Synthesis and biological activities of novel structural analogues of 2-arachidonoylglycerol, an endogenous cannabinoid receptor ligand

Article abstract of DOI:10.1016/S0960-894X(01)00350-X

Novel analogues of 2-arachidonoylglycerol (2-AG), an endogenous cannabinoid receptor ligand, were developed. Chemical synthesis of these analogues (2-AGA105 and 2-AGA109) was accomplished starting from 2-octyn-1-ol and diethyl malonate and employing Wittig coupling of triene phosphonate with an aldehyde intermediate in a convergent and stereoselective manner. These analogues should be useful lead compounds for the development of novel 2-AG mimetics.

Full text of DOI:10.1016/S0960-894X(01)00350-X

Bioorganic & Medicinal Chemistry Letters 11 (2001) 1985–1988
Synthesis and Biological Activities of Novel Structural
Analogues of 2-Arachidonoylglycerol, An Endogenous
Cannabinoid Receptor Ligand
Yoshitomo Suhara,a Shinji Nakane,b Shunsuke Arai,b Hiroaki Takayama,a
Keizo Waku,b Yoshio Ishimac and Takayuki Sugiurab,*
aDepartment of Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Teikyo University,
Sagamiko, Kanagawa 199-0195, Japan
bDepartment of Hygienic Chemistry and Nutrition, Faculty of Pharmaceutical Sciences, Teikyo University,
Sagamiko, Kanagawa 199-0195, Japan
cIshima Institute for Neurosciences, Kunitachi, Tokyo 186-0002, Japan
Received 28 March 2001; accepted 12 May 2001
Abstract—Novel analogues of 2-arachidonoylglycerol (2-AG), an endogenous cannabinoid receptor ligand, were developed. Che-
mical synthesis of these analogues (2-AGA105 and 2-AGA109) was accomplished starting from 2-octyn-1-ol and diethyl malonate
and employing Wittig coupling of triene phosphonate with an aldehyde intermediate in a convergent and stereoselective manner.
These analogues should be useful lead compounds for the development of novel 2-AG mimetics. # 2001 Elsevier Science Ltd. All
rights reserved.
2-Arachidonoylglycerol (2-AG) (1) is a unique mole-
cular species of monoacylglycerol identified as an endo-
genous cannabinoid receptor ligand by us1 and by
Mechoulam et al.2 several years ago. 2-AG binds to
both central and peripheral cannabinoid receptors (CB1
and CB2)1,2 and exhibits a variety of cannabimimetic
activities in vitro and in vivo.3 Evidence is accumulating
that 2-AG is the intrinsic natural ligand for the canna-
binoid receptors and that the cannabinoid receptors are
primarily 2-AG receptors.4À8
roles, such as the attenuation of neurotransmission and
vascular tone and the modulation of immune response;3
it is apparent that the endogenous ligand 2-AG is an
essential molecule in various mammalian tissues where
the cannabinoid receptors are expressed. Nevertheless,
the physiological roles of 2-AG in mammalian tissues
still remain to be fully clarified, as does the significance
of the cannabinoid receptor.
2-AG is a metabolically labile molecule, like acetylcho-
line, and therefore it is sometimes difficult to evaluate
precisely its biological activities, especially in vivo. It is
therefore important to develop metabolically stable
Cannabinoid receptors are expressed in diverse mamma-
lian tissues, and are assumed to play important regulatory
Figure 1. 2-Arachidonoylglycerol (2-AG) (1) and its structural analogues, 2-AG ether (2), 2-AGA105 (3), and 2-AGA109 (4).
*Corresponding author. Tel.: +81-426-85-3748; fax: +81-426-85-1345; e-mail: sugiurat@pharm.teikyo-u.ac.jp
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00350-X

