Synthesis, radiosynthesis and biological evaluation of 1,4-dihydroquinoline derivatives as new carriers for specific brain delivery

Article abstract of DOI:10.1039/b909650g

In spite of numerous reports dealing with the use of 1,4-dihydropyridines as carriers to deliver biological active compounds to the brain, this chemical delivery system (CDS) suffers from poor stability of the 1,4-dihydropyridine derivatives towards oxidation and hydration reactions seriously limiting further investigations in vivo. In an attempt to overcome these limitations, we report herein the first biological evaluation of more stable annellated NADH models in the quinoline series as relevant neuroactive drug-carrier candidates. The radiolabeled 1,4-dihydroquinoline [¹¹C]1a was prepared to be subsequently peripherally injected in rats. The injected animals were sacrificed and brains were collected. The radioactivity measured in rat brain indicated a rapid penetration of the carrier [¹¹C]1a into the CNS. HPLC analysis of brain homogenates showed that oxidation of [¹¹C]1a into the corresponding quinolinium salt [¹¹C]4a was completed in less than 5 min. An in vivo evaluation in mice is also reported to illustrate the potential of such 1,4-dihydroquinoline derivatives to transport a neuroactive drug in the CNS. For this purpose, γ-aminobutyric acid (GABA), well known to poorly cross the brain blood barrier (BBB) was connected to this 1,4-dihydroquinoline- type carrier. After i.p. injection of 1,4-dihydroquinoline-GABA derivative 1b in mice, a significant alteration of locomotor activity (LMA) was observed presumably resulting from an enhancement of central GABAergic activity. These encouraging results give strong evidence for the capacity of carrier-GABA derivative 1b to cross the BBB and exert a pharmacological effect on the CNS. This study paves the way for further progress in designing new redox chemical delivery systems.

Full text of DOI:10.1039/b909650g

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www.rsc.org/obc | Organic & Biomolecular Chemistry
Synthesis, radiosynthesis and biological evaluation of 1,4-dihydroquinoline
derivatives as new carriers for specific brain delivery†
Le´na¨ıg Foucout,a Fabienne Gourand,b Martine Dhilly,b Pierre Bohn,a,c Georges Dupas,a Jean Costentin,c
Ahmed Abbas,d Francis Marsais,a Louisa Barre´*b and Vincent Levacher*a
Received 15th May 2009, Accepted 15th June 2009
First published as an Advance Article on the web 10th July 2009
DOI: 10.1039/b909650g
In spite of numerous reports dealing with the use of 1,4-dihydropyridines as carriers to deliver
biological active compounds to the brain, this chemical delivery system (CDS) suffers from poor
stability of the 1,4-dihydropyridine derivatives towards oxidation and hydration reactions seriously
limiting further investigations in vivo. In an attempt to overcome these limitations, we report herein the
first biological evaluation of more stable annellated NADH models in the quinoline series as relevant
neuroactive drug-carrier candidates. The radiolabeled 1,4-dihydroquinoline [11C]1a was prepared to be
subsequently peripherally injected in rats. The injected animals were sacrificed and brains were
collected. The radioactivity measured in rat brain indicated a rapid penetration of the carrier [11C]1a
into the CNS. HPLC analysis of brain homogenates showed that oxidation of [11C]1a into the
corresponding quinolinium salt [11C]4a was completed in less than 5 min. An in vivo evaluation in mice
is also reported to illustrate the potential of such 1,4-dihydroquinoline derivatives to transport a
neuroactive drug in the CNS. For this purpose, g-aminobutyric acid (GABA), well known to poorly
cross the brain blood barrier (BBB) was connected to this 1,4-dihydroquinoline-type carrier. After i.p.
injection of 1,4-dihydroquinoline-GABA derivative 1b in mice, a significant alteration of locomotor
activity (LMA) was observed presumably resulting from an enhancement of central GABAergic
activity. These encouraging results give strong evidence for the capacity of carrier-GABA derivative 1b
to cross the BBB and exert a pharmacological effect on the CNS. This study paves the way for further
progress in designing new redox chemical delivery systems.
The BBB being essentially impermeable to hydrophilic and/or
Introduction
charged compounds, one of the most straightforward strategies
The delivery of drugs to the brain remains a major challenge in the
to surmount this transport problem is to administrate prodrugs
treatment of central nervous system (CNS) disorders. The blood-
sufficiently lipophilic to facilitate their passive diffusion. Among
brain barrier (BBB), a physiological barrier between blood and
the various pharmacological strategies to target drugs to the brain,
brain, constitutes the main obstacle for CNS therapy and a serious
one can mention Bodor’s chemical delivery systems (CDS). In this
bottleneck in drug development for CNS diseases. The BBB resides
approach, the drug is covalently coupled to a lipophilic redox
mainly within the endothelial cells of cerebral capillaries that
carrier; i.e. a 1,4-dihydropyridine derived from nicotinic acid. The
are connected with tight continuous circumferential junctions
first task of this 1,4-dihydropyridine is to increase sufficiently
hampering the passage of hydrophilic drugs to the brain. As a
the lipophilicity of the drug to facilitate crossing of the BBB
result, more than 98% of small molecules do not cross the BBB
via passive diffusion. Once in the brain, the 1,4-dihydropyridine
while transport of nutriments and hormones that are essential
moiety is converted into the corresponding pyridinium salt by
for brain functions are regulated by specific BBB transporters.
oxido-reductases. The so-obtained permanently charged carrier-
drug can no longer cross back the BBB by passive diffusion
aLaboratoire de Chimie Organique Fine et He´te´rocyclique, UMR 6014,
and remains locked in the CNS. Subsequently, the parent drug
IRCOF, CNRS, Universite´ et INSA de Rouen, B.P. 08 F-76131, Mont-
is released from the carrier by enzymatic cleavage providing
Saint-Aignan Cedex, (France). E-mail: vincent.levacher@insa-rouen.fr;
brain-specific release of the active drug while reducing putative
peripheral effects. This redox CDS has been explored for brain
targeting of a wide variety of drug classes including steroids,
antiviral agents, anti-tumor agents, antibiotics, anticonvulsants
and neuropeptides.1 A serious limitation observed with the 1,4-
dihydropyridine carrier arose from its too high sensitivity to
oxidation and/or hydration on the enamine 5,6-double bond of the
dihydropyridine ring.2 Although some attempts have been made
to address this problem through chemical modifications of the 1,4-
dihydopyridine ring,3 the modest stability of this class of carriers
remains an obstacle to further development of this CDS approach.
