Simple synthesis of carbon-11 labeled styryl dyes as new potential PET RNA-specific, living cell imaging probes

Article abstract of DOI:10.1016/j.ejmech.2008.02.033

A new type of styryl dyes have been developed as RNA-specific, live cell imaging probes for fluorescent microscopy technology to study nuclear structure and function. This study was designed to develop carbon-11 labeled styryl dyes as new probes for biome

Full text of DOI:10.1016/j.ejmech.2008.02.033

Available online at www.sciencedirect.com
European Journal of Medicinal Chemistry 44 (2009) 2300e2306
http://www.elsevier.com/locate/ejmech
Laboratory note
Simple synthesis of carbon-11 labeled styryl dyes as new potential
PET RNA-specific, living cell imaging probes
Min Wang a, Mingzhang Gao a, Kathy D. Miller b, George W. Sledge b,
Gary D. Hutchins a, Qi-Huang Zheng a,
*
a Department of Radiology, Indiana University School of Medicine, 1345 West 16th Street, L3-208, Indianapolis, IN 46202-2111, USA
b Department of Medicine, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Received 18 February 2008; received in revised form 27 February 2008; accepted 28 February 2008
Available online 7 March 2008
Abstract
A new type of styryl dyes have been developed as RNA-specific, live cell imaging probes for fluorescent microscopy technology to study
nuclear structure and function. This study was designed to develop carbon-11 labeled styryl dyes as new probes for biomedical imaging
technique positron emission tomography (PET) imaging of RNA in living cells. Precursors (E )-2-(2-(1-(triisopropylsilyl)-1H-indol-3-yl)vinyl)-
quinoline (2), (E )-2-(2,4,6-trimethoxystyryl)quinoline (3) and (E )-4-(2-(6-methoxyquinolin-2-yl)vinyl)-N,N-diemthylaniline (4), and standards
styryl dyes E36 (6), E144 (7) and F22 (9) were synthesized in multiple steps with moderate to high chemical yields. Precursor 2 was labeled by
[11C]CH3OTf, trapped on a cation-exchange CM Sep-Pak cartridge following a quick deprotecting reaction by addition of (n-Bu)4NF in THF,
and isolated by solid-phase extraction (SPE) purification to provide target tracer [11C]E36 ([11C]6) in 40e50% radiochemical yields, decay
corrected to end of bombardment (EOB), based on [11C]CO2. The target tracers [11C]E144 ([11C]7) and [11C]F22 ([11C]9) were prepared by
N-[11C]methylation of the precursors 3 and 4, respectively, using [11C]CH3OTf and isolated by SPE method in 50e70% radiochemical yields
at EOB. The specific activity of the target tracers [11C]6, [11C]7 and [11C]9 was in a range of 74e111 GBq/mmol at the end of synthesis (EOS).
Ó 2008 Elsevier Masson SAS. All rights reserved.
Keywords: Positron emission tomography; Carbon-11; Styryl dyes; RNA; Live cell; Imaging
1. Introduction
available. Commercial cyanine dyes are widely used for RNA
imaging in living cell, but their photostability and RNA selecti-
vity are limited, making them difficult to use for cellular
time-lapse RNA imaging [8,9]. Recently, fluorescent styryl
dyes E36 [(E )-2-(2-(1H-indol-3-yl)vinyl)-1-methylquinoli-
nium iodide (6)], E144 [(E )-1-methyl-2-(2,4,6-trimethoxystyr-
yl)quinolinium iodide (7)] and F22 [(E )-2-(4-(dimethylamino)
styryl)-6-methoxy-1-methylquinolinium iodide (9)] have
been identified as RNA-specific, live cell imaging probes
for fluorescent microscopy technology to study nuclear struc-
ture and function [7,10]. These probes are more selective for
RNA and more photostable than cyanine agents and are
particularly advantageous for visualizing RNA sites in live
cell nuclei. These RNA probes possess N-methyl position
amenable to labeling with a positron emitting radioisotope
such as carbon-11 as RNA radioligands. The same properties
are often beneficial in a diagnostic radiotracer. RNA also
There is great interest in imaging of RNA in human dis-
eases such as various neurological and psychiatric disorders,
heart and cancer diseases [1e6]. However, finding a RNA-se-
lective probe for living cell imaging has proved to be difficult
[7], and only a few RNA visualization agents are currently
Abbreviations: PET, positron emission tomography; SPE, solid-phase
extraction; EOB, end of bombardment; EOS, end of synthesis; rt, room
temperature; HPLC, high pressure liquid chromatography; SPECT, single
photon emission computed tomography; MRI, magnetic resonance imaging;
TMS, tetramethylsilane; HRMS, high resolution mass spectra; TLC, thin-layer
chromatography; RDS, radionuclide delivery system; INGEN, Indiana
genomics initiative.
