Design and synthesis of cyclopropane-based conformationally restricted GABA analogues as selective inhibitors for betaine/GABA transporter 1

Article abstract of DOI:10.1016/j.bmcl.2018.08.031

We previously designed and synthesized a series of cyclopropane-based conformationally restricted analogues of γ-aminobutyric acid (GABA). The study demonstrated that the critical conformation of the analogues that selectively active to betaine/GABA transporter 1 (BGT1) subtype is the trans-syn-form, in which the amino and carboxyl groups are in trans-configuration and the cyclopropane ring and the carboxyl group are in syn-arrangement. In this study, we designed and synthesized cyclopropane-based GABA analogues, which were conformationally restricted in the trans-syn-form by cyclopropylic strain based on the stereochemistry of the carbon adjacent to cyclopropane. Their conformation was confirmed as the syn-form by calculations and NMR studies, and their pharmacological evaluation clarified that compounds 11a and 11d had the BGT1 selectivity, although their inhibitory effects were insufficient.

Full text of DOI:10.1016/j.bmcl.2018.08.031

Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design and synthesis of cyclopropane-based conformationally restricted
GABA analogues as selective inhibitors for betaine/GABA transporter 1
Akihiro Suemasa, Mizuki Watanabe⁎, Takaaki Kobayashi, Hiroe Suzuki, Hayato Fukuda,
Masabumi Minami, Satoshi Shuto⁎
Faculty of Pharmaceutical Sciences, Hokkaido University, Kita-12, Nishi-6, Kita-ku, Sapporo 060-0812, Japan
A R T I C L E I N F O
A B S T R A C T
Keywords:
GABA
BGT1
Cyclopropane
Conformational restriction
Selective inhibitor
We previously designed and synthesized a series of cyclopropane-based conformationally restricted analogues of
γ-aminobutyric acid (GABA). The study demonstrated that the critical conformation of the analogues that se-
lectively active to betaine/GABA transporter 1 (BGT1) subtype is the trans-syn-form, in which the amino and
carboxyl groups are in trans-configuration and the cyclopropane ring and the carboxyl group are in syn-ar-
rangement. In this study, we designed and synthesized cyclopropane-based GABA analogues, which were con-
formationally restricted in the trans-syn-form by cyclopropylic strain based on the stereochemistry of the carbon
adjacent to cyclopropane. Their conformation was confirmed as the syn-form by calculations and NMR studies,
and their pharmacological evaluation clarified that compounds 11a and 11d had the BGT1 selectivity, although
their inhibitory effects were insufficient.
A major inhibitory neurotransmitter in the central nervous system
of mammals is γ-aminobutyric acid (GABA, 1, Fig. 1).1 GABA that is
released in the synaptic cleft acts on the GABAA and GABAB receptors to
transmit signals.2,3 This neurotransmission of GABA is terminated by
rapid reuptake of GABA through the GABA transporters (GATs), which
are expressed on the cell membrane of pre- and post-synapse and as-
trocytes.4,5 Four subtypes of GATs, i.e., GAT1, GAT2, GAT3, and be-
taine/GABA transporter 1 (BGT1), have been cloned. Each subtype
shows different distribution and physiological actions in the central
nervous system.6,7 Among the GAT subtypes, the functions of GAT1 and
its potential as a therapeutic target for neurological diseases have been
and therefore, development of such selective inhibitors, which can
work as effective tools to elucidate the role of GATs and also provide a
new therapeutic strategy for many neurological and psychiatric dis-
eases involved in the GABAergic system, is required.
As X-ray crystal structure analysis of GATs has not been succeeded,
their detailed three-dimensional structure information is not available.
