
nylcarbamate (12) a versatile usable precursor was obtained.
J. Label Compd. Radiopharm 2008, 52 13–22
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U. Mu¨hlhausen et al.
COOH
OCH3
10
O
NHBoc
conc. HCl
Bn
N
Bn
N
N
H
NH2
100 °C
PyBOP, TEA
CH2Cl2, RT
100 %
87 %
9
2
HCOONH4
Pd black
O
O
OMe
OMe
Bn
N
HN
N
H
N
MeOH
65 °C
44 %
H
11
12
NHBoc
NHBoc
F
O
Br
F
O
4
O
K2CO3, KI
DMF, 80 °C
53 %
OMe
N
N
H
13
NHBoc
Scheme 2. Syntheses of labelling precursors and standard compounds of R91150.
It proved useful for the alkylation with different procedure via 4-[18F]fluorophenol ([18F]14) as a secondary
phenolarylethers as well as for radioactive labelling using 4- labelling precursor was chosen (cf. Scheme 3).
[18F]fluorophenol ([18F]14). Reaction of 12 with 1-(3-bromopro-
The preparation of 4-[18F]fluorophenol ([18F]14) was adopted
poxy)-4-fluorobenzene (4) gave the N-Boc protected derivative from Ludwig et al.22 reproducibly obtaining an overall radio-
tert-butyl 4-(1-(3-(4-fluorophenoxy)propyl)-4-methylpiperidin-4- chemical yield (RCY) of 50–69% with a very good radiochemical
ylcarbamoyl)-3-methoxyphenylcarbamate (13) in 53% yield, an purity of 498% for isolated [18F]14. By optimization the overall
intermediate of the radiofluorination process. In contrast to the reaction time for synthesis of [18F]14 could be reduced by
alkylation of (4-methylpiperidin-4-yl)acetamide (3), which gave 10–50 min and because all reactions were conducted at the
only moderate yields independent of the base used for same temperature handling of the procedure was simplified.
deprotonation, the nucleophilic substitution of different bro-
mopropyl derivatives with 12 gave good to excellent results.
The next step on the way to [18F]R91150 ([18F]8) was the
synthesis of 1-(3-bromopropoxy)-4-[18F]fluorobenzene ([18F]4), a
Since the radiofluorination of R91150 via the synthesis of compound also described in the literature.22 The obtained
4-[18F]fluorophenol ([18F]14) as a secondary labelling precursor sodium 4-[18F]fluorophenolate was reacted with 1,3-dibromo-
is a possible way, N,N,N-trimethyl-4-(4-(trifluoromethyl)benzoyl)- propane in a two-phase system (sodium hydroxide solution/2-
phenylammonium trifluoromethylsulfonate (15) was prepared, butanone). In HPLC analysis the formation of four different
which was shown to be the best precursor for its formation.22 products besides [18F]4 was observed. One of them was
The preparation was conducted exactly as described earlier22 identified as the elimination product 1-(allyloxy)-4-[18F]fluoro-
aside from the first reaction step to (4-fluorophenyl)-(4- benzene (HPLC (condition II, see ‘Materials and methods’):
(trifluoromethyl)phenyl)methanone, for which a commercially k0 = 3.35), which was formed in varying amounts between 2 and
available fluorophenylmagnesium bromide solution in THF was 15%. The yield of [18F]4 was quite irreproducible under identical
reacted with 4-trifluoromethylbenzonitrile.
reaction conditions (30–80%). The only dependence observed
was that reaction times above 20 min resulted in lower yields.
For further reaction [18F]4 had to be purified and isolation by
HPLC afforded an RCY of 19–40% (relative to [18F]14) and a
radiochemical purity of 499%.
In order to avoid the time-consuming effort of a HPLC isolation
of [18F]4, first attempts of an alkylation of the piperidine
derivative 12 were made with the non-purified radiofluorinated
compound. A great deal of different bases, organic as well as
inorganic, were used for deprotonation of 12 with or without the
addition of potassium iodide. Whatever base was used, however,
the RCY of tert-butyl 4-(1-(3-(4-fluorophenoxy)propyl)-4-methylpi-
peridin-4-ylcarbamoyl)-3-methoxyphenylcarbamate ([18F]13) was
Radiochemistry
For the radiosynthesis of R91150 different synthetic strategies
are conceivable. Generally, it is desirable to have only a few
radioactive reaction steps. However, given the half-life of
fluorine-18 (109.7 min) it is in fact possible to conduct several
radioactive reaction steps. This became necessary here since a
direct [18F]fluoro-for-nitro exchange on the precursor tert-butyl
4-(1-(3-(3-formyl-4-nitrophenoxy)propyl)-4-methylpiperidin-4-yl-
carbamoyl)-3-methoxyphenylcarbamate for labelling of R91150
was not successful. Therefore, the more complex labelling
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U. Mu¨hlhausen et al.
