Synthesis of radiolabeled stilbene derivatives as new potential PET probes for aryl hydrocarbon receptor in cancers

Article abstract of DOI:10.1016/j.bmcl.2006.08.088

New carbon-11 and fluorine-18 labeled stilbene derivatives, cis-3,5-dimethoxy-4′-[¹¹C]methoxystilbene (4′-[¹¹C]8a), cis-3,4′,5-trimethoxy-3′-[¹¹C]methoxystilbene (3′-[¹¹C]8b), trans-3,5-dimethoxy-4′-[¹¹C]methoxystilbene (4′-[¹¹C]10a), trans-3,4′,5-trimethoxy-3′-[¹¹C]methoxystilbene (3′-[¹¹C]10b), cis-3,5-dimethoxy-4′-[¹⁸F]fluorostilbene (4′-[¹⁸F]12a), and trans-3,5-dimethoxy-4′-[¹⁸F]fluorostilbene (4′-[¹⁸F]13a), were designed and synthesized as potential PET probes for aryl hydrocarbon receptor (AhR) in cancers.

Full text of DOI:10.1016/j.bmcl.2006.08.088

Bioorganic & Medicinal Chemistry Letters 16 (2006) 5767–5772
Synthesis of radiolabeled stilbene derivatives as new potential
PET probes for aryl hydrocarbon receptor in cancers
Mingzhang Gao,a Min Wang,a Kathy D. Miller,b George W. Sledge,b
Gary D. Hutchinsa and Qi-Huang Zhenga,*
aDepartment of Radiology, Indiana University School of Medicine, Indianapolis, IN 46202, USA
bDepartment of Medicine, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Received 31 July 2006; revised 16 August 2006; accepted 17 August 2006
Available online 6 September 2006
Abstract—New carbon-11 and fluorine-18 labeled stilbene derivatives, cis-3,5-dimethoxy-40-[11C]methoxystilbene (40-[11C]8a), cis-
3,40,5-trimethoxy-30-[11C]methoxystilbene (30-[11C]8b), trans-3,5-dimethoxy-40-[11C]methoxystilbene (40-[11C]10a), trans-3,40,5-tri-
methoxy-30-[11C]methoxystilbene (30-[11C]10b), cis-3,5-dimethoxy-40-[18F]fluorostilbene (40-[18F]12a), and trans-3,5-dimethoxy-40-
[18F]fluorostilbene (40-[18F]13a), were designed and synthesized as potential PET probes for aryl hydrocarbon receptor (AhR) in
cancers.
ꢀ 2006 Elsevier Ltd. All rights reserved.
Fluorine-18 labeled stilbene derivatives have been devel-
oped by Kung and coworkers as non-invasive biomarkers
for biomedical imaging technique positron emission
tomography (PET) to image brain amyloid in Alzheimer’s
disease.1,2 Stilbene derivatives exhibit a variety of useful
biological properties such as anti-leukemic, anti-bacte-
rial, anti-fungal, anti-platelet aggregation, and coronary
vasodilator activities.3–7 They also have strong anti-oxi-
dative and anti-inflammatory activities as potential can-
cer chemopreventive agents based on their striking
inhibitory effects on cellular events associated with cancer
initiation, promotion, and progression. Resveratrol,
3,40,5-trihydroxy-trans-stilbene, is a stilbene-based phy-
toalexin with multiple potencies in various pathologies
including anti-oxidant potency, estrogenic potency, and
antagonistic activity against the aryl hydrocarbon recep-
tor (AhR), and resveratrol has been identified as an AhR
mixedagonist/antagonist.3 Therefore, stilbene derivatives
of resveratrol are AhR mixed agonist/antagonists and
could serve as new selective aryl hydrocarbon modula-
tors. AhR is an intracellular, ligand-dependent, basic he-
lix–loop–helix/PAS (per-arnt-sim) transcription factor
and modulates the expression of various genes in a wide
range of tissues and species.3 AhR provides an attractive
target for the development of receptor-based PET cancer
imaging agents. Stilbene derivatives labeled with a posi-
tron emitting radionuclide carbon-11 or fluorine-18 may
enable non-invasive monitoring AhR expression in can-
cers and cancer response to AhR agonist/antagonist ther-
apy. Here, we report the design andsynthesis of carbon-11
and fluorine-18 stilbene derivatives as PET probes for
AhR in cancers.
