[11C]GSK2126458 and [18F]GSK2126458, the first radiosynthesis of new potential PET agents for imaging of PI3K and mTOR in cancers

Article abstract of DOI:10.1016/j.bmcl.2011.12.136

GSK2126458 is a highly potent inhibitor of phosphoinositide 3-kinase (PI3K) and mammalian target of rapamycin (mTOR) with low picomolar to subnanomolar activity. [11C]GSK2126458 and [18F]GSK212 6458, new potential PET agents for imaging of PI3K and mTOR in cancer, were first designed and synthesized in 40-50% and 20-30% decay corrected radiochemical yield, and 370-740 and 37-222 GBq/lmol specific activity at end of bombardment (EOB), respectively.

Full text of DOI:10.1016/j.bmcl.2011.12.136

Bioorganic & Medicinal Chemistry Letters 22 (2012) 1569–1574
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
[11C]GSK2126458 and [18F]GSK2126458, the first radiosynthesis of new
potential PET agents for imaging of PI3K and mTOR in cancers
Min Wang a, Mingzhang Gao a, Kathy D. Miller b, George W. Sledge b, Qi-Huang Zheng a,
⇑
a Department of Radiology and Imaging Sciences, Indiana University School of Medicine, 1345 West 16th Street, Room 202, Indianapolis, IN 46202, USA
b Department of Medicine, Indiana University School of Medicine, Indianapolis, IN 46202, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
GSK2126458 is a highly potent inhibitor of phosphoinositide 3-kinase (PI3K) and mammalian target of
rapamycin (mTOR) with low picomolar to subnanomolar activity. [11C]GSK2126458 and [18F]GSK212
6458, new potential PET agents for imaging of PI3K and mTOR in cancer, were first designed and synthe-
Received 6 December 2011
Revised 27 December 2011
Accepted 29 December 2011
Available online 10 January 2012
sized in 40–50% and 20–30% decay corrected radiochemical yield, and 370–740 and 37–222 GBq/
specific activity at end of bombardment (EOB), respectively.
lmol
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
[
[
11C]GSK2126458
18F]GSK2126458
Phosphoinositide 3-kinase (PI3K)
Mammalian target of rapamycin (mTOR)
Positron emission tomography (PET)
Cancer
The phosphoinositide 3-kinase (PI3K) is a critical regulator of cell
growth and transformation.1 The mammalian target of rapamycin
(mTOR) is a key component of PI3K pathway and is also a central reg-
ulator of cell growth.2 Protein kinase B (PKB), also known as Akt, is a
serine/threonine protein kinase.3 The PI3K/Akt/mTOR pathway is an
intracellular signaling pathway that is involved in several cell func-
tions including growth, proliferation, apoptosis and autophagy, and
is among the most commonly activated pathways in human cancers.4,5
PI3K has emerged as an attractive target for cancer therapeutics, and
several PI3K inhibitors are currently under evaluation in human clini-
cal trials, such as GSK2126458 (GlaxoSmithKline), BEZ235 (Novartis),
GDC-0941 (Genentech), PX-866 (ProlX), and XL765 (Exelixis).1 Among
these PI3K inhibitors, GSK2126458(2,4-difluoro-N-(2-methoxy-5-(4-
(pyridazin-4-yl)quinolin-6-yl)pyridin-3-yl)benzenesulfonamide) is a
highly potent inhibitor of the PI3K/Akt/mTOR signaling pathway
with low picomolar to subnanomolar biological activity against
PI3K/Akt/mTOR signaling pathway has become an attractive target
for cancer imaging, however, no specific imaging agents have been
developed so far.6,7 Carbon-11 and fluorine-18 labeled GSK2126458
compounds may serve as new probes for the biomedical imaging
technique positron emission tomography (PET), and enable non-
invasive monitoring of PI3K/Akt/mTOR signaling pathway in can-
cers, since most PET imaging agents currently used for evaluation
of oncologic drug treatment are 2-[18F]fluoro-2-deoxyglucose
([18F]FDG), 30-[18F]fluoro- -thymidine ([18F]FLT), and [18F]fluoromi-
L
sonidazole ([18F]FMISO).8 To radiolabel therapeutic agents as diag-
nostic agents for imaging of PI3K/Akt/mTOR signaling pathway
and monitoring of therapeutic efficacy of PI3K/Akt/mTOR inhibitors,
we have designed and synthesized [11C]GSK2126458 {2,4-difluoro-
N-(2-[11C]methoxy-5-(4-(pyridazin-4-yl)quinolin-6-yl)pyridin-3-
yl)benzenesulfonamide} and [18F]GSK2126458 {2-[18F]fluoro-4-flu-
oro-N-(2-methoxy-5-(4-(pyridazin-4-yl)quinolin-6-yl)pyridin-3-
yl)benzenesulfonamide and 2-fluoro-4-[18F]fluoro-N-(2-methoxy-
5-(4-(pyridazin-4-yl)quinolin-6-yl)pyridin-3-yl)benzenesulfon-
amide} as new potential PET agents, for the first time. Due to the
important role of PI3K/Akt/mTOR signaling pathway in cancer pro-
gression, many pharmaceutical companies and academic laborato-
ries are actively developing its inhibitors and many of these such
as LY294002, wortmannin, or rapamycin analogues are already used
in later clinical trials (II/III) through mono-targeted therapy or in
combination with other therapy for cancer treatment. Although
GSK2126458 is a relatively new developed PI3K/Akt/mTOR inhibitor
and being evaluated in phase I clinical trial through multiple-targeted
PI3K, mTOR and Akt, Ki values: 0.019 nM (PI3K
0.024 nM (PI3Kd), 0.06 nM (PI3K ), 0.18 nM (mTORC1) and 0.3 nM
(mTORC2); and IC50 values: 0.04 nM (PI3K ), 0.41 nM (pAkt-S473/
a), 0.13 nM (PI3Kb),
c
a
T47D) and 0.18 nM (pAkt-S473/BT474), originally developed and
recently reported by GlaxoSmithKline.1 GSK2126458 acting on
more than one target has resulted in a better biological response
and in enhanced therapeutic potential, and this multiple-target
inhibitor results of great interest as potential antitumor agent.5
⇑
Corresponding author. Tel.: +1 317 278 4671.
E-mail address: qzheng@iupui.edu (Q.-H. Zheng).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.12.136

1570
M. Wang et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1569–1574
therapy, it displays superior PI3K/Akt/mTOR biological activities to
other more established or matured development inhibitors.1 There-
fore, we selected GSK2126458 as based model compound for label-
ing, without changing the GSK2126458 structure. These
radiolabeled GSK2126458 compounds as potential PET probes will
provide great help to accelerate GSK2126458 clinical development
and will monitor the PI3K/Akt/mTOR pathway and understanding
the pharmacology of GSK2126458 in vivo. On the other hand, these
PET probes will be ease getting approval as IND (Investigational
New Drug) from FDA (Food and Drug Administration), and will help
to select right patient for PI3K/Akt/mTOR targeted therapy if
GSK2126458 continues success in clinical trial in the near future.