Fax: +33(0)235522962
bGroupe de De´veloppements Me´thodologiques en Tomographie par Emis-
sion de Positons, CEA/DSV/I2BM/CI-NAPS UMR6232, Universite´
de Caen Basse Normandie, Caen, (France). E-mail: barre@cyceron.fr;
Fax: +33(0)231470275
cLaboratoire de Neuropharmacologie Expe´rimentale associe´ au CNRS
(FRE-2735). Faculte´ de Me´decine et de pharmacie, Universite´ de Rouen,
F-76000, (France)
dInserm-EPHE-Universite´ de Caen Basse-Normandie, Unite´ U923, GIP
Cyceron, CHU Coˆte de Nacre, Caen, (France)
† Electronic supplementary information (ESI) available: 1H NMR spectra
of 1a, 1b and 1d, HPLC chromatograms of 1a and 1b. See DOI:
10.1039/b909650g
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In this context, our expertise in the preparation of more stable
annellated NADH models in the quinoline series and their use in
biomimetic reductions4 prompted us to evaluate their potential as
carriers to target drugs in the CNS.5 A survey of the literature
reveals that little attention has been paid to the evaluation of 1,4-
dihydroquinoline derivatives as potential carriers.6 However, the
use of 1,4-dihydroquinolines may display attractive advantages
over the more conventional 1,4-dihydropyridines: (1) annelation of
the 1,4-dihydopyridine ring protects the enamine 5,6-double bond
from hydration, (2) additional electron withdrawing or donating
functional groups (FG) may be easily installed on the phenyl
ring, offering the possibility to tune the redox potential of the
carrier to find a good balance between stabilization of the 1,4-
dihydroquinoline at the periphery and its oxidation in the CNS
(Fig. 1).
of BBB penetration and central activities of administrated 1,4-
dihydroquinoline-GABA derivatives by investigating a putative
diminution of the locomotor activity (LMA) (Fig. 2b).8
Results and discussion
Design of the redox chemical delivery system (lipophilicity, redox
potential, linker)
The redox CDS was designed to satisfy several physicochemical
parameters regarding the lipophilicity and redox properties of the
system and implementation of an appropriate covalent connection
between the carrier and GABA compatible with enzymatic
cleavage in the brain (Fig. 3). For a given neuropharmacological
agent, its ability to gain access to the brain through the BBB by
passive diffusion is roughly related to its overall lipophilicity and is
reflected by its n-octanol-water partition coefficient (log P value).
Log P values around 2 are usually considered as satisfactory to
cross the BBB. Consequently, the log P values of compounds 1a,
1b and 1e were calculated.9 As depicted in Fig 3, log P values
ranged from 1.88 to 2.2 indicating that compounds 1a, 1b and 1e
are lipophilic enough to cross the BBB. One can anticipate that
the presence of the stabilizing phenyl ring, installed to protect
the 5,6-double bond from hydration, previously observed with
1,4-dihydropyridines, may also hamper the required bio-oxidation
step of the carrier once in the brain. To overcome this problem,
two electron-donating methoxy groups were installed to restore
adequate reducing properties of the carrier. Two strategies were
investigated to connect GABA to the quinoline moiety leading
either to an amide or ester linkage between the drug and the
carrier. In such systems, release of GABA derivatives from 1b or
1d would be ensured via in vivo hydrolysis in the brain by amidase
or esterase respectively (Fig 3).
Fig. 1 Bodor’s chemical delivery system (CDS) by means of 1,4-
dihydropyridines and 1,4-dihydroquinolines as new candidates.
To further explore the potential of such 1,4-dihydroquinolines
as carriers, we report herein the labelling of 1a with carbon-11,
a radionuclide used in Positron Emission Tomography (PET).
PET is an imaging technique that has become a powerful
scientific tool probing biochemical processes in vivo. For this
purpose, we achieved the radiosynthesis of N-[11C]methyl-1,4-
dihydroquinoline 1a and studied its biological behavior in rats
(Fig. 2a). This approach has consisted in evaluating the ability of
[11C]1a to cross the BBB and to be subsequently enzymatically
oxidized and “locked-in” in the brain as its corresponding
quinolinium salt. As an illustrative application of this new 1,4-
dihydroquinoline-type redox carrier system, we also reported
its coupling with g-aminobutyric acid (GABA) derivatives, an
inhibitory neurotransmitter well-known for not crossing the BBB
owing to its hydrophilic nature and the absence of active transport
system.7 In vivo studies in mice are reported herein to give evidence
Fig. 3 Design of the redox CDS (lipophilicity, redox potential, linker).
Preparation of 1,4-dihydroquinoline carrier 1a and
1,4-dihydroquinoline-GABA derivatives 1b and 1d
Hydrolysis of the available 6,7-dimethoxyquinoline-3-
carbonitrile10 furnished the corresponding carboxylic acid 2
in 85% yield. The latter was subsequently involved in the coupling
with benzylamine, GABA ethyl ester and valinol by means of
oxalyl chloride to yield respectively amides 3a–c in fair to good
yields. The amidoalcohol intermediate 3c was further coupled
with N-Boc GABA in the presence of DCC/DMAP to afford 3d
in 72% yield (Scheme 1).
Fig. 2 Design of a new 1,4-dihydroquinoline carrier: (a) radiolabeled
model [11C]1a, (b) illustrative application to brain delivery of GABA.
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of the quinoline 3a with [11C]methyl triflate12 afforded the qua-
ternary salt [11C]4a with an incorporation of [11C]CH3OTf up to
90%. Reduction of [11C]4a by using sodium dithionite and sodium
carbonate led to the corresponding 1,4-dihydroquinoline [11C]1a
with a radiochemical yield ranging from 35 to 66% based on
initial [11C]methyl triflate (Fig. 4). After HPLC purification and
formulation, [11C]1a was obtained with a radiochemical purity
higher than 99%.
Scheme 1 Preparation of quinoline amides 3a–d. Reagents and condi-
tions: (a) 25% aqueous NaOH/EtOH/reflux/20h (85%); (b) (COCl)2/
CH2Cl2/few drops of DMF/8h/rt/then benzylamine/NEt3/-10◦C to
rt/5h (90%); (c) (COCl)2/CH2Cl2/few drops of DMF/2h/rt/then
GABA ethyl ester HCl/NEt3/CH2Cl2/rt (67%); (d) (COCl)2/CH2Cl2/few
drops of DMF/2h/rt/then valinol/NEt3/-10◦C to rt/16h (70%);
(e) DCC/DMAP/CH2Cl2/rt/16h (75%).
The synthesis was completed by quaternization of quinoline
amides 3a,b and 3d followed by regioselective 1,4-reduction of
the corresponding quinolinium salts 4a,b and 4d (Scheme 2).
The quaternization step occurred smoothly at room temperature
in the presence of methyltrifluoromethanesulfonate as a methy-
lating agent to afford quinolinium salts 4a,b and 4d in almost
quantitative yields. The reduction took place regioselectively by
using sodium dithionite under basic conditions affording the
desired 1,4-dihydroquinolines 1a,b in excellent yields and 1d in
somewhat lower yield (60%). It is worth noting that both 1,4-
dihydroquinolines 1a,b could be stored several days either at
the solid state or in chloroform without any degradation being
observed. The higher chemical stability of 1,4-dihydroquinoline
derivatives compared to their 1,4-dihydropyridine analogues has
already been reported11 and may constitute a real advantage for
further in vivo development of these potential drug carriers.
