1,2,3-Triazolyl amino acids as AMPA receptor ligands

Article abstract of DOI:10.1016/j.bmcl.2010.09.139

The central nervous system glutamate receptors are an important target for drug discovery. Herein we report initial investigations into the synthesis and glutamate receptor activity of 1,2,3-triazolyl amino acids. Two compounds were found to be selective

Full text of DOI:10.1016/j.bmcl.2010.09.139

Bioorganic & Medicinal Chemistry Letters 20 (2010) 7512–7515
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
1,2,3-Triazolyl amino acids as AMPA receptor ligands
N. J. Stanley a, D. Sejer Pedersen b, B. Nielsen b, T. Kvist b, J. M. Mathiesen b, H. Bräuner-Osborne b,
D. K. Taylor c, A. D. Abell a,
⇑
a Discipline of Chemistry, The University of Adelaide, Adelaide 5005, Australia
b Department of Medicinal Chemistry, University of Copenhagen, 2100 Copenhagen, Denmark
c School of Agriculture, Food and Wine, The University of Adelaide, Glen Osmond 5005, Australia
a r t i c l e i n f o
a b s t r a c t
Article history:
The central nervous system glutamate receptors are an important target for drug discovery. Herein we
report initial investigations into the synthesis and glutamate receptor activity of 1,2,3-triazolyl amino
acids. Two compounds were found to be selective AMPA receptor ligands, which warrant further
investigation.
Received 3 September 2010
Revised 27 September 2010
Accepted 28 September 2010
Available online 14 October 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Synthesis
Glutamate receptor
Triazole
(S)-Glutamate (1) is the primary excitatory neurotransmitter in
the central nervous system (CNS) and its receptors are an important
target for drug discovery. Dysfunction of glutamatergic systems has
been implicated in a wide range of neurological disorders including
chronic pain, schizophrenia, Alzheimer’s disease, epilepsy, drug
dependence and depression.1–4 There are two distinct types of gluta-
mate receptor: ionotropic and metabotropic. The ionotropic
glutamate receptors (iGluRs) are a class of heteromeric ligand-gated
agonist ACPA (5), and the antagonists AMOA (6), ATOA (7) and ATPO
(8) have an appended acid substituent (carboxylic acid or phos-
phonic acid), which is tethered via an ether linkage for 6–8.8–12
One class of heterocycle, that is, yet to be investigated in struc-
tures of this type is the triazole. These derivatives are attractive
candidates because of their ease of preparation via a copper or
ruthenium-catalysed cycloaddition (‘Click’) reaction of suitably
functionalized azides and alkynes. Such a ‘Click’ synthetic strategy
provides the potential to prepare a library of compounds from sim-
ple and readily available fragments. Here we report the synthesis
and assay of a series of homologues (11–16, Fig. 2) as a first step
toward investigating the potential of this new class.
ion channels comprising three subtypes: N-methyl-D-aspartate
(NMDA), (R,S)-2-amino-3-(3-hydroxy-5-methylisoxazol-4-yl)pro-
pionic acid (AMPA) and 2-carboxy-3-carboxymethyl-4-isopropenyl
pyrrolidine (kainate), whereas the metabotropic glutamate recep-
tors (mGluRs) are part of the G-protein coupled receptor (GPCR)
super-family comprising eight receptor subtypes.5 The AMPA recep-
tor is found as a heteromeric tetramer, consisting of a varying com-
bination of four subunits, labelled GluR1–4, which exist in two
distinct, alternatively spliced isoforms, flip and flop.6
There has been an abundance of research into the development
of new compounds targeting AMPA receptors, which has resulted
in many potent competitive ligands. However, compounds pos-
sessing both potency and receptor subunit specificity remain rela-
tively few.7 Figure 1 depicts several known AMPA receptor ligands,
with the structure of (S)-Glutamate given for reference.
The
p-excessive heterocycles of HIBO (2), AMPA (3) and TDPA
(4) are known to make important contacts with key amino acid
residues within the binding pocket of the corresponding recep-
tor.13 We anticipated that the triazole of the new ligands (11–16)
would be able to do likewise. The structures 11–16 also contain a
carboxylic acid substituent as found in the known ligands 5–8.
The analogues 13 and 16 contain an additional tethered phenyl
group, as found in APPA (9) and 2-Py-AMPA (10). This series pro-
vides some insight into possible binding interactions as a first step
toward establishing some preliminary SAR.14,15
Given the structural similarity between the proposed triazoles
and known competitive ionotropic glutamate receptor ligands
3–10, we anticipated that they may also show affinity for gluta-
mate receptors. To test this hypothesis, compounds 11–16 were
screened in vitro against both native ionotropic (NMDA, AMPA
and kainate) and recombinant metabotropic (mGluR1, mGluR2
and mGluR4) glutamate receptors. Finally, in silico molecular
docking was carried out in an attempt to rationalise the in vitro
receptor binding results.
All the examplesin Figure 1 contain a keysubstituted p-excessive
heterocyclic core. The agonists HIBO (2), AMPA (3), TDPA (4), APPA
(9) and 2-Py-AMPA (10) have a hydroxyl group appended to the het-
erocycle, with 9 and 10 having an additional aryl substituent. The
⇑
Corresponding author. Tel.: +61 8 8303 5652; fax: +61 8 8303 4358.
E-mail address: andrew.abell@adelaide.edu.au (A.D. Abell).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.09.139

