Synthesis of an 18F-labeled cyclin-dependent kinase-2 inhibitor

Article abstract of DOI:10.1002/jlcr.1922

The radiosynthesis of N-(5-(((5-(tert-butyl)oxazol-2-yl)methyl)thio) thiazol-2-yl)-4-[¹⁸F]fluoro-benzamide [¹⁸F]2 as a potential radiotracer for molecular imaging of cyclin-dependent kinase-2 (CDK-2) expression in vivo by positron em

Full text of DOI:10.1002/jlcr.1922

Research Article
Received 10 June 2011,
Revised 22 July 2011,
Accepted 25 July 2011
Published online 12 September 2011 in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.1922
18
Synthesis of an F-labeled cyclin-dependent
kinase-2 inhibitor
Frieda Svensson,a Torsten Kniess,a Ralf Bergmann,a Jens Pietzsch,a and
Frank Wuesta,b
*
The radiosynthesis of N-(5-(((5-(tert-butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-4-[18F]fluoro-benzamide [18F]2 as a potential
radiotracer for molecular imaging of cyclin-dependent kinase-2 (CDK-2) expression in vivo by positron emission tomography
is described. Two different synthesis routes were envisaged. The first approach followed direct radiofluorination of respec-
tive nitro- and trimethylammonium substituted benzamides as labeling precursors with no-carrier-added (n.c.a.) [18F]fluo-
ride. A second synthesis route was based on the acylation reaction of 2-aminothiazole derivative with labeling agent [18F]
SFB. Direct radiofluorination afforded 18 F-labeled CDK-2 inhibitor in very low yields of 1%–3%, whereas acylation reaction
with [18F]SFB gave 18 F-labeled CDK-2 inhibitor [18 F]2 in high yields of up to 85% based upon [18 F]SFB during the optimiza-
tion experiments. Large scale preparation afforded radiotracer [18 F]2 in isolated radiochemical yields of 37%–44% (n = 3,
decay-corrected) after HPLC purification within 75 min based upon [18 F]SFB. This corresponds to a decay-corrected radiochem-
ical yield of 13%–16% based upon [18F]fluoride. The radiochemical purity exceeded 95% and the specific activity was deter-
mined to be 20 GBq/mmol.
Keywords: cyclin-dependent kinase 2; fluorine-18; radiofluorination; [18F]SFB
potential CDK-targeting therapeutic compounds. This led to the
Introduction
discovery and evaluation of a broad variety of small molecule inhibi-
Loss of growth control through an aberrant regulation of cell cycle
tors targeting CDKs. Although much biochemical evidence supports
machinery is one of the distinguishing hallmarks of cancer and is
targeting CDK4/6 as the most important CDK for regulating cell pro-
therefore a major focus of biomedical research and various pharma-
liferation, most drug development programs have produced com-
cological intervention strategies.1,2 The cell cycle is divided into
pounds which inhibit multiple CDKs, with CDK-2 and CDK-1 being
four distinct phases: DNA synthesis (S phase), mitosis (M phase),
particularly common targets. CDK-2 and CDK-4 are of particular
and the gaps of varying length between these periods called G1
interest because of their frequent dysregulated activity in a variety
and G2. Nondividing cells remain in a resting or quiescence stage
of human cancers. CDK-2 activity is required for progression
named G0 before they re-enter into phase G1. As in all signaling
through G1 to S phase of the cell cycle, and CDK-2 is one of
the key components of the G1 checkpoint.4 First generation
pathways, regulation of cell cycle progression is achieved by
sequential phosphorylation events of various key regulation
of CDK-2 inhibitors are so-called pan-CDK inhibitors. They
enzymes, called cyclins, and their catalytic partners, the cyclin-
lack selectivity and inhibit multiple kinases. Typical examples
dependent kinases (CDKs). The CDK family is a group of serine-
threonine protein kinases, and CDKs are the driving force behind
the cell cycle and cell proliferation. Because of their critical role in
the regulation of the cell cycle and the observed expression/
include flavopiridol, staurosporine UCN-01, olomoucine,
and roscovitine. The second generation involves selective
and potent CDK-2 inhibitors like various N-acyl and N-aryl-2-
aminothiazoles. Over 100 analogs have been developed, exhibiting
IC50 values in the 1–10 nm range.5,6 A prominent 2-aminothiazole
activity pattern in most human cancers, considerable effort has
been focused on the development of small molecule CDK inhibi-
compound is small molecule CDK inhibitor SNS-032, which
tors as potential therapeutic anticancer agents. To date, 11 CDKs
has entered phase I clinical trials for the treatment of B-cell
have been identified, but only CDKs 1, 2, 3, 4, 5, and 6 are directly
involved in the cell cycle progression. The CDKs are frequently
overactive in human cancer resulting in a loss of checkpoint
integrity and therefore leading to uncontrolled proliferation.