Fig.
radiosynthesis.
4
Radiochromatogram from HPLC analysis after [11C]1a
Biological investigations of [11C]1a in rats
In vitro study. The chemical stability of [11C]1a towards
oxidation was firstly investigated in vitro on blood samples to
establish whether it is stable enough to start developing an ex vivo
evaluation. To carry out this investigation, [11C]1a was added to rat
blood and after treatment, the plasma samples were analyzed using
reversed phase HPLC analysis. The chromatograms obtained at
various time points (5, 10 and 20 min) did not show any trace
of the quaternary form [11C]4a. Additionally, no by-products
which could have arisen from hydration reactions were detected
indicating that [11C]1a exhibits good stability in rat blood.
Ex vivo evaluation. To evaluate the potential of 1,4-
dihydroquinoline 1a as a carrier in vivo, rats were injected into
the tail vein with [11C]1a and sacrificed at different time intervals
(1, 2.5, 5, 10 and 20 minutes) post injection. Then, the brain
penetration of [11C]1a through the BBB and its oxidation to the
corresponding “quaternary form” [11C]4a expected in vivo were
examined. The results indicated a moderate but appreciable and
rapid penetration of [11C]1a into the CNS wherein the radioactivity
brain uptake ranged from 0.78% 0.19 ID/g at 1 min to 0.24%
0.04 ID/g at 20 min (Fig. 5). The oxidation rate of [11C]1a to the
Scheme 2 Access to 1,4-dihydroquinolines 1a–b and 1d. Reagents
and conditions: (a) MeOTf/CH2Cl2/rt/2h. (b) Na2S2O4/NaHCO3/
MeOH/H2O/rt/2h.
Radiosynthesis of [11C]1a. The radiosynthesis of [11C]1a was
carried out as illustrated in Scheme 3. The quaternization reaction
Scheme
3
Radiosynthesis of [11C]1a. Reagents and conditions:
(a) [11C]MeOTf/CH3CN/rt/7min (b) Na2S2O4/H2O/rt/5min then
Na2CO3/H2O/rt/3min.
Fig. 5 Time course for radioactivity expressed as %ID/g in brain after
i.v. injection of [11C]1a in rat.
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corresponding quinolinium salt [11C]4a was monitored by HPLC
analysis from brain homogenates over a period of 20 min. Our
findings showed the disappearance of [11C]1a beyond 5 min post
injection in favour of the quaternary form [11C]4a present at 100%
which revealed a rapid in vivo oxidation of the carrier in the brain
(Fig. 6).
inhibition of LMA could be observed after administration of
GABA at a dose of 150 mg/kg. These findings provide further
evidences that 1,4-dihydroquinoline-GABA derivative 1b, after
i.p. administration, is able to reach the brain and exert a central
activity after having passed the BBB.
In vitro study. Two hypotheses may be put forward to account
for the observed enhancement of central GABAergic activity; (i)
1,4-dihydroquinoline-GABA derivative 1b or the corresponding
quinolinium salt 4b act as true ligands for GABA receptors; (ii) a
delivery mechanism whereby, once in the brain, GABA is released
from the quinoline carrier through enzymatic hydrolysis at the
amide function. To partly address this question, in vitro assays to
assess the interaction of 1,4-dihydroquinoline-GABA derivative
1b and the corresponding quinolinium salt 4b with GABAA
receptors were performed by measuring their ability to displace
[3H] muscimol from GABAA receptors of rat cortical membrane
preparations. It was found that both compounds 1b and 4b failed
to displace [3H] muscimol at the two concentrations tested (i.e.
1 mM and 10 mM). At this stage, although these preliminary in vitro
assays do not rule out binding of 1b and/or 4b to other GABA
receptors which could account for the observed enhancement of
GABAergic activity, the failure of 1b and 4b to displace [3H]
muscimol binding argue for a release of GABA from the redox
CDS in the CNS.
Fig. 6 Percentage of [11C]1a and [11C]4a in rat brain.
A control experiment was deemed necessary to confirm that the
oxidized form [11C]4a does not cross the BBB. As expected, after
peripheral administration of [11C]4a in rats, no radioactivity could
be detected in the brain confirming that [11C]4a is unable to cross
the BBB by passive diffusion or by means of specific transporters.13
Conclusion
To conclude, we reported the first biological evaluation of 1,4-
dihydroquinoline derivatives as new carriers for specific brain
delivery. The radiosynthesis of [11C]1a and an ex vivo evaluation in
rats showed not only that a moderate but significant penetration
of the carrier in the CNS was observed, but also that once in
the brain, the 1,4-dihydroquinoline [11C]1a was rapidly oxidized
to the quaternary form [11C]4a. An in vivo evaluation of 1,4-
dihydroquinoline-GABA derivative 1b, based on alterations of
locomotor activity (LMA) in mice, demonstrated the potential
of such 1,4-dihydroquinolines to transport neuroactive drugs in
the CNS at a sufficiently high concentration and duration to
exert a pharmacological effect. These promising findings designate
1,4-dihydroquinoline derivatives as appealing chemical tools for
targeting neuroactive drugs to the brain. The potential of such
1,4-dihydroquinolines is currently exploited in the design of
original “bio-oxidative” prodrugs for specific delivery of acetyl-
cholinesterase inhibitors to the CNS14 and in the targeting of PET
radioligands for brain imaging.15
Biological investigations of 1b in mice
Locomotor activity. The next aspect of this study was to evalu-
ate the CNS activity of the 1,4-dihydroquinoline-GABA derivative
1b by examining diminution of locomotor activity (LMA) in
mice associated with enhancement of central GABAergic activity
(Fig. 7). Compound 1d was revealed to be insoluble in water and
was unfortunately excluded from this in vivo assay. Compound 1b
was injected intraperitoneally (i.p.) into mice at doses from 50–
150 mg/kg and after 30 min the vertical and horizontal LMA was
recorded during 1 hour. While a slight but significant decrease
in LMA was noted at 50 and 100 mg/kg, LMA was severely
affected at 150 mg/kg. At this dose, although not substantial, the
diminution in LMA was still evident after 90 min, while LMA was
completely restored after 120 min. As expected, no significant
Experimental
General methods
Melting points (◦C) were determined with a WME Ko¨fler
apparatus. IR spectra were recorded on a PERKIN ELMER
IRFT 1650 spectrometer. Liquids were applied as a film between
KBr windows and solids were dispersed in KBr tablets. Absorption
bands are given in cm-1. NMR spectra were recorded on BRUKER
1
AVANCE-300 (300 MHz) spectrometer. H at 300 MHz, 13C at
75 MHz at 282.5 MHz using CDCl3 or DMSO-d6 as solvents
and with the residual solvent signals as internal standards unless
otherwise indicated. The following abbreviations are used to
Fig. 7 Effect of 1,4-dihydroquinoline-GABA 1b on locomotor activity
(LMA) in mice.
