Synthesis and SAR studies of a novel class of S1P1 receptor antagonists

Article abstract of DOI:10.1016/j.bmc.2007.02.048

A series of Sodium 4-[(4-butoxyphenyl)thio]-2′-substituted-1,1′-biphenyl-3- sulfonates were identified as functional sphingosine-1-phosphate (S1P) antagonists with selectivity for the S1P₁ receptor subtype starting from chemical lead 2, which w

Full text of DOI:10.1016/j.bmc.2007.02.048

Bioorganic & Medicinal Chemistry 15 (2007) 3548–3564
Synthesis and SAR studies of a novel class of
S1P1 receptor antagonists
Tsuyoshi Nakamura,a Kiyoaki Yonesu,b Yumiko Mizuno,a Chie Suzuki,a
Yuki Sakata,b Yoh Takuwa,c Futoshi Narab and Susumu Satoha,*
aMedicinal Chemistry Research Laboratories, Sankyo Co., Ltd, 1-2-58 Hiromachi, Shinagawa-ku, Tokyo 140-8710, Japan
bLead Discovery Research Laboratories, Sankyo Co., Ltd, 1-2-58 Hiromachi, Shinagawa-ku, Tokyo 140-8710, Japan
cDepartment of Physiology, Kanazawa University Graduate School of Medicine, 13-1 Takara-machi, Kanazawa,
Ishikawa 920-8640, Japan
Received 27 September 2006; revised 22 February 2007; accepted 23 February 2007
Available online 3 March 2007
Abstract—A series of Sodium 4-[(4-butoxyphenyl)thio]-20-substituted-1,10-biphenyl-3- sulfonates were identified as functional sphin-
gosine-1-phosphate (S1P) antagonists with selectivity for the S1P1 receptor subtype starting from chemical lead 2, which was found
while screening our in-house compound library. We performed chemical modifications on each regional structure of compound 2,
for example, on the three ring compartments, the benzyl substituents, and the long alkyl chain part. The introduction of a biphenyl
skeletal structure and the installation of a hydroxyl group onto the terminal carbon in the side-chain region resulted in the potent
derivative 35c, which showed >500-fold more potent S1P1 inhibitory activity than lead compound 2. We report herein the synthesis
and structure–activity relationships of structurally novel S1P1 receptor antagonists.
ꢀ 2007 Elsevier Ltd. All rights reserved.
1. Introduction
agents for the cardiovascular disease and angiogenesis
caused by mitogenic cell growth.5
Sphingosine-1-phosphate (S1P) receptors were identified
as a family of G-protein coupled receptors and were sub-
divided into five subtypes (S1P1, S1P2, S1P3, S1P4, S1P5),
Herein, we report on the synthesis of a novel class of
S1P receptor antagonists, particularly S1P1 antagonists,
and their structure–activity relationships.
which are coupled differentially via Gi, Gq, G12/13
,
and Rho to multiple effector systems whose signaling
pathways are linked to transcription factor activation,
cytoskeletal proteins, adhesion molecule expression, and
caspase activities.1 Through these signaling pathways,
S1P receptors can affect diverse biological responses,
including mitogenesis, differentiation, migration, and
apoptosis, and thus are supposed to be involved in a
variety of pathological conditions such as angiogenesis,
inflammation, and cardiovascular diseases.2 Among the
widespread investigations to seek seeds for various
therapeutic agents targeting S1P receptors,3 S1P1 recep-
tor antagonists4 are expected to be effective therapeutic
Our discovery process for a subtype-selective S1P1
receptor antagonist started with the screening of our
in-house compound library. Sodium 2-(4-ethoxyphen-
oxy)-5-(3-octadecyl-5-oxo-4,5-dihydro-1H-pyrazol-1-yl)
benzenesulfonate (2) was found as the new chemical
lead during the screening. Chemical lead 2 possesses
three ring compartments (defined as the A, B, and C
rings, as shown in Fig. 1), a long saturated alkyl chain
region, and a sulfonic acid group on the B ring part as
representative structural features. Its lipid-like struc-
ture, featuring both a long alkyl chain part and a neg-
ative ionizable sulfonic acid group, recalls a structural
resemblance to sphingosine-1-phosphate (S1P) 1. Com-
pound 2 showed moderate human S1P1 antagonistic
activity (IC50 = 17.0 lM) in cAMP assays in human
S1P1 receptors stably expressed in Chinese hamster
ovary (CHO) cell membranes. Our initial effort was
Keywords: Sphingosine-1-phosphate; S1P receptor; Antagonist; Struc-
ture–activity relationships.
*
Corresponding author. Tel.: +81 3 3492 3131; fax: +81 3 5436
8563; e-mail: ssatoh@sankyo.co.jp
0968-0896/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2007.02.048

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3549
Table 2. S1P1 antagonistic activities of the variation of the C ring part
OH
CH3(CH2)9
SO3Na
R
OP(O)(OH)2
OH
NH2
Sphingosine 1-phosphate (1)
SO3Na
O
B
C
Compound
R
IC50 (lM)
N
A
N
OEt
CH3(CH2)16
14a
0.40
O
OC2H5
OC2H5
O
2
14b
14c
20a
20b
20c
20d
20e
20f
>17.0
8.9
Figure 1. Structures of sphingosine-1-phosphate and chemical lead 2.
O
O
directed at improving the S1P1 antagonist activity
along with defining the key structural elements
required for this activity.
C2H5O
0.70
0.40
0.19
0.19
0.52
>2.4
S
S
S
S
S
S
OCH3
2. Chemistry
OC2H5
The test compounds listed in Tables 1–4 were synthe-
sized as outlined in Schemes 1–4.
O(CH2)2CH3
O(CH2)3CH3
O(CH2)5CH3
O(CH2)7CH3
The synthesis of 11a, 12a, 13a, and 14a–c is described in
Scheme 1. To begin, sulfonic acid 3 was treated with eth-
oxyphenol in the presence of LiOH in DMSO to afford
biphenyl ether 4a–c. The nitro group of 4a–c was re-
duced to an amino group by using Pd on carbon as a
catalyst in MeOH under a hydrogen atmosphere. The
obtained amine 5a–c was treated with NaNO2 and aque-
ous HBF4 in EtOH, which afforded diazo species 6a–c,
and successive treatments of various aryl boric acids
and a catalytic amount of Pd(OAc)2 each gave the alde-
Table 1. S1P1 antagonistic activities of A ring and alkyl chain
variations 11a–14a
Table 3. S1P1 antagonistic activities of the variation in the substituent
on the benzyl position
SO3Na
O
CH3(CH2)9
SO3Na
S
R2
R1
R
OCH2CH3
O(CH2)3CH3
Compound
R
IC50 (lM)
Compound
R1
R2
IC50 (lM)
N
CH3(CH2)16
N
20d
25a
25b
25c
H
OH
NH2
H
0.19
0.37
2
17.0
H
O
OH
H
CH3
>8.1
1.0
S
OH
11a
12a
13a
5.5
1.3
4.6
5
2
CH3(CH2)9
CH3(CH2)9
OH
2S
hydes 7a, 8a, 9a, and 10a–c. The creation of a side-chain
moiety via a 1,2-addition reaction with 1-dodec-1-yn-1-
yllithium proceeded smoothly to afford secondary
alcohols, which were purified by silica gel column chro-
matography to give 11a, 12a, 13a, and 14a–c.
3
OH
m
CH3(CH2)9
CH3(CH2)9
OH
To investigate the SAR of the 4-ethoxyphenoxy moiety
of compound 14a, we turned our attention to the prep-
aration of thiophenylether derivatives. The synthetic
route of compounds 20a–f is shown in Scheme 2. Phenyl
14a
0.40
o

3550
T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
Table 4. S1P1 antagonistic activities of side-chain variations
mate group by treatment with a combination of trifluo-
roacetic acid and triethylsilane, or BF3ÆOEt2 and
phenylcarbamate in CH2Cl2, respectively. On the other
hand, compound 19d was oxidized with Dess–Martin
periodinane in CH2Cl2 to give ketone 23, which was
treated with methyl magnesium bromide in THF to af-
ford tertiary alcohol 24. The obtained phenyl sulfonates
22a,b and 24 were saponified with aqueous NaOH in
dioxane to afford the corresponding sodium salt of sul-
fonic acid 25a–c.
SO3Na
X
OH
S
n( )
O(CH2)3CH3
R
Compound
X
n
R
IC50 (lM)
20d
34
CH2
CH2
O
6
6
1
3
5
7
9
CH3
OH
OH
OH
OH
OH
OH
0.19
0.080
>20
0.34
35a
35b
35c
35d
35e
The synthetic route of the variations of the side-chain
moiety is shown in Scheme 4. The alkyne 26 was treated
with 3,4-dihydro-2H-pyran in CH2Cl2 in the presence of
p-TsOH to give 27. The propargyl ethers 29a–e were also
easily synthesized by alkylation of the hydroxy group of
propargylalcohol 28 with various alkyl bromides.7 Each
prepared alkyne part was treated with n-BuLi in THF,
and the resulting alkynyl lithium solution was treated
with aldehyde 18d to obtain the secondary alcohols 30
and 31a–e. The tetrahydropyranyl group on the terminal
hydroxyl group was deprotected by treatment with
PPTS in MeOH to afford diols 32 and 33a–e, whose phe-
nyl sulfonate groups were hydrolyzed by treatment with
aqueous NaOH in dioxane to afford the corresponding
sodium salt of sulfonic acid 34 and 35a–e.
