Synthesis and initial in vivo studies with [11C]SB-216763: The first radiolabeled brain penetrative inhibitor of GSK-3

Article abstract of DOI:10.1021/acsmedchemlett.5b00044

Quantifying glycogen synthase kinase-3 (GSK-3) activity in vivo using positron emission tomography (PET) imaging is of interest because dysregulation of GSK-3 is implicated in numerous diseases and neurological disorders for which GSK-3 inhibitors are bei

Full text of DOI:10.1021/acsmedchemlett.5b00044

Letter
pubs.acs.org/acsmedchemlett
Synthesis and Initial in Vivo Studies with [11C]SB-216763: The First
Radiolabeled Brain Penetrative Inhibitor of GSK‑3
Lei Li,† Xia Shao,‡ Erin L. Cole,‡ Stephan A. Ohnmacht,† Valentina Ferrari,† Young T. Hong,§
David J. Williamson,§ Tim D. Fryer,§ Carole A. Quesada,‡ Phillip Sherman,‡ Patrick J. Riss,†
Peter J. H. Scott,*,‡,∥ and Franklin I. Aigbirhio*,†,§
†Molecular Imaging Chemistry Laboratory, Wolfson Brain Imaging Centre, University of Cambridge, Cambridge CB2 1TN, U.K.
‡Division of Nuclear Medicine, Department of Radiology, The University of Michigan Medical School, Ann Arbor, Michigan 48109,
United States
§Laboratory for Molecular Imaging, Wolfson Brain Imaging Centre, University of Cambridge, Cambridge CB2 1TN, U.K.
∥The Interdepartmental Program in Medicinal Chemistry, The University of Michigan, Ann Arbor, Michigan 48109, United States
S
* Supporting Information
ABSTRACT: Quantifying glycogen synthase kinase-3 (GSK-
3) activity in vivo using positron emission tomography (PET)
imaging is of interest because dysregulation of GSK-3 is
implicated in numerous diseases and neurological disorders for
which GSK-3 inhibitors are being considered as therapeutic
strategies. Previous PET radiotracers for GSK-3 have been
reported, but none of the published examples cross the blood−
brain barrier. Therefore, we have an ongoing interest in
developing a brain penetrating radiotracer for GSK-3. To this
end, we were interested in synthesis and preclinical evaluation of [11C]SB-216763, a high-affinity inhibitor of GSK-3 (Ki = 9 nM;
IC50 = 34 nM). Initial radiosyntheses of [11C]SB-216763 proved ineffective in our hands because of competing [3 + 3]
sigmatropic shifts. Therefore, we have developed a novel one-pot two-step synthesis of [11C]SB-216763 from a 2,4-
dimethoxybenzyl-protected maleimide precursor, which provided high specific activity [11C]SB-216763 in 1% noncorrected
radiochemical yield (based upon [11C]CH3I) and 97−100% radiochemical purity (n = 7). Initial preclinical evaluation in rodent
and nonhuman primate PET imaging studies revealed high initial brain uptake (peak rodent SUV = 2.5 @ 3 min postinjection;
peak nonhuman primate SUV = 1.9 @ 5 min postinjection) followed by washout. Brain uptake was highest in thalamus, striatum,
cortex, and cerebellum, areas known to be rich in GSK-3. These results make the arylindolemaleimide skeleton our lead scaffold
for developing a PET radiotracer for quantification of GSK-3 density in vivo and ultimately translating it into clinical use.
KEYWORDS: Positron emission tomography, glycogen synthase kinase (GSK-3), carbon-11, neuroimaging, brain PET
lycogen synthase kinase-3 (GSK-3), which consists of two
engagement by new GSK-3 inhibitors in therapeutic discovery
Gisoforms (GSK3α and GSK3β), has been identified as an pipelines and, ultimately, aid in developing dosage regimens.
Positron emission tomography (PET) is a widely used in vivo
imaging technique, which can be applied to quantify enzyme
levels and activity in the living brain. The former can be
accomplished using a radiolabeled inhibitor of the enzyme,
while the latter would require a radiolabeled substrate.