OH
18F
18F]14
[
1,3-dibromopropane
NaOH, 2-butanone, 80 °C
O
Br
O
OMe
HN
Cs2CO3, KI
N
+
H
DMF, 80 °C
18F
12
NHBoc
[
18F]4
18F
O
O
OMe
N
TFA
N
H
CH2Cl2, 80 °C
[
18F]13
NHBoc
18F
O
O
OMe
N
N
H
[
18F]8
18F]R91150
NH2
[
Scheme 3. Radiosynthesis of [18F]R91150 via 4-[18F]fluorophenol and 1-(3-bromopropoxy)-4-[18F]fluorobenzene.
only about 5% as determined by HPLC analysis. However, the use complex reaction procedure including six radioactive steps is
of cesium carbonate/potassium iodide increased the yield to 12% not desirable, a reliable radiosynthesis of [18F]R91150 was
after 10min reaction time. Raising the reaction temperature repetitively possible and sufficient to perform first preclinical
above 801C did not result in higher yields but had a contrary evaluation experiments.
effect.
Because of the very moderate yields the alkylation reaction
was then attempted with purified [18F]4. In the presence of
Lipophilicity
Using the HPLC method corresponding to the OECD guideline
for the testing of chemicals,24 the lipophilicity of R91150 as
well as of altanserin and MDL 100907 was determined. Table 1
gives the log D7.4 values, calculated from HPLC retention
times, and a comparison with the available literature data.
A comparison of k0 values of 5-I-R91150 and R91150 (8 and
2.5, respectively, on a reversed-phase column)12 is in agreement
with a calculation26 of a 1.27 units higher log P value for the
iodinated analogue; i.e. the original compound is more than
10-fold less lipophilic, suggesting the possibility of lower non-
specific binding. The log D7.4 value of 2.53 for R91150
determined here is comparable to MDL 100907, beneficial for
its use in vivo.
cesium carbonate/potassium iodide, alkylation resulted in an
RCY of 48–68% of [18F]13 as determined by HPLC analysis
(condition I: k0 = 6.24). This compound was not isolated but
deprotected similar to the literature procedure23 using trifluoro-
acetic acid (TFA) with almost quantitative yield. Isolation by
reversed-phase HPLC gave [18F]R91150 ([18F]8) in an RCY of
34–38% (relative to [18F]4) with a very good radiochemical
purity of 499%. The overall reaction time starting from
[18F]fluoride was 190 min and the overall RCY was 1.8–5.7%.
[18F]R91150 ([18F]8) was obtained with specific activities of at
least 335 GBq/mmol end of synthesis as appraised by HPLC using
a UV mass calibration curve of non-radioactive R91150 (8) for
the determination of the carrier content. Even though such a
J. Label Compd. Radiopharm 2008, 52 13–22
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U. Mu¨hlhausen et al.
cartridges were purchased from Waters (Eschborn, Germany)
and LiChrolut RP-18e cartridges from Merck (Darmstadt,
Germany). Analytical TLC was performed on aluminum-backed
sheets (Silica gel 60 F254) unless otherwise noted and normal-
phase column chromatography was performed using Silica gel
60, both from Merck.
HPLC was performed on the following system from Dionex
(Idstein, Germany): an Ultimate 3000 LPG-3400A HPLC pump, an
Ultimate 3000 VWD-3100 UV/VIS detector (272 nm), a UCI-50
chromatography interface, an injection valve P/N 8215, analysis
of HPLC data with Chromeleon 6.80 software; radioactivity was
detected with a Gabi Star NaI(Tl) radioactivity detector from
Raytest (Straubenhardt, Germany).
Table 1. Log D7.4 values determined by HPLC and compar-
ison with the literature values
a
Compound
Log D7.4
Literature log P
R91150
Altanserin
MDL 100907
2.53
3.08
1.99
–
3.57
1.9–3.87,10,25
aSDo3%.
Biological evaluation
For the affinity as well as the selectivity of R91150 as a 5-HT2A
ligand, IC50 values are available from the literature.13 Compar-
ison with 5-I-R91150, which is used as a SPECT ligand, promises
better in vivo properties for the original compound. Therefore,
we conducted first ex vivo studies with radiolabelled [18F]R91150
in mice in order to determine the uptake into the brain and the
stability of the tracer.
Reversed-phase HPLC was carried out using a Gemini 5 mm
C18 110A column, for analytical separations with a dimension of
250 mm ꢀ 4.6 mm (flow 1 mL/min) and for semi-preparative
applications with 250 mm ꢀ 10 mm (flow 5 mL/min) from
Phenomenex (Aschaffenburg, Germany). For the separation of
18F-labelled compounds three different conditions were used:
(I) isocratic elution with acetonitrile, water and triethylamine (TEA)
60:40:0.1 (v/v/v) at a pH of 9.0 (phosphoric acid); (II) isocratic
elution with acetonitrile and water 65:35 (v/v); (III) gradient elution
using acetonitrile (0.1% TFA) and water: starting with 10%
acetonitrile (0.1% TFA) the gradient was increased to 100%
acetonitrile (0.1% TFA) over 20min and kept there for 5 min.
NMR spectra (1H-400 MHz) were obtained on an Inova
400 MHz spectrometer (Varian, Darmstadt, Germany). Chemical
shifts are reported in parts per million, solvent peaks were
referenced appropriately. ESI mass spectra were obtained on an
Automass Multi III instrument (Thermoquest, Biberach, Ger-
many). HRMS spectra were obtained on an FTICR ‘LTQ FT Ultra’
(Thermo Fisher Scientific, Dreieich, Germany). Melting points
were measured in a Melting Point B-540 apparatus (Bu¨chi
Labortechnik GmbH, Essen, Germany).
