Retinoids and related compounds. Part 19. Syntheses of 9E- and 9Z-locked retinoic acid analogues and their transcriptional activities as ligands for retinoic acid receptors and retinoid X receptors

Article abstract of DOI:10.1039/a606719k

Novel retinoic acid (RA) analogues, 9E-locked-RA 3 and 9Z-locked RA 4, have been synthesized in order to prohibit geometrical isomerization at the C(9)-C(10) double bond when these ligands interact with the retinoic acid receptor (RAR) and retinoid X rece

Full text of DOI:10.1039/a606719k

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Retinoids and related compounds. Part 19.1 Syntheses of 9E- and
9Z-locked retinoic acid analogues and their transcriptional activities
as ligands for retinoic acid receptors and retinoid X receptors
Yuko Katsuta, Yukiko Aoyama, Hisa Osone, Akimori Wada and
Masayoshi Ito*
Kobe Pharmaceutical University, 4-19-1 Motoyamakita-machi, Higashinada-ku, Kobe 658,
Japan
N ovel retinoic acid (R A) analogues, 9E-locked-R A 3 and 9Z -locked R A 4, have been synthesized in order
to prohibit geometrical isomerization at the C(9)᎐C(10) double bond when these ligands interact with the
retinoic acid receptor (R AR ) and retinoid X receptor (R XR ). Transcriptional activities of these analogues
for R AR and R XR are also described.
Introduction
Results and discussion
Retinoic acid (RA) is a derivative of vitamin A which plays a
key role in vertebrate vitamin A actions including proliferation,
differentiation, growth and development, apart from those
actions associated with vision.2 Recently, receptors for RA have
been identified as important members of the steroid/thyroid
nuclear receptor superfamily, which act as ligand-dependent
transcription factors. These receptors have been classified into
two types, the retinoic acid receptors (RARs)3 and the retinoid
X receptors (RXRs),4 and both have three kinds of subtypes (α,
β and γ) respectively. The difference in these two receptors is
due to the stereochemistry of the signalling molecules. Thus,
the ligand of RARs is all-E-RA 1 and that of RXRs is 9Z-RA
2,5 respectively. Interestingly, 9Z-RA 2 is also capable of bind-
ing directly to RARs, and their complexes have also been active
in the regulation of gene transcription. This implies that, unlike
all-E-RA 1, 9Z-RA 2 is a biologically active ligand for both
members of the RAR and RXR subfamilies. In addition, it is
suggested that the isomerization of retinoids is involved in con-
trolling signal-transduction pathways. A number of derivatives
of RA have hitherto been synthesized and their transcriptional
activities have been investigated. The structure of most
analogues prepared previously was aromatic and seemed to be
rather different from RA; therefore, a few of them behaved as
agonists or antagonists only for RARs.6 Further, RA is known
to isomerize around the double bonds rapidly in vivo. Hence we
prepared locked RA analogues 3 and 4, whose C(9)᎐C(10)
double bonds are prevented from isomerizing, to investigate
whether these enzyme analogues bind the receptors and exhibit
the transcriptional activities. Here we describe a full account of
the syntheses of the RA analogues 3 and 4 which are reported
briefly in the previous communication.7
Synthesis of 9E-locked RA 3 (Scheme 1)
Our first attempt to prepare 9E-locked trienone 5 according to
Albeck’s method 8 was unsuccessful due to the low yield in the
Wittig reaction between the triphenylphosphonium bromide of
β-cyclocitrol and 3-formylcyclohex-1-enone. Hence trienone 5
was synthesized by our original procedure as shown in Scheme
1. Dione 7, derived from the reaction of the lithium salt of
dithiane 69 with 5-chloropentan-2-one ethylene ketal (Aldrich)
and subsequent deprotection of the thioketal and ketal groups
(57% in 3 steps), was easily cyclized 10 in the presence of
MeONa to afford trienone 5 (100%). Condensation of the
trienone 5 with the C5-phosphonate did not proceed and the
desired retinoate analogue was not obtained. Therefore,
trienone 5 was converted to the retinoate analogue by stepwise
elongation of the side-chain. Condensation of the trienone 5
with propan-2-one N,N-dimethylhydrazone in the presence of
lithium diisopropylamide (LDA) afforded a mixture (1:1 ratio)
of tetraenones 8 and 9 in 72% yield after mild deprotection.11
The mixture was separated by low-pressure column chrom-
1
atography (LPCC) and these structures were confirmed by H
NMR spectroscopy; the 11Z-geometry was identified from the
downfield shift of the 10-H signal (δ 7.55) in compound 9 in
comparison with that (δ 5.98 or 6.02) in isomer 8 and the 11E-
geometry from the downfield shift of the 11a-H2 signal (δ 2.94)
in compound 8 compared with that (δ 2.38) in isomer 9, both
owing to anisotropic effects of the ketone groups. Peterson ole-
fination (92%) of 11E-isomer 8 with ethyl trimethylsilyl acetate
followed by preparative high-pressure liquid chromatography
(PHPLC) gave 13Z-isomer 10 and all-E-isomer 11 of the retin-
oate analogues in a 1:1 ratio. These structures were confirmed
1
on the basis of H NMR spectroscopy data; the 12-H signal (δ
6.93) in compound 10 was observed downfield in comparison
with that (δ 5.79) in isomer 11. Assignment of the 13C NMR
data of isomers 10 and 11 was performed by comparison with
9
7
11
13
14
CO2H
1
1
those of the known ethyl retinoates12 and by H–13C hetero-
12
8
10
nuclear shift-correlation (HETCOR) experiments, and com-
plete assignment of these H NMR data was obtained by 2D
5
1
1
2
homonuclear chemical-shift-correlation (COSY) experiments.