To establish a methodology for the identification of the bioactive con-
formation of ligands even if the structural information of the target
protein is unknown, we previously presented the three-dimensional
structural diversity-oriented strategy,15 for which the chiral cyclopro-
pane units with two readily convertible substituents in a trans or cis
configuration (7a,b, and their enantiomers ent-7a,b; Fig. 2) were de-
veloped.16 These units can be used efficiently to synthesize a series of
conformationally restricted derivatives with three-dimensional di-
versity.17,18 Because of the three-dimensional structural diversity, one
of these isomers can be identical or analogous to the bioactive form of
the conformationally flexible prototype ligand, resulting in the dis-
covery of a hit that is superior to the prototype ligand in its target se-
lectivity and/or binding potency. We applied this strategy to GABA and
synthesized a series of cyclopropane-based conformationally restricted
GABA analogues,19 that is the eight regio- and stereoisomers consisting
of trans- and cis-α,β-methano- or β,γ-methano-GABAs (8a,b and 9a,b)
and their enantiomers (ent-8a,b and ent-9a,b) from the chiral cyclo-
propane units (Fig. 2). As a result, 9a, one of the β,γ-methano-GABAs,
well studied using highly selective and potent inhibitors of GAT1.8
A
selective GAT1 inhibitor, tiagabine (2, Fig. 1), in fact, has been clini-
cally used as an antiepileptic drug. Inhibitors for the three GAT sub-
types other than GAT1 have also been reported9; however, the potency
and selectivity have been insufficient, which causes the poorly detailed
studies for the three GAT subtypes.10 In recent years, a GAT3 selective
inhibitor 3 (Fig. 1) and a noncompetitive BGT1 selective inhibitor
BPDBA (4, Fig. 1) were identified through a chemical library screening
by Clausen and Wellendorph et al.11,12 They have also reported a tia-
gabine derivative, RPC425 (5, Fig. 1), with an improved subtype se-
lectivity for BGT1 among the GATs,13 and a BPDBA derivative 6 (Fig. 1)
with the enhanced solubility and specificity.14 However, genuinely
potent and selective inhibitors of non-GAT1 subtypes are still limited,10
⁎
Corresponding authors.
E-mail addresses: mwatanab@pharm.hokudai.ac.jp (M. Watanabe), shu@pharm.hokudai.ac.jp (S. Shuto).
https://doi.org/10.1016/j.bmcl.2018.08.031
Received 22 July 2018; Received in revised form 25 August 2018; Accepted 25 August 2018
0960-894X/©2018ElsevierLtd.Allrightsreserved.
Pleasecitethisarticleas:Suemasa,A.,Bioorganic&MedicinalChemistryLetters(2018),https://doi.org/10.1016/j.bmcl.2018.08.031

A. Suemasa et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Fig. 3. The anti- and syn-form of 9a and their conformationally restricted
analogues 10a and 10b with the rigid bicyclic ring system.
Fig. 1. Chemical structures of GABA and representative GATs selective in-
hibitors.
Fig. 4. Designed GABA analogues 11a–f whose conformation is regulated in the
trans-syn.
was identified as the GAT3/BGT1 dual inhibitor (IC50 = 14 μM for
GAT3, IC50 = 5.5 μM for BGT1), but the inhibitory activity and subtype
selectivity remained moderate. Although the conformation of 9a is re-
stricted by cyclopropane, the rotatable bond between cyclopropane ring
carbon (C1) and its adjacent carbon (C1′) can make the carboxyl group
orient to anti-form (in the side opposite to cyclopropane ring) or syn-
form (in the side same with cyclopropane ring) (Fig. 3). According to
the conformational analysis using MacroModel 10.9, the stability of
both conformers indicated similar relative potential energy (ΔE <
0.24 kcal/mol). We thought that the rotation around the C1–C1′ bond
might cause the insufficient selectivity. Therefore, we designed and
synthesized bicyclo[3.1.0]hexane derivatives 10a and 10b, in which
the orientation of the carboxyl group is fixed in the anti- or syn-form,
respectively, by the rigid bicyclic skeleton (Fig. 3).20 Their pharmaco-
logical evaluation showed that compound 10b, in which spatial ar-
rangement between the cyclopropane and carboxyl group was fixed in
the syn-form, was identified as the first highly potent and selective in-
hibitor for BGT1 (IC50 = 76 μM for GAT3, IC50 = 0.59 μM for BGT1).
Thus, we proposed that the bioactive conformation of GABA for BGT1 is
the trans-syn-form.