* Corresponding author. Tel.: þ1 317 278 4671; fax: þ1 317 278 9711.
E-mail address: qzheng@iupui.edu (Q.-H. Zheng).
0223-5234/$ - see front matter Ó 2008 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2008.02.033

M. Wang et al. / European Journal of Medicinal Chemistry 44 (2009) 2300e2306
2301
provides an attractive target for the in vivo biomedical
imaging technique positron emission tomography (PET) to
map RNA and its related diseases. Compared to fluorescent
microscopy technology, only PET has sufficient sensitivity
and quantitation to measure the expression of genes in vivo
[1]. We are interested in the development of PET RNA
probes. In our previous works, we have developed radiola-
beled O6-benzylguanine derivatives for PET imaging of
DNA repair protein [11e14], radiolabeled penciclovir and
ganciclovir analogues as PET reporter probes for herpes sim-
plex virus thymidine kinase (HSV-tk) gene [15e17], and
radiolabeled D-luciferin derivatives as PET reporter probes
for luciferase gene [18]. In this ongoing study, we have de-
signed and synthesized new carbon-11 labeled styryl dyes,
(E )-2-(2-(1H-indol-3-yl)vinyl)-1-[11C]methylquinolinium tri-
flate ([11C]E36, [11C]6), (E )-1-[11C]methyl-2-(2,4,6-trime-
thoxystyryl)quinolinium triflate ([11C]E144, [11C]7), and
(E )-2-(4-(dimethylamino)styryl)-6-methoxy-1-[11C]methyl-
quinolinium triflate ([11C]F22, [11C]9), as potential PET
RNA-specific, living cell imaging probes.
E-2-styrylquinolines (1, 3 and 4) [19,20] in 32e73% yields.
Triisopropylsilyl group for protection of indole nitrogen of
compound 1 was employed since its silyl moiety could be
removed easily by tetrabutylammonium fluoride [(n-Bu)4NF]
in THF at room temperature (rt) [21,22]. Thus, compound 1
was reacted with sodium hydride and triisopropylsilyl chloride
in DMF to give precursor 2 in 59% yield.
Synthesis of standard compounds E36 (6), E144 (7) and
F22 (9) according to literature procedures with modifications
[7,10] is outlined in Scheme 2. N-methylation of 2-methylqui-
noline or 6-methoxy-2-methylquinoline with iodomethane in
acetonitrile provided 1,2-dimethylquinolinium iodide 5 and
6-methoxy-1,2-dimethylquinolinium iodide 8 in 60 and 62%
yields, respectively. Compound 6 could be prepared by
straightforward condensation of compound 5 and indole-3-
aldehyde, instead of the acetylated indole-3-aldehyde [7],
using catalytic amounts of pyrrolidine in EtOH in 39% yield.
Compounds 7 and 9 were prepared in a manner similar to
compound 6 from corresponding aromatic aldehydes by reac-
tion with methylquinolinium iodides (5 and 8) with catalytic
amounts of pyrrolidine or piperidine in 72 and 99% yields,
respectively.
