Solid-phase synthesis of quinol fatty alcohols, design of N/O-substituted quinol fatty alcohols and comparative activities on axonal growth

Article abstract of DOI:10.1016/j.bmcl.2006.05.027

Following the promising activity of Q₂FA15 on axonal growth, two new series of N/O-substituted QFAs were synthesized, based on a S_N2-type reaction. O-alkylated QFA bearing 14 carbon atoms on the side chain (n = 14) shows a very potent activity on axonal growth though lowered when compared to Q₂FA15. While O-alkylation allows good retention of the biological activity, N-alkylation abolishes it nonetheless. A solid-phase-supported synthesis of Q₂FA15 allowing the conception of new hybrid compounds is also described.

Full text of DOI:10.1016/j.bmcl.2006.05.027

Bioorganic & Medicinal Chemistry Letters 16 (2006) 3917–3920
Solid-phase synthesis of quinol fatty alcohols, design of
N/O-substituted quinol fatty alcohols and comparative
activities on axonal growth
Mazen Hanbali,a Dominique Bagnardb and Bang Luua,*
aLaboratoire de Chimie Organique des Substances Naturelles, LC3-UMR7177 CNRS-ULP,
Universite´ Louis Pasteur, 67084 Strasbourg Cedex, France
`
´
Laboratoire de Physiologie du Systeme Nerveux, U575 INSERM, Universite Louis Pasteur, 67084 Strasbourg Cedex, France
b
Received 1 April 2006; revised 10 May 2006; accepted 11 May 2006
Available online 30 May 2006
Abstract—Following the promising activity of Q2FA15 on axonal growth, two new series of N/O-substituted QFAs were synthe-
sized, based on a SN2-type reaction. O-alkylated QFA bearing 14 carbon atoms on the side chain (n = 14) shows a very potent activ-
ity on axonal growth though lowered when compared to Q2FA15. While O-alkylation allows good retention of the biological
activity, N-alkylation abolishes it nonetheless. A solid-phase-supported synthesis of Q2FA15 allowing the conception of new hybrid
compounds is also described.
Ó 2006 Elsevier Ltd. All rights reserved.
Regeneration of the CNS after injury is very limited due
to the presence in the lesion site of many inhibitory
factors such as myelin-associated proteins (MAG,
OMGp and NOGO),1 Sema3A2 or chondroitin sulfate
proteoglycan.3,4 In a prior publication, we have shown
that quinol fatty alcohols have the ability to promote
axonal growth especially on inhibitory substrates pres-
ent within the CNS.5 Specifically, Q2FA15 is able to pro-
mote axonal growth at 181% relative to control
conditions, while allowing proper axonal growth on
myelin proteins and Sema3A substrates.
Though Q2FA15 is a very active compound, its chemical
synthesis is rather tiresome. Based on a convergent syn-
thetic route, four steps are needed for the synthesis of
the side terminal alkyne chain, while an additional three
steps are required for obtaining the final compound. In
addition, the coupling reaction between the quinol and
the alkyne moieties is based on Sonogashira cross-
coupling9,10 therefore using palladium catalysts.11
Palladium is known to interact with organic compounds
and common purification procedures are often not suffi-
cient to eliminate the excess metal catalyst.12 In order to
circumvent that problem, pharmaceutical industries are
using techniques such as distillation, adsorption,13 crys-
tallization14 and extraction15 thus reducing palladium
levels to acceptable amounts.
X-Alkanol derivatives have proven in recent years to be
very active towards various targets in the CNS.6–8 The
need of a synthetic strategy applicable to differently
substituted x-alkanols is nowadays a crucial aspect of
our research. To address that, a synthetic strategy using
solid-phase organic synthesis was designed in order to
obtain Q2FA15, based on the Sonogashira cross-cou-
pling reaction between an arylbromide and a resin-sup-
ported terminal alkyne derivative.
In this paper, we thereby describe the design of two new
series of O-alkylated and N-alkylated quinol fatty alco-
hols where the carbon–carbon bond between the quinol
and the x-alkanol moieties was replaced by a C–O and
C–N bond, respectively. O/N-alkylated QFAs were test-
ed on a cellular model of axonal growth in order to com-
pare their activity to lead compound Q2FA15, while
their free radical scavenging activity was also evaluated.
Keywords: Quinol fatty alcohols; Solid-phase organic synthesis; SN2-
type reactions; Axonal growth; Myelin proteins; Sema3A.
*
Solid-phase Sonogashira cross-coupling reactions are
usually accomplished between a supported arylhalide/
Corresponding author. Tel.: +33 3 88 41 16 72; fax: +33 3 88 60 74
64; e-mail: luu@chimie.u-strasbg.fr
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.05.027