The synthesis of cis- and trans-stilbene derivative refer-
ence standards and phenolic hydroxyl precursors for car-
bon-11 radiolabeling was performed using a modified
method of the literature procedures.1–7 The synthetic ap-
proach is outlined in Scheme 1. The commercially
available starting material, 1-(bromomethyl)-3,5-dimeth-
oxybenzene (1), was reacted with triphenylphosphine to
provide
3,5-dimethoxybenzyltriphenylphosphonium
bromide (2) in 90% yield. Starting materials 4-hydroxy-
benzaldehyde (3a) and 3-hydroxy-4-methoxybenzalde-
hyde (3b) were reacted with tert-butyldimethylsilyl
chloride to afford corresponding products, 4-(tert-butyl-
dimethylsilyloxy)benzaldehyde
(4a)
and
3-(tert-
butyldimethylsilyloxy)-4-methoxybenzaldehyde (4b), in
about 92% yield. The Wittig reactions of compounds 2
and 4a,b were carried out to give cis-3,5-dimethoxy-
40-tert-butyldimethylsilyloxystilbene (5a) and cis-3,40,5-
trimethoxy-30-tert-butyldimethylsilyloxystilbene (5b) in
30–50% yield, and trans-3,5-dimethoxy-40-tert-butyldim-
ethylsilyloxystilbene (6a) and trans-3,40,5-trimethoxy-
30-tert-butyldimethylsilyloxystilbene (6b) in 30–60%
Keywords: Stilbene derivatives; Carbon-11; Fluorine-18; Positron em-
ission tomography (PET); Aryl hydrocarbon receptor (AhR); Cancer.
*
Corresponding author. Tel.: +1 317 278 4671; fax: +1 317 278
9711; e-mail: qzheng@iupui.edu
0960-894X/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.08.088

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M. Gao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5767–5772
CH2Br
CH2PPh3Br
R3
PPh3, toluene
reflux
R3
R4
R4
CH3O
OCH3
CH3O
OCH3
n-BuLi, THF
1
2
-78oC
CHO
CHO
OCH3
CH3O
t-Bu(CH3)2SiCl
Et3N, DMF
R3
R1
OCH3
OCH3
R4
R2
5a, R3=H, R4=OTBDMS
6a, R3=H, R4=OTBDMS
6b, R3=OTBDMS, R4=OCH3
3a, R1=H, R2=OH
3b, R1=OH, R2=OCH3
4a, R3=H, R4=OTBDMS
4b, R3=OTBDMS, R4=OCH3
5b, R3=OTBDMS, R4=OCH3
R5
R1
R6
R2
CH3I, K2CO3
acetone
Bu4NF, THF
5a,5b
CH3O
CH3O
OCH3
OCH3
7a, R5=H, R6=OH
8a, R1=H, R2=OCH3
7b, R5=OH, R6=OCH3
8b, R1=OCH3, R2=OCH3
R5
R1
R6
R2
CH3I, K2CO3
acetone
Bu4NF, THF
6a,6b
OCH3
OCH3
OCH3
9a, R5=H, R6=OH
9b, R5=OH, R6=OCH3
OCH3
10a, R1=H, R2=OCH3
10b, R1=OCH3, R2=OCH3
Scheme 1. Synthesis of cis- and trans-stilbene derivatives for carbon-11 radiolabeling.
yield, respectively. cis- and trans-Stilbene derivatives were
separable by column chromatography. The deprotection
reaction of silyloxy-protected stilbenes (5a,b and 6a,b)
using tetrabutylammonium fluoride gave cis-3,5-dime-
thoxy-40-hydroxystilbene (7a) and cis-3,40,5-trimethoxy-
30-hydroxystilbene (7b), and trans-3,5-dimethoxy-40-
hydroxystilbene (9a) and trans-3,40,5-trimethoxy-30-
hydroxystilbene (9b), in about 90% yield, as precursors
for radiolabeling. The methylation reaction of com-
pounds 7a,b and 9a,b using methyl iodide produced
cis-3,40,5-trimethoxystilbene (8a) and cis-3,30,40,5-tetra-
methoxystilbene (8b), and trans-3,40,5-trimethoxystil-
bene (10a) and trans-3,30,40,5-tetramethoxystilbene
(10b), in about 92% yield, as reference standards.
The synthesis of cis- and trans-stilbene derivative refer-
ence standards and nitro precursors for fluorine-18
radiolabeling was performed using a modified method
of the literature procedures.1–7 The synthetic approach
is outlined in Scheme 2. The Wittig reactions of
compounds 2 with 4-fluorobenzaldehyde (11a) and
4-nitrobenzaldehyde (11b) were carried out to give
R
R
CHO
CH2PPh3Br
n-BuLi, THF
-78oC
CH3O
OCH3
CH3O
OCH3
R
2
11a, R=F
OCH3
12a, R=F
12b, R=NO2
OCH3
13a, R=F
13b, R=NO2
11b, R=NO2
Scheme 2. Synthesis of cis- and trans-stilbene derivatives for fluorine-18 radiolabeling.

M. Gao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5767–5772
5769
corresponding cis-3,5-dimethoxy-40-fluorostilbene (12a)
18F
and trans-3,5-dimethoxy-40-fluorostilbene (13a) as refer-
ence standards, and cis-3,5-dimethoxy-40-nitrostilbene
(12b) and trans-3,5-dimethoxy-40-nitrostilbene (13b) as
radiolabeling precursors, in 30–60% yield. Likewise,
cis- and trans-stilbene derivatives were separated by
column chromatography.
