Synthesis and evaluation of novel 18f labeled 2-pyridinylbenzoxazole and 2-pyridinylbenzothiazole derivatives as ligands for positron emission tomography (PET) imaging of β-amyloid plaques

Article abstract of DOI:10.1021/jm300973k

A series of fluoro-pegylated (FPEG) 2-pyridinylbenzoxazole and 2-pyridinylbenzothiazole derivatives were synthesized and evaluated as novel β-amyloid (Aβ) imaging probes for PET. They displayed binding affinities for Aβ_1-42 aggregates that varied from 2.7 to 101.6 nM. Seven ligands with high affinity were selected for ¹⁸F labeling. In vitro autoradiography results confirmed the high affinity of these radiotracers. In vivo biodistribution experiments in normal mice indicated that the radiotracers with a short FPEG chain (n = 1) displayed high initial uptake into and rapid washout from the brain. One of the 2-pyridinylbenzoxazole derivatives, [¹⁸F]-5-(5-(2-fluoroethoxy)benzo[d]oxazol-2-yl)-N- methylpyridin-2-amine ([¹⁸F]32) (K_i = 8.0 ± 3.2 nM) displayed a brain_2min/brain_60min ratio of 4.66, which is highly desirable for Aβ imaging agents. Target specific binding of [ ¹⁸F]32 to Aβ plaques was validated by ex vivo autoradiographic experiment with transgenic model mouse. Overall, [¹⁸F]32 is a promising Aβ imaging agent for PET and merits further evaluation in human subjects.

Full text of DOI:10.1021/jm300973k

Article
pubs.acs.org/jmc
18
Synthesis and Evaluation of Novel F Labeled
2‑Pyridinylbenzoxazole and 2‑Pyridinylbenzothiazole Derivatives as
Ligands for Positron Emission Tomography (PET) Imaging of
β‑Amyloid Plaques
Mengchao Cui,*,†,∥ Xuedan Wang,†,∥ Pingrong Yu,† Jinming Zhang,†,‡ Zijing Li,† Xiaojun Zhang,‡
Yanping Yang,† Masahiro Ono,§ Hongmei Jia,† Hideo Saji,§ and Boli Liu*,†
†Key Laboratory of Radiopharmaceuticals, Ministry of Education, College of Chemistry, Beijing Normal University, Beijing 100875, P.
R. China
‡Department of Nuclear Medicine, Chinese PLA General Hospital, Beijing 100853, P. R. China
§Department of Patho-Functional Bioanalysis, Graduate School of Pharmaceutical Sciences, Kyoto University, 46-29 Yoshida
Shimoadachi-cho, Sakyo-ku, Kyoto 606-8501, Japan
S
* Supporting Information
ABSTRACT: A series of fluoro-pegylated (FPEG) 2-
pyridinylbenzoxazole and 2-pyridinylbenzothiazole derivatives
were synthesized and evaluated as novel β-amyloid (Aβ)
imaging probes for PET. They displayed binding affinities for
Aβ1−42 aggregates that varied from 2.7 to 101.6 nM. Seven
ligands with high affinity were selected for 18F labeling. In vitro
autoradiography results confirmed the high affinity of these
radiotracers. In vivo biodistribution experiments in normal
mice indicated that the radiotracers with a short FPEG chain
(n = 1) displayed high initial uptake into and rapid washout
from the brain. One of the 2-pyridinylbenzoxazole derivatives, [18F]-5-(5-(2-fluoroethoxy)benzo[d]oxazol-2-yl)-N-methylpyr-
idin-2-amine ([18F]32) (Ki = 8.0 3.2 nM) displayed a brain2min/brain60min ratio of 4.66, which is highly desirable for Aβ imaging
agents. Target specific binding of [18F]32 to Aβ plaques was validated by ex vivo autoradiographic experiment with transgenic
model mouse. Overall, [18F]32 is a promising Aβ imaging agent for PET and merits further evaluation in human subjects.
crucial step that ultimately leads to AD.1−5 Therefore, the
INTRODUCTION
■
development of novel imaging agents specifically targeting Aβ
Alzheimer’s disease (AD) is the most common form of
plaques may lead to early diagnosis of AD and monitoring of
dementia accounting for between 50% and over 70% of all cases
the effectiveness of novel therapies for this devastating
among elderly people and is becoming an extensive health
disease.6−8
problem with the ever-increasing aging population. Clinically,
In past decades, a number of radiolabeled Aβ imaging agents
AD is a lethal, progressive neurodegenerative disorder that
based on highly conjugated Aβ specific dyes, such as Congo
leads to a decline in memory and many cognitive deficits, such
Red (CR) and thioflavin T (Th-T), have been developed and
as irreversible memory loss, impaired judgment, and problems
reported for single photon emission computed tomography
with language. At present there are no treatments available to
(SPECT) and positron emission tomography (PET),9,10 some
reverse or halt the progression of the disease. Although some
of which have been tested in humans (Figure 1). Among
drugs may provide temporary relief from symptoms, the benefit
dozens of radioiodinated liagnds, [123I]-6-iodo-2-(4′-
dimethylamino)phenylimidazo[1,2]pyridine ([123I]1, IMPY), a
is limited. The best hope for progress might be to detect the
disease at an early stage and initiate treatment before the brain
damage is widespread. However, clinical diagnosis of AD is very
unique Th-T derivative with a [1,2,a]imidazopyridine ring, was
the only SPECT tracer tested in humans. However, the signal-
to-noise ratio for plaque labeling of [123I]1 in AD and healthy
difficult, and only possible or probable AD can be routinely
diagnosed, while a definite diagnosis can only be made on
controls was not robust, while it was believed that the in vivo
instability and fast metabolism of [123I]1 may ultimately lead to
biopsy and post-mortem brain tissue to confirm the existence of
β-amyloid (Aβ) plaques and neurofibrillary tangles (NFTs).
Although the etiology of AD has not been definitively
established, the amyloid cascade hypothesis is the most
Special Issue: Alzheimer's Disease
prevailing theory to explain the pathogenesis of AD, which
Received: July 6, 2012
posits that the deposition of the Aβ peptide in the brain is a
© XXXX American Chemical Society
A
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Figure 1. Structures of Aβ imaging agents for SPECT and PET that have been evaluated in human subjects.
a decreasing of signal.11−13 After that, there is no report of Aβ
imaging candidate for SPECT moving into clinical trial. In
contrast, the development of radiotracers for PET imaging was
more successful. [11C]-2-(4-(Methylamino)phenyl)-6-hydrox-
ybenzothiazol ([11C]2, PIB), to date, is the most suitable and
extensively studied PET radioligand for amyloid imaging and
can distinguish AD from control cases clearly.14−16 Recently,
Andersson et al. reported a close analogue of [11C]2, [11C]-2-
[6-(methylamino)pyridin-3-yl]-1,3-benzothiazol-6-ol ([11C]3,
AZD2184), by replacing the phenyl group with a pyridyl
group, which displayed a decreased nonspecific binding in
white matter.17,18 Additionally, the stilbene scaffold, which
could be considered as a more abbreviated form of CR, has also
been selected for developing Aβ imaging agents. Kung et. al
reported the first 11C-labeled stilbene derivative, [11C]-4-N-
methylamino-4′-hydroxystilbene ([11C]6, SB-13). Initial PET
imaging in vivo with [11C]6 demonstrated potential usefulness
in detecting Aβ plaques in the human brain.19,20 However, the
short half-life of 11C (T1/2 = 20 min) of the 11C-labeled tracers
limits their usefulness for widespread clinical application. Thus,
great efforts have been directed to developing imaging agents
labeled with 18F (T1/2 = 110 min). Two 18F-labeled PIB
analogues, [18F]-2-(3-fluoro-4-methyaminophenyl)-
benzothiazol-6-ol ([18F]4, GE-067)21 and [18F]-2-(2-fluoro-6-
(methylamino)pyridin-3-yl)benzofuran-6-ol ([18F]5,
AZD4694),22 are currently under phase II clinical testing in
Europe. Meanwhile, two 18F-labeled stilbene derivatives with a
short length of polyethylene glycol (PEG) units, [18F]-4-(N-
methylamino)-4′-(2-(2-(2-fluoroethoxy)ethoxy)ethoxy)stilbene
([18F]7, BAY94-9172)23 and [18F]-(E)-4-(2-(6-(2-(2-(2-
fluoroethoxy)ethoxy)ethoxy)pyridin-3-yl)vinyl)-N-methylani-
line ([18F]8, AV-45),24−26 are under commercial development.
More recently, [18F]8, brand-named Amyvid by Eli Lilly, has
been approved by the U.S. Food and Drug Administration
(FDA).
glycol chain with a fluorine atom [fluoropolyethylene glycol
(FPEG)], which will provide a flexible tool to adjust
lipophilicity and to maintain a relatively small size;28 (2)
displacing one of the benzene rings of ligand with one pyridine
ring. Previous work indicates that the in vivo pharmacokinetics
of the fluoro-pegylated pyridylbenzofuran derivatives ([18F]11,
[18F]12)29,30 were greatly improved compared with the
phenylbenzofuran derivatives ([18F]9, [18F]10).31 Tracer
[18F]12 displayed the best pharmacokinetics of radioactivity
in the brain, with the brain2min/brain60min ratio being 2.34. Kung
et. al also reported a series of fluoro-pegylated phenyl-
benzothiazole derivatives ([18F]15−17) with improved proper-
ties compared with the original radioiodinated ligand28 (Figure
2).
Figure 2. (A) Structures of radiolabeled benzofuran, benzoxazole, and
benzothiazole derivatives. (B) Structures of the designed 18F-labeled 2-
pyridinylbenzoxazole and 2-pyridinylbenzothiazole derivatives.
Despite the fact that great success has been achieved on 18F-
labeled tracers for amyloid imaging, it is clear that all 18F-
labeled tracers show high levels of nonspecific white matter
retention than [11C]2. In AD patients, the average cortical
binding is similar to or less than the uptake in white matter and
may limit the sensitivity of PET imaging.27 The mechanism of
white matter retention seems to be owing to nonspecific
binding that to some degree is governed by the lipophilicity of
the ligand.22 Thus, further modifications in order to decrease
lipophilicity are needed. Two approaches were used to decrease
the lipophilicity of 18F-labeled ligand: (1) introducing a short
length of PEG (n = 2−12) and capping the end of the ethylene
In an attempt to further explore more promising 18F-labeled
tracers, we extended our fluoro-pegylation approach from
benzofuran to benzoxazole. The strategy of FPEG was
successfully applied to the benzoxazole scaffold, and it has
been demonstrated that the fluoro-pegylated phenylbenzox-
azole derivatives ([18F]13, [18F]14) showed more promising in
vivo kinetics than the corresponding radioiodinated ligand and
the benzofuran derivatives.32 Tracer [18F]13 with a mono-
methylamino group exhibited high affinity for Aβ aggregates (Ki
= 9.3 nM) and high initial uptake into the brain and displayed
excellent binding to Aβ plaques in ex vivo autoradiographic
experiments with Tg2576 mice. What’s more, small-animal
B
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Scheme 1
a
Reagents and conditions: (a) TsCl, KOH, CH2Cl2, 0 °C to rt; (b) TBAF (1 M in THF), THF, reflux; (c) Pd/C, MeOH, rt; (d) 2-amino-5-
(trifluoromethyl)pyridine, NaOH (1 M), H2O, reflux; (e) (1) NaOMe, MeOH, (CH2O)n, reflux; (2) NaBH4, rt; (f) NaBH3CN, (CH2O)n,
CH3COOH, rt; (g) BBr3 (1 M in CH2Cl2), CH2Cl2, −78 °C to rt; (h) K2CO3, DMF, 110 °C; (i) 1-bromo-2-fluoroethane, K2CO3, DMF, 110 °C;
(j) KOH, H2O, reflux.
a
Scheme 2
a
Reagents and conditions: (a) K2CO3, 18-crown-6, acetone, reflux; (b) (Boc)2O, THF, reflux.
PET studies demonstrated significant differences between
Tg2576 and wild-type mice in the clearance profiles after the
administration of [18F]13. Although the preliminary results
indicate that [18F]13 is a promising PET tracer for imaging Aβ
plaques, the brain2min/brain60min ratio of [18F]13 (2.67) was still
lower than that of [18F]8 (3.90). To further improve the
pharmacokinetic profile of the fluoro-pegylated phenylbenzo-
thiazole and phenylbenzoxazole derivatives, we decided to use
the strategy of bioisosteric replacement by displacing the
phenyl group with a pyridyl group to reduce nonspecific
binding in white matter and enhance the signal-to-noise ratio.
We report herein the synthesis and the initial biological
evaluation of fluoro-pegylated pyridylbenzothiazole and pyr-
idylbenzoxazole derivatives as novel PET imaging probes
targeting Aβ plaques in the brain.
