3-(15-hydroxypentadecyl)-2,4,4-trimethyl-2-cyclohexen-1-one and its effect on neuropeptide secretion

Article abstract of DOI:10.1016/S0960-894X(00)00508-4

The aim of the present study was to describe the synthesis of a trimethyl cyclohexenonic long chain fatty alcohol (t-CFA), and analyze its biological activity. Specifically, 3-(15-hydroxypentadecyl)-2,4,4-trimethyl-2-cyclohexen-1-one, the t-CFA containing

Full text of DOI:10.1016/S0960-894X(00)00508-4

Bioorganic & Medicinal Chemistry Letters 10 (2000) 2537±2539
3-(15-Hydroxypentadecyl)-2,4,4-trimethyl-2-cyclohexen-1-one
and its E€ect on Neuropeptide Secretion
Celine Girlanda-Junges,a Bernadette Lutz-Bucher,b Jose-Luis Gonzalez de Aguilar,a,b
Jean-Philippe Loe‚erb and Bang Luua,*
a
Â
Laboratoire de Chimie Organique des Substances Naturelles, UMR CNRS 7509, Universite Louis Pasteur, Strasbourg, France
b
Â
Laboratoire de Neurophysiologie Cellulaire et Integree, UMR CNRS 7519, Universite Louis Pasteur, Strasbourg, France
Â
Â
Received 5 July 2000; revised 4 September 2000; accepted 5 September 2000
AbstractÐThe aim of the present study was to describe the synthesis of a trimethyl cyclohexenonic long chain fatty alcohol (t-
CFA), and analyze its biological activity. Speci®cally, 3-(15-hydroxypentadecyl)-2,4,4-trimethyl-2-cyclohexen-1-one, the t-CFA
containing 15 carbon atoms on the side chain (t-CFA n=15) stimulated arginine vasopressin secretion in nerve terminals of the
neurohypophysis. This e€ect was inhibited by extracellular calcium depletion, which suggets that t-CFA n=15 stimulates
neuropeptide secretion through a calcium-dependent exocytosis mechanism. # 2000 Published by Elsevier Science Ltd.
Introduction
to hypothesize that t-CFA n=15 could act at the mem-
brane level.
Since the demonstration of in vitro neurotrophic activ-
ity of n-hexacosanol, a long chain primary alcohol con-
taining 26 carbon atoms,1 we have focused our attention
on long chain fatty alcohols that contain a functional-
ized nucleus in order to improve their physicochemical
and bioavailability properties. Our aim is the search of
substances which are able to promote the development
of the nervous system, and in particular the establishment
of neuronal networks.
Chemistry
The compound 3-(15-hydroxypentadecyl)-2,4,4-tri-
methyl-2-cyclohexen-1-one 1 was synthesized in seven
steps from commercially available geranyl bromide 2.
In an methanol medium, 2 reacted with benzenesul®nic
acid sodium salt to give the corresponding ger-
anylphenylsulfone 3, which was cyclized as described by
Krishna et al.6 and Torri et al.7 to a€ord a mixture of 4
and 5 (85:15) in 90% yield (Scheme 1). The two isomers
were separated by successive recrystallizations in hex-
ane, and the desired 4 was obtained in 74% yield with a
purity over 99%.
Previous studies showed that monomethyl and dimethyl
cyclohexenonic long chain fatty alcohols with an
appropriate length of the side chain (n=14 carbon
atoms for the most active in these series) are potent
inducers of neurite outgrowth.2 In the present report,
we show that 3-(15-hydroxypentadecyl)-2,4,4-trimethyl-
2-cyclohexen-1-one, a trimethyl cyclohexenonic long
chain fatty alcohol containing 15 carbon atoms on the
side chain (t-CFA n=15, 1), with neuritogenic activity,3
is a potent inducer of arginine vasopressin secretion in
nerve terminals of the neurohypophysis, a well-char-
acterized example of neuropeptide exocytosis.4
Although the cellular mechanism underlying this e€ect
is still unclear, the structural homology of these com-
pounds with endogenous long chain fatty alcohols,
which are involved in ether lipid biosynthesis,5 allows us
As shown in Scheme 2, sulfone 4 was coupled with the
unprotected 14-bromo-tetradecan-1-ol in a one-pot pro-
cedure using n-butyllithium in the presence of HMPA to
give 6. After reductive desulfonation by means of 6%
Na/Hg amalgam,8 the resulting alcohol 7 was protected
by an acetate to give 8 which was oxidized to the corre-
sponding a,b-unsaturated ketone 9 using RuCl3 as cata-
lyst (0.7% mol) and 70% tBuOOH.9 The deprotection of
the alcohol by K2CO3 in methanol gave 1 in 91% yield.10
In the same manner, compounds with di€erent side
chain lengths (12 to 18 carbon atoms) were synthesized,
reactivities and yields being similar to those of 1.
*Corresponding author. Tel.: +33-3-88-411672; fax: +33-3-88-607464;
e-mail: luu@chimie.u-strasbg.fr
0960-894X/00/$ - see front matter # 2000 Published by Elsevier Science Ltd.
PII: S0960-894X(00)00508-4