These retinoate analogues were carefully hydrolysed to RA
analogues 12 and 3, respectively. δ-Values of the olefinic
protons in the retinoates and the RAs (Table 1) show that no
geometrical isomerization occurred during the final hydrolysis.
CO2H
9
CO2H
3
Synthesis of 9Z-locked RA 4 (Scheme 2)
4
9Z-Locked trienone 16 was synthesized by a modified pro-
cedure used for the preparation of 11Z-locked retinal.13 Aldol
CO2H
J. Chem. Soc., Perkin Trans. 1, 1997
1405

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O
OH
O
CHO
S
i
S
i, ii, iii
O
O
13
14
6
7
ii, iii
iv
NOE
OH
9
7
iv, v
5
O
O
O
16
15
5
vi, vii, viii
v, vi, vii
9
9
10
11
10
11
O
11a
11
+
11a
11
O
10
12
10
12
12
12
O
O
(more polar)
9Z, 11Z-18
1 1Z-9
9Z, 11E-17
(more polar)
(less polar)
11E-8
(less polar)
ix, x
viii, ix
9
9
13
13
CO2R
13
9Z-20 R = Et
CO2R
9Z, 13Z-19 R = Et
CO2R
(more polar)
13Z-10
(less polar)
R = Et
CO2R
xi
(less polar)
xi
x
all-E-11
9Z, 13Z-21 R = H
9Z-4 R = H
R = Et
13Z-12
R = H
(more polar)
Scheme 2 Reagents and conditions: i, 3-(1-methylpropoxy)cyclohex-2-
enone, LDA, THF, Ϫ78 ЊC (61%); ii, MeLi, THF, Ϫ78 to 0 ЊC; iii, 15%
H2SO4, rt (60%); iv, Ac2O, Et3N, DMAP, CH2Cl2, rt (83%); v, DBU,
x
toluene, reflux (83%); vi, LDA, Me2NN᎐CMe2, THF, rt; vii, AcOH–
᎐
all-E-3
R = H
THF–water–AcONa (5:2:2:1), rt (41% [57%]); viii, PHPLC; ix, LDA,
Me3SiCH2CO2Et, THF, Ϫ78 ЊC (100%); x, PHPLC (in the dark);
xi, 25% NaOH, EtOH, 50 ЊC (92%, 100%)
Scheme 1 Reagents and conditions: i, BuLi, 5-chloropentan-2-one
ethylene ketal, THF, Ϫ78 ЊC; ii, HgO, HgCl2, 97% MeOH, rt; iii, p-
TsOH, acetone, rt (57%); iv, MeONa, THF, rt (100%); v, LDA, Me2N-
1
Table 2 UV–VIS and H NMR data for 9Z-locked retinoids
N᎐CMe2, THF, rt; vi, AcOH–THF–water–NaOAc (5:2:2:1), rt (72%
᎐
[94%]); vii, LPCC; viii, LDA, Me3SiCH2CO2Et, THF, Ϫ78 ЊC (92%); ix,
PHPLC (in the dark); x, 25% NaOH, EtOH, 50 ЊC (92%, 81%)
9Z,13Z-19 9Z-20 9Z,13Z-21a 9Z-4a
1
Table 1 UV–VIS and H NMR data for 9E-locked retinoids
λmax/nm
348
342
335
334, 250
UV–VIS
13Z-10
All-E-11 13Z-12a
All-E-3a
1H NMR
(200 MHz)
(δ, CDCl3)
1,1-Me2
5-Me
9-Me
13-Me
7-H
10-H
12-H
14-H
0.95
1.46
1.96
2.07
6.02
6.11
6.89
5.61
0.96
1.46
1.97
2.28
6.04
5.97
5.76
5.68
0.95
1.46
1.97
2.11
6.05
6.11
6.87
5.63
0.96
1.47
1.98
2.30
6.06
5.98
5.79
5.72
UV–VIS
1H NMR
λmax/nm
358
355
345, 246
353
1,1-Me2
1.00
1.69
2.10
6.21
6.07
6.17
6.93
5.61
1.01
1.70
2.29
6.24
6.07
6.04
5.79
5.72
1.01
1.70
2.13
6.23
6.10
6.18
6.90
5.64
1.01
1.70
2.31
6.26
6.09
6.05
5.82
5.75
(200 MHz) 5-Me
(δ, CDCl3)
13-Me
7-H
8-H
10-H
12-H
14-H
a
No signal for the carboxy proton was observed.
resulting diol was treated with acid to give compound 15 which,
by acetylation and subsequent elimination (1,8-diazabicyclo-
[5.4.0]undec-7-ene, DBU) was converted to 7E-trienone 16
exclusively. The stereostructure of trienone 16 was confirmed
by a nuclear Overhauser and exchange spectroscopy (NOESY)
a
No signal for the carboxy proton was observed.
condensation between β-cyclocitral 1314 and easily prepared
3-(1-methylpropoxy)cyclohex-2-enone15 afforded hydroxy
ketone 14. After addition of methyllithium to ketone 14, the
1406
J. Chem. Soc., Perkin Trans. 1, 1997

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Merck Art. 7734. LPCC was conducted on a Yamazen Low
δ 1.02 (s)
δ 0.97 (br s)
Pressure Liquid Chromatography System using a Lobar
Merck LiChroprep Si-60 column. Preparative HPLC was con-
ducted on a Shimadzu LC-10AS instrument with a Shimadzu
UV–VIS detector, SPD-10A, using a Merck LiChrosorb Si-60
(7 µm), 1.0 × 25 cm column. Unless otherwise stated, solvent
extracts were dried over anhydrous sodium sulfate and all
operations were carried out under nitrogen or argon. The
extract or the filtrate was concentrated under reduced pressure
at <30 ЊC using a rotary evaporator. NMR assignments follow
the retinoic acid numbering system.