RESULTS AND DISCUSSION
■
Chemistry. The synthesis of fluoro-pegylated pyridylbenzo-
thiazole and pyridylbenzoxazole derivatives is outlined in
Scheme 1. The intermediates 20 and 21 were obtained by
ready fluorination of the oligoethylene glycol ditosylate ester
(18, 19) using anhydrous tetra-n-butylammonium fluoride in
THF. The desired 2-aminopyridylbenzothiazole and 2-amino-
pyridylbenzoxazole core structures (23 and 35) were obtained
by using a previously reported one-step reaction between 5-
(trifluoromethyl)pyridin-2-amine and 2-amino-4-methoxyphe-
nol (22) or 2-amino-5-methoxybenzenethiol (34) in aqueous
NaOH solution with excellent yields (89 and 50%,
respectively).33 Conversion of 23 and 35 to the monomethy-
lamino derivatives 24 and 36 was achieved by the
monomethylation of the amino group by using a method
C
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Scheme 3
a
Reagents and conditions: (a) 18F−, K2CO3, Kryptofix 222, acetonitrile, 100 °C; (b) (1) 18F−, K2CO3, Kryptofix 222, acetonitrile, 100 °C; (2) HCl
(1 M), 100 °C.
previously reported in yields of 98% and 98%, respectively.34
The dimethylamino derivatives 25 and 37 were prepared by an
efficient dimethylation method with paraformaldehyde, sodium
cyanoborohydride, and acetic acid (89% and 91%, respectively).
The O-methyl groups of 24, 25, 36, and 37 were removed by
reacting with BBr3 in CH2Cl2 to give 26, 27, 38, and 39 in
yields of 97%, 89%, 88%, and 68%, respectively. Then the
hydroxy groups were coupled with 1-bromo-2-fluoroethane to
give the fluorinated derivatives (32, 33, 44, and 45) with one
ethoxy unit as the PEG linkage. The corresponding fluorinated
derivatives with two or three ethoxy units (28−31 and 40−43)
were prepared by the hydroxyl compounds with K2CO3 and the
intermediate 20 or 21 in DMF (yield, 40−75%).
the retention time with that of the nonradioactive compound
(see Supporting Information) (Scheme 3).
In Vitro Binding Studies. The affinities of these fluoro-
pegylated pyridylbenzothiazole and pyridylbenzoxazole deriva-
tives (28−33 and 40−45) for Aβ1−42 aggregates were examined
with competition binding assays using [125I]1 as the competing
radioligand. IMPY and PIB were also screened using the same
system for comparison. As shown in Table 1, all ligands with a
Table 1. Inhibition Constants of Fluorinated 2-
Pyridinylbenzoxazole and 2-Pyridinylbenzothiazole
Derivatives for the Binding of [125I]IMPY to Aβ1‑42
Aggregates
The synthesis of tosylate precursors is outlined in Scheme 2.
The free hydroxy groups present in 26, 27, 38, and 39 were
coupled with ethane-1,2-diyl bis(4-methylbenzenesulfonate) or
19 in acetone catalyzed by 18-crown-6 to give 46−49 and 52−
54 (yield, 28−49%). For the monomethylated precursors, the
methylamino groups of intermediates 48, 49, 53, and 54 were
protected by a butyloxycarbonyl (BOC) group to give the
tosylate precursors 50, 51, 55, and 56 in yields of 67%, 23%,
80%, and 63%, respectively. Compared with the synthesis
routes for the precursors of benzofuran and benzoxazole
derivatives, the procedures reported here are much simpler,
while some protection and deprotection steps were not needed.
Radiolabeling. To make the desired 18F-labeled dimethy-
lamino tracers [18F]33, [18F]31, and [18F]43, tosylate
precursors 46, 47, and 52 were reacted with [18F]fluoride/
potassium carbonate and Kryptofix 222 in acetonitrile with
radiochemical yields of 53%, 57%, and 46%, respectively. For
the monomethylamino tracers [18F]32, [18F]29, [18F]44, and
[18F]41, the N-Boc-protected tosylates 50, 51, 55, and 56 were
employed as the precursors. After [18F]fluorination, the mixture
was treated with aqueous HCl to remove the N-BOC-
protecting group (radiochemical yields, 41%, 21%, 33%, and
27%, respectively). The radiochemical purity of these radio-
tracers was greater than 98% after purification by high
performance liquid chromatography (HPLC), and their specific
activity was estimated as approximately 200 GBq/μmol. The
identity of 18F-labeled tracers was verified by a comparison of
a
compd
Ki SEM (nM)
28
101.6 15.3
76.9 15.5
7.3 0.8
9.9 0.5
8.0 3.2
2.7 0.7
29.7 7.1
9.3 0.1
4.6 0.6
5.8 1.8
10.1 2.4
2.7 0.6
29
30
31
32
33
40
41
42
43
44
45
b
IMPY
PIB
10.5 1.0
b
9.0 1.3
a
The Ki values were determined in three independent experiments (n
= 3) unless otherwise noted. Data from ref 30.
b
dimethylamino group (30, 31, 33, 42, 43, and 45) displayed
high binding affinity to Aβ1−42 aggregates (Ki < 10.0 nM).
Similar to the findings reported previously,30 the tertiary N,N-
dimethylamino analogues were found to have higher affinities
than their secondary N-methylamino analogues, especially for
the pyridylbenzoxazole derivatives (e.g., 28 vs 30, 29 vs 31, 32
vs 33). The length of the FPEG chain did not bring about an
appreciable progress in the binding affinity to amyloid
D
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Figure 3. In vitro autoradiography of 18F-labeled 2-pyridinylbenzoxazole derivatives ([18F]29, [18F]31, [18F]32, and [18F]33) on a Tg model mouse
(C57BL6, APPswe/PSEN1, 12 months old, male) (A, E, I, M) and normal control mouse (C57BL6, 12 months old, male) (C, G, K, O). The
presence and distribution of plaques in the sections were confirmed by fluorescence staining using thioflavin S on the same sections with a filter set
for GFP (B, D, F, H, J, L, N, P).
Figure 4. In vitro autoradiography of 18F-labeled 2-pyridinylbenzothiazole derivatives ([18F]41, [18F]43, and [18F]44) on a Tg model mouse
(C57BL6, APPswe/PSEN1, 12 months old, male) (A, E, I) and normal control mouse (C57BL6, 12 months old, male) (C, G, K). The presence and
distribution of plaques in the sections were confirmed by fluorescence staining using thioflavin S on the same sections with a filter set for GFP (B, D,
F, H, J, L).
aggregates for the pyridylbenzothiazole derivatives (40−45).
However, the length of the FPEG chain is very important for
the secondary N-methylamino analogues of pyridylbenzoxazole.
The binding affinities were reduced dramatically with the
addition of the FPEG chain [e.g., 28 (Ki = 101.6 15.3 nM)
and 29 (Ki = 76.9 15.5 nM)], while 32 retained high binding
affinity for the n = 1 FPEG conjugates (Ki = 8.0 3.2 nM). On
the basis of the high binding affinity to Aβ1−42 aggregates,
ligands 29, 31, 32, 33, 41, 43, and 44 were selected for 18F
labeling.
In Vitro Autoradiography Studies. In vitro autoradio-
graphic studies of the 18F-labeled tracers were first performed
with sections of Tg mice (C57BL6, APPswe/PSEN1, 12
months old) and an age-matched control mice. As shown in
Figures 3 and 4, all of the 18F-labeled tracers displayed excellent
labeling of Aβ plaques in the hippocampus and cortical regions
of the Tg mice and lack of any notable Aβ labeling in control
mice. The distribution of Aβ plaques was consistent with the
results of fluorescent staining with thioflavin S. A previous
report suggested that the configuration/folding of Aβ plaques
in Tg mice might be different from the tertiary/quaternary
structure of Aβ plaques in AD brains.35 Thus, the binding
information for Aβ plaques in human AD brain is important.
Tracers [18F]29, [18F]32, [18F]41, and [18F]44 with a
monomethylamino group were selected for autoradiographic
studies in sections of human brain tissue. As shown in Figure 5,
a highly dense labeling of plaques was observed in AD brain
sections. In contrast, no apparent labeling was observed in
normal adult brain sections. Although the binding affinity of
[18F]29 to Aβ aggregates was not potent (Ki = 76.9
15.5
nM), [18F]29 was still able to label the plaques in sections of Tg
mice and AD.
In Vivo Biodistribution Studies. Biodistribution study of
the radiolabeled tracers in normal mice is often used as a test to
measure the initial brain uptake [ability to penetrate intact
blood−brain barrier (BBB)] and washout kinetics from the
normal brain. The time−activity data on the brain permeation
as well as the other organ distribution of the 18F-labeled tracers
acquired in normal ICR mice after intravenous administration
are summarized in Tables 2−4. [18F]32, [18F]33, and [18F]44
E
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
The high initial uptakes in the liver and the kidneys were
followed by a moderate washout, while the uptake in the
intestines showed an accumulation of radioactivity over time. In
summary, two of the 18F-labeled pyridylbenzoxazole derivatives
([18F]32 and [18F]33) meet the preliminary requirements for a
potential PET imaging agent for in vivo detection of Aβ plaque.
Compared with the 18F-labeled pyridylbenzofuran, phenyl-
benzoxazole, and phenylbenzothiazol derivatives, the 18F-
labeled pyridylbenzoxazole derivatives had greatly improved
pharmacokinetics. In addition, the brain2min/brain60min ratio of
[18F]32 and [18F]33 was superior than that of [18F]8 (3.90).
Ex Vivo Autoradiography Studies. A previous report
suggested that a rapid metabolism in vivo of the dimethylamino
group in the tracer may occur,36 and most of the radiotracers
for Aβ plaques in clinical trials have a monomethylamino group.
For the same reason, [18F]32 was selected for further ex vivo
autoradiography studies in Tg mice (C57BL6, APPswe/
PSEN1, 10 months old) and wild-type mice (C57BL6, 10
months old). As shown in Figure 6A and Figure 6C, the Aβ
plaques in the cortical regions of Tg mice were clearly labeled
by [18F]32, while wild-type mouse brain showed no such
labeling (Figure 6E). The labeling of the plaques was confirmed
by costaining with thioflavin S on the same sections (Figure 6B
and Figure 6D).
In Vitro Stability Studies. In vitro stability study of [18F]
32 in saline solution indicated that [18F]32 solutions were
stable during storage for at least 6 h at room temperature.
Moreover, [18F]32 was found to have high in vitro stability in
mice plasma; more than 95% of the activity was identified as the
intact tracer after 60 min of incubating with mice plasma at 37
°C (see Supporting Information).
Figure 5. In vitro autoradiography of [18F]29 (A), [18F]32 (C), [18F]
41 (E), and [18F]44 (G) on human brain sections: (A, C, E, G) post-
mortem AD cases; (B, D, F, H) normal control.
CONCLUSIONS
■
A series of fluoro-pegylated (FPEG) 2-pyridinylbenzoxazole
and 2-pyridinylbenzothiazole derivatives had been successfully
prepared and evaluated as PET imaging tracers for Aβ plaques.
In binding studies, they displayed binding affinities for Aβ
aggregates that varied from 2.7 to 101.6 nM. Seven ligands with
high affinity (29, 31, 32, 33, 41, 43, and 44) were selected for
18F labeling. In vitro autoradiography with sections of post-
mortem AD brain and Tg mouse brain confirmed the affinity of
these tracers. In biodistribution experiments using normal mice,
the radiotracers with a short FPEG chain (n = 1) displayed high
initial uptake into and rapid washout from the brain in normal
mice. However, the BBB penetration ability of the 18F-labeled
tracers with a long FPEG chain (n = 3) was not robust.
Compared with the 18F-labeled pyridylbenzofuran, phenyl-
benzoxazole, and phenylbenzothiazol derivatives, the 18F-
labeled pyridylbenzoxazole derivatives had greatly improved
pharmacokinetics. On the basis of the above results, one of the
with a short FPEG chain (n = 1) displayed high uptakes into
the brain (7.23, 7.27, and 7.87% ID/g, respectively) at 2 min
postinjection and fast washout kinetics from the healthy brain
(1.55, 1.47, and 2.83% ID/g at 60 min, respectively). These
values were comparable to those reported for the 18F-labeled
pyridylbenzofuran ([18F]10, [18F]12)30 and phenylbenzoxazole
([18F]13)32 derivatives as well as [18F]7 and [18F]8, which are
under commercial development. The ratio brain2min/brain60min
is considered as an important index with which to select tracers
with appropriate kinetics in vivo. These three tracers showed
brain2min/brain60min ratios of 4.66, 4.95, and 2.78, respectively,
indicating that the pyridylbenzoxazole scaffold is more potent
than pyridylbenzothiazole. However, the BBB penetration
ability of the 18F-labeled tracers with a long FPEG chain (n =
3) ([18F]29, [18F]31, [18F]41, and [18F]43) was decreased
(4.05, 3.79, 2.23, and 3.26% ID/g at 2 min, respectively). The
FPEG strategy was used to modulate the lipophilicity of 18F-
labeled tracers. As reflected in log D values, the lipophilicity of
the tracers with a long FPEG chain decreased slightly, which
may explain the lower brain uptake. Moreover, the flexiblility
and polar properties of the long FPEG chain may also interfere
and limit brain penetration.28 It was also observed that in vivo
defluorination was likely occurring with [18F]31 and [18F]44
because the bone uptakes increased with time (6.39 and 6.70%
dose/g at 1 h, respectively). However, they were minimal
compared to other 18F-labeled tracers, so the interference with
the imaging is expected to be relatively minor. These 18F-
labeled tracers were also distributed to several other organs.