O
O
0.031
0.098
0.13
O
O
2,5-dibromobenzene sulfonate 15 was the starting mate-
rial6 and was treated with 4-hydroxythiophenol in the
presence of triethylamine in DMF to afford thioether
16, whose phenolic hydroxyl group was alkylated by
treatment with alkylhalide and DBU in DMF to provide
17a–f. A Suzuki coupling reaction of bromide 17a–f
with 2-formylbenzeneboric acid proceeded smoothly in
the presence of a catalytic amount of Pd(PPh3)4 and
aqueous K2CO3 in DME to give the key intermediate
aldehyde 18a–f. The 1,2-addition reaction of 18a–f with
1-dodec-1-yn-1-yllithium afforded propargyl alcohol
19a–f smoothly, and saponification with aqueous NaOH
in dioxane afforded the corresponding sodium salt of
sulfonic acid 20. The variations in each structural region
could be synthesized using key reactive intermediates
such as compounds 18 and 19.
3. Results and discussion
The compounds listed in Tables 1–4 were biologi-
cally evaluated for their antagonistic activities in
cAMP assays8 in human S1P1 receptors stably ex-
pressed in Chinese hamster ovary (CHO) cell mem-
branes. During the course of our in-house
compound library screening, compound 2 was found
to show moderate activity. Replacement of the long
alkyl chain substituted pyrazolone ring of 2 with
thiophene and a benzene ring afforded 11–14a, which
showed more potent inhibitory activities, as shown
The variation on the benzyl position of the A ring
moiety could be synthesized in the manner shown in
Scheme 3. The secondary hydroxyl group of 19d was
esterified by treatment with Ac2O and Et3N in CH2Cl2
to afford acetate 21. The acetoxy group of 21 could be
converted alternatively to a hydrogen or phenylcarba-
SO3H
SO3H
SO3H
SO3H
Cl
O
O
O
c
a
b
OC2H5
OC2H5
OC2H5
N+
O2N
H2N
O2N
-
N
5a-c
BF4
4a-c
3
6a-c
a: 4-ethoxy
b: 3-ethoxy
c: 2-ethoxy
SO3H
O
CH3(CH2)9
OH
SO3Na
O
e
OHC
OC2H5
d
OC2H5
Ar
Ar
11a: Ar = 5-(1-hydroxytridec-2-yn-1-yl)thien-2-yl
12a: Ar = 2-(1-hydroxytridec-2-yn-1-yl)thien-3-yl
13a: Ar = 3-(1-hydroxytridec-2-yn-1-yl)phenyl
14a-c: Ar = 2-(1-hydroxytridec-2-yn-1-yl)phenyl
7a: Ar = 5-formylthien-2-yl
8a: Ar = 2-formylthien-3-yl
9a: Ar = 3-formylphenyl
10a-c: Ar = 2-formylphenyl
Scheme 1. Reagents: (a) ethoxyphenol, LiOH, DMSO; (b) H2, Pd–C, MeOH; (c) NaNO2, 48% aq HBF4, EtOH; (d) arylboric acid, Pd(OAc)2,
MeOH; (e) 1-dodecyne, n-BuLi, THF.

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3551
SO3Ph
S
SO3Ph
Br
SO3Ph
S
SO3Ph
S
CHO
c
a
b
OR
Br
Br
OH
Br
OR
18a-f
15
16
17a-f
a: R = CH3
d: R = (CH2)3CH3
e: R = (CH2)5CH3
f : R = (CH2)7CH3
b: R = CH2CH3
c: R = (CH2)2CH3
CH3(CH2)9
SO3Ph
CH3(CH2)9
e
SO3Na
OH
S
OH
S
d
OR
OR
20a-f
19a-f
Scheme 2. Reagents: (a) 4-hydroxythiophenol, Et3N, DMF; (b) alkylhalide, DBU, DMF; (c) 2-formylbenzeneboric acid, Pd(PPh3)4, aq K2CO3,
DME; (d) 1-dodecyne, n-BuLi, THF; (e) aq NaOH, dioxane.
CH3(CH2)9
a
SO3Ph
S
CH3(CH2)9
SO3Ph
S
R
OH
O(CH2)3CH4
O(CH2)3CH4
f
19d
21: R = OAc
22a: R = H
22b: R = NHCO2Ph
b
CH3(CH2)9
SO3Na
S
c
R1
R2
d
O(CH2)3CH4
25a: R1 = H, R2 = H
25b: R1 = H, R2 = NH2
25c: R1 = CH3, R2 = OH
CH3(CH2)9
SO3Ph
CH3
CH3(CH2)9
SO3Ph
S
f
S
O
e
OH
O(CH2)3CH4
O(CH2)3CH4
24
23
Scheme 3. Reagents: (a) Ac2O, Et3N, DMAP, CH2Cl2; (b) triethylsilane, CF3CO2H, CH2Cl2; (c) H2NCO2Ph, BF3ÆOEt2, CH2Cl2; (d) Dess–Martin
periodinane, CH2Cl2; (e) MeMgBr, THF; (f) aq NaOH, dioxane.
in Table 1. Due not only to the enhanced effect of
S1P1 affinity, but also to the synthetical advantage
of alkynyl side-chain derivatives, we shifted focus
to propargyl alcohol analogs. Thiophene analogs
11a and 12a and benzene analogs 13a and 14a
showed the same tendency to display structure–activ-
ity relationships in the A ring. Namely, thiophene
analog 12a, whose thiophene ring bridges between
the B ring part and side-chain part at the 1- and
2-positions, showed 4-fold higher affinity than the
2- and 5-position bridging thiophene analog 11a.
Furthermore, the ortho bridging benzene analog
14a also showed 11-fold more potent IC50 values
than the meta conjoining benzene derivative 13a.
These observations suggested that the conjoining po-
tive 14a, which exhibited 40-fold more potent S1P1
antagonist activity than lead compound 2.
Next, investigations around the SAR of the 4-ethoxy-
phenoxy moiety of compound 14a were performed and
are summarized in Table 2. The transposition of the eth-
oxy group onto the meta- or ortho-position decreased
the affinity to the S1P1 receptor (compound 14a vs 14b
and 14c). Oxo-ether 14a and thio-ether 20b showed al-
most equal IC50 values, suggesting that the tethering
atom between the B and C ring parts did not affect
S1P1 affinity. In the variation of the alkoxy group on
para-position 20a–f, the S1P1 receptor affinity was mod-
erately affected by the alkyl chain length of the alkoxy
group, and was maximized at a chain length of 3–4 car-
bon atoms (compounds 20c and 20d). The synthetic
advantage of the thio-ether analog prompted us to fix
the C ring moiety as a p-butoxy phenylthio group. We
sition of the side-chain region on the A ring
strongly affected the S1P1 affinity, and it was then
maximized with the ortho-substituted benzene deriva-

3552
T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
a
HO
THPO
( )8
( )8
26
SO3Ph
27
X
OH
S
c
d
n( )
OTHP
O(CH2)3CH3
THPO
b
HO
( )n
O
30: X = CH2, n = 6
31a-e: X = O
28
29a-e
a: n = 2 d: n = 8
b: n = 4 e: n = 10
c: n = 6
SO3Ph
S
SO3Na
X
X
OH
OH
S
e
n( )
OH
n( )
OH
O(CH2)3CH3
O(CH2)3CH3
34: X = CH2, n = 6
35a-e: X = O
32: X = CH2, n = 6
33a-e: X = O
Scheme 4. Reagents: (a) 3,4-dihydro-2H-pyran, p-TsOH, CH2Cl2; (b) bromide, NaH, DMF; (c) n-BuLi, THF then aldehyde 18d; (d) PPTS, MeOH;
(e) aq NaOH, dioxane.
then turned our attention to defining the effect of the key
polar functional groups in the structure of compound
20d.
Table 5. Antagonistic activities of compounds 20d and 35c for S1P1,
S1P2, S1P3, and S1P4
Compound
IC50 (lM)
S1P1
S1P2
S1P3
S1P4
In the scaffold of compound 20d, the effects of the substit-
uents on the benzyl position are summarized in Table 3.
The amino derivative 25a was found to retain its S1P1
inhibitory activity, although it did not improve the activ-
ity compared to the hydroxy analog 20d. The S1P1 antag-
onistic activity of compound 25b, which possesses a
hydrogenated structure on the benzyl position, decreased
drastically. The tertiary alcohol 25c also showed moder-
ately weaker activity than 20d, confirming that the sec-
ondary hydroxy group on the benzyl position of the A
ring part of 20d was the accommodating structure.
20d
35c
1.0
0.5
24
11
13
5
40
6
4. Conclusions
Starting from the novel sulfonic acid derivative 2, iden-
tified via high-throughput screening, a SAR was
established and the key structural requirements for
activity were determined. Exploratory derivatization,
focusing on each regional structure of compound 2, re-
vealed that the conjunction pattern between the long al-
kyl chain and the A ring part directly affected the
antagonistic activity for S1P1 receptor. We could then
obtain the biphenyl analog 20d, which exhibited 80-fold
more potent S1P1 antagonistic activity than compound
2. Further SAR studies on the scaffold of compound
20d clarified that the hydroxyl group on the terminal po-
sition of the long alkyl chain evenly enhanced the affin-
ity for each of the S1P receptors, and its activity was
slightly S1P1 selective. The most potent S1P1 antagonist
in this article, 35c, showed sub-micro order S1P1 antag-
onistic activity. Further derivatization based on the
structure of the compounds described in this article is
in progress in order to improve their pharmacokinetic
profiles and enhance their S1P1 antagonistic activities.
