3-Substituted phenylalanines as selective AMPA- and kainate receptor ligands

Article abstract of DOI:10.1016/j.bmc.2009.07.021

On the basis of X-ray structures of ionotropic glutamate receptor constructs in complex with amino acid-based AMPA and kainate receptor antagonists, a series of rigid as well as flexible biaromatic alanine derivatives carrying selected hydrogen bond acceptors and donors have been synthesized in order to investigate the structural determinants for receptor selectivity between AMPA and the GluR5 subtype of kainate receptors. Compounds selective for either GluR5 or AMPA receptors were identified. One particular substituent position appeared to be of special importance for control of ligand selectivity. Using molecular modeling the observed structure-activity relationships at AMPA and GluR5 receptors were deduced.

Full text of DOI:10.1016/j.bmc.2009.07.021

Bioorganic & Medicinal Chemistry 17 (2009) 6390–6401
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
3-Substituted phenylalanines as selective AMPA- and kainate receptor ligands
a
b
a
a,
*
´
Ewa Szymanska , Darryl S. Pickering , Birgitte Nielsen , Tommy N. Johansen
a Department of Medicinal Chemistry, Faculty of Pharmaceutical Sciences, University of Copenhagen, 2 Universitetsparken, DK-2100 Copenhagen, Denmark
b Department of Pharmacology and Pharmacotherapy, Faculty of Pharmaceutical Sciences, University of Copenhagen, 2 Universitetsparken, DK-2100 Copenhagen, Denmark
a r t i c l e i n f o
a b s t r a c t
Article history:
On the basis of X-ray structures of ionotropic glutamate receptor constructs in complex with amino acid-
based AMPA and kainate receptor antagonists, a series of rigid as well as flexible biaromatic alanine
derivatives carrying selected hydrogen bond acceptors and donors have been synthesized in order to
investigate the structural determinants for receptor selectivity between AMPA and the GluR5 subtype
of kainate receptors. Compounds selective for either GluR5 or AMPA receptors were identified. One par-
ticular substituent position appeared to be of special importance for control of ligand selectivity. Using
molecular modeling the observed structure–activity relationships at AMPA and GluR5 receptors were
deduced.
Received 4 August 2008
Revised 27 June 2009
Accepted 12 July 2009
Available online 16 July 2009
Keywords:
Ionotropic glutamte receptors
Antagonist
Suzuki cross-coupling
Competitive
Ó 2009 Elsevier Ltd. All rights reserved.
AMPA
GluR5
1. Introduction
nate between homomeric AMPA and kainate receptors. Only
recently the available crystal structures of the iGluR binding core
Ionotropic glutamate receptors (iGluRs) are the major excit-
atory amino acid neurotransmitter receptors in the vertebrate cen-
tral nervous system and are separated into three functionally
in complex with agonists or antagonists have provided more de-
tailed information on ligands’ binding modes and insight into the
molecular mechanisms of ligand selectivity. Well known examples
are unselective AMPA/GluR5 antagonist (S)-2-amino-3-[5-tert-
butyl-3-(phosphono-methoxy)-4-isoxazolyl]-propionic acid [(S)-
ATPO (1)], co-crystallized with GluR2 as well as GluR5 binding
cores9–11 and complexes of the selective GluR5 antagonists (S)-1-
(2-amino-2-carboxyethyl)-3-(2-carboxybenzyl)-pyrimidine-2,4-
dione (UBP302, 3) and (S)-1-(2-amino-2-carboxyethyl)-3-(2-carb-
oxythiophene-3-yl)methyl-5-methyl-pyrimidine-2,4-dione (UBP310,
4)12–15 co-crystallized with the GluR5 construct (Fig. 1). UBP302
and UBP310 belong to the group of willardiine-based antagonists
and show nanomolar antagonist potency at human recombinant
GluR5 receptors with high selectivity against cloned human
GluR1-4 and GluR6 receptors.15,16
distinct subclasses: N-methyl-D-aspartic acid (NMDA), (RS)-2-ami-
no-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA)
and (2S,3S,4S)-3-carboxymethyl-4-isopropenyl-pyrrolidine-2-car-
boxylic acid (kainate, KA) receptors. All three classes play an
important role in controlling synaptic plasticity, underlying learn-
ing and memory processes and other central nervous system (CNS)
functions, such as synaptogenesis and the development of func-
tional neurons.1–4 AMPA receptors, which mediate the majority
of the fast excitatory amino acid synaptic transmission in the
CNS and affect short-term changes in synaptic strength, are cat-
ion-selective tetrameric heterooligomers formed by combinations
of the subunits GluR1-4, whereas kainate receptors are tetrameric
assemblies of GluR5-7, KA1 and KA2 subunits. Their role is not fully
understood; however, studies suggest that both pre- and postsyn-
aptic kainate receptors can regulate neurotransmission at many
synapses and seem to be involved in short- and long-term plastic
phenomena.5 Recently GluR5 receptors were reported to play an
important role in nociception and central sensitization.3,6–8
Competitive non-NMDA iGluR amino acid-based antagonists
typically consist of an a-amino part and a distal acidic part linked
together through a heterocyclic ring system. Analysis of the crystal
structures of the (S)-ATPO, UBP302 and UBP310 complexes9,11,15
revealed that the spacer helps stabilize the receptor in an open
conformation.
There is a clear pharmacological distinction between NMDA and
other iGluRs. However, most of AMPA receptor agonists and antag-
onists bind also kainate receptors and only few ligands discrimi-
In the present study we have set out to replace the linker parts
of the UBP compounds with two aromatic ring systems and to
investigate the influence of a carboxylic acid substituent in the dis-
tal phenyl ring on iGluR affinity (Fig. 1b). With replacement of ura-
cil by phenyl it is possible to introduce substituents (R1) in the
proximal phenyl ring exploiting various regions of the binding
* Corresponding author. Tel.: +45 35 33 64 12; fax: +45 35 33 60 01.
E-mail address: tnj@farma.ku.dk (T.N. Johansen).
0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.07.021

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E. Szymanska et al. / Bioorg. Med. Chem. 17 (2009) 6390–6401
Figure 1. (a) Structures of selected AMPA and kainate receptor antagonists. (b) Structures of target compounds.
pocket and potentially increasing the affinity to the receptor. Based
2. Results and discussion
2.1. Chemistry
on the analysis of the crystal structures, introduction of a phenyl
ring instead of the heterocyclic parts of the ligands is expected to
be accepted both by AMPA as well as GluR5 receptors. In analogy
with the UBP compounds a second phenyl/benzyl/thiophenyl frag-
ment bearing carboxyl or methyl carboxyl groups has been placed
in position 3 of the phenylalanine ring, to some extent mimicking
the substitution pattern of the UBP compounds. The position of the
carboxylic substituent R2 in the distal phenyl has also been
examined.
In addition, we also decided to investigate analogues in which
the terminal acidic group was replaced by a series of hydrogen
bond acceptors or donors, such as nitro, tetrazolyl and hydroxyl
substituents (R2). All new target compounds were pharmacologi-
cally characterized at native (NMDA, AMPA, KA) and at recombi-
nant (GluR5, GluR6, GluR7) receptors. Finally, in attempt to
explain the observed selectivity towards either AMPA-R or GluR5,
molecular docking of selected target compounds was performed,
using as templates the (S)-ATPO-GluR2 and UBP302-GluR5 binding
core complexes (PDB ID: 1N0T and 2F35), respectively.
The synthetic pathways for the new phenylalanine derivatives
are illustrated on the Schemes 1–5.
Most of the target amino acids were prepared as a result of a
number of Suzuki cross-coupling reactions of aryl halides with
boronic acids. Hence, synthesis of the key intermediates 7, 16,
24, 29 and 30 containing iodo-, bromo- or methylbromo- substitu-
ents, was crucial for the strategy.
Compounds 6 and 15 were obtained as a result of benzylic bro-
mination of 5 and 14, respectively, with NBS under free radical
conditions (Scheme 1). 3-Iodotoluene (5) is commercially avail-
able; 1,2-dichloro-3-iodo-5-methyl-benzene (14) was synthesized
according to the literature methods from 2-chloro-4-methylaniline
(10). Nitration into the position 6 of the phenyl ring with 96% nitric
acid, following the Sandmayer reaction using CuCl2 gave 1,2-di-
chloro-5-methyl-3-nitrobenzene (12). Reduction of the nitro group
with Fe in the presence of hydrochloric acid afforded 13, which
Scheme 1. Reagents and conditions: (i) (1) (CH3COO)2O, 96% HNO3, <15 °C, (2) HCl, reflux; (ii) (1) CuCl2, tert-BuNO2, acetonitrile, 65 °C, (2) 20% HCl; (iii) Fe, HCl, ethanol,
reflux; (iv) (1) NaNO2, H2SO4, acetic acid, 25 °C ? 75 °C, (2) urea, KI; (v) NBS, benzoyl peroxide, CCl4, reflux; (vi) NaH 1.5 equiv, diethyl acetamidomalonate 1.5 equiv, DMF, rt;
(vii) substituted phenylboronic acid 1.5 equiv, PdCl2(PPh3)2 5 mol %, triethylamine, DME:water (1:1), rt or 50 °C; (viii) 48% HBr, reflux.

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E. Szymanska et al. / Bioorg. Med. Chem. 17 (2009) 6390–6401
Scheme 2. Reagents and conditions: (i) NaN3, TEA hydrochloride, DME, reflux; (ii) HCl/H2O/AcOH (1:1:1), reflux.
Scheme 3. Reagents and conditions: (i) 5-cyanothiophene-2-boronic acid 1.5 equiv, Pd(PPh3)4 5 mol %, K2CO3 3 equiv, toluene/ethanol/water (10:3:3), rt ? 50 °C; (ii) HCl/
H2O/AcOH (1:1:1), reflux.
Scheme 4. Reagents and conditions: (i) NaH 1.1 equiv, diethyl acetamidomalonate 1.1 equiv, DMF, rt; (ii) phenylboronic acid 1.5 equiv, [1,10-(PPh2)2Fe]PdCl2 5 mol %, 3 M
NaOH, TBAB, THF:water (5:1), 50 °C; (iii) 48% HBr, reflux.
was converted into the iodide 14 through the corresponding diazo-
nium salt. The key intermediates 7 and 16 were prepared from the
benzyl bromides 6 and 15, respectively, through substitutions with
the sodium salt of diethyl acetamidomalonate.
be ineffective. However, using an electron-rich and hindered
PdCl2(dppf) complex ([1,10-bis(diphenylphosphino)ferrocene]
dichloropalladium(II)) in a biphasic solvent system (THF and water
5:1) at 50 °C and under the presence of tetrabutylammonium bro-
mide (TBAB), the conversions were completed within 2 h to give
25a–c in good yields. The phase transfer catalyst, TBAB, has re-
cently been used successfully in Suzuki couplings performed under
biphasic conditions,17–19 enhancing the rate of the coupling reac-
tion by activating the boronic acid through formation of a boronate
complex [ArB(OH)3]ꢀ [R4N]+.
Deprotection of the prepared biphenyl-, thiophenyl- and ben-
zylphenyl derivatives in 48% aqueous hydrobromic acid or—under
milder conditions—in a mixture of acetic acid and hydrochloric
acid followed by reverse-phase HPLC purification afforded the tar-
get amino acids 9a–e, 18a–h, 20a–c, 22 and 26a–c.
In order to prepare the S- and R-enantiomers of 9b (Scheme 5)
commercially available (S)- and (R)-N-Boc-3-bromo-phenylala-
nine, respectively, were treated with ethyl iodide in the presence
of sodium bicarbonate in DMF,20 followed by reaction with 3-cya-
no-phenylboronic acid under Suzuki coupling conditions as de-
scribed for 8b. Deprotection in 48% aqueous hydrobromic acid
gave the desired amino acids 33 (S, ee = 99.2%) and 34 (R,
ee = 98.4%).
Treatment of 7 and 16 with phenylboronic acids in the presence
of palladium complexes afforded the biphenyl derivatives 8a–e
and 17a–h. (Scheme 1). A short optimization of Suzuki cross-cou-
pling conditions was performed: various palladium sources, sol-
vents and bases were examined; however, as most of the
reactions were conducted at elevated temperature and because
of the basic conditions, partial deprotection of the diethyl acetam-
idomalonate moiety was observed, which led to problems with iso-
lation of the product and low yield. Eventually, use of a mixture of
dimethoxyethane (DME) and water (1:1) as solvent, PdCl2(PPh3)
and triethylamine as base turned out to produce the most effective
conditions, under which most reactions could be completed at
room temperature.
To introduce the tetrazolyl ring system into the distal phenyl
ring, the corresponding nitriles 17a–c were heated for 48 h with
sodium azide in DME in the presence of triethylammonium hydro-
chloride (Scheme 2).
In the case of 5-cyanothiophene-2-boronic acid however, the
Suzuki cross-coupling did not lead to the coupled product. Instead,
de-iodination was observed. After optimizing the conditions, com-
pound 21 was obtained—only in low yield though—using a mixture
of toluene–ethanol–water (in ratio 10:3:3) as solvent and
Pd(PPh3)4 as catalyst (Scheme 3).
2.2. Pharmacology
All target compounds were tested for in vitro pharmacological
activity at recombinant rat GluR5(Q)1b, GluR6(V,C,R) and GluR7A
homomeric kainate receptors expressed in Sf9 insect cell mem-
In the case of benzylbromide 24 (Scheme 4) the Suzuki coupling
reaction conditions used for the preparation of 8a–e turned out to