However, to date there are no known small molecule substrates
for GSK-3. Hence, we are interested in development of a PET
radiotracer based upon a GSK-3 inhibitor that will allow us to
noninvasively quantify the density of GSK-3 in the living animal
brain. A number of GSK-3 inhibitors from various chemical
classes have been identified, show varying potencies and
isoform selectivity, as reported in recent literature.2,8,14−22
However, previous reports of PET radiotracers for GSK-3 are
enzyme involved in glycogen metabolism, with the highest
abundance in the brain.1 Recent research has shown that GSK-
3β acts as a key component of many cellular and physiological
events including cellular growth, apoptosis, and neuropatho-
logical events,2,3 but its dysregulation is known to play a role in
multiple diseases such as diabetes,4 neurodegenerative con-
ditions such as Alzheimer’s disease,5,6 Parkinson disease,7
neurological disorders,8 pain,9,10 and cancer.11−13 In light of
this, GSK-3 inhibitors are being developed as therapeutics
across this disease spectrum.2,8,14−22
Reflecting its role in the pathogenesis of this disease
spectrum, there is an important need for tools that enable
quantification of GSK-3 in vivo. From a diagnostic perspective,
early detection of the dysregulation of GSK-3 could prove to be
a prodromal biomarker for these conditions enabling early
diagnosis and treatment initiation, while the ability to quantify
GSK-3/ligand interactions can be used to understand target
Received: January 28, 2015
Accepted: March 10, 2015
© XXXX American Chemical Society
A
DOI: 10.1021/acsmedchemlett.5b00044
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limited (Figure 1).23−27 The first example was [11C]N-(4-
methoxybenzyl)-N′-(5-nitro-1,3-thiazole-2-yl)urea ([11C]AR-
a
Scheme 1. Initial Attempts to Synthesize [11C]SB-216763
a
Figure 1. PET Radiotracers for GSK-3.
Reagents and conditions: (i) 5 N KOH, EtOH, reflux, overnight
(21%); (ii)[11C]CH3I, NaH, 45 °C, 4 min; (iii) LiHMDS, MeOH/
DMF, 80 °C, 8 min.
A014418), reported by Vasdev and colleagues.24,25 Although
the central nervous system (CNS) effect following peripheral
administration of AR-A014418 into rats has been demon-
strated,28 the radioactive [11C]AR-A014418 showed no brain
penetration, according to ex vivo biodistribution studies
conducted by the Vasdev group. We recently reported the
synthesis and evaluation of [11C]PyrATP-1,26 a GSK-3β
inhibitor based upon the pyrazine scaffold initially reported
by Berg and colleagues21 and demonstrated specific binding in
rat brain ex vivo autoradiography experiments. However,
despite predictions of good CNS permeability from bovine
endothelial cell assays,21 [11C]PyrATP-1 also did not enter the
brain and is unsuitable for neuroimaging purposes.
In our continuing efforts to develop a brain penetrative GSK-
3 radiotracer, we next turned our attention to (3-(2,4-
dichlorophenyl)-4-(1-[11C]methyl-1H-indol-3-yl)-1H-pyrrole-
2,5-dione) (SB-216763), an arylindolemaleimide with high
affinity for both isoforms of GSK-3 (Ki = 9 nM; IC50 = 34 nM)
that was originally reported by Coghlan and colleagues.29 SB-
216763 inhibits the enzyme in an ATP competitive manner, has
good selectivity for GSK-3 over other protein kinases, and has
been widely used in preclinical studies with promising results to
date.30 While a synthesis of [11C]SB-216763 was reported in
2011 by Zheng and co-workers,27 no biological evaluation of
the radiotracer has been reported to date. The goal of the
current work was therefore evaluation of the imaging properties
of [11C]SB-216763, including brain uptake and washout, in
rodents and nonhuman primates.