Ex vivo biodistribution
In first studies the ex vivo biodistribution of [18F]R91150 ([18F]8)
in female Navel Medical Research Institute (NMRI) mice was
evaluated 30 and 60 min post injection. Brain uptake was
0.5470.27% ID/g (30 min p.i.) and 1.0970.17% ID/g (60 min p.i.).
The brain to plasma activity ratio, which is a very important
factor for possible PET applications, increased from 1.50 (30 min
p.i.) to 2.64 (60 min p.i.).
Radiometabolites of [18F]R91150
In addition to the ex vivo tissue distribution, the stability of
[18F]R91150 ([18F]8) in the plasma and the brain of mice 30 min
p.i. was examined. Use of acetonitrile as an extraction solvent for
the plasma samples (n = 11) and the brain homogenates (n = 2)
resulted in 9674 and 10076% recovery of radioactivity,
respectively. In plasma, unchanged [18F]R91150 (Rf = 0.60)
accounted for 64% of the measured radioactivity at that time
point. At least two radiometabolites were detected with Rf
values of 0.84 and 0.96, indicating their higher polarity, which
amounted to 20 and 14% of radioactivity measured in the
plasma, respectively. This is a faster metabolism than that of 5-
Chemistry
1-Benzyl-4-methylpiperidin-4-ol (1)
Similar to the literature procedure15 N-benzyl-piperid-4-one
(5.68 g, 30.0 mmol) was dissolved in dry diethyl ether (60 mL)
under an atmosphere of argon and cooled to ꢁ781C. A solution
of methyllithium in diethyl ether (1.6 M, 26 mL, 40.0 mmol) was
added dropwise. The resulting mixture was stirred for 70 min at
room temperature and the reaction terminated by dropwise
addition of water (20 mL) under cooling. The water phase was
extracted with diethyl ether (3 ꢀ 70 mL) and the organic phase
dried over Na2SO4. After evaporation of the solvent the crude
product was purified by silica gel chromatography using 3:1
n-hexane:acetone to give 1 (3.95 g, 19.2 mmol) in 64% yield as a
yellow oil. Comparison with the literature NMR data15,17 proved
the identity of 1. 1H NMR (CDCl3): 7.24–7.17 (m, 5H, Bn), 3.54
(s, 2H, CH2Ph), 2.49 (m, 2H, P2,6), 2.31 (m, 2H, P2,6), 1.61 (m, 2H,
P3,5), 1.53 (m, 2H, P3,5), 1.16 (s, 3H, Me). ESIMS: m/z (% int.) 206.1
(100) [M1H]1.
[
123I]I-R91150 in man where nearly 80% of the tracer was found
unmetabolized in the plasma even after 8 h using radio-HPLC
analysis.27 However, metabolite analysis of 5-[123I]I-R91150 in
baboons showed rapid decomposition of the tracer in the
plasma, leaving only 24710% of intact compound 120 min p.i.28
In the mouse brain, intact [18F]R91150 ([18F]8) accounted for
498% of the radioactivity, indicating that up to 30 min no
labelled radiometabolites crossed the blood–brain barrier or
were formed in the brain.
Thus, first ex vivo studies of [18F]R91150 confirm sufficient in
vivo uptake into the brain without interference from radio-
metabolites.
Materials and methods
N-(1-Benzyl-4-methylpiperidin-4-yl)acetamide (2)
General
N-(1-benzyl-4-methylpiperidin-4-yl)acetamide (2) was prepared
similarly to a previously described procedure.16,17 1-Benzyl-4-
methylpiperidin-4-ol (1) (2.77 g, 13.5 mmol) was dissolved in
absolute acetonitrile (20 mL) under an atmosphere of argon and
The chemicals were analytical grade or better and were used
without further purification (Sigma-Aldrich, Steinheim, Ger-
many). Sep-Pak plus C18, Alumina N and Oasis HLB 1 cc
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J. Label Compd. Radiopharm 2008, 52 13–22

U. Mu¨hlhausen et al.
cooled to 01C. To this solution concentrated sulfuric acid 1.46 (m, 4H, Pip3,5), 1.07 (s, 3H, Me). ESIMS: m/z (% int.) 267.3
(13.4 mL) was added dropwise so that the temperature did not (100) [M1H]1.
exceed 301C. After the addition the mixture was stirred at room
temperature for 24 h. For termination of the reaction the N-[1-(3-(4-Fluorophenoxy)propyl)-4-methylpiperidin-4-yl]-2-meth-
oxy-4-nitrobenzamide (7)
resulting oil was poured onto ice (100 g) and adjusted to pH 10
with a 50% KOH solution. The water phase was extracted with
dichloromethane (4 ꢀ 100 mL), dried over Na2SO4 and the
solvent was evaporated. The resulting yellow solid was levigated
with n-hexane. Product 2 (3.09 g, 12.5 mmol) was obtained as a
yellow solid in 93% yield and its identity verified by comparison
with the literature NMR data.17 1H NMR (CDCl3): 7.26–7.20
(m, 5H, Bn), 3.45 (s, 2H, CH2Ph), 2.49 (m, 2H, Pip2,6), 2.31
(m, 2H, Pip2,6), 1.90 (s, 3H, MeCO), 1.61 (m, 2H, Pip3,5), 1.53
(m, 2H, Pip3,5), 1.16 (s, 3H, Me). ESIMS: m/z (% int.) 247.1
(100) [M1H]1.