The results of the derivatization on the side-chain region
are summarized in Table 4.
Surprisingly, introducing a hydroxy group onto the ter-
minal carbon in the side-chain region clearly enhanced
S1P1 antagonistic activity. Diol derivative 34 showed
2-fold more potent S1P1 antagonistic activity than basic
compound 20d. In the derivatives of propargyl ether
35a–e, the alkyl chain length significantly affected their
activities, which were maximized at n = 5 (compound
35c).
Both compound 20d and hydroxylated analog 35c were
further profiled for selectivity among S1P receptors. The
antagonistic activities of these two compounds for S1P1,
S1P2, S1P3, and S1P4 in FLIPR assays9 in S1P receptors
stably expressed in Chinese hamster ovary (CHO) cell
membranes are summarized in Table 5.
5. Experimental
All melting points were measured with a Yanaco MP-
500D micro melting point apparatus and are uncor-
rected. The IR spectra were measured with a JASCO
FT/IR 8900 spectrophotometer, and the peaks were re-
Both compounds 20d and 35c possess slightly S1P1
selective inhibitory profiles. Comparisons with com-
pound 35c clearly showed that the terminal hydroxy
group on the side-chain part evenly enhanced the inhib-
itory activity for each receptor.
1
corded in cmꢀ1. H NMR spectra were recorded on a
Varian Mercury 400 or 500 spectrometer, with tetra-

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3553
methylsilane as an internal reference. The mass spectra
were recorded using a JEOL JMS-BU 20 or JMS-700
spectrometer. High-resolution mass (HRMS) spectros-
copy was carried out with a JEOL JMS-700V mass
spectrometer. Thin-layer chromatography (TLC) was
used routinely to monitor the progress and the purity
of the compounds and was performed on Merck Kiesel-
gel 60 F254 plates. For the flash column chromatogra-
phy, silica gel (Kieselgel 60, 230–400 mesh) was
employed.
the title compound (52.4 g, 95%) as a white crystalline
solid. Mp 285–300 ꢁC (dec); 1H NMR (400 MHz,
DMSO-d6) d 1.30 (t, 3H, J = 7.0 Hz), 3.95 (q, 2H,
J = 7.0 Hz), 6.46–6.49 (m, 2H), 6.78–6.84 (m, 4H),
7.11 (d, 1H, J = 2.9 Hz); IR (KBr) 3453, 2980, 2930,
2623, 1616, 1567, 1478 cmꢀ1; MS (FAB) m/z: 308
(MꢀH)ꢀ.
5.5. 5-Amino-2-(3-ethoxyphenoxy) benzenesulfonic acid (5b)
The title compound was synthesized in the same manner
as the general procedure for compound 5a. H NMR
1
5.1. 2-(4-Ethoxyphenoxy)-5-nitrobenzenesulfonic acid (4a)
(400 MHz, CDCl3) d 1.38 (t, 3H, J = 7.0 Hz), 3.73 (br
s, 2H), 3.98 (q, 2H, J = 7.0 Hz), 6.55–6.60 (m, 2H),
6.62–6.67 (m, 1H), 6.85 (dd, 1H, J = 8.8 Hz, 2.8 Hz),
6.92 (d, 1H, J = 8.8 Hz), 7.15–7.26 (m, 5H), 7.27–7.34
(m, 2H); IR (thin film) 3475, 3385, 1627, 1588, 1487,
LiOHÆH2O (17.2 g, 712 mmol) was added to a solution
of 2-chloro-5-nitrobenzenesulfonic acid
3
(57.0 g,
240 mmol) and 4-ethoxyphenol (49.7 g, 360 mmol) in
DMSO (215 ml) at room temperature. After stirring
for 0.5 h at 110 ꢁC, the reaction mixture was evaporated
in vacuo. Concd HCl (30 ml) and water (430 ml) were
added to the obtained residue, and the resulting mixture
was purified by ODS (Cosmosil 75C18-PREP) column
chromatography (eluent, H2O/MeOH = 9:1–65:35) to
afford the title compound (60.6 g, 75%) as a white crys-
talline solid. Mp 338–350 ꢁC (dec); 1H NMR (400 MHz,
DMSO-d6) d 1.34 (t, 3H, J = 7.0 Hz), 4.04 (q, 2H,
J = 7.0 Hz), 6.78 (d, 1H, J = 6.8 Hz), 6.99–7.06 (m,
4H), 8.13 (dd, 1H, J = 6.8 Hz, 2.9 Hz), 8.58 (d, 1H,
J = 2.9 Hz).
1368 cmꢀ1
.
5.6. 5-Amino-2-(2-ethoxyphenoxy) benzenesulfonic acid (5c)
The title compound was synthesized in the same manner
as the general procedure for compound 5a. H NMR
1
(400 MHz, CDCl3) d 1.29 (t, 3H, J = 7.0 Hz), 3.59 (br
s, 2H), 4.08 (q, 2H, J = 7.0 Hz), 6.64 (d, 1H,
J = 8.8 Hz), 6.77 (dd, 1H, J = 8.8 Hz, 2.9 Hz), 6.94 (td,
1H, J = 7.9 Hz, 1.5 Hz), 7.01 (dd, 1H, J = 7.9 Hz,
1.5 Hz), 7.08 (dd, 1H, J = 7.9 Hz, 1.7 Hz), 7.12 (d, 1H,
J = 2.9 Hz), 7.15 (td, 1H, J = 7.9 Hz, 1.7 Hz), 7.20–
7.26 (m, 1H), 7.28–7.35 (m, 4H); IR (KBr) 3386, 1625,
5.2. 2-(3-Ethoxyphenoxy)-5-nitrobenzenesulfonic acid (4b)
1490, 1371 cmꢀ1
.
The title compound was synthesized in the same manner
as the general procedure for compound 4a. H NMR
1
5.7. Sodium 4-(4-ethoxyphenoxy)-20-(1-hydroxytridec-2-
yn-1-yl)biphenyl-3-sulfonate (14a)
(400 MHz, CD3OD) d 1.43 (t, 3H, J = 7.0 Hz), 4.04 (q,
2H, J = 7.0 Hz), 6.69 (t, 1H, J = 2.3 Hz), 6.72 (dd, 1H,
J = 8.1 Hz, 2.3 Hz), 6.87 (dd, 1H, J = 8.4 Hz, 2.3 Hz),
7.04 (d, 1H, J = 9.2 Hz), 7.20–7.24 (m, 2H), 7.27–7.32
(m, 1H), 7.33–7.40 (m, 3H), 8.36 (dd, 1H, J = 9.2 Hz,
2.8 Hz), 8.81 (d, 1H, J = 2.8 Hz); IR (thin film) 1607,
48% aqueous HBF4 solution (5 ml) was added to the
solution of 5-amino-2-(4-ethoxyphenoxy) benzenesulf-
onic acid 5a (1.0 g, 3.2 mmol) in EtOH (5 ml), and
NaNO2 (0.89 g, 13.0 mmol) was added to the resulting
solution at 0 ꢁC. After stirring for 0.5 h, the precipitate
was filtered off to afford the crude compound 6a, which
was dissolved in MeOH (3.5 ml). 2-Formylphenylboron-
ic acid (0.63 g, 4.2 mmol) and Pd(OAc)2 (94 mg,
0.42 mmol) were added to the reaction mixture at room
temperature. After stirring for 2 h, the reaction mixture
was filtered through Celite and the filtrate was evapo-
rated in vacuo. The residue obtained was purified by
short column chromatography on silica gel (eluent,
MeOH/CH2Cl2 = 1:5) to afford the crude compound
10a (0.18 g). A solution of crude compound 10a in
THF (1.0 ml) was added to a mixture of 1-dodecyne
(0.20 ml, 1.2 mmol) and 1.6 M solution of n-BuLi in
hexane (0.85 ml, 1.4 mmol) in THF (5 ml) at 0 ꢁC. After
stirring for 0.5 h at room temperature, 1 M aqueous
NaOH (2.0 ml) was added to the reaction mixture and
evaporated in vacuo. The obtained residue was purified
by thin layer chromatography on silica gel (MeOH/
CH2Cl2 = 1:5) to afford the title compound (77 mg,
30%) as an amorphous solid. 1H NMR (400 MHz,
CD3OD) d 0.89 (t, 9H, J = 6.6 Hz), 1.26–1.35 (brs,
14H), 1.40 (t, 3H, J = 7.3 Hz), 1.43–1.49 (m, 2H), 2.20
(dt, 2H, J = 2.2 Hz, 7.3 Hz), 4.03 (q, 2H, J = 6.6 Hz),
1579, 1528, 1488, 1470 cmꢀ1
.