We therefore abandoned this strategy and, in need of a new
synthesis of [11C]SB-216763, investigated the possibility of
direct carbon-11 methylation of a maleimide precursor.
It has been reported that the lower pKa of the maleimide NH
(∼10) when compared to that of the indole NH (∼21) would
favor 11C-methylation at the maleimide if unprotected
precursor 1 was radiolabeled directly.27 However, given (1)
that the indole NH is expected to be more nucleophilic than
the maleimide NH because of the adjacent carbonyl groups on
maleimide, (2) that treatment with methyl iodide is a classical
strategy for methylating indoles,31,32 and (3) that a mixture of
the two possible labeled products should be separable by
semipreparative HPLC, we decided to test direct labeling of
precursor 1 with [11C]CH3I (Scheme 1). Unfortunately, one
radiolabeled product that did not correspond to [11C]SB-
216763 was obtained, as well as varying amounts of a
nonradioactive side product. These labeling results were
disappointing and somewhat perplexing. While the radioactive
species was likely the labeled maleimide, the identity of the
other nonradioactive side product was not clear. To understand
the cause of these problems further, we exposed precursor 1
and unlabeled 3 to reaction conditions (NaH/[12C]CH3I) on a
larger scale so that stability of both the precursor and product
to the reaction conditions, as well as the identity of any side
products, could be examined using traditional analytical
methods (NMR, mass spectrometry). To our surprise, in
each case we obtained significant quantities of cyclized side
products 6 or 7 (Scheme 2), which we expect arise from [3 +
3] sigmatropic shifts, followed by elimination of HCl to
rearomatize.
To explore these pericyclic side reactions further, we
performed an array of reactions subjecting both precursor 1
and reference standard 3 to a range of conditions (acid, base,
heat) to understand the origins of the side products during
labeling. Interestingly, cyclized products 6 and 7 were found in
every case, albeit in varying amounts up to 32%. As a side note,
we also subjected the monochloro unmethylated 4 and
methylated 5 analogues to the matrix of conditions to identify
whether the monochloro analogue of SB-216763 offered an
alternate scaffold for radiolabeling that could circumvent this
issue. Surprisingly, this did not improve the situation, and
Work commenced with attempts to synthesize [11C]SB-
216763 using Zheng’s procedure by 11C-methylation of the
maleic anhydride precursor 2.27 The synthesis of precursor 2
proceeded in 21% yield from 1 when synthesized on gram scale,
although no product was obtained when conducted on a
smaller scale. Unlabeled SB-216763 reference standard
(necessary for confirming identity of the carbon-11 labeled
radiotracer) could either be prepared using literature
procedures27 or purchased directly (Sigma-Aldrich). Precursor
2 was subjected to Zheng’s procedure (Scheme 1): treatment
with [11C]CH3I and subsequent conversion to the maleimide
by treatment with lithium hexamethyldisilazane (LiHMDS).
Regrettably, this approach yielded no product in either of our
laboratories, but rather complex mixtures of unidentified
byproducts (see Supporting Information (SI) for more details).
B
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b
Scheme 2. [3 + 3] Sigmatropic Shifts
Scheme 3. Radiosynthesis of [11C]SB-216763 (3) from
c
Precursor 8
b
Reagents and conditions: acid, base, or heat (see SI).
analogues 4 and 5 also underwent the [3 + 3] sigmatropic shift
followed by elimination of H2 to provide comparable yields of 6
and 7, respectively (see SI for full details of these experiments).