K[18F]F/K2.2.2
CH3CN
CH3O
12b
OCH3
4'-[18F]12a
Compounds 8a,b, 10a,b, 12a, and 13a are AhR antago-
nists with high receptor binding activity, Ki (AhR, nM)
75 3.2, 7.7 0.2, 96 3.4, and 3.1 0.8 for com-
pounds 8a, 10a, 12a, and 13a, respectively,3 and nano-
molar IC50 anti-tumor activity for compounds 8b and
10b.6,8
18F
K[18F]F/K2.2.2
CH3CN
OCH3
13b
Synthesis of target radiotracers carbon-11 stilbenes is
shown in Scheme 3. The phenolic hydroxyl precursors
(7a,b and 9a,b) were labeled with [11C]methyl triflate
(11CH3OTf)9 under basic conditions through
O-[11C]methylation and isolated by solid-phase extrac-
tion (SPE) purification procedure using a C18 Sep-Pak
cartridge10 to give corresponding carbon-11 stilbene
derivatives, cis-3,5-dimethoxy-40-[11C]methoxystilbene
(40-[11C]8a) and cis-3,40,5-trimethoxy-30-[11C]methoxy-
stilbene (30-[11C]8b), and trans-3,5-dimethoxy-40-
[11C]methoxystilbene (40-[11C]10a) and trans-3,40,5-tri-
4'-[18F]13a
OCH3
Scheme 4. Synthesis of fluorine-18 labeled stilbene derivatives.
to afford corresponding fluorine-18 stilbene derivatives,
cis-3,5-dimethoxy-40-[18F]fluorostilbene (40-[18F]12a)
(40-
and
trans-3,5-dimethoxy-40-[18F]fluorostilbene
[18F]13a), in 15–20% radiochemical yield at EOB. The
specific activity was 1.0–1.2 Ci/lmol at EOS.
methoxy-30-[11C]methoxystilbene
(30-[11C]10b),
in
30–40% radiochemical yields based on [11C]CO2,
15–20 min overall synthesis time from end of bom-
bardment (EOB), >95% radiochemical purity, and
1.0–2.0 Ci/lmol specific activity at end of synthesis
(EOS) measured by analytical HPLC method.11
The experimental details and characterization data for
compounds 2, 4a,b, 5a,b, 6a,b, 7a,b, 8a,b, 9a,b, 10a,b,
12a,b, and 13a,b, and new tracers 40-[11C]8a, 30-
[11C]8b, 40-[11C]10a, 30-[11C]10b, 40-[18F]12a, and 40-
[18F]13a are given.13
In summary, an efficient and convenient chemical and
radiochemical synthesis of the precursors, reference
standards, and target tracers has been well developed.
The synthetic methodology of carbon-11 and fluorine-
18 labeled cis- and trans-stilbene derivatives employed
Synthesis of target radiotracers fluorine-18 stilbenes is
shown in Scheme 4. The nitro precursors (12b and
13b) were labeled by a conventional nucleophilic substi-
tution with K18F/Kryptofix 2.2.2 in acetonitrile at
120 ꢁC for 15–20 min and purified by HPLC method12
O11CH3
11CH3O
CH3O
11CH3OTf, 3 N NaOH
7a
7b
CH3CN
CH3O
CH3O
OCH3
4'-[11C]8a
OCH3
3'-[11C]8b
O
11CH3
11CH3O
CH3O
OCH3
11CH3OTf, 3 N NaOH
CH3CN
9a
9b
OCH3
OCH3
OCH3
4'-[11C]10a
3'-[11C]10b
Scheme 3. Synthesis of carbon-11 labeled stilbene derivatives.

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M. Gao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5767–5772
The 11CH3OTf was made according to a literature
procedure.8 Melting points were determined on a MEL-
TEMP II apparatus and are uncorrected. 1H NMR
spectra were recorded on a Bruker QE 300 FT-NMR
spectrometer using tetramethylsilane (TMS) as an internal
standard. Chemical shift data for the proton resonances
were reported in parts per million (ppm, d scale) relative to
internal standard TMS (d 0.0), and coupling constants (J)
are reported in hertz (Hz). Chromatographic solvent
proportions are expressed on a volume: volume basis.