2-pyridinylbenzoxazole derivatives, [18F]32 (Ki = 8.0
3.2
nM), displayed a brain2min/brain60min ratio of 4.66, which is
highly desirable for Aβ imaging agents. Target specific binding
of [18F]32 to Aβ plaques was validated by ex vivo autoradio-
graphic experiments with Tg mouse. Furthermore, [18F]32
displayed high in vitro stability in saline solution and mice
plasma. Overall, [18F]32 is a promising PET imaging agent for
cerebral Aβ plaques and merits further evaluation in human
subjects.
EXPERIMENTAL SECTION
General Remarks. All reagents used in the synthesis were
commercial products and were used without further purification
■
F
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Table 2. Biodistribution in Normal ICR Mice after Intravenous Injection of the 18F-Labeled 2-Pyridinylbenzoxazole Tracers
organ
blood
2 min
10 min
30 min
60 min
[18F]29 (log D = 2.86 0.12)
5.07 0.41
4.05 0.36
5.50 0.70
14.22 2.65
4.13 0.32
5.30 0.31
8.51 0.97
5.08 0.98
10.81 0.86
1.75 0.25
3.04 0.29
3.16 0.26
2.06 0.29
3.16 0.28
6.47 0.72
2.18 0.11
2.81 0.24
3.73 0.20
12.93 1.25
28.65 2.66
2.46 0.33
2.74 0.24
1.78 0.21
2.50 0.27
4.21 0.66
1.79 0.16
2.34 0.19
2.54 0.33
12.02 1.82
25.16 0.97
3.93 0.48
brain
2.05 0.27
heart
3.35 0.37
liver
9.32 0.79
spleen
lung
2.64 0.35
3.15 0.46
kidney
stomach
4.79 0.57
b
b
12.32 0.85
22.90 1.90
1.92 0.65
intestine
bone
[18F]31 (log D = 3.26 0.04)
blood
brain
heart
liver
5.47 0.56
3.79 0.41
5.24 0.63
13.25 3.13
3.00 0.63
3.03 0.63
8.27 2.03
2.66 0.60
8.13 0.79
1.79 0.36
3.05 0.25
2.74 0.14
1.85 0.06
2.97 0.18
7.24 2.05
2.41 0.10
2.64 0.29
4.20 0.42
9.26 2.63
28.15 3.11
4.71 0.34
2.87 0.12
1.82 0.07
2.73 0.13
3.46 1.52
2.34 0.26
1.87 0.66
3.21 0.46
6.45 1.79
28.54 4.18
6.39 0.57
1.97 0.14
3.51 0.11
5.41 0.67
spleen
lung
2.69 0.65
2.34 0.66
kidney
5.14 0.30
b
stomach
intestine
bone
5.10 0.80
b
14.45 2.65
2.70 0.47
[18F]32 (log D = 3.52 0.13)
blood
brain
heart
liver
3.36 0.15
7.23 0.04
4.66 0.34
10.48 1.65
3.32 0.50
4.14 0.24
7.45 0.35
3.24 0.62
9.87 0.97
1.18 0.21
2.55 0.15
2.55 0.17
1.69 0.17
1.84 0.19
3.95 0.66
1.63 0.28
2.29 0.09
2.17 0.22
3.11 0.94
33.66 5.26
2.34 0.88
3.20 0.48
1.55 0.17
2.05 0.21
2.64 0.86
1.64 0.13
2.57 0.40
3.04 0.99
3.89 1.58
39.05 6.70
2.37 0.86
2.24 0.12
2.34 0.17
8.50 0.72
spleen
lung
2.13 0.24
2.80 0.48
kidney
3.52 0.52
b
b
stomach
5.59 1.45
intestine
bone
19.61 2.42
1.11 0.36
[18F]33 (log D = 3.53 0.04)
2.05 0.10
blood
brain
heart
liver
3.57 0.39
7.27 0.19
5.95 0.95
15.70 2.71
3.22 0.52
5.53 0.50
8.48 0.77
2.44 0.76
8.41 1.83
2.08 0.73
1.89 0.17
1.76 0.16
1.83 0.24
4.39 0.80
1.60 0.14
1.85 0.46
2.44 0.59
2.54 0.48
38.88 2.94
1.56 0.53
2.06 0.26
1.47 0.25
1.93 0.21
3.19 0.61
1.32 0.26
1.79 0.38
1.82 0.34
2.79 0.57
37.02 6.37
1.88 0.39
3.41 0.36
2.44 0.18
10.72 1.44
2.02 0.25
spleen
lung
2.58 0.31
kidney
3.82 0.37
b
stomach
intestine
bone
2.55 0.82
b
21.45 6.22
1.27 0.15
a
b
Expressed as % injected dose per gram unless otherwise indicated. Data are the average for five mice standard deviation. Expressed as % injected
dose per organ.
unless otherwise indicated. The 1H NMR spectra were obtained at 400
MHz on Bruker spectrometer in CDCl3 or DMSO-d6 solutions at
room temperature with TMS as an internal standard. Chemical shifts
were reported as δ values relative to the internal TMS. Coupling
constants were reported in hertz. Multiplicity is defined by s (singlet),
d (doublet), t (triplet), and m (multiplet). Mass spectra were acquired
under a SurveyorMSQ Plus (ESI) instrument. Radiochemical purity
was determined by HPLC performed on a Shimadzu system SCL-20
AVP equipped with a SPD-20A UV detector (λ = 254 nm) and
Bioscan flow count 3200 NaI/PMT γ-radiation scintillation detector.
HPLC separations were achieved on a Venusil MP C18 reverse phase
column (Agela Technologies, 5 μm, 10 mm × 250 mm), and elution
was with a binary gradient system at a flow rate of 4.0 mL/min. HPLC
analyses were achieved on a Venusil MP C18 reverse phase column
(Agela Technologies, 5 μm, 4.6 mm × 250 mm), and elution was with
a binary gradient system at a flow rate of 1.0 mL/min. Mobile phase A
was water, while mobile phase B was acetonitrile. The purity of the
synthesized key compounds (28−33 and 40−45) was determined
using analytical HPLC and was found to be more than 96%.
Fluorescent observation was performed by the Oberver Z1 (Zeiss,
Germany) equipped with GFP filter set (excitation, 505 nm). Normal
ICR mice (5 weeks, male) were used for biodistribution experiments.
Transgenic mice (C57BL6, APPswe/PSEN1, 12 months old), used as
an Alzheimer’s model, were purchased from the Institute of Laboratory
Animal Science, Chinese Academy of Medical Sciences. All protocols
requiring the use of mice were approved by the Animal Care
Committee of Beijing Normal University.
Chemistry. 2,2′-Oxybis(ethane-2,1-diyl) Bis(4-methylbenze-
nesulfonate) (18). Intermediate 18 was synthesized according to the
literature.37 Briefly, to a solution of tosyl chloride (19.8 g, 10 mmol)
G
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Table 3. Biodistribution in Normal ICR Mice after Intravenous Injection of the 18F-Labeled 2-Pyridinylbenzothiazole Tracers
organ
blood
2 min
10 min
30 min
60 min
[18F]41 (log D = 2.85 0.07)
6.92 0.60
2.23 0.33
3.53 0.36
15.22 2.34
2.68 0.35
5.53 0.26
6.02 0.98
2.96 0.62
5.69 0.98
1.19 0.42
4.25 0.32
3.49 0.11
1.31 0.17
1.96 0.13
8.16 0.46
1.74 0.07
3.11 0.06
3.12 0.25
11.46 3.60
21.68 2.74
1.33 0.32
2.94 0.19
1.26 0.15
1.85 0.13
4.12 0.65
1.49 0.15
1.93 0.43
2.66 0.23
8.78 1.46
18.60 4.73
1.42 0.17
brain
1.63 0.15
heart
2.16 0.16
liver
9.38 0.90
spleen
lung
2.05 0.24
3.36 0.50
kidney
stomach
4.04 0.39
b
b
9.44 1.92
intestine
bone
10.06 0.62
0.86 0.11
[18F]43 (log D = 3.08 0.01)
blood
brain
heart
liver
9.79 0.81
3.26 0.21
4.50 0.67
12.90 0.62
3.14 0.32
6.69 0.53
6.82 0.61
2.55 0.54
4.68 0.23
1.86 0.27
5.44 0.16
4.96 0.87
1.98 0.20
2.62 0.11
6.79 0.80
2.59 0.55
3.28 0.95
3.88 0.89
14.13 0.61
19.57 1.68
1.64 0.28
3.79 0.37
1.84 0.22
2.02 0.98
4.75 0.39
1.47 0.77
2.33 0.94
3.01 0.61
11.34 0.98
26.66 2.90
1.58 0.28
2.64 0.19
3.30 0.17
11.60 0.71
2.97 0.23
spleen
lung
4.59 0.34
kidney
5.45 0.32
b
stomach
intestine
bone
8.98 0.14
b
10.4 1.27
0.95 0.48
[18F]44 (log D = 3.59 0.02)
blood
brain
heart
liver
4.31 0.34
7.87 0.48
5.59 0.69
10.40 0.83
4.52 0.46
5.85 0.68
9.57 0.68
3.61 0.45
12.06 0.35
2.04 0.33
3.94 0.46
4.45 0.28
3.38 0.25
3.33 0.43
4.28 0.47
2.87 0.23
3.57 0.21
3.62 0.19
4.63 0.69
25.23 3.23
3.77 0.97
4.45 0.51
2.83 0.15
3.70 0.43
3.06 0.55
2.86 0.51
2.72 0.59
3.45 0.88
3.67 1.11
19.00 4.14
6.70 1.21
4.54 0.37
3.11 0.81
5.94 0.97
spleen
lung
3.29 0.75
3.02 0.60
kidney
4.42 0.53
b
b
stomach
5.61 0.81
intestine
bone
11.15 1.30
2.43 0.60
a
b
Expressed as % injected dose per gram unless otherwise indicated. Data are the average for five mice standard deviation. Expressed as % injected
dose per organ.
Table 4. Comparison of Inhibition Constants (Ki, nM) and Brain Kinetics between 18F-Labeled Tracers in This Work and Other
Tracers
a
a
compd
Ki (nM)
2 min
60 min
brain2min/brain60min
b
[18F]7
6.7 0.3
2.87 0.17
2.0 0.5
3.9 0.2
1.0 0.2
2.4 0.1
9.5 1.3
3.9 0.2
2.2 0.5
3.8 0.5
4.7 0.9
76.9 15.5
9.9 0.5
8.0 3.2
2.7 0.7
9.3 0.1
5.8 1.8
8.14 2.03
7.33 1.54
2.88 0.46
8.18 0.59
5.16 0.30
7.38 0.84
8.12 0.51
5.29 0.19
10.27 1.30
5.53 0.56
2.57 0.12
4.05 0.36
3.79 0.41
7.23 0.04
7.27 0.19
2.23 0.33
3.26 0.21
7.87 0.48
2.60 0.22
1.88 0.14
2.80 0.06
3.87 0.42
2.44 0.36
3.15 0.10
3.04 0.16
2.12 0.08
3.94 0.04
2.18 0.09
1.80 0.25
1.78 0.21
1.82 0.07
1.55 0.17
1.47 0.25
1.26 0.15
1.84 0.22
3.13
3.90
1.03
2.11
2.11
2.34
2.67
2.50
2.61
2.54
1.43
2.28
2.08
4.66
4.95
1.77
1.77
2.78
c
[18F]8
d
[18F]11
[18F]12
[18F]13
[18F]14
[18F]16
[18F]17
[18F]19
[18F]20
[18F]21
[18F]29
[18F]31
[18F]32
[18F]33
[18F]41
[18F]43
[18F]44
d
d
d
e
e
f
f
f
10.1 2.4
2.83 0.15
a
b
c
d
e
f
Expressed as % injected dose per gram. Data from ref 40. Data from ref 25. Data from ref 30. Data from ref 32. Data from ref 28.
H
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
(s, 3H). MS (ESI): m/z calcd for C13H11N3O2 241.09; found 241.9 (M
+ H)+.