N. J. Stanley et al. / Bioorg. Med. Chem. Lett. 20 (2010) 7512–7515
7513
Figure 1. Representative structures of the endogenous ligand, glutamate (1) and other known AMPA receptor ligands.
Figure 2. Series of potential glutamate receptor ligands.
Ph
N3
N3
N3
CO2But
CO2Me
BocHN
CO2R
R = H
R = Me
17
18
19:
20:
BocHN
CO2H
BocHN
CO2Me
CO2But
Scheme 1. Synthesis of 1,4-disubstituted triazolyl amino acids by copper-catalysed
cycloaddition. Reagents and conditions: (a) CuSO4Á5H2O (5 mol %), sodium ascor-
bate (20 mol %), t-BuOH/H2O (2:1); (b) 6 M HCl, 1,4-dioxane; (c) 1 M LiOH(aq), THF.
21
22
23
Figure 3. Azide and alkyne building blocks.
26 in 85% and 64% yield, respectively. The N-Boc and ester protect-
ing groups of 24, 25, and 26 were removed as shown in Scheme 1
to give the desired 1,2,3-triazolyl amino acids 11–13 in quantita-
tive yield.
The synthesis of the alternative 1,5-regioisomers 14–16, was
achieved using the ruthenium catalyst, CpÃRu(PPh3)2Cl.16,17 The
protected azide 20 underwent ruthenium-catalysed cycloaddition
with alkyne 21 to afford the fully protected 1,5-triazole 27 in
33% yield. The protected azides 17 and 18 were similarly and
Synthesis: The key step in the synthesis of the 1,2,3-triazolyl
amino acids involved the copper (I) and ruthenium (II) catalysed
cycloaddition of azides (17–20) with alkynes (21–23) (Fig. 3). In
particular, azide 19 was reacted with alkyne 21 in the presence
of copper sulfate and sodium ascorbate to give the protected triaz-
olyl amino acid 24 in 96% yield (Scheme 1). The azides 17 and 18
were separately reacted with alkyne 22 under copper cycloaddi-
tion conditions to afford protected triazolyl amino acids 25 and