5.3. 2-(2-Ethoxyphenoxy)-5-nitrobenzenesulfonic acid (4c)
The title compound was synthesized in the same manner
as the general procedure for compound 4a. H NMR
1
(400 MHz, CD3OD) d 1.25 (t, 3H, J = 7.0 Hz), 4.08 (q,
2H, J = 7.0 Hz), 6.85 (d, 1H, J = 9.2 Hz), 7.06 (td, 1H,
J = 7.8 Hz, 1.4 Hz), 7.09 (dd, 1H, J = 7.8 Hz, 1.4 Hz),
7.23 (dd, 1H, J = 1.7 Hz, 7.8 Hz), 7.26–7.36 (m, 6H),
8.31 (dd, 1H, J = 9.2 Hz, 2.8 Hz), 8.72 (d, 1H,
J = 2.8 Hz); IR (thin film) 1605, 1526, 1498, 1471,
1380 cmꢀ1
.
5.4. 5-Amino-2-(4-ethoxyphenoxy) benzenesulfonic acid (5a)
7.5% Pd–C (8.7 g) was added to a solution of 2-(4-eth-
oxyphenoxy)-5-nitrobenzenesulfonic acid 4a (60.6 g,
196 mmol) in H2O (350 ml) and MeOH (350 ml), and
the air was replaced with hydrogen. After stirring for
2.5 h at room temperature, the reaction mixture was fil-
tered through Celite, and the filtrate was evaporated in
vacuo. The obtained residue was recrystallized from
MeOH (90 ml) and acetonitrile (1000 ml) to afford

3554
T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
5.33 (d, 1H, J = 2.2 Hz), 6.93 (d, 2H, J = 9.5 Hz), 7.13
(d, 2H, J = 9.5 Hz), 7.21 (dd, 1H, J = 1.5 Hz, 8.8 Hz),
7.33–7.39 (m, 3H), 7.79 (dd, 1H, J = 1.5 Hz, 8.8 Hz),
7.97 (d, 1H, J = 2.2 Hz); IR (KBr) 3392, 2925, 1473,
J = 8.5 Hz, 2.4 Hz), 7.77 (s, 1H), 8.25 (d, 1H,
J = 2.4 Hz); IR (KBr) 1587, 1606, 2854, 2926,
3409 cmꢀ1
;
HRMS (ESI, positive) calcd for
C33H39Na2O6S (M+Na)+ 609.2263; obsd 609.2260.
1232 cmꢀ1
;
HRMS (ESI, negative) calcd for
C33H39O6S (MꢀH)ꢀ 563.2467; obsd 563.2465.
5.12. Sodium 4-(2-ethoxyphenoxy)-20-(1-hydroxytridec-2-
yn-1-yl)biphenyl-3-sulfonate (14c)
5.8. Sodium 2-(4-ethoxyphenoxy)-5-[5-(1-hydroxytridec-
2-yn-1-yl)-2-thienyl]benzenesulfonate (11a)
The title compound (15 mg, 26% from 5c) was synthe-
sized in the same manner as the general procedure for
compound 14a. H NMR (400 MHz, CD3OD) d 0.88
1
The title compound (76 mg, 45% from 5a) was synthe-
sized in the same manner as the general procedure for
(t, 3H, J = 6.9 Hz), 1.25 (t, 3H, J = 7.0 Hz), 1.26 (brs,
12H), 1.37–1.46 (m, 2H), 1.48–1.57 (m, 2H), 2.27 (dt,
2H, J = 6.9 Hz, 2.0 Hz), 4.10 (q, 2H, J = 7.0 Hz), 5.43
(d, 1H, J = 2.0 Hz), 6.70 (d, 1H, J = 8.5 Hz), 6.97 (dt,
1H, J = 7.6 Hz, 1.6 Hz), 7.20 (dd, 1H, J = 8.1 Hz,
1.6 Hz), 7.14-7.21 (m, 2H), 7.41 (t, 1H, J = 7.7 Hz),
7.48 (d, 1H, J = 7.7 Hz), 7.53–7.58 (m, 2H), 7.75 (s,
1H), 8.24 (d, 1H, J = 2.4 Hz); IR (KBr) 1475, 1498,
1604, 2854, 2926, 3393 cmꢀ1; HRMS (ESI, positive)
calcd for C30H40Na2O6S (M+Na)+ 587.2444; obsd
587.2451.
1
compound 14a. H NMR (400 MHz, CD3OD) d 0.88
(t, 3H, J = 6.8 Hz), 1.20–1.35 (m, 15H), 1.38 (t, 3H,
J = 6.8 Hz), 1.40–1.60 (m, 4H), 2.29 (dt, 2H,
J = 6.8 Hz, 2.0 Hz), 4.03 (q, 2H, J = 6.8 Hz), 5.56 (br,
1H), 6.76 (d, 1H, J = 8.8 Hz), 6.92 (d, 2H, J = 8.8 Hz),
7.06 (d, 1H, J = 3.7 Hz), 7.09 (d, 2H, J = 8.8 Hz), 7.17
(d, 1H, J = 3.7 Hz), 7.55 (dd, 1H, J = 8.8 Hz, 2.0 Hz),
8.19 (d, 1H, J = 2.0 Hz).
5.9. Sodium 2-(4-ethoxyphenoxy)-5-[3-(1-hydroxytridec-
2-yn-1-yl)-2-thienyl]benzenesulfonate (12a)
5.13. Phenyl 2,5-dibromobenzenesulfonate (15)
The title compound (59 mg, 35% from 5a) was synthe-
sized in the same manner as the general procedure for
compound 14a. H NMR (400 MHz, CD3OD) d 0.90
(t, 3H, J = 6.8 Hz), 1.24–1.35 (m, 15H), 1.40 (t, 3H,
J = 6.8 Hz), 1.50 (m, 2H), 2.16 (s, 1H), 2.23 (t, 2H,
J = 6.8 Hz), 4.04 (q, 2H, J = 6.8 Hz), 5.57 (s, 1H), 6.80
(d, 1H, J = 8.8 Hz), 6.94 (d, 2H, J = 8.8 Hz), 7.05 (d,
1H, J = 4.9 Hz), 7.11 (d, 2H, J = 8.8 Hz), 7.38 (d, 1H,
J = 4.9 Hz), 7.48 (dd, 1H, J = 8.8 Hz, 2.0 Hz), 8.07 (d,
1H, J = 2.0 Hz).
Chlorosulfonic acid (50 ml) was added to a solution of
1,4-dibromobenzene (50 g, 212 mmol) in chloroform
(150 ml) at room temperature. After stirring for 3 h at
80 ꢁC, the reaction mixture was cooled to room temper-
ature and poured into crushed ice, and extracted with
ether. The combined organic layer was washed with
brine, dried over MgSO4, and filtered. The filtrate was
evaporated in vacuo to give a crude product of sulfonyl
chloride. This residual crude product was dissolved in
CH2Cl2 (200 ml), to which were added PhOH (16.4 g,
174 mmol) and Et3N (29 ml, 209 mmol), and the mix-
ture was stirred for 1 h at 0 ꢁC. After the addition of
satd NaHCO3 (20 ml), the resulting mixture was poured
into water and extracted with ether. The combined or-
ganic layer was washed with brine, dried over MgSO4,
and filtered. The filtrate was evaporated in vacuo to give
the crude product. Recrystallization from MeOH gave
the title compound (54.1 g, 65%) as a white crystalline
1
5.10. Sodium 4-(4-ethoxyphenoxy)-30-(1-hydroxytridec-2-
yn-1-yl)biphenyl-3-sulfonate (13a)
The title compound (172 mg, 53% from 5a) was synthe-
sized in the same manner as the general procedure for
1
compound 14a. H NMR (400 MHz, CD3OD) d 0.88
(t, 3H, J = 6.8 Hz), 1.16–1.33 (m, 15H), 1.39 (t, 3H,
J = 7.8 Hz), 1.53 (quintet, 2H, J = 6.8 Hz), 2.27 (t, 2H,
J = 5.9 Hz), 4.03 (q, 2H, J = 7.3 Hz), 4.58 (brs, 1H),
5.43 (brs, 1H), 6.83 (d, 2H, J = 8.8 Hz), 6.93 (d, 2H,
J = 8.8 Hz), 7.11 (d, 2H, J = 8.8 Hz), 7.40 (d, 2H,
J = 8.1 Hz), 7.48 (d, 1H, J = 7.3 Hz), 7.54 (d, 1H,
J = 7.3 Hz), 7.59 (d, 1H, J = 7.3 Hz), 7.60 (d, 1H,
J = 8.1 Hz), 7.75 (s, 1H).
1
solid. Mp 104–106 ꢁC; H NMR (500 MHz, CDCl3) d
7.14 (d, 2H, J = 7.8 Hz), 7.27–7.36 (m, 3H), 7.59 (dd,
1H, J = 2.9 Hz, 8.8 Hz), 7.69 (d, 1H, J = 8.8 Hz), 8.08
(d, 1H, J = 2.9 Hz); IR (KBr) 1485, 1446, 1387, 1197,
1023 cmꢀ1
;
HRMS (FAB, positive) calcd for
C12H8Br2O3S (M)+ 398.8561; obsd 398.8566.