5-(5-Methoxybenzo[d]oxazol-2-yl)-N-methylpyridin-2-
amine (24). To a mixture of 23 (480 mg, 2 mmol) and
paraformaldehyde (240 mg, 8 mmol) in MeOH (20 mL) was added
CH3ONa (1.08 g, 20 mmol). The mixture was stirred under reflux for
1 h. After the mixture was cooled, NaBH4 (160 mg, 4 mmol) was
added, and the mixture was brought to reflux again for 2 h. The
reaction mixture was poured onto ice−water, and the precipitate was
collected by filtration to obtain 24 as a white solid (500 mg, 98.0%).
Mp: 178.6−179.8 °C. 1H NMR (400 MHz, DMSO-d6): δ 8.78 (d, J =
1.9 Hz, 1H), 8.04 (d, J = 8.5 Hz, 1H), 7.59 (d, J = 8.8 Hz, 1H), 7.35
(d, J = 4.5 Hz, 1H), 7.25 (d, J = 2.3 Hz, 1H), 6.91 (dd, J = 6.5, 2.3 Hz,
1H), 6.61 (d, J = 8.9 Hz, 1H), 3.81 (s, 3H), 2.87 (d, J = 4.7 Hz, 3H).
MS (ESI): m/z calcd for C14H13N3O2 255.1; found 255.9 (M + H)+.
5-(5-Methoxybenzo[d]oxazol-2-yl)-N,N-dimethylpyridin-2-
amine (25). To a solution of 23 (480 mg, 2 mmol) and
paraformaldehyde (300 mg, 10 mmol) in acetic acid (50 mL) was
added NaCNBH3 (189 mg, 6 mmol) in one portion at room
temperature. The resulting mixture was stirred at room temperature
overnight. After neutralization with NH4OH, water was added, and the
precipitate was collected by filtration to give 25 as a white solid (481
mg, 89.4%). Mp: 140.0−141.6 °C. 1H NMR (400 MHz, DMSO-d6): δ
8.84 (d, J = 2.1 Hz, 1H), 8.14 (dd, J = 6.9, 2.2 Hz, 1H), 7.59 (d, J = 8.8
Hz, 1H), 7.26 (d, J = 2.3 Hz, 1H), 6.92 (dd, J = 6.4, 2.4 Hz, 1H), 6.80
(d, J = 9.1 Hz, 1H), 3.81 (s, 3H), 3.14 (s, 6H). MS (ESI): m/z calcd
for C15H15N3O2 269.12, found 270.0 (M + H)+.
Figure 6. Autoradiography of [18F]32 ex vivo using Tg model mouse
(C57BL6, APPswe/PSEN1, 10 months old, male) (A, C) and wild-
type controls (C57BL6, 10 months old) (E). Plaques were also
confirmed by the staining of the same sections with thioflavin S.
and 2,2′-oxidiethanol (5.0 g, 5 mmol) in CH2Cl2 (200 mL) was added
KOH (21.1 g, 40 mmol). The mixture was stirred at 0 °C for 6 h. After
reaction, the mixture was washed with water (3 × 100 mL). The
organic layer was dried over anhydrous MgSO4. After removal of the
solvent, the residue was purified by recrystallization in anhydrous
2-(6-(Methylamino)pyridin-3-yl)benzo[d]oxazol-5-ol (26). To
a solution of 24 (480 mg, 1.88 mmol) in CH2Cl2 (30 mL) was added
BBr3 (10 mL, 1 M solution in CH2Cl2) dropwise at −78 °C in a liquid
nitrogen−ethanol bath. The mixture was allowed to warm to room
temperature and was stirred overnight. Water was added to quench the
reaction. After neutralization with NH4OH, the mixture was extracted
with CH2Cl2. The solvent was removed, and 26 was obtained as a
1
MeOH to give 18 (18.1 g, 92.7%). H NMR (400 MHz, CDCl3): δ
7.78 (d, J = 8.3 Hz, 4H), 7.34 (d, J = 8.2 Hz, 4H), 4.09 (t, J = 4.6 Hz,
4H), 3.61 (t, J = 4.7 Hz, 4H), 2.45 (s, 6H).
(Ethane-1,2-diylbis(oxy))bis(ethane-2,1-diyl) Bis(4-methyl-
benzenesulfonate) (19). The same reaction as described above to
prepare 18 was used, and 19 was obtained as a white crystal (24.7 g,
71.5%). 1H NMR (400 MHz, CDCl3): δ 7.79 (d, J = 8.3 Hz, 4H), 7.34
(d, J = 8.2 Hz, 4H), 4.14 (t, J = 4.7 Hz, 4H), 3.65 (t, J = 4.9 Hz, 4H),
3.53 (s, 4H), 2.44 (s, 6H).
1
white solid (442 mg, 97%). Mp: 237.1−238.6 °C. H NMR (400
MHz, DMSO-d6): δ 9.41 (s, 1H), 8.76 (d, J = 1.6 Hz, 1H), 8.02 (dd, J
= 7.2, 1.4 Hz, 1H), 7.47 (d, J = 8.7 Hz, 1H), 7.32 (d, J = 4.4 Hz, 1H),
7.00 (d, J = 2.1 Hz, 1H), 6.75 (dd, J = 6.4, 2.2 Hz, 1H), 6.60 (d, J = 8.8
Hz, 1H), 2.87 (d, J = 4.7 Hz, 3H). MS (ESI): m/z calcd for
C13H11N3O2 241.1, found 241.8 (M + H)+.
2-(2-Fluoroethoxy)ethyl 4-Methylbenzenesulfonate (20).
Intermediate 20 was synthesized according to the literature.38 Briefly,
to a solution of 18 (2.3 g, 5 mmol) in dry THF (30 mL) was added
anhydrous TBAF (10 mL, 10 mmol, 1 M in THF). The solution was
stirred at 70 °C for 5 h. After removal of solvent, the residue was
purified by silica gel chromatography to give 20 (540 mg, 37.1%) as a
colorless oil. 1H NMR (400 MHz, CDCl3): δ 7.81 (d, J = 8.2 Hz, 2H),
7.35 (d, J = 8.1 Hz, 2H), 4.58−4.52 (m, 1H), 4.46−4.40 (m, 1H),
4.21−4.16 (m, 2H), 3.75−3.62 (m, 4H), 2.45 (s, 3H).
2-(6-(Dimethylamino)pyridin-3-yl)benzo[d]oxazol-5-ol (27).
The same reaction as described above to prepare 26 was used, and
27 was obtained as a white solid (348 mg, 88.5%). Mp: 285.2−286.7
1
°C. H NMR (400 MHz, DMSO-d6): δ 9.43 (s, 1H), 8.82 (d, J = 2.1
Hz, 1H), 8.13 (dd, J = 9.0, 2.3 Hz, 1H), 7.48 (d, J = 8.7 Hz, 1H), 7.01
(d, J = 2.2 Hz, 1H), 6.80 (d, J = 9.1 Hz, 1H), 6.76 (dd, J = 8.7, 2.3 Hz,
1H), 3.14 (s, 6H). MS (ESI): m/z calcd for C14H13N3O2 255.1, found
255.9 (M + H)+.
2-(2-(2-Fluoroethoxy)ethoxy)ethyl 4-Methylbenzenesulfo-
nate (21). The same reaction as described above to prepare 20 was
1
5-(5-(2-(2-Fluoroethoxy)ethoxy)benzo[d]oxazol-2-yl)-N-
methylpyridin-2-amine (28). A solution of 26 (48 mg, 0.2 mmol),
2-(2-fluoroethoxy)ethyl 4-methylbenzenesulfonate (60 mg, 0.2 mmol),
and K2CO3 (28 mg, 0.2 mmol) in DMF (6 mL) was stirred at 110 °C
for 3 h. The aqueous portion was extracted with dichloromethane (3 ×
15 mL), and the organic layer was dried over anhydrous MgSO4.
CH2Cl2 was removed in vacuum. The residue was purified by column
chromatography (petroleum ether/AcOEt, 1/2) to give 28 as a white
used, and 21 was obtained as a colorless oil (309.2 mg, 20.2%). H
NMR (400 MHz, CDCl3): δ 7.80 (d, J = 8.1 Hz, 2H), 7.34 (d, J = 8.1
Hz, 2H), 4.59 (t, J = 3.8 Hz, 1H), 4.48 (t, J = 3.8 Hz, 1H), 4.17 (t, J =
4.6 Hz, 2H), 3.74 (t, J = 4.2 Hz, 1H), 3.61−3.72 (m, 3H), 3.61 (s,
4H), 2.45 (s, 3H).
2-Amino-4-methoxyphenol (22). Intermediate 22 was synthe-
sized according to the literature.32 Briefly, a mixture of 4-methoxy-2-
nitrophenol (3.5 g, 20.7 mmol) and Pd/C (10%, 0.5 g) in absolute
methanol (100 mL) under a hydrogen atmosphere (balloon) was
vigorously stirred for 24 h at room temperature. The mixture was
filtered and the filtrate washed with methanol (20 mL). Evaporation of
the solvent under reduced pressure gave 22 as brown flake crystal (2.8
g, 97.3%). Mp: 133.4−135.7 °C.
5-(5-Methoxybenzo[d]oxazol-2-yl)pyridin-2-amine (23). A
solution of 22 (1.0 g, 7 mmol) and 5-(trifluoromethyl)pyridin-2-
amine (1.7 g, 10.8 mmol) in NaOH (1 M in water, 20 mL) was stirred
at 90 °C for 3 h. After the mixture was cooled to room temperature,
the precipitate was collected by filtration. Compound 23 (1.5 g,
88.9%) was obtained as a white solid. Mp: 204.1−205.9 °C. 1H NMR
(400 MHz, DMSO-d6): δ 8.71 (d, J = 2.1 Hz, 1H), 8.05 (dd, J = 6.6,
2.2 Hz, 1H), 7.58 (d, J = 8.8 Hz, 1H), 7.25 (d, J = 2.3 Hz, 1H), 6.92
(dd, J = 6.5, 2.4 Hz, 1H), 6.80 (s, 2H), 6.59 (d, J = 8.8 Hz, 1H), 3.81
1
solid (41 mg, 62.2%). Mp: 134.8−135.6 °C. H NMR (400 MHz,
CDCl3): δ 8.92 (s, 1H), 8.21 (dd, J = 7.0, 1.7 Hz, 1H), 7.41 (d, J = 8.8
Hz, 1H), 7.21 (d, J = 2.2 Hz, 1H), 6.93 (dd, J = 6.6, 2.2 Hz, 1H), 6.48
(d, J = 8.8 Hz, 1 H), 5.12 (d, J = 4.4 Hz, 1H), 4.61 (dt, J = 47.6, 4.0
Hz, 2H), 4.19 (t, J = 4.5 Hz, 2H), 3.92 (t, J = 4.8 Hz, 2H), 3.87 (t, J =
4.0 Hz, 1H), 3.80 (t, J = 4.1 Hz, 1H), 3.02 (d, J = 5.1 Hz, 3H). HRMS
(EI): m/z calcd for C17H19N3O3F 332.1410; found 332.1412 (M +
H)+.
5-(5-(2-(2-(2-Fluoroethoxy)ethoxy)ethoxy)benzo[d]oxazol-
2-yl)-N-methylpyridin-2-amine (29). The same reaction as
described above to prepare 28 was used, and 29 was obtained as a
white solid (30 mg, 40.2%). Mp: 87.2−89.1 °C. 1H NMR (400 MHz,
CDCl3): δ 8.92 (s, 1H), 8.21 (dd, J = 7.0, 1.8 Hz, 1H), 7.40 (d, J = 8.8
Hz, 1H), 7.21 (d, J = 2.2 Hz, 1H), 6.92 (dd, J = 6.6, 2.2 Hz, 1H), 6.49
I
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
(d, J = 8.8 Hz, 1H), 5.12 (d, J = 4.4 Hz, 1H), 4.57 (dt, J = 47.7, 4.0 Hz,
2H), 4.18 (t, J = 4.5 Hz, 2H), 3.90 (t, J = 4.8 Hz, 2H), 3.82−3.72 (m,
6H), 3.80 (t, J = 4.1 Hz, 1H), 3.02 (d, J = 5.0 Hz, 3H). HRMS (EI):
m/z calcd for C19H23N3O4F 376.1673; found 376.1674 (M + H)+.
5-(5-(2-(2-Fluoroethoxy)ethoxy)benzo[d]oxazol-2-yl)-N,N-di-
methylpyridin-2-amine (30). The same reaction as described above
to prepare 28 was used, and 30 was obtained as a white solid (30 mg,
3.18 (s, 6H). MS (ESI): m/z calcd for C15H15N3OS 285.1; found
285.6 (M + H)+.
2-(6-(Methylamino)pyridin-3-yl)benzo[d]thiazol-6-ol (38).