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E. Szymanska et al. / Bioorg. Med. Chem. 17 (2009) 6390–6401
Scheme 5. Reagents and conditions: (i) C2H5I 5 equiv, NaHCO3 2 equiv, DMF, rt; (ii) 3-cyanophenylboronic acid 1.5 equiv, PdCl2(PPh3)2 5 mol %, triethylamine, DME:water
(1:1), rt; (iii) 48% HBr, reflux.
branes as well as at the native AMPA, kainate and NMDA receptors
in rat cortical membranes (Table 1).
In the AMPA receptor binding assay to native receptors, all tar-
get compounds containing carboxyl groups showed either no or
(7.9 lM and 2.8 lM, respectively); comparable to that of (S)-ATPO.
Adding two chlorines as R1 substituents to the phenylalanine ring
improved GluR5 potency, making analogue 18b the most active
GluR5 ligand among the obtained compounds. Interestingly, nei-
ther 9b nor 18b showed measurable affinity towards native iGluR:
AMPA, kainate or NMDA receptors.
To examine, which isomer of the active target 3-carboxy substi-
tuted biphenyl compounds is responsible for GluR5 kainate recep-
tor activity, both the S- and R-enantiomers of 9b (compounds 33
and 34, respectively) were synthesized and tested. As expected
on the basis of similarity of structures 9b and UBP302, the
GluR5-subtype activity was located in the S-isomer of 9b (33),
while the corresponding R-enantiomer (34) showed no measurable
affinity (Ki > 1 mM).
The position of the carboxyl substituent seems to be imperative
for high affinity, as none of the positional isomers of 9b and 18b
displayed any significant affinity. In addition, replacement of the
acidic group with an hydroxy group (18d–f), nitro group (18g, h)
or tetrazole ring (20a-c), or prolongation of the terminal acidic
chain (9d,e) or exchange of the whole carboxybenzene fragment
into 2-carboxythiophene-5-yl resulted in a marked drop of affinity
at GluR5 compared to 9b/18b.
very low affinity (IC50 higher than or close to 100 lM). However,
only analogues containing a hydroxyl substituent showed measur-
able affinities, 18e (the dichloro-substituted biphenyl analogue
possessing an hydroxyl group in position 3 of the distal phenyl
ring) being the most active (IC50 = 4.6 lM); more potent than (S)-
ATPO. Change of substituent position resulted in a decrease of
affinity; the 4-hydroxy analogue (18f) was approximately seven-
fold less potent than the parent compound, while the 2-hydroxy
analogue (18d) was almost inactive in this assay. Weak micromo-
lar affinity towards AMPA receptors was also expressed by 4-car-
boxy substituted biphenyl analogues 9c and 18c and
carboxythiophene analogue 22. None of the amino acids active in
the AMPA receptor binding assay displayed measurable affinity
to GluR5s.
At homomeric GluR5, compounds with structures based on the
biphenyl core with a carboxylic acid group attached to the distal
phenyl ring in position 3 (9b and 18b) showed the highest affinity
In addition, the three analogues possessing the methylene lin-
ker between the two phenyl rings (26a–c) showed almost no affin-
ity at GluR5, regardless of the position of the carboxylic group.
None of the target amino acids expressed measurable affinity to
native NMDA or KA receptors, GluR6 or GluR7 rat homomeric re-
combinant receptors.
Table 1
Binding affinities at native AMPA receptors and homomeric GluR5 kainate receptorsa
IC50
(
l
M)
Ki (lM)
GluR5
[3H]AMPA
(S)-ATPO
UBP302c
9a
9b
9c
16 1b
nd
2.90 0.26
nd
>100
>100
>100
7.9 0.8
>100
2.3. Molecular modeling study
41
2
9d
9e
>100
>100
>100
>100
P100
>100
>100
2.80 0.78
>100
>100
>100
>100
>100
>100
>100
P100
>100
P100
P100
50–100
P100
4.08 0.45
>1000
To understand recombinant homomeric GluR5 affinity data ob-
tained for the new compounds, molecular docking of selected com-
pounds was undertaken. The series of structures with biphenyl or
phenylbenzyl core acting as a spacer between amino acid part and
the distal carboxyl group (9a–c, and 18a–c) were docked into the
ligand-binding domain of UBP302-GluR5 complex (PDB ID: 2F35).
Molecular modeling was performed using Schrödinger Suite
2007 environment.21 The initial ligand structures were prepared
with MacroModel v. 9.5 (MMFFs forcefield including water solva-
tion, carboxy and amino groups were ionized). Only S-enantiomers
were taken into account. Protein preparation and docking of li-
gands was performed using Glide 4.5 (all water molecules were re-
moved from the protein before docking). The obtained poses, in
which amino acid chain was bound in the way corresponding to
the binding mode of UBP302, were ranked according to highest
scores (GlideScore function) and lowest energies.
18a
18b
18c
18d
18e
18f
18g
18h
20a
20b
20c
22
42
85
6
1
4.6 0.2
33
4
>100
>100
>100
>100
>100
61
8
26a
26b
26c
33
>100
>100
>100
nd
34
nd
The most effective poses were observed for the structures 9b
and 18b (the top pose of 18b is presented in Fig. 2). As expected,
nd = Not determined.
a
Values come from three independent experiments and are expressed as the
mean SEM.
apart from contacts between
a-amino acid and residues Arg508,
b
(S)-ATPO affinity data at AMPA receptors taken from Ref. 10; affinity at GluR5
Thr503, Pro501; 18b forms also hydrogen bond interactions be-
tween the distal carboxylic acid group and residue Thr675 of do-
main 2, resembling the binding mode of UBP302. The
homomeric receptors determined in current study.
c
UBP302 binding affinity at GluR2 S1S2 ligand-binding core Kd = 146 33 lM
and at GluR5 S1S2 ligand-binding core Kd = 3.94 0.7 l
M.15

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6394
chlorine—this effect most likely contributes to lower the affinity
of the phenylalanine amino acids at GluR5.
Docking of 2- and 4-carboxy isomers of 9b/18b resulted in low-
er scored poses with the distal acidic group interacting with Ser674
and/or Thr675. However, no clear connection between docking
scores and the binding affinity of compounds can be observed.
Low or no activity of 9a, c and 26a–c in the GluR5 assay is most
likely connected to the absence of substituents in positions 4
and/or 5 of phenylalanine.
To explain the AMPA receptor affinity data, molecular docking
of compound 18e (3-hydroxy-substituted analogue) was at-
tempted, using (S)-ATPO-GluR2 binding core complex (PDB ID:
1N0T) as a template. Preparation of the protein and ligands (both
S- and R-enantiomers) as well as the docking procedure was per-
formed in analogy to the experiment described above for the GluR5
binding core (water molecules were removed). The results were
clear-cut, with ligands given highest scores and lowest energies
in binding poses with amino acid chain bound in the way corre-
sponding to the binding mode of (S)-ATPO antagonist.
Even though certain reservations should be made, docking
experiments indicated that both enantiomers of 18e are likely to
be active at AMPA receptors. In the top ranked poses (Fig. 3a and
b), despite the difference in chiral center configuration, the amino
acidic part of both isomers makes contacts with the protein in a
way resembling the ATPO binding mode and involving Arg485,
Thr480, Pro478 and Glu705 residues. The rigid biphenyl linker be-
tween the amino acidic moiety and the 3-hydroxy group, interact-
ing with Thr655 of domain 2, stabilizes, like in case of ATPO, the
open conformation of the receptor. Chlorine atoms of phenylala-
nine are located in partly polar, partly hydrophobic cavities; the
exchange of Ser726 in GluR5 into lipophilic Met708 in GluR2
receptor has a big influence on accommodation of 18e by GluR2,
while in the case of 18b in the GluR5 binding core, a polar substi-
tuent in position 4 seems to be more suitable. The presence of chlo-
rine in this position probably also contributes to higher affinity of
18e at AMPA receptors compared to ATPO.
Figure 2. Compound 18b (blue) docked to UBP302-GluR5 S1S2 complex. Residues
of UBP302-GluR5 binding core—green, UBP302 ligand—orange. Ion pair interactions
and hydrogen bonds within the distance lower than 3.5 Å between 18b and the
protein are represented by dashes. Prepared by Pymol.22
phenylalanine ring is almost coplanar with the uracil ring of the
willardiine antagonist; position 5 in 18b, substituted with a chlo-
rine atom, overlaps with position 5 of uracil. It has recently been
reported for willardiine antagonists series that substitution in this
position of the uracil ring with methyl (i.e., in UBP310 compound)
or halogen improved both potency and selectivity for kainate
receptors.13,16 The comparison of UBP310-GluR5 with other crystal
structures of antagonists with either the GluR2 or GluR5 binding
core reveals that the methyl group of UBP310, t-butyl group in
ATPO structures, and one of the nitro groups in DNQX (Fig. 1) occu-
py a similar space of the GluR2/GluR5 binding pocket. The chlorine
atom in position 5 of phenylalanine in 18b seems to fill this partly
polar and partly hydrophobic cavity, which likely increases the
affinity to GluR5 compared to the non-substituted analogue 9b.
On the other hand, the second chlorine atom in position 4 is lo-
cated in a cavity formed by Ser726, Thr725, hydroxy groups of
Tyr749 and Tyr429 and methylene carbons of the Glu723 side
chain; polar residues surrounding this space tolerate better the
oxygen atom of the carbonyl group in UBP302 than hydrophobic
3. Conclusion
In efforts to investigate the structural determinants for receptor
selectivity between AMPA and the GluR5 receptors, a series of rigid
as well as flexible biaromatic alanine derivatives carrying selected
hydrogen bond acceptors and donors have been synthesized. The
Figure 3. Compound 18e docked to (S)-ATPO-GluR2 S1S2 complex. Residues of ATPO-GluR2 binding core—green, (S)-ATPO ligand—orange. (a) Top ranked pose of the S-
enantiomer (navy blue); (b) top ranked pose of the R-enantiomer (purple). Ion pair interactions and hydrogen bonds within the distance lower than 3.5 Å between 18e and
the protein are represented by dashes. Prepared by Pymol.22