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N. J. Stanley et al. / Bioorg. Med. Chem. Lett. 20 (2010) 7512–7515
Table 1
In vitro ionotropic glutamate receptor binding data
Compounds
NMDA Ki (
l
M)
AMPA IC50
0.04a
(
l
M)
Kainate IC50 (lM)
AMPA (3)
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
11
12
13
14
15
16
63 [54;74]b
>100
>100
49 [44;55]b
>100
>100
Values are expressed as the antilog to the log mean of three-four individual
experiments.
a
Data from Vogensen et al. (2004).18
Numbers in parentheses indicate maximum and minimum SEM.
b
11–16 was carried out using Autodock 4 to gain some insight into
their possible mode of binding. The compounds were automati-
cally docked into the rigid structure of the GluR2flop ligand binding
domain. A representative docked structure for the most active li-
gand 14 is shown in Figure 4B, along with an overlay with docked
glutamate and the ligand bound X-ray structure of AMPA19 for
comparison (Fig. 4A). All numbering is taken directly from the Pro-
tein Data Bank file.
Scheme 2. Synthesis of 1,5-disubstituted triazolyl amino acids by ruthenium-
catalysed cycloaddition. Reagents and conditions: (a) CpÃRu(PPh3)2Cl (2 mol %),
THF, N2, reflux; (b) 1 M LiOH(aq), THF; (c) 6 M HCl, 1,4-dioxane.
separately reacted with alkyne 23 to give the triazoles 28 and 29 in
92% and 33% yield, respectively (Scheme 2). Attempted ruthenium-
catalysed reaction using the free acids 19 or 22 gave poor yields of
the corresponding triazole, with considerable product decomposi-
tion. The N-Boc and ester protecting groups of 27–29 were re-
moved as detailed is Scheme 2 to give the amino acids 14–16 in
high yield.
Pharmacology: The in vitro receptor binding potency of com-
pounds 11–16 was determined for native ionotropic glutamate
receptors (NMDA, AMPA, kainate) and in functional assays for re-
combinant metabotropic glutamate receptors. The data obtained
from the in vitro competitive binding assays at the ionotropic glu-
tamate receptors indicates that compounds 11 and 14 exhibited
significant competitive binding with IC50 values of 63
49 M, respectively (Table 1). Compared to AMPA (3), with an
IC50 value of 0.04 M, these compounds show weak binding affin-
lM and
l
l
ity, however, they are comparable to the binding affinity for the
antagonists AMOA (6), ATOA (7) and ATPO (8).11,18 It must be noted
that these assays only indicate binding affinity. Further work is re-
quired to characterise the mode of receptor activation, that is, ago-
nist or antagonist, of the active compounds 11 and 14.
The functional assays at the metabotropic glutamate receptors
1, 2 and 4 reveal that compounds 11–13, 15 and 16 (14 not tested)
were inactive, with an EC50 >100
unexpected, considering ligands 11–16 share a related amino
acid-substituted -excessive heterocylic core to the known AMPA
lM. This result is not altogether
p
ligands shown in Figure 1. Of these isoxazole ligands only HIBO
shows any metabotropic receptor activity. This is possibly due to
the separation of the distal acid and amino acid groups.
Figure 4. (A) Overlap of triazole 14 (white) with (S)-AMPA (3) (magenta) and
(S)-glutamate (1) (cyan), as derived from in silico docking against the GluR2flop
AMPA receptor subunit (PDB: 1ftm); (B) docked triazolyl amino acid 14 (white)
showing possible hydrogen bonding interactions with selected amino acid residues.
Only selected amino acid residues are shown. Figure produced in PyMOL.
Molecular Docking: In silico molecular docking of the GluR2flop
ligand binding subunit (PDB: 1ftm)19 with glutamate, AMPA and

N. J. Stanley et al. / Bioorg. Med. Chem. Lett. 20 (2010) 7512–7515
7515
Figure 4(A) reveals a good overlap of 14 with docked glutamate
Supplementary data
and the AMPA X-ray crystal structure. Hydrogen bonding interac-
tions of 14 are evident with key amino acid residues Pro89,
Thr91, Arg96, Gly141, Ser142 and Glu193 (Fig. 4B). These results
suggest a good computational model for the 3D orientation of
bound 14. AMPA was also docked into the GluR2flop ligand binding
subunit (results not shown) and this complex was calculated to
have a mean docking energy of À9.92 kcal/mol, compared to
À9.35 kcal/mol for triazole 11 and À9.14 kcal/mol for triazole 14.
These values are in agreement with the observed potency of these
three derivatives (see in vitro data in Table 1). However, unlike
AMPA, neither compound 11 nor 14 make polar contacts with
the hydroxyl group of Thr143 (left-hand edge, Fig. 4) and com-
Supplementary data (full compound characterisation data and
in vitro assay details) associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.09.139.
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pound 11 also lacks
a contact with Glu193 (bottom edge,
Fig. 4).13 This may contribute to the lower potency of 11 and 14
compared to AMPA. The distal carboxylic acid groups in docked
12 and 15 (not shown) occupy a region of the receptor not occu-
pied by AMPA. Triazoles 13 and 16 docked with high docking ener-
gies, possibly due to the bulky phenyl rings interacting in a
sterically or electrostatically disfavoured manner. This is consis-
tent with inactivity of these derivatives. (See Table 1).
Summary: This work highlights a convenient and versatile syn-
thesis of the 1,4- and 1,5-disubstituted 1,2,3-triazolyl amino acid
glutamate homologues 11–16. In vitro screening indicated selective
binding for 11 and 14 at AMPA receptors, albeit with low potency.
Further structural elaboration and pharmacological investigation
is needed in order to establish the full potential of the 1,2,3-triazolyl
amino acid core structure.
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This work was generously funded by the Australian Research
Council under Grant DP0771901. N.J.S. is grateful to be the recipient
of a University of Adelaide Divisional Scholarship. T.K. and M.B.-O.
were funded by the GluTarget Center of Excellence and J.M.M. was
funded by the Danish Council for Independant Research.
19. Armstrong, N.; Gouaux, E. Neuron 2000, 28, 165.