Thin-layer chromatography was run using Analtech silica
gel GF uniplates (5 · 10 cm2). Plates were visualized by
UV light. Normal phase flash chromatography was carried
out on EM Science silica gel 60 (230–400 mesh) with a
forced flow of the indicated solvent system in the
proportions described below. All moisture- and/or air-
sensitive reactions were performed under a positive
pressure of nitrogen maintained by a direct line from a
nitrogen source. Analytical HPLC was performed using a
Prodigy (Phenomenex) 5 lm C18 column, 4.6 ·ꢀ250 mm;
3:1:1 CH3CN/MeOH 20 mM, pH 6.7, KHPO4 (buffer
solution) mobile phase, flow rate 1.5 mL/min, and UV
(254 nm) and c-ray (NaI) flow detectors. Semi-preparative
HPLC was performed using a Prodigy (Phenomenex)
5 lm C-18 column, 10 · 250 mm; 3:1:1 CH3CN/MeOH
readily available benzyl bromides and benzyl aldehydes,
featuring a stereo-divergent Wittig olefination, O-meth-
ylation with 11CH3OTf, and nucleophilic aromatic sub-
stitution with K18F/Kryptofix 2.2.2 under phase
transfer catalysis. These reactions are mostly high yield,
and the resulting stilbene derivatives were shown to have
excellent radiochemical yields, and thus, given that they
could serve as potential AhR antagonists, these agents
should be useful as potential PET probes for imaging
AhR in tumors. The chemistry result with reported in vi-
tro binding data provides the foundation for further
in vivo biological evaluation of carbon-11 and fluo-
rine-18 labeled stilbene derivatives as new potential
PET cancer AhR imaging agents.
Acknowledgments
This work was partially supported by the Susan G. Ko-
men Breast Cancer Foundation, Breast Cancer Research
Foundation, NIH/NINDS 5P50NS052606 and Indiana
Genomics Initiative (INGEN) of Indiana University,
which is supported in part by Lilly Endowment Inc.
The authors thank Dr. Bruce H. Mock, Barbara E.
Glick-Wilson, and Michael L. Sullivan for their assis-
tance in production of 11CH3OTf and K18F/Kryptofix
2.2.2. The referees’ criticisms and editor’s comments
for the revision of the manuscript are greatly
appreciated.
ꢀ
20 mM, pH 6.7, KHPO4 mobile phase, 5.0 mL/min flow
rate, UV (254 nm) and c-ray (NaI) flow detectors. Semi-
prep C18 silica guard cartridge column 1 · 1 cm was
obtained from E. S. Industries, Berlin, NJ, and part
number 300121-C18-BD 10 l. Semi-prep SiO2 Sep-Pak
type cartridge was obtained from Waters Corporate
Headquarters, Milford, MA. Sterile vented Millex-GS
0.22 lm filter unit was obtained from Millipore Corpora-
tion, Bedford, MA; (b) Compound 2. Compound 1
(11.6 g, 5.0 mmol) in toluene (180 mL) was added triphe-
nylphosphine (13.2 g, 5.0 mmol). The mixture was heated
at reflux for 14 h, and then the mixture was cooled down
to room temperature. The resulting precipitate was
filtered, washed with toluene, and recrystallized from
ethanol to give a pure product 2 as a colorless solid
References and notes
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Kung, H. F. Nucl. Med. Biol. 2005, 32, 799.
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Carboni, I.; Placanica, G.; Prota, G.; Rapposelli, S.;
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48, 6783.
6. Roberti, M.; Pizzirani, D.; Simoni, D.; Rondanin,
R.; Baruchello, R.; Bonora, C.; Buscemi, F.; Gri-
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9. Mock, B. H.; Mulholland, G. K.; Vavrek, M. T. Nucl.
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1
(22.3 g, 90% yield). Mp 273–274 ꢁC. H NMR (300 MHz,
DMSO-d6): d 3.50 (s, 6H, 2 · CH3), 5.10 (t, J = 12.9 Hz,
2H, CH2), 6.12 (dd, J = 2.5, 4.0 Hz, 2H, Ph-H), 6.42 (d,
J = 2.2 Hz, 1H, Ph-H), 7.65–7.81 (m, 12H, Ph-H), 7.92 (t,
J = 7.0 Hz, 3H, Ph-H); (c) General procedure for synthesis
of compounds 4a,b. To a solution of compound 3a or 3b
(10 mmol) in DMF (120 mL) was added triethylamine
(12.1 g, 12 mmol). Then tert-butyldimethylsilyl chloride
(16.5 g, 11 mmol) in DMF (50 mL) was added slowly, and
the reaction solution was stirred for overnight at room
temperature. The reaction mixture was poured into water
and extracted with ethyl acetate for three times, washed
with brine for two times, and dried over MgSO4. The
solvent was removed under vacuum, and the crude
product was purified by flash chromatography (1:19–1:4
EtOAc/hexanes) to give oily pure compound 4a or 4b in
about 92% yield. Compound 4a. Rf = 0.78 (1:6 EtOAc/
1
hexanes). H NMR (300 MHz, CDCl3): d 0.25 (s, 6H, Si–
CH3), 0.98 (s, 9H, C–CH3), 6.93 (d, J = 8.5 Hz, 2H, Ph-
H), 7.77 (d, J = 8.5 Hz, 2H, Ph-H), 9.88 (s, 1H, CHO).