Procedures used by Knight et al. for the synthesis of
GSK2126458 (22)1 were adapted to synthesize its nitro precursors
2-nitro-GSK2126458 (23) and 4-nitro-GSK2126458 (24) for fluo-
rine-18 labeling, and desmethyl-GSK2126458 (25) for carbon-11
labeling. The synthetic strategy included an in situ borylation
and standard palladium-catalyzed cross-coupling reaction.1 In par-
ticular, the use of 6-bromo-4-(pyridazin-4-yl)quinoline (7) as the
precursor for the corresponding pinacol boronic ester 7a via cou-
pling with bis(pinacolato)diboron was pursued, as the pinacol
boronic ester 7a would be suitably poised to be coupled with aryl
and heteroaryl halides. Thus, the key intermediate 7 was synthe-
sized according to the procedures outlined in Scheme 1. Meth-
oxymethylene Meldrum’s acid (2) in situ was formed by
treatment Meldrum’s acid (1) with trimethyl orthoformate, which
was condensed with 4-bromoaniline to afford enamine intermedi-
ate 3. Thermal cyclization of 3 in Ph2O was accomplished by spon-
taneous elimination of CO2 and propanone to give quinoline-4-one
derivative 4 in overall 73% yield.9–12 Compound 4 was converted to
6-bromo-4-chloroquinoline (5) by treatment with POCl3 using
DMF as catalyst in 70% yield. The 4-chloro group of 5 was substi-
tuted by iodo via the formation of the hydrochloride salt of 5,
followed by treatment with NaI in propionitrile to afford 6-
bromo-4-iodoquinoline (6) in 90% yield.13 Palladium-catalyzed
cross-coupling reaction between 6 and 4-(tributylstannyl)pyrida-
zine in dioxane furnished the desired compound 7 in 45% yield.
Syntheses of synthons fluoronitrobenzenesulfonyl chlorides 12
and 13 were carried out in Scheme 2. Selective nucleophilic
displacement of the fluorine at ortho- and para-position activated
by the nitro group in difluoronitrobenzenes 8 and 9 was achieved
via an equimolar amount of phenylmethanethiol in the presence of
SO2Cl
NO2
F
S
a
b
b
F
F
NO2
F
NO2
10, 87%
12, 71%
8
F
F
S
SO2Cl
F
a
O2N
O2N
F
O2N
11, 96%
9
13, 76%
Scheme 2. Synthesis of synthons 12 and 13. Reagents and conditions: (a)
phenylmethanethiol, K2CO3, DMF, 0 °C to rt; (b) 1,3-dichloro-5,5-dimethylhydan-
toin, CH3CN–HOAc–H2O, 0 °C.
K2CO3 in DMF to form fluoronitrophenylbenzylthioethers 10 and
11 in 87% and 96% yield, respectively.14 Based on a recent report
in which a simple and highly effective oxidative chlorination pro-
tocol for the preparation of arenesulfonyl chlorides was de-
scribed,15 1,3-dichloro-5,5-dimethylhydantoin as oxidative
chlorination agent was used to convert 10 and 11 to their corre-
sponding sulfonyl chlorides 12 and 13 in 71% and 76% yield,
respectively.
Scheme 3 describes the synthesis of another synthon 5-bromo-
2-methoxy-3-pyridinamine (17). 5-Bromo-2-methoxy-3-nitropyr-
idine (16) from 5-bromo-2-hydroxy-3-nitropyridine (14) was
accomplished by two synthetic routes. Treatment of 14 with POCl3
using DMF as catalyst gave 5-bromo-2-chloro-3-nitropyridine 15
in 81% yield, and subsequent reaction with NaOMe in MeOH affor-
ded 16 in 94% yield. Compound 16 could also be achieved by direct
methylation of 14 with CH3I in the presence of Ag2CO3 in CH3Cl in
34% yield.16 The reduction of 16 was carried out with SnCl2ꢀ2H2O in
EtOAc to give 17 in 76% yield.
Synthesis of GSK2126458, 2-nitro-GSK2126458, 4-nitro-
GSK2126458 and desmethyl-GSK2126458 is shown in Scheme 4.
Coupling benzenesulfonyl chlorides 18, 12 and 13 with 17 in pyr-
idine provided the corresponding aryl bromides derivatives 19, 20
and 21 in 46%, 28% and 26% yield, respectively. The palladium-
catalyzed cross-coupling reaction of bis(pinacolato)diboron with
aryl bromide 7 gave arylboronic ester 7a in situ. The reaction
was catalyzed by PdCl2(dppf)-CH2Cl2 in the presence of KOAc in
dioxane. Arylboronic ester 7a was coupled with aryl bromides
derivatives 19, 20 and 21 to yield the desired standard 22, nitro
precursors 23 and 24 in 71%, 21% and 24% yield, respectively. Di-
rect demethylation was performed by treatment of GSK2126458
22 with trimethylsilyl iodide in CH3CN to afford the desmethylated
precursor 25 in 49% yield.
O
O
Br
O
O
Br
O
O
H
a
b
O
O
c
Synthesis of the target tracer [11C]GSK2126458 ([11C]22) is
indicated in Scheme 5. Desmethyl-GSK2126458 precursor 25 was
O
O
N
N
H
O
O
H
O
OMe
1
2
4, 73%
3
labeled by
[
11C]methyl triflate ([11C]CH3OTf)17,18 through O-
N
N
[
11C]methylation19 at 80 °C under basic condition (2 N NaOH)
Cl
I
and isolated by a semi-preparative high performance liquid chro-
matography (HPLC) method (C-18 column) and a solid-phase
extraction (SPE) method (C-18 Plus Sep-Pak cartridge) (a second
purification or isolation process)20 to produce the corresponding
pure radiolabeled compound [11C]22 in 40–50% radiochemical
yield, decay corrected to end of bombardment (EOB), based on
Br
Br
e
d
Br
f
N
N
N
6, 90%
5, 70%
7, 45%
N
N
N
O
B
g
O
MeO
O2N
N
MeO
H2N
N
Cl
N
HO
N
a
b
d
Br
7a
Br
O2N
Br
O2N
Br
14
16, 94% (b); 34% (c)
17, 76%
15, 81%
Scheme 1. Synthesis of key intermediate 7. Reagents and conditions: (a) trimethyl
orthoformate, reflux; (b) 4-bromoaniline, trimethyl orthoformate, reflux; (c) Ph2O,
250 °C; (d) POCl3, DMF (cat.), reflux; (e) 2 M HCl in Et2O, THF, room temperature
(rt); NaI, propiontrile, reflux; (f) 4-(tributylstannanyl)pyridazine, PdCl2(dppf)2-
CH2Cl2, dioxane, reflux; (g) bis(pinacolato)diboron, PdCl2(dppf)2-CH2Cl2, KOAc,
dioxane, reflux.
c
Scheme 3. Synthesis of synthon 17. Reagents and conditions: (a) POCl3, DMF (cat.),
reflux; (b) NaOMe, MeOH, 0 °C to rt; (c) MeI, Ag2CO3, CH3Cl; (d) SnCl2ꢀ2H2O, EtOAc,
reflux.

M. Wang et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1569–1574
1571
MeO
N
N
N
N
N
O
N
N
O
O
O
MeO
H2N
N
S
a
S
b
MeO
O
N
MeO
O
S
N
H
N
Cl
N
H
Br
+
O
O
a
Br
N
S
N
H
R2
R1
R2
R1
18, R1=R2=F
19, R1=R2=F, 46%
17
12, R1=NO2, R2=F
13, R1=F, R2=NO2
F
18F
F
NO2
20, R1=NO2, R2=F, 28%
21, R1=F, R2=NO2, 26%
2-[18F]22 (2-[18F]GSK2126458), 20-30%
23 (2-Nitro-GSK2126458)
N
N
N
N
N
N
N
N
N
MeO
O
S
N
H
N
MeO
O
S
N
H
N
HO
O
N
MeO
N
O
O
O
O
O
a
S
F
S
N
H
c
N
H
R2
R1
N
N
O2N
F
18F
F
F
4-[18F]22 (4-[18F]GSK2126458), 20-30%
24 (4-Nitro-GSK2126458)
22 (GSK2126458), R1=R2=F, 71%
25 (Desmethyl-GSK2126458), 49%
23 (2-Nitro-GSK2126458), R1=NO2, R2=F, 21%
24 (4-Nitro-GSK2126458), R1=F, R2=NO2, 24%
Scheme 6. Synthesis of [18F]GSK2126458. Reagents and conditions: (a) K[18F]F/
Kryptofix 2.2.2, DMSO, 140 °C.