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M. Hanbali et al. / Bioorg. Med. Chem. Lett. 16 (2006) 3917–3920
triflate and soluble terminal alkynes.16 In our case, sup-
ported terminal alkynes were needed in order to be cou-
pled to a variety of arylhalides/triflates. To do so, we
used a trichloroacetamidate17 Wang resin 5 which was
subsequently substituted by 13-bromotridecan-1-ol in a
mixture of methylene chloride and cyclohexane in acidic
conditions to give resin 6. Terminal alkyne 7 was ob-
tained with lithium acetylide in DMSO.18 The Sono-
gashira cross-coupling reaction was followed by a
cleavage of our compound from the Wang resin with
10% TFA in methylene chloride and water. A final cat-
alytic hydrogenation gave Q2FA15 1 (Scheme 1).
N-alkylated QFAs 3a–d were obtained by a SN2-type
reaction between commercial 2,5-dimethoxyaniline 15
and protected bromoalcohols 12a–d with n-BuLi
in THF,21 followed by deprotection with TBAF22
(Scheme 2).
The C10 and C12 diols were commercially available.
Diol C14 was obtained by reduction of the correspond-
ing diacid with lithium aluminium hydride. Diol C16 is
obtained by reduction of the corresponding lactone with
the same reducing agent.6 Bromoalcohols 11a–d were
obtained by monobromination of the corresponding
diols in a mixture of HBr–cyclohexane.6 The use of
tert-butyldimethylsilyl chloride in methylene chloride23
gave silylated bromoalkanols 12a–d.
O-alkylated QFAs 2a–d were obtained by a SN2-type
reaction between 2,5-dimethoxyphenol 14 and bromoal-
cohols 11a–d with potassium carbonate in acetone.19
2,5-dimethoxyphenol 14 was obtained through
Baeyer–Villiger oxidation of 2,5-dimethoxycarboxalde-
a
In an attempt to study the biological activity of our
heteroatom-alkylated QFAs, we analyzed the ability
of each compound to promote axonal growth on
mouse cortical neurons (Table 1). The experimental
protocol has been described in a prior publication.24
Briefly, mice embryos at the 15th day of gestation
were dissected and cortical extracts were dissociated
with trypsin. The neurons obtained were plated in 6-
well plates and cultured on poly-L-lysine coverslips
for 24 h at 37 °C, 5% CO2. On day 2, neurons were
incubated with the test compound and grown for
24 h at 37 °C, 5% CO2.
hyde 13.20
NH
a
b
OH
O
CCl3
O
13Br
5
6
4
OMe
O
13
c
d
O
13
7
OMe
8
OMe
OMe
On day 3, neurons were fixed and immunostained with a
primary anti-phosphoneurofilament antibody (SMi312,
Sternberger) and a secondary Alexa488-coupled anti-
body. Coverslips were then mounted on plates with
aqua-polymount.
e
f
OH
15
OH
OMe
OMe
13
9
1
A complete screening of all compounds at 10ꢀ9 M
showed that O-alkylated compound (O-QFA14) with a
14-carbon side chain was the most effective, while its
N-alkylated homologue had no apparent biological
activity (Fig. 1).
Scheme 1. Reagents and conditions: (a) CCl3CN, DBU, 0 °C, 67%; (b)
BF3 Æ OEt2, 13-bromotridecan-1-ol, cyclohexane, CH2Cl2, 90%; (c)
lithium acetylide (ethylene diamine complex), 0 °C to rt, 95%; (d) 1-
bromo-2,5-dimethoxyphenol, Pd(PPh3)4, piperidine, 80 °C, 90%; (e)
TFA 10%, CH2Cl2, H2O, rt, 80%; (f) H2, Pd/C 10%, EtOH, 90%.
a
b
Br
OTBDMS
HO
OH
Br
OH
d
n-2
n-2
n-2
10a-d
11a-d
12a-d
OMe
OMe
OMe
OMe
c
a: n=10
b: n=12
c: n=14
d: n=16
H
OH
O
OH
n-2
OMe
OMe O
13
14
2a-d
OMe
OMe
OMe
f
e
N
OH
N
OTBDMS
NH2
n-2
n-2
H
H
OMe
OMe
OMe
15
16a-d
3a-d
Scheme 2. Reagents and conditions: (a) HBr 47%, cyclohexane, reflux, 6 h, 79–89%; (b) TBDMS-Cl, Imid., CH2Cl2, rt, 4 h, 89–95%; (c) i—mCPBA,
CH2Cl2, 0 °C to rt, ii—NaOH 10%, MeOH, 95%; (d) K2CO3, 11a–d, acetone, reflux, 6 h, 62–70%; (e) i—n-BuLi, THF, 0 °C, ii—12a–d, THF, 60 °C,
iii—dioxane, 60 °C, 14 h, 54–65%; (f) TBAF, THF, rt, 4 h, 93–95%.