2. Results and discussion
2.1. Chemistry
2.2. Radiochemistry
Synthesis of precursors (2e4) is shown in Scheme 1. As
illustrated in Scheme 1, condensation of 2-methylquinoline
or 6-methoxy-2-methylquinoline with aromatic aldehydes in
the presence of catalytic amounts of piperidine afforded
Synthesis of target probes [11C]E36 ([11C]6), [11C]E144
([11C]7) and [11C]F22 ([11C]9) is indicated in Scheme 3. Pre-
cursor 2 was labeled by a reactive [11C]methylating agent,
[11C]methyl triflate ([11C]CH3OTf) [23,24] prepared from
CHO
(i-Pr)3SiCl
NH
NaH, DMF
NH
N
piperidine
N
CH3
H3CO
OHC
1
73
OCH3
H3CO
N
piperdine
Si(i-Pr)3
N
2
59
OCH3
N
3
43
H3CO
OCH3
H3CO
OHC
N(CH3)2
H3CO
N
piperdine
N
CH3
4
32
CH3
N
CH3
Scheme 1. Synthesis of styryl dye precursors.

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M. Wang et al. / European Journal of Medicinal Chemistry 44 (2009) 2300e2306
CHO
CH3I, CH3CN
N
NH
NH
CH3
N
I
CH3
CH3
N
pyrrolidine, EtOH
CH3
I
6
5
60
39
H3CO
OHC
OCH3
OCH3
H3CO
N
I
pyrrolidine, EtOH
H3CO
CH3
H3CO
OCH3
7
72
N
CH3
CH3I, CH3CN
H3CO
H3CO
OHC
N(CH3)2
N
piperidine, MeOH
N
I
CH3
I
CH3
CH3
N
8
62
CH3
9
99
CH3
Scheme 2. Synthesis of styryl dyes as standards.
[11C]CO2, in acetonitrile through the N-[11C]methylation and
trapped on a cation-exchange CM Sep-Pak cartridge to release
the non-reacted indole nitrogen protected tertiary amine pre-
cursor with ethanol and to retain the pure N-[11C]-methylated
quaternary ammonium intermediate, (E )-2-(2-(1-triisopropyl-
silyl)-1H-indol-3-yl)vinyl-1-[11C]methylquinolinium triflate
([11C]10), on the same CM Sep-Pak. The 11C-labeled
intermediate underwent the deprotecting reaction [21,22] by
addition of a 1.0 M solution of (n-Bu)4NF in THF to the
same cartridge. After 2 min, non-reacted (n-Bu)4NF was re-
moved from the cartridge by rinsing with ethanol. The final
carbon-11 labeled product [11C]E36 was then eluted from
the cartridge with saline in 40e50% radiochemical yields,
decay corrected to end of bombardment (EOB), based on
(n-Bu)4NF
THF, rt
11C]CH3OTf, CH3CN
N
[
N
N
Si(i-Pr)3
2
11CH3
OTf
NH
11CH3
OTf
[
11C]10
[
11C]6
40-50
%
OCH3
[
11C]CH3OTf, CH3CN
N
3
OTf
11CH3
H3CO
OCH3
[
11C]7
50-70%
H3CO
[
11C]CH3OTf, CH3CN
4
N
OTf
11CH3
11C]9
50-70%
CH3
N
[
CH3
Scheme 3. Synthesis of carbon-11 labeled styryl dyes.

M. Wang et al. / European Journal of Medicinal Chemistry 44 (2009) 2300e2306
2303
[11C]CO2. The synthesis was performed in an automated multi-
purpose 11C-radiosynthesis module, allowing measurement of
specific activity during synthesis [25e27]. The specific activity
of [11C]E36 was in a range of 74e111 GBq/mmol at the end of
synthesis (EOS). The purification technique we used in the ra-
diosynthesis of [11C]E36 is the solid-phase extraction (SPE)
method [28,29], and the key part in this technique is a CM
Sep-Pak cartridge. Similarly, tertiary amine precursors 3 and
4 were labeled with [11C]CH3OTf and purified by SPE
technique to provide target quaternary ammonium tracers
[11C]E144 and [11C]F22, respectively, in 50e70% radiochem-
ical yields, decay corrected to EOB, based on [11C]CO2. The
large polarity difference between the tertiary amine precursor
and the labeled quaternary ammonium product permitted the
use of SPE technique for purification of labeled product from
radiolabeling reaction mixture. A cation-exchange CM Sep-
Pak cartridge was used in SPE purification technique. The
reaction mixture was loaded onto the cartridge by gas pressure.
