Synthesis of a new carbon-11-labeled sulfamate derivative as a potential PET tracer for imaging of breast cancer aromatase and steroid sulfatase expression

Article abstract of DOI:10.1080/00397911003797825

A carbon-11-labeled sulfamate derivative was designed and synthesized as a new potential positron-emission-tomography dual aromatase-steroid sulfatase inhibitor radiotracer for imaging of aromatase and steroid sulfatase expression in breast cancer. The ta

Full text of DOI:10.1080/00397911003797825

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Synthesis of a New Carbon-11–Labeled
Sulfamate Derivative as a Potential PET
Tracer for Imaging of Breast Cancer
Aromatase and Steroid Sulfatase
Expression
Min Wang a , Mingzhang Gao a , Kathy D. Miller b & Qi-Huang Zheng a
a Department of Radiology and Imaging Sciences, Indiana University
School of Medicine, Indianapolis, Indiana, USA
b Department of Medicine, Indiana University School of Medicine,
Indianapolis, Indiana, USA
Published online: 21 Mar 2011.
To cite this article: Min Wang , Mingzhang Gao , Kathy D. Miller & Qi-Huang Zheng (2011)
Synthesis of a New Carbon-11–Labeled Sulfamate Derivative as a Potential PET Tracer for Imaging
of Breast Cancer Aromatase and Steroid Sulfatase Expression, Synthetic Communications: An
International Journal for Rapid Communication of Synthetic Organic Chemistry, 41:8, 1127-1140, DOI:
10.1080/00397911003797825
To link to this article: http://dx.doi.org/10.1080/00397911003797825
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Synthetic Communications1, 41: 1127–1140, 2011
Copyright # Taylor & Francis Group, LLC
ISSN: 0039-7911 print=1532-2432 online
DOI: 10.1080/00397911003797825
SYNTHESIS OF A NEW CARBON-11–LABELED
SULFAMATE DERIVATIVE AS A POTENTIAL PET
TRACER FOR IMAGING OF BREAST CANCER
AROMATASE AND STEROID SULFATASE EXPRESSION
Min Wang,1 Mingzhang Gao,1 Kathy D. Miller,2 and
Qi-Huang Zheng1
1Department of Radiology and Imaging Sciences, Indiana University
School of Medicine, Indianapolis, Indiana, USA
2Department of Medicine, Indiana University School of Medicine,
Indianapolis, Indiana, USA
GRAPHICAL ABSTRACT
Abstract A carbon-11-labeled sulfamate derivative was designed and synthesized as a new
potential positron-emission-tomography dual aromatase–steroid sulfatase inhibitor radio-
tracer for imaging of aromatase and steroid sulfatase expression in breast cancer. The
target tracer 2-chloro-4-(((4-cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)methyl)-6-[11C]
methoxyphenyl sulfamate ([11C]7) was prepared from its corresponding precursor
2-chloro-4-(((4-cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)methyl)-6-hydroxyphenyl sul-
famate (18) with [11C]CH3OTf under basic conditions through the O-[11C]methylation
and isolated by the reversed-phase high-performance liquid chromatography in 40–45%
radiochemical yields based on [11C]CO2 and decay corrected to end of bombardment.
The specific activity at end of synthesis was 111-185 GBq=lmol.
Keywords Aromatase; cancer imaging; positron emission tomography; radiotracer;
steroid sulfatase; sulfamate derivative
Received December 5, 2009.
Address correspondence to Qi-Huang Zheng, Department of Radiology and Imaging Sciences,
Indiana University School of Medicine, 1345 West 16th Street, L3-208, Indianapolis, IN 46202-2111,
USA. E-mail: qzheng@iupui.edu
1127

1128
M. WANG ET AL.
INTRODUCTION
The enzymes aromatase and steroid sulfatase (STS) are particularly attractive
targets in the treatment of estrogen receptor (ER)–positive breast cancer, and a novel
series of sulfamate derivatives have been recently developed as potent dual aroma-
tase–steroid sulfatase inhibitors (DASSIs).[1] The enzymes aromatase and STS also
provide attractive targets for the development of enzyme-based breast cancer ima-
ging agents for the biomedical imaging technique positron emission tomography
(PET).[2] Sulfamate derivatives labeled with a positron-emitting radionuclide such
as carbon-11 or fluorine-18 may enable noninvasive monitoring of the enzymes aro-
matase and STS and breast cancer response to DASSI treatment using PET. The
selected title compound, 2-chloro-4-(((4-cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)-
methyl)-6-methoxyphenyl sulfamate (7), exhibited excellent biological activity:
in vitro inhibition of aromatase and STS activity assessed using intact monolayers
of JEG-3 cells with IC50 (nM) 2.9 ꢀ 0.2 and 536 ꢀ 38, respectively.[1] Also, it has
an O-methyl position amenable to labeling with carbon-11. These properties are
often applied to development of a diagnostic radiotracer. We are interested in the
development of enzyme- and=or receptor-based PET breast cancer imaging agents.
A series of PET breast cancer imaging agents have been developed in this laboratory,
including carbon-11-labeled aromatase inhibitors (NS-398 derivatives) for imaging
of aromatase enzyme,[3] carbon-11-labeled cyclofenil derivatives as nonsteroidal
estrogen radioligands for imaging of ER,[4] carbon-11-labeled tetrahydroisoquino-
line derivatives as selective estrogen receptor modulator (SERM) radioligands for
imaging of ER expression,[5] and carbon-11-labeled sulfamate derivatives as new
DASSI radiotracers for imaging of aromatase and STS expression.[2] This ongoing
study was to develop a new carbon-11-labeled chloro-containing sulfamate deriva-
tive, because this compound displayed better in vitro biological activity than other
compounds previously reported[2] and opened an avenue to label sulfamate deriva-
tives with another positron emitting radionuclide fluorine-18 through a halogen
exchange reaction. Here, we report the design and synthesis of 2-chloro-4-(((4-
cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)methyl)-6-[11C]methoxyphenyl sulfamate
([11C]7), a new potential enzyme-based PET agent for imaging of breast cancer
aromatase and STS expression.
RESULTS AND DISCUSSION
The synthesis of the reference standard 7 is outlined in Scheme 1 using slight mod-
ifications of the literature method.[1] The improvements included modified synthetic
approaches with moderate to excellent chemical yields, more complete experimental
procedures, and detailed spectral data. The phenolic hydroxyl group of 3-chloro-4-
hydroxy-5-methoxybenzaldehyde was protected as benzyl ether with benzyl bromide
in CH3CN using K2CO3 as a base to give 1 in 91% yield.[6] The benzyl protected alde-
hyde 1 was reduced to alcohol 2 using NaBH4 in a mixture of CH2Cl2 and MeOH in
99% yield. The benzyl alcohol 2 was converted to benzyl chloride 3 with SO2Cl2 in
CH2Cl2 in 71% yield. Coupling of the protected hydroxybenzyl chloride 3 and
4-((4-cyanophenyl)amino)-4H-1,2,4-triazole 4, which was prepared from 4-fluoroben-
zonitrile and 4-amino-4H-1,2,4-triazole essentially following the literature method[7] in