9
O
O
δ 1.49
δ 1.68
9Z-β-Ionylidene
acetaldehyde
16
λ
max(EtOH)/nm 321
λ
max(EtOH)/nm 296
δ 1.02 (br s)
Synthesis of 9E-locked RA 3. (E)-8-(2,6,6-Trimethylcyclohex-
1-enyl)oct-7-ene-2,6-dione 7
To a solution of the dithiane 6 (2.0 g, 7.46 mmol) in dry tetra-
hydrofuran (THF) (20 cm3) was added a solution of BuLi
(1.70 mol dmϪ3; 4.83 cm3, 8.21 mmol) in hexane at Ϫ78 ЊC.
The mixture was stirred at room temperature (rt) for 1 h and
5-chloropentan-2-one ethylene ketal (1.35 cm3, 8.96 mmol)
was then added dropwise at Ϫ78 ЊC. After being stirred at rt for
3 h, the reaction mixture was quenched by addition of water.
After evaporation off of the THF, the residue was extracted
with ether. The extract was washed with brine, dried and evap-
orated to give an oil, which was dissolved with 97% methanol
(100 cm3). HgCl2 (8.12 g, 29.9 mmol) and HgO (4.85 g, 22.4
mmol) were added to this solution and the reaction mixture was
stirred at rt for 15 min. Evaporation off of the methanol gave a
residue, which was dissolved in ether and the solution was fil-
tered through Celite. The filtrate was washed with brine, dried
and evaporated to afford a residue, which was dissolved with
acetone (50 cm3). To the solution was added toluene-p-sulfonic
acid (TsOHؒ2H2O) (142 mg, 0.75 mmol) at 0 ЊC and the reac-
tion mixture was stirred at rt for 1 h before being neutralized
by the addition of saturated aq. NaHCO3 and the acetone was
evaporated off to give a residue, which was extracted with
ether. The extract was washed with brine, dried and evaporated
to give an oil, which was purified by CC (AcOEt–hexane, 1:9).
This afforded the title compound 7 (1.11 g, 57%) as an oil
(Found: C, 77.66; H, 9.71%; Mϩ, 262.1915. C17H26O2 requires
C, 77.82; H, 9.99%; M, 262.1934); νmax(CHCl3)/cmϪ1 1712
7
O
δ 1.52
7Z-β-Ionylidene
acetaldehyde
λ
max(EtOH)/nm 272 (ε 14800)
313 (ε 9180)
Fig. 1
experiment (observed crosspeak between 7-H and 9-Me). The
broad singlet at δ 0.97 (1,1-Me2) indicates that rotational bar-
riers exist in the two rings in trienone 16 and hence the twisted
6s-cis conformation is consistent with the unusual high-field
shift of the 5-Me group. In such a conformation, the 5-Me
group is in the shielding cone of the C(7)᎐C(8) double bond and
similar phenomena were found in 7Z-retinoids16 (Fig. 1). The
significantly shorter absorption maximum (296 nm) of trienone
16 than that of 9Z-β-ionylideneacetaldehyde (321 nm) also
revealed this highly twisted 6s-cis conformation (Fig. 1). Final
transformation of trienone 16 into 9Z-locked RAs was
achieved by means of the route used for 9E-locked RAs 12 and
1
3. Their H NMR data in Table 2 show that no geometrical
isomerization occurred during the final hydrolysis.
(nonconj. C᎐O), 1657 (conj. C᎐O) and 1603 (C᎐C);
᎐
᎐
᎐
λmax(EtOH)/nm 295 and 219sh; δH (200 MHz; CDCl3) 1.04 (6
H, s, gem-Me), 1.74 (3 H, s, 5-Me), 1.89 (2 H, quin, J 7, 9b-
H2), 2.12 (3 H, s, COMe), 2.50 and 2.59 (each 2 H, each t, J 7,
CH2CH2CH2COMe), 6.09 (1 H, d, J 16.5, 8-H) and 7.30 (1 H,
d, J 16.5, 7-H).
Transcriptional activities of 9E- and 9Z-locked RAs for RAR
and RXR
Transcriptional activities of synthesized RA analogues 3, 12, 4
and 21 were compared with those of all-E-RA 1 and 9Z-RA 2
by chloramphenicol acetyltransferase (CAT) assay. All anal-
ogues bound to both mRARα and mRXRα, and had activities
which were significant but weaker than those of all-E-RA 1 and
9Z-RA 2 (3; 1/10 of 1 or 2: 12, 4 and 21; 1/100 of 1 or 2). The
results suggest that the synthesized analogues exhibited agon-
istic actions towards both mRARα and mRXRα receptors, but
it was supposed that the artificial 6-membered ring hindered the
interaction between ligands and receptors.
(E)-3-[2-(2,6,6-Trimethylcyclohex-1-enyl)ethenyl]cyclohex-2-
enone 5
To a solution of dione 7 (1.10 g, 4.20 mmol) in dry CH2Cl2 (10
cm3) was added dropwise a solution of NaOMe (261 mg, 4.83
mmol) in methanol (2 cm3) at 0 ЊC. After being stirred at rt for
3 h, the reaction mixture was poured into water and extracted
with CH2Cl2. The extract was washed with brine, dried and
evaporated. The residue was purified by CC (AcOEt–hexane,
1:4) to give the title compound 5 (1.02 g, 100%) as an oil,
νmax(CHCl3)/cmϪ1 1651 (C᎐O) and 1610 (C᎐C); λmax(EtOH)/nm
᎐
᎐
Experimental
322 and 271; δH (200 MHz; CDCl3) 1.02 (6 H, s, gem-Me), 1.70
(3 H, s, 5-Me), 2.41 and 2.51 (each 2 H, each t-like, J 6.5 and 6,
9a-H2 and 11a-H2), 5.90 (1 H, s, 10-H), 6.16 (1 H, d, J 16, 8-H)
and 6.64 (1 H, d, J 16, 7-H) (Found: Mϩ, 244.1833. C17H24O
requires M, 244.1826).
