Design and synthesis of (2R,3S)-iodoreboxetine analogues for SPECT imaging of the noradrenaline transporter

Article abstract of DOI:10.1016/j.bmcl.2009.07.064

A stereoselective 10-step synthesis of iodophenoxy analogues of (2R,3S)-reboxetine has been developed with the aim of generating a new SPECT imaging agent for the noradrenaline transporter (NAT). In vitro testing of these compounds against various mono-am

Full text of DOI:10.1016/j.bmcl.2009.07.064

Bioorganic & Medicinal Chemistry Letters 19 (2009) 4996–4998
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design and synthesis of (2R,3S)-iodoreboxetine analogues for SPECT imaging
of the noradrenaline transporter
Nicola K. Jobson a, Andrew R. Crawford b, Deborah Dewar b, Sally L. Pimlott c, Andrew Sutherland a,
*
a WestCHEM, Department of Chemistry, The Joseph Black Building, University of Glasgow, Glasgow G12 8QQ, UK
b Division of Clinical Neuroscience, Faculty of Medicine, University of Glasgow, Glasgow G12 8QQ, UK
c West of Scotland Radionuclide Dispensary, University of Glasgow and North Glasgow University Hospital NHS Trust, Glasgow G11 6NT, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
A stereoselective 10-step synthesis of iodophenoxy analogues of (2R,3S)-reboxetine has been developed
with the aim of generating a new SPECT imaging agent for the noradrenaline transporter (NAT). In vitro
testing of these compounds against various mono-amine transporters showed an ortho-iodophenoxy
analogue to have excellent affinity (Ki 8.4 nM) and good selectivity for NAT.
Received 11 June 2009
Revised 9 July 2009
Accepted 10 July 2009
Available online 16 July 2009
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Reboxetine
Stereoselective synthesis
SPECT
Imaging
Noradrenaline transporter
The noradrenaline transporter(NAT)isa membranespanningpro-
tein situated on the pre-synaptic terminal of noradrenergic neurons.
Its main function is to regulate the concentration of noradrenaline
in the synaptic cleft via a re-uptake mechanism.1,2 Recent research
has shown that a reduction in the synaptic levels of noradrenaline is
associated with various neuropsychiatric and neurodegenerative dis-
orders such as attention-deficit/hyperactivity disorder, anxiety, Alz-
heimer’s disease and depression and thus, NAT is a major target for
the development of drugs for these conditions.3–6
the synthesis and in vitro testing of four stereoisomers of an
iodophenylreboxetine analogue.10,11 The results of the biological
testing showed that while the (2S,3S)-iodophenylreboxetine ana-
logue 3 had as expected, significant affinity for NAT (Ki 53.8 nM),
the (2R,3S)-analogue 4 was of similar potency (Ki 58.2 nM). The
outcome of this work revealed that (2R,3S)-analogues of reboxetine
should also be considered when designing high affinity molecular
probes for NAT.
Non-invasive imaging techniques such as positron emission
tomography (PET) and single photon emission computed tomogra-
phy (SPECT) have been proposed as important tools for determin-
ing the link between NAT and these neuropsychiatric and
neurodegenerative disorders. These techniques could also be used
for the assessment of drug occupancy of the transporter in vivo. To
utilise PET and SPECT in this way requires the development of
highly selective radiolabelled molecular probes. Recent work has
focused on the development of probes based on (2S,3S)-reboxetine
1, the well-characterised NAT inhibitor.7 Development of small
libraries of iodinated analogues of reboxetine, by the Saji and
Tamagnan groups identified analogue 2 (Ki 0.84–4.22 nM) as hav-
ing significant potential as an imaging agent of NAT.8,9 With the
aim of understanding the stereochemical pharmacology of NAT
for the development of novel SPECT tracers, we recently reported
O
O
O
O
OEt
I
N
H
N
H
2
1
I
I
O
O
O
O
OEt
OEt
N
H
N
H
3
4
* Corresponding author. Tel.: +33 141 330 5936; fax: +33 141 330 4888.
E-mail address: andrews@chem.gla.ac.uk (A. Sutherland).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.07.064