The same reaction as described above to prepare 26 was used, and
38 was obtained as a white solid (653 mg, 88.4%). Mp: 238.5−244.2
1
°C. H NMR (400 MHz, DMSO-d6): δ 9.72 (s, 1H), 8.61 (s, 1H),
7.94 (d, J = 8.3 Hz, 1H), 7.74 (d, J = 8.6 Hz, 1H), 7.36 (s, 1H), 7.17
(d, J = 3.8 Hz, 1H), 6.93 (d, J = 6.8 Hz, 1H), 6.57 (d, J = 8.7 Hz, 1H),
2.86 (d, J = 4.2 Hz, 3H). MS (ESI): m/z calcd for C13H11N3OS 257.1,
found 257.9 (M + H)+.
2-(6-(Dimethylamino)pyridin-3-yl)benzo[d]thiazol-6-ol (39).
The same reaction as described above to prepare 26 was used, and 39
was obtained as a white solid (0.61 g, 68.2%). Mp: 287.2−288.7 °C.
1H NMR (400 MHz, DMSO-d6): δ 9.74 (s, 1H), 8.68 (d, J = 2.1 Hz,
1
43.5%). Mp: 88.0−89.7 °C. H NMR (400 MHz, CDCl3): δ 8.98 (s,
1H), 8.20 (dd, J = 6.8, 2.2 Hz, 1H), 7.40 (d, J = 8.8 Hz, 1H), 7.21 (d, J
= 2.3 Hz, 1H), 6.92 (dd, J = 6.4, 2.4 Hz, 1H), 6.59 (d, J = 9.0 Hz, 1H),
4.61 (dt, J = 47.7, 4.1 Hz, 2H), 4.19 (t, J = 4.5 Hz, 2H), 3.92 (t, J = 4.9
Hz, 2H), 3.88 (t, J = 4.2 Hz, 1H), 3.80 (t, J = 4.2 Hz, 1H), 3.19 (s,
6H). HRMS (EI): m/z calcd for C18H21N3O3F 346.1567; found
346.1559 (M + H)+.
1H), 8.04 (dd, J = 9.0, 2.2 Hz, 1H), 7.75 (d, J = 8.7 Hz, 1H), 7.36 (d, J
= 2.1 Hz, 1H), 6.94 (dd, J = 8.7, 2.1 Hz, 1H), 6.76 (d, J = 9.0 Hz, 1H),
3.12 (s, 6H). MS (ESI): m/z calcd for C14H13N3OS 271.1; found
271.5 (M + H)+.
5-(5-(2-(2-(2-Fluoroethoxy)ethoxy)ethoxy)benzo[d]oxazol-
2-yl)-N,N-dimethylpyridin-2-amine (31). The same reaction as
described above to prepare 28 was used, and 31 was obtained as a
white solid (40 mg, 50%). Mp: 120.0−121.8 °C. 1H NMR (400 MHz,
CDCl3): δ 8.98 (s, 1H), 8.19 (d, J = 8.9 Hz, 1H), 7.39 (d, J = 8.8 Hz,
1H), 7.27 (d, J = 0.9 Hz, 1H), 6.91 (d, J = 8.6 Hz, 1H), 6.59 (d, J = 9.0
Hz, 1H), 4.50−4.63 (m, 2H), 4.18 (t, J = 4.1 Hz, 2H), 3.90 (t, J = 4.0
Hz, 2H), 3.62−3.82 (m, 6H), 3.19 (s, 6H). HRMS (EI): m/z calcd for
C20H25N3O4F 390.1829; found 390.1829 (M + H)+.
5-(6-(2-(2-Fluoroethoxy)ethoxy)benzo[d]thiazol-2-yl)-N-
methylpyridin-2-amine (40). The same reaction as described above
to prepare 28 was used, and 40 was obtained as a white solid (40 mg,
1
55.2%). Mp: 155.7−157.5 °C. H NMR (400 MHz, CDCl3): δ 8.72
(d, J = 2.0 Hz, 1H), 8.10 (dd, J = 6.5, 2.2 Hz, 1H), 7.86 (d, J = 8.9 Hz,
1H), 7.35 (d, J = 2.3 Hz, 1H), 7.08 (dd, J = 6.5, 2.4 Hz, 1H), 6.46 (d, J
= 8.8 Hz, 1H), 4.98 (d, J = 4.6 Hz, 1H), 4.61 (dt, J = 47.6, 4.0 Hz,
2H), 4.21 (t, J = 4.5 Hz, 2H), 3.93 (t, J = 4.9 Hz, 2H), 3.87 (t, J = 4.1
Hz, 1H), 3.80 (t, J = 4.2 Hz, 1H), 2.99 (d, J = 5.1 Hz, 3H). HRMS
(EI): m/z calcd for C17H19N3O2FS 348.1182; found 348.1179 (M +
H)+.
5-(5-(2-Fluoroethoxy)benzo[d]oxazol-2-yl)-N-methylpyridin-
2-amine (32). The same reaction as described above to prepare 28
was used, and 32 was obtained as a white solid (30 mg, 52.5%). Mp:
1
212.2−212.8 °C. H NMR (400 MHz, DMSO-d6): δ 8.78 (d, J = 1.9
Hz, 1H), 8.04 (d, J = 7.5 Hz, 1H), 7.60 (d, J = 8.8 Hz, 1H), 7.36 (d, J
= 4.5 Hz, 1H), 7.30 (d, J = 2.3 Hz, 1H), 6.95 (dd, J = 6.4, 2.4 Hz, 1H),
6.61 (d, J = 8.9 Hz, 1 H), 4.77 (dt, J = 47.8, 3.5 Hz, 2H), 4.33 (t, J =
3.7 Hz, 1H), 4.25 (t, J = 3.8 Hz, 1H), 2.87 (d, J = 4.7 Hz, 3H). HRMS
(EI): m/z calcd for C15H15N3O2F 288.1148; found 288.1147 (M +
H)+.
5-(6-(2-(2-(2-Fluoroethoxy)ethoxy)ethoxy)benzo[d]thiazol-
2-yl)-N-methylpyridin-2-amine (41). The same reaction as
described above to prepare 28 was used, and 41 was obtained as a
1
white solid (45 mg, 57.3%). Mp: 126.4−128.7 °C. H NMR (400
MHz, CDCl3): δ 8.71 (d, J = 2.0 Hz, 1H), 8.10 (dd, J = 6.5, 2.2 Hz,
1H), 7.86 (d, J = 8.9 Hz, 1H), 7.34 (d, J = 2.3 Hz, 1H), 7.08 (dd, J =
6.5, 2.4 Hz, 1H), 6.46 (d, J = 8.8 Hz, 1H), 4.98 (d, J = 4.6 Hz, 1H),
4.57 (dt, J = 47.7, 4.0 Hz, 2H), 4.20 (t, J = 4.6 Hz, 2H), 3.90 (t, J = 4.9
Hz, 2H), 3.69−3.81 (m, 6H), 2.99 (d, J = 5.1 Hz, 3H). HRMS (EI):
m/z calcd for C19H23N3O3FS 392.1444; found 392.1446 (M + H)+.
5-(6-(2-(2-Fluoroethoxy)ethoxy)benzo[d]thiazol-2-yl)-N,N-
dimethylpyridin-2-amine (42). The same reaction as described
above to prepare 28 was used, and 42 was obtained as a white solid
(50 mg, 75%). Mp: 140.3−141.1 °C. 1H NMR (400 MHz, CDCl3): δ
8.76 (s, 1H), 8.12 (dd, J = 6.6, 2.3 Hz, 1H), 7.86 (d, J = 8.9 Hz, 1H),
7.34 (d, J = 2.4 Hz, 1H), 7.07 (dd, J = 6.5, 2.4 Hz, 1H), 6.57 (d, J = 9.0
Hz, 1H), 4.60 (dt, J = 47.7, 4.0 Hz, 2H), 4.21 (t, J = 4.5 Hz, 2H), 3.93
(t, J = 4.9 Hz, 2H), 3.87 (t, J = 4.2 Hz, 1H), 3.80 (t, J = 4.1 Hz, 1H),
3.10 (s, 6H). HRMS (EI): m/z calcd for C18H21N3O2FS 362.1339;
found 362.1338 (M + H)+.
5-(5-(2-Fluoroethoxy)benzo[d]oxazol-2-yl)-N,N-dimethylpyr-
idin-2-amine (33). The same reaction as described above to prepare
28 was used, and 33 was obtained as a white solid (54 mg, 89.7%).
1
Mp: 167.0−168.4 °C. H NMR (400 MHz, CDCl3): δ 8.98 (s, 1H),
8.20 (d, J = 8.0 Hz, 1H), 7.42 (d, J = 8.4 Hz, 1H), 7.21 (s, 1H), 6.93
(d, J = 7.5 Hz, 1H), 6.60 (d, J = 8.4 Hz, 1H), 4.85 (s, 1H), 4.73 (s,
1H), 4.30 (s, 1H), 4.23 (s, 1H), 3.20 (s, 6H). HRMS (EI): m/z calcd
for C16H17N3O2F 302.1305; found 302.1307 (M + H)+.
2-Amino-5-methoxybenzenethiol (34). Intermediate 34 was
synthesized according to the literature.14 Briefly, a mixture of 6-
methoxybenzo[d]thiazol-2-amine (9 g, 50 mmol) and KOH (28 g, 500
mmol) in H2O (100 mL) was stirred at 120 °C overnight. After
removal of the scraps by filtering, the filtrate was neutralized by acetic
acid (30% in water), and the precipitate was collected by filtration to
give 34 (7.75 g, 88.9%) as a light yellow solid. Mp: 81.6−82.8 °C.
5-(6-Methoxybenzo[d]thiazol-2-yl)pyridin-2-amine (35). The
same reaction as described above to prepare 23 was used, and 35 was
5-(6-(2-(2-(2-Fluoroethoxy)ethoxy)ethoxy)benzo[d]thiazol-
2-yl)-N,N-dimethylpyridin-2-amine (43). The same reaction as
described above to prepare 28 was used, and 43 was obtained as a
1
obtained (1.29 g, 50.2%). Mp: 224.7−225.4 °C. H NMR (400 MHz,
1
DMSO-d6): δ 8.58 (s, 1H), 7.98 (d, J = 8.2 Hz, 1H), 7.84 (d, J = 8.8
Hz, 1H), 7.65 (s, 1H), 7.08 (d, J = 8.2 Hz, 1H), 6.70 (s, 2H), 6.57 (d, J
= 8.5 Hz, 1H), 3.84 (s, 3H). MS (ESI): m/z calcd for C13H11N3OS
white solid (32.4 mg, 40%). Mp: 109.9−111.3 °C. H NMR (400
MHz, CDCl3): δ 8.76(s, 1H), 8.11 (dd, J = 6.6, 2.4 Hz, 1H), 7.85 (d, J
= 8.9 Hz, 1H), 7.34 (d, J = 2.3 Hz, 1H), 7.07 (dd, J = 6.5, 2.4 Hz, 1H),
6.57 (d, J = 9.0 Hz, 1H), 4.56 (dt, J = 47.7, 4.0 Hz, 2H), 4.20 (t, J = 4.6
Hz, 2H), 3.90 (t, J = 4.9 Hz, 2H), 3.71−3.81 (m, 6H), 3.17 (s, 6H).
HRMS (EI): m/z calcd for C20H25N3O3FS 406.1601; found 406.1590
(M + H)+.
+
257.1; found 257.5 (M + H) .
5-(6-Methoxybenzo[d]thiazol-2-yl)-N-methylpyridin-2-
amine (36). The same reaction as described above to prepare 24 was
used, and 36 was obtained as a white solid (798 mg, 98%). Mp:
1
194.7−195.6 °C. H NMR (400 MHz, DMSO-d6): δ 8.64 (s, 1H),
5-(6-(2-Fluoroethoxy)benzo[d]thiazol-2-yl)-N-methylpyri-
din-2-amine (44). The same reaction as described above to prepare
28 was used, and 44 was obtained as a white solid (40 mg, 64.6%).
Mp: 182.8−184.6 °C. 1H NMR (400 MHz, CDCl3): δ 8.72 (d, J = 2.0
Hz, 1H), 8.11 (dd, J = 6.5, 2.3 Hz, 1H), 7.88 (d, J = 8.9 Hz, 1H), 7.36
(d, J = 2.4 Hz, 1H), 7.09 (dd, J = 6.4, 2.4 Hz, 1H), 6.47 (d, J = 8.8 Hz,
1H), 4.80 (dt, J = 47.4, 4.0 Hz, 2H), 4.32 (t, J = 4.2 Hz, 1H), 4.25 (t, J
= 4.2 Hz, 1H), 3.00 (d, J = 5.1 Hz, 3H). HRMS (EI): m/z calcd for
C15H15N3OFS 304.0920; found 304.0924 (M + H)+.