Mps are uncorrected. Ether refers to diethyl ether and hexane
to n-hexane. BuLi was used as a solution in hexane. UV–VIS
spectra were recorded on a JASCO Ubest-55 instrument and IR
or FT-IR spectra on a Shimadzu IR-27G or Shimadzu FT-IR-
1
4200 spectrometer. H NMR spectra at 60, 200, 300 or 500
MHz were measured on a JOEL JNM-PMX 60, a Varian
Gemini-200, a Varian Gemini-300 or a Varian VXR-500 super-
conducting FT-NMR spectrometer using chloroform (δ 7.25)
as internal reference and 13C NMR spectra at 75 MHz or 125
MHz were measured on a Varian Gemini-300 or a Varian
VXR-500 spectrometer. Mass spectra were determined on
a Hitachi M-4100 double-focusing GC mass spectrometer.
Column chromatography (CC) was performed on silica gel
Propan-2-one N,N-dimethylhydrazone17
A mixture of acetone (19.7 g, 0.34 mol), N,N-dimethyl-
hydrazine (10 g, 0.17 mol) and benzene (100 cm3) was refluxed
for 1 h using a Dean–Stark separator and was then distilled to
give a solution of the title compound in benzene (50% w/w,
1
the concentration was determined by H NMR); δH (60 MHz;
CDCl3) 1.92 (6 H, d, J 2, CMe2), 2.41 (6 H, s, NMe2) and 7.25 (6
H, s, benzene).
J. Chem. Soc., Perkin Trans. 1, 1997
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(E,E/Z)-3-{3-[2-(2,6,6-Trimethylcyclohex-2-enyl)ethenyl]cyclo-
hex-2-enylidene}propan-2-one 8 and 9
129.73 (C-12), 129.86 (C-5), 131.25 (C-10), 135.33 (C-8), 137.68
(C-6), 141.14 and 142.04 (C-9 and -11), 153.35 (C-13) and
167.21 (C-15) (Found: Mϩ, 354.2559. C24H34O2 requires M,
354.2557).
To a solution of LDA (8.20 mmol, prepared from 1.15 cm3 of
diisopropylamine and 4.82 cm3 of 1.70 mol dmϪ3 BuLi) in dry
THF (8 cm3) was added a solution of propan-2-one N,N-
dimethylhydrazone (50% w/w; 1.46 g, 8.20 mmol) in benzene at
0 ЊC. After the reaction mixture had been stirred at rt for 30
min, a solution of trienone 5 (400 mg, 1.64 mmol) in dry THF
(4 cm3) was added at 0 ЊC and the mixture was stirred at rt for
2 h. The reaction was quenched by the addition of water and
the THF was evaporated off to give a residue, which was
extracted with AcOEt. The extract was washed with brine, dried
and evaporated off to afford a crude oil, which was dissolved
with a mixture of AcOH–THF–water–NaOAc (5:2:2:1) and
the reaction mixture was stirred at rt for 5 h and extracted with
AcOEt. The extract was washed with brine, dried and evapor-
ated to give a residue, which was purified by CC (AcOEt–
hexane, 1:9). This afforded a mixture of geometrical isomers
8 and 9 (335 mg, 72%) as a pale yellow oil, together with
recovered starting material 5 (95 mg, 24%). The isomers were
separated by LPCC (AcOEt–hexane, 1:19) to give the more
polar compound 8 and the less polar compound 9 each in a
pure state, in the ratio ~1:1.
13Z-Isomer 10: νmax(KBr)/cmϪ1 1707 (CO2Et) and 1597 and
1570 (C᎐C); λmax(EtOH)/nm 358; δH (200 MHz; CDCl3) 1.00 (6
᎐
H, s, gem-Me), 1.25 (3 H, t, J 7, CO2CH2CH3), 1.69 (3 H, s, 5-
Me), 2.10 (3 H, s, 13-Me), 2.31 and 2.52 (each 2 H, each t-like, J
5.5, 9a- and 11a-H2), 4.13 (2 H, q, J 7, CO2CH2CH3), 5.61 (1 H,
s, 14-H), 6.07 (1 H, d, J 16, 8-H), 6.17 (1 H, s, 10-H), 6.21 (1 H,
d, J 16, 7-H) and 6.93 (1 H, s, 12-H); δH (500 MHz; CDCl3) 1.01
(6 H, s, gem-Me), 1.25 (3 H, t, J 7, CO2CH2CH3), 1.69 (3 H, d,
J 1, 5-Me), 1.77 (2 H, quintet, J 6, 9b-H2), 2.10 (3 H, d, J 1,
13-Me), 2.31 (2 H, t, J 6, 9a-H2), 2.52 (2 H, m, 11a-H2), 4.13
(2 H, q, J 7, CO2CH2CH3), 5.61 (1 H, s, 14-H), 6.08 (1 H, d,
J 16, 8-H), 6.17 (1 H, s, 10-H), 6.19 (1 H, d, J 16, 7-H) and 6.93
(1 H, s, 12-H) (Found: Mϩ, 354.2556).