Selective inhibition of CDKs is thought to limit progression of
malignancies and advanced solid tumors. Preclinical studies
have shown that SNS-032 is a specific andpotent inhibitor of
tumor cells through the cell cycle and to facilitate the induction
aInstitute of Radiopharmacy, Helmholtz-Zentrum Dresden-Rossendorf, Dresden,
Germany
of apoptotic pathways to support therapeutic interventions.3,4
Over the past 2 decades, the therapeutic value of targeting
members of the cyclin-dependent kinase (CDK) protein family
bDepartment of Oncology, University of Alberta, Edmonton, Canada
has been extensively studied. A growing body of structural infor-
mation about CDKs in complex with their regulatory cyclin sub-
*Correspondence to: Frank Wuest, Department of Oncologic Imaging , Cross
Cancer Institute, University of Alberta, Canada.
units and synthetic inhibitors has facilitated the development of E-mail: wuest@ualberta.ca
J. Label Compd. Radiopharm 2011, 54 769–774
Copyright © 2011 John Wiley & Sons, Ltd.

F. Svensson et al.
CDK-2. SNS-032 also shows inhibitory potency to CDK-7 and CDK-9. fluorobenzoylated 2-aminothiazole 2 as potential PET radiotracer
Based on the 2-aminothiazole backbone, various N-acylated com- when labeled with 18F (Figure 2).
pounds with high CDK-2 inhibitory potency have been
described. A selection of various CDK-2 inhibitors is given in of 18 F-labeled fluorobenzamide [18 F]2.
Figure 1. The first approach was based on direct radiofluorination of cor-
Two different synthesis routes were explored for the preparation
Recently, we have described the design and synthesis of responding nitro- and trimethylammonium precursor 5 and 7
radiolabeled selective CDK-4/6 inhibitors for molecular using the prominent nucleophilic radiofluorination agent [18F]KF.
imaging of CDK-4/6 expression in vivo by means of positron A second approach exploited the acylation reaction of 2-aminothia-
emission tomography (PET).7 Within the arsenal of molecular zole 3 with labeling agent succinimidyl-4-[18F]fluorobenzoate ([18F]
imaging methodologies, PET represents an important, noninva- SFB).
sive technique for the development and evaluation of anticancer
The synthesis of labeling precursors 5 and 7 and reference
strategies through visualization and assessment of tissue compound 2 is depicted in Figure 3.
function and quantification of metabolic pathways in vivo.