Compound 4b. Rf = 0.60 (1:4, EtOAc/hexane). H NMR
1
10. Gao, M.; Miller, K. D.; Sledge, G. W.; Zheng, Q.-H.
Bioorg. Med. Chem. Lett. 2005, 15, 3865.
11. Zheng, Q.-H.; Mock, B. H. Biomed. Chromatogr. 2005, 19,
671.
12. Wang, J.-Q.; Miller, K. D.; Sledge, G. W.; Zheng, Q.-H.
Bioorg. Med. Chem. Lett. 2005, 15, 4380.
13. (a) Experimental details and characterization data. Gen-
eral: all commercial reagents and solvents were used
without further purification unless otherwise specified.
(300 MHz, CDCl3): d 0.17 (s, 6H, Si–CH3), 1.00 (s, 9H, C–
CH3), 3.89 (s, 3H, OCH3), 6.94 (d, J = 8.5 Hz, 1H, Ph-H),
7.36 (d, J = 2.2 Hz, 1H, Ph-H), 7.46 (d, J = 8.5 Hz, 1H,
Ph-H), 9.82 (s, 1H, CHO); (d) General procedure for
synthesis of compounds 5a,b, 6a,b, 12a,b, and 13a,b. To
the compound 2 (2.46 g, 5.0 mmol) in anhydrous THF
(60 mL) at ꢀ78 ꢁC was added n-butyllithium (2.1 mL,
2.5 M, 5.25 mmol), and the resulting red solution was

M. Gao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5767–5772
5771
stirred for 3 h. A solution of the aldehyde (5.0 mmol,
1.0 equiv) in THF was added dropwise over 30 min. The
reaction temperature was allowed to slowly rise to room
temperature, and the mixture was stirred for another 5 h.
The resulting cream suspension was poured into water and
extracted with dichloromethane for three times. The
organic phase was washed brine, dried over MgSO4, and
the solvent was removed under vacuum to afford crude
product, which was separated and purified by flash
chromatography (1:200–1:20 EtOAc/hexanes) to give two
pure cis- and trans- products. The cis-stilbene eluted first
in 30–50% yield, followed by the trans-isomer in 30–60%
1H NMR (300 MHz, CDCl3): d 3.66 (s, 6H, OCH3), 5.10
(s, 1H, OH), 6.32 (t, J = 2.2 Hz, 1H, Ph-H), 6.43 (d,
J = 2.2 Hz, 2H, Ph-H), 6.41 (d, J = 12.5 Hz, 1H, CH@C),
6.49 (d, J = 12.5 Hz, 1H, C@CH), 6.66 (dd, J = 2.2,
6.6 Hz, 2H, Ph-H), 7.13 (d, J = 8.8 Hz, 2H, Ph-H).
Compound 9a. Mp 87–88 ꢁC, Rf = 0.68 (1:1 EtOAc/
hexanes). 1H NMR (300 MHz, CDCl3): d 3.82 (s, 6H,
OCH3), 5.05 (s, 1H, OH), 6.38 (t, J = 2.2 Hz, 1H, Ph-H),
6.65 (d, J = 2.2 Hz, 2H, Ph-H), 6.81 (dd, J = 2.2, 6.62 Hz,
2H, Ph-H), 6.86 (d, J = 16.5 Hz, 1H, CH@C), 6.99 (d,
J = 16.5 Hz, 1H, C@CH), 7.38 (d, J = 8.8 Hz, 2H, Ph-H).
1
Compound 7b. Rf = 0.46 (1:4 EtOAc/hexanes). H NMR
1
yield. Compound 5a. Rf = 0.52 (1:19 EtOAc/hexanes). H
(300 MHz, CDCl3): d 3.66 (s, 6H, OCH3), 3.84 (s, 3H,
OCH3), 5.56 (s, 1H, OH), 6.31 (t, J = 2.2 Hz, 1H, Ph-H),
6.40–6.50 (m, 4H, CH@CH and Ph-H), 6.68 (d,
J = 8.8 Hz, 1H, Ph-H), 6.76 (dd, J = 2.2, 8.80 Hz, 1H,
Ph-H), 6.87 (d, J = 2.2, 1H, Ph-H). Compound 9b. Mp 89–
90 ꢁC, Rf = 0.42 (1:4 EtOAc/hexanes). 1H NMR
(300 MHz, CDCl3): d 3.82 (s, 6H, OCH3), 3.90 (s, 3H,
OCH3), 5.62 (s, 1H, OH), 6.37 (t, J = 2.2 Hz, 1H, Ph-H),
6.64 (d, J = 2.2 Hz, 2H, Ph-H), 6.81–6.91 (m, 2H), 6.96–
7.01 (m, 2H), 7.13 (d, J = 2.2, 1H, Ph-H); (f) General
procedure for synthesis of compounds 8a,b and 10a,b. To
a solution of compounds 7a, 7b, 9a or 9b (1 equiv) in
acetone (30 mL), potassium carbonate (1.2 equiv) and
iodomethane (1.5 equiv) were added. The reaction mixture
was heated at reflux for 5 h, and then the mixture was
cooled down to room temperature and filtered. The
organic phase was evaporated under vacuum, and the
residue was purified by flash chromatography to obtain
pure products 8a, 8b, 10a or 10b in about 92% yield.