(E,E,E)-3-Methyl-4-{3-[2-(2,6,6-trimethylcyclohex-1-enyl)-
ethenyl]cyclohex-2-enylidene}but-2-enoic acid 3
A mixture of all-E-retinoate analogue 11 (6.6 mg, 0.019 mmol),
ethanol (0.5 cm3) and aq. NaOH (25% w/w; 0.15 cm3) was
stirred at rt for 3 h and then at 50 ЊC for 30 min. The reaction
mixture was acidified by the addition of dil. HCl and extracted
with AcOEt. The extract was washed with brine, dried and
evaporated to give a residue, which was purified by CC
(AcOEt–hexane, 1:4) to afford all-E-RA analogue 3 (274.9
mg, 81%) as a pale yellow solid (mp 121–123 ЊC), νmax(KBr)/
cmϪ1 3200–2500, 1678 (CO H) and 1564 (C᎐C); λmax(EtOH)/
11E-Isomer 8: νmax(CHCl3)/cmϪ1 1658 (C᎐O) and 1554
᎐
(C᎐C); λmax(EtOH)/nm 346; δH (200 MHz; CDCl3) 1.01 (6 H, s,
᎐
gem-Me), 1.70 (3 H, s, 5-Me), 2.19 (3 H, s, 13-Me), 2.36 (2 H, t-
like, J 6, 9a-H2), 2.94 (2 H, m, 11a-H2), 5.98 and 6.02 (each 1 H,
each s, 10- and 12-H), 6.12 (1 H, d, J 16, 8-H) and 6.39 (1 H,
d, J 16, 7-H) (Found: Mϩ, 284.2133. C20H28O requires M,
284.2138).
᎐
2
nm 353; δH (200 MHz; CDCl3) 1.01 (6 H, s, gem-Me), 1.70 (3 H,
s, 5-Me), 2.31 (3 H, s, 13-Me), 2.34 and 2.61 (each 2 H, each
t-like, J 5, 9a- and 11a-H2), 5.75 (1 H, s, 14-H), 5.82 (1 H, s, 12-
H), 6.05 (1 H, s, 10-H), 6.09 (1 H, d, J 16, 8-H) and 6.26
(1 H, d, J 16, 7-H) (Found: Mϩ, 326.2237. C22H30O2 requires M,
326.2244).
11Z-Isomer 9: νmax(CHCl3)/cmϪ1 1660 (C᎐O) and 1560
᎐
(C᎐C); λmax(EtOH)/nm 349; δH (200 MHz; CDCl3) 1.01 (6 H, s,
᎐
gem-Me), 1.71 (3 H, s, 5-Me), 2.18 (3 H, s, 13-Me), 2.38 (4 H, m,
9a- and 11a-H2), 5.86 (1 H, s, 12-H), 6.22 (1 H, d, J 16, 8-H),
6.40 (1 H, d, J 16, 7-H) and 7.55 (1 H, s, 10-H) (Found: Mϩ,
284.2142).
(E,E,Z)-3-Methyl-4-{3-[2-(2,6,6-trimethylcyclohex-1-enyl)-
ethenyl]cyclohex-2-enylidene}but-2-enoic acid 12
Ethyl (E,E,Z/E)-3-methyl-4-{3-[2-(2,6,6-trimethylcyclohex-1-
enyl)ethenyl]cyclohex-2-enylidene}but-2-enoate 10 and 11
To a solution of LDA (0.35 mmol, prepared from 0.049 cm3 of
diisopropylamine and 0.219 cm3 of 1.60 mol dmϪ3 BuLi) in dry
THF (1 cm3) was added a solution of ethyl trimethylsilylacetate
(0.064 cm3, 0.35 mmol) in dry THF (1 cm3) at Ϫ78 ЊC. After the
reaction mixture had been stirred at Ϫ78 ЊC for 15 min, a solu-
tion of tetraenone 8 (20 mg, 0.070 mmol) in dry THF (2 cm3)
was added at Ϫ78 ЊC and the mixture was stirred for 30 min at
rt. The whole was concentrated to give a residue, which was
purified by CC (AcOEt–hexane, 1:19) to afford a mixture of
geometrical isomers 10 and 11 (23 mg, 92%). The isomers were
separated by PHPLC [LiChrosorb Si-60 (7 µm), 1 × 25 cm,
Et2O–hexane, 3:97] to give the less polar compound 10 and the
more polar compound 11, each in a pure state and each as a
pale yellow oil, in the ratio ~1:1.
In the same manner as described for the preparation of all-E-
RA analogue 3 from all-E-retinoate analogue 11, hydrolysis of
13Z-retinoate analogue 10 (7.7 mg, 0.022 mmol) by NaOH gave
13Z-RA analogue 12 (6.5 mg, 92%) as a pale yellow amorphous
solid, νmax(KBr)/cmϪ1 3200–2500, 1674 (CO2H) and 1593 and
1564 (C᎐C); λmax(EtOH)/nm 345 and 246; δH (200 MHz; CDCl3)
᎐
1.01 (6 H, s, gem-Me), 1.70 (3 H, s, 5-Me), 2.13 (3 H, s, 13-Me),
2.32 and 2.54 (each 2 H, each t-like, J 6 and 5, 9a-H2 and 11a-
H2), 5.64 (1 H, s, 14-H), 6.10 (1 H, d, J 16.5, 8-H), 6.18 (1 H, s,
10-H), 6.23 (1 H, d, J 16.5, 7-H) and 6.90 (1 H, s, 12-H) (Found:
Mϩ, 326.2245).