Scheme 4. Synthesis of standard GSK2126458 22 and precursors 2-nitro-
GSK2126458 23, 4-nitro-GSK2126458 24, and desmethyl-GSK2126458 25.
Reagents and conditions: (a) pyridine, 0 °C to rt; (b) 7, bis(pinacolato)diboron,
PdCl2(dppf)2-CH2Cl2, KOAc, dioxane, reflux; PdCl2(dppf)2-CH2Cl2, 2 M Na2CO3,
reflux; (c) iodotrimethylsilane, CH3CN, reflux.
labeled with K[18F]F/Kryptofix 2.2.2 through nucleophilic substitu-
tion and isolated by a semi-preparative HPLC method with C-18 col-
umn and a SPE method with a C-18 Plus Sep-Pak cartridge (a second
purification or isolation process)20,26 to produce the corresponding
pure radiolabeled compound 2-[18F]22 or 4-[18F]22 in 20–30%
radiochemical yield, decay-corrected to EOB, based on H[18F]F. Like-
wise, addition of NaHCO3 to quench the radiolabeling reaction and
to dilute the radiolabeling mixture prior to the injection onto the
semi-preparative HPLC column for purification gave better separa-
tion of labeled product 2-[18F]GSK2126458 or 4-[18F]GSK2126458
from its corresponding 2-nitro or 4-nitro precursor.20,22 The radio-
synthesis was performed using a self-designed automated multi-
purpose [18F]-radiosynthesis module.20,26 The overall synthesis,
purification and formulation time was 50–60 min from EOB. The
N
N
N
N
H311CO
N
HO
O
N
O
O
O
a
S
S
F
N
H
N
H
N
F
F
N
F
[
11C]22 ([11C]GSK2126458), 40-50%
25 (Desmethyl-GSK2126458)
Scheme 5. Synthesis of
[
[
11C]GSK2126458. Reagents and conditions: (a)
11C]CH3OTf, 2 N NaOH, CH3CN, 80 °C.
specific radioactivity was 37–222 GBq/lmol at EOB. No-carrier-
[
11C]CO2. [11C]CH3OTf is a proven methylation reagent with greater
added [18F]fluoride ion in [18O]water was trapped without a QMA
cartridge. This way20,26,27 significantly increased the specific activ-
ity of the prepared F-18 labeled product. As indicated in the litera-
ture,27 when the cyclotron-produced [18F]fluoride ion was dried
without the use of a cartridge, but through cycles of evaporation
with added acetonitrile, the specific radioactivity of the prepared
2-[18F]GSK2126458 and 4-[18F]GSK2126458 was substantially
higher, and was similar to that we achieved in the radiosynthesis
of [18F]fallypride and [18F]PBR06.20,26 The reason was that there
was a low-level contamination of QMA anionic resins with fluoride
ion.27 The amounts of 2-nitro or 4-nitro precursor used were ꢁ1 mg.
A large amount of precursor would increase the radiochemical yield
of 2-[18F]GSK2126458 or 4-[18F]GSK2126458, but decrease the
chemical purity of the 2-[18F]22 or 4-[18F]22 tracer solution due to
precursor contamination. To our F-18 labeling experiences on
reactivity than commonly used [11C]methyl iodide ([11C]CH3I),21
and thus, the radiochemical yield of [11C]GSK2126458 was rela-
tively high. Addition of NaHCO3 to quench the radiolabeling reac-
tion and to dilute the radiolabeling mixture prior to the injection
onto the semi-preparative HPLC column for purification gave bet-
ter separation of [11C]22 from its heteroaryl hydroxyl precur-
sor.20,22 The radiosynthesis was performed in
a home-built
automated multi-purpose [11C]-radiosynthesis module, allowing
measurement of specific radioactivity during synthesis.23,24 The
overall synthesis, purification and formulation time was 30–
40 min from EOB. The specific radioactivity was in a range of
370–740 GBq/lmol at EOB. Chemical purity and radiochemical
purity were determined by analytical HPLC.25 The chemical purity
of the precursor desmethyl-GSK2126458 and reference standard
GSK2126458 was >96%. The radiochemical purity of the target tra-
cer [11C]GSK2126458 was >99% determined by radio-HPLC through
[
18F]fallypride and [18F]PBR06,20,26 although the HPLC systems we
employed have shown good separation from precursor and prod-
ucts, there always was a co-elution of the F-18 labeled product with
its corresponding precursor from the HPLC column. 2-[18F]GSK-
2126458 and 4-[18F]GSK2126458 were also in the same case. In
addition, a large amount of precursor would also decrease the
specific activity of final labeled product due to potential F-18/F-19
exchange during the radiolabeling. The reaction solvent and tem-
perature were either CH3CN/120 °C or DMSO/140 °C. Radiolabeling
procedure with DMSO at 140 °C resulted in higher radiochemical
yield.20,26,28 Chemical purity and radiochemical purity were
determined by analytical HPLC.25 The chemical purity of 2-nitro-
GSK2126458 (23) and 4-nitro-GSK2126458 precursors and refer-
ence standard GSK2126458 was >96%. The radiochemical purity of
the target tracers 2-[18F]GSK2126458 and 4-[18F]GSK2126458 was
>99%, and the chemical purity of 2-[18F]GSK2126458 and
4-[18F]GSK2126458 was >90% determined by HPLC methods. Most
impurity of 2-[18F]GSK2126458 and 4-[18F]GSK2126458 was their
corresponding precursor contamination from the HPLC co-elution.
Likewise, a C-18 Plus Sep-Pak cartridge was used to significantly im-
prove the chemical purity of the tracer solution as aforemen-
c
[
-ray (PIN diode) flow detector, and the chemical purity of
11C]GSK2126458 was >93% determined by reverse-phase HPLC
through UV flow detector. A C-18 Plus Sep-Pak cartridge was used
to significantly improve the chemical purity of the tracer solution.
Initial HPLC purification was employed to separate the labeled
product from its un-reacted excess precursor and other labeled
by-products. The second SPE purification with Sep-Pak20,25 was
employed, instead of rotary evaporator, to remove potential impu-
rities from the HPLC co-elution with the precursor and from the
residual solvents including HPLC mobile phase solvents and mod-
ule set-up cleaning solvents. Rotary evaporation was unable to per-
form in this regard. Moreover, it could result in the decomposition
of the labeled product such as desmethylation during the heating.
The chemical purity of the [11C]GSK2126458 tracer solution with
Sep-Pak purification was increased higher 10–20% than that with-
out Sep-Pak purification.20,26
Synthesis of the target tracers 2-[18F]GSK2126458 (2-[18F]22)
and 4-[18F]GSK2126458 (4-[18F]22) is outlined in Scheme 6. 2-Ni-
tro-GSK2126458 (23) or 4-nitro-GSK2126458 (24) precursor was

1572
M. Wang et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1569–1574
tioned.20,26 The chemical purity of the 2-[18F]GSK2126458 or 4-
18F]GSK2126458 tracer solution with Sep-Pak purification was in-
Zhang, W.-G.; Hollander, I.; Gibbons, J. J.; Abraham, R. T.; Ayral-Kaloustian, S.;
Mansour, T. S.; Yu, K. J. Med. Chem. 2009, 52, 5013.
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6. Fuereder, T.; Wanek, T.; Pflegerl, P.; Jaeger-Lansky, A.; Hoeflmayer, D.;
Strommer, S.; Kuntner, C.; Wrba, F.; Werzowa, J.; Hejna, M.; Müller, M.;
Langer, O.; Wacheck, V. Clin. Cancer Res. 2011, 17, 5322.
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Cullinane, C.; Hicks, R. J.; Johnstone, R. W.; McArthur, G. A. Mol. Cancer Ther.
2011, 10, 1440.
8. Figueiras, R. G.; Padhani, A. R.; Goh, V. J.; Vilanova, J. C.; González, S. B.; Martín,
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[
creased higher 10–20% than that without Sep-Pak purification.