The cartridge column was washed with ethanol and water to
remove non-reacted [11C]CH3OTf, precursor and reaction
solvent, and then final labeled product was eluted with saline
from the CM Sep-Pak. SPE technique is fast, efficient and
convenient and works very well for the quaternary ammonium
tracer production. The specific activity for [11C]E144 and
[11C]F22 was in a range of 74e111 GBq/mmol at EOS under
the same targetry conditions which have been used in the radio-
synthesis of [11C]E36, determined by analytical high pressure
liquid chromatography (HPLC) method [30].
points were determined on a MEL-TEMP II capillary tube ap-
paratus and were uncorrected. 1H NMR spectra were recorded
on a Bruker QE 300 NMR spectrometer using tetramethylsi-
lane (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 ) were reported in hertz (Hz). The high
resolution mass spectra (HRMS) were obtained using
a Thermo MAT 95XP-Trap spectrometer. Chromatographic
solvent proportions are indicated in a volume:volume ratio.
Thin-layer chromatography (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). 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 KHPOꢁ4 (buffer solution) mo-
bile phase; flow rate 1.5 mL/min; and UV (254 nm) and g-ray
(NaI) flow detectors. Semi-prep cation-exchange CM Sep-Pak
cartridges were obtained from Waters Corporate Headquarters,
Milford, MA. Sterile vented Millex-GS 0.22 mm filter unit was
obtained from Millipore Corporation, Bedford, MA.
3.2. (E )-2-(2-(1H-Indol-3-yl)vinyl)quinoline (1)
A
mixture of indole-3-aldehyde (1.0 g, 6.89 mmol),
Methods available to image gene expression in living
animals are rapidly increasing. Several techniques, including
radionuclide approaches PET and single photon emission
computed tomography (SPECT), magnetic approach magnetic
resonance imaging (MRI), and optical approaches green fluo-
rescent protein and luciferase, are all under active investiga-
tion [1]. Fluorescent styryl molecules have been synthesized
and screened for an in vitro RNA response and live cell
nuclear imaging [7,10]. However, fluorescent microscopy
technology is only available for small animals in preclinical
study and has not sufficient sensitivity and quantitation to
measure the expression of genes in vivo. PET coupled with
appropriate radiotracers has the unique capability of non-inva-
sively measuring biochemical and metabolic processes, is par-
ticularly suited for quantitatively imaging animals and humans
with a relatively high sensitivity, and has become a clinically
valuable and accepted diagnostic tool to image diseases. The
further in vivo evaluation of PET imaging of transgene expres-
sion in animal models with the probes [11C]E36, [11C]E144
and [11C]F22 is currently underway, and the results will be
reported in due course.
2-methylquinoline (1.18 g, 8.25 mmol), and a catalytic amount
of piperidine (80 mL) in a tube was heated at 158 ꢂC overnight.
After cooling, the resulting solid was triturated with i-PrOH,
filtered, and recrystallized from MeOH to afford 1 (1.35 g,
73%) as a yellow solid, mp 209e210 ꢂC (lit [19], 210e
1
211 ꢂC). H NMR (CDCl3): d 8.24 (d, J ¼ 8.7 Hz, 1H, Ar-H),
8.10e8.06 (m, 1H, Ar-H), 7.96 (d þ d, J ¼ 8.6, 16.5 Hz,
2H, Ar-H and CH]CH), 7.91 (d, J ¼ 8.7 Hz, 1H, Ar-H),
7.85 (d, J ¼ 8.1 Hz, 1H, Ar-H), 7.74e7.68 (m, 1H, Ar-H),
7.64 (s, 1H, Ar-H), 7.52e7.38 (m þ d, J ¼ 16.5 Hz, 3H,
Ar-H þ CH]CH), 7.23e7.18 (m, 2H, Ar-H).