Synthesis of 9Z-locked RA 4. 3-(1-Methylpropoxy)-6-[hydroxy-
(2,6,6-trimethylcyclohex-1-enyl)methyl]cyclohex-2-enone 14
To a solution of LDA (32.9 mmol, prepared from 4.60 cm3 of
diisopropylamine and 20.2 cm3 of 1.63 mol dmϪ3 BuLi) in dry
THF (14 cm3) was added a solution of 3-(1-methylpropoxy)-
cyclohex-2-enone (6.15 g, 36.2 mmol) in dry THF (14 cm3) at
Ϫ78 ЊC. After the reaction mixture had been stirred at Ϫ78 ЊC
for 30 min, a solution of β-cyclocitral 13 (5.0 g, 32.9 mmol) in
dry THF (10 cm3) was added at Ϫ78 ЊC and the mixture was
stirred at Ϫ78 ЊC for 1 h. The reaction mixture was quenched by
addition of water. After evaporation off of the THF, the resi-
due was extracted with ether. The extract was washed with
brine, dried and evaporated to give a residue, which was purified
by CC (AcOEt–hexane, 1:9) to afford the title compound 14
(6.42 g, 61%) as a pale yellow oil (Found: C, 74.80; H, 9.82%;
M, 320.2369. C20H32O3 requires C, 74.96; H, 10.17%; M,
All-E-isomer 11: νmax(KBr)/cmϪ1 1709 (CO2Et) and 1571
(C᎐C); λmax(EtOH)/nm 355; δH (200 MHz; CDCl3) 1.01 (6 H, s,
᎐
gem-Me), 1.28 (3 H, t, J 7, CO2CH2CH3), 1.70 (3 H, s, 5-Me),
2.29 (3 H, s, 13-Me), 2.33 and 2.60 (each 2 H, t-like and m, J 6,
9a- and 11a-H2), 4.15 (2 H, q, J 7, CO2CH2CH3), 5.72 (1 H, s,
14-H), 5.79 (1 H, s, 12-H), 6.04 (1 H, s, 10-H), 6.07 (1 H, d, J
16.5, 8-H) and 6.24 (1 H, d, J 16.5, 7-H); δH (500 MHz; CDCl3)
1.01 (6 H, s, gem-Me), 1.28 (3 H, t, J 7, CO2CH2CH3), 1.45 (2
H, t, J 6, 2-H2), 1.59 (2 H, quintet, J 6.5, 3-H2), 1.70 (3 H, s, 5-
Me), 1.77 (2 H, quintet, J 6, 9b-H2), 2.00 (2 H, t, J 6, 4-H2), 2.29
(3 H, s, 13-Me), 2.33 (2 H, t, J 6, 9a-H2), 2.58 (2 H, m, 11a-H2),
4.15 (2 H, q, J 7, CO2CH2CH3), 5.72 (1 H, s, 14-H), 5.78 (1 H, s,
12-H), 6.04 (1 H, s, 10-H), 6.08 (1 H, d, J 16, 8-H) and 6.23 (1
H, d, J 16, 7-H); δC(125 MHz; CDCl3) 14.39 (CO2CH2CH3),
19.22 (C-3), 19.82 (C-20), 21.73 (C-18), 22.41 (C-21), 24.50 (C-
19), 27.59 (C-22), 28.94 (1,1-gem-Me), 33.09 (C-4), 34.24 (C-1),
39.61 (C-2), 59.55 (CH2CH2CH3), 117.65 (C-14), 127.48 (C-7),
320.2350); νmax(CHCl3)/cmϪ1 3420 (OH), 1617sh (C᎐O) and
᎐
1598 (C᎐C); νmax(KBr)/cmϪ1 3440 (OH), 1630 (C᎐O) and 1601
᎐
᎐
(C᎐C); λmax(EtOH)/nm 253; δH (500 MHz; CDCl3) 0.91 [6 H, m,
᎐
1-Me and OCH(CH3)CH2CH3], 1.13 (3 H, s, 1-Me), 1.23 and
1408
J. Chem. Soc., Perkin Trans. 1, 1997

View Article Online
1.26 [total 3 H, each d, J 6, OCH(CH3)CH2CH3], 1.83 (3 H, s,
5-Me), 2.39 (1 H, m, 8a-H), 2.84 (1 H, octet, J 5, 8-H), 4.19 [1
H, m, OCH(CH3)CH2CH3], 4.49 (1 H, br s, 7-H), 4.69 (1 H, s,
OH) and 5.34 (1 H, s, 10-H) (Found: Mϩ, 320.2369. C20H32O3
requires M, 320.2350).
11E-Isomer 17: νmax(CHCl3)/cmϪ1 1659 (C᎐O) and 1562
᎐
(C᎐C); λmax(EtOH)/nm 339 and 239; δH (200 MHz; CDCl3) 0.95
᎐
(6 H, br s, gem-Me), 1.46 (3 H, s, 5-Me), 2.01 (3 H, s, 9-Me),
2.18 (3 H, s, 13-Me), 5.95 and 5.97 (each 1 H, each s, 10- and
12-H) and 6.14 (1 H, s, 7-H) (Found: Mϩ, 284.2137. C20H28O
requires M, 284.2138).
4-[Hydroxy-(2,6,6-trimethylcyclohex-1-enyl)methyl]-3-methyl-
cyclohex-2-enone 15
11Z-Isomer 18: νmax(CHCl3)/cmϪ1 1662 (C᎐O) and 1579
᎐
(C᎐C); λmax(EtOH)/nm 341; δH (200 MHz; CDCl3) 0.95 (6 H, br
᎐
To a solution of keto alcohol 14 (2.83 g, 8.85 mmol) in dry THF
s, gem-Me), 1.45 (3 H, s, 5-Me), 2.04 (3 H, s, 9-Me), 2.17 (3 H, s,
13-Me), 5.82 (1 H, s, 12-H), 6.18 (1 H, s, 7-H) and 7.42 (1 H, s,
10-H) (Found: Mϩ, 284.2130).