5.11. Sodium 4-(3-ethoxyphenoxy)-20-(1-hydroxytridec-2-
yn-1-yl)biphenyl-3-sulfonate (14b)
5.14. Phenyl 5-bromo-2-[(4-hydroxyphenyl)thio]benzene-
sulfonate (16)
The title compound (19 mg, 41% from 5b) was synthe-
sized in the same manner as the general procedure for
compound 14a. H NMR (400 MHz, CD3OD) d 0.88
(t, 3H, J = 6.9 Hz), 1.26 (brs, 12H), 1.37 (t, 3H,
J = 7.0 Hz), 1.38–1.46 (m, 2H), 1.48–1.57 (m, 2H), 2.28
(dt, 2H, J = 6.9 Hz, 2.0 Hz), 4.01 (q, 2H, J = 7.0 Hz),
4.56 (brs, 1H), 5.44 (s, 1H), 6.67–6.74 (m, 2H), 6.75 (t,
1H, J = 2.3 Hz), 6.93 (d, 1H, J = 8.5 Hz), 7.24 (t, 1H,
J = 8.2 Hz), 7.42 (t, 1H, J = 7.7 Hz), 7.49 (t, 1H,
J = 7.7 Hz), 7.56 (1H, J = 7.7 Hz), 7.62 (dd, 1H,
N,N-diisopropylethylamine (57.0 ml, 336 mmol) and 4-
hydroxythiophenol (21.2 g, 168 mmol) were added to a
solution of compound 15 (44.0 g, 112 mmol) in DMF
(100 ml). After stirring for 6 h at 80 ꢁC, satd NaHCO3
(20 ml) was added to the reaction mixture. The resulting
mixture was poured into water and extracted with ether.
The combined organic layer was washed with 3 M aque-
ous HCl (200 ml) and brine, dried over MgSO4, and fil-
tered. The filtrate was evaporated in vacuo to give the
crude product, which was purified by silica gel column
1

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3555
chromatography. Elution with EtOAc/hexane (1:4–2:3)
afforded the title compound (39.1 g, 77%) as a pale yel-
low crystalline solid. Mp 148–150 ꢁC; 1H NMR
(500 MHz, CDCl3) d 5.63 (brs, 1H), 6.76 (d, 1H,
J = 8.8 Hz), 6.95 (d, 2H, J = 8.8 Hz), 7.20 (d, 2H,
J = 7.8 Hz), 7.29 (t, 1H, J = 7.8 Hz), 7.36 (t, 2H,
J = 7.8 Hz), 7.42 (dd, 1H, J = 2.9 Hz, 8.8 Hz), 7.45 (d,
2H, J = 8.8 Hz), 7.96 (d, 1H, J = 2.9 Hz); IR (KBr)
1.90 (m, 2H), 3.98 (t, 2H, J = 6.5 Hz), 6.75 (d, 1H,
J = 8.6 Hz), 6.99 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.18–
7.23 (m, 2H), 7.26–7.32 (m, 1H), 7.33–7.38 (m, 2H),
7.40 (dd, 1H, J = 2.2 Hz, 8.6 Hz), 7.48 (dd, 2H,
J = 2.1 Hz, 8.8 Hz), 7.96 (d, 1H, J = 2.2 Hz); IR (thin
film) 1592, 1489, 1443, 1384, 1250 cmꢀ1; HRMS (ESI,
positive) calcd for C21H19O4BrS2Na (M+Na)+
500.9806; obsd 500.9781.
3472, 1599, 1584, 1495, 1487, 1443, 1379, 1361 cmꢀ1
;
HRMS (FAB, positive) calcd for C18H13BrO4S2 (M)+
453.9439; obsd 453.9436.
5.19. Phenyl 5-bromo-2-[(4-hexyloxyphenyl)thio]ben-
zenesulfonate (17e)
5.15. Phenyl 5-bromo-2-[(4-butoxyphenyl)thio]benzene-
sulfonate (17d)
The title compound was synthesized in the same manner
as the general procedure for compound 17d. H NMR
1
(400 MHz, CDCl3) d 0.95 (t, 3H, J = 7.1 Hz), 1.35–
1.52 (m, 4H), 1.78–1.87 (m, 2H), 4.01 (t, 2H,
J = 6.6 Hz), 6.75 (d, 1H, J = 8.6 Hz), 6.99 (dd, 2H,
J = 2.1 Hz, 8.8 Hz), 7.18–7.23 (m, 2H), 7.26–7.32 (m,
1H), 7.33–7.38 (m, 2H), 7.40 (dd, 1H, J = 2.2 Hz,
8.6 Hz), 7.48 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.96 (d, 1H,
J = 2.2 Hz); IR (KBr) 1594, 1489, 1442, 1383,
1-Bromobutane (3.2 ml, 30 mmol) and DBU (4.6 ml,
30 mmol) were added to a solution of compound 16
(8.7 g, 20.0 mmol) in DMF (20 ml). After stirring for 4 h
at 60 ꢁC, the reaction mixture was cooled to room temper-
atureandpouredintowater(50 ml)followedbyextraction
with ether. The combined organic layer was washed with
brine, dried over MgSO4, and filtered. The filtrate was
evaporated in vacuo to give the crude product. Recrystal-
lization from AcOEt/hexane(1:9)gave thetitlecompound
(8.1 g, 82%) as a white crystalline solid. Mp 80–82 ꢁC; 1H
NMR (500 MHz, CDCl3) d 1.00 (t, 3H, J = 6.8 Hz), 1.51
(sextet, 2H, J = 6.8 Hz), 1.80 (quintet, 2H, J = 6.8 Hz),
4.01 (t, 2H, J = 6.8 Hz), 6.75 (d, 1H, J = 8.8 Hz), 6.98 (d,
2H, J = 8.8 Hz), 7.20 (d, 2H, J = 7.8 Hz), 7.28 (t, 1H,
J = 6.8 Hz), 7.35 (t, 2H, J = 8.8 Hz), 7.40 (dd, 1H,
J = 2.0 Hz, 8.8 Hz), 7.47 (d, 2H, J = 8.8 Hz), 7.96 (d, 1H,
J = 2.0 Hz); IR (KBr) 1592, 1489, 1443, 1384,
1252 cmꢀ1
;
HRMS (ESI, positive) calcd for
C23H23O4BrS2Na (M+Na)+ 529.0119; obsd 529.0155.
5.20. Phenyl 5-bromo-2-[(4-octyloxyphenyl)thio]benzene-
sulfonate (17f)
The title compound was synthesized in the same manner
as the general procedure for compound 17d. H NMR
1
(400 MHz, CDCl3) d 0.90 (t, 3H, J = 6.9 Hz), 1.24–
1.41 (m, 8H), 1.44–1.53 (m, 2H), 1.77–1.86 (m, 2H),
4.00 (t, 2H, J = 6.5 Hz), 6.75 (d, 1H, J = 8.6 Hz), 6.99
(dd, 2H, J = 8.8 Hz, 2.1 Hz), 7.18–7.23 (m, 2H), 7.26–
7.32 (m, 1H), 7.33–7.38 (m, 2H), 7.40 (dd, 1H,
J = 8.8 Hz, 2.1 Hz), 7.48 (dd, 2H, J = 8.6 Hz, 2.2 Hz),
7.96 (d, 1H, J = 2.2 Hz); IR (thin film) 2854, 1592,
1490, 1442, 1379 cmꢀ1; MS (FAB) m/z: 548 (M)+.
1249 cmꢀ1
;
HRMS (FAB, positive) calcd for
C22H21BrO4S2 (M)+ 492.0065; obsd 492.0054.
5.16. Phenyl 5-bromo-2-[(4-methoxyphenyl)thio]benzene-
sulfonate (17a)
The title compound was synthesized in the same manner
as the general procedure for compound 17d. H NMR
5.21. Phenyl 4-[(4-butoxyphenyl)thio]-20-formylbiphenyl-
3-sulfonate (18d)
1
(400 MHz, CDCl3) d 3.87 (s, 3H), 6.75 (d, 1H,
J = 8.6 Hz), 7.00 (dd, 2H, J = 8.8 Hz, 2.1 Hz), 7.19–
7.24 (m, 2H), 7.27–7.38 (m, 3H), 7.41 (dd, 1H,
J = 8.6 Hz, 2.2 Hz), 7.50 (dd, 2H, J = 8.8 Hz, 2.1 Hz),
7.97 (d, 1H, J = 2.2 Hz); IR (thin film) 1588, 1492,
1443, 1377 cmꢀ1; MS (FAB) m/z: 473 (M+Na)+.