While pericyclic reactions of this kind are known in the
literature for related compounds, they are normally mediated
by light (in the presence of 2,3-dichloro-5,6-dicyano-1,4-
benzoquinone, DDQ, or iodine), or achieved using Pd-
catalyzed Heck reactions.33−36 Such reactions being a
complicating factor in this case were therefore surprising but
nevertheless, it was clear that they were negatively impacting
radiolabeling through consumption of precursor and/or base (6
was confirmed in the crude radiochemical reaction mixture (see
SI), but we never observed formation of [11C]7). We reasoned
that inhibiting the [3 + 3] sigmatropic shift would improve 11C-
methylation. As the competing pericyclic side reaction is
governed by strict orbital symmetry requirements, we
postulated that it could be eliminated by disrupting the
conformation of the molecule. Our strategy for this was to
install a bulky protecting group on the maleimide nitrogen that
could be easily removed under acidic conditions after
radiolabeling. We initially considered installing a Boc group,
but all of our attempts to Boc-protect 1 resulted in complex
mixtures of products (cyclized products were again apparent in
NMR spectra of crude reaction mixtures). We next considered
using the bulky 2,4-dimethyl benzyl group as it can also be
easily removed under acidic conditions, and the precursor (8)
could be accessed from maleic anhydride 2, rather than
maleimide 1. Thus, refluxing maleic anhydride 2 with 2,4-
dimethoxyl benzyl amine in acetic acid for 2 h yielded
desmethyl precursor 8 in 79% yield, ready for radiolabeling
(Scheme 3).
c
Reagents and conditions: (i) 2,4-dimethoxybenzyl amine, AcOH, 140
°C, 2 h (79%); (ii) [11C]CH3I, NaH, 45 °C, 4 min; (iii) 50%
CH3SO3H in THF, 110 °C, 6 min (1% RCY).
cartridge to provide doses suitable for preclinical evaluation
(radiochemical yields = 5−5.5 mCi, 185−200 MBq (1%
noncorrected radiochemical yield (RCY) at end-of-synthesis
(EOS) based upon [11C]CH3I), radiochemical purity = 97−
100%, specific activity = 4−9 Ci/μmol, 150−350 GBq/μmol at
EOS, n = 7).
Having successfully overcome our synthesis issues, we turned
our attention to examining the imaging properties of [11C]SB-
216763 in rodents (Figure 2A) and nonhuman primates
(Figure 2B) using preclinical PET imaging. Following imaging,
regions-of-interest (ROIs) were drawn over brain regions on
multiple planes of summed images. The volumetric ROIs were
then applied to the full dynamic data set to generate time−
radioactivity curves. Both rodent (Figure 2C) and nonhuman
primate (Figure 2D) curves confirmed good initial brain uptake
With the requisite precursor and reference standard in hand,
we revisited the radiosynthesis of [11C]SB-216763 (Scheme 3).
[11C]CO2 (∼3 Ci, 111 GBq) was produced in a cyclotron via
the 14N(p,α)11C nuclear reaction and delivered to the synthesis
module where it was first reduced to [11C]CH4 by treating with
hydrogen over a nickel catalyst at 350 °C. [11C]CH4 was then
reacted with iodine at 750 °C to yield [11C]CH3I (∼750 mCi,
27.8 GBq, ∼26% yield from [11C]CO2). Gratifyingly, treatment
of precursor 8 with NaH and subsequent methylation with
[11C]CH3I provided protected [11C]SB-216763. Subsequent
deprotection proceeded smoothly upon treatment with 50%
methanesulfonic acid solution in THF at 110 °C (Note:
cleavage of the 2,4-dimethoxylbenzyl group could not be
accomplished using Ceric ammonium nitrate due to its poor
solubility in THF). Purification of the crude reaction mixture by
semipreparative HPLC yielded [11C]SB-216763, which was
reconstituted into ethanolic saline using an Oasis HLB sep-pak
Figure 2. [11C]SB-216763 data: (A) rodent SUV map (60−120 min
p.i.); (B) summed primate PET image (0−90 min p.i..); (C) rodent
time−radioactivity curves; (D) primate time−radioactivity curves.