7.96 (d, J = 7.4 Hz, 1H), 7.83 (d, J = 8.8 Hz, 1H), 7.64 (d, J = 1.6 Hz,
1H), 7.20 (d, J = 4.6 Hz, 1H), 7.07 (dd, J = 6.8, 2.1 Hz, 1H), 6.57 (d, J
= 8.8 Hz, 1H), 3.84 (s, 3H), 2.86 (d, J = 4.6 Hz, 3H). MS (ESI): m/z
calcd for C14H13N3OS 271.1, found 271.8 (M + H)+.
5-(6-Methoxybenzo[d]thiazol-2-yl)-N,N-dimethylpyridin-2-
amine (37). The same reaction as described above to prepare 25 was
1
used, and 37 was obtained (0.65 g, 91.1%). Mp: 175.4−176.7 °C. H
NMR (400 MHz, CDCl3): δ 8.76 (d, J = 2.3 Hz, 1H), 8.13 (dd, J =
9.0, 2.3 Hz, 1H), 7.87 (d, J = 8.9 Hz, 1H), 7.32 (d, J = 2.4 Hz, 1H),
7.05 (dd, J = 8.9, 2.4 Hz, 1H), 6.58 (d, J = 9.0 Hz, 1H), 3.88 (s, 3H),
5-(6-(2-Fluoroethoxy)benzo[d]thiazol-2-yl)-N,N-dimethyl-
pyridin-2-amine (45). The same reaction as described above to
J
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
prepare 28 was used, and 45 was obtained as a white solid (30 mg,
78.9%). Mp: 166.1−168.5 °C. H NMR (400 MHz, CDCl3): δ 8.69
(d, J = 2.3 Hz, 1H), 8.05 (dd, J = 6.6, 2.4 Hz, 1H), 7.80 (d, J = 8.9 Hz,
1H), 7.28 (d, J = 2.4 Hz, 1H), 7.01 (dd, J = 6.4, 2.4 Hz, 1H), 6.51 (d, J
= 9.0 Hz, 1H), 4.72 (dt, J = 47.4, 4.0 Hz, 2H), 4.24 (t, J = 4.2 Hz, 1H),
4.17 (t, J = 4.2 Hz, 1H), 3.10 (s, 6H). HRMS (EI): m/z calcd for
C16H17N3OFS 318.1076; found 318.1075 (M + H)+.
1H), 6.98 (dd, J = 6.7, 2.2 Hz, 1H), 4.15−4.19 (m, 4H), 3.86 (t, J = 4.8
Hz, 2H), 3.67−3.69 (m, 4H), 3.63 (t, J = 5.2 Hz, 2H), 3.49 (s, 3H),
2.42 (s, 3H), 1.56 (s, 9H). MS (ESI): m/z calcd for C31H37N3O9S
627.23, found 628.0 (M + H)+.
2-(2-(2-(2-(6-(Dimethylamino)pyridin-3-yl)benzo[d]thiazol-
6-yloxy)ethoxy)ethoxy)ethyl 4-Methylbenzenesulfonate (52).
The same reaction as described above to prepare 46 was used, and 52
was obtained as a white solid (87 mg, 48.7%). Mp: 107.6−108.7 °C.
1H NMR (400 MHz, CDCl3): δ 8.76 (d, J = 2.2 Hz, 1H), 8.12 (dd, J =
6.6, 2.4 Hz, 1H), 7.85 (d, J = 8.9 Hz, 1H), 7.79 (d, J = 8.2 Hz, 2H),
7.30−7.34 (m, 3H), 7.05 (dd, J = 6.4, 2.5 Hz, 1H), 6.58 (d, J = 9.0 Hz,
1H), 4.15−4.18 (m, 4H), 3.86 (t, J = 4.8 Hz, 2H), 3.61−3.72 (m, 6H),
3.17 (s, 6H), 2.42 (s, 3H). MS(ESI): m/z calcd for C27H31N3O6S2
557.17, found 558.0 (M + H)+.
1
2-(2-(6-(Dimethylamino)pyridin-3-yl)benzo[d]oxazol-5-
yloxy)ethyl 4-Methylbenzenesulfonate (46). To a solution of 27
(60 mg, 0.24 mmol) in acetone (30 mL) was added ethane-1,2-diyl
bis(4-methylbenzenesulfonate) (131 mg, 0.35 mmol), K2CO3 (124
mg, 0.9 mmol), and 18-crown-6 in catalytic amount. The mixture was
stirred at 75 °C for 3 h. Acetone was removed in vacuum. The residue
was purified by column chromatography (petroleum ether/AcOEt, 1/
1) to give 46 as a white solid (38.5 mg, 35.4%). Mp: 179.1−179.5 °C.
1H NMR (400 MHz, CDCl3): δ 8.97 (s, 1H), 8.19 (d, J = 8.9 Hz, 1H),
7.83 (d, J = 8.3 Hz, 2H), 7.37 (d, J = 8.8 Hz, 1H), 7.34 (d, J = 8.1 Hz,
2H), 7.06 (d, J = 2.4 Hz, 1H), 6.77 (dd, J = 8.8, 2.5 Hz, 1H), 6.60 (d, J
= 9.0 Hz, 1 H), 4.17−4.42 (m, 4H), 3.21 (s, 6H), 2.44 (s, 3H). MS
(ESI): m/z calcd for C23H23N3O5S 453.14, found 454.11 (M + H)+.
2-(2-(2-(2-(6-(Dimethylamino)pyridin-3-yl)benzo[d]oxazol-5-
yloxy)ethoxy)ethoxy)ethyl 4-Methylbenzenesulfonate (47).
The same reaction as described above to prepare 46 was used, and
47 was obtained as a white solid (60 mg, 27.7%). Mp: 86.3−87.9 °C.
1H NMR (400 MHz, CDCl3): δ 8.98(s, 1H), 8.22 (d, J = 7.7 Hz, 1H),
2-(2-(6-(Methylamino)pyridin-3-yl)benzo[d]thiazol-6-yloxy)-
ethyl 4-Methylbenzenesulfonate (53). The same reaction as
described above to prepare 46 was used, and 53 was obtained as a
1
white solid (55 mg, 40.3%). Mp: 153.0−155.4 °C. H NMR (400
MHz, CDCl3): δ 8.70 (s, 1H), 8.11 (dd, J = 8.8, 2.3 Hz, 1H), 7.81−
7.85 (m, 3H), 7.33 (d, J = 8.0 Hz, 2H), 7.20 (d, J = 2.5 Hz, 1H), 6.92
(dd, J = 8.9, 2.6 Hz, 1H), 6.47 (d, J = 8.8 Hz, 1H), 5.02 (d, J = 4.5 Hz,
1H), 4.20−4.43 (m, 4H), 3.01 (d, J = 5.2 Hz, 3H), 2.44 (s, 3H). MS
(ESI): m/z calcd for C22H21N3O4S2 455.1, found 456.06 (M + H)+.
2-(2-(2-(2-(6-(Methylamino)pyridin-3-yl)benzo[d]thiazol-6-
yloxy)ethoxy)ethoxy)ethyl 4-Methylbenzenesulfonate (54).
The same reaction as described above to prepare 46 was used, and
54 was obtained as a white solid (63 mg, 38.4%). Mp: 110.1−110.9
°C. 1H NMR (400 MHz, CDCl3): δ 8.71 (d, J = 2.2 Hz, 1H), 8.10 (dd,
J = 6.4, 2.3 Hz, 1H), 7.86 (d, J = 8.9 Hz, 1H), 7.79 (d, J = 8.2 Hz, 2H),
7.30−7.35 (m, 3H), 7.06 (dd, J = 6.4, 2.5 Hz, 1H), 6.46 (d, J = 8.8 Hz,
1H), 4.98 (d, J = 4.6 Hz, 1H), 4.15−4.20 (m, 4H), 3.86 (t, J = 4.9 Hz,
2H), 3.63−3.72 (m, 4H), 3.61 (t, J = 3.7 Hz, 2H), 3.00 (d, J = 5.1 Hz,
3H), 2.42 (s, 3H). MS (ESI): m/z calcd for C26H29N3O6S2 543.15,
found 543.9 (M + H)+.
7.79 (d, J = 8.2 Hz, 2H), 7.40 (d, J = 8.8 Hz, 1H), 7.32 (d, J = 8.1 Hz,
2H), 7.20 (d, J = 2.4 Hz, 1H), 6.91 (dd, J = 6.6, 2.2 Hz, 1H), 6.62 (d, J
= 8.5 Hz, 1H), 4.16 (m, 4H), 3.85 (t, J = 4.9 Hz, 2H), 3.61−3.73 (m,
6H), 3.22 (s, 6H), 2.42 (s, 3H). MS (ESI): m/z calcd for
C27H31N3O7S 541.2, found 542.0 (M + H)+.
2-(2-(6-(Methylamino)pyridin-3-yl)benzo[d]oxazol-5-yloxy)-
ethyl 4-Methylbenzenesulfonate (48). The same reaction as
described above to prepare 46 was used, and 48 was obtained as a
1
white solid (40 mg, 30.5%). Mp: 169.7−170.7 °C. H NMR (400
2-(2-(6-(tert-Butoxycarbonyl)pyridin-3-yl)benzo[d]thiazol-6-
yloxy)ethyl 4-Methylbenzenesulfonate (55). The same reaction
as described above to prepare 50 was used, and 55 was obtained as a
MHz, CDCl3): δ 8.90 (s, 1H), 8.22 (dd, J = 8.8, 2.0 Hz, 1H), 7.83 (d, J
= 8.2 Hz, 2H), 7.38 (d, J = 8.8 Hz, 1H), 7.34 (d, J = 8.1 Hz, 2H), 7.07
(d, J = 2.4 Hz, 1H), 6.78 (dd, J = 8.8, 2.4 Hz, 1H), 6.51 (d, J = 8.8 Hz,
1 H), 5.37 (s, 1H), 4.40 (t, J = 4.4 Hz, 2H), 4.18 (t, J = 5.0 Hz, 2H),
3.02 (d, J = 5.0 Hz, 3H), 2.44 (s, 3H). MS (ESI): m/z calcd for
C22H21N3O5S 439.12, found 440.09 (M + H)+.
1
white solid (53 mg, 79.6%). Mp: 163.4−165.4 °C. H NMR (400
MHz, CDCl3): δ 8.97 (d, J = 2.2 Hz, 1H), 8.27 (dd, J = 8.8, 2.4 Hz,
1H), 7.92 (dd, J = 8.8, 7.2 Hz, 2H), 7.82 (d, J = 8.3 Hz, 2H), 7.34 (d, J
= 8.1 Hz, 2H), 7.25 (d, J = 2.6 Hz, 1H), 6.98 (dd, J = 9.0, 2.6 Hz, 1H),
4.22−4.44 (m, 4H), 3.48 (s, 3H), 2.44 (s, 3H), 1.56 (s, 9H). MS
(ESI): m/z calcd for C27H29N3O6S2 555.15, found 556.13 (M + H)+.
2-(2-(2-(2-(6-(tert-Butoxycarbonyl)pyridin-3-yl)benzo[d]-
thiazol-6-yloxy)ethoxy)ethoxy)ethyl 4-Methylbenzenesulfo-
nate (56). The same reaction as described above to prepare 50 was
used, and 56 was obtained as a white solid (45 mg, 63.4%). Mp: 88.2−
2-(2-(2-(2-(6-(Methylamino)pyridin-3-yl)benzo[d]oxazol-5-
yloxy)ethoxy)ethoxy)ethyl 4-Methylbenzenesulfonate (49).
The same reaction as described above to prepare 46 was used, and
49 was obtained as a white solid (75 mg, 47.8%). Mp: 91.3 − 92.2 °C.
1H NMR (400 MHz, CDCl3): δ 8.90 (d, J = 2.0 Hz, 1H), 8.16 (dd, J =
6.7, 2.1 Hz, 1H), 7.77 (d, J = 8.2 Hz, 2H), 7.37 (d, J = 8.8 Hz, 1H),
7.29 (d, J = 8.1 Hz, 2H), 7.17 (d, J = 2.4 Hz, 1H), 6.88 (dd, J = 6.4, 2.4
Hz, 1H), 6.45 (d, J = 8.8 Hz, 1H), 5.25 (d, J = 4.8 Hz, 1H), 4.11−4.17
(m, 4H), 3.82 (t, J = 4.9 Hz, 2H), 3.59−3.70 (m, 6H), 2.98 (d, J = 5.1
Hz, 3H), 2.39 (s, 3H). MS (ESI): m/z calcd for C26H29N3O7S 527.17,
found 528.19 (M + H)+.