Fig. 5. Conformational restriction based on the cyclopropylic strain.
Fig. 6. Conformational restriction of 11a–f to the syn-form by the cyclopropylic
strain.
local stable conformations other than the syn-type within 1.5 kcal/mol
of relative potential energy from the global minimum. The most stable
syn-form of 11a (R = Me) obtained by the calculations is shown as a
representative example in Fig. 7a. The calculation results show that the
orientation of the carboxyl group can be effectively regulated due to the
cyclopropylic strain, as we expected. In 11c and 11e, the most stable
conformations were the syn-form, while other local stable conforma-
tions with the anti-form were present within energy difference of
0.50 kcal/mol. Besides, when the most stable conformation of 11a was
superimposed on the most stable conformation of bicyclic compound
10b (Fig. 7b), the three-dimensional positioning of the essential amino
group and carboxyl group were overlapped very well (Fig. 7c). There-
fore, the conformation of 11a–f is restricted to the trans-syn-form si-
milar to 10b, which seems to be essential for exhibiting the BGT1 se-
lectivity. Furthermore, in the designed compounds, the substituent that
is introduced to regulate the conformation can fill the space around the
anti-direction and might interact with the binding site of GATs.
Therefore, we thought that the new interaction, which is impossible in
the previous compound 10b, might improve the selectivity/inhibitory
activity against BGT1.
In this study, we designed 9a derivatives, 11a–f, having a sub-
stituent on the adjacent carbon (C1′) of cyclopropane (Fig. 4). The
conformation of the derivatives was regulated to the trans-syn-form due
to “cyclopropylic strain” instead of the rigid bicyclic ring system in 10b.
Adjacent substituents in cis-configuration on the cyclopropane ring are
fixed in eclipse orientation, so sizable steric repulsion like 1,3-allylic
strain occurs between the two substituents, that is “cyclopropylic
strain.”21 By the cyclopropylic strain, the conformation where the
smallest group on the C1’ orients toward the cyclopropane side is
22,23
stable, as shown in Fig. 5
Thus, depending on the C1’-stereo-
chemistry, the orientation of the side chain can be controlled. That is, if
a substituent is introduced at the C1’-position in the stereochemistry
shown in Fig. 4, the orientation of the carboxyl group should prefer the
syn-form to the anti-form (Fig. 6). The conformation of the designed
compounds 11a–f was investigated by calculations using MacroMolel
10.9, which showed that the most stable conformation of all of the
compounds was the syn-form. In 11a, 11b, 11d, and 11f, there were no
Chiral Cyclopropane Units
α,β-methano-GABAs
β,γ-methano-GABAs
OTBDPS
NH2
8b
CO2H
CO2H
H2N
CO2H
OHC
OHC
OTBDPS
HO2C
HO2C
NH2
CHO
CO2H
NH2
NH2
9a
GAT3/BGT1 inhibitor
7a
7b
OTBDPS
8a
9b
NH2
OTBDPS
NH2
H2N
CO2H
CHO
CO2H
ent-7a
ent-7b
ent-8a
ent-8b
ent-9a
ent-9b
Fig. 2. Chiral cyclopropane units and the conformationally restricted GABA analogues.
2

A. Suemasa et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Fig. 7. The most stable conformation of 11a (a) and
10b (b) by the conformational search using
MacroModel 10.9. (Force field: MMFFs, solvent:
H2O); (c) superimposition of the stable conformation
of 11a and 10b.
conditions28 to provide carboxylic acid 20a. Conversion to a benzyl
ester and subsequent removal of the TBDPS group afforded alcohol 21a.
Treatment of 19b with lithium benzyl oxide prepared in situ gave a
benzyl ester, followed by TBAF treatment led alcohol 21b. Removal of
the chiral auxiliary of 19c and 19d by methanolysis using MeMgBr
premixed in MeOH29 produced methyl esters 20b and 20c, respectively.
Deprotection of a hydroxyl group of 20b gave alcohol 21c. Acetylation
of the secondary hydroxy group of 20c and subsequent removal of the
TBDPS group afforded primary alcohol 21d, which was subjected to the
next step without purification (Scheme 4).