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describe peak pattern: s (singlet), d (doublet), t (triplet), q
(quartet), m (multiplet), br (broad). Elemental analyses were
carried out at the University of Rouen (Microanalytical Service
Laboratory) on a CARLO ERBA 1160. Measurement accuracy
is around 0.4% on carbon. Mass spectra (EI, CI, FAB) were
recorded on JEOL JMS AX-500. Analytical thin layer chro-
matography (TLC) was performed on silica gel plates (Kieselgel
60F254) from Merck laboratory. Flash chromatography on silica
gel was carried out using silica (70–230 mesh ASTM) from Merck
laboratory. In the radiochemistry, the labelled compounds were
isolated on a HPLC system equipped with a Waters 501 pump, a
spectrophotometer (l = 254 nm) coupled with a Geiger-Mu¨ller
probe (SDS). Radiochemical purity analyses were performed on
a Merck HPLC system coupled with a NaI radioactive detector
(Novelec). For each labeled compound, the characteristics of the
column and solvent used will be described. The identities of the
labeled compounds were confirmed by their co-elution with
the unlabeled reference compound in the HPLC system.
4-aminobutyrate hydrochloride (0.62 g, 3.7 mmol) and triethy-
lamine (1 mL, 7.1 mmol). Purification of the crude product by flash
chromatography on silica gel (eluent: dichloromethane/ethanol
9:1) yielded 3b as a white solid (0.79 g, 67%); mp 129 ◦C; 1H NMR
(CDCl3, 300 MHz) d 9.07 (1H, s), 8.47 (1H, s), 7.44 (1H, s), 7.09
(1H, s), 6.91 (1H, brs), 4.16 (2H, q, J = 7.1 Hz), 4.05 (3H, s),
4.02 (3H, s), 3.60 (2H, q, J = 7.1 Hz), 2.50 (2H, t, J = 7.1 Hz),
2.02 (2H, quint, J = 7.1 Hz), 1.25 (3H, t, J = 7,1 Hz); 13C NMR
(CDCl3, 75 MHz) d 166.3, 159, 9, 154.2, 150.8, 147.1, 146.2, 134.2,
125.8, 123.1, 108.2, 106.0, 61.2, 56.6, 56.5, 40.4, 32.6, 24.6, 14.6.
IR (KBr) n = 3282, 2915, 1730, 1664, 1547, 1505, 1430, 1250,
1186, 1008 cm-1; MS(ESI+) m/z: 715 [2M + Na+], 692.93 [2M +
H+], 347.33 [M + H+]; Anal. Calcd for C18H22N2O5: C, 62.42; H,
6.40; N, 8.09. Found: C, 62.39; H, 6.54; N, 8.08.
N-(1-Hydroxy-3-methylbutan-2-yl)-6,7-dimethoxyquinoline-3-
carboxamide (3c). Prepared according to procedure A from
oxalyl chloride (2.92 mL, 34 mmol), carboxylic acid 2 (2.0 g,
8.58 mmol), rac-valinol (1.32 g, 12.9 mmol) and triethylamine
(1.32 mL, 9.44 mmol). Purification of the crude product by
flash chromatography on silica gel (dichloromethane/ethanol 9:1)
afforded 3c (1.89 g, 70%) as a white solid; mp: 191 ◦C; 1H NMR
(DMSO-d6, 300 MHz) d 9.07 ( 1H, d, J= 1.9Hz), 9.64 (1H, d, J=
1.9 Hz), 8.21 (1H, d, J= 9.0 Hz), 7.43 (2H, s), 4.63 (1H, t, J=
5.6 Hz), 3.96 (3H, s), 3.92 (3H, s), 3.90 (1H, m), 3.53 (2H, m), 1.90
(1H, m), 0.92 (6H, t); 13C NMR (DMSO-d6, 75 MHz) d 165.7,
153.6, 150.2, 146.9, 146.1, 133.9, 126.2, 122.4, 107.7, 106.6, 61.6,
57.0, 56.2, 56.1, 28.9, 20.0, 19.0; Anal. Calcd for C17H22N2O4: C,
64.13; H, 6.97; N, 8.80; Found: C, 64.34; H, 6.56; N, 8.67.
6,7-Dimethoxyquinoline-3-carboxylic acid (2). A mixture of
6,7-dimethoxyquinoline-3-carbonitrile10 (2.55 g, 11.9 mmol) and
25% aqueous sodium hydroxide (1.47 mL) in ethanol (50 mL) was
refluxed with stirring for 20h. After cooling, the reaction mixture
was acidified with diluted hydrochloric acid. The precipitate was
filtered, dried and purified by recrystallization from EtOH/H2O
(1:1) to yield 2 as a white solid (2.4 g, 85%); mp 262 ◦C; 1H NMR
(DMSO-d6, 300 MHz) d 8.96 (1H, s), 8.61 (1H, s), 7.42 (1H, s),
7.31 (1H, s), 3.83 (3H, s), 3.79 (3H, s); 13C NMR (DMSO-d6,
75 MHz) d 167.0, 154.4, 150.3, 147.8, 147.1, 136.5, 122.5, 121.9,
107.7, 107.0, 56.2, 56.1. IR (KBr) n = 3378, 3062, 1727, 1657,
1614, 1509, 1439, 1381, 1259, 1150, 993 cm-1.
4-tert-Butyloxycarbonylaminobutyric acid 2-(6,7-dimethoxy-3-
carbonylaminoquinoline)-3-methylbutyl ester (3d). BocGABA
(13.0 mg, 0.06 mmol), DMAP (7.7 mg, 0.06 mmol) and DCC
(13.0 mg, 0.06 mmol) were successively added to a solution of
compound 3c (16 mg, 0.05 mmol) in dichloromethane and the
resulting mixture was stirred overnight at room temperature. The
reaction mixture was subsequently filtered. Flash chromatography
using ethyl acetate as eluent provided the desired compound 3d
(19 mg, 75%) as a colorless oil; 1H NMR (CDCl3, 300 MHz) d 9.09
(1H, d, J= 1.3 Hz), 8.46 (1H, d, J= 1.3 Hz), 7.39 (1H, s), 7.33 (1H,
d, J= 8.5 Hz), 7.04 (1H, s), 4.78 (1H, brs), 4.49 (1H, d, J= 9.6 Hz),
4.23 (2H, m), 4.01 (3H, s), 3.98 (3H, s), 3.22 (1H, m), 2.97 (1H,
m), 2.31 (2H, m), 2.04 (1H, m), 1.70 (2H, m), 1.36 (9H, s), 1.04
(6H, d, J= 6.0 Hz); 13C NMR (CDCl3, 75 MHz) d 173.6, 166.5,
156.7, 154.1, 150.6, 147.0, 146.8, 134.3, 126.0, 123.0, 108.1, 106.1,
79.8, 64.6, 56.6, 56.5, 55.2, 39.7, 31.3, 29.4, 28.7, 26.0, 19.9; IR
(KBr) n = 3321, 2968, 1743, 1692, 1641, 1502, 1245, 1171, 1007,
755 cm-1; MS(ESI+) m/z: 1045.00 [2MK+], 1029.13 [2MNa+],
1007.13 [2MH+], 526.33 [MNa+], 504.27 [MH+]; HRMS (IC+) m/z
[M + H+]: calcd for C26H38N307, 504.2710; found, 504.2698.