SYNTHESIS OF A NEW PET TRACER
1129
Scheme 1. Synthesis of a sulfamate derivative 7.
42% yield, in dimethylformamide (DMF) with NaH as a base afforded the tertiary
amine 5 in 56% yield. Deprotection of benzyl group of 5 by catalytic hydrogenation
with 10% Pb=C in a mixture of MeOH and tetrahydrofuran (THF) provided phenol
6 in 65% yield. The resulting phenol 6 was reacted with an excess of sulfamoyl chloride
in dimethylamine (DMA) to give the target compound 7 in 52% yield.
To obtain a phenolic precursor, we envisioned that 18 could be prepared by
O-demethylation of compound 7. A variety of protocols[8] were screened for this
purpose, including protic acid (HBr), Lewis acids (BBr3, AlCl3=EtSH, LiCl=DMF),
and base (EtSNa). However, in all cases, the demethylated product was obtained
with very poor yield. As a result of these unsuccessful attempts, an alternative strat-
egy was first investigated as we were interested in the development of the precursor
chemistry with readily available starting material. As shown in Scheme 2, demethyla-
tion of 3-chloro-4-hydroxy-5-methoxybenzaldehyde with BBr3 in CH2Cl2 gave
3-chloro-4,5-dihydroxybenzaldehyde 8 in 91% yield.[9,10] To generate differentially
protected hydroxyl groups at 4- and 5-positions, selective benzylation[11] of the more
acidic 4-hydroxyl group by pretreatment of catechol 8 with Li2CO3 as a base in
DMF prior to addition of benzyl bromide was achieved to provide 9 in 89% yield.
Silylation of the 5-hydroxyl group of 9 as tert-butyldimethylsilyl (TBDMS) ether
with tert-butyldimethylsilyl chloride, triethylamine, and dimethylaminopyradine
(DMAP) in CH2Cl2 afforded 10 in 89% yield. Redution of the protected aldehyde
10 with NaBH4 in MeOH provided the resulting alcohol 11 in 84% yield. Bromin-
ation of compound 11 with PBr3 in Et2O gave benzyl bromide 12 in 89% yield. Ben-
zyl bromide 12 was reacted with 4 in CH3CN using K2CO3 as a base to afford a key
intermediate tertiary amine 13 in 13% yield. As indicated in Scheme 3, debenzylation

1130
M. WANG ET AL.
Scheme 2. Synthesis of a key intermediate 13.
Scheme 3. Synthesis of a sulfamate derivative precursor 18 and its regioisomer 19.