GSK2126458 was 18F-labeled at 2- and 4-positions to be 2-
[
18F]GSK2126458 and 4-[18F]GSK2126458. No significantly differ-
ent results were identified. This could be that both ortho-nitro pre-
cursor and para-nitro precursor have similar reactivity for F-18
labeling.
The synthetic information of GSK2126458 was limited in the lit-
erature.1 Thus, the experimental details and characterization data
for compounds 4–7, 10–13, 15–17, and 19–25, and for the tracers
[
11C]22, 2-[18F]22 and 4-[18F]22 are given.29
In summary, [11C]GSK2126458 and [18F]GSK2126458 were first
designed and synthesized as new potential PET agents for imaging
of PI3K and mTOR in cancers. New desmethyl-GSK2126458 precur-
sor (for C-11 labeling) and 2-nitro-GSK2126458 and 4-nitro-
GSK2126458 precursors (for F-18 labeling) have been designed
and synthesized for the first time. Desmethyl-GSK2126458 was la-
beled with [11C]CH3OTf, and isolated by semi-preparative HPLC
combined with SPE purification to provide [11C]GSK2126458, a car-
bon-11 labeled form of GSK2126458, in high radiochemical yield
with excellent specific activity and shorter reaction times. 2-Ni-
tro-GSK2126458 and 4-nitro-GSK2126458 were labeled with
K[18F]F/Kryptofix 2.2.2 through nucleophilic substitution, and iso-
lated by semi-preparative HPLC combined with SPE purification to
produce 2-[18F]GSK2126458 and 4-[18F]GSK2126458, fluorine-18
labeled forms of GSK2126458, in moderate radiochemical yield,
high chemical purity and specific activity. Automated self-designed
multi-purpose [11C]- and [18F]-radiosynthesis modules for the syn-
thesis of [11C]GSK2126458 and [18F]GSK2126458 have been built,
featuring the measurement of specific activity by the on-the-fly
technique. New and improved results in the synthetic methodology,
radiolabeling, preparative separation and analytical details for
GSK2126458, desmethyl-GSK2126458, 2-nitro-GSK2126458 and
4-nitro-GSK2126458, [11C]GSK2126458 and [18F]GSK2126458 have
been presented. These methods are efficient and convenient. It is
anticipated that the approaches for the design, synthesis and auto-
mation of new tracer and radiolabeling precursors, and improve-
ments to increase radiochemical yield, chemical purity and
specific activity of the tracers described here can be applied with
advantage to the synthesis of other 11C- and 18F-radiotracers for
PET imaging. These chemistry results warrant future preclinical
and clinical PET studies of [11C]GSK2126458 and [18F]GSK2126458
in animals and humans to image cancer. This work will provide use-
ful information for other investigators who will perform in vitro and
in vivo biological evaluations of these [11C] and [18F] tracers and de-
velop new PET tracers for imaging the PI3K and mTOR in vivo.
10. Ghosh, B.; Antonio, T.; Reith, M. E. A.; Dutta, A. K. J. Med. Chem. 2010, 53, 2114.
11. da Silva, L. E.; Joussef, A. C.; Pacheco, L. K.; Albino, D. B. L.; Duarte, A. M. C.;
Steindel, M.; Rebelo, R. A. Lett. Drug Des. Disc. 2007, 4, 154.
12. Walz, A. J.; Sundberg, R. J. J. Org. Chem. 2000, 65, 8001.
13. Wolf, C.; Tumambac, G. E.; Villalobos, C. N. Synlett 2003, 1801.
14. Zhersh, S.; Lukin, O.; Matvienko, V.; Tolmachev, A. Tetrahedron 2010, 66, 5982.
15. Pu, Y.-M.; Christesen, A.; Ku, Y.-Y. Tetrahedron Lett. 2010, 51, 418.
16. Blomgren, P. A.; Currie, K. S.; Kropf, J. E.; Lee, T.; Darrow, J. W.; Mitchell, S. A.;
Xu, J.; Schmitt, A. C. WO Patent No. 033854 A1, 2008.
17. Jewett, D. M. Int. J. Radiat. Appl. Instrum. A 1992, 43, 1383.
18. Mock, B. H.; Mulholland, G. K.; Vavrek, M. J. Nucl. Med. Biol. 1999, 26, 467.
19. Wang, M.; Yoder, K. K.; Gao, M.; Mock, B. H.; Xu, X.-M.; Saykin, A. J.; Hutchins,
G. D.; Zheng, Q.-H. Bioorg. Med. Chem. Lett. 2009, 19, 5636.
20. Gao, M.; Wang, M.; Mock, B. H.; Glick-Wilson, B. E.; Yoder, K. K.; Hutchins, G.
D.; Zheng, Q.-H. Appl. Radiat. Isot. 2010, 68, 1079.
21. Allard, M.; Fouquet, E.; James, D.; Szlosek-Pinaud, M. Curr. Med. Chem. 2008, 15,
235.
22. Gao, M.; Wang, M.; Hutchins, G. D.; Zheng, Q.-H. Appl. Radiat. Isot. 2008, 66,
1891.
23. Mock, B. H.; Zheng, Q.-H.; DeGrado, T. R. J. Labelled Compd. Radiopharm. 2005,
48, S225.
24. Mock, B. H.; Glick-Wilson, B. E.; Zheng, Q.-H.; DeGrado, T. R. J. Labelled Compd.
Radiopharm. 2005, 48, S224.
25. Zheng, Q.-H.; Mock, B. H. Biomed. Chromatogr. 2005, 19, 671.
26. Wang, M.; Gao, M.; Miller, K. D.; Zheng, Q.-H. Steroids 2011, 76, 1331.
27. Lu, S.; Giamis, A. M.; Pike, V. W. Curr. Radiopharm. 2009, 2, 49.
28. Kuhnast, B.; Hinnen, F.; Dolle, F. J. Labelled Compd. Radiopharm. 2009, 52, S286.
29. (a) All commercial reagents and solvents were purchased from Sigma–Aldrich
and Fisher Scientific and used without further purification. [11C]CH3OTf was
prepared according to
determined on MEL-TEMP II capillary tube apparatus and were
a
literature procedure.18 Melting points were
a
uncorrected. 1H NMR spectra were recorded on Bruker Avance II 500 MHz
NMR spectrometers using tetramethylsilane (TMS) as an internal standard.
Chemical shift data for the proton resonances were reported in parts per
million (ppm, d scale) relative to internal standard TMS (d 0.0), and coupling
constants (J) were reported in hertz (Hz). The high resolution mass spectra
(HRMS) were obtained using a Thermo Electron Corporation MAT 95XP-Trap
spectrometer. Chromatographic solvent proportions are indicated as
volume:volume ratio. Thin-layer chromatography (TLC) was run using
Analtech silica gel GF uniplates (5 ꢂ 10 cm2). Plates were visualized under
UV light. Preparative TLC was run using Analtech silica gel UV 254 plates
(20 ꢂ 20 cm2). Normal phase flash column chromatography was carried out on
EM Science Silica Gel 60 (230–400 mesh) with a forced flow of the indicated
solvent system in the proportions described below. All moisture- and/or air-
sensitive reactions were performed under a positive pressure of nitrogen
maintained by a direct line from a nitrogen source. Analytical HPLC was
Acknowledgments
performed using a Prodigy (Phenomenex) 5
3:1:3 CH3CN/MeOH/20 mM, pH 6.7 phosphate (buffer solution) mobile phase;
flow rate 1.5 mL/min; and UV (254 nm) and -ray (PIN diode) flow detectors.