4.6 M aqueous solution of K2CO3 (10 ml, 46 mmol)
was added to a solution of compound 17d (7.6 g,
15.4 mmol), 2-formylbenzeneboric acid (2.8 g,
18.5 mmol), and Pd(PPh3)4 (0.80 g, 0.7 mmol) in
dimethoxyethane (20 ml). After stirring for 5 h at
60 ꢁC, the reaction mixture was poured into water
(30 ml) and extracted with ether. The combined organ-
ic layer was washed with brine, dried over MgSO4,
and filtered. The filtrate was evaporated in vacuo to
give the crude product, which was purified by silica
gel column chromatography. Elution with EtOAc/hex-
ane (1:9) afforded the title compound (6.8 g, 90%) as a
white crystalline solid. Mp 95–97 ꢁC; 1H NMR
(500 MHz, CDCl3) d 1.00 (t, 3H, J = 6.8 Hz), 1.51
(sextet, 2H, J = 6.8 Hz), 1.80 (quintet, 2H,
J = 6.8 Hz), 4.00 (q, 2H, J = 6.8 Hz), 6.98–7.01 (m,
3H), 7.18–7.29 (m, 3H), 7.34 (t, 2H, J = 6.8 Hz),
7.52–7.60 (m, 4H), 7.73 (d, 1H, J = 7.8 Hz), 7.87 (d,
1H, J = 7.8 Hz), 7.96 (s, 1H), 8.10 (s, 1H), 10.05 (s,
1H); IR (KBr) 1700, 1594, 1489, 1250, 1194, 1174,
5.17. Phenyl 5-bromo-2-[(4-ethoxyphenyl)thio]benzene-
sulfonate (17b)
The title compound was synthesized in the same manner
as the general procedure for compound 17d. H NMR
1
(400 MHz, CDCl3) d 1.46 (t, 3H, J = 7.0 Hz), 4.08 (q,
2H, J = 7.0 Hz), 6.73 (d, 1H, J = 8.6 Hz), 6.98 (d, 2H,
J = 8.6 Hz), 7.20 (d, 2H, J = 8.6 Hz), 7.26–7.43 (m,
4H), 7.48 (d, 2H, J = 8.6 Hz), 7.95 (d, 1H, J = 2.0 Hz).
5.18. Phenyl 5-bromo-2-[(4-propoxyphenyl)thio]benzene-
sulfonate (17c)
The title compound was synthesized in the same manner
as the general procedure for compound 17d. H NMR
1
1145 cmꢀ1
C27H22O5S2 (M)+ 490.0909; obsd 490.0897.
;
HRMS (FAB, positive) calcd for
(400 MHz, CDCl3) d 1.07 (t, 3H, J = 7.4 Hz), 1.80–

3556
T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
5.22. Phenyl 4-[(4-methoxyphenyl)thio]-20-formylbiphenyl-
3-sulfonate (18a)
(7.7 ml, 36 mmol) in THF (40 ml) at ꢀ45 ꢁC. After stir-
ring for 10 min, a solution of compound 18d (9.8 g,
20 mmol) in THF (10 ml) was added to the reaction mix-
ture. After stirring for 1 h, satd NH4Cl (10 ml) was
added to the reaction mixture. The resulting mixture
was poured into water and extracted with ether. The
combined organic layer was washed with brine, dried
over MgSO4, and filtered. The filtrate was evaporated
in vacuo to give the crude product, which was purified
by silica gel column chromatography. Elution with
EtOAc/hexane (1:3–1:1) afforded the title compound
(11.4 g, 83%) as a colorless oil. 1H NMR (500 MHz,
CDCl3) d 0.88 (t, 3H, J = 6.8 Hz), 1.00 (t, 3H,
J = 7.8 Hz), 1.20–1.38 (m, 14H), 1.45–1.55 (m, 4H),
1.81 (quintet, 2H, J = 8.8 Hz), 2.20 (dt, 2H, J = 2.0 Hz,
6.8 Hz), 4.03 (t, 2H, J = 6.8 Hz), 5.01 (s, 1H), 6.93 (d,
1H, J = 8.8 Hz), 7.01 (d, 2H, J = 8.8 Hz), 7.06 (d, 1H,
J = 7.8 Hz), 7.21 (d, 2H, J = 7.8 Hz), 7.25–7.42 (m,
6H), 7.55 (d, 1H, J = 7.8 Hz), 7.80 (d, 1H, J = 7.8 Hz),
7.91 (d, 1H, J = 2.0 Hz); IR (liquid film) 3535, 1593,
1489, 1458, 1388, 1367, 1248 cmꢀ1; HRMS (ESI, posi-
tive) calcd for C41H48NaO5S2 (M+Na)+ 707.2841; obsd
707.2836.
The title compound was synthesized in the same manner
as the general procedure for compound 18d. H NMR
1
(400 MHz, CDCl3) d 3.87 (s, 3H), 6.75 (d, 1H,
J = 8.6 Hz), 7.00 (dd, 2H, J = 8.8 Hz, 2.1 Hz), 7.19–
7.24 (m, 2H), 7.27–7.38 (m, 3H), 7.41 (dd, 1H,
J = 8.6 Hz, 2.2 Hz), 7.50 (dd, 2H, J = 8.8 Hz, 2.1 Hz),
7.97 (d, 1H, J = 2.2 Hz), 8.08 (s, 1H), 10.04 (s, 1H).
5.23. Phenyl 4-[(4-ethoxyphenyl)thio]-20-formylbiphenyl-
3-sulfonate (18b)
The title compound was synthesized in the same manner
as the general procedure for compound 18d. H NMR
1
(400 MHz, CDCl3) d 1.47 (t, 3H, J = 6.8 Hz), 4.11 (q,
2H, J = 6.8 Hz), 6.98–7.03 (m, 3H), 7.22–7.29 (m, 3H),
7.34 (t, 2H, J = 6.8 Hz), 7.52–7.60 (m, 4H), 7.71 (d,
1H, J = 7.8 Hz), 7.85 (d, 1H, J = 7.8 Hz), 7.95 (s, 1H),
8.08 (s, 1H), 10.04 (s, 1H); IR (KBr) 1700, 1594, 1489,
1250, 1194, 1174, 1145 cmꢀ1
.
5.24. Phenyl 4-[(4-propoxyphenyl)thio]-20-formylbiphenyl-
3-sulfonate (18c)
5.28. Phenyl 4-[(4-methoxyphenyl)thio]-20-(1-hydroxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (19a)
The title compound was synthesized in the same manner
as the general procedure for compound 18d. H NMR
1
The title compound was synthesized in the same manner
as the general procedure for compound 19d. H NMR
1
(400 MHz, CDCl3) d 1.07 (t, 3H, J = 7.4 Hz), 1.80–
1.90 (m, 2H), 3.98 (t, 2H, J = 6.5 Hz), 6.98–7.03 (m,
3H), 7.22–7.29 (m, 3H), 7.34 (t, 2H, J = 6.8 Hz), 7.52–
7.60 (m, 4H), 7.71 (d, 1H, J = 7.8 Hz), 7.85 (d, 1H,
J = 7.8 Hz), 7.95 (s, 1H), 8.08 (s, 1H), 10.04 (s, 1H).
(400 MHz, CDCl3) d 0.89 (t, 3H, J = 6.9 Hz), 1.27 (br
s, 12H), 1.32–1.39 (m, 2H), 1.46–1.55 (m, 2H), 1.88 (d,
1H, J = 5.2 Hz), 2.22 (dt, 2H, J = 7.2 Hz, 1.9 Hz), 3.90
(s, 3H), 5.00–5.03 (m, 1H), 6.94 (d, 1H, J = 8.1 Hz),
7.04 (dd, 2H, J = 8.9 Hz, 2.1 Hz), 7.09 (dd, 1H,
J = 7.6 Hz, 1.2 Hz), 7.22–7.25 (m, 2H), 7.27–7.44 (m,
6H), 7.59 (dd, 2H, J = 2.1 Hz, 8.9 Hz), 7.82 (dd, 1H,
J = 7.8 Hz, 1.3 Hz), 7.93 (d, 1H, J = 2.0 Hz); IR (thin
film) 2925, 2854, 1591, 1493, 1458, 1387 cmꢀ1; MS
(ESI) m/z: 665 (M+Na)+.
5.25. Phenyl 4-[(4-hexyloxyphenyl)thio]-20-formylbiphenyl-
3-sulfonate (18e)
The title compound was synthesized in the same manner
as the general procedure for compound 18d. H NMR
1
(400 MHz, CDCl3) d 0.95 (t, 3H, J = 7.1 Hz), 1.35–
1.52 (m, 4H), 1.78–1.87 (m, 2H), 4.01 (t, 2H,
J = 6.6 Hz), 7.00 (dd, 2H, J = 8.8 Hz, 2.1 Hz), 7.19–
7.24 (m, 2H), 7.27–7.38 (m, 3H), 7.41 (dd, 1H,
J = 8.6 Hz, 2.2 Hz), 7.50 (dd, 2H, J = 8.8 Hz, 2.1 Hz),
7.97 (d, 1H, J = 2.2 Hz), 8.08 (s, 1H), 10.04 (s, 1H).
5.29. Phenyl 4-[(4-ethoxyphenyl)thio]-20-(1-hydroxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (19b)
The title compound was synthesized in the same manner
as the general procedure for compound 19d. H NMR
1
(400 MHz, CDCl3) d 0.88 (t, 3H, J = 6.8 Hz), 1.47 (t,
3H, J = 7.8 Hz), 1.20–1.38 (m, 14H), 1.81 (quintet, 2H,
J = 8.8 Hz), 2.20 (dt, 2H, J = 2.0 Hz, 6.8 Hz), 4.08 (t,
2H, J = 6.8 Hz), 5.01 (s, 1H), 6.93 (d, 1H, J = 8.8 Hz),
7.01 (d, 2H, J = 8.8 Hz), 7.06 (d, 1H, J = 7.8 Hz), 7.21
(d, 2H, J = 7.8 Hz), 7.25–7.42 (m, 6H), 7.55 (d, 1H,
J = 7.8 Hz), 7.80 (d, 1H, J = 7.8 Hz), 7.91 (d, 1H,
J = 2.0 Hz).