SYNTHESIS OF A NEW PET TRACER
1131
of compound 13 by catalytic hydrogenation yielded two phenols as a 3:1 mixture of
regioisomers 14 and 15 in 86% yield. It was difficult to separate and purify the mix-
ture of two regioisomers by silica-gel thin-layer chromatography (TLC) or flash col-
umn chromatography, and the ratio of the resulting regioisomers was determined by
1H NMR. The phenol 15 is the product of 1,2-O,O-TBDMS migration of the phenol
14.[12] Without separation, the mixture was sulfamoylated in DMA to give two sul-
famates 16 and 17 in 51% yield. Likewise, these two regioisomers were not separable
by silica gel. Initial removal of TBDMS groups (desilylation) of compounds 16 and
17 with tetra-n-butylammonium fluoride (TBAF) was unsuccessful. However, 6 N
HCl in THF[13] proved to cleanly cleave the silicon–oxygen bond to give the separ-
able alcohols 18 and 19 in 47% and 13% yields, respectively.
It is noteworthy that desilylation of amine 13 with TBAF in THF took place
rapidly to give phenol 20 in 35% yield. Sulfamoylation of phenol 20 with sulfamoyl
chloride in DMA afforded sulfamate 21 in 68% yield, which in turn was deprotected
by catalytic hydrogenation to give free phenol 19 in 75% yield. The synthetic
approach was outlined in Scheme 4. This also confirmed that the compound 19
was formed due to 1,2-O,O-TBDMS migration.
Synthesis of the target radiotracer [11C]7 is indicated in Scheme 5. Precursor 18
was labeled by
a
reactive [11C]methylating agent, [11C]methyl triflate
([11C]CH3OTf)[14,15] prepared from [11C]CO2, in the presence of 2 N NaOH in acet-
onitrile through the O-[11C]methylation and isolated by semipreparative
reversed-phase high-performance liquid chromatography (HPLC) to provide target
tracer [11C]7 in 40–45% radiochemical yields, decay corrected to end of bombard-
ment (EOB), based on [11C]CO2. The synthesis was performed in an automated mul-
tipurpose 11C-radiosynthesis module, allowing measurement of specific activity
during synthesis.[16,17] The specific activity of [11C]7 was in a range of 222–
370 GBq=mmol at EOB measured by the on-the-fly technique using semipreparative
HPLC during synthesis[17] and 111–185 GBq=mmol at the end of synthesis (EOS)
determined by analytical HPLC,[18] respectively. Chemical purity and radiochemical
purity were determined by analytical HPLC.[18] The chemical purity of the precursor
18 and reference standard 7 was >96%. The radiochemical purity of the target tracer
[11C]7 was >99% determined by radio-HPLC through c-ray (PIN diode) flow detec-
tor, and the chemical purity of the target tracers [11C]7 was >95% determined by
reversed-phase HPLC through an ultraviolet (UV) flow detector.
The precursor has the potential to methylate either at the nitrogen or at the
oxygen position. The radio-HPLC chromatogram of the radiosynthesis did show
Scheme 4. Synthesis of a sulfamate derivative regioisomer 19.

1132
M. WANG ET AL.
Scheme 5. Synthesis of a carbon-11–labeled sulfamate derivative [11C]7.
other radiolabeled minor by-products, because the methylation of the precursor at
the nitrogen position is a potential competing reaction. However, the evidence pro-
vided by the radio-HPLC chromatogram of the radiosynthesis showed that the pre-
cursor is mainly methylated at the oxygen position to produce the major target
radiotracer compared to the retention time of the reference standard. These results
are consistent with the theoretical explanation that the deprotonization at the oxygen
position of the precursor is easier than at the nitrogen position under basic con-
ditions (2 N NaOH) because the acidity of HO- of the precursor is greater than
the acidity of H2NO2SO- of the precursor, and the methylation of the precursor will
prefer to occur at the oxygen position rather than at the nitrogen position. More
strongly basic conditions such as NaH may increase the yield of the methylated pro-
duct at the nitrogen position.[19] Compared to the acidic HO- of the precursor,
H2NO2SO- of the precursor tends to be basic.
CONCLUSIONS
In summary, an efficient and convenient synthesis of a new carbon-11-labeled
sulfamate derivative has been developed. The new precursor synthetic methodology
employed organic reactions such as selective benzylation and debenzylation,
reduction, bromination, coupling reaction, protecting and deprotecting reactions,
and sulfamoylation to prepare a series of new sulfamate derivative precursors. The
target radiotracer was prepared by the O-[11C]methylation of its corresponding phe-
nolic precursor using a reactive [11C]methylating agent, [11C]CH3OTf, and isolated
by a HPLC procedure in high radiochemical yields, short overall synthesis time,
and high specific radioactivities. These chemistry results combined with the reported
in vitro biological data[1] encourage further in vivo biological evaluation of this new
carbon-11-labeled DASSI as a candidate PET radiotracer for imaging of enzymes
aromatase and STS in breast cancer.
EXPERIMENTAL
All commercial reagents and solvents from Sigma-Aldrich and Fisher Scientific
were used without further purification. [11C]CH3OTf was prepared according to a
literature procedure.[15] Melting points were determined on a Mel-Temp II capillary