Scheme 1. Reagents and conditions. (a) EtMgBr, THF, 97%; (b) (1)
MeOCH2PPh3Cl, NaHMDS, THF, 0 °C; (2) aq. HCl, THF, 0 °C; (3) NaClO2,
NaH2PO4, 2-methyl-2-butene, aq. tBuOH; (4) CbzCl, Et3N, DMAP, CH2Cl2; (5)
TBAF, THF, 11% (14) and 7% (15) for 5 steps.
To synthesize 11f having a vinyl group, we conducted a stereo-
selectively benzyloxymethylation32 of 22 with a chiral auxiliary using
benzyl chloromethyl ether (BOMCl) to give 23 (Scheme 3). Reductive
removal of the oxazolidinone of 23 provided alcohol 24, which was
oxidized to aldehyde 25. After construction of a vinyl group and sub-
sequent removal of the benzyl group, alcohol 27 was converted to
benzyl ester 28, which was treated with TBAF to afford alcohol 29.
Alcohols 14, 21a–d, and 29 were subjected to the stepwise oxida-
tions and the subsequent conversion to the corresponding acid azides,
followed by Curtius rearrangement gave N-Boc-amines 30a–f (Scheme
4). Stepwise removal of the benzyl group and the Boc group of 30a and
30b provided final compounds 11a and 11b, and simultaneous dees-
terification and N-Boc removal of 30c–f afforded final compounds
11c–f, respectively. The absolute configuration of the C1′ of 11a, 11b,
11d, and 11e was determined by the phenylglycine methyl ester
(PGME) method30 or the modified Mosher method.31 Also, that of 11c
was presumed from the chemical shift values of the 1H NMR spectra,
and that of 11f was determined by the conversion of 29 to the corre-
sponding ethyl compound, of which NMR spectra agreed with those of
All of the designed conformationally restricted GABA analogues
(11a–f, Fig. 4) was synthesized from trans-cyclopropane Weinreb amide
12,17 which was converted to ketone 13 (Scheme 1). Wittig reaction of
13 using MeOCH2PPh3Cl, followed by acidic hydrolysis, Pinnick oxi-
dation, subsequent treatment with CbzCl, removal of the TBDPS group
and final separation by silica gel chromatography, afforded diaster-
eomers 14 and 15, respectively.
The substituents (Me, F, N3, and OH) at the C1′-position were ste-
reoselectively introduced using Evans’ chiral auxiliary (Scheme 2). Al-
dehyde 1624 was oxidized to carboxylic acid 17, which was condensed
with a chiral auxiliary (4R)-4-benzyl-2-oxazolidinone to afford common
intermediate 18. The substituent was successfully introduced to the C1′
of 18 using CH3I to give 19a (R = Me), N-fluorobenzenesulfonimide
(NFSI)25 to give 19b (R = F), 2,4,6-triisopropylbenzenesulfonyl azide26
to give 19c (R = N3), or 3-phenyl-2-(phenylsulfonyl)-1,2-oxaziridine
(Davis' oxaziridine)27 to give 19d (R = OH), respectively. The chiral
auxiliary of 19a was removed by perhydrolysis under the LiOH/H2O2
Scheme 2. Reagents and conditions. (a) NaClO2,
NaH2PO4, 2-methyl-2-butene, aq. tBuOH, 96%; (b)
(1) PivCl, Et3N, CH2Cl2, 0 °C; (2) (4R)-4-benzyl-2-
oxazolidinone, nBuLi, THF, −78 °C, 82% for 2 steps;
(c) CH3I, NaHMDS, CH2Cl2, −78 °C, 83% for 19a;
(d) NFSI, NaHMDS, THF, −78 °C, 44% for 19b; (e)
2,4,6-triisopropylbenzenesulfonyl azide, KHMDS,
THF, −78 °C, then AcOH, for 19c; (f) Davis’ ox-
aziridine, NaHMDS, THF, −78 °C, for 19d; (g) LiOH,
H2O2, aq·THF, 0 °C, 97% for 20a from 19a; (h) (1)
CbzCl, Et3N, DMAP, CH2Cl2, 0 °C to rt; (2) NH4F,
MeOH, reflux, 59% for 21a from 20a; (i) (1) BnOH,
nBuLi, THF, 0 °C; (2) TBAF, AcOH, THF, 76% for
21b from 19b; (j) MeMgBr, MeOH, 0 °C, 38% for
20b and 41% for 20c from 18, respectively; (k) NH4F, MeOH, 95% for 21c from 20b; (l) (1) Ac2O, Et3N, DMAP, CH2Cl2; (2) NH4F, MeOH, reflux, for 21d from 20c
(not isolated).