NMR (300 MHz, CDCl3): d 0.16 (s, 6H, Si–CH3), 0.94 (s,
9H, C–CH3), 3.65 (s, 6H, OCH3), 6.31 (t, J = 2.2 Hz, 1H,
Ph-H), 6.41 (d, J = 6.1 Hz, 1H, CH@C), 6.50 (d,
J = 6.1 Hz, 1H, C@CH), 6.4 (d, J = 2.2 Hz, 2H, Ph-H),
6.99 (dd, J = 2.0, 6.6 Hz, 2H, Ph-H), 7.13 (d, J = 8.8 Hz,
2H, Ph-H). Compound 6a. Rf = 0.38 (1:19 EtOAc/hex-
anes). 1H NMR (300 MHz, CDCl3): d 0.21 (s, 6H, Si–
CH3), 0.99 (s, 9H, C–CH3), 3.82 (s, 6H, OCH3), 6.37 (t,
J = 2.2 Hz, 1H, Ph-H), 6.64 (d, J = 2.2 Hz, 2H, Ph-H),
6.81 (d, J = 8.8 Hz, 2H, Ph-H), 6.86 (d, J = 16.2 Hz, 1H,
CH@C), 6.92 (d, J = 16.2 Hz, 1H, C@CH), 7.36 (d,
J = 8.8, 2H, Ph-H). Compound 5b. Rf = 0.32 (1:19
1
EtOAc/hexanes). H NMR (300 MHz, CDCl3): d 0.05 (s,
6H, Si–CH3), 0.92 (s, 9H, C–CH3), 3.67 (s, 6H, OCH3),
3.77 (s, 3H, OCH3), 6.30 (t, J = 2.2 Hz, 1H, Ph-H), 6.41 (d,
J = 2.2 Hz, 2H, Ph-H), 6.45 (d, J = 3.0 Hz, 2H, Ph-H),
6.70 (d, J = 8.8 Hz, 1H, CH@C), 6.75 (d, J = 2.2 Hz, 1H,
Ph-H), 6.81 (dd, J = 2.2, 8.8 Hz, 1H, C@CH). Compound
6b. Yellow solid, mp 53–54 ꢁC, Rf = 0.24 (1:19 EtOAc/
1
Compound 8a. Rf = 0.74 (1:3 EtOAc/hexanes). H NMR
1
hexanes). H NMR (300 MHz, CDCl3): d 0.18 (s, 6H, Si–
(300 MHz, CDCl3): d 3.65 (s, 6H, OCH3), 3.76 (s, 3H,
OCH3), 6.31 (t, J = 2.2 Hz, 1H, Ph-H), 6.43 (d, J = 2.2 Hz,
2H, Ph-H), 6.41 (d, J = 11.8 Hz, 1H, CH@C), 6.49 (d,
J = 11.8 Hz, 1H, C@CH), 6.74 (d, J = 8.8 Hz, 2H, Ph-H),
7.19 (d, J = 8.8 Hz, 2H, Ph-H). Compound 10a. Mp 57 ꢁC,
Rf = 0.68 (1:3 EtOAc/hexanes). 1H NMR (300 MHz,
CDCl3): d 3.83 (s, 9H, OCH3), 6.37 (t, J = 2.2 Hz, 1H,
Ph-H), 6.65 (d, J = 2.2 Hz, 2H, Ph-H), 6.87 (d, J = 8.8 Hz,
2H, Ph-H), 6.87 (d, J = 16.2 Hz, 1H, CH@C), 7.01 (d,
J = 16.2 Hz, 1H, C@CH), 7.42 (d, J = 8.8 Hz, 2H, Ph-H).