3.3. (E )-2-(2-(1-(Triisopropylsilyl)-1
H-indol-3-yl)vinyl)quinoline (2)
Compound 1 (300 mg, 1.11 mmol) was added to a stirred
suspension of NaH (32 mg, 60% dispersion in mineral oil,
1.33 mmol) in anhydrous DMF (3 mL) at 0 ꢂC. After
20 min, at this temperature, triisopropylsilyl chloride
(0.35 mL, 1.67 mmol) was added dropwise and the stirring
was continued for 3 h. Water was added and the resulting mix-
ture was extracted with CHCl3. The organic layer was washed
with H2O, brine and dried over anhydrous Na2SO4. The sol-
vent was evaporated and the crude product was purified by
preparative TLC plate with CH2Cl2eMeOH (100:1) to afford
2 (280 mg, 59%) as a foam yellow solid, mp 134e135 ꢂC. 1H
NMR (CDCl3): d 8.15 (d, J ¼ 7.7 Hz, 2H, Ar-H), 8.04 (d,
J ¼ 8.6, 16.0 Hz, 1H, CH]CH), 7.78 (d, J ¼ 8.0 Hz, 1H,
Ar-H), 7.75e7.72 (m, 2H, Ar-H), 7.63 (s, 1H, Ar-H), 7.54
3. Materials and methods
3.1. General
All commercial reagents and solvents from Aldrich and
Sigma were used without further purification. [11C]CH3OTf
was prepared according to a literature procedure [23]. Melting

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M. Wang et al. / European Journal of Medicinal Chemistry 44 (2009) 2300e2306
(d, J ¼ 7.6 Hz, 2H, Ar-H), 7.52e7.46 (m, 1H, Ar-H), 7.31e
7.22 (m þ d, J ¼ 15.3 Hz, 3H, Ar-H þ CH]CH), 1.74 (sept,
3H, J ¼ 7.4 Hz, CH), 1.17 (d, J ¼ 7.4 Hz, 18H, 6 ꢀ CH3).
HRMS (CI) m/z calculated for C28H24N2Si ([M]þ),
426.2486; found, 426.2485.
(25 mL) was added pyrrolidine (12 mL). The solution was
heated to reflux overnight. After cooling, the resulting precip-
itate was filtered, and recrystallized from anhydrous EtOH to
1
provide 6 (56 mg, 39%) as a red solid. H NMR (DMSO-
d6): d 12.30 (s, 1H, NH), 8.81 (d, J ¼ 9.1 Hz, 1H, Ar-H),
8.63 (d, J ¼ 16.3 Hz, 1H, CH]CH), 8.59 (d, J ¼ 9.6 Hz,
1H, Ar-H), 8.43 (d, J ¼ 8.9, 1H, Ar-H), 8.37 (s, 1H, Ar-H),
8.26e8.18 (m, 2H, Ar-H), 8.07(t, J ¼ 8.0 Hz, 1H, Ar-H),
7.83 (t, J ¼ 8.0 Hz, 1H, Ar-H), 7.58e7.53 (d þ m,
J ¼ 15.8 Hz, 2H, CH]CH þ Ar-H), 7.31e7.25(m, 2H, Ar-H),
4.45 (s, 3H, NCH3).