Scheme 3. Reagents and conditions. (a) (1)
PivCl, Et3N, CH2Cl2, 0 °C; (2) (4S)-4-benzyl-2-
oxazolidinone, nBuLi, THF, −78 °C, 88% for 2
steps; (b) BOMCl, LHMDS, LiI, THF, −78 °C,
91%; (c) LiBH4, THF, MeOH, 0 °C, 89%; (d)
Dess–Martin periodinane, CH2Cl2, 89%; (e)
Ph3PMeBr, nBuLi, THF, −78 °C to 0 °C, 83%; (f)
DDQ, H2O, CH2Cl2, 92%; (g) (1) Dess–Martin
periodinane, CH2Cl2; (2) NaClO2, NaH2PO4, 2-
methyl-2-butene, aq. tBuOH; (3) BnOH, EDC, DMAP, CH2Cl2, 0 °C, 79% for 3 steps; (h) TBAF, AcOH, THF, 85%.
3

A. Suemasa et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Table 1
Inhibitory effects (%) of GABA analogues (100 µM) on GABA uptake in GAT
subtypes.a
R
GAT1
GAT2
GAT3
BGT1
11a
11b
11c
11d
11e
11f
Me
Et
F
N3
OH
vinyl
–
4.3
8.6
5.4
13
−5.2
3.3
45.3
6.8
20.1
−9.5
66.8
29.0
2.3
4.2
2.1
4.4
1.3
9.3
−2.4
5.2
11.8
8.3
−3.6
92.6
48.2
1.3
4.7
2.4
8.9
2.0
25
2.2
62.7
13.4
22.2
57.2
44.7
−32.5
92.8
97.6
6.3
6.4
3.4
3.3
1.1
NTb
NTb
NTb
NTb
25.1
12.2
36
1.1
30
0.7
9ac
8.1
10bc
–
Scheme 4. Reagents and conditions. (a) (1) Dess–Martin periodinane, CH2Cl2;
(2) NaClO2, NaH2PO4, 2-methyl-2-butene, aq. tBuOH; (3) (PhO)2P(O)N3, Et3N,
CH2Cl2; (4) tBuOH, reflux, 71% for 30a from 21a, 28% for 30b from 14, 50%
for 30c from 21b, 75% for 30d from 21c, 46% for 30e from 21d, 59% for 30f
from 29; (b) (1) H2, Pd/C, THF; (2) HCl, dioxane, 91% for 11a from 30a, 82%
for 11b from 30b; (c) aq. HCl, reflux, 98% for 11c from 30c, 99% for 11d from
30d, 99% for 11e from 30e, 26% for 11f from 30f.
a
Data are expressed as means
Not tested.
Data were taken from Refs. 19 (c) and 20 (d).
SEM (n = 3).
b
c
described above. These results mean that the sufficient conformational
restriction by the methyl group or the azide group at the C1′ caused the
BGT1 selectivity among GATs and that the bioactive conformation for
the BGT1 binding is the syn-form,35 consistent with the previous study
results. On the other hand, 11c failed to have the BGT1 selectivity. The
cause of this result might be that 11c can take a conformation other
than the syn-form due to insufficient cyclopropylic strain by the rather
small fluorine substituent and the fluorine substitution can modulate
the physicochemical properties of the molecule affecting the binding
affinity to the protein.33,36 However, the inhibitory effects of all the
compounds were significantly lower than those of the lead compounds
9a and 10b. These results indicate that the substituent that was in-
troduced to restrict the conformation did not interact with BGT1 but
was a steric hindrance for the binding to BGT1, decreasing the in-
hibitory effect. Notably, despite their assuming the same conformation,
while 11a (R = Me) and 11d (R = N3) and still showed the weak in-
hibitory effects on BGT1, 11b (R = Et) and 11f (R = vinyl) lost them.