CH3), 1.02 (s, 9H, C–CH3), 3.81 (s, 3H, OCH3), 3.82 (s,
6H, OCH3), 6.37 (t, J = 2.2 Hz, 1H, Ph-H), 6.64 (d,
J = 2.2 Hz, 2H, Ph-H), 6.81–6.87 (m, 2H, Ph-H and
CH@C), 7.00–7.07 (m, 3H, Ph-H and C@CH). Compound
12a. Rf = 0.48 (1:19 EtOAc/hexanes). 1H NMR (300 MHz,
CDCl3): d 3.65 (s, 6H, OCH3), 6.32 (t, J = 2.2 Hz, 1H, Ph-
H), 6.37 (d, J = 2.2 Hz, 2H, Ph-H), 6.48 (d, J = 12.5 Hz,
1H, CH@C), 6.53 (d, J = 12.5, 1H, C@CH), 6.91 (t,
J = 8.8 Hz, 2H, Ph-H), 7.20–7.25 (m, 2H, Ph-H). Com-
pound 13a. Colorless solid, mp 42–44 ꢁC, Rf = 0.34 (1:19
1
Compound 8b. Rf = 0.56 (1:4 EtOAc/hexanes). H NMR
1
EtOAc/hexanes). H NMR (300 MHz, CDCl3): d 3.83 (s,
(300 MHz, CDCl3): d 3.65 (s, 3H, OCH3), 3.67 (s, 6H,
OCH3), 3.85 (s, 3H, OCH3), 6.32 (t, J = 2.2 Hz, 1H, Ph-
H), 6.45 (d, J = 2.2 Hz, 2H, Ph-H), 6.48 (d, J = 11.1 Hz,
1H, CH@C), 6.50 (d, J = 11.1 Hz, 1H, C@CH), 6.73 (d,
J = 8.8 Hz, 1H, Ph-H), 6.83 (d, J = 8.8 Hz, 2H, Ph-H).
Compound 10b. Rf = 0.50 (1:4 EtOAc/hexanes). 1H NMR
(300 MHz, CDCl3): d 3.82 (s, 6H, OCH3), 3.89 (s, 3H,
OCH3), 3.94 (s, 3H, OCH3), 6.37 (t, J = 2.2 Hz, 1H, Ph-
H), 6.65 (d, J = 2.2 Hz, 2H, Ph-H), 6.84 (d, J = 8.1 Hz,
1H, Ph-H), 6.92 (d, J = 17.6 Hz, 1H, CH@C), 7.00–7.06
(m, 3H, C@CH and Ph-H); (g) Typical experimental
procedure for the radiosynthesis of C-11 tracer 40-[11C]8a,
30-[11C]8b, 40-[11C]10a, or 30-[11C]10b. The precursor (7a,
7b, 9a or 9b) (0.3–0.5 mg) was dissolved in CH3CN
(300 lL). To this solution was added 3 N NaOH (2–3 lL).
The mixture was transferred to a small volume, three-
necked reaction tube. 11CH3OTf was passed into the air-
cooled reaction tube at ꢀ15 to ꢀ20 ꢁC, which was
generated by a Venturi cooling device powered with
100 psi compressed air, until radioactivity reached a
maximum (ꢁ3 min), then the reaction tube was heated
at 70–80 ꢁC for 3 min. The contents of the reaction tube
were diluted with NaHCO3 (1 mL, 0.1 M). This solution
was passed onto a C18 cartridge by gas pressure. The
cartridge was washed with H2O (2·3 mL), and the
aqueous washing was discarded. The product was eluted
6H, OCH3), 6.40 (t, J = 2.2 Hz, 1H, Ph-H), 6.65 (d,
J = 2.2 Hz, 2H, Ph-H), 6.91 (d, J = 16.2 Hz, 1H, CH@C),
6.96–7.07 (m, 3H, C@CH and Ph-H), 7.44–7.49 (m, 2H,
Ph-H). Compound 12b. Yellow solid, mp 71–72 ꢁC,
Rf = 0.22 (1:19 EtOAc/hexanes). 1H NMR (300 MHz,
CDCl3): d 3.84 (s, 6H, OCH3), 6.46 (t, J = 2.2 Hz, 1H, Ph-
H), 6.69 (d, J = 2.2 Hz, 2H, Ph-H), 7.08 (d, J = 16.2 Hz,
1H, CH@C), 7.17 (d, J = 16.2 Hz,1H, C@CH), 7.61 (d,
J = 8.8 Hz, 2H, Ph-H), 8.20 (d, J = 8.8 Hz, 2H, Ph-H).
Compound 13b. Yellow solid, mp 134–135 ꢁC, Rf = 0.15
(1:19 EtOAc/hexanes). 1H NMR (300 MHz, CDCl3): d
3.67 (s, 6H, OCH3), 6.34 (d, J = 2.2 Hz, 2H, Ph-H), 6.36 (t,
J = 2.2 Hz, 1H, Ph-H), 6.58 (d, J = 12.1 Hz, 1H, CH@C),
6.73 (d, J = 12.1 Hz, 1H, C@CH), 7.38 (d, J = 8.8 Hz, 2H,
Ph-H), 8.07 (d, J = 8.8 Hz, 2H, Ph-H); (e) General
procedure for synthesis of compounds 7a,b and 9a,b. To
a solution of compound 5a, 5b, 6a or 6b (1 equiv) in
anhydrous THF (20 mL), tetrabutylammonium fluoride
(1 M in THF, 3 equiv) was added. The yellow solution was
stirred for 1 h at room temperature. Then the reaction
mixture was poured into water, extracted with dichloro-
methane. Solvent was removed under vacuum to afford
the crude product, which was purified by flash chroma-
tography to give pure product 7a, 7b, 9a or 9b in about
90% yield. Compound 7a. Rf = 0.74 (1:1 EtOAc/hexanes).