2-Methoxy-4-nitrobenzoic acid (130 mg, 0.66 mmol) was dis-
solved in absolute dichloromethane (6 mL) and TEA (103 mL)
under an atmosphere of argon. After 1 min (benzotriazol-1-
yloxy)tripyrrolidinophosphonium hexafluorophosphate (PyBOP)
(343 mg, 0.66 mmol) was added and upon 40 min of stirring at
room temperature 4-amino-1-(3-(4-fluorophenoxy)propyl)-4-
methylpiperidine (6) (176 mg, 0.66 mmol), dissolved in absolute
dichloromethane (2 mL) and TEA (103 mL), was added dropwise.
The resulting solution was stirred for 18 h at room temperature.
Then the organic phase was washed successively with water
(2 ꢀ 10 mL), 5% sodium hydroxide solution (3 ꢀ 10 mL) and
water (2 ꢀ 10 mL). The organic phase was dried over Na2SO4 and
the solvent evaporated. Purification by column chromatography
(4:1 chloroform:methanol) yielded 7 (297 mg) slightly contami-
nated with pyrrolidine derivatives in about 100% as a yellow oil.
The substance was used without further purification.
(4-Methylpiperidin-4-yl)acetamide (3)
Ammonium formate (0.59 g, 9.3 mmol) was dried in vacuo
(1 ꢀ 10ꢁ3 mbar) for 30 min. N-(1-benzyl-4-methylpiperidin-4-yl)a-
cetamide (2) (0.50 g, 2.0 mmol) was dissolved in absolute
methanol (12 mL) under an atmosphere of argon and ammo-
nium formate was added. After the addition of palladium black
(0.10 g) the resulting suspension was heated to 651C and stirred
for 1 h. The mixture was filtered (pore width 1.0 mm) and the
solvent removed. The product was used without further
purification. (4-Methylpiperidin-4-yl)acetamide (3) (362 mg) was
obtained as a white solid with small impurities in about 100%
yield.
4-Amino-N-[1-(3-(4-fluorophenoxy)propyl)-4-methylpiperidin-4-yl]-
2-methoxy-benzamide (8)
Ammonium formate (170 mg, 2.70 mmol) was dried in vacuo
(1 ꢀ 10ꢁ3 mbar) for 30 min. It was added to a solution of 7
(257 mg, approx. 0.58 mmol) in absolute methanol (4 mL) under
an atmosphere of argon and palladium black (30 mg, 0.28 mmol)
was added slowly. The suspension was heated to 651C and
stirred for 75 min. After removing the catalyst by filtration, the
crude product was adsorbed on silica gel. Column chromato-
N-[1-(3-(4-Fluorophenoxy)propyl)-4-methyl-4-piperidinyl]-
acetamide (5)
graphy (4:1 chloroform:methanol) resulted in
8 (132 mg,
(4-Methylpiperidin-4-yl)acetamide (3) (360 mg, 2.3 mmol) was
dissolved in absolute DMF (4 mL) under an atmosphere of argon
and potassium carbonate (320 mg, 2.3 mmol), potassium iodide
(380 mg, 2.3 mmol) and 1-(3-bromopropoxy)-4-fluorobenzene
(4)18 (630 mg, 2.7 mmol) were added. The suspension was
heated to 801C and stirred for 6 h. After removing the solvent
in vacuo the residue was taken up in saturated sodium
carbonate solution (10 mL) and extracted with dichloromethane
(4 ꢀ 20 mL). The organic phase was dried over Na2SO4 and
the solvent evaporated. Purification by silica gel chromatogra-
phy (4:1 chloroform:methanol) gave 5 (182 mg, 0.59 mmol)
in 26% yield as a yellow solid. For comparison with the literature,
the melting point of 5 was determined to be 104.31C
(literature: 104–10629 and 103.41C13). 1H NMR (DMSO-d6):
7.02 (m, 2H, Ph3,5), 6.85 (m, 2H, Ph2,6), 3.87 (t, 2H, A3;
J = 6.30), 2.48 (m, 2H, Pip2,6), 2.40 (t, 2H, A1; J = 7.59), 2.19
(m, 2H, Pip2,6), 1.97 (m, 2H, Pip3,5), 1.79 (m, 2H, A2), 1.73 (s, 3H,
MeCO), 1.40 (m, 2H, Pip3,5), 1.16 (s, 3H, Me). ESIMS: m/z (% int.)
309.2 (100) [M1H]1.