2-(2-(6-(tert-Butoxycarbonyl)pyridin-3-yl)benzo[d]oxazol-5-
yloxy)ethyl 4-Methylbenzenesulfonate (50). To a solution of 48
(40 mg, 0.09 mmol) in THF (30 mL) was added excessive (Boc)2O.
The mixture was stirred at 85 °C for 28 h. After solvent was removed
in vacuum, the residue was purified by column chromatography
(petroleum ether/AcOEt, 1/1) to give 50 as a white solid (32.7 mg,
66.7%). Mp: 172.1−173.2 °C. 1H NMR (400 MHz, CDCl3): δ 9.16 (s,
1H), 8.37 (dd, J = 8.9, 6.6 Hz, 1H), 8.02 (d, J = 8.8 Hz, 1H), 7.83 (d, J
= 8.3 Hz, 2H), 7.44 (d, J = 8.9 Hz, 1H), 7.35 (d, J = 8.0 Hz, 2H), 7.12
(d, J = 2.4 Hz, 1H), 6.86 (dd, J = 8.9, 2.5 Hz, 1H), 4.19−4.43 (m, 4H),
3.51 (s, 3H), 2.45 (s, 3H), 1.57 (s, 9H). MS (ESI): m/z calcd for
C27H29N3O7S 539.17, found 540.18 (M + H)+.
1
90.1 °C. H NMR(400 MHz, CDCl3): δ 8.96 (s, 1H), 8.25 (dd, J =
6.4, 2.4 Hz, 1H), 7.92 (t, J = 8.7 Hz, 2H), 7.79 (d, J = 8.2 Hz, 2H),
7.34 (d, J = 2.5 Hz, 1H), 7.31 (d, J = 8.1 Hz, 2H), 7.11 (dd, J = 6.6, 2.3
Hz, 1H), 4.15−4.22 (m, 4H), 3.87 (t, J = 4.7 Hz, 2H), 3.62−3.73 (m,
6H), 3.47 (s, 3H), 2.42 (s, 3H), 1.56 (s, 9H). MS (ESI): m/z calcd for
C31H37N3O8S2 643.20, found 644.18 (M + H)+.
Radiolabeling. [18F]Fluoride trapped on a QMA cartridge was
eluted with 1 mL of (Kryptofix 222)/K2CO3 solution (13 mg of
Kryptofix 222 and 1.1 mg of K2CO3 in CH3CN/H2O, 0.8:0.2). The
solvent was removed at 120 °C under a stream of nitrogen gas. The
residue was azeotropically dried with 1 mL of anhydrous acetonitrile
three times at 120 °C under a stream of nitrogen gas.
For [18F]29, [18F]32, [18F]41, and [18F]44, a solution of the
tosylate precursors 51, 50, 56, and 55 (1.0 mg) in CH3CN (0.5 mL)
was added to the reaction vessel containing the 18F− activity,
respectively. The mixture was heated at 100 °C for 5 min. After the
solution was cooled to room temperature, HCl (1 M aqueous solution,
0.2 mL) was added and the mixture was heated at 100 °C again for 5
min. An aqueous solution of NaHCO3 was added to adjust to pH 8−9.
Water (10 mL) was added, and the mixture was passed through a Sep-
Pak C18 cartridge (Waters). The cartridge was washed with 10 mL of
water, and the labeled compound was eluted with 2 mL of acetonitrile.
After the solvent was removed, the residue was dissolved in CH3CN
2-(2-(2-(2-(6-(tert-Butoxycarbonyl)pyridin-3-yl)benzo[d]-
oxazol-5-yloxy)ethoxy)ethoxy)ethyl 4-Methylbenzenesulfo-
nate (51). The same reaction as described above to prepare 50 was
used, and 51 was obtained as a white solid (20 mg, 22.5%). Mp: 85.0−
1
85.5 °C. H NMR (400 MHz, CDCl3): δ 9.16 (s, 1H), 8.37 (dd, J =
6.8, 2.0 Hz, 1H), 7.99 (d, J = 8.9 Hz, 1H), 7.79 (d, J = 8.0 Hz, 2H),
7.45 (d, J = 8.8 Hz, 1H), 7.32 (d, J = 8.0 Hz, 2H), 7.24 (d, J = 2.0 Hz,
K
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
and subjected to HPLC for purification (Agela Technologies, 5 μm, 10
mm × 250 mm, CH3CN/water = 6/4; flow rate = 4 mL/min). The
retention times of [18F]29, [18F]32, [18F]41, and [18F]44 were 7.72,
6.36, 5.28, and 7.81 min, respectively, in this HPLC system. The
preparation took 60 min, and the radiochemical yield was 20−30%
(decay not corrected). The radiochemical purity of all tracers was
greater than 98%.
(0.2 M, pH 7.4) for 30 min. After that, they were incubated with 18F-
labeled tracers (370 KBq per 100 μL) for 1 h at room temperature.
They were then washed in 40% EtOH before being rinsed with water
for 1 min. After drying, the sections were exposed to a phosphorus
plate (PerkinElmer, U.S.) for 2 h. In vitro autoradiographic images
were obtained using a phosphor imaging system (Cyclone, Packard).
After autoradiographic examination, the same mouse brain sections
were stained by thioflavin S to confirm the presence of Aβ plaques. For
the staining of thioflavin S, sections were immersed in a 0.125%
thioflavin S solution containing 10% EtOH for 3 min and washed in
40% EtOH. After drying, fluorescent observation was performed using
the Oberver Z1 (Zeiss, Germany) equipped with GFP filter set
(excitation, 505 nm).
Ex Vivo Autoradiography Studies. The ex vivo evaluation was
performed using a Tg (C57BL6-APP/PS1, 10 months old, male)
mouse, which was used as an Alzheimer’s model, and an age-matched
control (C57BL6, 10 months, male). A saline solution of the labeled
agent [18F]32 (19.2 MBq) was injected directly into the tail vein. The
mice were sacrificed by decapitation at 30 min after intravenous
injection. The brains were immediately removed and frozen in a liquid
nitrogen bath. Sections of 20 μm were cut and exposed to a
phosphorus plate (PerkinElmer, U.S.) for 5 h. Ex vivo film
autoradiograms were thus obtained using a phosphor imaging system
(Cyclone, Packard). After autoradiographic examination, the same
sections were stained with thioflavin S to confirm the presence of
amyloid plaques.
For [18F]31, [18F]33, and [18F]43, a solution of the tosylate
precursors 47, 46, and 52 (1.0 mg) in CH3CN (0.5 mL) was added to
the reaction vessel containing the 18F− activity, respectively. The
mixture was heated at 100 °C for 5 min. Water (10 mL) was added,
and the mixture was passed through a Sep-Pak C18 cartridge (Waters).
The cartridge was washed with 10 mL of water, and the labeled
compound was eluted with 2 mL of acetonitrile. The eluted compound
was purified by HPLC (Agela Technologies, 5 μm, 10 mm × 250
mm). The retention times of [18F]31, [18F]33, and [18F]43 were 9.09,
5.93, and 8.23 min, respectively, in this HPLC system (CH3CN/water
= 7/3; flow rate = 4 mL/min). The preparation took 40 min, and the
radiochemical yield was 40−50% (decay not corrected). The
radiochemical purity of all tracers was greater than 98%. Specific
activity, estimated by comparing the UV peak intensity of purified 18F-
labeled compounds with reference nonradioactive compounds, was
approximately 200 GBq/μmol.
Binding Assay Using Aβ1−42 Aggregates. Inhibition experi-
ments were carried out in 12 mm × 75 mm borosilicate glass tubes
according to procedures described previously with some modifications.
The radioligand [125I]IMPY was prepared according to procedures
described previously.12 After HPLC purification, the radiochemical
purity was greater than 95%. The reaction mixture contained 100 μL
of aggregated Aβ1−42 fibrils, 100 μL of radioligands ([125I]IMPY,
60000−100000 cpm/100 μL), 100 μL of inhibitors (10−4−10−8.5 M in
ethanol), and 700 μL of 0.05% bovine serum albumin solution in a
final volume of 1 mL. The mixture was incubated at 37 °C for 2 h, and
the free radioactivity was separated by vacuum filtration through
Whatman GF/B filters using a Brandel Mp-48T cell harvester followed
by 3 × 4 mL washes with PBS (0.02 M, pH 7.4) at room temperature.
Filters with the bound [125I]IMPY were counted in a γ counter
(WALLAC/Wizard 1470, U.S.) with 70% efficiency. Inhibition
experiments were repeated three times, and the half maximal inhibitory
concentration (IC50) was determined using GraphPad Prism 4.0, and
the inhibition constant (Ki) was calculated using the Cheng−Prusoff
equation:39 Ki = IC50/(1 + [L]/Kd).
In Vitro Stability Studies. The in vitro stability of [18F]32 in
mouse plasma was determined by incubating 1.85 MBq purified [18F]
32 with 100 μL of mouse plasma at 37 °C for 2, 10, 30, and 60 min.
Proteins were precipitated by adding 200 μL of acetonitrile after
centrifugation at 5000 rpm for 5 min at 4 °C. The supernatant was
collected. Approximately 0.1 mL of the supernatant solution was
analyzed using HPLC.
ASSOCIATED CONTENT
■
S
* Supporting Information
Purity of key target compounds together with HPLC
1
chromatograms, H NMR spectra of compounds, and HRMS
data of key target compounds. This material is available free of
charge via the Internet at http://pubs.acs.org.
Partition Coefficient Determination. The determination of
partition coefficients of radiofluorinated tracers was performed
according to the procedure previously reported.32 A solution of 18F-
labeled tracer (1.5 MBq) was added to premixed suspensions
containing 3.0 g of n-octanol and 3.0 g of PBS (0.05 M, pH 7.4) in
a test tube. The test tube was vortexed for 3 min at room temperature,
followed by centrifugation for 5 min at 3000 rpm. Two samples from
the n-octanol (50 μL) and water (500 μL) layers were measured. The
partition coefficient was expressed as the logarithm of the ratio of the
count per gram from n-octanol versus PBS. Samples from the n-
octanol layer were repartitioned until consistent partition coefficient
values were obtained. The measurement was done in triplicate and
repeated three times.
Biodistribution Studies. The biodistribution experiments were
performed in normal male mice (5 weeks, male, average weight, about
20 g) and approved by the Animal Care Committee of Beijing Normal
University. A saline solution containing the 18F-labeled tracers (370
KBq per 100 μL) was injected directly into the tail. The mice (n = 5
for each time point) were sacrificed at 2, 10, 30, and 60 min after
injection. The organs of interest were removed and weighed, and
radioactivity was measured with an automatic γ counter. The percent
dose per gram of wet tissue was calculated by a comparison of the
tissue counts to suitably diluted aliquots of the injected material.
In Vitro Autoradiography Studies. Paraffin-embedded brain
sections were deparaffinized with 2 × 20 min washes in xylene, 2 × 5
min washes in 100% ethanol, a 5 min wash in 90% ethanol/H2O, a 5
min wash in 80% ethanol/H2O, a 5 min wash in 60% ethanol/H2O,
and a 10 min wash in running tap water and then incubated in PBS
AUTHOR INFORMATION
■
Corresponding Author
*For M.C.: phone, +86-10-58808891; fax, +86-10-58808891; e-
mail, cmc@bnu.edu.cn. For B.L.: phone, +86-10-58808891; e-
mail, liuboli@bnu.edu.cn.
Author Contributions
∥These authors contributed equally.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
The authors thank Dr. Jin Liu (College of Life Science, Beijing
Normal University) for assistance in the in vitro neuro-
pathological staining. This work was supported by National
Natural Science Foundation of China (Grants 21201019,
21071023, and 30670586).
ABBREVIATIONS USED
■
AD, Alzheimer’s disease; Aβ, β-amyloid; NFT, neurofibrillary
tangle; CR, Congo Red; Th-T, thioflavin T; SPECT, single
photon emission computed tomography; PET, positron
emission tomography; IMPY, 2-(4′-dimethylaminophenyl)-6-
imidazo[1,2-a]pyridine; PEG, polyethylene glycol; FPEG,
L
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Alzheimer disease β-amyloid with [11C]SB-13 PET. Am. J. Geriatr.
Psychiatry 2004, 12, 584−595.
(21) Koole, M.; Lewis, D. M.; Buckley, C.; Nelissen, N.;
Vandenbulcke, M.; Brooks, D. J.; Vandenberghe, R.; Van Laere, K.
Whole-body biodistribution and radiation dosimetry of 18F-GE067: a
radioligand for in vivo brain amyloid imaging. J. Nucl. Med. 2009, 50,
818−22.
(22) Jureus, A.; Swahn, B. M.; Sandell, J.; Jeppsson, F.; Johnson, A.
E.; Johnstrom, P.; Neelissen, J. A.; Sunnemark, D.; Farde, L.; Svensson,
S. P. Characterization of AZD4694, a novel fluorinated Aβ plaque
neuroimaging PET radioligand. J. Neurochem. 2010, 114, 784−794.