2-Aminothiazole 3 as key intermediate and labeling precursor
We hypothesize that CDK inhibitors labeled with the short- for acylation with [18F]SFB was prepared according to published
lived positron emitters iodine-124 (124I, t1/2 = 4.2 d) and fluorine- literature procedures.5 Acylation of 2-aminothiazole 3 with ben-
18 (18F, t1/2 = 109.8 min) are a novel promising class of PET zoic acid derivatives 4 (R = F, NO2, and NMe2) in the presence
radiotracers for imaging and functional characterization of of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochlo-
components of the cell cycle machinery in tumors in vivo by ride afforded acylated derivatives 2 (R = F), 5 (R = NO2), and 6
PET.7–9
(R = NMe2) in good chemical yields of 78%, 73%, and 80%, respec-
In the present work, we describe the design and synthesis tively, after purification using column chromatography. Conver-
of an 18 F-labeled 2-aminothiazole containing a fluorobenza- sion of dimethylamino group into trimethylammonium group in
mide motif as potential radiotracer for molecular imaging compound 7 was achieved through treatment of compound 6 with
of CDK-2 expression in vivo.
a two-fold excess of MeOTf in nitromethane as the solvent. Removal
of the solvent and washing of the residue with diethyl ether afforded
compound 7 in 90% yield and high chemical purity greater 95%.
Results and discussion
Radiosynthesis of 18 F-labeled 2-aminothiazole [18 F]2
according to a direct radiofluorination reaction
Design concept and synthesis of labeling precursors and
reference compound
Based on the high inhibitory potency of benzamide-containing
2-aminothiazole 1 (IC50 (CDK/E) = 26 nm)5 we propose directed
bioisosteric substitution of hydrogen with fluorine to afford
In a first set of reactions, we explored direct incorporation of
no-carrier-added (n.c.a.) [18 F]fluoride into the aromatic ring
through aromatic nucleophilic substitution on labeling precursors
Figure 1. Selection of first and second generation CDK-2 inhibitors.
Figure 2. Bioisosteric substitution to design fluorobenzamide 2.
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Copyright © 2011 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2011, 54 769–774

F. Svensson et al.
Figure 3. Synthesis of labeling precursors and reference compound.
5 and 7 possessing a NO2 or NMe3 group, respectively, as leaving 5-HT1A receptor ligands p-[18F]MPPF, p-[18F]FBP, and 18 F-labeled
group. The reaction scheme is shown in Figure 4.
Org 13063.10–12
Direct incorporation of [18 F]fluoride into benzamides is known
to be challenging. This type of reaction usually requires high tem-
perature (150–200 ꢀC), and reactions are preferentially performed
in polar aprotic solvents with a high boiling point like dimethyl sulf-
Radiosynthesis of 18 F-labeled 2-aminothiazole [18 F]2
according to acylation reaction with [18 F]SFB
oxide (DMSO), dimethylformamide (DMF), and N-methyl-2-pyrro- The very low radiochemical yields obtained using direct incor-
lidone (NMP). Prominent examples include radiosynthesis of poration of n.c.a. [18 F]fluoride into benzamides 5 or 7 prompted
5-HT1A receptor ligands p-[18 F]MPPF and p-[18 F]FBP.10,11 Synth- us to envisage an alternative synthesis route for the preparation
esis of both radiotracers requires high reaction temperature of of 4-[18 F]fluorobenzamide [18 F]2.
150–200 ꢀC resulting in relatively low radiochemical yields between
The second synthesis approach is based on acylation of 2-ami-
5% and 8%. Better results could be obtained using microwave acti- nothiazole 3 with labeling agent [18 F]SFB. [18 F]SFB has been
vation for heating. Microwave activation increased radiochemical widely used as versatile prosthetic group for the radiolabeling
yields for
[
18 F]fluoride incorporation into nitro-substituted of peptides and proteins under mild conditions.13–15 Only very
benzamides to 42% compared with 8% radiochemical yield few reports are known for the application of [18 F]SFB as building
using conventional heating at 150 ꢀC as exemplified for 18 F-labeled block for the radiosynthesis of nonpeptide small molecules via
5-HT1A agonist Org 13063.12
acylation reaction. A prominent example includes 4-[18 F]fluoro-
Table 1 summarizes the results of direct radiofluorination with benzoylated urea derivatives as imaging agents for prostate-specific
labeling precursors 5 and 7 using conventional heating at 150 membrane antigen.16
and 180 ꢀC.