C
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(peak rodent standardized uptake value (SUV) = 2.5 @ 3 min
postinjection (p.i.); peak primate SUV = 1.9 @ 5 min p.i. (n =
2)), followed by washout. Washout was faster in rodents than
primates, and we attribute this to increased tissue volumes and
lower blood flow in the primates. Brain uptake was fairly
uniform throughout all brain regions in both species, which
would be consistent with the ubiquitous distribution of GSK-
3,37 although the highest uptake was observed in thalamus,
striatum, cortex, and cerebellum, areas of the mammalian brain
known to be rich in GSK-3β.37
In summary, we have developed a novel one-pot two-step
method for the radiosynthesis of [11C]SB-216763 and
demonstrated that it is the first radiotracer for GSK-3 able to
cross the BBB and enter the CNS in both rodents and
nonhuman primates. The arylindolemaleimide skeleton repre-
sents our lead scaffold for developing a radiotracer for
quantification of GSK-3 in vivo using brain PET and ultimately
translating it into clinical use.
injection; NMR, nuclear magnetic resonance; RCY, radio-
chemical yield; ROI, regions-of-interest; SUV, standardized
uptake value; TLC, thin layer chromatography
REFERENCES
■
(1) Woodget, J. R. Molecular cloning and expression of glycogen
synthase kinase-3/factor A. EMBO J. 1990, 9, 2431−2438.
(2) Cohen, P.; Goedert, M. GSK3 inhibitors: development and
therapeutic potential. Nat. Rev. Drug Discovery 2004, 3, 479−487.
(3) Jope, R. S.; Johnson, G. V. W. The glamour and gloom of
glycogen synthase kinase-3. Trends Biochem. Sci. 2004, 29, 95−102.
(4) Eldar-Finkelman, H.; Schreyer, S. A.; Shinohara, M. M.; LeBoeuf,
R. C.; Krebs, E. G. Increased glycogen synthase kinase-3 activity in
diabetes- and obesity-prone C57BL/6 J mice. Diabetes 1999, 48,
1662−1666.
(5) Hooper, C.; Killick, R.; Lovestone, S. The GSK3 hypothesis of
Alzheimer’s disease. J. Neurochem. 2008, 104, 1433−1439.
́
(6) Avila, J.; Wandosell, F.; Hernandez, F. Role of glycogen synthase
kinase-3 in Alzheimer’s disease pathogenesis and glycogen synthase
kinase-3 inhibitors. Expert Rev. Neurother. 2010, 10, 703−710.
ASSOCIATED CONTENT
* Supporting Information
■
́
(7) Morales-García, J. A.; Susín, C.; Alonso-Gil, S.; Perez, D. I.;
S
Palomo, V.; Per
́
ez, C.; Conde, S.; Santos, A.; Gil, C.; Martínez, A.;
Full experimental details and copies of NMR spectra and/or
HPLC chromatograms for all compounds synthesized; HRMS
for standards and precursors; procedures for radiochemical
syntheses and quality control; protocols for animal PET
imaging studies. This material is available free of charge via
the Internet at http://pubs.acs.org.
Perez-Castillo, A. Glycogen synthase kinase-3 inhibitors as potent
́
therapeutic agents for the treatment of Parkinson disease. ACS Chem.
Neurosci. 2013, 4, 350−360.
(8) Jope, R. S.; Roh, M.-S. Glycogen synthase kinase-3 (GSK3) in
psychiatric diseases and therapeutic interventions. Curr. Drug Targets
2006, 7, 1421−1434.
(9) Martins, D. F.; Rosa, A. O.; Gadotti, V. M.; Mazzardo-Martins, L.;
́
Nascimento, F. P.; Egea, J.; Lopez, M. G.; Santos, A. R. The
AUTHOR INFORMATION
Corresponding Authors
*E-mail: fia20@medschl.cam.ac.uk. Tel: +44(1223)331823.
Fax: +44(1223)331826.
*E-mail: pjhscott@umich.edu. Tel: +1 (734) 615-1756. Fax: +1
(734) 615-2557.
Author Contributions
The manuscript was written through contributions of all
authors and all authors have given approval to the final version
of the manuscript.
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antinociceptive effects of AR-A014418, a selective inhibitor of
glycogen synthase kinase-3 beta, in mice. J. Pain 2011, 12, 315−322.