3.4. (E )-2-(2,4,6-Trimethoxystyryl)quinoline (3)
A mixture of 2,4,6-trimethoxybenzaldehyde (200 mg,
1.02 mmol), 2-methylquinoline (175 mg, 1.22 mmol), and
a catalytic amount of piperidine (30 mL) in a tube was heated
at 150 ꢂC overnight. After cooling, the resulting solid was trit-
urated with i-PrOH, filtered, and recrystallized from MeOH to
3.8. (E )-1-Methyl-2-(2,4,6-trimethoxystyryl)
quinolinium iodide (E144, 7)
1
afford 3 (141 mg, 43%) as a green solid, mp 99e101 ꢂC. H
NMR (DMSO-d6): d 8.25 (d, J ¼ 8.6 Hz, 1H, Ar-H), 8.00 (d,
J ¼ 16.6 Hz, 1H, CH]CH), 7.93 (d, J ¼ 8.5 Hz, 1H, Ar-H),
7.88 (d, J ¼ 8.9 Hz, 1H, Ar-H), 7.72e7.65 (m, 2H, Ar-H),
7.59 (d, J ¼ 16.6 Hz, 1H, CH]CH), 7.48 (t, J ¼ 7.4 Hz, 1H,
Ar-H), 6.31(s, 2H, Ar-H), 3.90 (s, 6H, 2 ꢀ OCH3), 3.83 (s,
3H, OCH3). HRMS (CI) m/z calculated for C20H19NO3
([M]þ), 321.1359; found, 321.1351.
To a solution of compound 5 (200 mg, 0.70 mmol) and
2,4,6-trimethoxybenzaldehyde (147 mg, 0.75 mmol) in anhy-
drous EtOH (20 mL) was added pyrrolidine (30 mL). The solu-
tion was heated to reflux for 7 h. After cooling, the resulting
precipitate was filtered, and recrystallized from anhydrous
1
EtOH to provide 7 (233 mg, 72%), as a red solid. H NMR
(DMSO-d6): d 8.89 (d, J ¼ 9.0 Hz, 1H, Ar-H), 8.47 (d,
J ¼ 9.0 Hz, 1H, Ar-H), 8.38 (d, J ¼ 9.1 Hz, 1H, Ar-H),
8.33e8.29 (m, 1H, Ar-H), 8.17 (d, J ¼ 15.8 Hz, 1H,
CH]CH), 8.12e8.10 (m, 1H, Ar-H), 7.94 (d, J ¼ 16.4 Hz,
1H, CH]CH), 7.87 (d, J ¼ 7.5 Hz, 1H, Ar-H), 6.38 (s, 2H,
Ar-H), 4.39 (s, 3H, NCH3), 3.97 (s, 6H, 2 ꢀ OCH3), 3.89
(s, 3H, OCH3).
3.5. (E )-4-(2-(6-Methoxyquinolin-2-yl)vinyl)-N,
N-dimethylaniline (4)
A mixture of p-dimethylaminobenzaldehyde (200 mg,
1.34 mmol),
6-methoxy-2-methylquinoline
(278 mg,
1.61 mmol), and a catalytic amount of piperidine (40 mL) in
a tube was heated at 160 ꢂC overnight. After cooling, the re-
sulting solid was triturated with i-PrOH, filtered, and recrystal-
lized from MeOH to afford 4 (130 mg, 32%) as a green solid,
mp 198 ꢂC (dec.). 1H NMR (DMSO-d6): d 8.15 (d, J ¼ 7.3 Hz,
1H, Ar-H), 7.83 (d, J ¼ 8.1 Hz, 1H, Ar-H), 7.72 (d, J ¼ 6.6 Hz,
2H, Ar-H), 7.62 (d, J ¼ 16.6 Hz, 1H, CH]CH), 7.36e7.30
(m, 3H, Ar-H), 7.14 (d, J ¼ 16.1 Hz, 1H, CH]CH), 6.73 (d,
J ¼ 6.8 Hz, 2H, Ar-H), 3.87 (s, 3H, OCH3), 2.94 (s, 6H,
N(CH3)2). HRMS (CI) m/z calculated for C20H20N2O
([M]þ), 304.1570; found, 304.1585.