5.26. Phenyl 4-[(4-octyloxyphenyl)thio]-20-formylbiphe-
nyl-3-sulfonate (18f)
The title compound was synthesized in the same manner
as the general procedure for compound 18d. H NMR
1
(400 MHz, CDCl3) d 0.90 (t, 3H, J = 6.9 Hz), 1.24–1.41
(m, 8H), 1.44–1.53 (m, 2H), 1.77–1.86 (m, 2H), 4.00 (t,
2H, J = 6.5 Hz), 7.00 (dd, 2H, J = 8.8 Hz, 2.1 Hz),
7.19–7.24 (m, 2H), 7.25–7.36 (m, 3H), 7.42 (dd, 1H,
J = 8.6 Hz, 2.2 Hz), 7.48 (dd, 2H, J = 8.8 Hz, 2.1 Hz),
7.99 (d, 1H, J = 2.2 Hz), 8.08 (s, 1H), 10.03 (s, 1H).
5.30. Phenyl 4-[(4-propoxyphenyl)thio]-20-(1-hydroxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (19c)
The title compound was synthesized in the same manner
as the general procedure for compound 19d. H NMR
(400 MHz, CDCl3) d 0.88 (t, 3H, J = 6.9 Hz), 1.07 (t,
3H, J = 7.4 Hz), 1.25 (brs, 12H), 1.30–1.38 (m, 2H),
1.43–1.52 (m, 2H), 1.81–1.92 (m, 3H), 2.20 (td, 2H,
J = 1.9 Hz, 7.1 Hz), 3.99 (t, 2H, J = 6.5 Hz), 4.98–5.02
1
5.27. Phenyl 4-[(4-butoxyphenyl)thio]-20-(1-hydroxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (19d)
1.6 M solution of n-butyllithium in hexane (19.0 ml,
30 mmol) was added to a solution of 1-dodecyne

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3557
(m, 1H), 6.93 (d, 1H, J = 8.3 Hz), 7.02 (dd, 2H,
J = 2.0 Hz, 8.8 Hz), 7.08 (dd, 1H, J = 1.2 Hz, 7.6 Hz),
7.20–7.25 (m, 2H), 7.27–7.44 (m, 6H), 7.56 (dd, 2H,
J = 2.0 Hz, 8.8 Hz), 7.81 (dd, 1H, J = 1.1 Hz, 7.8 Hz),
7.91 (d, 1H, J = 1.9 Hz); IR (thin film) 3530, 2926,
1592, 1489, 1458, 1248 cmꢀ1; HRMS (ESI, positive)
calcd for C40H46O5S2Na (M+Na)+ 693.2684; obsd
693.2707.
5.34. Sodium 20-(1-hydroxytridec-2-yn-1-yl)-4-[(4-meth-
oxyphenyl)thio]biphenyl-3-sulfonate (20a)
The title compound (50 mg, 65% from 19a) was synthe-
sized in the same manner as the general procedure for
20d. 1H NMR (400 MHz, CD3OD) d 0.89 (t, 3H,
J = 6.9 Hz), 1.20–1.33 (m, 12H), 1.30–1.38 (m, 2H),
1.41–1.50 (m, 2H), 2.19 (td, 2H, J = 6.9 Hz, 2.0 Hz),
3.85 (s, 3H), 5.29 (t, 1H, J = 2.1 Hz), 6.82 (d, 1H,
J = 8.2 Hz), 7.02 (dd, 2H, J = 8.8 Hz, 2.1 Hz), 7.19
(dd, 1H, J = 7.6 Hz, 1.4 Hz), 7.22 (dd, 1H, J = 8.2 Hz,
2.1 Hz), 7.32 (td, 1H, J = 7.6 Hz, 1.4 Hz), 7.38 (td, 1H,
J = 7.6 Hz, 1.4 Hz), 7.57 (dd, 2H, J = 8.8 Hz, 2.1 Hz),
7.79 (dd, 1H, J = 7.6 Hz, 1.4 Hz), 7.99 (d, 1H,
J = 2.1 Hz); IR (KBr) 1444, 1458, 1495, 1592, 2854,
2926, 3423 cmꢀ1; HRMS (ESI, positive) calcd for
C32H37Na2O5S2 (M+Na)+ 611.1878; obsd 611.1871.
5.31. Phenyl 4-[(4-hexyloxyphenyl)thio]-20-(1-hydroxytri-
dec-2-yn-1-yl)biphenyl-3-sulfonate (19e)
The title compound was synthesized in the same manner
as the general procedure for compound 19d. H NMR
1
(400 MHz, CDCl3) d 0.88 (t, 3H, J = 6.6 Hz), 0.95 (t,
3H, J = 7.1 Hz), 1.25 (brs, 12H), 1.30–1.37 (m, 2H),
1.37–1.53 (m, 6H), 1.79–1.87 (m, 2H), 1.88 (d, 1H,
J = 5.2 Hz), 2.20 (dt, 2H, J = 2.1 Hz, 7.2 Hz), 4.02 (t,
2H, J = 6.5 Hz), 4.98–5.03 (m, 1H), 6.93 (d, 1H,
J = 8.3 Hz), 7.01 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.08
(dd, 1H, J = 1.3 Hz, 7.6 Hz), 7.21–7.25 (m, 2H), 7.27–
7.44 (m, 6H), 7.56 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.80
(dd, 1H, J = 1.2 Hz, 7.8 Hz), 7.91 (d, 1H, J = 2.0 Hz);
5.35. Sodium 4-[(4-ethoxyphenyl)thio]-20-(1-hydroxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (20b)
The title compound (16 mg, 38% from 19b) was synthe-
sized in the same manner as the general procedure for
20d. 1H NMR (500 MHz, CD3OD) d 0.88 (t, 3H,
J = 7.8 Hz), 1.20–1.33 (m, 12H), 1.37–1.44 (m, 4H),
1.51 (quintet, 2H, J = 7.8 Hz), 2.26 (dt, 2H, J = 2.0 Hz,
6.8 Hz), 4.08 (q, 2H, J = 6.8 Hz), 5.41 (s, 1H), 6.87 (d,
1H, J = 8.8 Hz), 6.99 (d, 2H, J = 8.8 Hz), 7.38–7.49
(m, 3H), 7.53 (d, 2H, J = 8.8 Hz), 7.74 (s, 1H), 8.24 (d,
1H, J = 2.0 Hz); IR (KBr) 1197, 1243, 1459, 1494,
1594, 3412 cmꢀ1; HRMS (ESI, positive) calcd for
C33H39Na2O5S2 (M+Na)+ 625.2034; obsd 625.2050.
IR (thin film) 3536, 2926, 2855, 1593, 1489, 1458 cmꢀ1
;
HRMS (ESI, positive) calcd for C45H56O5S2Na
(M+Na)+ 763.3466; obsd 763.3456.
5.32. Phenyl 4-[(4-octyloxyphenyl)thio]-20-(1-hydroxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (19f)
The title compound was synthesized in the same manner
as the general procedure for compound 19d. H NMR
1
(400 MHz, CDCl3) d 0.89 (t, 3H, J = 6.9 Hz), 1.27 (br
s, 12H), 1.32–1.39 (m, 2H), 1.46–1.55 (m, 2H), 1.88 (d,
1H, J = 5.2 Hz), 2.22 (dt, 2H, J = 7.2 Hz, 1.9 Hz), 3.90
(s, 3H), 5.00–5.03 (m, 1H), 6.94 (d, 1H, J = 8.1 Hz),
7.04 (dd, 2H, J = 8.9 Hz, 2.1 Hz), 7.09 (dd, 1H,
J = 7.6 Hz, 1.2 Hz), 7.22–7.25 (m, 2H), 7.27–7.44 (m,
6H), 7.59 (dd, 2H, J = 2.1 Hz, 8.9 Hz), 7.82 (dd, 1H,
J = 7.8 Hz, 1.3 Hz), 7.93 (d, 1H, J = 2.0 Hz); IR (thin
film) 2925, 2854, 1591, 1493, 1458, 1387 cmꢀ1; MS
(ESI, positive) m/z: 665 (M+Na)+.
5.36. Sodium 20-(1-hydroxytridec-2-yn-1-yl)-4-[(4-
propoxyphenyl)thio]biphenyl-3-sulfonate (20c)
The title compound (35 mg, 60% from 19c) was synthe-
sized in the same manner as the general procedure for
20d. 1H NMR (400 MHz, CD3OD) d 0.89 (t, 3H,
J = 6.9 Hz), 1.06 (t, 3H, J = 7.4 Hz), 1.25 (brs, 12H),
1.31–1.39 (m, 2H), 1.40–1.50 (m, 2H), 1.77–1.88 (m,
2H), 2.19 (td, 2H, J = 1.9 Hz, 6.9 Hz), 3.99 (t, 2H,
J = 6.5 Hz), 5.29 (t, 1H, J = 1.9 Hz), 6.83 (d, 1H,
J = 8.2 Hz), 7.01 (dd, 2H, J = 2.0 Hz, 8.8 Hz), 7.19
(dd, 1H, J = 1.3 Hz, 7.5 Hz), 7.22 (dd, 1H, J = 2.0 Hz,
8.2 Hz), 7.32 (td, 1H, J = 1.3 Hz, 7.5 Hz), 7.38 (td, 1H,
J = 1.3 Hz, 7.5 Hz), 7.55 (dd, 2H, J = 2.0 Hz, 8.8 Hz),
7.79 (dd, 1H, J = 1.3 Hz, 7.5 Hz), 7.99 (d, 1H,
J = 2.0 Hz); IR (KBr) 3368, 1593, 1494, 1458,
5.33. Sodium 4-[(4-butoxyphenyl)thio]-20-(1-hydroxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (20d)
NaOH (0.12 g, 2.9 mmol) was added to a solution of com-
pound 19d (0.25 g, 0.36 mmol) in a mixed solvent of diox-
ane (1 ml) and water (0.3 ml). After stirring for 8 h at
90 ꢁC, the resulting mixture was purified by silica gel col-
umn chromatography. Elution with MeOH/CH2Cl2 (1:4)
afforded the title compound (0.19 g, 84%) as an amor-
1241 cmꢀ1
;
HRMS (ESI, negative) calcd for
C34H41O5S2 (MꢀNa)ꢀ 593.2396; obsd 593.2375.