N. K. Jobson et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4996–4998
4997
OH
In an effort to identify new high affinity SPECT tracers for the imag-
ing of NAT, we were interested in preparing the (2R,3S)-stereoiso-
mer 5 of the iodophenoxyreboxetine analogue 2 reported by the
Saji and Tamagnan groups.8,9 Our aim was to develop a flexible syn-
thetic route to 5 that would also permit the preparation of meta-
and para-iodophenoxy isomers 6 and 7, thus allowing us to probe
the structure–activity-relationship of the iodophenoxy moiety.
Herein, we now report a 10-step stereoselective synthesis of 5, 6
and 7. In vitro testing of all three compounds for affinity with
NAT, the serotonin transporter (SERT) and the dopamine trans-
porter (DAT) is also described.
a
Cl
Cl
OH
8
9
b
c
OH
OH
N3
O
OH
11
10
d
OH
O
OH
O
e
NH
I
NH2
N
OH
H
O
5
OH
Cl
13
12
I
f
O
O
I
O
O
N
H
N
H
g, h
6
7
O
O
N
HO
HO
N
H
O
Boc
In our previous synthesis of a (2R,3S)-reboxetine analogue, a Sharp-
less asymmetric epoxidation12 was used to establish the key stere-
ogenic centres followed by the preparation of an amino alcohol
which was eventually used to form the morpholine ring.10 While
this approach did produce the target compound, introduction of
diversity occurred early in the route, effectively restricting the
number of analogues which could be produced. In this current
study, we have developed an alternative, more flexible approach
to (2R,3S)-reboxetine analogues using a Sharpless asymmetric
dihydroxylation13 to create the key stereogenic centres followed
by subsequent formation of the morpholine ring. The final stage
uses an activated mesylate intermediate for the concurrent intro-
duction of the iodophenoxy moieties and inversion of the stereo-
chemical configuration at the C-3 position to give the desired
(2R,3S)-analogues. Thus, cinnamyl chloride 8 was subjected to a
Sharpless asymmetric dihydroxylation using AD-mix-b to give 9
in 66% yield and in an excellent 99% ee (Scheme 1).14 Diol 9 was
then treated with sodium hydroxide, forming the epoxide 10 in
98% yield. During our previous synthesis of (2S,3S)- and (2R,3R)-
iodophenylreboxetine analogues, it was shown that 2,3-epoxy-1-
(4-iodophenyl)propan-1-ols could react with ammonia in a highly
regioselective manner, resulting in attack at the least hindered
end of the epoxide to give the corresponding amino alcohol.11 How-
ever, a similar reaction with epoxide 10 gave both amino alcohol
regioisomers as observed by 1H NMR spectroscopy and while this
mixture could undergo a transformation with chloroacetyl chloride
to give desired intermediate 13, the isolated yield of 13 was only
25% over the two steps. Srinivasan and co-workers in their synthesis
of (2S,3S)-reboxetine, prepared an analogous amino alcohol via an
azide intermediate.15 In a similar manner, 9 was reacted with so-
dium azide to give 11 in 85% yield. Catalytic hydrogenation of azide
11 with 10% palladium on carbon at atmospheric pressure gave the
key amino alcohol 12 in 90% yield. Amine 12 was then acetylated
with chloroacetyl chloride and the morpholinone ring was formed
by subsequent reaction with potassium tert-butoxide. The
14
15
Scheme 1. Reagents and conditions: (a) AD-Mix-b, MeSO2NH2, NaHCO3, t-BuOH,
H2O, 0 °C, 66%; (b) NaOH, THF, 0 °C, 98%; (c) NaN3, DMF, , 85%; (d) 10% Pd/C, H2,
EtOAc, 90%; (e) chloroacetyl chloride, Et3N, MeCN, À10 °C to rt, 52%; (f) sodium tert-
butoxide, t-BuOH, 40 °C, 58%; (g) BH3.Me2S, THF, 0 °C to rt, 78%; (h) Boc2O, Et3N,
DMAP (cat.), CH2Cl2, 56%.
D
morpholinone ring was reduced using the borane–dimethylsulfide
complex and the resulting amine was Boc-protected to give 15 in
good yield over the two steps.
The last stage of the synthesis of 5, 6 and 7 required the intro-
duction of the iodophenol moieties with inversion of configuration
at the C-3 position. This was achieved by activating alcohol 15 as a
mesylate under standard conditions (Scheme 2). Mesylate 16 was
then reacted with 2-iodo, 3-iodo and 4-iodophenol in the presence
of caesium carbonate to give the desired (2R,3S)-coupled products
17–19 with complete inversion of configuration at the C-3 posi-
tion.16 Removal of the Boc-protecting groups of 17–19 with TFA
then gave the desired targets 5–7 in good overall yields.
In vitro competition binding of compounds 5, 6 and 7 at NAT,
SERT and DAT was carried out using homogenates of rat brain
(Table 1).17 The ortho-iodophenoxy analogue 5 showed excellent
affinity for NAT with a Ki value of 8.4 nM and with a 6-fold and
63-fold selectivity compared to SERT and DAT, respectively. This
result again shows the suitability of (2R,3S)-analogues of reboxe-
tine for the development of high affinity agents of NAT. The meta-
and para-iodophenoxy analogues, 6 and 7, showed considerably
less affinity for NAT compared to 5. The high affinity of 5 relative
to 6 and 7 is predictable as reboxetine 1 and other analogues that
show high affinity for NAT all have ortho-phenoxy substitutents.7–
9,18,19
The significantly high difference in affinity between 5 and 6
(202-fold difference) clearly shows that a binding pocket is avail-
able to bind specifically ortho-substituents and even a slight shift
to the meta-position results in a dramatic loss of affinity. While