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E. Szymanska et al. / Bioorg. Med. Chem. 17 (2009) 6390–6401
established structure–activity relationships revealed that among
the series one particular substituent position appeared to be essen-
tial for control of ligand selectivity. Among the new series of com-
pounds both GluR5-selective (18b) as well as AMPA-selective (18e)
compounds were identified. For compound 18b and for the corre-
sponding positional isomers 18a, c docking to the UBP302-GluR5
complex was in good agreement with the observed GluR5 receptor
affinity. Likewise for compound 18e, good agreement between
AMPA receptor affinity and docking to the (S)-ATPO-GluR2 com-
plex was observed. For further improvements on potency and sub-
type selectivity at AMPA/kainate receptors, compounds 18b, e
provide good starting points for the design of ligands.
reaction was followed by 1H NMR. After cooling and filtration,
dichloromethane (50 mL) was added; the organic phase was washed
with 2 M NaOH, dried (MgSO4) and evaporated. Purification by flash
column chromatography (heptane) afforded a white solid, which
consisted of a mixture of 6 and the corresponding dibromo deriva-
tive in ratio 4.3:1. Yield of 6, calculated on the basis of 1H NMR spec-
tra, 4.13 g, 70%; 1H NMR (300 MHz, CDCl3): d 4.40 (s, 2H, CH2Br), 7.08
(t, J = 7.6 Hz, 1H, Ar), 7.35 (dd, J1 = 7.6 Hz, J2 = 1.2 Hz, 1H, Ar), 7.63 (d,
1H, J = 7.9 Hz, Ar), 7.75 (s, 1H, Ar); 13C NMR (75 MHz, CDCl3): d 32.2,
94.4, 128.3, 130.5, 137.4, 137.9, 138.8.
4.3. Diethyl 2-acetamido-2-(3-iodobenzyl)malonate (7)
A 60% suspension of sodium hydride in mineral oil (0.78 g,
19.5 mmol) in dry DMF (10 mL) was cooled to 5 °C, and a solution
of diethyl acetamidomalonate (4.23 g, 19.5 mmol) in dry DMF
(20 mL) was added under nitrogen atmosphere. After stirring for
30 min the solution of 6 (mixed with 1,1-(dibromomethyl)-3-iodo-
benzene in ratio 4.3:1, recalculated amount of 6—3.86 g,
13.0 mmol) in DMF (20 mL) was added. The mixture was stirred
for 16 h at ambient temperature under nitrogen atmosphere,
3 mL of glacial acetic acid was added and the solvent was evapo-
rated under reduced pressure. The residue was suspended in tolu-
ene (80 mL), stirred for 10 min, filtered and washed with water
(80 mL) and brine (80 mL). After drying (MgSO4), evaporation
and flash column chromatography, 7 was isolated as a white solid.
Yield 3.87 g, 69%; MS: m/z 433 [M+]; 1H NMR (300 MHz, CDCl3): d
1.30 (t, J = 7.1 Hz, 6H, OCH2CH3), 2.05 (s, 3H, NHCOCH3), 3.59 (s, 2H,
ArCH2), 4.27 (q, J = 7.1 Hz, 4H, OCH2CH3), 6.54 (s, 1H, NH), 6.90–
7.03 (m, 2H, Ar), 7.37 (s, 1H, Ar), 7.57 (d, J = 7.1 Hz, 1H, Ar); 13C
NMR (75 MHz, CDCl3): d 14.3, 23.2, 37.4, 63.0, 67.2, 94.3, 129.1,
130.0, 136.2, 137.7, 139.0, 167.2 169.2.
4. Experimental
4.1. Chemistry. General procedures
Analytical thin-layer chromatography (TLC) was performed on
Silica Gel 60 F254 aluminum plates (Merck). All compounds were
detected using UV light and a KMnO4 spraying reagent, the target
amino acids were also visualized using a ninhydrin spraying re-
agent. Flash column chromatography (FC) was performed using
CombiFlash Companion System (Teledyne Isco, Inc.) on RediSep
columns with silica gel (average particle size 35–70 lm). The mix-
ture of heptane fraction and ethyl acetate (both HPLC purity) were
used as eluents, unless otherwise stated. 1H NMR spectra were re-
corded on a 300 MHz Varian Mercury 300BB or 300 MHz Varian
Gemini 2000BB spectrometer. 13C NMR spectra were recorded on
the Varian Gemini spectrometer. CDCl3 (with TMS as internal stan-
dard), D2O and DMSO-d6 (dimethyl-d6-sulfoxide) were used as sol-
vents. Chemical shifts (d) are given in parts per million (ppm), and
coupling constants (J) are given in hertz (Hz). GC–MS were re-
corded on a Shimadzu QP 5050A mounted with a Supelco MDN-
5S column and using chemical ionization.
4.4. General procedure for diethyl 2-acetamido-2-(biphenyl-3-
ylmethyl)malonates (8a–e)
Elemental analyses were performed by J. Theiner, Microanalyt-
ical Laboratory, Department of Physical Chemistry, University of
Vienna, Austria, and are within 0.4% of the theoretical values.
Unless otherwise stated, preparative chromatography was per-
formed using HPLC system consisting of a Jasco 880-PU pump, a
Rheodyne 7125 injector equipped with a 5.0 mL sample loop, a Shi-
madzu SPD-6A detector (220 nm or 250 nm) and a Merck-Hitachi
D-2000 Chromato-Integrator. Purification of the target amino acids
Compounds 8a–e were prepared in analogy to a literature meth-
od.25 Suspension of 7 (433 mg, 1 mmol) and bis(triphenylphos-
phine)palladium(II) dichloride PdCl2(PPh3)2 (35 mg, 0.05 mmol)
was stirred in DME (25 mL) under nitrogen atmosphere at room
temperature for 15 min. Phenylboronic acid (1.5 mmol), triethyl-
amine (5.58 mL, 40 mmol) and water (25 mL) were added and the
mixture was stirred under nitrogen at room temperature for several
hours, controlled by GC–MS. If the starting material was not con-
verted after 24 h of stirring, the temperature was raised to 50 °C.
When the reaction was finished, the mixture was filtrated and water
(30 mL) was added. After extraction with ethyl acetate (3 ꢁ 50 mL)
the combined extracts were washed with 2 M NaOH (50 mL) follow-
ing water (50 mL), dried (MgSO4) and evaporated. The raw product
was purified by flash column chromatography.
was performed on
a reverse-phase XTerra MS C18 column
(10 ꢁ 300 mm 10 m, Waters). The column was eluted at 8 mL/
l
min with 15 mM acetic acid containing methanol in varying con-
centrations depending on the structure of the target amino acid.
The enantiomeric purity expressed by the enantiomeric excess
(ee) was determined by chiral HPLC using an analytical Chirobiotic
T column (4.6 ꢁ 150 mm, ASTEC) equipped with a Chirobiotic T
guard column (4.6 ꢁ 50 mm, ASTEC) and connected to the HPLC
system described above (detection at 250 nm). Elution was per-
formed with 60% (v/v) ethanol in water adjusted with acetic acid
to pH 4 (0.5 mL/min). Optical rotation was measured on a Per-
kin–Elmer 241 polarimeter. CD spectra were recorded at ambient
temperature in 1.0-cm cuvettes on an OLIS DSM (Digital Subtrac-
tive Method) 10 CD spectrophotometer.
4.4.1. Diethyl 2-acetamido-2-((20-cyanobiphenyl-3-yl)methyl)-
malonate (8a)
Yield 183 mg, 45%; MS: m/z 408 [M+]; 1H NMR (300 MHz,
CDCl3): d 1.28 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.12 (s, 3H, NHCOCH3),
3.73 (s, 2H, ArCH2), 4.20–4.30 (m, 4H, OCH2CH3), 6.99 (s, 1H,
NH), 7.11–7.20 (m, 1H, Ar), 7.25 (s, 1H, Ar), 7.25–7.47 (m, 4H,
Ar), 7.64 (dt, J1 = 7.7 Hz, J2 = 1.4 Hz, 1H, Ar), 7.76 (dd, J1 = 8.3 Hz,
J2 = 1.4 Hz, 1H, Ar); 13C NMR (75 MHz, CDCl3): d 14.3, 23.4, 38.2,
62.8, 67.5, 111.3, 118.8, 127.7, 128.8, 129.9, 130.3, 130.5, 132.9,
133.6, 135.9, 138.3, 145.4, 167.3, 169.5.
4.2. 1-(Bromomethyl)-3-iodobenzene (6)
Compounds 623 and 7 were prepared according to the literature
procedure.24 A mixture of commercially available 1-iodo-3-methyl-
benzene 5 (4.36 g, 20 mmol), NBS (N-bromosuccinimide 5.34 g,
30 mmol) and benzoyl peroxide (0.727 g, 3 mmol) in tetrachloro-
methane (100 mL) was refluxed for 4 h. NBS and benzoyl peroxide
were added in adequate amounts each hour. The progress of the
4.4.2. Diethyl 2-acetamido-2-((30-cyanobiphenyl-3-yl)methyl)-
malonate (8b)
Yield 325 mg, 80%; MS: m/z 408 [M+]; 1H NMR (300 MHz,
CDCl3): d 1.30 (t, J = 7.1 Hz, 6H, OCH2CH3), 2.03 (s, 3H, NHCOCH3),

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6396
3.74 (s, 2H, ArCH2), 4.29 (dq, J1 = 7.1 Hz, J2 = 2.4 Hz, 4H, OCH2CH3),
6.56 (s, 1H, NH), 7.06 (d, J = 7.3 Hz, 1H, Ar), 7.18 (s, 1H, Ar), 7.37 (t,
J = 7.3 Hz, 1H, Ar), 7.44 (d, J = 7.6 Hz, 1H, Ar), 7.54 (t, J = 7.6 Hz, 1H,
Ar), 7.63 (d, J = 7.9 Hz, 1H, Ar), 7.72–7.78 (m, 2H, Ar); 13C NMR
(75 MHz, CDCl3): d 14.3, 23.3, 38.0, 63.0, 67.3, 113.1, 118.8,
126.1, 128.7, 129.2, 129.7, 129.9, 130.6, 130.9, 131.4, 136.4,
139.0, 142.2, 167.5, 169.1.
5.24; N, 4.34; 1H NMR (300 MHz, DMSO-d6 + 50
lL D2O): d 2.99
(dd, J1 = 14.3 Hz, J2 = 8.3 Hz, 1H, ArCH2), 3.21 (dd, J1 = 14.3 Hz,
J2 = 4.7 Hz, 1H, ArCH2), 3.61–3.65 (m, 1H, ArCH2CH), 7.27 (d,
J = 7.7 Hz, 1H, Ar), 7.40 (t, J = 7.4 Hz, 1H, Ar), 7.51–7.59 (m, 3H,
Ar), 7.86–7.90 (m, 2H, Ar), 8.14 (s, 1H, Ar).
4.6.2. (RS)-30-(2-Amino-2-carboxyethyl)biphenyl-4-carboxylic
acid (9c)
4.4.3. Diethyl 2-acetamido-2-((40-cyanobiphenyl-3-yl)methyl)-
malonate (8c)
Yield 37 mg, 26%; Anal. Calcd for C16H15NO4ꢂ0.3 C2HF3O2: C,
62.40; H, 4.83; N, 4.38. Found: C, 62.11; H, 4.92; N, 4.22; 1H
Yield 263 mg, 64%; 1H NMR (300 MHz, CDCl3): d 1.29 (t,
J = 7.1 Hz, 6H, OCH2CH3), 2.01 (s, 3H, NHCOCH3), 3.74 (s, 2H,
ArCH2), 4.28 (dq, J1 = 7.1 Hz, J2 = 2.4 Hz, 4H, OCH2CH3), 6.56 (s,
1H, NH), 7.06 (d, J = 7.6 Hz, 1H, Ar), 7.21 (s, 1H, Ar), 7.37 (t,
J = 7.6 Hz, 1H, Ar), 7.46 (d, J = 7.9 Hz, 1H, Ar), 7.61 (d, J = 8.2 Hz,
2H, Ar), 7.71 (d, J = 8.2 Hz, 2H, Ar); 13C NMR (75 MHz, CDCl3): d
14.3, 23.3, 38.0, 63.0, 67.3, 111.2, 118.9, 126.2, 127.7, 128.8,
129.2, 130.1, 132.7, 136.4, 139.3, 145.4, 167.4, 169.1.
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 2.98 (dd, J1 = 14.3 Hz,
J2 = 8.0 Hz, 1H, ArCH2), 3.21 (dd, J1 = 14.3 Hz, J2 = 4.4 Hz, 1H,
ArCH2), 3.61–3.65 (m, 1H, ArCH2CH), 7.28 (d, J = 7.7 Hz, 1H, Ar),
7.41 (t, J = 7.4 Hz, 1H, Ar), 7.56–7.59 (m, 2H, Ar), 7.77 (d,
J = 8.3 Hz, 2H, Ar), 8.03 (d, J = 8.3 Hz, 2H, Ar).
4.7. General procedure for compounds 9d,e
A mixture of 8d, e (297 mg, 0.7 mmol), acetic acid (2 mL), 12 M
HCl (2 mL), and water (2 mL) was refluxed for 16 h. The mixture of
acids was removed in vacuo; to the solid residue water (10 mL)
was added and evaporated again. The raw product was refluxed
with water (10 mL) for 5 h. After cooling the mixture was evapo-
rated under reduced pressure and purified by reverse-phase flash
column chromatography using R-18 Silica Gel and a gradient mo-
bile phase consisting of 15 mM aqueous acetic acid and methanol.
4.4.4. Diethyl 2-acetamido-2-((30-(cyanomethyl)biphenyl-3-yl)-
methyl)malonate (8d)
Yield 355 mg, 84%; MS: m/z 422 [M+]; 1H NMR (300 MHz,
CDCl3): d 1.31 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.05 (s, 3H, NHCOCH3),
3.74 (s, 2H, ArCH2), 3.82 (s, 2H, CH2CN), 4.30 (dq, J1 = 7.2 Hz,
J2 = 1.7 Hz, 4H, OCH2CH3), 6.58 (s, 1H, NH), 7.02 (d, J = 7.7 Hz, 1H,
Ar), 7.21 (s, 1H, Ar), 7.28–7.32 (m, 1H, Ar), 7.35 (d, J = 7.7 Hz, 1H,
Ar), 7.41–7.48 (m, 4H, Ar); 13C NMR (75 MHz, CDCl3): d 14.3,
23.4, 23.9, 38.1, 63.0, 67.4, 118.9, 126.2, 126.6, 126.8, 126.9,
128.8, 128.9, 129.3, 129.7, 130.5, 136.0, 140.4, 142.0, 167.5, 169.2.
4.7.1. (RS)-2-Amino-3-(30-(carboxymethyl)biphenyl-3-yl)prop-
anoic acid (9d)
Yield 191 mg, 91%; Anal. Calcd for C17H17NO4 0.7H2O: C, 65.46;
4.4.5. Diethyl 2-acetamido-2-((40-(cyanomethyl)biphenyl-3-yl)-
methyl)malonate (8e)
H, 5.95; N, 4.49. Found: C, 65.35; H, 5.77; N, 4.38; 1H NMR
(300 MHz, DMSO-d6 + 50 lL D2O): d 2.96 (dd, J1 = 14.3 Hz,
Yield 329 mg, 78%; MS: m/z 422 [M+]; 1H NMR (300 MHz, CDCl3):
d 1.29 (t, J = 7.1 Hz, 6H, OCH2CH3), 2.02 (s, 3H, NHCOCH3), 3.72 (s, 2H,
ArCH2), 3.79 (s, 2H, CH2CN), 4.28 (dq, J1 = 7.1 Hz, J2 = 2.1 Hz, 4H,
OCH2CH3), 6.57 (s, 1H, NH), 7.01 (d, J = 7.6 Hz, 1H, Ar), 7.20 (s, 1H,
Ar), 7.32 (d, J = 7.6 Hz, 1H, Ar), 7.36–7.40 (m, 2H, Ar), 7.45 (d,
J = 7.6 Hz, 1H, Ar), 7.52 (d, J = 8.2 Hz, 2H, Ar); 13C NMR (75 MHz,
CDCl3): d 14.5, 23.5, 23.7, 38.2, 63.1, 67.5, 119.0, 126.2, 127.9,
128.6, 128.9, 129.1, 129.2, 129.3, 136.2, 140.4, 141.0, 167.7, 169.3.
J2 = 8.0 Hz, 1H, ArCH2), 3.21 (dd, J1 = 14.3 Hz, J2 = 4.4 Hz, 1H,
ArCH2), 3.60 (s, 2H, CH2COOH), 3.62–3.67 (m, 1H, ArCH2CH),
7.19–7.23 (m, 2H, Ar), 7.35–7.40 (m, 2H, Ar), 7.46–7.51 (m, 4H, Ar).
4.7.2. (RS)-2-Amino-3-(40-(carboxymethyl)biphenyl-3-yl)prop-
anoic acid (9e)
Yield 186 mg, 88%; Anal. Calcd for C17H17NO4 H2O: C, 64.34; H,
6.03; N, 4.41. Found: C, 64.35; H, 5.72; N, 4.36; 1H NMR (300 MHz,
DMSO-d6 + 50 lL D2O): d 2.99 (dd, J1 = 14.3 Hz, J2 = 7.7 Hz, 1H,
4.5. (RS)-30-(2-Amino-2-carboxyethyl)biphenyl-2-carboxylic
acid (9a)
ArCH2), 3.18 (dd, J1 = 14.3 Hz, J2 = 4.7 Hz, 1H, ArCH2), 3.57 (s, 2H,
CH2COOH), 3.71–3.75 (m, 1H, ArCH2CH), 7.20 (d, J = 7.4 Hz, 1H,
Ar), 7.29 (d, J = 8.0 Hz, 2H, Ar), 7.37 (t, J = 7.7 Hz, 1H, Ar), 7.48–
7.59 (m, 2H, Ar), 7.56 (d, J = 8.0 Hz, 2H, Ar).
A mixture of 8a (102 mg, 0.25 mmol), acetic acid (2 mL), 12 M
HCl (2 mL), and water (2 mL) was refluxed for 20 h. The reaction
mixture was evaporated and purified by HPLC followed by recrystal-
lization from water to give 9a as white crystals. Yield 34 mg, 48%;
Anal. Calcd for C16H15NO4ꢂH2O: C, 63.36; H, 5.65; N, 4.62. Found: C,
4.8. 2-Chloro-4-methyl-6-nitroaniline (11)
Compound 1126 was prepared according to the literature proce-
dure.27 While stirring, 2-chloro-4-methylaniline 10 (14.16 g,
100 mmol) was slowly added to acetic anhydride (110 mL,
1.16 mol) and cooled to 5–7 °C. To this mixture 96% nitric acid
(9.63 mL, 0.22 mol) was added dropwise over a period of 1 h, keep-
ing the temperature below 15 °C. The reaction mixture was stirred
for additional 10 min, and then poured into ice-water (400 mL).
The resulting yellowish solid was filtered; the wet product was
treated with 12 M HCl (20 mL) and EtOH (40 mL). The mixture
was refluxed for 4 h. Upon cooling ethanol was removed by evap-
oration, and the residue was poured into the mixture of ammonia
(40 mL) and water (200 mL). The orange solid was filtered, washed
with water (3 ꢁ 100 mL) and dried to afford 11, which was directly
used in next step. Yield 10.82 g, 58%; MS: m/z 186 [M+]; 1H NMR
(300 MHz, CDCl3): d 2.27 (s, 3H, CH3), 6.41 (br s, 2H, NH2), 7.38
(s, 1H, Ar), 7.90 (s, 1H, Ar); 13C NMR (75 MHz, CDCl3): d 20.4,
121.9, 124.9, 125.9, 127.4, 136.7, 139.6.
63.33; H, 5.39; N, 4.45; 1H NMR (300 MHz, DMSO-d6 + 50
lL D2O):
d
2.91 (dd, J1 = 14.0 Hz, J2 = 8.5 Hz, 1H, ArCH2), 3.18 (dd,
J1 = 14.3 Hz, J2 = 3.9 Hz, 1H, ArCH2), 3.53–3.57 (m, 1H, ArCH2CH),
7.14–7.26 (m, 3H, Ar), 7.30 (t, J = 7.7 Hz, 1H, Ar), 7.38–7.43 (m, 2H,
Ar), 7.53 (t, J = 7.4 Hz, 1H, Ar), 7.64 (d, J = 8.3 Hz, 1H, Ar).
4.6. General procedure for compounds 9b, c
Compounds 8b, c (204 mg, 0.5 mmol) were deprotected using
the procedure described for 9a. The reaction mixture was evapo-
rated and purified by HPLC using a gradient mobile phase consist-
ing of aqueous trifluoroacetic acid and acetonitrile.
4.6.1. (RS)-30-(2-Amino-2-carboxyethyl)biphenyl-3-carboxylic
acid (9b)
Yield 45 mg, 32%; Anal. Calcd for C16H15NO4ꢂ0.3
C2HF3O2ꢂ0.4H2O: C, 61.03; H, 4.97; N, 4.29. Found: C, 61.20; H,