3.9. 6-Methoxy-1,2-dimethylquinolinium iodide (8)
To a solution of 6-methoxy-2-methylquinoline (3.0 g,
17.32 mmol) in acetonitrile (6 mL) was added iodomethane
(2.70 mL, 43.30 mmol). The solution was heated under reflux
for 2 h. After cooling, the resulting precipitate was filtered,
washed with cooled acetonitrile and dried to yield 8 (3.36 g,
62%) as a yellow solid. 1H NMR (DMSO-d6): d 8.94
(d, J ¼ 8.4 Hz, 1H, Ar-H), 8.54e8.49 (m, 1H, Ar-H), 8.05
(d, J ¼ 8.8 Hz, 1H, Ar-H), 7.88e7.81(m, 2H, Ar-H), 4.42
(s, 3H, NCH3), 3.99 (s, 3H, OCH3), 3.02 (s, 3H, CH3).
3.6. 1,2-Dimethylquinolinium iodide (5)
To a solution of 2-methylquinoline (5.0 g, 34.92 mmol) in
acetonitrile (10 mL) was added iodomethane (4.36 mL,
69.84 mmol). The solution was heated to reflux for 1 h. After
cooling, the resulting precipitate was filtered, washed with
cooled acetonitrile and dried to yield 5 (6.02 g, 60%) as
3.10. (E )-2-(4-(Dimethylamino)styryl)-6-methoxy-1-
methylquinolinium iodide (F22, 9)
To a solution of compound 8 (2.0 g, 6.35 mmol) and p-di-
methylaminobenzaldehyde (1.42 g, 9.53 mmol) in anhydrous
MeOH (20 mL) was added piperidine (0.2 mL). The solution
was heated to reflux for 5 h. After cooling, the resulting pre-
cipitate was filtered, washed with EtOAc and dried to provide
1
a yellow solid. H NMR (DMSO-d6): d 9.11 (d, J ¼ 8.4 Hz,
1H, Ar-H), 8.60 (d, J ¼ 8.8 Hz, 1H, Ar-H), 8.41 (dd, J ¼ 2.5,
8.1 Hz, 1H, Ar-H), 8.28e8.19 (m, 1H, Ar-H), 8.13(d,
J ¼ 8.4 Hz, 1H, Ar-H), 7.80 (t, J ¼ 7.5 Hz, 1H, Ar-H), 4.45
(s, 3H, NCH3), 3.09 (s, 3H, CH3).
1
9 (2.8 g, 99%) as a purple solid. H NMR (DMSO-d6): d 8.69
(d, J ¼ 9.1 Hz, 1H, Ar-H), 8.45 (d, J ¼ 9.3 Hz, 1H, Ar-H), 8.36
(d, J ¼ 9.1 Hz, 1H, Ar-H), 8.11 (d, J ¼ 15.5 Hz, 1H,
CH]CH), 7.80 (d, J ¼ 8.9 Hz, 2H, Ar-H), 7.71e7.67 (m,
2H, Ar-H), 7.50 (d, J ¼ 15.5 Hz, 1H, CH]CH), 6.79 (d,
J ¼ 8.9 Hz, 2H, Ar-H), 4.42 (s, 3H, NCH3), 3.95 (s, 3H,
OCH3), 3.04 (s, 6H, N(CH3)2).
3.7. (E )-2-(2-(1H-Indol-3-yl)vinyl)-1-
methylquinolinium iodide (E36, 6)
To a solution of compound 5 (100 mg, 0.35 mmol) and in-
dole-3-aldehyde (153 mg, 1.05 mmol) in anhydrous EtOH

M. Wang et al. / European Journal of Medicinal Chemistry 44 (2009) 2300e2306
2305
3.11. (E )-2-(2-(1H-Indol-3-yl)vinyl)-1-
radioactivity was assayed and the total volume was noted.
The overall synthesis time was 15e20 min from EOB. The ra-
diochemical yields decay corrected to EOB, from [11C]CO2,
were 50e70%, the radiochemical purity was >99%, and the
chemical purity of the target tracer was >95%. Retention
times in the analytical HPLC system were: tR 3 ¼ 2.60 min,
tR 7 ¼ 1.80 min, tR [11C]7 ¼ 1.80 min; and tR 4 ¼ 2.71 min,
tR 9 ¼ 1.83 min, tR [11C]9 ¼ 1.83 min.