SYNTHESIS OF A NEW PET TRACER
1133
1
tube apparatus and were uncorrected. H NMR spectra were recorded on Bruker
Avance II 500-MHz NMR spectrometers using tetramethylsilane (TMS) as an inter-
nal 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 con-
stants (J) were reported in hertz (Hz). The low-resolution mass spectra (LRMS) were
obtained using a Bruker Biflex III matrix-assisted laser desorption=ionization
time-of-flight (MALDI-TOF) mass spectrometer, and the high resolution mass spec-
tra (HRMS) were obtained using a Thermo MAT 95XP-Trap spectrometer. Chroma-
tographic solvent proportions are indicated as volume=volume ratio. TLC was run
using Analtech silica-gel GF uniplates (5 ꢁ 10 cm2). Plates were visualized under
UV light. Preparative TLC was run using Analtech silica gel UV 254 plates
(20 ꢁ 20 cm2). Normal phase flash column 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 next. 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 mm C-18 column, 4.6 ꢁ 250 mm; 3:1:1 CH3CN = MeOH = 20 mM, pH 6.7 phosphate
(buffer solution) mobile phase; flow rate 1.5 mL=min; and UV (254 nm) and c-ray
(PIN diode) flow detectors. Semipreparative HPLC was performed using a
YMC-Pack ODS-A, S-5 mm, 12 nm, 10 ꢁ 250 mm C-18 column; 3:1:1 CH3CN =
MeOH = 20 mM, pH 6.7 phosphate (buffer solution) mobile phase; 5.0 mL=min flow
rate; UV (254 nm) and c-ray (PIN diode) flow detectors. Sterile Millex-GS 0.22-mm
vented filter unit was obtained from Millipore Corporation, Bedford, MA.
4-(Benzyloxy)-3-chloro-5-methoxybenzaldehyde (1)
Benzyl bromide (15.3 mL, 129 mmol) was added to a mixture of 3-chloro-4-
hydroxy-5-methoxybenzaldehyde (10.0 g, 53.6 mmol) and K2CO3 (18.6 g, 134 mmol)
in CH3CN (160 mL). The mixture was stirred, heated at reflux for 6 h, and then
allowed to cool to ambient temperature. After the inorganic powder was filtered
off, the filtrate was concentrated in vacuo. Water was added to the residue and
extracted with CH2Cl2. The combined organic layer was washed with brine, dried
(Na2SO4), filtered, concentrated in vacuo, and then purified with column chromato-
graphy (1:4 EtOAc=hexanes) to give 1 (13.4 g, 91%) as a pale yellow solid, mp
1
42–43 ꢂC (lit.[6] mp 43–44 ꢂC). H NMR (CDCl3): d 9.84 (s, 1H), 7.51–7.50 (m,
3H), 7.39–7.33 (m, 4H), 5.17 (s, 2H), 3.94 (s, 3H).
(4-(Benzyloxy)-3-chloro-5-methoxyphenyl)methanol (2)
NaBH4 (2.2 g, 58.3 mmol) was added to a stirred solution of 1 (13.0 g, 47.1 mmol)
in CH2Cl2 (8 mL) and MeOH (80 mL) portionwise at 0 ꢂC. After stirring at room tem-
perature for 5 h, the reaction mixture was concentrated in vacuo. Cooled water was
added to the residue and extracted with EtOAc. The combined organic layer was
washed with brine and dried (Na2SO4). Concentration in vacuo afforded 2 (13.0 g,
99%) as a white solid, which was used for the next step in the reaction without further
1
purification. H NMR (CDCl3): d 7.53–7.52 (m, 2H), 7.39–7.31 (m, 3H), 6.96 (d,
J ¼ 1.5 Hz, 1H), 6.85 (d, J ¼ 1.5 Hz, 1H), 5.02 (s, 2H), 4.61 (s, 2H), 3.87 (s, 3H).

1134
M. WANG ET AL.
4-(Benzyloxy)-3-chloro-5-methoxybenzyl chloride (3)
SOCl2 (9.0 mL, 123.7 mmol) was added to a stirred solution of 2 (13.0 g,
46.8 mmol) in CH2Cl2 (75 mL) dropwise. After stirring at room temperature for
4 h, the reaction mixture was concentrated in vacuo. The oily residue was poured
onto ice and extracted with Et2O. The combined organic layer was washed with satu-
rated NaHCO3, water, and brine; dried (Na2SO4); filtered; and concentrated in
vacuo. The crude product was purified by trituration from CH2Cl2 by addition of
hexanes to afford 3 (9.9 g, 71%) as a white solid, mp 39–40 ꢂC (lit.[1] mp
1
39–41 ꢂC). H NMR (CDCl3): d 7.53–7.52 (m, 2H), 7.40–7.32 (m, 3H), 7.02 (d,
J ¼ 2.0 Hz, 1H), 6.86 (d, J ¼ 2.0 Hz, 1H), 5.04 (s, 2H), 4.51 (s, 2H), 3.88 (s, 3H).
4-((4-Cyanophenyl)amino)-4H-1,2,4-triazole (4)
4-Amino-4H-1,2,4-triazole (16.8 g, 200 mmol) was added portionwise to a stir-
red suspension of potassium tert-butoxide (22.4 g, 200 mmol) in DMSO (100 mL) at
0 ꢂC. After stirring at room temperature for 0.5 h, a solution of 4-fluorobenzonitrile
(12.2 g, 100 mmol) in DMSO (30 mL) was added dropwise below 30 ꢂC. The mixture
was stirred for another 0.5 h at room temperature, then poured into water, and neu-
tralized with 1 N HCl. The precipitate was filtered and recrystallized from water to
1
afford 4 (7.8 g, 42%) as a white solid, mp 202–204 ꢂC (lit.[7] mp 206–208 ꢂC). H
NMR (DMSO-d6): d 10.22 (s, 1H), 8.84 (s, 2H), 7.69 (d, J ¼ 9.0 Hz, 2H), 6.56 (d,
J ¼ 9.0 Hz, 2H).
4-((4-(Benzyloxy)-3-chloro-5-methoxybenzyl)(4H-1,2,4-triazole-4-
yl)amino)benzonitrile (5)
Compound 4 (4.0 g, 21.6 mmol) was added portionwise to a stirred suspension
of NaH (60% dispersion in mineral oil, 950 mg, 23.8 mmol) in DMF (45 mL) at 0 ꢂC.
After stirring at room temperature for 0.5 h, a solution of 3 (7.06 g, 23.8 mmol) in
DMF (15 mL) was added, and the mixture was heated at 90 ꢂC overnight. The reac-
tion mixture was cooled, poured into water, and extracted with EtOAc. The com-
bined organic layer was washed with brine and dried (Na2SO4). Concentration in
vacuo gave an orange residue, which was recrystallized from i-PrOH to afford 5
(5.2 g, 56%) as a white solid, mp 178–180 ꢂC (lit.[1] mp 177–178 ꢂC). 1H NMR
(DMSO-d6): d 8.85 (s, 2H), 7.78 (d, J ¼ 9.0 Hz, 2H), 7.44–7.42 (m, 2H), 7.39–7.31
(m, 3H), 6.97 (s, 2H), 6.78 (d, J ¼ 9.0 Hz, 2H), 5.00 (s, 2H), 4.96 (s, 2H), 3.81 (s, 3H).
4-((3-Chloro-4-hydroxy-5-methoxybenzyl)(4H-1,2,4-triazol-4-
yl)amino)benzonitrile (6)
To a stirred solution of 5 (4.0 g, 9.3 mmol) in THF (200 mL) and MeOH
(100 mL) was added 10% Pd=C (400 mg). The flask was charged with a balloon filled
with H2 gas and stirred at room temperature overnight. The catalyst was removed by
filtration through celite. The filtrate was concentrated in vacuo to give a beige resi-
due, which was recrystallized from EtOH to give 6 (2.1 g, 65%) as a white solid, mp
1
235 ꢂC (dec) [lit.[1] mp > 230 ꢂC (dec)]. H NMR (DMSO-d6): d 9.50 (s, 1H), 8.79