0.32 mmol) as a white porous solid in 55% yield. Comparison
of the melting point of 8 with the literature (76.11C, literature:
75.51C13) confirmed the identity. 1H NMR (DMSO-d6): 7.52 (d, 1H,
Ph60; J = 8.34), 7.05 (m, 2H, Ph3,5), 6.87 (m, 2H, Ph2,6), 6.19
(d, 1H, Ph30; J = 1.61), 6.13 (dd, 1H, Ph50; J = 8.44, 1.73), 3.93
(t, 2H, A1; J = 6.35), 3.80 (s, 3H, OMe), 2.66 (m, 2H, Pip2,6), 2.48
(t, 2H, A3; J = 7.24), 2.21 (m, 2H, Pip2,6), 2.09 (m, 2H, P3,5), 1.83
(m, 2H, A2), 1.51 (m, 2H, Pip3,5), 1.32 (s, 3H, Me). ESIMS: m/z
(% int.) 416.2 (100) [M1H]1.
1-Benzyl-4-methyl-piperidin-4-amine (9)
The deacetylation of N-(1-benzyl-4-methylpiperidin-4-yl)aceta-
mide (2) was conducted as described for the synthesis of 4-
amino-1-(3-fluorophenoxy)propyl-4-methylpiperidine (6) and
similar to the literature procedure.17 Compound 2 (800 mg,
3.25 mmol) was heated at 1001C in concentrated hydrochloric
acid (8.1 mL) for 94 h. Purification by column chromatography
(4:1 chloroform:methanol) gave 9 (580 mg, 2.84 mmol) as a
yellow oil in 87% yield. Comparison with the literature NMR
1
data17 proved the identity of 9. H NMR (DMSO-d6): 7.35–7.26
1-(3-(4-Fluorophenoxy)propyl)-4-methylpiperidin-4-amine (6)
(m, 5H, Bn), 3.54 (s, 2H, CH2Ph), 2.44 (m, 4H, Pip2,6), 1.96 (s, 2H,
NH2), 1.57 (m, 4H, Pip3,5), 1.14 (s, 3H, Me). ESIMS: m/z (% int.)
205.1 (98) [M1H]1.
1-(3-(4-Fluorophenoxy)propyl)-4-methylpiperidin-4-amine
(6)
was prepared similarly to a previously described procedure13,29
extending the reaction time to 72 h. Purification by column
chromatography (4:1 chloroform:methanol) gave 6 as a yellow
oil in 50% yield. 1H NMR (DMSO-d6): 7.03 (m, 2H, Ph3,5), 6.87
(m, 2H, Ph2,6), 3.89 (t, 2H, A1; J = 6.52), 2.45 (m, 2H, Pip2,6),
2.36 (t, 2H, A3; J = 7.14), 2.20 (m, 2H, Pip2,6), 1.78 (m, 2H, A2),
4-(tert-Butoxycarbonylamino)-2-methoxybenzoic acid (10)
To a solution of 4-amino-2-methoxybenzoic acid21 (1.54 g,
9.21 mmol) in absolute methanol (15 mL), TEA (1.28 mL,
J. Label Compd. Radiopharm 2008, 52 13–22
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U. Mu¨hlhausen et al.
9.21 mmol) and di-tert-butyl dicarbonate (2.03 g, 9.21 mmol) (4-Fluorophenyl)-(4-(trifluoromethyl)phenyl)methanone/N,N,
N-trimethyl-4-(4-(trifluoromethyl)benzoyl)phenylammonium
trifluoromethylsulfonate (15)
were added. The mixture was stirred for 7.5 h at room
temperature and then the solvent evaporated. Drying
in vacuo (1 ꢀ 10ꢁ3 mbar) resulted in 10 (1.83 g, 6.85 mmol) as a
beige solid in 74% yield. 1H NMR (DMSO-d6): 7.51 (d, 1H,
Ph6; J = 8.20), 7.26 (d, 1H, Ph3; J = 1.38), 6.95 (dd, 1H, Ph5;
J = 8.46, 1.49), 3.69 (s, 3H, OMe), 1.43 (s, 9H, MeBoc). ESIMS:
m/z (% int.) 268.1 (100) [M1H]1, 212.0 (58) [M-tBu1H]1,
168.2 (53) [M-Boc1H]1. HRMS: calculated for C13H18O5N1:
268.11795, found: 268.11808.
To a solution of 4-fluorophenylmagnesium bromide (15 mL,
1 M in THF, 15.0 mmol) in absolute THF (20 mL) under an
atmosphere of argon a solution of 4-trifluoromethylbenzonitrile
(2.56 g, 15 mmol) in absolute THF (10 mL) was added
dropwise. Then the mixture was heated to 671C for 6 h.