Semi-preparative HPLC was performed using a Prodigy (Phenomenex) 5 m,
10 ꢂ 250 mm C-18 column; 3:1:3 CH3CN/MeOH/20 mM, pH 6.7 phosphate
(buffer solution) mobile phase; 5.0 mL/min flow rate; UV (254 nm) and -ray
(PIN diode) flow detectors. C18 Plus Sep-Pak cartridges were obtained from
lm C-18 column, 4.6 ꢂ 250 mm;
This work was partially supported by the Breast Cancer Re-
search Foundation. 1H NMR spectra were recorded on a Bruker
Avance II 500 MHz NMR spectrometer in the Department of Chem-
istry and Chemical Biology at Indiana University Purdue University
Indianapolis (IUPUI), which is supported by a NSF-MRI grant CHE-
0619254.
c
l
c
Waters Corporation (Milford, MA). Sterile Millex-FG 0.2
obtained from Millipore Corporation (Bedford, MA).
lm filter units were
(b) 6-Bromoquinolin-4(1H)-one (4). A solution of Meldrum’s acid (1) (32.4 g,
225.0 mmol) in trimethyl orthoformate (250 mL) was stirred and heated at
reflux for 3 h under N2 to give 2. The solution was allowed to cool to ꢁ50 °C,
and 4-bromoaniline (25.8 g, 150.0 mmol) in trimethyl orthoformate (80 mL)
was added dropwise. The reaction mixture was heated at reflux for 2 h. After
removal of the solvent in vacuo, the residue was diluted with hexanes, filtered,
and the solid was washed with hexanes and dried to give Meldrum’s acid
derivative 3 as a yellow solid. Compound 3 was added slowly in small portions
(ꢁ1.0 g each) to preheated (245 °C) Ph2O (500 mL). Caution not to perform
this addition too rapidly must be taken as gas violently evolves! The
reaction mixture was stirred and heated at 250 °C for 15 min, then allowed to
cool to room temperature (rt) and diluted with hexanes, filtered. The solid was
washed with hexanes, then 30% Et2O/hexanes. The crude product was purified
References and notes
1. Knight, S. D.; Adams, N. D.; Burgess, J. L.; Chaudhari, A. M.; Darcy, M. G.;
Donatelli, C. A.; Luengo, J. I.; Newlander, K. A.; Parrish, C. A.; Ridgers, L. H.;
Sarpong, M. A.; Schmidt, S. J.; Van Aller, G. S.; Carson, J. D.; Diamond, M. A.;
Elkins, P. A.; Gardiner, C. M.; Garver, E.; Gilbert, S. A.; Gontarek, R. R.; Jackson, J.
R.; Kershner, K. L.; Luo, L.; Raha, K.; Sherk, C. S.; Sung, C.-M.; Sutton, D.;
Tummino, P. J.; Wegrzyn, R. J.; Auger, K. R.; Dhanak, D. ACS Med. Chem. Lett.
2010, 1, 39.
2. Zask, A.; Verheijen, J. C.; Curran, K.; Kaplan, J.; Richard, D. J.; Nowak, P.;
Malwitz, D. J.; Brooijmans, N.; Bard, J.; Svenson, K.; Lucas, J.; Toral-Barza, L.;

M. Wang et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1569–1574
1573
by silica gel column chromatography (16:1–10:1 CH2Cl2/MeOH) to afford 4
(24.6 g, 73%) as a pale brown solid, mp 282–284 °C (lit30 282–284 °C). 1H NMR
(DMSO-d6) d 11.93 (br s, 1H, NH), 8.17 (d, J = 2.0 Hz, 1H, Ar-H), 7.96 (dd, J = 7.5,
6.0 Hz, 1H), 7.79 (dd, J = 9.0, 2.5 Hz, 1H, Ar-H), 7.53 (d, J = 8.5 Hz, 1H), 6.08 (d,
J = 2.5 Hz, 1H, Ar-H).
(c) 6-Bromo-4-chloroquinoline (5). A mixture of 4 (17.0 g, 131.5 mmol), POCl3
(180 mL) and anhydrous DMF (18 mL) was stirred and heated at reflux for 2 h.
After removal of POCl3, the residue was poured into ice water and extracted
with EtOAc. The combined organic layers were washed with saturated aqueous
NaHCO3, brine, dried over anhydrous Na2SO4, filtered and concentrated in
vacuo. The crude product was purified by silica gel column chromatography
(4:1 hexanes/EtOAc) to afford 5 (12.8 g, 70%) as a white solid, mp 111–112 °C
(lit31 111–112 °C). 1H NMR (CDCl3) d 8.79 (d, J = 5.0 Hz, 1H, Ar-H), 8.40 (d,
J = 2.5 Hz, 1H, Ar-H), 8.00 (d, J = 9.0 Hz, 1H, Ar-H), 7.84 (dd, J = 9.0, 2.0 Hz, 1H,
Ar-H), 7.52 (d, J = 5.0 Hz, 1H, Ar-H).
(d) 6-Bromo-4-iodoquinoline (6). To a solution of 5 (11.0 g, 45.6 mmol) in
anhydrous THF (150 mL) was added 2 M HCl in Et2O (29 mL, 58.0 mmol)
dropwise. After stirring at rt for 30 min, the solvent was removed in vacuo and
the solid was dried to afford 6-bromo-4-chloroquinoline hydrochloride as an
off-white solid. The hydrochloride salt and anhydrous NaI (34.2 g, 228.3 mmol)
were suspended in propionitrile (300 mL). After the reaction mixture was
stirred and heated at reflux for 96 h, it was cooled to rt. 10% aqueous K2CO3
(200 mL) was added, followed by 5% aqueous Na2SO3 (80 mL), and the mixture
was extracted with CH2Cl2. The combined organic layers were washed with
brine, dried over anhydrous Na2SO4, filtered and concentrated in vacuo. The
crude product was purified by silica gel column chromatography (100:10:1–
100:20:1 hexanes/EtOAc/Et3N) to afford 6 (13.7 g, 90%) as a white solid, mp
140–142 °C. 1H NMR (CDCl3) d 8.45 (d, J = 4.5 Hz, 1H, Ar-H), 8.21 (d, J = 2.0 Hz,
1H, Ar-H), 8.01 (d, J = 4.5 Hz, 1H, Ar-H), 7.93 (d, J = 9.0 Hz, 1H, Ar-H), 7.81 (dd,
J = 9.0, 2.5 Hz, 1H, Ar-H).
(k) 5-Bromo-2-methoxy-3-nitropyridine (16). Method A: to
a
cooled
suspension of 15 (15.0 g, 63.2 mmol) in anhydrous MeOH (60 mL) was added
20% NaOMe in MeOH (15 mL, 66.4 mmol) dropwise at 0 °C. After the reaction
mixture was allowed to warm to rt and stirred overnight, the pale yellow
precipitate was filtered off to give the crude product. The crude product was
diluted with water and stirred for 1 h. The solid was collected by filtration,
washed with water and dried to afford 16 (9.22 g, 62%) as a pale yellow solid.
The original filtrate was concentrated in vacuo and diluted with water.
Saturated aqueous NH4Cl was added and the mixture was stirred for 1 h. The
solid was collected by filtration, washed with water and dried in a vacuum
oven (40 °C) to give the second crop of 16 (4.74 g, 32%) as a pale yellow solid,
mp 88–90 °C. 1H NMR (CDCl3) d 8.45 (d, J = 2.5 Hz, 1H, Ar-H), 8.39 (d, J = 2.0 Hz,
1H, Ar-H), 4.11 (s, 3H, OCH3). Method B: to
a suspension of 14 (5.0 g,
22.8 mmol) in anhydrous CH3Cl (50 mL) under N2 in the dark (wrapped in
aluminum foil) was added Ag2CO3 (7.55 g, 28.0 mmol), followed by CH3I
(14.2 mL, 230.0 mmol). After stirring at rt for 48 h, the reaction mixture was
filtered through
a pad of Celite, washed with abundant CH2Cl2, and
concentrated in vacuo. The crude product was purified by silica gel column
chromatography (10:1 hexanes/EtOAc) to afford 16 (1.86 g, 34%) as a pale
yellow solid. The analytical data were same as aforementioned.