(10) Maixner, D. W.; Weng, H.-R. The role of glycogen synthase
kinase 3 beta in neuroinflammation and pain. J. Pharm. Pharmacol.
2013, 1, 1−24.
(11) Luo, J. Glycogen synthase kinase 3β (GSK3β) in tumorigenesis
and cancer chemotherapy. Cancer Lett. 2009, 273, 194−200.
(12) Martinez, A.; Castro, A.; Dorronsoro, I.; Alonso, M. Glycogen
synthase kinase 3 (GSK-3) inhibitors as new promising drugs for
diabetes, neurodegeneration, cancer, and inflammation. Med. Res. Rev.
2002, 22, 373−384.
Funding
(13) Eldar-Finkelman, H. Glycogen synthase kinase 3: an emerging
therapeutic target. Trends Mol. Med. 2002, 8, 126−132.
This work was supported by a Medical Research Council (UK),
Collaborative Programme grant award RG46503 (FIA), and the
National Institute of Health Research (UK) Cambridge
Biomedical Research Unit in Dementia. Additional financial
support of this work from the National Institute of Biomedical
Imaging and Bioengineering, part of the National Institutes of
Health (under Award Number T32-EB005172) is also
gratefully acknowledged (PJHS). The content of this article is
solely the responsibility of the authors and does not necessarily
represent the official views of the National Institutes of Health.
(14) Rix, L. L. R.; Kuenzi, B. M.; Luo, Y.; Remily-Wood, E.; Kinose,
F.; Wright, G.; Li, J.; Koomen, J. M.; Haura, E. B.; Lawrence, H. R.;
Rix, U. GSK3 alpha and beta are new functionally relevant targets of
tivantinib in lung cancer cells. ACS Chem. Biol. 2014, 9, 353−358.
(15) Frame, S.; Cohen, P. GSK3 takes centre stage more than 20
years after its discovery. Biochem. J. 2001, 359, 1−16.
(16) Wagman, A. S.; Johnson, K. W.; Bussiere, D. E. Discovery and
development of GSK3 inhibitors for the treatment of type 2 diabetes.
Curr. Pharm. Des. 2004, 10, 1105−1137.
(17) O’Neill, D. J.; Shen, L.; Prouty, C.; Conway, B. R.; Westover, L.;
Xu, J. Z.; Zhang, H.-C.; Maryanoff, B. E.; Murray, W. V.; Demaresta,
K. T.; Kuo, G.-H. Design, synthesis, and biological evaluation of novel
7-azaindolyl-heteroaryl-maleimides as potent and selective glycogen
synthase kinase-3β (GSK-3β) inhibitors. Bioorg. Med. Chem. 2004, 12,
3167−3185.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
We thank Robert Smith (UK) and Paul Burke (UK) for
technical assistance.
■
(18) Smalley, T. L., Jr.; Peat, A. J.; Boucheron, J. A.; Dickerson, S.;
Garrido, D.; Preugschat, F.; Schweiker, S. L.; Thomson, S. A.; Wang,
T. Y. Synthesis and evaluation of novel heterocyclic inhibitors of GSK-
3. Bioorg. Med. Chem. Lett. 2006, 16, 2091−2094.
ABBREVIATIONS
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BBB, blood−brain barrier; Boc, tert-butyloxycarbonyl; CNS,
central nervous system; DDQ, 2,3-dichloro-5,6-dicyano-1,4-
benzoquinone; GSK-3, glycogen synthase kinase-3; HMDS,
hexamethyldisilazane; HPLC, high-performance liquid chroma-
tography; PET, positron emission tomography; p.i., post
(19) Monte, F. L.; Kramer, T.; Bolander, A.; Plotkin, B.; Eldar-
̈
Finkelman, H.; Fuertes, A.; Dominguez, J.; Schmidt, B. Synthesis and
biological evaluation of glycogen synthase kinase 3 (GSK-3) inhibitors:
an fast and atom efficient access to 1-aryl-3-benzylureas. Bioorg. Med.