SYNTHESIS OF A NEW PET TRACER
1135
(s, 2H), 7.77 (d, J ¼ 9.0 Hz, 2H), 6.84 (s, 1H), 6.82 (s, 1H), 6.79 (d, J ¼ 9.0 Hz, 2H),
4.92 (s, 2H), 3.77 (s, 3H).
2-Chloro-4-(((4-cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)methyl)-6-
methoxyphenyl Sulfamate (7)
Sulfamoyl chloride (729 mg, 6.31 mmol) was added to a stirred solution of 6
(660 mg, 1.86 mmol) in dimethylamine (DMA) (6 mL) at 0 ꢂC under a nitrogen
atmosphere. After stirring at room temperature overnight, the mixture was poured
into ice water and extracted with EtOAc. The combined organic layer was washed
with brine, dried (Na2SO4), filtered, and concentrated in vacuo. The crude product
was purified with column chromatography (1:7 MeOH=CHCl3) to give 7 (420 mg,
52%) as a white solid, mp 217 ꢂC (dec) [lit.[1] mp > 210 ꢂC (dec)]. 1H NMR
(DMSO-d6): d 8.91 (s, 2H), 7.98 (s, 2H), 7.78 (d, J ¼ 9.0 Hz, 2H), 7.08 (s, 1H),
7.05 (s, 1H), 6.75 (d, J ¼ 9.0 Hz, 2H), 5.06 (s, 2H), 3.77 (s, 3H).
3-Chloro-4,5-dihydroxybenzaldehyde (8)
BBr3 (12.1 mL, 130 mmol) was added to a stirred solution of 3-chloro-4-
hydroxy-5-methoxybenzaldehyde (20.0 g, 110 mmol) in CH2Cl2 (350 mL) at 0 ꢂC
under a nitrogen atmosphere. After stirring at 0 ꢂC for 0.5 h and at room tempera-
ture for 3 h, the mixture was cooled to 0 ꢂC, and MeOH (100 mL) was added care-
fully. The solvent was concentrated in vacuo, and the residue was added MeOH
(100 mL). This process was repeated three times. The residue was diluted with
CH2Cl2, and the precipitate was filtered and washed with CH2Cl2 to give 8 (16.8 g,
1
91%) as a pink solid, mp 235 ꢂC (dec). H NMR (DMSO-d6): d 10.4 (s, 2H), 9.70
(s, 1H), 7.43 (d, J ¼ 2.0 Hz, 1H), 7.22 (d, J ¼ 2.0 Hz, 1H).
4-(Benzyloxy)-3-chloro-5-hydroxybenzldehyde (9)
Li2CO3 (13.4 g, 180.8 mmol) was added to a stirred solution of 8 (12.0 g,
69.54 mmol) in DMF (120 mL). After stirring and heating atꢂC for 1 h, benzyl bro-
mide (21.5 mL, 180.8 mmol) was added dropwise. The mixture was stirred at 45 ꢂC
for another 1 h and then cooled to 0 ꢂC. The reaction was quenched with 1 N HCl
and extracted with EtOAC. The combined organic layer was washed with water
and brine, dried (Na2SO4), filtered, concentrated in vacuo, and then purified with
column chromatography (1:9 hexanes=CH2Cl2) to give 9 (16.2 g, 89%) as an
1
off-white solid, mp 45–47 ꢂC. H NMR (CDCl3): d 9.80 (s, 1H), 7.49 (d, J ¼ 2.0 Hz,
1H), 7.44–7.37 (m, 5H), 7.32 (d, J ¼ 2.0 Hz, 1H), 5.19 (s, 2H). HRMS (CI, m=z): calc.
for C14H12O3Cl ([M þ H]þ), 263.0469; found, 263.0459.
4-(Benzyloxy)-3-(tert-butyldimethylsilyloxy)-5-
chlorobenzaldehyde (10)
Triethylamine (44.6 mL, 320 mmol) was added to a stirred solution of 9 (12.0 g,
45.7 mmol) and DMAP (1.68 g, 13.7 mmol) in CH2Cl2 (200 mL), followed by tert-
butyldimethylsilyl chloride (20.7 g, 137.1 mmol), at 0 ꢂC under a nitrogen atmos-
phere. After stirring at 0 ꢂC for 1 h and at room temperature for 2 h, the reaction