N-Benzyl-6,7-dimethoxyquinoline-3-carboxamide (procedure A)
(3a). Oxalyl chloride (0.54 mL, 6.44 mmol) was added to a
solution of carboxylic acid 2 (0.5 g, 2.15 mmol) in dichloromethane
(30 mL) under nitrogen followed by one drop of freshly distilled
DMF. After stirring at room temperature for 2h, dichloromethane
was removed under vacuum to afford the acyl chloride interme-
diate. The residue was dissolved in dichloromethane (30 mL) and
was added to a solution of benzylamine (0.3 mL, 2.15 mmol)
in dichloromethane pre-cooled to -10 ◦C. Triethylamine (0.3 mL,
2.15 mmol) was then added and the solution was stirred for 5 hours
at room temperature after which solvent was removed. Water was
added and the pH of the aqueous layer was adjusted to 5. The
organic layer was then washed with water (3 ¥ 100 mL) before
being dried over MgSO4 and concentrated. Flash chromatography
(eluent: dichloromethane/ethanol 9:1) provided the pure product
3a (0.62 g, 90%) as a white solid; mp: 168 ◦C; 1H NMR (CDCl3,
300 MHz) d 9.06 (1H, d, J= 1.9 Hz), 8.47 (1H, d, J= 1.9 Hz),
7.43 (1H, s), 7.37 (5H, m), 7.08 (1H, s), 6.60 (1H, brs), 4.71 (2H,
d, J= 5.6 Hz), 4.04 (3H, s), 4.01 (3H, s). 13C NMR (CDCl3,
75 MHz) d 166.3, 154.3, 150.8, 147.1, 146.1, 138.4, 134.3, 129.2,
128.4, 128.1, 125.7, 123.0, 108.1, 106.0, 56.6, 56.5, 44.6; IR (KBr)
n = 3265, 1657, 1501, 1246, 1146, 1012, 701 cm-1; Anal. Calcd for
C19H18N2O3: C, 70.79; H, 5.63; N, 8.69. Found: C, 70.72; H, 5.87;
N, 8.62.
3-(Benzylaminocarbonyl)-6,7-dimethoxy-1-methylquinolinium
triflate (procedure B) (4a). To a solution of quinoline 3a
(0.50 g, 1.5 mmol) in dichloromethane was added methyl trifluo-
romethanesulfonate (0.19 mL, 1.7 mmol) previously treated with
K2CO3 to avoid the presence of trifluoromethane sulfonic acid.
The mixture was stirred at room temperature for 2 hours. The
solvent was then removed under vacuum. Ether or heptane was
added to a solution of the quinolinium salt in dichloromethane.
The precipitate was filtered and dried to yield the pure quinolinium
Ethyl 4-(6,7-dimethoxyquinoline-3- carbonylamino)butanoate
(3b). Prepared according to procedure A from oxalyl chloride
(1.15 mL, 13 mmol), carboxylic acid 2 (0.79 g, 3.4 mmol), ethyl
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◦
1
salt 4a (0.70 g, 98%) as a white solid; mp: 191 C; H NMR
(CDCl3, 300 MHz) d 9.32 (1H, s), 9.31 (1H, s), 8.70 (1H, brs, J=
5.6 Hz), 7.52 (1H, s), 7.31 (6H, m), 4.62 (2H, d, J= 5.6 Hz), 4.51
(3H, s), 4.23 (3H, s), 4.06 (3H, s); 13C NMR (CDCl3, 75 MHz) d
161.7, 159.6, 152.9, 142.9, 138.5, 137.7, 128.9, 128.6, 127.7, 126.5,
126.2, 108.4, 98.0, 90.0, 58.2, 57.3, 46.3, 44.5; IR (KBr) n = 3344,
3064, 1660, 1511, 1274, 1167, 1032, 640 cm-1; Anal. Calcd for
C21H21F3N2O6S: C, 51.85; H, 4.35; N, 5.76; S, 6.59. Found: C,
51.76; H, 4.36; N, 5.72; S, 6.38.
148.2, 144.7, 140.6, 139.2, 132.9, 128.8, 128.0, 127.5, 113.1, 98.2,
97.6, 56.4, 56.3, 43.8, 39.0, 26.6; IR (CHCl3) n = 3392, 1665, 1602,
1578, 1519, 1237, 1101, 1032 cm-1; HRMS (IC+) m/z [M + H+]:
calcd for C20H23N203, 339.1709; found, 339.1715
Ethyl 4-(1,4-dihydro-6,7-dimethoxy-1-methylquinoline-3-carbo-
nylamino)butanoate (1b). Prepared according to procedure C
from NaHCO3 (0.29 g, 3.44 mmol), Na2S2O4 (0.43 g, 2.86 mmol)
and quinolinium salt 4b (0.29 g, 0.57 mmol). Compound 1b (0.20 g,
97%) was obtained as a yellow oil; 1H NMR (CDCl3, 300 MHz)
d 7.20 (1H, s), 6.58 (1H, s), 6.32 (1H, s), 5.55 (1H, brs), 4.11
(2H, q, J = 7.2 Hz), 3.87 (3H, s), 3.82 (3H, s), 3.72 (2H, s), 3.39
(2H, q, J = 7.2 Hz) 3.21 (3H, s), 2.39 (2H, t, J = 7.2 Hz), 1.89
(2H, quint, J = 7.2 Hz), 1.25 (3H, t, J = 7.2 Hz); 13C NMR
(CDCl3, 75 MHz) d 173.1, 167.2, 148.0, 144.3, 139.1, 133.4, 113.7,
113.3, 99.2, 99.0, 60.1, 56.2, 56.1, 51.6, 41.4, 31.1, 26.4, 25.2,
14.5. MS (ESI+) m/z 363 [M + H+], 333, 247, 203, 188, 160, 115;
UV/vis (methanol) 348 nm; HRMS (IC+) m/z [M + H+]: calcd for
C19H27N205, 363.1920; found, 363.1927
3-(Ethoxycarbonylpropylaminocarbonyl)-6,7-dimethoxy-1-me-
thylquinolinium triflate (4b). Quinolinium salt 4b (0.56 g, 97%)
was obtained from quinoline 3b (0.40 g, 1.15 mmol) and methyl
trifluoromethanesulfonate (143 mL, 1.26 mmol) according to
◦
1
procedure B. mp 152 C; H NMR (CDCl3, 300 MHz) d 9.43
(1H, s), 9.30 (1H, s), 8.31 (1H, brt, J = 5.3 Hz), 7.55 (1H, s),
7.51 (1H, s), 4.69 (3H, s), 4.25 (3H, s), 4.12 (2H, m), 4.06 (3H,
s), 3.48 (2H, m), 2.41 (2H, t, J = 7.5 Hz), 1.98 (2H, quint, J =
7.2 Hz), 1.24 (3H, t, J = 7.1 Hz); 13C NMR (CDCl3, 75 MHz) d
173.6, 161.8, 159.6, 153.0, 144.3, 142.9, 137.9, 126.6, 126.3, 108.5,
98.1, 60.8, 58.2, 57.3, 45.5, 40.2, 32.0, 24.7, 14.5. IR (KBr) n =
3408, 2940, 1722, 1678, 1511, 1266, 1028 cm-1; Anal. Calcd for
C20H25F3N2O8S: C, 47.06; H, 4.94; N, 5.49; S, 6.28. Found: C,
47.03; H, 4.96; N, 5.54; S; 6.30; UV/vis (methanol) 262, 362 nm;
MS(ESI+) m/z: 361.33 [M-TfO-]+.