The radiosynthesis of 18 F-labeled 2-aminothiazole [18 F]2 using
[
18 F]SFB was explored in different solvents with or without the
The results reported in Table 1 clearly demonstrate the chal-
lenge to prepare 4-[18F]fluorobenzamide [18F]2 through direct addition of an auxiliary base like Triethylamine (TEA), NaH, and
incorporation of n.c.a. [18F]fluoride into nitro-precursor 5 or NaOBut. All reactions were carried out at low activity levels of
trimethylammonium-precursor 7. The latter precursor hardly
[
18 F]SFB (20–30 MBq) during the optimization experiments. The
demonstrated any product formation, presumably because of reaction scheme is displayed in Figure 5 and the results are sum-
decomposition of compound 7 at elevated temperatures of 150– marized in Table 2.
180 ꢀC in different solvents (DMSO, DMF, NMP) (entries 7–12).
Acylation of amino group in 2-aminothiazole 3 with [18 F]SFB
Most activity was related to nonreacted [18F]fluoride along in the absence of an auxiliary base did not result in any product
with numerous unidentified side product representing 5–10% formation as indicated by radio-TLC analysis independent of the
of the total activity area using radio-thin layer chromatography used solvent (acetonitrile, THF or diethyl ether) (entries 1–3). The
(TLC) analysis of the reaction mixture. The use of nitro-precursor addition of amine base triethylamine (TEA, 25 mL) did also not
5 gave less side products, but product formation was also result in the formation of the desired product in all solvents
very small. Best results were achieved by the reaction of nitro- tested. However, the use of stronger bases like NaH (pKa = 37)
precursor 5 in DMSO at 180 ꢀC to afford 1%–3% of the desired and NaOt-Bu (pKa = 19) resulted in product formation employing
compound [18F]2 as indicated by radio-TLC analysis (entry 2). THF or diethyl ether as solvents (entries 7–10). Because of the
All other solvents (DMF, NMP) gave even lower yields (entries potential acidity of acetonitrile (pKa = 25), this solvent was not
1, 3–6).
tested when NaH and NaOt-Bu were used as auxiliary bases.
This finding is consistent with previously reported low radio- Highest and most consistent radiochemical yields (70–85) were
chemical yields on direct incorporation of n.c.a. [18F]fluoride achieved with NaH as the base (entries 7 and 8). Diethyl ether
into benzamides using conventional heating as exemplified for as the solvent gave slightly higher yields compared with THF
Figure 4. Direct radiofluorination to form 4-[18 F]fluorobenzamide [18 F]2.
J. Label Compd. Radiopharm 2011, 54 769–774
Copyright © 2011 John Wiley & Sons, Ltd.
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F. Svensson et al.
Table 1. Reaction conditions for direct radiofluorination using labeling precursors 5 and 7a
Entry
Precursor
Solvent
Reactiontemperature (ꢀC)
Yield (%)b
1
2
3
4
5
6
7
8
5
5
5
5
5
5
7
7
7
7
7
7
DMSO
DMSO
DMF
150
180
150
180
150
180
150
180
150
180
150
180
~1
1–3
<1
<1
~1
DMF
NMP
NMP
DMSO
DMSO
DMF
DMF
NMP
NMP
~1
<1
<1
<1
<1
<1
<1
9
10
11
12
DMSO, dimethyl sulfoxide; DMF, dimethylformamide; NMP, N-methyl-2-pyrrolidone.
aAll reactions were performed at least two times with 15 mg of labeling precursor (5 or 7) for 20 min at the indicated temperature.
bYield was determined by radio-TLC representing the percentage of radioactivity area of 18 F-labeled product related to the total
radioactivity area.
Figure 5. Synthesis of 4-[18 F]fluorobenzamide [18 F]2 through acylation reaction with [18 F]SFB.