1986
Y. Suhara et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1985–1988
Scheme 1. (a) EtMgBr, CuI; (b) Ph2PCH2CH2PPh2ꢀ 2Br2; (c) EtMgBr, CuI, THPOCH2CH2CꢁCH, 85%; (d) H2/Lindlar catalyst, 62%; (e) Ph3P,
reflux 36 h, 92%.
analogues of 2-AG as novel experimental tools or ther-
apeutic agents. Previously, we synthesized an ether
analogue of 2-AG, 2-(5(Z),8(Z),11(Z),14(Z)-eicosate-
traenyl) glycerol (2-AG ether) (2).6,8 This compound
was found to exhibit appreciable agonistic activities at
the cannabinoid receptors,6À8 though its activity was
weak compared with that of 2-AG. Independently and
concurrently, Mechoulam et al.9 also reported the
development of 2 (HU-310). However, no other stable
analogues have been reported so far. Here, we report
the synthesis and biological activities of novel stable
analogues of 2-AG, 2-AGA105 (3) and 2-AGA109 (4).
hydropyranyl ether of butynyl alcohol to yield the triyne
6 in 85% yield. The triyne 6 was then subjected to con-
trolled hydrogenation employing the Lindlar catalyst to
afford the crude triene 7. Traces of overreduced or
underreduced products were removed by argentation
chromatography utilizing 10% silver nitrate on silica
gel, to give pure 7 in 62% yield.10,11 The resultant pure
cis-triene 7 was deprotected and brominated in one step
with 1,2-bis(diphenylphosphino) ethane tetrabromide
and finally treated with triphenyl phosphine to afford 8
in 92% yield as a colorless oil.11
The key aldehyde synthon 13 was prepared from com-
mercially available 5-hexen-1-ol (9) as illustrated in
Scheme 2. Protection of the alcohol 9 with benzyl bromide
using NaH as a base followed by oxidation with
osmium tetroxide gave the diol 10 in an overall yield of
72%. Introduction of tosylate at the primary alcohol of
10 (74% yield), and subsequent protection of the sec-
ondary hydroxyl group with a tert-butyldimethylsilyl
(TBS) group afforded 11 in good yield. Then conversion
of the tosylate 11 to iodide with lithium iodide in dry
acetone, followed by alkylation with diethyl malonate
employing NaH gave the dicarboxylate 12 (88% yield in
each case). The benzyl group of 12 was hydrogenated
using Pd/C in ethanol, and then Swern oxidation affor-
ded the desired aldehyde 13 in 89% yield after purifica-
tion by column chromatography.
Figure 1 illustrates the chemical structures of 1, 2 and
the newly developed structurally related stable analogues
of 2-AG, 2-hydroxymethyl-tricosa-8(Z),11(Z),14(Z),
17(Z)-tetraene-1,4-diol (3) and 1-hydroxy-2-hydroxy-
methyl-tricosa-8(Z),11(Z),14(Z),17(Z)-tetraen-4-one (4).
Compound 3 is a hydroxyl group-containing and an ester
linkage-lacking analogue of 1, and 4 is a ketone analogue
of 1, which also lacks an ester linkage. These compounds
are resistant to hydrolyzing enzymes such as lipase.
The outline of our synthetic plan for 3 and 4 is as fol-
lows. First, we prepared phosphonium salt (8) and
aldehyde synthon (13). Then these components were
combined in a convergent and stereoselective manner to
obtain 14, a key intermediate. Step-wise chemical mod-
ification of 14 gave the desired compounds 3 and 4.
Synthesis of 3 and 4 from 8 and 13 was carried out as
follows (Scheme 3). Wittig coupling of the anion derived
from the phosphonate 8 with the aldehyde 13 gave a
69% yield of adduct 14. No trace of the trans isomer
was found by 400 MHz NMR spectroscopy. Reduction
of the diethyl ester of 14 with lithium aluminum hydride
afforded the diol 15 in 81% yield. The TBS protecting
group of 15 was removed by treatment with TBAF to
produce the triol 3 in 72% yield.12 Selective protection
of the primary alcohol of 3 with a TIPS group, followed
by oxidation of secondary alcohol 16 gave 17 in 74%
3(Z),6(Z),9(Z)-Tetradecatrienyl phosphonate (8), an
intermediate in the total synthesis of the above 2-AG
analogues, was prepared as illustrated in Scheme 1.
Commercially available 2-octyn-1-ol was brominated
with phosphorus tribromide. The resultant halide was
then coupled with propargyl alcohol to give 2,4-undeca-
diyl-1-ol (5) according to the method of Kerdesky et
al.10 The diyne 5 was subsequently halogenated with
1,2-bis(diphenylphosphophino) ethane tetrabromide
and the resultant bromide was coupled with the tetra-
Scheme 2. (a) NaH, BnBr; (b) OsO4, NMO, 72% (two steps); (c) TsCl, pyridine, 74%; (d) TBSCl, imidazole, 91%; (e) LiI, acetone, 88%; (f) NaH
(CO2Et)2, 88%; (g) H2, Pd/C, 97%; (h) Swern oxidation, 89%.