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E. Szymanska et al. / Bioorg. Med. Chem. 17 (2009) 6390–6401
4.9. 1,2-Dichloro-5-methyl-3-nitrobenzene (12)
reduced pressure. Purification with column chromatography (hep-
tane) afforded a white solid, the mixture of 15 and dibromo deriv-
ative in ratio 2.5:1, respectively. Yield of 15, calculated on the basis
of the proton NMR spectra, 6.06 g, 64%; MS: m/z 364 [M+]; 1H NMR
(300 MHz, CDCl3): d 4.33 (s, 2H, CH2Br), 7.48 (d, J = 2.1 Hz, 1H, Ar),
7.70 (d, 1H, J = 2.1 Hz, Ar); 13C NMR (75 MHz, CDCl3): d 30.2, 99.3,
128.7, 130.9, 132.5, 136.2, 138.8.
Compound 1228 was prepared according to the literature proce-
dure.29 To a mixture of tert-butyl nitrite (7.75 g, 75 mmol) and
anhydrous copper(II) chloride (8.0 g, 60 mmol) in dry acetonitrile
(40 mL) at 65 °C, a solution of 11 (9.30 g, 50 mmol) in dry acetoni-
trile (60 mL) was added dropwise under vigorous stirring during
30 min. Then stirring was continued for 1 h. Upon cooling, the mix-
ture was poured into 20% hydrochloric acid (200 mL) and extracted
with diethyl ether (3 ꢁ 120 mL). The combined organic phases
were dried (MgSO4) and evaporated. Based on 1H NMR and GC–
MS analyses, the resulting yellow crystals proved to be 12, and
could be used in the following step without further purification.
Yield 8.85 g, 89%; MS: m/z 205 [M+]; 1H NMR (300 MHz, CDCl3):
d 2.57 (s, 3H, CH3), 7.67 (s, 2H, Ar); 13C NMR (75 MHz, CDCl3): d
20.9, 123.9, 134.2, 134.9, 135.0, 138.8, 149.2.
4.13. Diethyl 2-acetamido-2-(3,4-dichloro-5-iodobenzyl)-
malonate (16)
Compound 16 was prepared from 15 (5.49 g, 15 mmol) by the
method described for compound 7. After purification by flash col-
umn chromatography compound 16 was isolated as a white solid.
Yield 5.88 g, 78%; MS: m/z 501 [M+]; 1H NMR (300 MHz, CDCl3): d
1.32 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.08 (s, 3H, NHCOCH3), 3.57 (s, 2H,
Ar), 4.28 (q, J = 7.2 Hz, 4H, OCH2CH3), 6.57 (s, 1H, NH), 7.08 (d,
J = 1.8 Hz, 1H, Ar), 7.43 (d, J = 1.8 Hz,1H, Ar); 13C NMR (75 MHz,
CDCl3): d 14.3, 23.2, 36.5, 63.3, 67.0, 99.1, 131.7, 132.1, 135.7,
136.7, 139.9, 166.9, 169.4.
4.10. 2,3-Dichloro-5-methylaniline (13)
Compound 1328 was prepared according to the literature proce-
dure.30 Iron powder (48 g, 86 mmol), water (80 mL) and 12 M
hydrochloric acid (1.57 mL, 19 mmol) were added consecutively
to a stirred solution of 12 (8.24 g, 40 mmol) in ethanol (310 mL).
The reaction mixture was refluxed for 90 min, filtered hot and eth-
anol was removed under reduced pressure. The residue was
washed with dichloromethane (80 mL). To the aqueous phase
2 M NaOH (50 mL) was added, then the solution was extracted
with dichloromethane (3 ꢁ 80 mL), the combined organic extracts
were dried (MgSO4), and the solvent was removed in vacuo. Ob-
tained brown oil without further purification could be used in
the next step. Yield 7.04 g, 86%; MS: m/z 175 [M+]; 1H NMR
(300 MHz, CDCl3): d 2.21 (s, 3H, CH3), 4.08 (br s, 2H, NH2), 6.47
(s, 1H, Ar), 6.67 (s, 1H, Ar); 13C NMR (75 MHz, CDCl3): d 21.1,
114.4, 114.5, 120.6, 132.6, 138.0, 144.3.
4.14. General procedure for compounds 17a–h
Compounds 17a–h were prepared from 16 (502 mg, 1 mmol)
and the relevant substituted phenylboronic acids according to
the procedure described for compounds 8a–e.
4.14.1. Diethyl 2-acetamido-2-((5,6-dichloro-20-cyanobiphenyl-
3-yl)methyl)malonate (17a)
Yield 0.234 g, 49%; MS: m/z 476 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.29 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.08 (s, 3H, NHCOCH3),
3.65 (s, 2H, ArCH2), 4.26 (q, J = 7.2 Hz, 4H, OCH2CH3), 6.89 (s, 1H,
NH), 6.93 (d, J = 1.9 Hz, 1H, Ar), 7.22 (d, J = 1.9 Hz, 1H, Ar), 7.40
(dt, J1 = 7.7 Hz, J2 = 1.4 Hz, 1H, Ar), 7.51 (dt, J1 = 7.7 Hz, J2 = 1.4 Hz,
1H, Ar), 7.65 (dt, J1 = 7.7 Hz, J2 = 1.4 Hz, 1H, Ar), 7.76 (dt,
J1 = 7.7 Hz, J2 = 1.4 Hz, 1H, Ar); 13C NMR (75 MHz, CDCl3): d 14.4,
23.3, 37.2, 63.2, 67.2, 112.5, 117.7, 128.7, 130.4, 130.7, 131.0,
132.1, 132.5, 132.9, 133.6, 135.4, 138.6, 142.5, 166.9, 169.6.
4.11. 1,2-Dichloro-3-iodo-5-methylbenzene (14)
Compound 14 was prepared according to the known literature
procedure.27 The suspension of 13 (5.28 g, 30 mmol) in glacial ace-
tic acid (30 mL) was heated to form a clear solution, then cooled to
25 °C. A pre-cooled solution of sodium nitrite (3.0 g, 48 mmol) in
cold concentrated sulfuric acid (16.5 mL) was slowly added over
a period of 30 min, stirred for another 30 min, then heated to
75 °C, stirred for 1 h and cooled to 25 °C. The mixture was poured
into ice-water (130 mL) while stirring and successively treated
with of urea (3.0 g, 50 mmol) and aqueous KI solution (7.47 g,
45 mmol, in 35 mL of water). After KI addition was completed, so-
lid NaHSO3 was added until the color of the mixture obviously
changed. The reaction mixture was extracted with dichlorometh-
ane (3 ꢁ 120 mL), the organic phases were dried over MgSO4, and
the solvent was removed under reduced pressure. Purification by
flash column chromatography (heptane) afforded 14 as a white
low melting solid. Yield 8.61 g, 54%; MS: m/z 286 [M+]; 1H NMR
(300 MHz, CDCl3): d 2.27 (s, 3H, CH3), 7.25 (s, 1H, Ar), 7.59 (s, 1H,
Ar); 13C NMR (75 MHz, CDCl3): d 20.4, 99.1, 131.1, 131.7, 133.5,
139.1, 139.2.
4.14.2. Diethyl 2-acetamido-2-((5,6-dichloro-30-cyanobiphenyl-
3-yl)methyl)malonate (17b)
Yield 0.343 g, 72%; MS: m/z 476 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.28 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.03 (s, 3H, NHCOCH3),
3.66 (s, 2H, ArCH2), 4.26 (dq, J1 = 7.2 Hz, J2 = 1.7 Hz, 4H, OCH2CH3),
6.58 (s, 1H, NH), 6.81 (d, J = 1.9 Hz, 1H, Ar), 7.16 (d, J = 1.9 Hz, 1H,
Ar), 7.53–7.58 (m, 2H, Ar), 7.61 (s, 1H, Ar), 7.66–7.69 (m, 1H, Ar);
13C NMR (75 MHz, DMSO-d6): d 13.9, 22.1, 36.8, 62.0, 66.7, 111.5,
118.3, 128.3, 129.6, 131.4, 131.8, 132.0, 132.6, 133.9, 135.9,
139.3, 139.4, 166.6, 169.3.
4.14.3. Diethyl 2-acetamido-2-((5,6-dichloro-40-cyanobiphenyl-
3-yl)methyl)malonate (17c)
Yield 0.329 g, 69%; MS: m/z 476 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.28 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.02 (s, 3H, NHCOCH3),
3.66 (s, 2H, ArCH2), 4.22–4.28 (m, 4H, OCH2CH3), 6.58 (s, 1H,
NH), 6.81 (d, J = 2.1 Hz, 1H, Ar), 7.16 (d, J = 2.1 Hz, 1H, Ar), 7.44
(d, J = 8.5 Hz, 2H, Ar), 7.71 (d, J = 8.5 Hz, 2H, Ar); 13C NMR
(75 MHz, CDCl3): d 14.2, 23.2, 37.0, 63.1, 67.0, 112.1, 118.5,
129.7, 129.9, 130.5, 131.7, 132.1, 133.7, 135.4, 140.5, 143.4,
167.0, 169.3.
4.12. 5-(Bromomethyl)-1,2-dichloro-3-iodobenzene (15)
Compound 15 was prepared according to the modified litera-
ture method described for 6. A mixture of 14 (7.47 g, 26 mmol),
NBS (6.946 g, 38 mmol) and benzoyl peroxide (0.945 g, 3.9 mmol)
in tetrachloromethane (150 mL) was refluxed for 10 h. The pro-
gress of the reaction was controlled by 1H NMR. After cooling
and filtration, dichloromethane (80 mL) was added and the liquid
was washed with 2 M NaOH, dried (MgSO4) and evaporated under
4.14.4. Diethyl 2-acetamido-2-((5,6-dichloro-20-methoxybi-
phenyl-3-yl)methyl)malonate (17d)
Yield 0.299 g, 62%; MS: m/z 481 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.27 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.02 (s, 3H, NHCOCH3),
3.62 (s, 2H, ArCH2), 3.76 (s, 3H, OCH3), 4.25 (dq, J1 = 7.2 Hz,