1136
M. WANG ET AL.
mixture was washed with saturated NH4Cl. The organic layer was washed with water
and brine, dried (Na2SO4), filtered, concentrated in vacuo, and then purified with
column chromatography (1:9 hexanes=CH2Cl2) to give 10 (15.4 g, 89%) as a pale
1
yellow oil. H NMR (CDCl3): d 9.82 (s, 1H), 7.52 (d, J ¼ 2.0 Hz, 1H), 7.49–7.48
(m, 2H), 7.39–7.33 (m, 3H), 7.30 (d, J ¼ 2.0 Hz, 1H), 5.12 (s, 2H), 1.02 (s, 9H),
0.23 (s, 6H). HRMS (CI, m=z): calc. for C20H26O3ClSi ([M þ H]þ), 377.1334; found,
377.1333.
(4-(Benzyloxy)-3-(tert-butyldimethylsilyloxy)-5-
chlorophenyl)methanol (11)
NaBH4 (2.6 g, 68.7 mmol) was added to a stirred solution of 10 (14.6 g,
38.7 mmol) in MeOH (40 mL) portionwise at 0 ꢂC. After stirring at room tempera-
ture for 5 h, the reaction mixture was concentrated in vacuo. Cooled water was
added to the residue and extracted with EtOAc. The combined organic layer was
washed with brine, dried (Na2SO4), filtered, concentrated in vacuo, and then purified
with column chromatography (1:100 MeOH=CH2Cl2) to give 11 (12.3 g, 84%) as a
1
white solid, mp 37–39 ꢂC. H NMR (CDCl3): d 7.51–7.50 (m, 2H), 7.39–7.31 (m,
3H), 7.01 (d, J ¼ 2.0 Hz, 1H), 6.81 (d, J ¼ 2.0 Hz, 1H), 5.00 (s, 2H), 4.58 (s, 2H),
1.00 (s, 9H), 0.19 (s, 6H). HRMS (CI, m=z): calc. for C20H27O3ClSi ([M]þ),
378.1415; found, 378.1413.
4-(Benzyloxy)-3-(tert-Butyldimethylsilyloxy)-5-chlorobenzyl
Bromide (12)
PBr3 (3.0 mL, 31.9 mmol) was added to a stirred solution of 11 (11.5 g,
30.3 mmol) in Et2O (150mL) dropwise at 0 ꢂC. After stirring at room temperature
for 7 h, the reaction mixture was poured into aqueous NaHCO3 and extracted with
CH2Cl2. The combined organic layer was washed with water and brine, dried
(Na2SO4), filtered, and concentrated in vacuo to afford 12 (11.9 g, 89%) as a pale
yellow oil. This material was used for the next step in the reaction without further puri-
fication. 1H NMR (CDCl3): d 7.51–7.49 (m, 2H), 7.39–7.33 (m, 3H), 7.04 (d, J ¼ 2.0 Hz,
1H), 6.83 (d, J ¼ 2.0 Hz, 1H), 5.01 (s, 2H), 4.37 (s, 2H), 1.00 (s, 9H), 0.20 (s, 6H).
HRMS (CI, m=z): calc. for C20H27O2BrClSi ([M]þ), 441.0647; found, 441.0641.
4-((4-(Benzyloxy)-3-(tert-butyldimethylsilyloxy)-5-chlorobenzyl)(4H-
1,2,4-triazole-4-yl)amino)benzonitrile (13)
A mixture of 4 (4.1 g, 22.3 mmol), 12 (9.8 g, 22.3 mmol), and K2CO3 (6.2 g,
44.5 mmol) in CH3CN (180 mL) was stirred overnight at room temperature. The
reaction mixture was diluted with water and extracted with CH2Cl2. The combined
organic layer was washed with water and brine, dried (Na2SO4), filtered, concen-
trated in vacuo, and then purified with column chromatography (1:30 MeOH=
CH2Cl2) to afford 13 (1.6 g, 13%) as a yellow solid, mp 58–60 ꢂC. 1H NMR
(CD3COCD3): d 8.58 (s, 2H), 7.74–7.71 (m, 2H), 7.55–7.33 (m, 5H), 7.15 (d,
J ¼ 2.0 Hz, 1H), 6.95 (d, J ¼ 2.0 Hz, 1H), 6.88–6.85 (m, 2H), 5.11 (s, 2H), 5.03 (s,
2H), 0.98 (s, 9H), 0.18 (s, 6H). HRMS (TOF ESþ, m=z): calc. for C29H33N5O2ClSi
([M þ H]þ), 546.2092; found, 546.2105.