After cooling 1 M sulfuric acid (100 mL) was added and the
organic solvent removed in vacuo. The residual aqueous
phase was extracted with diethyl ether (4 ꢀ 50 mL) and the
solvent was evaporated. (4-Fluorophenyl)-(4-(trifluoromethyl)-
phenyl)methanone (4.02 g, 15.0 mmol) was obtained as a
brownish solid in 100% yield. Spectroscopic data were identical
to those reported earlier.22
In order to get N,N,N-trimethyl-4-(4-(trifluoromethyl)benzoyl)-
phenylammonium trifluoromethylsulfonate (15), (4-fluorophe-
nyl)-(4-(trifluoromethyl)phenyl)methanone was first reacted with
dimethylamino hydrochloride and then with methyl trifluor-
omethanesulfonate to give 15 as described.22
tert-Butyl 4-(1-benzyl-4-methylpiperidin-4-ylcarbamoyl)-3-methox-
yphenylcarbamate (11)
As described for the synthesis of N-[1-(3-(4-fluorophenoxy)pro-
pyl)-4-methylpiperidin-4-yl]-2-methoxy-4-nitrobenzamide (7), 10
(432 mg, 1.62 mmol), dissolved in absolute dichloromethane
(15 mL) and TEA (252 mL), was reacted with PyBOP (842 mg,
1.62 mmol) and subsequently with 1-benzyl-4-methyl-piperidin-
amine (9) (330 mg, 1.62 mmol), dissolved in dichloromethane
(15 mL) and TEA (252 mL). Workup and column chromatography
(4:1 chloroform:methanol) yielded 11 (771 mg) slightly
contaminated with pyrrolidine derivatives as an orange porous
solid in about 100%. This product was used without further
purification.
Radiochemistry
N.c.a. [18F]fluoride was produced via the 18O(p,n)18F nuclear
reaction by the bombardment of an isotopically enriched
[18O] water target with 17 MeV protons at the JSW cyclotron
BC 1710 (FZ Ju¨lich).30 The [18F]fluoride solution was
azeotropically dried as described in the literature by the
addition of Kryptofixs 222 and potassium carbonate for
anion activation.22
tert-Butyl 3-methoxy-4-(4-methylpiperidin-4-ylcarbamoyl)phenyl-
carbamate (12)
As described for the synthesis of (4-methylpiperidin-4-yl)aceta-
mide (3), tert-butyl 4-(1-benzyl-4-methylpiperidin-4-ylcarba-
4-[18F]fluorophenol ([18F]14)
moyl)-3-methoxyphenylcarbamate
(11)
(1.69 g,
approx.
3.70 mmol) in absolute methanol (25 mL), ammonium formate
(1.11 g, 17.5 mmol) and palladium black (191 mg, 1.79 mmol)
were reacted for 3.5 h. Purification by column chromatography
(4:1 chloroform:methanol) gave 12 (585 mg, 1.61 mmol) in
The preparation of 4-[18F]fluorophenol ([18F]14) was carried out
with some modifications of the earlier described procedure.22
Labelling: Compound 15 (10 mg, 25.8 mmol) dissolved in dry
acetonitrile (1 mL) was added to the dry cryptate of [18F]KF and
the mixture stirred for 8 min at 801C. Then the solvent was
evaporated under a stream of argon at 700 mbar.
Oxidation: At 801C the precipitate was dissolved in
a mixture of acetic acid and acetic anhydride (3:2, 2 mL)
and hydrogen peroxide (30%, 0.6 mL) and concentrated
sulfuric acid (0.5 mL) were slowly added. The mixture was
stirred for 10 min before dilution with water (5 mL) and fixation
1
approximately 44% yield as a white solid. H NMR (DMSO-d6):
7.45 (d, 1H, Ph60; J = 8.46), 7.32 (d, 1H, Ph30; J = 1.26), 7.00 (dd, 1H,
Ph50; J = 8.55, 1.30), 3.81 (s, 3H, OMe), 3.08 (m, 2H, Pip2,6), 2.84
(m, 2H, Pip2,6), 2.32 (m, 2H, Pip3,5), 1.42 (s, 9H, MeBoc), 1.66 (m,
2H, Pip3,5), 1.33 (s, 3H, Me). ESIMS: m/z (% int.) 364.2 (100)
[M1H]1. HRMS: calculated for C19H30O4N3: 364.22308, found:
364.22305.
on
a conditioned Sep-Pak plus C18 cartridge (Waters).
tert-Butyl 4-(1-(3-(4-fluorophenoxy)propyl)-4-methylpiperidin-4-yl-
carbamoyl)-3-methoxyphenylcarbamate (13)
The cartridge was washed with water (5 mL) and dried under a
stream of argon. Elution was conducted over a conditioned
Sep-Pak plus Alumina N cartridge (Waters) with diethyl ether
(4 mL). The solvent was evaporated under a stream of argon at
251C and 700 mbar.