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6398
J2 = 1.4 Hz, 4H, OCH2CH3), 6.59 (s, 1H, NH), 6.81 (d, J = 2.2 Hz, 1H,
Ar), 6.93–7.08 (m, 4H, Ar), 7.36 (dt, J1 = 7.7 Hz, J2 = 1.9 Hz, 1H,
Ar); 13C NMR (75 MHz, CDCl3): d 14.3, 23.2, 37.1, 55.7, 63.1, 67.1,
111.1, 120.4, 128.1, 129.9, 130.5, 130.6, 131.4, 132.8, 134.5,
136.8, 139.8, 156.4, 167.2, 169.2.
4.15.2. (RS)-50-(2-Amino-2-carboxyethyl)-20,30-dichlorobiphe-
nyl-3-carboxylic acid (18b)
Yield 129 mg, 73%; Anal. Calcd for C16H13Cl2NO4 1.75H2O: C,
49.82; H, 4.31; N, 3.63. Found: C, 49.53; H, 3.94; N, 3.64; 1H
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 2.99 (dd, J1 = 14.3 Hz,
J2 = 7.4 Hz, 1H, ArCH2), 3.14 (dd, J1 = 14.3 Hz, J2 = 4.4 Hz, 1H,
ArCH2), 3.75–3.79 (m, 1H, ArCH2CH), 7.23 (d, J = 1.9 Hz, 1H, Ar),
7.52 (d, J = 1.9 Hz, 1H, Ar), 7.59 (t, J = 7.4 Hz, 1H, Ar), 7.99 (d,
J = 7.7 Hz, 1H, Ar), 7.95–7.98 (m, 2H, Ar).
4.14.5. Diethyl 2-acetamido-2-((5,6-dichloro-30-methoxy-
biphenyl-3-yl)methyl)malonate (17e)
Yield 0.357 g, 74%; MS: m/z 481 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.28 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.03 (s, 3H, NHCOCH3),
3.63 (s, 2H, ArCH2), 3.82 (s, 3H, OCH3), 4.26 (dq, J1 = 7.2 Hz,
J2 = 1.4 Hz, 4H, OCH2CH3), 6.58 (s, 1H, NH), 6.85–6.93 (m, 3H, Ar),
7.10 (d, J = 1.9 Hz, 1H, Ar), 7.24 (s, 1H, Ar), 7.32 (t, J = 7.7 Hz, 1H,
Ar); 13C NMR (75 MHz, CDCl3): d 14.3, 23.3, 37.1, 55.5, 63.1, 67.1,
113.6, 114.9, 121.5, 129.3, 130.1, 130.7, 130.9, 133.5, 134.9,
140.3, 142.4, 159.2, 167.2, 169.3.
4.15.3. (RS)-50-(2-Amino-2-carboxyethyl)-20,30-dichlorobiphe-
nyl-4-carboxylic acid (18c)
Yield 140 mg, 79%; Anal. Calcd for C16H13Cl2NO4ꢂ0.5H2O: C,
52.91; H, 3.89; N, 3.86. Found: C, 53.07; H, 3.66; N, 3.71; 1H
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 2.96 (dd, J1 = 14.4 Hz,
J2 = 7.7 Hz, 1H, ArCH2), 3.14 (dd, J1 = 14.4 Hz, J2 = 4.5 Hz, 1H,
ArCH2), 3.52–3.56 (m, 1H, ArCH2CH), 7.25 (d, J = 1.9 Hz, 1H, Ar),
7.54 (d, J = 8.5 Hz, 2H, Ar), 7.57 (d, J = 1.9 Hz, 1H, Ar), 7.99 (d,
J = 8.5 Hz, 2H, Ar).
4.14.6. Diethyl 2-acetamido-2-((5,6-dichloro-40-methoxy-
biphenyl-3-yl)methyl)malonate (17f)
Yield 0.328 g, 68%; MS: m/z 481 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.28 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.03 (s, 3H, NHCOCH3),
3.63 (s, 2H, ArCH2), 3.85 (s, 3H, OCH3), 4.26 (dq, J1 = 7.2 Hz,
J2 = 1.4 Hz, 4H, OCH2CH3), 6.57 (s, 1H, NH), 6.83 (d, J = 2.2 Hz, 1H,
Ar), 6.93 (d, J = 8.8 Hz, 2H, Ar), 7.07 (d, J = 2.2 Hz, 1H, Ar), 7.25 (d,
J = 8.8 Hz, 2H, Ar); 13C NMR (75 MHz, CDCl3): d 14.3, 23.3, 37.1,
55.5, 63.1, 67.1, 113.6, 130.2, 130.3, 130.5, 131.1, 131.4, 133.3,
134.8, 142.2, 159.3, 167.2, 169.3.
4.15.4. (RS)-2-Amino-3-(5,6-dichloro-20-hydroxybiphenyl-3-
yl)propanoic acid (18d)
Yield 108 mg, 67%; Anal. Calcd for C15H13Cl2NO3ꢂ0.5H2O: C,
53.75; H, 4.21; N, 4.18. Found: C, 53.72; H, 3.88; N, 4.04; 1H
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 2.91 (dd, J1 = 14.3 Hz,
J2 = 8.0 Hz, 1H, ArCH2), 3.12 (dd, J1 = 14.3 Hz, J2 = 5.5 Hz, 1H,
ArCH2), 3.47–3.50 (m, 1H, ArCH2CH), 6.82–6.89 (m, 2H, Ar), 7.07
(d, J = 6.6 Hz, 1H, Ar), 7.11 (s, 1H, Ar), 7.20 (dt, J1 = 7.7 Hz,
J2 = 1.7 Hz, 1H, Ar), 7.46 (s, 1H, Ar).
4.14.7. Diethyl 2-acetamido-2-((5,6-dichloro-30-nitrobiphenyl-
3-yl)methyl)malonate (17g)
Yield 0.258 g, 52%; MS: m/z 496 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.30 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.06 (s, 3H, NHCOCH3),
3.68 (s, 2H, ArCH2), 4.26 (dq, J1 = 7.2 Hz, J2 = 1.9 Hz, 4H, OCH2CH3),
6.62 (s, 1H, NH), 6.88 (d, J = 2.2 Hz, 1H, Ar), 7.19 (d, J = 2.2 Hz, 1H,
Ar), 7.61 (t, J = 8.0 Hz, 1H, Ar), 7.72 (dt, J1 = 7.7 Hz, J2 = 1.7 Hz, 1H,
Ar), 8.19 (t, J = 1.7 Hz, 1H, Ar), 8.24–8.28 (m, 1H, Ar); 13C NMR
(75 MHz, CDCl3): d 14.3, 23.3, 37.1, 63.3, 67.1, 123.1, 124.2, 129.3,
130.0, 131.8, 133.9, 135.4, 135.6, 139.9, 140.4, 148.1, 167.1, 169.4.
4.15.5. (RS)-2-Amino-3-(5,6-dichloro-30-hydroxybiphenyl-3-yl)-
propanoic acid (18e)
Yield 143 mg, 88%; Anal. Calcd for C15H13Cl2NO3ꢂ0.25H2O: C,
54.48; H, 4.11; N, 4.24. Found: C, 54.27; H, 3.89; N, 4.12; 1H
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 2.92 (dd, J1 = 14.0 Hz,
J2 = 7.7 Hz, 1H, ArCH2), 3.10 (dd, J1 = 14.0 Hz, J2 = 4.1 Hz, 1H,
ArCH2), 3.50–3.54 (m, 1H, ArCH2CH), 6.77–6.82 (m, 3H, Ar), 7.18
(d, J = 1.9 Hz, 1H, Ar), 7.23 (t, J = 8.8 Hz, 1H, Ar), 7.47 (d,
J = 1.9 Hz, 1H, Ar).
4.14.8. Diethyl 2-acetamido-2-((5,6-dichloro-40-nitrobiphenyl-
3-yl)methyl)malonate (17h)
4.15.6. (RS)-2-Amino-3-(5,6-dichloro-40-hydroxybiphenyl-3-yl)-
propanoic acid (18f)
Yield 0.278 g, 56%; MS: m/z 496 [M+ꢀ14]; 1H NMR (300 MHz,
CDCl3): d 1.28 (t, J = 7.2 Hz, 6H, OCH2CH3), 2.03 (s, 3H, NHCOCH3),
3.67 (s, 2H, ArCH2), 4.23–4.32 (m, 4H, OCH2CH3), 6.59 (s, 1H,
NH), 6.84 (d, J = 1.9 Hz, 1H, Ar), 7.18 (d, J = 1.9 Hz, 1H, Ar), 7.51
(dt, J1 = 8.8 Hz, J2 = 1.9 Hz, 2H, Ar), 8.28 (dt, J1 = 8.8 Hz, J2 = 1.9 Hz,
2H, Ar); 13C NMR (75 MHz, CDCl3): d 14.3, 23.3, 37.1, 63.2, 67.1,
123.6, 129.8, 130.2, 130.5, 131.9, 133.9, 135.5, 140.2, 145.3,
147.6, 167.1, 169.3.
Yield 127 mg, 78%; Anal. Calcd for C15H13Cl2NO3ꢂH2O: C, 52.34;
H, 4.39; N, 4.07. Found: C, 52.08; H, 4.13; N, 3.93; 1H NMR
(300 MHz, DMSO-d6 + 50 lL D2O): d 2.93 (dd, J1 = 14.3 Hz,
J2 = 7.7 Hz, 1H, ArCH2), 3.10 (dd, J1 = 14.3 Hz, J2 = 4.1 Hz, 1H,
ArCH2), 3.56–3.60 (m, 1H, ArCH2CH), 6.81 (d, J = 8.5 Hz, 2H, Ar),
7.17 (d, J = 1.9 Hz, 1H, Ar), 7.23 (d, J = 8.5 Hz, 2H, Ar), 7.40 (d,
J = 1.9 Hz, 1H, Ar).
4.15. General procedure for compounds 18a–h
4.15.7. (RS)-2-Amino-3-(5,6-dichloro-30-nitrobiphenyl-3-yl)-
propanoic acid (18g)
A mixture of 17a–h (0.5 mmol) in 48% aq HBr (6 mL) was re-
fluxed for 2 h. The reaction mixture was evaporated under reduced
pressure and purified by reverse-phase HPLC (mobile phase: a mix-
ture of methanol in 15 mM acetic acid) to give white crystals of the
target amino acid.
Yield 130 mg, 73%; Anal. Calcd for C15H12Cl2N2O4ꢂ1.2H2O: C,
47.82; H, 3.85; N, 7.43. Found: C, 48.05; H, 3.70; N, 7.14; 1H
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 2.95 (dd, J1 = 14.3 Hz,
J2 = 8.0 Hz, 1H, ArCH2), 3.14 (dd, J1 = 14.3 Hz, J2 = 4.7 Hz, 1H,
ArCH2), 3.52–3.56 (m, 1H, ArCH2CH), 7.30 (d, J = 1.9 Hz, 1H, Ar),
7.57 (d, J = 1.9 Hz, 1H, Ar), 7.75 (t, J = 8.8 Hz, 1H, Ar), 7.89 (d,
J = 8.0 Hz, 1H, Ar), 8.23–8.27 (m, 2H, Ar).
4.15.1. (RS)-50-(2-Amino-2-carboxyethyl)-20,30-
dichlorobiphenyl-2-carboxylic acid (18a)
Yield 92 mg, 52%; Anal. Calcd for C16H13Cl2NO4 2.25H2O: C,
4.15.8. (RS)-2-Amino-3-(5,6-dichloro-40-nitrobiphenyl-3-yl)-
propanoic acid (18h)
48.69; H, 4.47; N, 3.55. Found: C, 48.78; H, 4.61; N, 3.45; 1H
NMR (300 MHz, DMSO-d6 + 50
l
L D2O): d 2.91–3.00 (m, 1H,
Yield 138 mg, 78%; Anal. Calcd for C15H12Cl2N2O4ꢂ0.5H2O: C,
ArCH2), 3.06–3.18 (m, 1H, ArCH2), 3.59–3.63 (m, 1H, ArCH2CH),
7.10 (d, J = 1.9 Hz, 1H, Ar), 7.24–7.31 (m, 1H, Ar), 7.43–7.53 (m,
2H, Ar), 7.62 (t, J = 8.5 Hz, 1H, Ar), 7.89 (d, J = 8.5 Hz, 1H, Ar).
49.47; H, 3.60; N, 7.69. Found: C, 49.19; H, 3.24; N, 7.44; 1H NMR
(300 MHz, DMSO-d6 + 50
lL
D2O):
d
2.96 (dd, J1 = 14.3 Hz,
J2 = 7.4 Hz, 1H, ArCH2), 3.10 (dd, J1 = 14.3 Hz, J2 = 5.0 Hz, 1H, ArCH2),