These results suggested that, when a series of cyclopropane GABA de-
rivatives bind to BGT1, the space accommodating a hydrophobic sub-
stituent in the anti-direction at the C1′ seems to be rather limited.
In summary, we designed and synthesized cyclopropane-based
conformationally restricted GABA analogues of in which the relative
positioning of the cyclopropane and the carboxylic group is effectively
regulated in the syn-form by the cyclopropylic strain. Compounds 11a
and 11d exhibited the BGT1 selectivity due to their syn-conformation,
but their inhibitory activities against BGT1 were insufficient probably
due to the steric hindrance by the introduced substituents. The present
study on the conformational restriction of GABA provides the vital in-
formation about the bioactive conformation and about the space which
can be introduced substituents between GABA and BGT1.
Fig. 8. NOEs observed in compounds 11a–f (CD3OD, 400 MHz).
the compound derived from 14 (Schemes S1, S2 and Figs. S1, S2 in the
Supplementary data).
Stable conformations of all synthesized conformationally restricted
analogues were analyzed by NOE experiments in CD3OD (Fig. 8). In all
the compounds 11a–f, NOEs between H-1′ and H-3a at the cis-position to
the C1′ were observed. Also, NOEs between H-1′ and H-2 on cyclopro-
pane were observed in all the compounds except for 11f. These results
suggest that the smallest H-1′ at the C1′ is oriented to the cyclopropane-
ring side in all the compounds. Thus, as we expected, the conformations
of the compounds were regulated to the syn-form due to the cyclo-
propylic strain by the substituent introduced at the C1′-position, as in-
dicated in Fig. 6. However, only for 11c, an NOE between H-1′ and H-1
was also observed, suggesting that the fluorine substituent can be stably
oriented to the inside. This result means that not only syn-form but also
anti-form in 11c can be stable, and this is consistent with the results of
the calculations described above. The covalently bound fluorine is less
bulky than a methyl group, and the C–F bond length is relatively short,
although van der Waals radius of fluorine (1.47 Å) is larger than that of
hydrogen (1.20 Å).33,34 Thus, the steric repulsion between the fluorine
and the H-2/H-3a on the cyclopropane ring seems to be insufficient for
effectively restricting the conformation around the C1′.
Acknowledgements
This work was supported by MEXT/JSPS KAKENHI Grant Numbers
JP15H02495 (to SS), JP17K15476 (to MW), and a research grant from
Takeda Science Foundation (to MW), and partly by Platform Project for
Supporting Drug Discovery and Life Science Research (BINDS) from
AMED under Grant Number JP18am0101093. We are grateful to Sanyo
Fine Co., Ltd. for the gift of the chiral epichlorohydrin.
Appendix A. Supplementary data
The inhibitory effects of the conformationally restricted analogues
11a–f at 100 µM on GABA uptake in each GAT subtypes were evaluated
using CHO cells stably expressing GATs according to the reported
method19 (Table 1). As a result, 11a, 11d, and 11e showed approxi-
mately 5–7-fold selectivity for BGT1 over GAT3. 11a and 11d also
exhibited the BGT1 selectivity over GAT2. The stable conformation of
11a and 11d was strongly suggested to be the syn-form from the
computational analysis using MacroModel and the NOE experiments
Supplementary data associated with this article can be found, in the
online version, at https://doi.org/10.1016/j.bmcl.2018.08.031.
References
1. Ben-Ari Y, Gaiarsa JL, Tyzio R, Khazipov R. GABA: a pioneer transmitter that excites
immature neurons and generates primitive oscillations. Physiol Rev.
2007;87(4):1215–1284.