4-tert-Butyloxycarbonylaminobutyric acid 2-(1,4-dihydro-6,7-
dimethoxy-1-methyl-3-carbonylaminoquinoline)-3-methylbutyl ester
(1d). Prepared according to procedure C from NaHCO3 (206 mg,
2.4 mmol), Na2S2O4 (308 mg, 2.0 mmol) and quinolinium salt 4d
(273 mg, 0.4 mmol). Compound 1d (124 mg, 60%) was obtained
as a yellow oil; 1H NMR (CDCl3, 300 MHz) d 7.30 (1H, s), 6.64
(1H, s), 6.37 (1H, s), 5.53 (1H, d, J= 8.3 Hz), 4.84 (1H, brs),
4.23 (3H, m), 3.92 (3H, s), 3.88 (3H, s), 3.73 (2H, s), 3.27 (3H, s),
3.16 (2H, m), 2.40 (2H, m), 1.82 (3H, m), 1.47 (9H, s), 1.01 (6H,
m). HRMS (IC+) m/z [M + H+]: calcd for C27H42N307, 520.3023;
found, 520.3031
N-(1-tert-Butyloxycarbonylaminopropylcarbonyloxy-3-methyl-
butan-2-yl)-3-carbonylamino-6,7-dimethoxy-1-methyl quinolinium
triflate (4d). Quinolinium salt 4d (0.14 g, 82%) was obtained from
quinoline 3d (0.12 g, 0.25 mmol) and methyl trifluoromethane-
sulfonate (28 mL, 0.25 mmol) as a colourless translucent solid,
according to procedure B. mp: 87 ◦C; 1H NMR (DMSO-d6,
300 MHz) d 9.52 (1H, s), 9.32 (1H, s), 8.82 (1H, brd, J= 7.9 Hz),
7.87 (1H, s), 7.69 (1H, s), 6.80 (1H, brt, J= 5.0 Hz), 4.80 (3H, s),
4.30 (1H, m), 4.17 (3H, s), 4.14 (2H, m), 4.02 (3H, s), 2.86 (2H,
m), 2.26 (2H, t, J= 7.3 Hz), 1.94 (1H, m), 1.56 (2H, m), 1.30 (9H,
s), 0.99 (6H, d). 13C NMR (DMSO-d6, 75 MHz) d 173.0, 162.8,
158.0, 155.9, 151.9, 145.7, 141.8, 137.3, 126.0, 125.2, 108.6, 99.2,
77.8, 64.2, 57.8, 56.9, 55.2, 54.4, 45.8, 31.2, 29.4, 28.5, 25.2, 19.7,
19.0; IR (KBr) n = 3355, 2974, 1737, 1705, 1667, 1513, 1281, 1165,
1030 cm-1; Anal. Calcd for C28H40F3N3O10S: C, 50.37; H, 6.04; N,
6.29; S, 4.80. Found: C, 50.39; H, 6.02; N, 6.36; S, 4.74; MS(ESI+)
m/z: 518.33 [M-TfO-]+.
[11C]Methyl triflate12. [11C]Carbon dioxide was produced by
the 14N(p,a)11C nuclear reaction of a gas target filled with nitrogen
containing 0.5% oxygen at pressure of 20 bars irradiated with
16 MeV protons (IBA Cyclone 18/9 cyclotron). The [11C]carbon
dioxide formed was carried by a nitrogen flow through P2O5 and
collected in a stainless loop immersed in liquid argon. Then the
loop was removed from the liquid nitrogen and brought back
to room temperature. The [11C]carbon dioxide was carried by
nitrogen flow into a reactor maintained at 0 ◦C and containing
LiAlH4 in tetrahydrofuran (1M, 200 mL) the◦n tetrahydrofuran
◦
was removed at 140 C and after cooling at 0 C, hydriodic acid
(1.0 mL, 54%) was added. The [11C]methyl iodide was distilled
under stream of nitrogen at 140 ◦C and passed at 200 ◦C through
an oven containing silver triflate-impregnated graphitized carbon
and the [11C]methyl triflate was obtained.
N-Benzyl-1,4-dihydro-6,7-dimethoxy-1-methylquinoline-3-car-
boxamide (procedure C) (1a). A mixture of NaHCO3 (50 mg,
0.62 mmol) and Na2S2O4 (77 mg, 0.51 mmol) was added to a
solution of quinolinium salt 4a (50 mg, 0.10 mmol) dissolved in
a mixture of water (4 mL) and methanol (4 mL) deaerated with
nitrogen. The mixture was stirred in darkness, at room temperature
for 2h. The same amounts of NaHCO3 and Na2S2O4 were added
and the mixture was stirred for an additional time of 30 min.
After addition of ethyl acetate (10 mL) and phase separation, the
aqueous layer was extracted with ethyl acetate (3 ¥ 10 mL) and
the combined organic phases were washed with water (3 ¥ 10 mL).
After drying over MgSO4, solvent was removed under vacuum,
giving 1a (32.7 mg, 94%) as a yellow oil; 1H NMR (CDCl3,
300 MHz) d 7.29 (6H, m), 6.55 (1H, s), 6.34 (1H, s), 5.67 (1H,
brt, J = 5.7 Hz), 4.56 (2H, d, J = 5.7 Hz), 3.88 (3H, s), 3.81 (3H,
s), 3.75 (2H, s), 3.22 (3H, s); 13C NMR (CDCl3, 75 MHz) d 167.7,
N-Benzyl-1,4-dihydro-6,7-dimethoxy-1-[11C]methylquinoline-3-
carboxamide ([11C]1a). [11C]Methyl triflate was trapped at room
temperature into a reactor containing a solution of quinoline 3a
(1 mg, 3.1 mmol) in acetonitrile (150 mL). When all the radioactivity
was collected, a solution of Na2S2O4 (4 mg, 23 mmol) in water
(150 mL) was added in the reactor. After a time of 5 min, a solution
of Na2CO3 (2 mg, 18.9 mmol) in water (150 mL) was added in
the reactor. The resulting mixture was kept for 3 min at room
temperature. The purification was performed by reverse-phase
semi-preparative HPLC (Macherey Nagel nucleosil 100–5 protect1
column 10 ¥ 250 mm, 4 mL/min, l = 254 nm, CH3CN/H2O/Et3N
40:60:0.01). The fraction containing the labelled product [11C]1a
(tR = 16.5 min) was collected into a flask containing water (25 mL).