(l) 5-Bromo-2-methoxy-3-pyridinamine (17). To
a solution of 16 (12.0 g,
51.7 mmol) in EtOAc (200 mL) was added SnCl2ꢀ2H2O (50.0 g, 221.6 mmol).
The reaction mixture was stirred and heated at reflux for 4 h. After removal of
the solvent, the residue was treated with 2 N aqueous NaOH and extracted
with CH2Cl2. The combined organic layers were washed with brine, dried over
anhydrous Na2SO4, filtered and concentrated in vacuo. The crude product was
purified by silica gel column chromatography (9:1 hexanes/EtOAc) to afford 17
(8.0 g, 76%) as a pale yellow solid, mp 58–60 CH3. 1H NMR (CDCl3) d 7.58 (d,
J = 2.0 Hz, 1H, Ar-H), 6.98 (d, J = 2.0 Hz, 1H, Ar-H), 3.96 (s, 3H, OCH3).
(m)
N-(5-Bromo-2-methoxypyridin-3-yl)-2,4-difluorobenzenesulfonamide
(e) 6-Bromo-4-(pyridazin-4-yl)quinoline (7).
10.8 mmol), 4-(tributylstannyl)pyridazine
A
suspension of
6
(3.61 g,
and
(19). To a cooled solution of 17 (4.0 g, 19.8 mmol) in pyridine (40 mL) was
added 2,4-difluorobenzenesulfonyl chloride (18) (3.2 mL, 23.8 mmol) dropwise
at 0 °C. After the reaction mixture was allowed to warm to rt and stirred
overnight, it was diluted with cold water. The precipitate was collected by
filtration, washed with water, dried to afford 19 (3.42 g, 46%) as a pale pink
solid, mp 154–156 °C. 1H NMR (CDCl3) d 7.91–7.88 (m+d, J = 2.0 Hz, 2H, Ar-H),
7.82 (d, J = 2.0 Hz, 1H, Ar-H), 7.22 (br s, 1H, NH), 7.00–6.92 (m, 2H, Ar-H), 3.90
(s, 3H, OCH3).
(n) N-(5-Bromo-2-methoxypyridin-3-yl)-4-fluoro-2-nitrobenzenesulfonamide
(20). To a cooled solution of 17 (1.0 g, 5.0 mmol) in pyridine (6 mL) was added
a solution of 12 (1.19, 5.0 mmol) in pyridine (2 mL) dropwise at 0 °C. After the
reaction mixture was allowed to warm to rt and stirred overnight, it was
poured into ice water and extracted with EtOAC. The combined organic layers
were washed with brine, dried over anhydrous Na2SO4, filtered and
concentrated in vacuo. The crude product was purified by silica gel column
chromatography (3:1 hexanes/EtOAc) to afford 20 (561 mg, 28%) as a yellow
solid, mp 140–142 °C. 1H NMR (CDCl3) d 8.03 (dd, J = 9.0, 5.5 Hz, 1H, Ar-H), 8.01
(d, J = 2.0 Hz, 1H, Ar-H), 7.96 (d, J = 2.0 Hz, 1H, Ar-H), 7.83 (br s, 1H, NH), 7.68
(dd, J = 7.5, 2.5 Hz, 1H, Ar-H), 7.41–7.36 (m, 1H, Ar-H), 3.83 (s, 3H, OCH3).
HRMS (ESI, m/z): calcd for C12H10N3O5FSBr ([M+H]+) 405.9503, found 405.9504.
(o) N-(5-Bromo-2-methoxypyridin-3-yl)-2-fluoro-4-nitrobenzenesulfonamide
(21). A similar procedure for 20 was used to prepare 21 from 13 and 17 in 26%
yield as a yellow solid, mp 144–146 °C. 1H NMR (CDCl3) d 8.14–8.06 (m, 3H, Ar-
H), 7.93 (d, J = 2.5 Hz, 1H, Ar-H), 7.86 (d, J = 2.0 Hz, 1H, Ar-H), 7.30 (br s, 1H,
NH), 3.89 (s, 3H, OCH3). HRMS (CI, m/z): calcd for C12H10N3O5FSBr ([M+H]+)
405.9503, found 405.9497.
(4.0 g, 10.8 mmol)
PdCl2(dppf)-CH2Cl2 (632.0 mg, 0.8 mmol) in anhydrous 1,4-dioxane (50 mL)
was stirred and heated at reflux for overnight. After cooling to rt, the reaction
mixture was filtered through a pad of Celite, washed with abundant CH2Cl2,
and concentrated in vacuo. The crude product was purified by silica gel column
chromatography (50:1–20:1 EtOAc/MeOH) to afford 7 (1.38 g, 45%) as a pale
brown solid, mp 176 °C (dec.). 1H NMR (CDCl3) d 9.46 (d, J = 5.0 Hz, 1H, Ar-H),
9.39 (s, 1H, Ar-H), 9.06 (d, J = 3.0 Hz, 1H, Ar-H), 8.16 (d, J = 8.5 Hz, 1H, Ar-H),
7.90 (dd, J = 9.0, 2.0 Hz, 1H, Ar-H), 7.88 (d, J = 1.5 Hz, 1H, Ar-H), 7.67 (d,
J = 3.0 Hz, 1H, Ar-H), 7.43 (d, J = 3.5 Hz, 1H, Ar-H).
(f) 1-(Benzylsulfanyl)-4-fluoro-2-nitrobenzene (10). To a suspension of 2,5-
diflouoronitrobenzene (8) (15.0 g, 94.3 mmol), anhydrous K2CO3 (26.1 g,
138.2 mmol) in anhydrous DMF (100 mL) was added phenylmethanethiol
(11.7 g, 94.3 mmol) dropwise at 0 °C. After the reaction mixture was allowed to
warm to rt and stirred for 2 h, it was poured into ice-water. The precipitate was
filtered off, washed with water and dried. The crude product was purified by
silica gel column chromatography (2:1 hexanes/CHCl3) to afford 10 (21.6 g,
87%) as a yellow solid; mp 98–99 °C (lit15 104 °C). 1H NMR (CDCl3) d 7.91 (dd,
J = 8.5, 3.0 Hz, 1H, Ar-H), 7.43–7.40 (m, 3H, Ar-H), 7.35–7.32 (m, 2H, Ar-H),
7.30–7.25 (m, 2H, Ar-H), 4.39 (s, 2H, Ph-CH2).
(g) 1-(Benzylsulfanyl)-2-fluoro-4-nitrobenzene (11). To a suspension of 3, 4-
diflouoronitrobenzene (9) (15.0 g, 94.3 mmol), anhydrous K2CO3 (26.1 g,
138.2 mmol) in anhydrous DMF (100 mL) was added phenylmethanethiol
(11.7 g, 94.3 mmol) dropwise at 0 °C. After the reaction mixture was allowed to
warm to rt and stirred for 2 h, it was poured into ice-water. The precipitate was
filtered off, washed with water and dried to afford 11 (23.7 g, 96%) as a yellow
solid, mp 122–123 °C (lit15 126 °C). 1H NMR (CDCl3) d 7.94–7.91 (m, 1H, Ar-H),
7.88 (dd, J = 9.5, 2.5 Hz, 1H, Ar-H), 7.38–7.26 (m, 6H, Ar-H), 4.24 (s, 2H, Ph-
CH2).
(h) 4-Fluoro-2-nitrobenzene-1-sulfonyl chloride (12). To a cooled solution of
10 (10.5 g, 40.0 mmol) in CH3CN–HOAc–H2O (400 mL, 5 mL, 10 mL) was added
1,3-dichloro-5,5-dimethylhydantoin (15.8 g, 80.0 mmol) portionwise at 0 °C.