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M. Gao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5767–5772
from the column with EtOH (2·3 mL) and then passed
13a = 5.79 min, tR 40-[18F]13a = 5.79 min; and tR
K18F = 1.88 min. The reaction mixture was sealed and
heated at 120 ꢁC for 15–20 min and was subsequently
allowed to cool down, at which time the crude product
was passed through a semi-prep SiO2 Sep-Pak cartridge
to remove Kryptofix 2.2.2 and unreacted K18F. The
Sep-Pak column was eluted with 15% MeOH/CH2Cl2
(5.0 mL), and the fractions were passed onto a rotatory
evaporator. The solvent was removed by evaporation
under high vacuum (0.1–1.0 mmHg) to give a crude
product 40-[18F]12a or 40-[18F]13a. The mixture contain-
ing precursor and product was purified with semi-
preparative HPLC method. The contents of the mixture
residue were diluted with HPLC mobile phase 3:1:1
CH3CN/MeOH 20 mM, pH 6.7, KHPO4ꢀ, and injected
onto the semi-preparative HPLC column with 2 mL
injection loop. The product fraction was collected, the
solvent was removed by rotatory evaporation under
vacuum, and the final product 40-[18F]12a or 40-[18F]13a
was formulated in saline, sterile-filtered through a sterile
vented Millex-GS 0.22 lm cellulose acetate membrane,
and collected into a sterile vial. Total radioactivity was
assayed and total volume was noted. The overall
synthesis, purification, and formulation time was
60–70 min from EOB. Retention times in the semi-
preparative HPLC system were: tR 12b = 7.28 min, tR
12a = 9.89 min, tR 40-[18F]12a = 9.89 min; and tR 13b =
7.76 min, tR 13a = 9.35 min, tR 40-[18F]13a = 9.35 min.
The radiochemical yield of 40-[18F]12a or 40-[18F]13a was
15–20%. Chemical purity, radiochemical purity, and
specific radioactivity were determined by analytical
HPLC method. The chemical purities of precursors
12b and 13b, and standard samples 12a and 13a were
>96%, the radiochemical purity of target radiotracer
40-[18F]12a or 40-[18F]13a was >99%, and the chemical
purity of radiotracer 40-[18F]12a or 40-[18F]13a
was >94%.
onto a rotatory evaporator. The solvent was removed by
evaporation under high vacuum. The labeled product 40-
[11C]8a, 30-[11C]8b, 40-[11C]10a, or 30-[11C]10b was formu-
lated with saline, whose volume was dependent upon the
use of the labeled product in tissue biodistribution studies
(ꢁ6 mL, 3·2 mL) or in micro-PET imaging studies (1–
3 mL), sterile-filtered through a sterile vented Millex-GS
0.22 lm cellulose acetate membrane, and collected into a
sterile vial. Total radioactivity was assayed and total
volume was noted. The overall synthesis time was 15–
20 min. The decay corrected radiochemical yield, from
11CO2, was 30–40%, and the radiochemical purity was
>95% by analytical HPLC. Retention times in the
analytical HPLC system were: tR 7a = 3.50 min, tR
8a = 5.55 min, tR 40-[11C]8a = 5.55 min; tR 7b = 3.66 min,
tR 8b = 4.47 min, tR 30-[11C]8b = 4.47 min; tR 9a =
3.17 min, tR 10a = 5.39 min, tR 40-[11C]10a = 5.39 min;
and tR 9b = 3.28 min, tR 10b = 4.22 min, tR 30-
[11C]10b = 4.22 min. The chemical purities of the target
tracers 40-[11C]8a, 30-[11C]8b, 40-[11C]10a and 30-[11C]10b
were >93%; (h) Typical experimental procedure for the
radiosynthesis of F-18 tracer 40-[18F]12a or 40-[18F]13a.
No-carrier-added (NCA) aqueous H18F (0.5 mL) prepared
by 18O(p,n)18F nuclear reaction in a RDS-112 cyclotron
on an enriched H218O water (95+%) target was added to a
Pyrex vessel which contains K2CO3 (4 mg, in 0.2 mL H2O)
and Kryptofix 2.2.2 (10 mg, in 0.5 mL CH3CN). Azeotro-
pic distillation at 115 ꢁC with HPLC grade CH3CN
(3·1 mL) under a nitrogen steam efficiently removed
water to form anhydrous K18F-Kryptofix 2.2.2 complex.
The nitro-precursor 12b or 13b (2–3 mg, dissolved in
0.5 mL CH3CN) was introduced to the K18F-Kryptofix
2.2.2 complex. The radiolabeling reaction was monitored
by analytical radio-HPLC method. Retention times in the
analytical HPLC system were: tR 12b = 5.18 min, tR 12a =
6.09 min, tR 40-[18F]12a = 6.09 min; tR 13b = 5.31 min, tR