Chem. Lett. 2011, 21, 5610−5615.
D
DOI: 10.1021/acsmedchemlett.5b00044
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
Letter
(20) Schmole, A.-C.; Brennfuhrer, A.; Karapetyan, G.; Jaster, R.;
(35) Sanchez-Martinez, C.; Shih, C.; Faul, M. M.; Zhu, G.; Paal, M.;
Somoza, C.; Li, T.; Kumrich, C. A.; Winneroski, L. L.; Xun, Z.; Brooks,
H. B.; Patel, B. K. R.; Schultz, R. M.; DeHahn, T. B.; Spencer, C. D.;
Watkins, S. A.; Considine, E.; Dempsey, J. A.; Ogg, C. A.; Campbell, R.
M.; Anderson, B. A.; Wagner, J. Aryl[α]pyrrolo [3,4-c] carbazoles as
selective cyclin D1-CDK4 inhibitors. Bioorg. Med. Chem. Lett. 2003, 13,
3835−3839.
̈
̈
Pews-Davtyan, A.; Hubner, R.; Ortinau, S.; Beller, M.; Rolfs, A.; Frech,
̈
M. J. Novel indolylmaleimide acts as GSK-3β inhibitor in human
neural progenitor cells. Bioorg. Med. Chem. 2010, 18, 6785−6795.
(21) Berg, S.; Bergh, M.; Hellberg, S.; Hogdin, K.; Lo-Alfredsson, Y.;
̈
Soderman, P.; von Berg, S.; Weigelt, T.; Ormo, M.; Xue, Y.; Tucker, J.;
̈
̈
Neelissen, J.; Jerning, E.; Nilsson, Y.; Bhat, R. Discovery of novel
potent and highly selective glycogen synthase kinase-3β (GSK3β)
inhibitors for Alzheimer’s disease: design, synthesis, and character-
ization of pyrazines. J. Med. Chem. 2012, 55, 9107−9119.
́
(36) Routier, S.; Merour, J.-Y.; Dias, N.; Lansiaux, A.; Bailly, C.;
Lozach, O.; Meijer, L. Synthesis and biological evaluation of novel
phenylcarbazoles as potential anticancer agents. J. Med. Chem. 2006,
49, 789−799.
(37) Leroy, K.; Brion, J.-P. Developmental expression and local-
ization of glycogen synthase kinase-3β in rat brain. J. Chem. Neuroanat.
1999, 16, 279−293.
(22) Georgievska, B.; Sandin, J.; Doherty, J.; Mortberg, A.; Neelissen,
̈
J.; Andersson, A.; Gruber, S.; Nilsson, Y.; Schott, P.; Arvidsson, P. I.;
̈
Hellberg, S.; Osswald, G.; Berg, S.; Falting, J.; Bhat, R. V. AZD1080, a
̈
novel GSK3 inhibitor, rescues synaptic plasticity deficits in rodent
brain and exhibits peripheral target engagement in humans. J.
Neurochem. 2013, 125, 446−456.
(23) Karimi, F.; Langstrom B. 11C/18F-labeled inhibitors of glycogen
synthase kinase-3. WO2007138408A1, 2007.
(24) Vasdev, N.; Garcia, A.; Stableford, W. T.; Young, A. B.; Meyer, J.
H.; Houle, S.; Wilson, A. A. Synthesis and ex vivo evaluation of
carbon-11 labelled N-(4-methoxybenzyl)-N′-(5-nitro-1,3-thiazol-2-yl)-
urea ([11C]AR-A014418): a radiolabelled glycogen synthase kinase-3β
specific inhibitor for PET studies. Bioorg. Med. Chem. Lett. 2005, 15,
5270−5273.
(25) Hicks, J. W.; Wilson, A. A.; Rubie, E. A.; Woodgett, J. R.; Houle,
S.; Vasdev, N. Towards the preparation of radiolabeled 1-aryl-3-benzyl
ureas: radiosynthesis of [11C-carbonyl] AR-A014418 by [11C]CO2
fixation. Bioorg. Med. Chem. Lett. 2012, 22, 2099−2101.