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The radioactive solution was transferred onto a Macherey Nagel
chromabond C18ec previously conditioned with methanol (2 mL)
and water (5 mL). The 1,4-dihydroquinoline [11C]1a was eluted
with ethanol (200 mL). To the ethanolic solution was added
1 mL of an isotonic saline solution containing 10% phosphate
buffer (25 mM) pH = 7.4. The final solution was filtered through
a 0.22 mm filter into a sterile vial. The radiochemical and
chemical purities were determined by reverse-phase analytical
HPLC (Macherey Nagel nucleosil 100–5 protect1 column 4.6 ¥
250 mm, 1 mL/min, l = 254 nm, CH3CN/H2O/Et3N 40:60:0.01,
tR = 12.5 min). Radiochemical purity exceeded 99% and [11C]1a
was ready for injection in about 50 min in batches of 185–740 MBq.
The specific activity ranges from 14 to 30GBq/mmol at EOS.
the precipitate was measured to quantify the acetonitrile extraction
efficiency. Supernatants were filtered and injected onto HPLC
(Macherey Nagel nucleosil 100–5 protect1 column 4.6 ¥ 250 mm,
1 mL/min, l = 254 nm, CH3CN/H2O/Et3N 40:60:0.01, [11C]4a :
tR = 3.5 min, [11C]1a : tR = 12.5 min).
Biological investigations of 1b in mice
Animals. Male CD1 mice with body masses of 30–40 g were
housed in groups of 15–20 in Makrolon cages (38 ¥ 24 ¥ 18 cm),
with free access to water and food and kept in a ventilated room at
temperature of 21◦ 1 ◦C, under 12 h light/dark cycles (light on
between 7 a.m. and 7 p.m.). Experiments were carried out between
9 a.m. and 7 p.m. The animals were isolated in small individual
cages (27 ¥ 13 ¥ 13 cm) for 15 min prior testing. In these assays,
8 mice were used in each group (N = 8).
Biological investigations of [11C]1a in rats
Animals
Drugs. Dihydroquinoline-GABA
derivative
1b
(50–
150 mg/kg) was dissolved in dimethylsulfoxide and then
diluted in Cremophor EL and NaCl 0.9% (final concentration:
20% DMSO; 10% Cremophor; 70% NaCl 0.9%). The solutions
of drugs were prepared fresh and injected i.p. in a volume of
10 mL kg-1.
Animal experimental procedures were in accordance with the
recommendations of the EEC (86/609/EEC) and the French
National Committee (decret 87/848) for the care and use of
laboratory animals and were approved by the Normandy Regional
Animal Ethics Committee (saisine N/05–04-05–05). Sprague-
Dawley male rats weighing 250 to 300 g were used in all
experiments. The animals were kept at constant temperature
(22 ◦C) and humidity (50%) with 12 h light/dark cycles and
were allowed free access to food and water until experiment time.
Anesthesia was induced with 5% isoflurane in a gas mixture of
nitrous oxide/oxygen (70/30%) and maintained with 1.5–2.5%
isoflurane during the entire surgical procedure. Body temperature
was monitored rectally and kept close to 37.5 ◦C. A catheter was
inserted into the tail vein.
Locomotor activity. Locomotor activity was measured with a
Digiscan Animal Activity Monitor system (Ommitech Electronics
Inc., Columbus, OH, USA) which monitored horizontal (locomo-
tion) and vertical (rearing) movements of the animals. The digiscan
analyzer was interfaced with a IBM-PC compatible computer
using Digipro software. The individual compartments (L = 20;
W = 20; H = 30 cm) were put in a dimly lit and quiet room.
Horizontal i.e. locomotion and vertical movements i.e. rearing
were expressed as a number of beams crossed over four 15 min
periods of testing. Mice were injected with increasing doses of
dihydroquinoline 1b (50–150 mg/kg i.p.) or its vehicle and placed
in the actimeters 30 min after injection, for a 60 min test.
In vitro stability in rat blood. Compound [11C]1a (14 MBq)
formulated in a solution containing NaCl 0.9%/phosphate buffer
25 mM pH= 7.4/ethanol (v/v/v 80/10/10) was added to rat
blood. After incubation for 5, 10 and 20 min at 37 ◦C, blood
samples (1 mL) were centrifuged (2000 g, 4 min) and plasma
was mixed with an equivalent volume of acetonitrile containing
0.01% Et3N and centrifuged (2000 g, 10 min). Supernatants
were filtered and injected onto HPLC (Macherey Nagel nucleosil
100–5 protect1 column 4.6 ¥ 250 mm, 1 mL/min, l = 254 nm,
CH3CN/H2O/Et3N 40:60:0.01).
In vitro experiments
Evaluation of the binding of 1b and 4b to GABAA receptors.
The incubation mixture consisted of [3H]-muscimol (94 Ci/mmol,
3 nM final concentration, 100 mL), rat cortical membrane sus-
pension as a GABAA receptor source (2 g/L, 250 mL), phosphate
buffer (550 mL) and a solution of compound 1b or 4b (100 mL).
After a 60.0 min incubation time at room temperature, the reaction
was stopped by rapid vacuum filtration through Whatman GF/C
filter paper (pre-soaked in a solution of polyethylenimine (0.1%)
to reduce binding to filters). Filters were subsequently washed
with ice-cold phosphate buffer (3 ¥ 1.5 mL) and placed overnight
in 3 mL of Ready-Safe scintillation cocktail (Beckman Coulter,
Inc.). Radioactivity was measured using a Wallac liquid scintil-
lation counter. Each experiment was carried out two times. Two
concentration points of compounds 1b and 4b were evaluated: 10-5
and 10-6 M.
The specific binding was determined by subtracting non-specific
binding (in the presence of 1 mM muscimol) from the total
binding (in the absence of 1 mM muscimol and compounds 1b and
4b). The occupancy percentage (or percentage of [3H]-muscimol
displacement) was determined by subtracting non-specific binding
from binding with compounds 1b and 4b at 10-5 or 10-6 M divided
by the specific binding. [3H]-muscimol displacement induced by
Brain uptake. Rats were injected with [11C]1a (18.5–70.3 MBq)
formulated in a solution containing NaCl 0.9%/phosphate buffer
25 mM pH = 7.4/ethanol (v/v/v 80/10/10) and were sacrificed
at 1, 2.5, 5, 10 and 20 min post injection (n = 3 per time).