Table 2. Reaction conditions for radiolabeling using [18 F]SFBa
Entry
Base
Solvent
Yield (%)b
1
2
No base
No base
Acetonitrile
THF
0
0
3
4
5
6
7
8
9
10
No base
Diethyl ether
Acetonitrile
THF
Diethyl ether
THF
Diethyl ether
THF
Diethyl ether
0
0
0
0
TEA (25 mL)
TEA (25 mL)
TEA (25 mL)
NaH (2–4 mg)
NaH (2–4 mg)
NaOt-Bu (2 mg)
NaOt-Bu (2 mg)
70–78
75–85
20–50
25–45
TEA, Triethanolamine.
aAll reactions were performed at least two times with 5 mg of 2-aminothiazole 3 as labeling precursor for 15 min at 60 ꢀC.
bYield was determined by radio-TLC representing the percentage of radioactivity area of 18 F-labeled product related to the total
radioactivity area.
as the solvent. The use of weaker base NaOt-Bu afforded
[
18 F]2 using higher activity levels of [18 F]SFB (0.8 - 1.7 GBq). This
product [18 F]2 in moderate but more inconsistent radiochem- also included HPLC purification of the final product. In a typical
ical yields of 20%–50% (entries 9 and 10). Based on these experiment, 1.4 GBq of [18 F]SFB could be converted into
findings, it seems to be obvious that primary amine group in 360 MBq of 18 F-labeled CDK-2 inhibitor [18 F]2 within 75 min
2-aminothiazoline 3 has to be deprotonated by strong bases like starting from [18 F]SFB. This represents a decay-corrected radio-
NaH and NaOt-Bu to allow successful acylation reaction with chemical yield of 39% based upon [18 F]SFB and a decay-
[
18 F]SFB.
corrected radiochemical yield of 14% based upon [18 F]fluoride.
Based on the promising results displayed in entries 7 and 8 of The radiochemical purity exceeded 95%, and the specific activity
Table 2, we set up the preparation of 4-[18 F]fluorobenzamide was determined to be greater 20 GBq/mmol.
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Copyright © 2011 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2011, 54 769–774

F. Svensson et al.
The stability of the radiolabeled compound was tested in 0.9% (s, 1H, CH), 7.52 (s, 1H, CH), 8.11 and 8.24(2d of AA’BB’ system,
saline at 1, 2, and 5 h. No decomposition was observed as indi- J = 9.0 Hz, 4H; Ar-H). LR-MS (ESI positive): m/z = 431.12 (M-OTf)+.
cated by radio-TLC analysis. Preliminary radiopharmacological
evaluation including small animal PET imaging of compound
Radiochemical syntheses
[18F]2 is currently in progress.
No-carrier-added aqueous [18F]fluoride was produced in an IBA
CYCLONE 18/9 cyclotron (IBA, Louvain-la-Neuve, Belgium) by
irradiation of [18O]H2O via the 18O(p,n)18F nuclear reaction. Reso-
lubilization of the aqueous [18 F]fluoride (300–700 MBq) was
accomplished as described by Coenen et al. with Kryptofix 2.2.2
and K2CO3 in a conical vial and azeotropical removal of water
Experimental
General
All commercial reagents and solvents were used without further
purification unless otherwise specified. 2-Aminothiazole 3 was
prepared according to literature procedures.5 Nuclear magnetic
resonance spectra were recorded on a Varian Unity 300 MHz
spectrometer. 1H-NMR chemical shifts were given in ppm and
were referenced with the residual solvent resonances relative to
tetramethylsilane. Mass spectra were obtained on a Quattro/LC
mass spectrometer (MICROMASS) by electrospray ionization.