(10 cm3) was added a solution of methyllithium (1.5 mol dmϪ3
;
17.7 cm3, 26.6 mmol) in THF at Ϫ78 ЊC. After the reaction
mixture had been stirred at Ϫ78 ЊC for 70 min, Ϫ50 ЊC for 20
min and at 0 ЊC for 20 min, it was treated with sulfuric acid (15%
w/v; 10 cm3) and the reaction mixture was stirred at 0 ЊC for a
further 20 min before being extracted with AcOEt. The extract
was washed with brine, dried and evaporated to give a residue,
which was purified by CC (AcOEt–hexane, 1:4) to provide the
title compound 15 (1.69 g, 60%) as an oil (Found: C, 77.52; H,
9.82%; Mϩ, 262.1931. C17H26O2 requires C, 77.82; H, 9.99%; M,
262.1931); νmax(CHCl3)/cmϪ1 3600 and 3440 (OH) and 1658
Ethyl (E,E,Z/E)-3-methyl-4-{3-methyl-4-[(2,6,6-trimethyl-
cyclohex-1-enyl)methylene]cyclohex-2-enylidene}but-2-enoate
19 and 20
In the same manner as described for the preparation of retin-
oate analogues 10 and 11 from tetraenone 8, tetraenone 17
(71 mg, 0.25 mmol) was converted to a mixture of geometrical
isomers 19 and 20 (88 mg, 100%), which was separated by
PHPLC [LiChrosorb Si-60 (7 µm), 1 × 25 cm, Et2O–hexane,
3:97] to give the less polar compound 19 and the more polar
compound 20, each in a pure state and each as a pale yellow oil,
in the ratio ~1:1.
(C᎐O); λmax(EtOH)/nm 240; δH (200 MHz; CDCl3) 1.01 and 1.11
᎐
(each 3 H, each s, gem-Me), 1.85 (3 H, s, 5-Me), 2.19 (3 H, s, 9-
Me), 2.51 (1 H, dq, J 17.5 and 5, 8Ј-H), 2.95 (1 H, dt, J 9 and 5,
8-H), 4.43 (1 H, br d, J 9, 7-H) and 5.91 (1 H, br s, 10-H).
9Z,13Z-Isomer 19: νmax(KBr)/cmϪ1 1709 (CO2Et) and 1597
and 1579 (C᎐C); λmax(EtOH)/nm 348; δH (300 MHz; CDCl3)
᎐
(E)-3-Methyl-4-[(2,6,6-trimethylcyclohex-1-enyl)methylene]-
cyclohex-2-enone 16
0.95 (6 H, br s, gem-Me), 1.25 (3 H, t, J 7, CO2CH2CH3), 1.46 (3
H, s, 5-Me), 1.96 (3 H, s, 9-Me), 2.08 (3 H, s, 13-Me), 4.13 (2 H,
q, J 7, CO2CH2CH3), 5.61 (1 H, s, 14-H), 6.02 (1 H, s, 7-H), 6.11
(1 H, s, 10-H) and 6.89 (1 H, s, 12-H); δH (200 MHz; CDCl3)
0.95 (6 H, br s, gem-Me), 1.25 (3 H, t, J 7, CO2CH2CH3), 1.46 (3
H, s, 5-Me), 1.96 (3 H, s, 9-Me), 2.07 (3 H, s, 13-Me), 4.12 (2 H,
q, J 7, CO2CH2CH3), 5.61 (1 H, s, 14-H), 6.02 (1 H, s, 7-H), 6.11
(1 H, s, 10-H) and 6.89 (1 H, s, 12-H) (Found: Mϩ, 354.2567.
C24H34O2 requires M, 354.2557).
Triethylamine (11.1 cm3, 79.6 mmol), 4-(dimethylamino)-
pyridine (DMAP) (3.88 g, 31.8 mmol) and acetic anhydride
(3.0 cm3, 31.8 mmol) were added to a solution of keto alcohol
15 (4.17 g, 15.9 mmol) in dry CH2Cl2 (30 cm3) at 0 ЊC and the
mixture was stirred at rt for 4 h. After being poured into
water, the reaction mixture was extracted with CH2Cl2. The
extract was washed with brine, dried and evaporated to give a
residue, which was purified by CC (AcOEt–hexane, 1:6) to
provide the acetate of compound 15 (4.02 g, 83%) as an oil,
9Z,13E-Isomer 20: νmax(KBr)/cmϪ1 1711 (CO2Et) and 1578
(C᎐C); λmax(EtOH)/nm 342; δH (300 MHz; CDCl3) 0.94 (6 H, br
᎐
νmax(CHCl3)/cmϪ1 1728 (OAc) and 1660 (C᎐O); λmax(EtOH)/
s, gem-Me), 1.27 (3 H, t, J 7, CO2CH2CH3), 1.47 (3 H, s, 5-Me),
1.97 (3 H, s, 9-Me), 2.28 (3 H, s, 13-Me), 4.15 (2 H, q, J 7,
CO2CH2CH3), 5.68 (1 H, s, 14-H), 5.76 (1 H, s, 12-H), 5.97 (1 H,
s, 10-H) and 6.04 (1 H, s, 7-H); δH (200 MHz; CDCl3) 0.96 (6 H,
br s, gem-Me), 1.27 (3 H, t, J 7, CO2CH2CH3), 1.46 (3 H, s, 5-
Me), 1.97 (3 H, s, 9-Me), 2.28 (3 H, s, 13-Me), 4.15 (2 H, q, J 7,
CO2CH2CH3), 5.68 (1 H, s, 14-H), 5.76 (1 H, s, 12-H), 5.97 (1 H,
s, 10-H) and 6.04 (1 H, s, 7-H) (Found: Mϩ, 354.2549).
᎐
nm 235; δH (200 MHz; CDCl3) 1.10 and 1.14 (each 3 H, each s,
gem-Me), 1.80 (3 H, s, 5-Me), 2.01 (3 H, s, OAc), 2.05 (3 H, s,
9-Me), 2.27 and 2.54 (each 1 H, each m, 8Ј-H2), 2.99 (1 H,
dt, J 9.5, 5, 8-H), 5.89 (1 H, s, 10-H) and 5.93 (1 H, d, J 9.5,
7-H) (Found: Mϩ Ϫ OAc, 245.1921. C17H25O requires m/z,
245.1905).