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E. Szymanska et al. / Bioorg. Med. Chem. 17 (2009) 6390–6401
3.52–3.56 (m, 1H, ArCH2CH), 7.28 (d, J = 1.9 Hz, 1H, Ar), 7.57 (d,
J = 1.9 Hz, 1H, Ar), 7.7 (d, J = 8.5 Hz, 2H, Ar), 8.28 (d, J = 8.5 Hz, 2H, Ar).
NMR (300 MHz, DMSO-d6 + 50
l
L D2O): d 2.93 (dd, J1 = 14.3 Hz,
J2 = 7.7 Hz, 1H, ArCH2), 3.10 (dd, J1 = 14.3 Hz, J2 = 4.7 Hz, 1H,
ArCH2), 3.89–3.97 (m, 1H, ArCH2CH), 7.32 (d, J = 1.7 Hz, 1H, Ar),
7.48–7.51 (m, 2H, Ar), 7.58 (t, J = 7.4 Hz, 1H, Ar), 8.05 (d,
J = 7.7 Hz, 1H, Ar), 8.11 (s, 1H, Ar).
4.16. General procedure for compounds 19a–c
Compounds 19a–c were prepared according to the known liter-
ature procedure.31 A suspension of 17a–c (239 mg, 0.5 mmol),
NaN3 (65 mg, 1 mmol) and TEA hydrochloride (138 mg, 1 mmol)
in 1,2-dimethoxyethane (5 mL) was boiled under reflux for 48 h.
The mixture was cooled and poured into water (4 mL). pH of the
aqueous phase was adjusted to ca. 1 by addition of 37% HCl (aq)
and the aqueous phase was extracted with diethyl ether. The or-
ganic extracts were dried (MgSO4) and the solvent was removed
in vacuo. Flash column chromatography gave crystals of the tetraz-
olyl derivatives 19a–c.
4.17.3. (RS)-2-Amino-3-(5,6-dichloro-40-(1H-tetrazol-5-yl)bi-
phenyl-3-yl)propanoic acid (20c)
Yield 93 mg, 82%; Anal. Calcd for C16H13Cl2N5O2ꢂ0.65H2O: C,
49.29; H, 3.70; N, 17.96. Found: C, 49.48; H, 3.87; N, 17.69; 1H
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 3.01 (dd, J1 = 14.3 Hz,
J2 = 7.7 Hz, 1H, ArCH2), 3.15 (dd, J1 = 14.3 Hz, J2 = 4.7 Hz, 1H,
ArCH2), 3.78–3.82 (m, 1H, ArCH2CH), 7.27 (d, J = 1.9 Hz, 1H, Ar),
7.51 (d, J = 1.9 Hz, 1H, Ar), 7.56 (d, J = 8.3 Hz, 2H, Ar), 8.06 (d,
J = 8.3 Hz, 2H, Ar).
4.16.1. Diethyl 2-acetamido-2-((5,6-dichloro-20-(1H-tetrazol-5-
yl)biphenyl-3-yl)methyl)malonate (19a)
4.18. Diethyl 2-acetamido-2-(3,4-dichloro-5-(5-cyanothiophen-
2-yl)benzyl)malonate (21)
Yield 159 mg, 61%; 1H NMR (300 MHz, DMSO-d6): d 1.11–1.22
(m, 6H, OCH2CH3), 1.99 (s, 3H, NHCOCH3), 3.44 (s, 2H, ArCH2),
4.14 (q, J = 7.1 Hz, 4H, OCH2CH3), 6.86 (s, 1H, Ar), 7.21 (s, 1H, Ar),
7.42 (d, J = 7.6 Hz, 1H, Ar), 7.66–7.74 (m, 2H, Ar), 7.89 (d,
J = 6.5 Hz, 1H, Ar), 8.24 (s, 1H, NH); 13C NMR (75 MHz, DMSO-d6):
d 13.9, 22.1, 36.8, 62.1, 66.7, 121.8, 128.3, 128.8, 129.0, 129.2,
130.8, 130.9, 131.7, 135.3, 137.9, 140.6, 144.1, 155.2, 166.6, 169.3.
Compound 21 was prepared in analogy to the literature proce-
dure.32 Compound 16 (602 mg, 1.2 mmol), 5-cyanothiophene-2-
boronic acid (275 mg, 1.8 mmol), Pd(PPh3)4 (70 mg, 0.06 mmol)
and potassium carbonate (498 mg, 3.6 mmol) were suspended in
the mixture of toluene (15 mL), ethanol (4.5 mL) and water
(4.5 mL). The mixture was stirred under nitrogen atmosphere for
2 days at room temperature and then for next 3 days at 50 °C.
The reaction mixture was then partitioned into water (50 mL)
and ethyl acetate (50 mL). The organic phase was dried (MgSO4),
filtered and concentrated under reduced pressure. The crude mate-
rial was purified by flash column chromatography to give a yellow-
ish oil 21. Yield 140 mg, 24%; MS: m/z 482 [M+]; 1H NMR (300 MHz,
CDCl3): d 1.29 (t, J = 7.1 Hz, 6H, OCH2CH3), 2.04 (s, 3H, NHCOCH3),
3.61 (s, 2H, ArCH2), 4.29 (q, J = 7.1 Hz, 4H, OCH2CH3), 6.60 (s, 1H,
NH), 7.02 (d, J = 2.1 Hz, 1H, Ar), 7.18 (d, J = 2.1 Hz, 1H, Ar), 7.27
(d, J = 3.8 Hz, 1H, thiophene), 7.61 (d, J = 3.8 Hz, 1H, thiophene);
13C NMR (75 MHz, CDCl3): d 14.3, 23.3, 37.0, 63.3, 67.1, 110.6,
113.9, 129.2, 130.3, 131.1, 131.8, 132.4, 132.8, 134.5, 135.7,
146.5, 167.0, 169.4.
4.16.2. Diethyl 2-acetamido-2-((5,6-dichloro-30-(1H-tetrazol-5-
yl)biphenyl-3-yl)methyl)malonate (19b)
Yield 196 mg, 75%; 1H NMR (300 MHz, DMSO-d6): d 1.14 (t,
J = 7.1 Hz, 6H, OCH2CH3), 1.9 (s, 3H, NHCOCH3), 3.47 (s, 2H, ArCH2),
4.15 (q, J = 7.1 Hz, 4H, OCH2CH3), 7.00 (s, 1H, Ar), 7.26 (s, 1H, Ar),
7.62 (d, J = 7.6 Hz, 1H, Ar), 7.73 (t, J = 7.6 Hz, 1H, Ar), 8.07–8.10
(m, 2H, Ar), 8.35 (s, 1H, NH); 13C NMR (75 MHz, DMSO-d6): d
13.9, 22.1, 36.9, 62.0, 66.7, 124.2, 126.6, 127.3, 128.3, 129.5,
129.8, 131.2, 131.7, 131.9, 135.9, 139.3, 140.4, 154.8, 166.6, 169.4.
4.16.3. Diethyl 2-acetamido-2-((5,6-dichloro-40-(1H-tetrazol-5-
yl)biphenyl-3-yl)methyl)malonate (19c)
Yield 207 mg, 80%; 1H NMR (300 MHz, CDCl3): d 1.27 (t,
J = 7.1 Hz, 6H, OCH2CH3), 2.11 (s, 3H, NHCOCH3), 3.22 (br s, 1H,
NH tetrazole), 3.67 (s, 2H, ArCH2), 4.28 (q, J = 7.1 Hz, 4H, OCH2CH3),
6.86 (s, 1H, NH), 6.88 (d, J = 2.1 Hz, 1H, Ar), 7.14 (d, J = 2.1 Hz, 1H,
Ar), 7.49 (d, J = 8.2 Hz, 2H, Ar), 8.18 (d, J = 8.2 Hz, 2H, Ar); 13C NMR
(75 MHz, CDCl3): d 14.3, 23.3, 37.2, 63.4, 67.2, 123.9, 127.4, 130.1,
130.8, 131.1, 132.1, 133.7, 134.9, 141.4, 141.8, 156.3, 167.0, 170.6.
4.19. (RS)-5-(5-(2-Amino-2-carboxyethyl)-2,3-dichloro-
phenyl)thiophene-2-carboxylic acid (22)
Compound 21 (102 mg, 0.2 mmol) was deprotected and puri-
fied by the method described for compounds 18a–h. Yield 33 mg,
43%; Anal. Calcd for C14H11Cl2NO4Sꢂ2H2O: C, 42.44; H, 3.82; N,
3.53. Found: C, 42.75; H, 3.54; N, 3.55; 1H NMR (300 MHz,
4.17. General procedure for compounds 20a–c
DMSO-d6 + 50 lL D2O): d 2.96 (dd, J1 = 14.8 Hz, J2 = 7.7 Hz, 1H,
The target amino acids 20a–c were obtained from 19a–c
(156 mg, 0.3 mmol) according to the procedure described for com-
pounds 18a–h.
ArCH2), 3.16 (dd, J1 = 14.8 Hz, J2 = 4.7 Hz, 1H, ArCH2), 3.75–3.80
(m, 1H, ArCH2CH), 7.42 (d, J = 3.9 Hz, 1H, thiophene), 7.51 (s, 1H,
Ar), 7.56 (s, 1H, Ar), 7.68 (d, J = 3.9 Hz, 1H, thiophene).
4.17.1. (RS)-2-Amino-3-(5,6-dichloro-20-(1H-tetrazol-5-yl)-
biphenyl-3-yl)propanoic acid (20a)
4.20. Diethyl 2-acetamido-2-(3-(bromomethyl)benzyl)
malonate (24)
Yield 58 mg, 51%; Anal. Calcd for C16H13Cl2N5O2ꢂ1.5H2O: C,
47.42; H, 3.98; N, 17.28. Found: C, 47.13; H, 4.04; N, 17.14; 1H
Compound 24 was prepared from 23 (2.11 g, 8 mmol) by the
method described for the compound 7. Purification by flash col-
umn chromatography afforded white solid of 24 (1.16 g, 36%);
NMR (300 MHz, DMSO-d6 + 50 lL D2O): d 2.96 (dd, J1 = 14.3 Hz,
J2 = 8.0 Hz, 1H, ArCH2), 3.16 (dd, J1 = 14.3 Hz, J2 = 4.4 Hz, 1H,
ArCH2), 3.74–3.80 (m, 1H, ArCH2CH), 7.24 (d, J = 2.2 Hz, 1H, Ar),
7.34–7.40 (m, 2H, Ar), 7.46 (dd, J1 = 7.7 Hz, J2 = 1.9 Hz, 1H, Ar),
7.54–7.61 (m, 3H, Ar), 7.78–7.81 (m, 1H, Ar).
MS: m/z 399 [M+]; 1H NMR (300 MHz, CDCl3):
d 1.23 (t,
J = 7.2 Hz, 6H, OCH2CH3), 1.97 (s, 3H, NHCOCH3), 3.57 (s, 2H,
ArCH2), 4.20 (q, J = 7.2 Hz, 4H, OCH2CH3), 4.35 (s, 2H, CH2Br), 6.43
(s, 1H, NH), 6.84 (dt, J1 = 6.6 Hz, J2 = 1.7 Hz, 1H, Ar), 6.97 (s, 1H,
Ar), 7.13–7.17 (m, 2H, Ar); 13C NMR (75 MHz, CDCl3): d 14.3,
23.3, 33.5, 37.7, 62.9, 67.3, 94.3, 127.8, 128.7, 129.8, 130.9, 136.0,
137.9, 167.3, 169.6.
4.17.2. (RS)-2-Amino-3-(5,6-dichloro-30-(1H-tetrazol-5-yl)-
biphenyl-3-yl)propanoic acid (20b)
Yield 88 mg, 78%; Anal. Calcd for C16H13Cl2N5O2ꢂ1.35H2O: C,
47.74; H, 3.93; N, 17.40. Found: C, 47.65; H, 4.08; N, 17.34; 1H