4

A. Suemasa et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
2. Bettler B, Kaupmann K, Mosbacher J, Gassmann M. Molecular structure and phy-
siological functions of GABA(B) receptors. Physiol Rev. 2004;84(3):835–867.
3. Farrant M, Nusser Z. Variations on an inhibitory theme: phasic and tonic activation of
GABA(A) receptors. Nat Rev Neurosci. 2005;6(3):215–229.
4. Heja L, Karacs K, Kardos J. Role for GABA and Glu plasma membrane transporters in
the interplay of inhibitory and excitatory neurotransmission. Curr Top Med Chem.
2006;6(10):989–995.
lead for potent inhibitors of GABA transports. Bioorg Med Chem.
2013;21(17):4938–4950.
20. Kobayashi T, Suemasa A, Igawa A, et al. Conformationally restricted GABA with
bicyclo[3.1.0]hexane backbone as the first highly selective BGT-1 inhibitor. ACS Med
Chem Lett. 2014;5(8):889–893.
21. Shuto S, Ono S, Hase Y, et al. Conformational restriction by repulsion between ad-
jacent substituents on a cyclopropane ring: design and enantioselective synthesis of
1-phenyl-2-(1-aminoalkyl)-N, N-diethylcyclopropanecarboxamides as potent NMDA
receptor antagonists. J Org Chem. 1996;61(3):915–923.
5. Heja L, Nyitrai G, Kekesi O, et al. Astrocytes convert network excitation to tonic
inhibition of neurons. BMC Biol. 2012;10:21.
6. Conti F, Minelli A, Melone M. GABA transporters in the mammalian cerebral cortex:
localization, development and pathological implications. Brain Res Rev.
2004;45(3):196–212.
7. Kempson SA, Zhou Y, Danbolt NC. The betaine/GABA transporter and betaine: roles
in brain, kidney, and liver. Front Physiol. 2014;5:16.
22. Watanabe M, Hirokawa T, Kobayashi T, et al. Investigation of the bioactive con-
formation of histamine H3 receptor antagonists by the cyclopropylic strain-based
conformational restriction strategy. J Med Chem. 2010;53(9):3585–3593.
23. Kawamura S, Unno Y, Tanaka M, et al. Investigation of the noncovalent binding
mode of covalent proteasome inhibitors around the transition state by combined use
of cyclopropylic strain-based conformational restriction and computational mod-
eling. J Med Chem. 2013;56(14):5829–5842.
8. Salat K, Kulig K. GABA transporters as targets for new drugs. Future Med Chem.
2011;3(2):211–222.
9. Thomsen C, Sørensen PO, Egebjerg J. 1-(3-(9H-Carbazol-9-yl)-1-propyl)-4-(2-meth-
oxyphenyl)-4-piperidinol, a novel subtype selective inhibitor of the mouse type II
GABA-transporter. Br J Pharmacol. 1997;120(6):983–985.
10. Damgaard M, Haugaard AS, Kickinger S, et al. Development of non-GAT1-selective
inhibitors: challenges and achievements. In: Ortega A, Schousboe A, eds. Glial amino
acid transporters. Cham, Switzerland: Springer; 2017:315–332.
24. Watanabe M, Kobayashi T, Hirokawa T, et al. Cyclopropane-based stereochemical
diversity-oriented conformational restriction strategy: histamine H3 and/or H4 re-
ceptor ligands with the 2,3-methanobutane backbone. Org Biomol Chem.
2012;10(4):736–745.
25. Kirk KL. Fluorination in medicinal chemistry: methods, strategies, and recent de-
velopments. Org Process Res Dev. 2008;12(2):305–321.
11. Damgaard M, Al-Khawaja A, Vogensen SB, et al. Identification of the first highly
subtype-selective inhibitor of human GABA transporter GAT3. ACS Chem Neurosci.
2015;6(9):1591–1599.
26. Evans DA, Britton TC. Electrophilic azide transfer to chiral enolates. A general ap-
proach to the asymmetric synthesis of.alpha.-amino acids. J Am Chem Soc.
1987;109(22):6881–6883.