M. Hanbali et al. / Bioorg. Med. Chem. Lett. 16 (2006) 3917–3920
Table 1. Compounds synthesized and tested
3919
Compound
Compound name
Molecular structure
HRMS (M+H+)
Antioxidant activity
ABTS assay IC50 (lM)
—
Axonal growth
Calculated Found
%/control (1 nM)
100
—
Ethanol
—
—
—
—
1
TroloxÒ
Q2FA15
—
C23H40O3
—
365.3050
—
365.3056
600
>10,000
—
181
2a
2b
2c
2d
O-QFA10
O-QFA12
O-QFA14
O-QFA16
C18H30O4
C
311.2217
339.2530
367.2843
395.3156
311.2211
339.2526
367.2848
395.3159
122
145
165
109
***
***
***
***
20H34O4
C22H38O4
C24H42O4
3a
3b
3c
3d
N-QFA10
N-QFA12
N-QFA14
N-QFA16
C18H31NO3
C20H35NO3
C22H39NO3
C24H43NO3
310.2377
338.2690
366.2847
394.3316
310.2380
338.2701
366.2831
394.3322
100
102
105
104
***
***
***
***
Each compound is tested on the ABTS competition assay. The IC50 values were determined as the concentration of the compound required for a 50%
diminution of the ABTSÅ+
stands for an antioxidant activity that did not reach the IC50. Axonal growth was measured with UTHSCSA Image
.
***
Tool 3.0 and is presented as the mean value of three different experiments. Variation was generally 5%.
In a parallel experiment, we evaluated the ability of
QFAs to scavenge hydroxyl radicals present in an etha-
nolic medium. To do so, we used the 2,20-azinobis-(3-
ethylbenzothiazoline-6-sulfonic acid) (ABTS) competi-
tion assay.25 In the presence of hydroxyl radicals, ABTS
is oxidized to the stable ABTS cation radical (ABTSÅ+)
observed by its absorbance at 405 nm.
ble outcome. A protonated compound would bear an
extra positive charge which would not allow proper
compound–membrane interaction (Fig. 2).
Furthermore, the nitrogen of N-QFA14 could hydrogen
bond to the methoxy moiety of the benzene ring,29 hence
blocking the compound in a certain configuration that
does not allow proper interaction with the cell
membrane.
This method measures the relative ability of C/O/N-
QFAs to scavenge hydroxyl radicals and thus inhibit
the formation of ABTSÅ+ as measured by the decrease
of its absorbance at 405 nm. Results are shown in Table 1.
In this paper, we describe a solid-phase synthesis of
Q2FA15. Though the overall yield is significantly
lower than that of the previously described synthesis5
(Table 2), this approach allows us to obtain our com-
pound in a very straightforward manner using signifi-
cantly less purification steps and less organic solvents.
This synthesis should be easily applicable to the design
of new potentially active hybrid compounds.
Though antioxidant activity of O-QFA1426 (2c) and
N-QFA14 (3c) seems to be higher than that of
Q2FA1527 (1), their biological activity towards regener-
ation of axon extension is affected. O-QFA14 seems to
exert a biological activity that is comparatively similar
to that of Q2FA15 though 10% lower. However,
N-QFA14 is totally inactive towards axonal growth.
This lack of biological activity could be due to proton-
ation of N-QFA14 in our aqueous cell-culture medium.
The syntheses of N/O-alkylated QFAs are on the other
hand overly simple and require fewer steps to obtain
the final compounds. Both syntheses are based on
SN2-type reactions therefore needing no transition metal
catalysts. Though the activity of N-QFAs on axonal
growth is nil, O-QFA14 appears to have the capacity
to promote axonal growth to comparable extents as
Q2FA15.
X-Alkanols are known to interact with the cell mem-
brane and slightly destabilize it.28 Diffusion through
the cell membrane through a flip-flop process is a possi-
Altogether, these data suggest that though C-alkylated
QFAs, and other hybrid compounds, are biologically
Figure 1. Promoting effect of O-QFAs and N-QFAs according to the
side-chain length on a model assay of axonal growth. Compounds are
tested at 10ꢀ9 M. Data are shown as means SEM. ***p < 0.001, ns,
not significant.
Figure 2. Possible configurations of O-QFA14 and protonated
N-QFA14. The difference in membrane position is the most plausible
explanation of why N-QFA14 is an inactive compound.

3920
M. Hanbali et al. / Bioorg. Med. Chem. Lett. 16 (2006) 3917–3920
Table 2. Comparison of the synthetic strategies and biological
activities of C-alkylated, O-alkylated and N-alkylated QFAs
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Number
of steps
Coupling
reaction
Overall
yield (%)
Biological
activity
Q2FA15
3
6
2
3
Sonogashira
Sonogashira
SN2
51
37
65
54
Active
Active
Active
Inactive
spQ2FA15
O-QFA14
N-QFA14
SN2
sp, solid phase.
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