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6400
4.21. General procedure for compounds 25a–c
J = 7.4 Hz, 1H, Ar), 7.03 (d, J = 7.7 Hz, 1H, Ar), 7.11–7.16 (m, 2H,
Ar), 7.24–7.28 (m, 2H, Ar), 7.41 (dt, J1 = 7.6 Hz, J2 = 1.7 Hz, 1H,
Ar), 7.71 (dd, J1 = 7.7 Hz, J2 = 1.7 Hz, 1H, Ar).
Compounds 25a–c were prepared according to the modified lit-
erature procedure.17 [1,10-bis(diphenylphosphino) ferrocene]dichlo-
ropalladium(II) (complex with dichloromethane) (41 mg,
0.05 mmol) and boronic acid (or boronic acid pinacol ester)
(1.5 mmol) were charged into a 10 mL Schlenk flask. The flask was
evacuated and backfilled with nitrogen before adding THF (5 mL),
3 M NaOH solution (1 mL), tetrabutylammonium bromide (TBAB,
65 mg, 0.2 mmol) and 24 (400 mg, 1 mmol). The mixture was stirred
at 50 °C under nitrogen atmosphere for 3 h, then washed with 1 M
NaOH and the aqueous phase was extracted with diethyl ether
(30 mL). The combined organic layers were washed with brine
(20 mL), dried (MgSO4) and concentrated in vacuo. The crude mate-
rial was purified by flash column chromatography.
4.23. General procedure for compounds 26b, c
The target amino acid 26b, c were obtained from 25b, c
(210 mg, 0.5 mmol) and purified according to the procedure de-
scribed for compounds 18a–h.
4.23.1. (RS)-3-(3-(2-Amino-2-carboxyethyl)benzyl)benzoic acid
(26b)
Yield 98 mg, 66%; Anal. Calcd for C17H17NO4ꢂ0.5H2O: C, 66.22; H,
5.88 N, 4.54. Found: C, 66.13; H, 5.51; N, 4.39; 1H NMR (300 MHz,
DMSO-d6): d 2.87 (dd, J1 = 14.0 Hz, J2 = 7.4 Hz, 1H, ArCH2), 3.09 (dd,
J1 = 14.0 Hz, J2 = 4.4 Hz, 1H, ArCH2), 3.46–3.49 (m, 1H, ArCH2), 3.93
(s, 2H, ArCH2Ar), 7.03 (d, J = 7.4 Hz, 1H, Ar), 7.08 (d, J = 7.4 Hz, 1H,
Ar), 7.15–7.20 (m, 2H, Ar), 7.34 (t, J = 7.4 Hz, 1H, Ar), 7.42 (d,
J = 7.7 Hz, 1H, Ar), 7. 71 (d, J = 7.4 Hz, 1H, Ar), 7.75 (s, 1H, Ar).
4.21.1. Diethyl 2-acetamido-2-(3-(2-(ethoxycarbonyl)benzyl)-
benzyl)malonate (25a)
Yield 390 mg, 83%; MS: m/z 469 [M+]; 1H NMR (300 MHz,
CDCl3): d 1.26 (t, J = 7.2 Hz, 6H, OCH2CH3), 1.31 (t, J = 7.2 Hz, 3H,
OCH2CH3), 1.94 (s, 3H, NHCOCH3), 3.56 (s, 2H, CH2), 4.11–4.30
(m, 6H, OCH2CH3), 4.34 (s, 2H, ArCH2Ar), 6.50 (s, 1H, NH), 6.72 (s,
1H, Ar), 6.82 (d, J = 7.4 Hz, 1H, Ar), 7.06 (d, J = 7.7 Hz, 1H, Ar),
7.13–7.18 (m, 2H, Ar), 7.28 (dt, J1 = 7.7 Hz, J2 = 1.4 Hz, 1H, Ar),
7.42 (dt, J1 = 7.7 Hz, J2 = 1.4 Hz, 1H, Ar), 7.88 (dd, J1 = 7.7 Hz,
J2 = 1.7 Hz, 1H, Ar); 13C NMR (75 MHz, CDCl3): d 14.2, 14.4, 23.2,
38.0, 39.7, 61.1, 62.7, 67.3, 126.4, 127.6, 128.0, 128.2, 130.3,
130.5, 130.7, 131.7, 131.9, 135.3, 141.1, 141.7, 167.4, 167.6, 169.0.
4.23.2. (RS)-4-(3-(2-Amino-2-carboxyethyl)benzyl)benzoic acid
(26c)
Yield 129 mg, 87%; Anal. Calcd for C17H17NO4ꢂ0.2H2O: C, 67.4;
H, 5.79 N, 4.62. Found: C, 67.1; H, 5.39; N, 4.42; 1H NMR
(300 MHz, DMSO-d6): d 2.79 (dd, J1 = 14.3 Hz, J2 = 8.5 Hz, 1H,
ArCH2), 3.13 (dd, J1 = 14.3 Hz, J2 = 4.1 Hz, 1H, ArCH2), 3.39 (dd,
J1 = 4.3 Hz, J2 = 8.5 Hz, 1H, ArCH2), 3.95 (s, ArCH2Ar), 7.03 (d,
J = 7.4 Hz, 1H, Ar), 7.09 (d, J = 7.7 Hz, 1H, Ar), 7.15–7.21 (m, 2H,
Ar), 7.31 (d, J = 8.0 Hz, 2H, Ar), 7. 71 (d, J = 8.0 Hz, 2H, Ar).
4.21.2. Diethyl 2-acetamido-2-(3-(3-cyanobenzyl)benzyl)-
malonate (25b)
4.24. General procedure for compounds 29 and 30
Yield 271 mg, 64%; MS: m/z 422 [M+]; 1H NMR (300 MHz, CDCl3):
d 1.25 (t, J = 7.1, 6H, OCH2CH3), 1.94 (s, 3H, NHCOCH3), 3.61 (s, 2H,
CH2), 3.95 (s, 2H, ArCH2Ar), 4.13–4.26 (m, 4H, OCH2CH3), 6.48 (s,
1H, NH), 6.76 (s, 1H, Ar), 6.89 (d, J = 7.6 Hz, 1H, Ar), 7.06 (d,
J = 7.6 Hz, 1H, Ar), 7.22 (t, J = 7.3 Hz, 1H, Ar), 7.38–7.41 (m, 3H, Ar),
7.49–7.52 (m, 1H, Ar); 13C NMR (75 MHz, CDCl3): d 14.3, 23.2, 37.9,
41.5, 62.8, 67.3, 112.6, 118.8, 127.9, 128.3, 128.8, 129.4, 130.0,
130.5, 132.3, 133.5, 135.9, 139.4, 142.5, 167.4, 169.0.
Compounds 29 and 30 were prepared in analogy to the litera-
ture procedure.20 To a suspension of N-Boc-(S)-3-bromophenylala-
nine 27 (or N-Boc-(R)-3-bromophenylalanine 28) (344 mg,
1 mmol) and sodium hydrogencarbonate (168 mg, 2 mmol) in
DMF (5 mL), ethyl iodide (0.4 mL, 5 mmol) was added and the mix-
ture was stirred at room temperature for 24 h. After this time the
mixture was partitioned into water and ethyl acetate. The organic
phase was washed with water, dried over Na2SO4 and evaporated.
Crude product was purified by flash column chromatography.
4.21.3. Diethyl 2-acetamido-2-(3-(4-cyanobenzyl)benzyl)-
malonate (25c)
4.24.1. Ethyl (S)-3-(3-bromophenyl)-2-(pivaloyloxyamino)-
propanoate (29)
Yield 266 mg, 63%; MS: m/z 422 [M+]; 1H NMR (300 MHz,
CDCl3): d 1.25 (t, J = 7.1, 6H, OCH2CH3), 1.92 (s, 3H, NHCOCH3),
3.60 (s, 2H, CH2), 3.97 (s, 2H, ArCH2Ar), 4.12–4.25 (m, 4H,
OCH2CH3), 6.45 (s, 1H, NH), 6.75 (s, 1H, Ar), 6.88 (d, J = 7.3 Hz,
1H, Ar), 7.06 (d, J = 7.6 Hz, 1H, Ar), 7.19–7.27 (m, 3H, Ar), 7.58 (d,
J = 7.9 Hz, 2H, Ar); 13C NMR (75 MHz, CDCl3): d 14.3, 23.2, 37.9,
42.0, 62.8, 67.3, 110.2, 118.9, 128.0, 128.3, 128.8, 129.7, 130.6,
132.4, 135.9, 139.4, 146.6, 167.4, 168.9.
Yield 331 mg, 89%; 1H NMR (300 MHz, CDCl3): d 1.25 (t,
J = 7.2 Hz, 3H, OCH2CH3), 1.44 (s, 9H, CH3), 2.97–3.14 (m, 2H,
ArCH2), 4.17 (q, J = 7.2 Hz, 2H, OCH2CH3), 4.53 (m, 1H, CHNH),
5.02 (d, J = 6.9 Hz, 1H, NH), 7.07 (d, J = 7.4 Hz, 1H, Ar), 7.15 (t,
J = 7.7 Hz, 1H, Ar), 7.28 (s, 1H, Ar), 7.36 (d, J = 8.0 Hz, 1H, Ar); 13C
NMR (75 MHz, CDCl3): d 14.4, 28.5, 38.2, 54.5, 61.7, 80.2, 122.5,
128.0, 130.0, 130.1, 132.5, 138.5, 155.0, 171.5.
4.22. (RS)-2-(3-(2-Amino-2-carboxyethyl)benzyl)benzoic acid
(26a)
4.24.2. Ethyl (R)-3-(3-bromophenyl)-2-(pivaloyloxyamino)-
propanoate (30)
Yield 320 mg, 86%; 1H NMR (300 MHz, CDCl3): d 1.26 (t,
J = 7.2 Hz, 3H, OCH2CH3), 1.44 (s, 9H, CH3), 2.96–3.14 (m, 2H,
ArCH2), 4.17 (q, J = 7.2 Hz, 2H, OCH2CH3), 4.53 (m, 1H, CHNH),
5.02 (d, J = 6.9 Hz, 1H, NH), 7.07 (d, J = 7.4 Hz, 1H, Ar), 7.15 (t,
J = 7.7 Hz, 1H, Ar), 7.28 (s, 1H, Ar), 7.36 (d, J = 8.0 Hz, 1H, Ar); 13C
NMR (75 MHz, CDCl3): d 14.4, 28.5, 38.2, 54.5, 61.7, 80.2, 122.5,
128.0, 130.0, 130.1, 132.5, 138.5, 155.0, 171.5.
A mixture of 25a (352 mg, 0.75 mmol), acetic acid (3 mL), 12 M
HCl (3 mL), and water (3 mL) was refluxed for 20 h. After cooling
the mixture of acids was removed under reduced pressure; the so-
lid residue was dissolved in water (8 mL) and purified by a reverse-
phase flash column chromatography using R-18 Silica Gel (mobile
phase: mixture of methanol and 15 mM acetic acid with the gradi-
ent of concentration), followed by recrystallization from water.
Yield 205 mg, 91%; Anal. Calcd for C17H17NO4ꢂH2O: C, 64.34; H,
6.03; N, 4.41. Found: C, 64.36; H, 6.15; N, 4.22; 1H NMR
(300 MHz, DMSO-d6): d 2.79 (dd, J1 = 14.3 Hz, J2 = 8.3 Hz, 1H,
ArCH2), 3.10 (dd, J1 = 14.3 Hz, J2 = 4.4 Hz, 1H, ArCH2), 3.44 (dd,
J1 = 4.7 Hz, J2 = 8.3 Hz, 1H, ArCH2), 4.28 (s, 2H, ArCH2Ar), 6.93 (d,
4.25. General procedure for compounds 31 and 32
Compounds 31 and 32 were prepared from 29 and 30, respec-
tively (242 mg, 0.65 mmol), and 3-cyano-phenylboronic acid
according to the procedure described for compounds 8a–e.