The whole brain was quickly dissected. The brain sample was
weighed and the radioactivity was measured in a g-counter (Cobra
2 gamma counter, Packard). Data were expressed as the percentage
of injected dose (decay-corrected) per gram of tissue (% ID/g).
Oxidation rate in brain homogenate. Rats were injected with
[11C]1a (18.5–70.3 MBq) formulated in a solution containing
NaCl 0.9%/phosphate buffer 25 mM pH = 7.4/ethanol (v/v/v
80/10/10) and were sacrificed at 1, 2.5, 5, 10 and 20 min post
injection (n = 3 per time). Whole brain was quickly removed and
analysis were carried out on half-brain crushed in acetonitrile
containing 0.01% Et3N (2.5 mL; UltraTurrax T25, Janke and
Kunkel) and centrifuged at 2000 g for 10 min. The radioactivity of
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Table 1
4 (a) S. Gaillard, C. Papamicael, F. Marsais, G. Dupas and V. Levacher,
Synlett, 2005, 441; (b) J-L. Vasse, V. Levacher, J. Bourguignon and
G. Dupas, Chem. Comm., 2002, 2256; (c) J-L. Vasse, V. Levacher, J.
Bourguignon and G. Dupas, Tetrahedron, 2003, 59, 4911; (d) C. Vitry,
J-L. Vasse, G. Dupas, V. Levacher, G. Que´guiner and J. Bourguignon,
Tetrahedron, 2001, 57, 3087; (e) V-L. Vasse, P. Charpentier, V. Levacher,
G. Dupas, G. Que´guiner and J. Bourguignon, Synlett, 1998, 10,
1144.
Solutions
Total
Blank 10-5
M
10-6
M
Phosphate buffer 650 mL 550 mL 550 mL
GABA membrane 250 mL 250 mL 250 mL
Sample
550 mL
250 mL
0
0
0
100 mL (10-4M) 100 mL (10-5 M)
0
Muscimol 10–3 M
100 mL
0
[3H]-muscimol
100 mL 100 mL 100 mL
100 mL
5 C. Patteux, L. Foucout, P. Bohn, G. Dupas, J. Leprince, M-C. Tonon, B.
Dehouck, F. Marsais, C. Papamicael and V. Levacher, Org. Bio. Chem.,
2006, 4, 817.
6 N. Bodor, H. Farag, M. D. C. Barros, W-M Wu and P. Buchwald, J. Dug
Targ., 2002, 10, 63.
compound 1b: 19% at 10-5 M and 6% at 10-6 M. [3H]-muscimol
displacement induced by compound 4b: 11% at 10-5 M and 8% at
10-6 M.
7 (a) For previous in vivo evaluations of 1,4-dihydropyridine-GABA
derivatives able to cross the BBB, see: P. A. Woodard, D. Winwood,
M. E. Brewster, K. S. Estes and N. Bodor, Drug Des. Deliv., 1990, 6,
15; (b) K. Matsuyama, C. Yamashita, A. Noda, S. Goto, H. Noda, Y.
Ichimaru and Y. Gomita, Chem. Pharm. Bull., 1984, 32, 4089; (c) V.
Carelli, F. Liberatore, L. Scipione, G. Giorgioni, A. Di Stefano, M.
Impicciatore, V. Ballabeni, F. Calcina, F. Magnanini and E. Barocelli,
Bioorg. Med. Chem. Lett., 2003, 13, 3765.
8 For correlation between alteration of locomotor activity and central
GABAergic activity, see: Sk. Jamaluddin and M. K. Poddar, Neuro-
chemical Research, 2001, 26, 439.
9 Prior to log P calculation, the geometry of the molecule was fully
optimized at the PM3 level of theory as implemented in the Cerius2
package (Accelrys inc.). The volume of the molecule was then computed
and used in the formula published by: N. Bodor and P. Buchwald,
J. Phys. Chem. B., 1997, 101, 3404.
Acknowledgements
We thank the CNRS, the CEA, the re´gion Haute-Normandie,
the re´gion Basse-Normandie and the CRIHAN for financial
and technical support. The authors thank Olivier Tirel, Me´ziane
Ibazize`ne and Je´roˆme Delamare for their technical assistance in
production of [11C]1a.
Notes and references
1 (a) For leading reviews on Bodor’s chemical delivery systems, see: N.
Bodor and P. Buchwald, Adv. Drug Deliv. Rev., 1999, 36, 229; (b) L.
Prokai, K. Prokai-Tatrai and N. Bodor, Med. Res. Chem., 2000, 20,
367; (c) N. Bodor and P. Buchwald, Burger’s Medicinal Chemistry and
Drug Discovery, 2, 534 chapiter 5; (d) N. Bodor and L. Prokai, Drug
Discov. Today, 2002, 7, 766.
2 (a) N. Bodor, H. Farrag and F. Omar, Bull. Pharm. Sci., 1993, 16, 175;
(b) E. Pop, T. Loftsson and N. Bodor, Pharm. Res., 1991, 8, 1044.
3 (a) M. Sheha, A. Al-Tayeb, H. El-Sherief and H. Farag, Bioorg. Med.
Chem., 2003, 11, 1865; (b) Magda A. El-Sherbeny, Huda S. Al-Salem,
Maha A. Sultan, MAhasen A. Radwan, Hassan A. Farag and Hussein
I. El-Subbagh, Arch. Pharm. Pharm. Med. Chem., 2003, 445; (c) H.
A. Hassan, M. Abdel-Aziz, Gamal El-Din A. A. Abuo-Rahma and
Hassan H. Farag, Bioorg. Med. Chem., 2009, 17, 1681.
10 P. Charpentier, V. Lobre´gat, V. Levacher, G. Dupas, J. Bourguignon
and G. Que´guiner, Tetrahedron. Lett., 1998, 39, 4013.
11 M. E. Brewster, A. Simay, K. Czako, D. Winwood, H. Farag and N.
Bodor, J. Org. Chem., 1989, 54, 3721.
12 D. M. Jewett, Int. J. Rad. Appl. Instrum. [A], 1992, 43, 1383.
13 (a) For previous work related to the active transport of permanently
charged pyridinium salts to the brain by means of the BBB choline
transporter, see: Z. Zhang, P. R. Lockman, R. K. Mittapalli, D. D.
Allen, L. P. Dwoskin and P. A. Crooks, Bioorg. Med. Chem. Lett.,
2008, 18, 5622; (b) D. D. Allen, P. R. Lockman, K. E. Roder, L. P.
Dwoskin and P. A. Crooks, Jpet, 2003, 304, 1268.
14 P. Bohn, N. Le Fur, G. Hagues, J. Costentin, N. Torquet, C. Papamicae¨l,
F. Marsais and V. Levacher, Org. Biomol. Chem., 2009, 7, 2612.
15 Thesis F. Gourand, University of Caen, 2007.
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©