After stirring at 0 °C for 3 h, the reaction mixture was concentrated to near
dryness in vacuo. The crude product was diluted with CH2Cl2 (500 mL), and the
solution was cooled down to ꢁ0 °C. 5% aqueous NaHCO3 (550 mL) was added
slowly at <10 °C, the mixture was stirred at 0 °C for 15 min, The separated
organic layer was washed with cooled brine, dried over anhydrous Na2SO4,
filtered and concentrated in vacuo. This liquid product was used without
further purification. Small amount of the analytical pure sample was obtained
by preparative TLC plate (9:1 hexanes/EtOAc) to afford 12 (71% yield) as a pale
yellow solid, mp 44–45 °C (lit15 45–47 °C). 1H NMR (CDCl3) d 8.32 (dd, J = 9.0,
5.0 Hz, 1H, Ar-H), 7.64 (dd, J = 7.5, 2.5 Hz, 1H, Ar-H), 7.61–7.57 (m, 1H, Ar-H).
(i) 2-Fluoro-4-nitrobenzene-1-sulfonyl chloride (13). A similar procedure for
12 was used to prepare 13 from 11 in 76% yield as a pale yellow solid, mp 69–
70 °C (lit15 70 °C). 1H NMR (CDCl3) d 8.34 (dd, J = 9.5, 2.0 Hz, 1H, Ar-H), 8.31–
8.27 (m, 2H, Ar-H).
(p) 2,4-Difluoro-N-(2-methoxy-5-(4-(pyridazin-4-yl)quinolin-6-yl)pyridin-3-
yl)benzenesulfonamide (GSK2126458, 22).
A suspension of 7 (570.0 mg,
2.0 mmol), bis(pinacolato)diboron (559.4 mg, 2.2 mmol), PdCl2(dppf)-CH2Cl2
(82.0 mg, 0.1 mmol) and KOAc (392.7 mg, 4.0 mmol) in anhydrous 1,4-dioxane
(20 mL) was stirred and heated at reflux for 3 h. The reaction mixture was
treated with 19 (790.0 mg, 2.1 mmol), 2 M Na2CO3 (4 mL), and another portion
of PdCl2(dppf)-CH2Cl2 (82.0 mg, 0.2 mmol), then heated at reflux overnight.
After the reaction mixture was allowed to cool to rt, it was filtered through a
pad of Celite, washed with abundant CH2Cl2, and concentrated in vacuo. The
crude product was purified by silica gel column (90:5:5 EtOAc/CH2Cl2/MeOH)
to afford 22 (717.0 mg, 71%) as a pale brown solid, mp 183–184 °C (lit1 187–
189 °C). 1H NMR (DMSO-d6) d 10.3 (br s, 1H, NH), 9.57–9.56 (m, 1H, Ar-H), 9.47
(dd, J = 5.0, 1.0 Hz, 1H, Ar-H), 9.05 (d, J = 4.5 Hz, 1H, Ar-H), 8.42 (d, J = 4.5 Hz,
1H, Ar-H), 8.25 (d, J = 8.5 Hz, 1H, Ar-H), 8.12 (dd, J = 9.0, 2.0 Hz, 1H, Ar-H), 8.07
(dd, J = 5.0, 2.0 Hz, 1H, Ar-H), 7.94 (dd, J = 4.0, 2.0 Hz, 2H, Ar-H), 7.74–7.70 (m,
1H, Ar-H), 7.67 (d, J = 4.5 Hz, 1H, Ar-H), 7.58–7.53 (m, 1H, Ar-H), 7.19 (td,
J = 8.5, 2.5 Hz, 1H, Ar-H), 3.64 (s, 3H, OCH3). HRMS (ESI, m/z): calcd for
C
25H18N5O3F2S ([M+H]+) 506.1098, found 506.1101.
(q) 4-Fluoro-N-(2-methoxy-5-(4-(pyridazin-4-yl)quinolin-6-yl)pyridin-3-yl)-
2-nitrobenzenesulfonamide (2-nitro-GSK2126458, 23). suspension of
(285.0 mg, 1.0 mmol), bis(pinacolato)diboron (279.7 mg, 1.1 mmol),
A
7
(j) 5-Bromo-2-chloro-3-nitropyridine (15). A mixture of 5-bromo-2-hydroxy-
3-nitropyridine (14) (28.8 g, 131.5 mmol), POCl3 (288 mL) and anhydrous DMF
(30 mL) was stirred and heated at reflux for 3 h. After removal of POCl3, the
residue was poured into ice water and extracted with EtOAc. The combined
organic layers were washed with saturated aqueous NaHCO3, brine, dried over
anhydrous Na2SO4, filtered and concentrated in vacuo. The crude product was
purified by silica gel column chromatography (4:1 hexanes/EtOAc) to afford 15
(25.4 g, 81%) as a pale yellow solid, mp 66–67 °C (lit32 68 °C). 1H NMR (CDCl3) d
8.70 (d, J = 2.0 Hz, 1H, Ar-H), 8.37 (d, J = 2.5 Hz, 1H, Ar-H).
PdCl2(dppf)-CH2Cl2 (41.0 mg, 0.05 mmol) and KOAc (196.3 mg, 2.0 mmol) in
anhydrous 1,4-dioxane (10 mL) was stirred and heated at reflux for 3 h. The
reaction mixture was treated with 20 (426.5 mg, 1.05 mmol), 2 M Na2CO3
(2 mL), and another portion of PdCl2(dppf)-CH2Cl2 (41.0 mg, 0.1 mmol), then
heated at reflux overnight. After the reaction mixture was allowed to cool to rt,
it was filtered through a pad of Celite, washed with abundant CH2Cl2, and
concentrated in vacuo. The crude product was purified by preparative TLC plate
(90:5:5 EtOAc/CH2Cl2/MeOH) to afford 23 (110.0 mg, 21%) as a pale yellow
solid, mp 119–120 °C. 1H NMR (DMSO-d6) d 10.4 (br s, 1H, NH), 9.57–9.56 (m,

1574
M. Wang et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1569–1574
1H, Ar-H), 9.46 (dd, J = 5.0, 1.0 Hz, 1H, Ar-H), 9.06 (d, J = 4.5 Hz, 1H, Ar-H), 8.44
reaction vial were diluted with 0.1 M NaHCO3 (1 mL), and injected onto the
semi-preparative HPLC column with 3 mL injection loop for purification. The
product fraction was collected in a recovery vial containing 30 mL water. The
diluted tracer solution was then passed through a C-18 Sep-Pak Plus cartridge,
and washed with water (5 mL ꢂ 4). The cartridge was eluted with EtOH
(1 mL ꢂ 2) to release [11C]GSK2126458 ([11C]22). The eluted product was then
(d, J = 1.5 Hz, 1H, Ar-H), 8.26 (d, J = 8.5 Hz, 1H, Ar-H), 8.12 (dd, J = 9.0, 2.0 Hz,
1H, Ar-H), 8.07–8.04 (m, 3H, Ar-H), 7.96 (d, J = 1.5 Hz, 1H, Ar-H), 7.91 (d,
J = 2.5 Hz, 1H, Ar-H), 7.77–7.73 (m, 1H, Ar-H), 7.68 (d, J = 4.0 Hz, 1H, Ar-H), 3.65
(s, 3H, OCH3). HRMS (ESI, m/z): calcd for C25H18N6O5FS ([M+H]+) 533.1043,
found 533.1019.