(23) Rowe, C. C.; Ackerman, U.; Browne, W.; Mulligan, R.; Pike, K.
L.; O’Keefe, G.; Tochon-Danguy, H.; Chan, G.; Berlangieri, S. U.;
Jones, G.; Dickinson-Rowe, K. L.; Kung, H. P.; Zhang, W.; Kung, M.
P.; Skovronsky, D.; Dyrks, T.; Holl, G.; Krause, S.; Friebe, M.;
Lehman, L.; Lindemann, S.; Dinkelborg, L. M.; Masters, C. L.;
Villemagne, V. L. Imaging of amyloid β in Alzheimer’s disease with
18F-BAY94-9172, a novel PET tracer: proof of mechanism. Lancet
Neurol. 2008, 7, 129−135.
(24) Lin, K. J.; Hsu, W. C.; Hsiao, I. T.; Wey, S. P.; Jin, L. W.;
Skovronsky, D.; Wai, Y. Y.; Chang, H. P.; Lo, C. W.; Yao, C. H.; Yen,
T. C.; Kung, M. P. Whole-body biodistribution and brain PET imaging
with [18F]AV-45, a novel amyloid imaging agent-a pilot study. Nucl.
Med. Biol. 2010, 37, 497−508.
fluoropolyethylene glycol; THF, tetrahydrofuran; BOC,
butyloxycarbonyl; TBAF, tetra-n-butylammonium fluoride
REFERENCES
■
(1) Selkoe, D. J. The origins of Alzheimer disease: a is for amyloid.
JAMA, J. Am. Med. Assoc. 2000, 283, 1615−1617.
(2) Selkoe, D. J. Alzheimer’s disease: genes, proteins, and therapy.
Physiol. Rev. 2001, 81, 741−766.
(3) Hardy, J.; Selkoe, D. J. The amyloid hypothesis of Alzheimer’s
disease: progress and problems on the road to therapeutics. Science
2002, 297, 353−356.
(4) Hardy, J. The amyloid hypothesis for Alzheimer’s disease: a
critical reappraisal. J. Neurochem. 2009, 110, 1129−1134.
(5) Reitz, C. Alzheimer’s disease and the amyloid cascade hypothesis:
a critical review. Int. J. Alzheimer's Dis. 2012, 2012, 369808.
(6) Mathis, C. A.; Wang, Y.; Klunk, W. E. Imaging β-amyloid plaques
and neurofibrillary tangles in the aging human brain. Curr. Pharm. Des.
2004, 10, 1469−1492.
(7) Cai, L.; Innis, R. B.; Pike, V. W. Radioligand development for
PET imaging of β-amyloid (Aβ)-current status. Curr. Med. Chem.
2007, 14, 19−52.
(8) Sugimoto, H. Development of anti-Alzheimer’s disease drug
based on β-amyloid hypothesis. Yakugaku Zasshi 2010, 130, 521−526.
(9) Valotassiou, V.; Archimandritis, S.; Sifakis, N.; Papatriantafyllou,
J.; Georgoulias, P. Alzheimer’s disease: spect and pet tracers for β-
amyloid imaging. Curr. Alzheimer Res. 2010, 7, 477−486.
(10) Herholz, K.; Ebmeier, K. Clinical amyloid imaging in
Alzheimer’s disease. Lancet Neurol. 2011, 10, 667−670.
(11) Kung, M. P.; Hou, C.; Zhuang, Z. P.; Zhang, B.; Skovronsky, D.;
Trojanowski, J. Q.; Lee, V. M.; Kung, H. F. IMPY: an improved
thioflavin-T derivative for in vivo labeling of β-amyloid plaques. Brain
Res. 2002, 956, 202−210.
(12) Zhuang, Z. P.; Kung, M. P.; Wilson, A.; Lee, C. W.; Plossl, K.;
Hou, C.; Holtzman, D. M.; Kung, H. F. Structure−activity relationship
of imidazo[1,2-a]pyridines as ligands for detecting β-amyloid plaques
in the brain. J. Med. Chem. 2003, 46, 237−243.
(13) Newberg, A. B.; Wintering, N. A.; Plossl, K.; Hochold, J.; Stabin,
M. G.; Watson, M.; Skovronsky, D.; Clark, C. M.; Kung, M. P.; Kung,
H. F. Safety, biodistribution, and dosimetry of 123I-IMPY: a novel
amyloid plaque-imaging agent for the diagnosis of Alzheimer’s disease.
J. Nucl. Med. 2006, 47, 748−54.
(14) Mathis, C. A.; Wang, Y.; Holt, D. P.; Huang, G. F.; Debnath, M.
L.; Klunk, W. E. Synthesis and evaluation of 11C-labeled 6-substituted
2-arylbenzothiazoles as amyloid imaging agents. J. Med. Chem. 2003,
46, 2740−2754.
(15) Klunk, W. E.; Engler, H.; Nordberg, A.; Wang, Y.; Blomqvist,
G.; Holt, D. P.; Bergstrom, M.; Savitcheva, I.; Huang, G. F.; Estrada,
S.; Ausen, B.; Debnath, M. L.; Barletta, J.; Price, J. C.; Sandell, J.;
Lopresti, B. J.; Wall, A.; Koivisto, P.; Antoni, G.; Mathis, C. A.;
Langstrom, B. Imaging brain amyloid in Alzheimer’s disease with
Pittsburgh compound-B. Ann. Neurol. 2004, 55, 306−319.
(16) Johnson, K. A. Amyloid imaging of Alzheimer’s disease using
Pittsburgh compound B. Curr. Neurol. Neurosci. Rep. 2006, 6, 496−
503.
(25) Choi, S. R.; Golding, G.; Zhuang, Z.; Zhang, W.; Lim, N.; Hefti,
F.; Benedum, T. E.; Kilbourn, M. R.; Skovronsky, D.; Kung, H. F.
Preclinical properties of 18F-AV-45: a PET agent for Aβ plaques in the
brain. J. Nucl. Med. 2009, 50, 1887−1894.
(26) Wong, D. F.; Rosenberg, P. B.; Zhou, Y.; Kumar, A.; Raymont,
V.; Ravert, H. T.; Dannals, R. F.; Nandi, A.; Brasic, J. R.; Ye, W.;
Hilton, J.; Lyketsos, C.; Kung, H. F.; Joshi, A. D.; Skovronsky, D. M.;
Pontecorvo, M. J. In vivo imaging of amyloid deposition in Alzheimer
disease using the radioligand 18F-AV-45 (florbetapir [corrected] F
18). J. Nucl. Med. 2010, 51, 913−920.
(27) Rowe, C. C.; Villemagne, V. L. Brain amyloid imaging. J. Nucl.
Med. 2011, 52, 1733−1740.
(28) Stephenson, K. A.; Chandra, R.; Zhuang, Z. P.; Hou, C.; Oya, S.;
Kung, M. P.; Kung, H. F. Fluoro-pegylated (FPEG) imaging agents
targeting Aβ aggregates. Bioconjugate Chem. 2007, 18, 238−246.
(29) Cheng, Y.; Ono, M.; Kimura, H.; Kagawa, S.; Nishii, R.; Saji, H.
A novel 18F-labeled pyridyl benzofuran derivative for imaging of β-
amyloid plaques in Alzheimer’s brains. Bioorg. Med. Chem. Lett. 2010,
20, 6141−6144.
(30) Ono, M.; Cheng, Y.; Kimura, H.; Cui, M.; Kagawa, S.; Nishii, R.;
Saji, H. Novel 18F-labeled benzofuran derivatives with improved
properties for positron emission tomography (PET) imaging of β-
amyloid plaques in Alzheimer’s brains. J. Med. Chem. 2011, 54, 2971−
2979.
(31) Cheng, Y.; Ono, M.; Kimura, H.; Kagawa, S.; Nishii, R.;
Kawashima, H.; Sajio, H. Fluorinated benzofuran derivatives for PET
imaging of β-amyloid plaques in Alzheimer’s disease brains. ACS Med.
Chem. Lett. 2010, 1, 321−325.
(32) Cui, M.; Ono, M.; Kimura, H.; Ueda, M.; Nakamoto, Y.;
Togashi, K.; Okamoto, Y.; Ihara, M.; Takahashi, R.; Liu, B.; Saji, H.
Novel (18)F-labeled benzoxazole derivatives as potential positron
emission tomography probes for imaging of cerebral β-amyloid
plaques in Alzheimer’s disease. J. Med. Chem. [Online early access].
DOI: 10.1021/jm300251n. Published Online: Jun 12, 2012.
(33) Qiao, J. X.; Wang, T. C.; Hu, C.; Li, J.; Wexler, R. R.; Lam, P. Y.
Transformation of anionically activated trifluoromethyl groups to
heterocycles under mild aqueous conditions. Org. Lett. 2011, 13,
1804−1807.
(34) Cui, M.; Ono, M.; Kimura, H.; Kawashima, H.; Liu, B. L.; Saji,
H. Radioiodinated benzimidazole derivatives as single photon emission
computed tomography probes for imaging of β-amyloid plaques in
Alzheimer’s disease. Nucl. Med. Biol. 2011, 38, 313−320.
(35) Toyama, H.; Ye, D.; Ichise, M.; Liow, J. S.; Cai, L.; Jacobowitz,
D.; Musachio, J. L.; Hong, J.; Crescenzo, M.; Tipre, D.; Lu, J. Q.;
Zoghbi, S.; Vines, D. C.; Seidel, J.; Katada, K.; Green, M. V.; Pike, V.
(17) Johnson, A. E.; Jeppsson, F.; Sandell, J.; Wensbo, D.; Neelissen,
J. A.; Jureus, A.; Strom, P.; Norman, H.; Farde, L.; Svensson, S. P.
AZD2184: a radioligand for sensitive detection of β-amyloid deposits.
J. Neurochem. 2009, 108, 1177−1186.
(18) Nyberg, S.; Jonhagen, M. E.; Cselenyi, Z.; Halldin, C.; Julin, P.;
Olsson, H.; Freund-Levi, Y.; Andersson, J.; Varnas, K.; Svensson, S.;
Farde, L. Detection of amyloid in Alzheimer’s disease with positron
emission tomography using [11C]AZD2184. Eur. J. Nucl. Med. Mol.
Imaging 2009, 36, 1859−1863.
(19) Ono, M.; Wilson, A.; Nobrega, J.; Westaway, D.; Verhoeff, P.;
Zhuang, Z. P.; Kung, M. P.; Kung, H. F. 11C-labeled stilbene
derivatives as Aβ-aggregate-specific PET imaging agents for
Alzheimer’s disease. Nucl. Med. Biol. 2003, 30, 565−571.
(20) Verhoeff, N. P.; Wilson, A. A.; Takeshita, S.; Trop, L.; Hussey,
D.; Singh, K.; Kung, H. F.; Kung, M. P.; Houle, S. In-vivo imaging of
M
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
W.; Cohen, R. M.; Innis, R. B. PET imaging of brain with the β-
amyloid probe, [11C]6-OH-BTA-1, in a transgenic mouse model of
Alzheimer’s disease. Eur. J. Nucl. Med. Mol. Imaging 2005, 32, 593−
600.
(36) Kung, H. F.; Choi, S. R.; Qu, W.; Zhang, W.; Skovronsky, D. 18F
stilbenes and styrylpyridines for PET imaging of A beta plaques in
Alzheimer’s disease: a miniperspective. J. Med. Chem. 2010, 53, 933−
941.
(37) Bonger, K. M.; van den Berg, R. J.; Heitman, L. H.; AP, I. J.;
Oosterom, J.; Timmers, C. M.; Overkleeft, H. S.; van der Marel, G. A.
Synthesis and evaluation of homo-bivalent GnRHR ligands. Bioorg.
Med. Chem. 2007, 15, 4841−4856.
(38) Kim, D. Y.; Kim, H. J.; Yu, K. H.; Min, J. J. Synthesis of [(18)F]-
labeled (2-(2-fluoroethoxy)ethyl)tris(4-methoxyphenyl)phosphonium
cation as a potential agent for positron emission tomography
myocardial imaging. Nucl. Med. Biol. 2012, 39, 1093−1098.
(39) Cheng, Y.; Prusoff, W. H. Relationship between the inhibition
constant (K1) and the concentration of inhibitor which causes 50 per
cent inhibition (I50) of an enzymatic reaction. Biochem. Pharmacol.
1973, 22, 3099−3108.
(40) Zhang, W.; Oya, S.; Kung, M. P.; Hou, C.; Maier, D. L.; Kung,
H. F. F-18 polyethyleneglycol stilbenes as PET imaging agents
targeting Aβ aggregates in the brain. Nucl. Med. Biol. 2005, 32, 799−
809.
N
dx.doi.org/10.1021/jm300973k | J. Med. Chem. XXXX, XXX, XXX−XXX