Flash chromatography was conducted according to Still et al.17
using MERCK silica gel (Sigma–Aldrich Co., USA) (mesh size
230–400 ASTM). TLC was performed on Merck silica gel F-254
aluminum plates with visualization under UV (254 nm).
with acetonitrile in a stream of nitrogen.18 18 F]SFB was also pre-
[
pared according to literature procedure in a TRACERlab FxFDG
module (GE Medical Systems).19
Radio-HPLC analyses were carried out with a SUPELCO Discov-
ery C18 column (Sigma–Aldrich Co., USA) (4.6 Â 150 mm, 5 m)
using an isocratic elution with CH3CN/H2O (55:45) at a flow
rate of 1.0 ml/min. Semipreparative HPLC was performed using
a
PhenomenexLuna column (Phenomenex Inc., CA, USA)
(10.0 Â 250 mm, 10 m) using isocratic elution with CH3CN/H2O
(55:45) at a flow rate of 5.0 ml/min. The products were monitored
by UV detector L4500 (Merck, Hitachi) at 254 nm and by g-detection
with a scintillation detector GABI (RAYTEST). Radio-TLC was per-
formed on Merck silica gel F-254 aluminum plates. For radioac-
tivity detection, a BAS 2000 scanner (FUJIX) (Fuji photofilm,
Tokyo, Japan) was used.
Chemical syntheses
General procedure for the preparation of benzamides 2, 5,
and 6. 5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-amine
1 (120 mg, 0.45 mmol), 1-ethyl-3-(3-dimethyl-aminopropyl)carbo-
diimide hydrochloride (100 mg, 0.53 mmol), and substituted ben-
zoic acid 4 (0.66 mmol) were stirred in methylene chloride (5 mL)
for 2 h at room temperature under a nitrogen atmosphere. Satu-
rated NaHCO3 solution (50 mL) was added, and the mixture was
extracted with EtOAc. The organic layer was dried over Na2SO4,
and the solvent was removed under reduced pressure. The residue
was purified by flash chromatography (50% EtOAc/petroleum
ether) to afford desired compounds 3–5.
N-(5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-
4-fluorobenzamide 2. Yield: 78%. 1H-NMR (400 MHz, DMSOd6): d
1.19 (s, 9H, C(CH3)3), 3.93 (s, 2H, CH2), 6.58 (s, 1H, CH), 7.04 (s, 1H,
CH), 7.19 (t, J = 8.6 Hz, 2H), 7.92-7.98 (m, 2H). 19F-NMR (376 MHz,
DMSOd6): d -109.1 (m), (ESI positive): m/z = 392.02 (M + H).
N-(5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-
Direct radiofluorination with n.c.a. [18 F]fluoride
Labeling precursor 5 or 7 (15 mg) was dissolved in 0.5 mL of sol-
vent (DMSO, DMF, or NMP), and the mixture was added to a vial
containing dried n.c.a. [18 F]fluoride (20–70 MBq). The vial was
sealed and placed into an oil bath at a temperature of 150 or
180 ꢀC. After 20 min, aliquots were taken for radio-TLC analysis.
N-(5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-
4-[18 F]fluorobenzamide [18 F]2. Radio-TLC: Rf = 0.8; 95% aceto-
nitrile/water.
Radiolabeling with [18 F]SFB
An aliquot of module-produced [18 F]SFB solution in acetonitrile
(50 MBq) was transferred into a conical reaction vial, and the sol-
vent was evaporated in a gentle stream of nitrogen at 60–70 ꢀC.
2-Aminothiazole 3 (5 mg, 0.018 mmol) was dissolved in the indi-
cated solvent (0.5 mL), and base (TEA, NaH, NaOBut) was added if
applicable. The reaction vial was sealed and heated at 60 ꢀC for
15 min. Aliquots were taken for radio-TLC and radio-HPLC
analysis.
1
4-nitrobenzamide 5. Yield: 73%. H-NMR (400 MHz, DMSOd6): d
1.20 (s, 9H, C(CH3)3), 4.12 (s, 2H, CH2), 6.73 (s, 1H, CH), 7.53 (s,
1H, CH), 8.27 and 8.36 (2d of AA’BB’ system, J = 8.9 Hz, 4H; Ar-H).