(26) Cole, E. L.; Shao, X.; Sherman, P.; Quesada, C.; Fawaz, M. V.;
Desmond, T. J.; Scott, P. J. H. Synthesis and evaluation of
[11C]PyrATP-1, a novel radiotracer for PET Imaging of glycogen
synthase kinsae-3β (GSK-3β). Nucl. Med. Biol. 2014, 41, 507−512.
(27) Wang, M.; Gao, M.; Miller, K. D.; Sledge, G. W.; Hutchins, G.
D.; Zheng, Q. H. The first synthesis of [11C]SB-216763, a new
potential PET agent for imaging of glycogen synthase kinase-3 (GSK-
3). Bioorg. Med. Chem. Lett. 2011, 21, 245−249.
(28) Gould, T. D.; Einat, H.; Bhat, R.; Manji, H. K. AR-A014418, a
selective GSK-3 inhibitor, produces antidepressant-like effects in the
forced swim test. Int. J. Neuropsychopharmacol. 2004, 7, 387−390.
(29) Coghlan, M. P.; Culbert, A. A.; Cross, D. A. E.; Corcoran, S. L.;
Yates, J. W.; Pearce, N. J.; Rausch, O. L.; Murphy, G. J.; Carter, P. S.;
Cox, L. R.; Mills, D.; Brown, M. J.; Haigh, D.; Ward, R. W.; Smith, D.
G.; Murray, K. J.; Reith, A. D.; Holder, J. C. Selective small molecule
inhibitors of glycogen synthase kinase-3 modulate glycogen metabo-
lism and gene transcription. Chem. Biol. 2000, 7, 793−803.
(30) For a recent review see: Eldar-Finkelman, H.; Martinez, A. GSK-
3 inhibitors: preclinical and clinical focus on CNS. Front. Mol. Neurosci.
2011, 4, article 32.
(31) Reineeke, M. G.; Sebastian, J. F.; Johnson, H. W., Jr.; Pyun, C.
Effect of solvent and cation on the reaction of organometallic
derivatives of indole with methyl iodide. J. Org. Chem. 1972, 37, 3066−
3068.
(32) Nunomoto, S.; Kawakami, Y.; Yamashita, Y.; Takeuchi, H.;
Eguchi, S. Regioselectivity control in alkylation reactions of indolyl
ambident anion. J. Chem. Soc., Perkin Trans. 1 1990, 111−114.
(33) Zhu, G.; Conner, S.; Zhou, X.; Shih, C.; Brooks, H. B.;
Considine, E.; Dempsey, J. A.; Ogg, C.; Patel, B.; Schultz, R. M.;
Spencer, C. D.; Teicher, B.; Watkins, S. A. Synthesis of quinolinyl/
isoquinolinyl[α]pyrrolo [3,4-c] carbazoles as cyclin D1/CDK4
inhibitors. Bioorg. Med. Chem. Lett. 2003, 13, 1231−1235.
(34) Engler, T.; Furness, K.; Malhotra, S.; Sanchez-Martinez, C.;
Shih, C.; Xie, W.; Zhu, G.; Zhou, X.; Conner, S.; Faul, M. M.; Sullivan,
K. A.; Kolis, S. P.; Broks, H. B.; Patel, B.; Schultz, R. M.; DeHahn, T.
B.; Kirmani, K.; Spencer, C. D.; Watkins, S. A.; Considine, E. L.;
Dempsey, J. A.; Ogg, C. A.; Stamm, N. B.; Anderson, B. D.; Campbell,
R. M.; Vasudevan, V.; Lytle, M. L. Novel, potent and selective cyclin
D1/CDK4 inhibitors: indolo[6,7-α]pyrrolo [3,4-c] carbazoles. Bioorg.
Med. Chem. Lett. 2003, 13, 2261−2267.
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DOI: 10.1021/acsmedchemlett.5b00044
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