´
6401
E. Szymanska et al. / Bioorg. Med. Chem. 17 (2009) 6390–6401
4.25.1. Ethyl (S)-3-(30-cyanobiphenyl-3-yl)-2-(pivaloyloxy-
Acknowledgments
amino)propanoate (31)
Yield 228 mg, 89%; 1H NMR (300 MHz, CDCl3): d 1.22 (t,
J = 7.2 Hz, 3H, OCH2CH3), 1.41 (s, 9H, CH3), 3.09–3.21 (m, 2H,
ArCH2), 4.16 (q, J = 7.2 Hz, 2H, OCH2CH3), 4.61 (m, 1H, CHNH),
5.03 (d, J = 7.1 Hz, 1H, NH), 7.06 (d, J = 7.6 Hz, 1H, Ar), 7.20 (s, 1H,
Ar), 7.37 (t, J = 7.6 Hz, 1H, Ar), 7.46 (d, J = 7.6 Hz, 1H, Ar), 7.61 (d,
J = 8.0 Hz, 2H, Ar), 7.70 (d, J = 8.0 Hz, 2H, Ar); 13C NMR (75 MHz,
CDCl3): d 14.4, 28.5, 38.6, 54.6, 61.6, 80.1, 111.1, 118.6, 126.1,
128.3, 129.4, 129.7, 129.9, 130.6, 130.9, 131.4, 136.4, 140.2,
143.4, 159.9, 171.8.
This work was supported by the Danish Medical Research Coun-
cil and the Alfred Benzon Foundation.
The expert technical assistance of Kaj Scherz Andersen is grate-
fully acknowledged.
References and notes
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3. Bleakman, D.; Alt, A.; Nisenbaum, E. S. Semin. Cell Dev. Biol. 2006, 17, 592.
4. Planells-Cases, R.; Lerma, J.; Ferrer-Montiel, A. Curr. Pharm. Des. 2006, 12, 3583.
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Neuropharmacology 1998, 37, 25.
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396.
8. Dominguez, E.; Iyengar, S.; Shannon, H. E.; Bleakman, D.; Alt, A.; Arnold, B. M.;
Bell, M. G.; Bleisch, T. J.; Buckmaster, J. L.; Castano, A. M.; Del Prado, M.;
Escribano, A.; Filla, S. A.; Ho, K. H.; Hudziak, K. J.; Jones, C. K.; Martinez-Perez, J.
A.; Mateo, A.; Mathes, B. M.; Mattiuz, E. L.; Ogden, A. M.; Simmons, R. M.; Stack,
D. R.; Stratford, R. E.; Winter, M. A.; Wu, Z.; Ornstein, P. L. J. Med. Chem. 2005,
48, 4200.
4.25.2. Ethyl (R)-3-(30-cyanobiphenyl-3-yl)-2-(pivaloyloxy-
amino)propanoate (32)
Yield 221 mg, 86%; 1H NMR (300 MHz, CDCl3): d 1.22 (t,
J = 7.2 Hz, 3H, OCH2CH3), 1.41 (s, 9H, CH3), 3.09–3.21 (m, 2H,
ArCH2), 4.16 (q, J = 7.2 Hz, 2H, OCH2CH3), 4.61 (m, 1H, CHNH),
5.03 (d, J = 7.1 Hz, 1H, NH), 7.06 (d, J = 7.6 Hz, 1H, Ar), 7.20 (s, 1H,
Ar), 7.37 (t, J = 7.6 Hz, 1H, Ar), 7.46 (d, J = 7.6 Hz, 1H, Ar), 7.61 (d,
J = 8.0 Hz, 2H, Ar), 7.70 (d, J = 8.0 Hz, 2H, Ar); 13C NMR (75 MHz,
CDCl3): d 14.4, 28.5, 38.6, 54.6, 61.6, 80.1, 111.1, 118.6, 126.1,
128.3, 129.4, 129.7, 129.9, 130.6, 130.9, 131.4, 136.4, 140.2,
143.4, 159.9, 171.8.
9. Hogner, A.; Greenwood, J. R.; Liljefors, T.; Lunn, M. L.; Egebjerg, J.; Larsen, I. K.;
Gouaux, E.; Kastrup, J. S. J. Med. Chem. 2003, 46, 214.
10. Møller, E. H.; Egebjerg, J.; Brehm, L.; Stensbøl, T. B.; Johansen, T. N.; Madsen, U.;
Krogsgaard-Larsen, P. Chirality 1999, 11, 752.
4.26. General procedure for compounds 33 and 34
11. Hald, H.; Naur, P.; Pickering, D. S.; Sprogøe, D.; Madsen, U.; Timmermann, D. B.;
Ahring, P. K.; Liljefors, T.; Schousboe, A.; Egebjerg, J.; Gajhede, M.; Kastrup, J. S.
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12. More, J. C.; Nistico, R.; Dolman, N. P.; Clarke, V. R.; Alt, A. J.; Ogden, A. M.;
Buelens, F. P.; Troop, H. M.; Kelland, E. E.; Pilato, F.; Bleakman, D.; Bortolotto, Z.
A.; Collingridge, G. L.; Jane, D. E. Neuropharmacology 2004, 47, 46.
13. Dolman, N. P.; Troop, H. M.; More, J. C.; Alt, A.; Knauss, J. L.; Nistico, R.; Jack, S.;
Morley, R. M.; Bortolotto, Z. A.; Roberts, P. J.; Bleakman, D.; Collingridge, G. L.;
Jane, D. E. J. Med. Chem. 2005, 48, 7867.
14. Dolman, N. P.; More, J. C.; Alt, A.; Knauss, J. L.; Troop, H. M.; Bleakman, D.;
Collingridge, G. L.; Jane, D. E. J. Med. Chem. 2006, 49, 2579.
15. Mayer, M. L.; Ghosal, A.; Dolman, N. P.; Jane, D. E. J. Neurosci. 2006, 26,
2852.
16. Dolman, N. P.; More, J. C.; Alt, A.; Knauss, J. L.; Pentikainen, O. T.; Glasser, C. R.;
Bleakman, D.; Mayer, M. L.; Collingridge, G. L.; Jane, D. E. J. Med. Chem. 2007, 50,
1558.
17. Chang, C. P.; Huang, Y. L.; Hong, F. E. Tetrahedron 2005, 61, 3835.
18. Botella, L.; Najera, C. J. Organometal. Chem. 2002, 663, 46.
19. Castanet, A. S.; Colobert, F.; Desmurs, J. R.; Schlama, T. J. Mol. Catal. A: Chem.
2002, 182, 481.
The target amino acids 33 and 34 were obtained from 31 and 32
(197 mg, 0.5 mmol) according to the procedure described for com-
pounds 18a–h, followed by two re-crystallizations from water.
4.26.1. (S)-30-(2-Amino-2-carboxyethyl)biphenyl-3-carboxylic
acid (33)
Yield 53 mg, 37%; ee = 99.2%; ½a D22
ꢃ
+26.2° (c 0.014, 1 M HCl:E-
tOH 1:1);
D
e
(214 nm) = +0.031 m2/mol; Anal. Calcd for
C16H15NO4ꢂ1.1H2O: C, 62.98; H, 6.68; N, 4.59. Found: C, 62.94; H,
5.40; N, 4.63; 1H NMR (300 MHz, DMSO-d6):
d
2.97 (dd,
J1 = 14.3 Hz, J2 = 8.3 Hz, 1H, ArCH2), 3.23 (dd, J1 = 14.3 Hz,
J2 = 4.1 Hz, 1H, ArCH2), 3.55 (dd, J1 = 8.3 Hz, J2 = 4.1 Hz, 1H,
ArCH2CH), 7.28 (d, J = 7.7 Hz, 1H, Ar), 7.38 (t, J = 7.7 Hz, 1H, Ar),
7.51–7.59 (m, 3H, Ar), 7.86–7.91 (m, 2H, Ar), 8.19 (s, 1H, Ar).
20. Aoki, S.; Cao, L. W.; Matsui, K.; Rachmat, R.; Akiyama, S.; Kobayashi, M.
Tetrahedron 2004, 60, 7053.
21. Schrödinger. Suite 2007; MacroModel v. 9.5; Induced Fit Docking protocol;
Glide version v. 4.5; Prime version v. 1.6: LLC, New York, NY, 2005.
22. Delano. Scientific, The PyMol Molecular Graphics System: San Carlos, CA, 2002.
23. Shoppee, C. W. J. Chem. Soc. C 1932, 696.
24. Bang-Andersen, B.; Ahmadian, H.; Lenz, S. M.; Stensbøl, T. B.; Madsen, U.;
Bøgesø, K. P.; Krogsgaard-Larsen, P. J. Med. Chem. 2000, 43, 4910.
25. Kaae, B. H.; Krogsgaard-Larsen, P.; Johansen, T. N. J. Org. Chem. 2004, 69, 1401.
26. Lemaire, M.; Guy, A.; Boutin, P.; Guette, J. P. Synthesis 1989, 761.
27. Liang, Y. X.; Gao, S.; Wan, H. H.; Wang, J. W.; Chen, H. L.; Zheng, Z.; Hu, X. Q.
Tetrahedron: Asymmetry 2003, 14, 1267.
4.26.2. (R)-30-(2-Amino-2-carboxyethyl)biphenyl-3-carboxylic
acid (34)
Yield 34 mg, 24%; ee = 98.4%; ½a D22
ꢀ23.7° (c 0.014, 1 M HCl:E-
ꢃ
tOH 1:1);
D
e
(219 nm) = ꢀ0.027 m2/mol; Anal. Calcd for
C16H15NO4ꢂ1.2H2O: C, 62.61; H, 5.71; N, 4.56. Found: C, 62.68; H,
5.43; N, 4.60; 1H NMR (300 MHz, DMSO-d6):
d 2.97 (dd,
J1 = 14.3 Hz, J2 = 8.3 Hz, 1H, ArCH2), 3.23 (dd, J1 = 14.3 Hz,
J2 = 4.1 Hz, 1H, ArCH2), 3.55 (dd, J1 = 8.3 Hz, J2 = 4.1 Hz, 1H,
ArCH2CH), 7.28 (d, J = 7.7 Hz, 1H, Ar), 7.38 (t, J = 7.7 Hz, 1H, Ar),
7.51–7.59 (m, 3H, Ar), 7.86–7.91 (m, 2H, Ar), 8.19 (s, 1H, Ar).
28. Godfrey, K. E.; Thrift, R. I. J. Chem. Soc. C 1967, 400.
29. Doyle, M. P.; Siegfried, B.; Dellaria, J. F. J. Org. Chem. 1977, 42, 2426.
30. Merlic, C. A.; Motamed, S.; Quinn, B. J. Org. Chem. 1995, 60, 3365.
31. Bang-Andersen, B.; Lenz, S. M.; Skj
ꢀrbꢀk, N.; Søby, K. K.; Hansen, H.
4.27. Pharmacology
O.; Ebert, B.; Bøgesø, K. P.; Krogsgaard-Larsen, P. J. Med. Chem. 1997, 40,
2831.
32. Terefenko, E. A.; Kern, J.; Fensome, A.; Wrobel, J.; Zhu, Y.; Cohen, J.; Winneker,
R.; Zhang, Z.; Zhang, P. Bioorg. Med. Chem. Lett. 2005, 15, 3600.
33. Ransom, R. W.; Stec, N. L. J. Neurochem. 1988, 51, 830.
34. Honoré, T.; Nielsen, M. Neurosci. Lett. 1985, 54, 27.
35. Braitman, D. J.; Coyle, J. T. Neuropharmacology 1987, 26, 1247.
36. Sills, M. A.; Fagg, G.; Pozza, M.; Angst, C.; Brundish, D. E.; Hurt, S. D.; Wilusz, E.
J.; Williams, M. Eur. J. Pharmacol. 1991, 192, 19.
37. Hermit, M. B.; Greenwood, J. R.; Nielsen, B.; Bunch, L.; Jørgensen, C. G.;
Vestergaard, H. T.; Stensbøl, T. B.; Sanchez, C.; Krogsgaard-Larsen, P.; Madsen,
U.; Bräuner-Osborne, H. Eur. J. Pharmacol. 2004, 486, 241.
38. Sagot, E.; Pickering, D. S.; Pu, X.; Umberti, M.; Stensbol, T. B.; Nielsen,
B.; Chapelet, M.; Bolte, J.; Gefflaut, T.; Bunch, L. J. Med. Chem. 2008, 51,
4093.
Rat brain membrane preparations used in the native receptor
binding experiments were prepared according to the method de-
scribed by Ransom and Stec.33 Affinity for AMPA,34 KA35 and
NMDA36 receptor sites was determined using 5 nM [3H]AMPA,
5 nM [3H]KA, and 2 nM [3H]CGP 39653 with some modifications
as previously described.37
Recombinant rat iGluR5(Q)1b, iGluR6(V,C,R) and iGluR7A were
expressed in Sf9 insect cells by baculoviral infection and receptor
binding assays carried out using [3H]-SYM 2081 radioligand as pre-
viously detailed.38