LR-MS (ESI positive): m/z = 418.98 (M + H).
N-(5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-
4-(dimethylamino)benzamide 6. Yield: 80%. 1H-NMR (400 MHz,
DMSOd6): d 1.19 (s, 9H, C(CH3)3), 3.03 (s, 6H, N(CH3)2), 4.07 (s, 2H,
CH2), 6.72 (s, 1H, CH), 7.42 (s, 1H, CH), 6.74 and 7.97 (2d of AA’BB’
system, J = 9.1 Hz, 4H; Ar-H). LR-MS (ESI positive): m/z = 417.08
(M + H).
N-(5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-
4-[18 F]fluorobenzamide
[
18 F]2. Radio-TLC: Rf = 0.8; 95%
acetonitrile/water. Radio-HPLC-analysis: CH3CN/H2O (55/45),
tR = 7.3 min.
Large scale preparation of [18 F]fluorobenzamide [18 F]2
N-(5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-
4-(trimethylammonium)benzamide triflate 7. Dimethylamino
benzamide 6 (60 mg, 0.14 mmol) was dissolved in nitromethane Module-produced [18 F]SFB solution in acetonitrile (0.8–1.7 GBq)
(1.5 mL) and trifluoromethanesulfonic acid methyl ester (30 mL, was transferred into a conical reaction vial, and acetonitrile (ca.
0.273 mmol) was added. The mixture was stirred at room tem- 3 mL) was evaporated a gentle stream of nitrogen at 60–70 ꢀC.
perature for 4 h. The solvent was evaporated under reduced 2-Aminothiazole 3 (5 mg, 0.018 mmol) was dissolved in diethyl
pressure and the residue was washed with diethyl ether ether (0.5 mL) and 2–4 mg of NaH (60% in mineral oil) was added
(20 mL). The product was filtered off and dried to afford 75 mg with a spatula. After 15 min at 60 ꢀC eluent (1.5 mL, CH3CN/H2O
1
(90%) of the desired product 7. H-NMR (400 MHz, DMSOd6): d (55/45)) was added, and the reaction mixture was applied to a
1.13 (s, 9H, C(CH3)3), 2.83 (s, 9H, N(CH3)3), 4.11 (s, 2H, CH2), 7.02 semipreparative HPLC purification using isocratic elution with
J. Label Compd. Radiopharm 2011, 54 769–774
Copyright © 2011 John Wiley & Sons, Ltd.
www.jlcr.org

F. Svensson et al.
CH3CN/H2O (55/45) at a flow rate of 5 mL/min. The peak eluting
between 18–19 min was collected. Water (25 mL) was added to
the eluate, and the mixture was passed through a Water SEP-
PAK tC18 cartridge (Waters Corporation, MA, USA). The cartridge
was washed with water, and the product was eluted with 1 mL of
ethanol to give 36%–43% (decay-corrected radiochemical yield,
based upon [18 F]SFB) of radiotracer [18 F]2. In a typical experi-
ment, 1.4 GBq of [18 F]SFB were converted into 360 MBq of radio-
tracer [18 F]2 within 75 min, including HPLC purification.
References
[1] An HX, Beckmann MW, Reifenberger G, Bender HG, Niederacher D.
Am J Pathol 1999; 154: 113–138.
[2] Ortega S, Malumbres M, Barbacid M. Biochim Biophys Acta. 2002;
1602: 73–87.
[3] Wadler S. Drug Resist Updat. 2001; 4: 347–367.
[4] Shapiro GI. J Clin Oncol. 2006; 24: 1770–1783.