SYNTHESIS OF A NEW PET TRACER
1137
4-((3-(tert-Butyldimethylsilyloxy)-5-chloro-4-hydroxybenzyl)(4H-
1,2,4-triazol-4-yl)amino)benzonitrile (14) and 4-((4-(tert-
Butyldimethylsilyloxy)-3-chloro-5-hydroxybenzyl)(4H-1,2,4-triazol-4-
yl)amino)benzonitrile (15)
To a stirred solution of 13 (500 mg, 0.92 mmol) in EtOH (5 mL) was added 10%
Pd=C (50 mg). The flask was charged with a H2 balloon and stirred at room tempera-
ture overnight. The catalyst was removed by filtration through celite. The filtrate was
concentrated in vacuo and then purified with preparative TLC plate (1:30 MeOH=
CH2Cl2) to afford a nonseparable mixture of regioisomers 14 and 15 (360 mg,
1
86%) as a yellow solid. H NMR (DMSO-d6): d 9.93, 9.07 (s, 1H), 8.74, 8.71 (s,
2H), 7.76 (d, J ¼ 9.0 Hz, 2H), 6.92–6.64 (m, 4H), 4.90 (s, 2H), 0.99, 0.92 (s, 9H),
0.17, 0.11 (s, 6H). HRMS (TOF ESþ, m=z): calc. for C22H27N5O2ClSi ([M þ H]þ),
456.1623; found, 456.1634.
2-(tert-Butyldimethylsilyloxy)-6-chloro-4-(((4-cyanophenyl)(4H-1,2,4-
triazol-4-yl)amino)methyl)phenyl Sulfamate (16) and 2-(tert-
Butyldimethylsilyloxy)-3-chloro-5-(((4-cyanophenyl)(4H-1,2,4-triazol-
4-yl)amino)methyl)phenyl Sulfamate (17)
Sulfamoyl chloride (227 mg, 1.97 mmol) was added to a stirred solution of a
mixture of two regioisomers 14 and 15 (300 mg, 0.66 mmol) in DMA (3 mL) at
0 ꢂC under a nitrogen atmosphere. After stirring at room temperature overnight,
ice water was added, and the mixture was extracted with EtOAc. The combined
organic layer was washed with brine, dried (Na2SO4), filtered, concentrated in vacuo,
and then purified with preparative TLC plate (1:30 MeOH=CH2Cl2) to afford a
nonseparable mixture of regioisomers 16 and 17 (179 mg, 51%) as a pale yellow solid.
1H NMR (DMSO-d6): d 8.83, 8.78 (s, 2H), 8.17, 8.02 (s, 2H), 7.77 (d, J ¼ 9.0 Hz,
2H), 7.34 (d, J ¼ 2.0 Hz, 1H), 7.30 (d, J ¼ 2.0 Hz, 1H), 6.76 (d, J ¼ 9.0 Hz, 2H),
4.97 (s, 2H), 0.98, 0.92 (s, 9H), 0.20, 0.13 (s, 6H). HRMS (TOF ESþ, m=z): calc.
for C22H28N6O4SClSi ([M þ H]þ), 535.1351; found, 535.1361.
2-Chloro-4-(((4-cyanophenyl)(4H-1,2,4-triazol-4-
yl)amino)methyl)-6-hydroxyphenyl Sulfamate (18) and 3-Chloro-5-
(((4-cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)methyl)-2-
hydroxyphenyl Sulfamate (19)
To a stirred solution of a mixture of two regioisomers 16 and 17 (38 mg,
0.07 mmol) in THF (0.5 mL) was added 6 N HCl (1 mL). After stirring at room tem-
perature for 1 h, the reaction mixture was diluted with water and extracted with
EtOAC. The combined organic layer was washed with brine, dried (Na2SO4), fil-
tered, concentrated in vacuo, and then purified with preparative TLC plate (1:7
MeOH=CH2Cl2) to afford 18 (14 mg, 47%) as a pale yellow solid and 19 (4 mg,
1
13%) as a pale yellow solid. Compound 18, mp 170 ꢂC (dec). H NMR (DMSO-
d6): d 8.85 (s, 2H), 7.77 (d, J ¼ 9.0 Hz, 2H), 6.94 (d, J ¼ 2.0 Hz, 1H), 6.83 (d,
J ¼ 2.0 Hz, 1H), 6.72 (d, J ¼ 9.0 Hz, 2H), 5.00 (s, 2H). HRMS (TOF ESþ, m=z): calc.
for C16H14N6O4SCl ([M þ H]þ), 421.0486; found, 421.0481. Compound 19, mp