[11C]methylquinolinium triflate ([11C]E36, [11C]6)
[11C]CO2 was produced by the 14N(p,a)11C nuclear reac-
tion in 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). Precursor 2 (0.1e0.3 mg) was dissolved in CH3CN
(300 mL). The mixture was transferred to a small reaction vial.
No-carrier-added (high specific activity) [11C]CH3OTf that
was produced by the gas-phase production method [23] from
[11C]CO2 through [11C]CH4 and [11C]CH3Br with silver tri-
flate (AgOTf) column was passed into the reaction vial, which
was cooled to w0 ꢂC, until radioactivity reached a maximum
(w2 min), and then the reaction mixture was heated at 80 ꢂC
for 2 min. The reaction vessel was connected to a CM Sep-Pak
cartridge. The labeled product mixture solution was passed
onto the Sep-Pak cartridge to release the non-reacted excess
precursor with ethanol and to retain the pure N-[11C-methyl-
ated quaternary ammonium intermediate ([11C]10), on the
same CM Sep-Pak. Then the 11C-labeled intermediate under-
went the deprotecting reaction by addition of a 1.0 M solution
of (n-Bu)4NF in THF (2 mL) to the same cartridge. After
2 min, the Sep-Pak cartridge was washed with ethanol
(5 mL) and water (2 mL) to remove non-reacted (n-Bu)4NF,
and the washing solution was discarded to a waste bottle.
The final product [11C]6 was eluted from the CM Sep-Pak
with saline (2e4 mL) and sterile-filtered through a 0.22 mm
cellulose acetate membrane and collected into a sterile vial.
Total radioactivity was assayed and the total volume was
noted. The overall synthesis time was 20e25 min from
EOB. The radiochemical yields decay corrected to EOB,
from [11C]CO2, were 40e50%, the radiochemical purity was
>99%, and the chemical purity of the target tracer was
>95% measured by analytical HPLC. Retention times in the
analytical HPLC system were: tR 2 ¼ 5.20 min, tR
6 ¼ 1.88 min, tR [11C]6 ¼ 1.88 min.
4. Conclusions
An efficient and convenient synthesis of new carbon-11
labeled styryl dyes has been well-developed. The synthetic
methodology employed classical organic chemistry such as
condensation, protecting and deprotecting, and methylation
reactions to synthesize unlabeled styrylquinoline derivatives.
Carbon-11 labeling at nitrogen position of the precursor
through N-[11C]methylation was incorporated efficiently using
[11C]CH3OTf, a signature reaction of carbon-11 radiochemis-
try from our laboratory. Radiosynthesis produced new probes
in amounts and purity suitable for the preclinical application
in animal studies using PET. Labeled products are suitable
for injection, with the higher specific radioactivities in a range
of 74e111 GBq/mmol at EOS, and can be obtained within
25 min from EOB including fast and efficient SPE purification
and formulation. These chemistry results combined with the
reported imaging data using fluorescent microscopy technol-
ogy encourage further PET imaging evaluation of carbon-11
labeled styryl dyes as new potential probes for imaging of
RNA in living cells.
Acknowledgments
This work was partially supported by the Susan G. Komen
for the Cure grant BCTR0504022, Breast Cancer Research
Foundation, and Indiana Genomics Initiative (INGEN) of Indi-
ana University, which is supported in part by Lilly Endowment
Inc. The authors would like to thank Dr. Bruce H. Mock and
Barbara E. Glick-Wilson for their assistance in production of
[11C]CH3OTf. The referees’ criticisms and editor’s comments
for the revision of the manuscript are greatly appreciated.
3.12. (E )-1-[11C]Methyl-2-(2,4,6-trimethoxystyryl)
quinolinium triflate ([11C]E144, [11C]7) and (E )-2-
(4-(dimethylamino)styryl)-6-methoxy-1-[11C]
methylquinolinium triflate ([11C]F22, [11C]9)
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[11C]7 or [11C]9 was eluted from the CM Sep-Pak with saline
(2e4 mL) and sterile-filtered through a 0.22 mm cellulose
acetate membrane and collected into a sterile vial. Total
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