4998
N. K. Jobson et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4996–4998
Sharpless asymmetric dihydroxylation to establish the key stereo-
genic centres. The final stage of this route involved the flexible and
stereospecific introduction of various iodophenoxy moieties. While
this approach has similar overall yields compared to our previous
stereoselective syntheses of iodinated reboxetine analogues, its
main advantage is that it permits the introduction of diversity at
a much later stage of the synthetic route (e.g., penultimate step)
and thus, allows easier preparation of small libraries of com-
pounds. More importantly, in vitro testing of these compounds
has identified ortho-iodophenoxy analogue 5 as having excellent
affinity for NAT and therefore, significant potential for develop-
ment as a SPECT tracer for this receptor. The preparation of meta-
and para-iodophenoxy analogues 6 and 7 has allowed us to probe
the structure–activity-relationship of the iodophenoxy moiety of
these compounds and the results have emphasised the require-
ment of ortho-substituted analogues for high affinity with this
receptor. The radiosynthesis and in vivo evaluation of 5 as a SPECT
imaging agent for NAT and the development of further analogues
of 7 for imaging SERT is currently underway.
a
O
O
HO
MsO
N
N
Boc
Boc
15
16
b
c
I
I
O
O
O
O
N
H
N
Boc
5-7
17-19
Acknowledgements
Scheme 2. Reagents and conditions: (a) MeSO2Cl, Et3N, DMAP (cat.), CH2Cl2, 88%;
(b) 2-iodophenol, 3-iodophenol or 4-iodophenol, Cs2CO3, , 1,4-dioxane, (17, 61%;
D
18, 32%; 19, 52%); (c) TFA, CH2Cl2 (5, 76%; 6, 80%; 7, 55%).
Financial support from EPSRC (DTA award to NKJ), BBSRC
(Industrial Studentship with GSK to ARC) and Medical Research
Scotland is gratefully acknowledged.
Table 1
Binding affinity of (2R,3S)-reboxetine analogues 5, 6 and 7 with NAT, SERT and DAT
Supplementary data
Compound
NAT Ki (nm)
SERT Ki (nm)
DAT Ki (nm)
Supplementary data (Experimental procedures and spectro-
scopic data for all compounds synthesised as well as details for
competition binding assays.) associated with this article can be
found, in the online version, at doi:10.1016/j.bmcl.2009.07.064.
8.4 1.7a
51.5 8.4
525.9 125.5
O
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O
I
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1100 200
34.5 1.7
2100 300
O
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N
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7
14. The enantiomeric excess of compound 9 was determined by chiral HPLC. See
Supplementary data for full details.
a
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A a
[3H]nisoxetine displacement assay was used for NAT, [3H]citalopram
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DAT. See Supplementary data for full details.
para-iodophenoxy analogue 7 showed low affinity for NAT, it did
show high affinity (Ki 34.5 nM) and high selectivity for SERT and
thus, further analogues of 7 may find application in the SPECT
imaging of this receptor.
In summary, a stereoselective 10-step synthesis of (2R,3S)-
iodophenoxyreboxetine analogues has been achieved using a
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