12. Kragholm B, Kvist T, Madsen KK, et al. Discovery of a subtype selective inhibitor of
the human betaine/GABA transporter 1 (BGT-1) with a non-competitive pharmaco-
logical profile. Biochem Pharmacol. 2013;86(4):521–528.
27. Davis FA, Lamendola J, Nadir U, et al. Chemistry of oxaziridines. 1. Synthesis and
structure of 2-arenesulfonyl-3-aryloxaziridines. A new class of oxaziridines. J Am
Chem Soc. 1980;102(6):2000–2005.
13. Vogensen SB, Jorgensen L, Madsen KK, et al. Selective mGAT2 (BGT-1) GABA uptake
inhibitors: design, synthesis, and pharmacological characterization. J Med Chem.
2013;56(5):2160–2164.
28. Evans DA, Britton TC, Ellman JA. Contrasteric carboximide hydrolysis with lithium
hydroperoxide. Tetrahedron Lett. 1987;28(49):6141–6144.
29. Williams DR, Patnaik S, Clark MP. Total synthesis of cystothiazoles A and C. J Org
14. Jorgensen L, Al-Khawaja A, Kickinger S, et al. Structure-activity relationship, phar-
macological characterization, and molecular modeling of noncompetitive inhibitors
of the betaine/gamma-aminobutyric acid transporter 1 (BGT1). J Med Chem.
2017;60(21):8834–8846.
Chem. 2001;66(25):8463–8469.
30. Yabuuchi T, Kusumi T. Phenylglycine methyl ester, a useful tool for absolute con-
figuration determination of various chiral carboxylic acids. J Org Chem.
2000;65(2):397–404.
15. Mizuno A, Matsui K, Shuto S. From peptides to peptidomimetics: a strategy based on
the structural features of cyclopropane. Chem Eur J. 2017;23(58):14394–14409.
16. Kazuta Y, Matsuda A, Shuto S. Development of versatile cis- and trans-dicarbon-
substituted chiral cyclopropane units: Synthesis of (1S,2R)- and (1R,2R)-2-amino-
methyl-1-(1H-imidazol-4-yl)cyclopropanes and their enantiomers as con-
formationally restricted analogues of histamine. J Org Chem. 2002;67(5):1669–1677.
17. Watanabe M, Kazuta Y, Hayashi H, Yamada S, Matsuda A, Shuto S. Stereochemical
diversity-oriented conformational restriction strategy. Development of potent hista-
mine H-3 and/or H-4 receptor antagonists with an imidazolylcyclopropane structure.
J Med Chem. 2006;49(18):5587–5596.
31. Ohtani I, Kusumi T, Kashman Y, Kakisawa H. High-field FT NMR application of
Mosher's method. The absolute configurations of marine terpenoids. J Am Chem Soc.
1991;113(11):4092–4096.
32. Ishikawa Y, Nishiyama S. Synthesis of the BCD ring system of azaspiracid: con-
struction of the trispiro ring structure by the thioether approach. Tetrahedron Lett.
2004;45(2):351–354.
33. Smart BE. Fluorine substituent effects (on bioactivity). J Fluor Chem.
2001;109(1):3–11.
34. O'Hagan D. Understanding organofluorine chemistry. An introduction to the C-F
bond. Chem Soc Rev. 2008;37(2):308–319.
18. Watanabe M, Yamaguchi K, Tang W, et al. Synthesis of a series of 3,4-metha-
noarginines as side-chain conformationally restricted analogues of arginine. Bioorg
Med Chem. 2011;19(20):5984–5988.
19. Nakada K, Yoshikawa M, Ide S, et al. Cyclopropane-based conformational restriction
of GABA by a stereochemical diversity-oriented strategy: identification of an efficient
35. The BGT1 selectivity of S6, which is the anti-isomer of 11a, was lower than that of
11a (Table S1 in the Supplementary data). This result also supported that the
bioactive conformation for the BGT1 binding is the syn-form.
36. Böhm H-J, Banner D, Bendels S, et al. Fluorine in medicinal chemistry.
ChemBioChem. 2004;5(5):637–643.
5