Y. Suhara et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1985–1988
1987
Scheme 3. (a) LiHMDS, HMPA, À98 to 0 ꢂC, 69%; (b) LiAlH4, 81%; (c) TBAF; (d) TIPSCl, imidazole, DMF, 60%; (e) PDC, 74%.
yield. Finally, deprotection of the TIPS group of 17
gave the ketone analogue 4 in 75% yield after the
removal of small amounts of by-products.13
In summary, we have developed two novel structural
analogues of 2-AG, 2-AGA105 (3) and 2-AGA109 (4).
These compound, as well as 2-AG ether (2), should be
useful lead compounds in the development of novel
stable analogues of 2-AG (1).
The biological activities of 3 and 4 as cannabinoid
receptor agonists were evaluated by estimating their
capacities to induce Ca2+ transients in NG108-15 cells,
which express the CB1 receptor, and in HL-60 cells,
which express the CB2 receptor, according to the
method described previously.4À8 As shown in Figures 2
and 3, 3 and 4 exhibited appreciable agonistic activities.
The agonistic activity of 4 toward NG 108-15 cells was
comparable to that of 2-AG ether (2). Also, the ago-
nistic activity of 3 toward HL-60 cells was nearly
comparable to that of 2-AG ether (2), yet it will be
necessary to determine in the future whether two
enantiomers of compound 3 possess similar biologi-
cal activities. The agonistic activities of these novel
Acknowledgements
The authors are grateful to Junko Shimode and Maroka
Kitsukawa for the spectroscopic measurements. This
study was supported in part by Special Coordination
Funds of the Ministry of Education, Culture, Sports,
Science and Technology, the Japanese Government.
References and Notes
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noda, A.; Itoh, K.; Yamashita, A.; Waku, K. Biochem. Bio-
phys. Res. Commun. 1995, 215, 89.
2-AG analogues (2,
approximately 100 times lower than that of 2-AG
3
and 4) were, however,
(1).6,7
Figure 2. Effects of 2-AG ether (2)(A), 2-AGA105 (3)(B) and 2-AGA109 (4)(C) on the intracellular free Ca2+ concentrations in NG108-15 cells
which express the CB1 receptor. The mean valuesÆSD were taken from six determinations.
Figure 3. Effects of 2-AG ether (2)(A), 2-AGA105 (3)(B) and 2-AGA109 (4)(C) on the intracellular free Ca2+ concentrations in HL-60 cells which
express the CB2 receptor. The mean valuesÆSD were taken from six determinations.

1988
Y. Suhara et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1985–1988
2. Mechoulam, R.; Ben-Shabat, S.; Hanus, L.; Ligumsky, M.;
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12. Data for 3: 1H NMR (400 MHz, CDCl3–D2O) d 5.41–5.33
(m, 8H), 3.77–3.63 (m, 5H), 2.85–2.78 (m, 6H), 2.07 (dt, 4H,
J=7.0, 7.0 Hz), 1.95 (dt, 1H, J=5.8, 5.8 Hz), 1.74–1.67 (m,
2H), 1.60–1.22 (m, 10H), 0.89 (t, 3H, J=6.7 Hz); 13C NMR
(100 MHz, CDCl3) d 130.5, 129.8, 128.6, 128.3, 128.2, 128.1,
127.9, 127.5, 70.2, 65.6, 65.3, 40.5, 37.8, 36.7, 31.5, 29.3, 27.2,
27.1, 25.7, 25.6, 25.5, 22.5, 14.0; MS 378 [M]+; HR-EIMS
calcd for [C24H42O3] 378.3134, found 378.3134.
13. Data for 4: 1H NMR (400 MHz, CDCl3–D2O) d 5.42–5.32
(m, 8H), 3.74 (dd, 2H, J=4.8, 10.8 Hz), 3.68 (dd, 2H, J=5.7,
10.8 Hz), 2.85–2.79 (m, 6H), 2.57 (d, 2H, J=6.7 Hz), 2.47 (t,
2H, J=7.3 Hz), 2.29 (ddd, 1H, J=4.8, 5.8, 6.7 Hz), 2.12–1.99
(m, 4H), 1.66 (dt, 2H, J=7.3, 7.3 Hz), 1.59–1.52 (m, 2H),
1.40–1.25 (m, 6H), 0.89 (t, 3H, J=6.8 Hz); 13C NMR
(100 MHz, CDCl3) d 210.9, 130.5, 129.1, 128.8, 128.6, 128.3,
128.2, 127.9, 127.6, 65.1, 42.7, 41.5, 38.3, 31.5, 29.3, 27.2, 26.5,
25.7, 23.6, 22.6, 14.1; MS 358 [MÀH2O]+; HR-EIMS calcd
for [C24H38O2] 358.2872, found 358.2867.
8. Suhara, Y.; Takayama, H.; Nakane, S.; Miyashita, T.;
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