Hydrolysis: At 801C aqueous sodium hydroxide (5 M, 0.7 mL)
and methanol (0.07 mL) were added into the reaction vial
and the solution stirred for 5 min. For further reactions the
alkaline phenolate solution was used. For purpose of analysis
the mixture was acidified with concentrated hydrochloric
acid (350 mL), diluted with water (5 mL), fixed on a conditioned
Sep-Pak plus C18 cartridge (Waters) and eluted with
As described for the synthesis of N-[1-(3-(4-fluorophenoxy)pro-
pyl)-4-methyl-4-piperidinyl]acetamide (5), 12 (80 mg, 0.22 mmol)
in dry DMF (1 mL), potassium carbonate (30 mg, 0.22 mmol),
potassium iodide (36 mg, 0.22 mmol) and 1-(3-bromopropoxy)-
4-fluorobenzene (4) (80 mg, 0.30 mmol) in absolute DMF (1 mL)
were reacted for 17 h. Purification by column chromatography
(4:1 chloroform:methanol) gave 13 (60 mg, 0.12 mmol) as an
1
orange porous solid in 53% yield. H NMR (DMSO-d6): 7.63 (d,
1H, Ph60; J = 8.62), 7.34 (d, 1H, Ph30; J = 1.20), 7.04 (m, 2H, Ph3,5),
7.00 (dd, 1H, Ph50; J = 8.54, 1.35), 6.88 (m, 2H, Ph2,6), 3.91 (t, 2H,
A3), 3.81 (s, 3H, OMe), 3.48 (m, 2H, A1), 2.61 (m, 2H, Pip2,6), 2.19
(m, 2H, Pip2,6), 2.11 (m, 2H, Pip3,5), 1.84 (m, 2H, A2), 1.50 (m, 2H,
Pip3,5), 1.42 (s, 9H, MeBoc), 1.33 (s, 3H, Me). ESIMS: m/z (% int.)
516.3 (100) [M1H]1. HRMS: calculated for C28H39F1O5N3:
516.28683, found: 516.28706.
acetonitrile
(1 mL).
Reversed-phase
analytical
HPLC
(condition I) identified the product as 4-[18F]fluorophenol
([18F]14) with elution at k0 = 1.65 and a radiochemical purity of
498%. [18F]14 was prepared in 50–69% RCY in a reaction
time of 50 min.
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J. Label Compd. Radiopharm 2008, 52 13–22

U. Mu¨hlhausen et al.
1-(3-Bromopropoxy)-4-[18F]fluorobenzene ([18F]4)
number of reference compounds (ascorbic acid, benzaldehyde,
anisole, toluene, 4-bromoanisole, 4-iodoanisole) with known
log P values (from ꢁ1.67 to 3.24) were determined and
the capacity factors k0 were calculated. Plotting log k0
against log P gave the reference curve used to determine the
log P values of R91150, altanserin and MDL 100907 by their
retention times.
The alkylation of [18F]14 was carried out with some modifica-
tions to the previously described procedure.22 To the alkaline 4-
[18F]fluorophenolate solution, a mixture of 2-butanone (2.2 mL)
and 1,3-dibromopropane (65 mL, 600 mmol) was added at 801C
and the two-phase system was stirred vigorously for 20 min.
Then the mixture was diluted with water (15 mL) and passed
through a conditioned LiChrolut RP-18e cartridge (Merck). The
cartridge was washed with water (5 mL) and eluted with
acetonitrile (1 mL). The solution was injected onto a reversed-
Ex vivo studies
phase HPLC (condition II). Elution occurred at k0 = 4.47. After Biodistribution
dilution with water (15 mL), the collected fraction was fixed on a
conditioned LiChrolut RP-18e cartridge (Merck), washed with
water (5 mL) and eluted through a glass column (LiChrolut
Female Navel Medical Research Institute (NMRI) mice (permis-
sion to perform animal experiments 50.203.2-KFA 12/02 for
mice) weighing 3974 g received about 110 kBq [18F]R91150
([18F]8) in a volume of 100 mL physiological saline containing 7%
ethanol by tail vein injection. The animals were sacrificed 30 min
(n = 4) and 60 min (n = 2) after tracer injection, the tissues of
interest removed, weighed and the radioactivity measured with
a gamma-counter.
˚
65 ꢀ 10 mm, Merck) filled with 4 A molecular sieves and sodium
sulfate (250 mg) with dry DMF (1 mL). Isolation led to the
labelled compound [18F]4 with an RCY of 19–40%, relative to
4-[18F]fluorophenol ([18F]14), and
of 499%.
a radiochemical purity
tert-Butyl 4-(1-(3-(4-[18F]fluorophenoxy)propyl)-4-methylpiperidin-
4-ylcarbamo-yl)-3-methoxyphenylcarbamate ([18F]13)/4-amino-
N-[1-(3-(4-[18F]fluorophenoxy)propyl)-4-methylpiperidin-4-yl]-2-
methoxy-benzamide ([18F]8)
Radiometabolite analysis
Integrity of the tracer in mice was determined according to an
earlier developed procedure.31 Briefly, blood samples (200 mL
each) of mice (n = 3) were taken 30 min after tracer injection.
Centrifugation (1250 ꢀ g, 3 min, 181C) separated the plasma.
Plasma aliquots (50 mL each) were mixed with an equal volume
of acetonitrile and shaken with a vortex mixer for 2 min. Mouse
brains (approx. 500 mg each) were homogenized by a potter
in ice cold 0.1 N Tris–HCl buffer, pH 7.4 (1500 mL), for 1 min.