1138
M. WANG ET AL.
112–114 ꢂC.1H NMR (DMSO-d6): d 8.78 (s, 2H), 7.76 (d, J ¼ 9.0 Hz, 2H), 7.25 (d,
J ¼ 2.0 Hz, 1H), 7.23 (d, J ¼ 2.0 Hz, 1H), 6.76 (d, J ¼ 9.0 Hz, 2H), 4.97 (s, 2H).
HRMS (TOF ESþ, m=z): calc. for C16H14N6O4SCl ([M þ H]þ), 421.0486; found,
421.0498.
4-((4-(Benzyloxy)-3-chloro-5-hydroxybenzyl)(4H-1,2,4-triazol-4-
yl)amino)benzonitrile (20)
To a stirred solution of 13 (252 mg, 0.46 mmol) in THF (2 mL) was added a
1.0 M solution of TBAF in THF (2.0 mL, 2.0 mmol) slowly at 0 ꢂC under nitrogen
atmosphere. After stirring at room temperature for 1 h, the reaction mixture was
poured into ice water and extracted with CH2Cl2. The combined organic layer was
washed with brine, dried (Na2SO4), filtered, and concentrated in vacuo, and then
purified with preparative TLC plate (1:8 MeOH=CH2Cl2) to afford 20 (70 mg,
1
35%) as a yellow solid, mp 225 ꢂC (dec). H NMR (DMSO-d6): d 10.04 (s, 1H),
8.80 (s, 2H), 7.77 (d, J ¼ 9.5 Hz, 2H), 7.46 (d, J ¼ 7.0 Hz, 2H), 7.37 (t, J ¼ 7.0 Hz,
2H), 7.34–7.31 (m, 1H), 6.86 (d, J ¼ 1.5 Hz, 1H), 6.77 (d, J ¼ 1.5 Hz, 1H), 6.74 (d,
J ¼ 9.0 Hz, 2H), 4.97 (s, 2H), 4.95 (s, 2H). HRMS (TOF ESþ, m=z): calc. for
C23H19N5O2Cl ([M þ H]þ), 432.1227; found, 432.1231.
2-(Benzyloxy)-3-chloro-5-(((4-cyanophenyl)(4H-1,2,4-triazol-4-
yl)amino)methyl)phenyl Sulfamate (21)
Sulfamoyl chloride (170 mg, 1.47 mmol) was added to a stirred solution of 20
(60 mg, 0.14 mmol) in DMA (0.5 mL) at 0 ꢂC under nitrogen atmosphere. After stir-
ring at room temperature overnight, ice water was added, and the mixture was
extracted with EtOAc. The combined organic layer was washed with brine, dried
(Na2SO4), filtered, concentrated in vacuo, and then purified with preparative TLC
plate (1:30 MeOH=CH2Cl2) to afford 21 (48 mg, 68%) as a pale yellow solid, mp
1
159–161 ꢂC. H NMR (DMSO-d6): d 8.82 (s, 2H), 8.35 (s, 2H), 7.78 (d, J ¼ 9.0 Hz,
2H), 7.48 (d, J ¼ 7.5 Hz, 2H), 7.41–7.34 (m, 5H), 6.75 (d, J ¼ 9.0 Hz, 2H), 5.07 (s,
2H), 5.06 (s, 2H). HRMS (TOF ESþ, m=z): calc. for C23H20N6O4SCl ([M þ H]þ),
511.0955; found, 511.0932.
3-Chloro-5-(((4-cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)methyl)-
2-hydroxyphenyl Sulfamate (19)
To a stirred solution of 21 (39 mg, 0.08 mmol) in MeOH (0.3 mL) and THF
(0.5 mL) was added 10% Pd=C (11 mg). The flask was charged with a hydrogen bal-
loon and stirred at room temperature for 3 h. The catalyst was removed by filtration
through celite. The filtrate was concentrated in vacuo and then purified with pre-
parative TLC plate (1:30 MeOH=CH2Cl2) to afford 19 (24 mg, 75%) as a pale yellow
solid, mp 110–112 ꢂC. 1H NMR (DMSO-d6): d 8.78 (s, 2H), 7.76 (d, J ¼ 9.0 Hz, 2H),
7.24 (d, J ¼ 2.0 Hz, 1H), 7.20 (d, J ¼ 2.0 Hz, 1H), 6.76 (d, J ¼ 9.0 Hz, 2H), 4.97 (s,
2H). LC-MS (ESI, m=z): calc. for C16H14N6O4SCl ([M þ H]þ), 421.0; found, 421.1.

SYNTHESIS OF A NEW PET TRACER
1139
2-Chloro-4-(((4-cyanophenyl)(4H-1,2,4-triazol-4-yl)amino)methyl)-
6-[11C]methoxyphenyl Sulfamate ([11C]7)
[11C]CO2 was produced by the 14N(p,a)11C nuclear reaction in a small-volume
(9.5-cm3) aluminum gas target (CTI) from 11-MeV proton cyclotron on research
purity nitrogen (þ1% O2) in a Siemens radionuclide delivery system (Eclipse
RDS-111). In a small reaction vial (5 mL), the precursor 18 (0.1 mg) was dissolved
in CH3CN (300 mL). To this solution was added 2 N NaOH (2 mL). No carrier-added
(high specific activity) [11C]CH3OTf that was produced by the gas-phase production
method[15] from [11C]CO2 through [11C]CH4 and [11C]CH3Br with a silver triflate
(AgOTf) column was passed into the reaction vial at room temperature, until radio-
activity reached a maximum (ꢃ2 min), and then the reaction vial was isolated and
reacted at 80 ꢂC for 3 min. The contents of the reaction vial were diluted with
NaHCO3 (1 mL, 0.1 M) and injected onto the semipreparative 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, [11C]7, was formu-
lated in saline, sterile-filtered through a sterile vented Millex-GS 0.22-mm cellulose
acetate membrane, and collected into a sterile vial. Total radioactivity was assayed,
and total volume was noted for tracer dose dispensing. The overall synthesis, puri-
fication, and formulation time was 20–25 min from EOB. Retention times in the ana-
lytical HPLC system were tR 18 ¼ 3.45 min, tR 7 ¼ 6.58 min, and tR [11C]7 ¼ 6.58 min.
Retention times in the semipreparative HPLC system were tR 18 ¼ 5.76 min, tR
7 ¼ 8.02 min, and tR [11C]7 ¼ 8.02 min. The radiochemical yields were 40–45% decay
corrected to EOB, based on [11C]CO2.
ACKNOWLEDGMENTS
This work was partially supported by the Susan G. Komen for the Cure, Breast
Cancer Research Foundation, and Indiana Genomics Initiative (INGEN) of Indiana
University, which is supported in part by Lilly Endowment Inc. The authors thank
Dr. Bruce H. Mock and Barbara E. Glick-Wilson for their assistance in production
1
of [11C]CH3OTf. H NMR spectra were recorded on a Bruker Avance II 500-MHz
NMR spectrometer in the Department of Chemistry and Chemical Biology at
Indiana University Purdue University Indianapolis (IUPUI), which is supported
by NSF-MRI Grant CHE-0619254. The referee’s criticisms and editor’s comments
for the revision of the manuscript are greatly appreciated.
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