[5] Kim KS, Kimball SD, Misra RN, Rawlins DB, Hunt JT, Xiao HY, Lu S,
Qian L, Han WC, Shan W, Mitt T, Cai ZW, Poss MA, Zhu H, Sack JS,
Tokarski JS, Chang CY, Pavletich N, Kamath A, Humphreys WG,
Marathe P, Bursuker I, Kellar KA, Roongta U, Batorsky R, Mulheron
JG, Bol D, Fairchild CR, Lee FY, Webster KR. J Med Chem. 2002; 45:
3905–3927.
N-(5-(((5-(tert-Butyl)oxazol-2-yl)methyl)thio)thiazol-2-yl)-
4-[18 F]fluorobenzamide
[
18 F]2. Radio-TLC: Rf = 0.8; 95%
acetonitrile/water. Radio-HPLC-analysis: CH3CN/H2O (55/45),
tR = 7.3 min.
[6] Graf F, Mosch B, Koehler L, Bergmann R, Wuest F, Pietzsch J. Mini Rev
Med Chem. 2010; 10: 527–539.
[7] Koehler L, Graf F, Bergmann R, Steinbach J, Pietzsch J, Wuest F. Eur J
Med Chem. 2010; 45: 727–737.
[8] Graf F, Koehler L, Kniess T, Wuest F, Mosch B, Pietzsch J. J Oncol.
2009; 2009: 1–12.
[9] Hwang DR, Ngo K, Savenkova L, Huang Y, Guo N, Zhu Z, Laruelle M.
J Label Compd Radiopharm. 2001; 44: S179-S179.
[10] Plenevaux A, Lemaire C, Aerts J, Lacan G, Rubins D, Melega WP,
Brihaye C, Degueldre C, Fuchs S, Salmon E, Maquet P, Laureys S,
Damhaut P, Weissmann D, Le Bars D, Pujol JF, Luxen A. Nucl Med Biol.
2000; 27: 467–71.
[11] Vandecapelle M, Dumont F, De Vos F, Strijckmans K, Leysen D,
Audenaert K, Dierckx, Slegers G. J Label Compd Radiopharm. 2004;
47: 531–542.
[12] Richter S, Bergmann R, Pietzsch J, Ramenda T, Steinbach J, Wuest F.
Biopolymers. 2009; 92: 479–488.
[13] Vaidyanathan G, Zalutsky MR. Nat Protoc. 2006; 1: 1655–1661.
[14] Li ZB, Wu Z, Chen K, Ryu EK, Chen X. J Nucl Med. 2008; 49: 453–461.
[15] Chen Y, Foss CA, Byun Y, Nimmagadda S, Pullambhatla M, Fox JJ,
Castanares M, Lupold SE, Babich JW, Mease RC, Pomper MG. J Med
Chem. 2008; 51: 7933–7943.
[16] Still WC, Kahn M, Mitra A. J Org Chem. 1978; 43: 2923–2925.
[17] Coenen HH, Klatte B, Knöchel A, Schüller M, Stöcklin G, J Label
Compd Radiopharm. 1986; 23: 455–462.
Conclusion
The present work describes the radiosynthesis of 18 F-labeled
benzamide [18 F]2 as potential radiotracer for molecular imaging
of CDK-2 in cancer. To the best of our knowledge, this is the first
synthesis of 18 F-labeled CDK-2 inhibitor. Direct radiofluorination
with n.c.a. [18 F]fluoride gave only low radiochemical yields,
whereas acylation reaction with labeling agent [18 F]SFB afforded
desired compound in reasonable radiochemical yields suitable
for comprehensive subsequent radiopharmacological evalua-
tion. The described work further expands the scope of using
prosthetic group
[
18 F]SFB as efficient building block for
the synthesis of [18 F]fluorobenzoyl-containing small molecules
beyond peptides.
Acknowledgements
The authors wish to thank S. Preusche for radioisotope produc-
tion and T. Krauss for technical assistance.
[18] Mäding P, Füchtner F, Wuest F. Appl Radiat Isot. 2005; 63: 329–332.
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J. Label Compd. Radiopharm 2011, 54 769–774