2538
C. Girlanda-Junges et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2537±2539
Biological Activity
described.11 t-CFA n=15 was dissolved in ethanol and
diluted to render a ®nal concentration of 0.05% etha-
nol. All NILs were incubated in the same conditions.
In order to determine whether t-CFA n=15 exhibits any
speci®c activity in neuropeptide secretion, mouse neuro-
intermediate lobes (NILs) were incubated in a static
culture system to quantify arginine vasopressine (AVP)
release. Brie¯y, pituitary glands from male FVB/N mice
were rapidly dissected, and further separated into ante-
rior lobe and NIL under a microscope. NILs were pre-
incubated for 1 h at 37 ꢀC in Krebs±Ringer bicarbonate
bu€er (KRB, pH 7.4) containing (in mM): NaCl 118.5,
KCl 4.75, CaCl2 2H2O 2.5, KH2PO4 1.2, MgSO4 2H2O
1.2, and supplemented with 0.2% glucose and 0.1%
bovine serum albumin. Tissues were then treated with
500 nM t-CFA n=15 in both normal and calcium-free
medium (KRB without CaCl2, and supplemented with
2.5 mM MgCl2, and 1 mM EGTA). After 20 min, med-
ium samples were collected, and stored at 20 ꢀC until
being radioimmunoassayed for AVP as previously
As illustrated in Figure 1, t-CFA n=15 induced a sig-
ni®cant increase (5.09-fold) in AVP secretion. To ascer-
tain whether this stimulatory e€ect was due to
activation of a regulated exocytotic pathway rather than
non-speci®c release, NILs were treated with t-CFA
n=15 in the absence of extracellular calcium. Interest-
ingly, AVP secretion was completely abolished by cal-
cium depletion. The crucial role of calcium in
neuropeptide exocytosis, and the stimulation of AVP
secretion through calcium-dependent mechanisms have
been largely demonstrated.12 Our present ®ndings
showing a calcium-dependent stimulatory e€ect suggest
that t-CFA n=15 could trigger an in¯ux of Ca2+ ions
from the extracellular medium, which in turn would
facilitate neuropeptide secretion.
.
.
Scheme 1. Reagents: (a) PhSO2Na, MeOH, 0 ꢀC, 1 h (80%); (b) H2SO4 32 equiv, AcOH, 12 ꢀC, 30 min (90%).
Scheme 2. Reagents: (a) (1) n-BuLi, THF, HMPA, CHPh3, 78 ꢀC to rt, 1 h; (2) Br(CH2)14OH, THF, 78 ꢀC to 20 ꢀC, 2 h (87%); (b) Na/Hg 6%,
MeOH, Na2HPO4, 0 ꢀC to rt, 3 h (90%); (c) Ac2O, pyridine, rt, 1 h (98%); (d) RuCl3 0.7% mol, tBuOOH 70%, cyclohexane, H2O, rt, 6 h (53%);
(e) K2CO3, MeOH, H2O, rt, 2 h (91%).

C. Girlanda-Junges et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2537±2539
2539
Acknowledgements
This study was partially supported by Meiji Milk Pro-
ducts Co., Ltd (Tokyo, Japan). Dr. J. L. Gonzalez de
Aguilar was the recipient of an Aide aux Etudes from
Association Francaise contre les Myopathies. We thank
Kuraray Co. Ltd (Japan) for supplying a sample of
cyclogeranylphenylsulfone.
References and Notes
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Figure 1. E€ect of t-CFA n=15 on AVP secretion. NILs were treated
with 500 nM t-CFA n=15 in the presence or absence of Ca2+ in the
culture medium. The data represent the meanÆSE (n=4), and they
are expressed as total pg/300 mL. ÃP<0.05 vs control, #P<0.05 vs
t-CFA n=15, determined by one-way ANOVA, followed by Student±
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The neurohypophysis is a key organ implicated,
through release of AVP and oxytocin, in numerous
physiological functions, including osmotic homeostasis,
pregnancy, and lactation.13 Thus, the utilization of tri-
methyl-CFAs as potent neural lobe secretagogues opens
new lines of investigation in biomedical research.
13. Acher, R.; Chauvet, J. Front. Neuroendocrinol. 1995, 16,
237.