Aliquots (500 mL each) were transferred into Eppendorf
vials, mixed with an equal volume of acetonitrile and shaken
with a vortex mixer for 5 min. After centrifugation (20 800 ꢀ g,
2 min, 181C) of the extracts, triplicate samples (5 mL each)
[
18F]4 (in 0.7 mL DMF) was added to a suspension of 12 (17 mg,
50 mmol), cesium carbonate (16.3 mg, 50 mmol) and potassium
iodide (8.3 mg, 50 mmol) in 50 mL dry DMF under an atmosphere
of argon. The mixture was stirred at 801C for 20 min and after
dilution with water (5 mL) passed through a conditioned Sep-
Pak plus C18 cartridge (Waters). The cartridge was washed with
water (5 mL) and eluted with dichloromethane (1 mL). For
analytical purposes aliquots of the reaction mixture were
analyzed using reversed-phase HPLC (condition I) identifying
[
18F]13 at k0 = 6.24.
of the supernatants were spotted on
a TLC plate (SIL
Deprotection was achieved with some modifications to the
literature procedure.23 To the dichloromethane phase, TFA
(150 mL) was added and the mixture evaporated to dryness
under a gentle stream of argon at 801C by adding acetonitrile
(0.5 mL) after removal of dichloromethane. The residue was
dissolved in acetonitrile (0.3 mL) and injected onto a reversed-
phase HPLC (condition III). Elution occurred at k0 = 3.71. After
dilution with water (30 mL), the collected fraction was passed
through a conditioned Oasis HLB 1 cc cartridge (Waters) and
washed with water (5 mL). Elution with ethanol (0.7 mL) led to
G/UV254, Macherey-Nagel, Du¨ren, Germany), the plate was
developed with methanol, air dried and immediately
scanned with an Instant Imager to measure the fluorine-18
activity.
Conclusion
The synthesis of [18F]R91150 ([18F]8), a high-affinity antagonist
of 5-HT2A receptors, was accomplished by a complex labelling
strategy via 4-[18F]fluorophenol ([18F]14) and 1-(3-bromopro-
poxy)-4-[18F]fluorobenzene ([18F]4). In a six-step radiosynthesis,
[18F]R91150 ([18F]8) could be prepared, isolated and formulated
within 190 min. The overall RCY was 1.8–5.7% and the specific
activity was at least 335 GBq/mmol (EOS). Even though such a
complex reaction route is not desirable, the radiofluorinated
compound could be synthesized reproducibly and a preliminary
pharmacological evaluation conducted.
Ex vivo studies with mice showed sufficient uptake of
[18F]R91150 into the brain. Radiometabolite studies in the
mouse brain revealed no radiometabolites, whereas in the
plasma at least two radiometabolites could be detected
30 min p.i. and here only 64% of the radioactivity accounted
for intact [18F]R91150 ([18F]8). The results obtained encourage
the efforts for an improved radiosynthesis and continued
evaluation of this new 5-HT2A antagonist as a radioligand for
PET studies.
[
18F]8 in an RCY of 34–48% relative to 1-(3-bromopropoxy)-
4-[18F]fluorophenol ([18F]4) and a radiochemical purity of
499%. For biological evaluation the ethanol was removed in
vacuo and [18F]8 dissolved in physiological saline containing 7%
ethanol. [18F]R91150 ([18F]8) was prepared in a reaction time of
190 min with an RCY of 1.8–5.7%, relative to [18F]fluoride. The
specific activity was at least 335 GBq/mmol (EOS) as determined
by HPLC using a UV mass calibration curve of the non-
radioactive R91150 (8).
Lipophilicity
Using the HPLC method corresponding to the OECD guideline
for the testing of chemicals,24 the lipophilicities were deter-
mined using a LiChrospher 100 RP-8 (5 mm) column (Merck).
As eluent Soerensen buffer was used (methanol and phosphate
buffer 75:25 (v/v) at a pH of 7.4). The retention times for a
J. Label Compd. Radiopharm 2008, 52 13–22
Copyright r 2008 John Wiley & Sons, Ltd.
www.jlcr.org

U. Mu¨hlhausen et al.
[14] U. Mu¨hlhausen, J. Ermert, H. H. Coenen, J. Labelled Compd.
Radiopharm. 2007, 50, S320.
Acknowledgement
[15] D. Nagarathnam, J. M. Wetzel, S. W. Miao, M. R. Marzabadi,
G. Chiu, W. C. Wong, X. Hong, J. Fang, C. Forray, T. A. Branchek,
W. E. Heydorn, R. S. L. Chang, T. Broten, T. W. Schorn,
C. Gluchowski, J. Med. Chem. 1998, 41, 5320–5333.
[16] F. Himmelsbach, G. Dahlmann, T. Von Ruden, T. Metz, PCT
International Application 1997, WO 9732880, 1997.
[17] I. Shima, A. Kuroda, T. Ohkawa, T. Kurosaki, Y. Sawada, A. Wada,
PCT International Application 2005, WO 2005042533, 2005.
[18] Y. Ohmomo, M. Hirata, K. Murakami, Y. Magata, C. Tanaka,
A. Yokoyama, Chem. Pharm. Bull. 1991, 39, 1038–1040.
[19] J. Coste, D. Le-Nguyen, B. Castro, Tetrahedron Lett. 1990, 31,
205–208.
The authors wish to thank Wiebke Sihver, Annette Schulze
and Dirk Bier for the radiopharmacological evaluation
studies, Marcus H. Holschbach and Sabine Willbold for the
NMR measurements and Diana Hofmann for the HRMS
measurements.
References
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