(r) 2-Fluoro-N-(2-methoxy-5-(4-(pyridazin-4-yl)quinolin-6-yl)pyridin-3-yl)-
4-nitrobenzenesulfonamide (4-nitro-GSK2126458, 24). A similar procedure
for 23 was used to prepare 24 from 7 and 21 in 24% yield as a yellow solid, mp
124–125 °C. 1H NMR (DMSO-d6) d 10.7 (br s, 1H, NH), 9.57 (q, 1H, J = 1.0 Hz, Ar-
H), 9.47 (dd, J = 5.0, 1.0 Hz, 1H, Ar-H), 9.06 (d, J = 4.5 Hz, 1H, Ar-H), 8.43 (d,
J = 2.0 Hz, 1H, Ar-H), 8.38 (dd, J = 10.0, 2.0 Hz, 1H, Ar-H), 8.26 (d, J = 9.0 Hz, 1H,
Ar-H), 8.16–8.14 (m, 2H, Ar-H), 8.07 (dd, J = 5.0, 2.0 Hz, 1H, Ar-H), 7.99 (d,
J = 2.0 Hz, 1H, Ar-H), 7.96–7.93 (m, 2H, Ar-H), 7.68 (d, J = 4.5 Hz, 1H, Ar-H), 3.60
(s, 3H, OCH3). HRMS (ESI, m/z): Calcd for C25H18N6O5FS ([M+H]+) 533.1043,
found 533.1028.
sterile-filtered through a Millex-FG 0.2 lm membrane into a sterile vial and
formulated with 10 mL saline. Total radioactivity was assayed and total
volume was noted for tracer dose dispensing. Retention times in the semi-
preparative HPLC system were: tR 25 = 4.07 min, tR 22 = 10.81 min, tR
[
11C]22 = 10.81 min. Retention times in the analytical HPLC system were: tR
25 = 2.51 min, tR 22 = 5.12 min, tR
radiochemical yields of [11C]22 from [11C]CO2 were 40–50%.
(u)
2-[18F]Fluoro-4-fluoro-N-(2-methoxy-5-(4-(pyridazin-4-yl)quinolin-
[
11C]22 = 5.12 min. The decay corrected
6-yl)pyridin-3-yl)benzenesulfonamide (2-[18F]GSK2126458, 2-[18F]22) and
2-fluoro-4-[18F]fluoro-N-(2-methoxy-5-(4-(pyridazin-4-yl)quinolin-6-yl)
pyridin-3-yl)benzenesulfonamide (4-[18F]GSK2126458, 4-[18F]22). No-carrier-
added (NCA) aqueous H[18F]F was produced by 18O(p,n)18F nuclear reaction
using a Siemens Eclipse RDS-111 cyclotron by irradiation of H218O (2.5 mL).
H[18F]F (7.4–18.5 GBq) in [18O]water plus 0.1 mL K2CO3 solution (1.7 mg) and
Kryptofix 2.2.2 (10 mg) in 1.0 mL CH3CN with additional 1 mL CH3CN were
placed in the fluorination reaction vial (10-mL V-vial) and repeated azeotropic
distillation (17 min) was performed at 110 °C to remove water and to form the
anhydrous K[18F]F-Kryptofix 2.2.2 complex. The precursor 2-nitro-
GSK2126458 (23) or 4-nitro-GSK2126458 (24) (1 mg) dissolved in DMSO
(1.0 mL) was introduced to the reaction vessel and heated at 140 °C for 15 min
to affect radiofluorination. After cooling to ꢁ90 °C, the contents of the reaction
vial were diluted with 0.1 M NaHCO3 (1 mL), and injected onto the semi-
preparative HPLC column with 3 mL injection loop for purification. The product
fraction was collected in a recovery vial containing 30 mL water. The diluted
tracer solution was then passed through a C-18 Sep-Pak Plus cartridge, and
washed with water (5 mL ꢂ 4). The cartridge was eluted with EtOH (1 mL ꢂ 2)
to release 2-[18F]GSK2126458 (2-[18F]22) or 4-[18F]GSK2126458 (4-[18F]22).
(s) 2,4-Difluoro-N-(2-hydroxy-5-(4-(pyridazin-4-yl)quinolin-6-yl)pyridin-3-
yl)benzenesulfonamide (desmethyl-GSK2126458, 25). To
GSK2126458 22 (283.0 mg, 0.56 mmol) in anhydrous CH3CN (5 mL) was
added iodotrimethylsilane (152 L, 1.12 mmol). After the reaction mixture was
a solution of
l
stirred and heated at reflux for 4 h, it was concentrated in vacuo and the
residue was purified by preparative TLC plate (1:9 CH2Cl2/MeOH) to afford 25
(135.0 mg, 49%) as a yellow solid: mp 179 °C (dec.). 1H NMR (DMSO-d6) d 12.3
(br s, 1H, OH), 9.87 (br s, 1H, NH), 9.56 (s, 1H, Ar-H), 9.48 (d, J = 5.0 Hz, 1H, Ar-
H), 9.02 (d, J = 4.0 Hz, 1H, Ar-H), 8.20 (d, J = 9.0 Hz, 1H, Ar-H), 8.06 (dd, J = 5.0,
2.0 Hz, 1H, Ar-H), 8.03 (dd, J = 8.5, 1.5 Hz, 1H, Ar-H), 7.85 (q, J = 1.0 Hz, 1H, Ar-
H), 7.78 (d, J = 1.5 Hz, 1H, Ar-H), 7.75 (d, J = 2.0 Hz, 1H, Ar-H), 7.71 (s, 1H, Ar-H),
7.65 (d, J = 4.5 Hz, 1H, Ar-H), 7.50 (td, J = 10.0, 2.0 Hz, 1H, Ar-H), 7.21 (td, J = 8.5,
2.0 Hz, 1H, Ar-H). HRMS (ESI, m/z): calcd for C24H16N5O3F2S ([M+H]+) 492.0942,
found 492.0928.
(t)
2,4-Difluoro-N-(2-[11C]methoxy-5-(4-(pyridazin-4-yl)quinolin-6-
11C]22). 11C]CO2
yl)pyridin-3-yl)benzenesulfonamide ([11C]GSK2126458,
[
[
was produced by the 14N(p, 11C nuclear reaction in the small volume
a)
(9.5 cm3) aluminum gas target provided with the Siemens RDS-111 Eclipse
cyclotron. The target gas consisted of 1% oxygen in nitrogen purchased as a
specialty gas from Praxair, Indianapolis, IN. Typical irradiations used for the
The eluted product was then sterile-filtered through
a Millex-FG 0.2 lm
membrane into sterile vial and formulated with 10 mL saline. Total
a
radioactivity was assayed and total volume was noted for tracer dose
dispensing. Retention times in the semi-preparative HPLC system were: tR
23 = 6.50 min, tR 24 = 5.87 min, tR 22 = 10.81 min, tR 2-[18F]22 = 10.81 min, tR
4-[18F]22 = 10.81 min. Retention times in the analytical HPLC system were: tR
23 = 2.62 min, tR 24 = 2.57 min, tR 22 = 5.12 min, tR 2-[18F]22 = 5.12 min, tR 4-
development were 50 lA beam current and 15 min on target. The production
run produced approximately 25.9 GBq of [11C]CO2 at EOB. In a small reaction
vial (5 mL), the desmethyl-GSK2126458 precursor 25 (0.5–1.0 mg) was
dissolved in CH3CN (400
carrier-added (high specific activity) [11C]CH3OTf that was produced by the
gas-phase production method18 from 11C]CO2 through 11C]CH4 and
11C]CH3Br with silver triflate (AgOTf) column was passed into the reaction
lL). To this solution was added 2 N NaOH (2 lL). No
[
18F]22 = 5.12 min. The decay corrected radiochemical yields of 2-[18F]22 and
[
[
4-[18F]22 from H[18F]F were 20–30%.
[
30. Huang, X.; Liu, Z. J. Org. Chem. 2002, 67, 6731.
31. Lin, A.; Loo, T. J. Med. Chem. 1978, 21, 268.
32. Jouve, K.; Bergman, J. J. Heterocycl. Chem. 2003, 40, 261.
vial at rt, until radioactivity reached a maximum (ꢁ2 min), and then the
reaction vial was isolated and heated at 80 °C for 3 min. The contents of the