A mixture of a solution of the acetate (3.98 g, 13.1 mmol) in
dry toluene (5 cm3) and a solution of DBU (3.98 g, 26.2 mmol)
in dry toluene (5 cm3) was refluxed for 1 h. After being cooled to
rt, the mixture was evaporated to remove toluene to give a resi-
due, which was purified by CC (AcOEt–hexane, 1:6) to afford
the title compound 16 (2.65 g, 83%) as an oil (Found: C, 83.27;
H, 10.17%; Mϩ, 244.1837. C17H24O requires C, 83.55; H, 9.90%;
(E,E,E)-3-Methyl-4-{3-methyl-4-[(2,6,6-trimethylcyclohex-1-
enyl)methylene]cyclohex-2-enylidene}but-2-enoic acid 4
In the same manner as described for the preparation of all-E-
RA analogue 3 from all-E-retinoate analogue 11, hydrolysis of
9Z,13E-retinoate analogue 20 (3.5 mg, 0.010 mmol) by NaOH
yielded 9Z,13E-RA analogue 4 (3.3 mg, 100%) as a pale yellow
solid, mp 140–141 ЊC; νmax(KBr)/cmϪ1 3250–2500, 1680 (CO2H)
M, 244.1826); νmax(CHCl3)/cmϪ1 1654 (C᎐O); λmax(EtOH)/nm
᎐
296; δH (300 MHz; CDCl3) 0.97 (6 H, br s, gem-Me), 1.49 (3 H, s,
5-Me), 2.10 (3 H, s, 9-Me), 2.42 and 2.52 (each 2 H, each m, 8a-
and 8b-H2), 5.87 (1 H, s, 10-H) and 6.40 (1 H, s, 7-H); NOESY,
see Scheme 2; δC(75 MHz; CDCl3) 19.8 (C-3), 21.1 (9-Me), 21.9
(5-Me), 27.3 (C-8a), 29.0 (gem-Me), 32.4 (C-4), 35.3 (C-1), 37.6
(C-8b), 39.5 (C-2), 127.1 (C-8), 129.9 (C-10), 131.3 (C-7), 135.7
and 136.7 (C-5 and -6), 156.3 (C-9) and 200.4 (C-11).
and 1572 (C᎐C); λmax(EtOH)/nm 334 and 250; δH (200 MHz;
᎐
CDCl3) 0.96 (6 H, br s, gem-Me), 1.47 (3 H, s, 5-Me), 1.98 (3 H,
s, 9-Me), 2.30 (3 H, s, 13-Me), 5.72 (1 H, s, 14-H), 5.79 (1 H, s,
12-H), 5.98 (1 H, s, 10-H) and 6.06 (1 H, s, 7-H) (Found: Mϩ,
326.2240. C22H30O2 requires M, 326.2244).
(E,E,Z)-3-Methyl-4-{3-methyl-4-[(2,6,6-trimethylcyclohex-1-
enyl)methylene]cyclohex-2-enylidene}but-2-enoic acid 21
(E,E/Z)-3-{3-Methyl-4-[(2,6,6-trimethylcyclohex-1-enyl)methyl-
ene]cyclohex-2-enylidene}propan-2-one 17 and 18
In the same manner as described for the preparation of all-E-
RA analogue 3 from all-E-retinoate analogue 11, hydrolysis of
9Z,13Z-retinoate analogue 19 (5 mg, 0.014 mmol) by NaOH
provided 9Z,13Z-RA analogue 21 (4.2 mg, 92%) as a pale yel-
low solid, mp 128–130 ЊC; νmax(KBr)/cmϪ1 3300–2400, 1680
In the same manner as described for the synthesis of tetra-
enones 8 and 9 from trienone 5, trienone 16 (410 mg, 1.68
mmol) was converted into a mixture of the title compounds 17
and 18 (197 mg, 41%) as a pale yellow oil and some starting
material 16 (113 mg, 28%) was recovered. The mixture of geo-
metrical isomers was separated by PHPLC [LiChrosorb Si-60
(7 µm), 1 × 25 cm, AcOEt–hexane, 3:197] to give the less polar
compound 17 and the more polar compound 18 each in a pure
state, in the ratio ~1:1.
(CO H) and 1580 (C᎐C); λmax(EtOH)/nm 335; δH (200 MHz;
᎐
2
CDCl3) 0.95 (6 H, br s, gem-Me), 1.46 (3 H, s, 5-Me), 1.97 (3 H,
s, 9-Me), 2.11 (3 H, s, 13-Me), 5.63 (1 H, s, 14-H), 6.05 (1 H, s,
10-H), 6.11 (1 H, s, 7-H) and 6.87 (1 H, s, 12-H) (Found: Mϩ,
326.2236).
J. Chem. Soc., Perkin Trans. 1, 1997
1409

View Article Online
7 Y. Katsuta, Y. Aoyama, H. Osone, A. Wada, S. Uchiyama,
T. Kitamoto, S. Masushige, S. Kato and M. Ito, Chem. Pharm. Bull.,
1994, 42, 2659.
8 A. Albeck, N. Friedman, M. Sheves and M. Ottolenghi, J. Am.
Chem. Soc., 1986, 108, 4614.
Acknowledgements
We thank Professors S. Masushige and S. Kato for the CAT
assay experiments and valuable discussions.
9 D. Seebach, Synthesis, 1969, 17; M. H. Park, T. Yamamoto and
K. Nakanishi, J. Am. Chem. Soc., 1989, 111, 4997.
10 T. Sato, Y. Wakahara, J. Otera and H. Nozaki, Tetrahedron, 1991,
47, 9773.
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Paper 6/06719K
Received 2nd October 1996
Accepted 17th December 1996
1410
J. Chem. Soc., Perkin Trans. 1, 1997