1
phous solid. H NMR (500 MHz, CD3OD) d 0.89 (t,
5.37. Sodium 4-[(4-hexyloxyphenyl)thio]-20-(1-hydroxy-
tridec-2-yl)biphenyl-3-sulfonate (20e)
3H, J = 7.3H), 0.99 (t, 3H, J = 7.3 Hz), 1.21–1.39 (m,
14H), 1.40–1.57 (m, 4H), 1.78 (quintet, 2H, J = 6.6 Hz),
2.18 (dt, 2H, J = 1.5 Hz, 6.6 Hz), 4.02 (t, 2H,
J = 6.6 Hz), 5.29 (s, 1H), 6.82 (d, 1H, J = 8.1 Hz), 7.00
(d, 2H, J = 8.8 Hz), 7.18 (d, 1H, J = 7.3 Hz), 7.22 (dd,
1H, J = 2.2 Hz, 8.1 Hz), 7.32 (t, 1H, J = 6.6 Hz), 7.38 (t,
1H, J = 7.3 Hz), 7.54 (d, 2H, J = 8.1 Hz), 7.78 (d, 1H,
J = 8.1 Hz), 7.99 (d, 1H, J = 2.2 Hz); IR (KBr) 3386,
1593, 1494, 1458, 1243 cmꢀ1; HRMS (ESI, positive) calcd
for C35H43Na2O5S2 (M+Na)+ 653.2347; obsd 653.2345.
The title compound (42 mg, 69% from 19e) was synthe-
sized in the same manner as the general procedure for
20d. 1H NMR (400 MHz, CD3OD) d 0.89 (t, 3H,
J = 6.9 Hz), 0.96 (t, 3H, J = 7.2 Hz), 1.25 (brs, 12H),
1.29-1.38 (m, 2H), 1.38–1.53 (m, 6H), 1.76–1.85 (m,
2H), 2.19 (td, 2H, J = 1.9 Hz, 6.9 Hz), 4.02 (t, 2H,
J = 6.5 Hz), 5.29 (t, 1H, J = 1.9 Hz), 6.82 (d, 1H,
J = 8.2 Hz), 7.01 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.19

3558
T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
(dd, 1H, J = 1.4 Hz, 7.5 Hz), 7.22 (dd, 1H, J = 2.0 Hz,
8.2 Hz), 7.32 (dt, 1H, J = 1.4 Hz, 7.5 Hz), 7.38 (dt, 1H,
J = 1.4 Hz, 7.5 Hz), 7.55 (dd, 2H, J = 2.1 Hz, 8.8 Hz),
7.78 (dd, 1H, J = 1.4 Hz, 7.5 Hz), 7.89 (d, 1H,
J = 2.0 Hz); IR (KBr) 3411, 1594, 1494, 1458,
1241 cmꢀ1; HRMS (ESI, positive) calcd for C36H45O5S2-
Na2 (M+Na)+ 667.2504; obsd 667.2493.
mixture was added satd NaHCO3 (0.2 ml). The
resulting mixture was poured into water and ex-
tracted with ether. The combined organic layer was
washed with brine, dried over MgSO4, and filtered.
The filtrate was evaporated in vacuo to give the
crude product, which was purified by silica gel col-
umn chromatography. Elution with EtOAc/hexane
(2:8) afforded the title compound (38 mg, 70%) as a
colorless oil. 1H NMR (500 MHz, CDCl3) d 0.88 (t,
3H, J = 7.8 Hz), 1.00 (t, 3H, J = 7.8 Hz), 1.20–1.38
(m, 14H), 1.42–1.59 (m, 4H), 1.81 (quintet, 2H,
J = 6.8 Hz), 2.12–2.18 (m, 2H), 3.15 (s, 2H), 4.03 (t,
2H, J = 5.9 Hz), 6.93 (d, 1H, J = 8.8 Hz), 7.00–7.04
(m, 3H), 7.79 (d, 1H, J = 2.0 Hz); IR (liquid film)
1369, 1388, 1458, 1490, 1593 cmꢀ1; MS (FAB, posi-
tive) m/z: 691 (M+Na)+.
5.38. Sodium 4-[(4-octyloxyphenyl)thio]-20-(1-hydroxy-
tridec-2-yn-1-yl)biphenyl-3-sulfonate (20f)
The title compound (54 mg, 79% from 19f) was syn-
thesized in the same manner as the general procedure
1
for 20d. H NMR (400 MHz, CD3OD) d 0.89 (t, 3H,
J = 6.3 Hz), 0.90 (t, 3H, J = 6.3 Hz), 1.25 (brs, 14H),
1.32 (brs, 8H), 1.40–1.54 (m, 4H), 1.75–1.84 (m,
2H), 2.19 (td, 2H, J = 6.9 Hz, 2.0 Hz), 4.02 (t, 2H,
J = 6.4 Hz), 5.29 (t, 1H, J = 2.0 Hz), 6.83 (d, 1H,
J = 8.2 Hz), 7.00 (dd, 2H, J = 8.8 Hz, 2.1 Hz), 7.19
(dd, 1H, J = 7.5 Hz, 1.4 Hz), 7.22 (d, 1H,
J = 8.2 Hz, 2.0 Hz), 7.32 (td, 1H, J = 7.5 Hz, 1.4 Hz),
7.38 (t, 1H, J = 7.5 Hz, 1.4 Hz), 7.55 (dd, 2H,
J = 8.8 Hz, 2.1 Hz), 7.85 (dd, 1H, J = 7.5 Hz,
1.4 Hz), 7.99 (d, 1H, J = 2.0 Hz); IR (KBr) 1571,
1593, 1634, 2855, 2926, 3447 cmꢀ1; HRMS (ESI, po-
sitive) calcd for C39H51Na2O5S2 (M+Na)+ 709.2974;
obsd 709.2993.
5.41. Sodium 4-[(4-butoxyphenyl)thio]-20-tridec-2-yn-1-
ylbiphenyl-3-sulfonate (25a)
NaOH (16 mg, 0.39 mmol) was added to a solution of
compound 22a (33 mg, 0.049 mmol) in a mixed solvent
of dioxane (0.30 ml) and H2O (0.10 ml). After stirring
for 24 h at 90 ꢁC, the resulting mixture was purified by
silica gel column chromatography. Elution with
MeOH/CH2Cl2 (1:4) afforded the title compound
(10 mg, 33%) as an amorphous solid. 1H NMR
(500 MHz, CDCl3) d 0.89 (t, 3H, J = 6.8 Hz), 1.00 (t,
3H, J = 7.8 Hz), 1.20–1.39 (m, 14H), 1.43 (quintet, 2H,
J = 6.8 Hz), 1.52 (sextet, 2H, J = 7.8 Hz), 1.78 (quintet,
2H, J = 6.8 Hz), 2.10–2.16 (m, 1H), 3.38 (s, 1H), 4.02
(t, 2H, J = 6.8 Hz), 6.84 (d, 1H, J = 8.8 Hz), 7.00 (d,
2H, J = 8.8 Hz), 7.11–7.32 (m, 8H), 7.51–7.57 (m, 3H),
7.89 (d, 1H, J = 2.0 Hz); IR (KBr) 1246, 1458, 1494,
1594 cmꢀ1; MS (FAB, positive) m/z: 615 (M+H)+, 637
(M+Na)+.
5.39. Phenyl 4-[(4-butoxyphenyl)thio]-20-(1-acetoxytridec-
2-yn-1-yl)biphenyl-3-sulfonate (21)
Acetic anhydride (0.43 ml, 4.5 mmol) was slowly added
to a solution of compound 19d (1.1 g, 1.5 mmol), 4-
dimethylaminopyridine (10 mg, 0.10 mmol), and Et3N
(1.0 ml, 7.5 mmol) in CH2Cl2 (3.0 ml) at 0 ꢁC. After
stirring for 3 h, satd NaHCO3 (1.0 ml) was added to
the reaction mixture. The resulting mixture was poured
into water and extracted with ether. The combined or-
ganic layer was washed with brine, dried over MgSO4,
and filtered. The filtrate was evaporated in vacuo to
give the crude product, which was purified by silica
gel column chromatography. Elution with EtOAc/hex-
ane (1:9) afforded the title compound (1.0 g, 95%) as
a colorless oil. 1H NMR (500 MHz, CDCl3) d 0.88
(t, 3H, J = 6.8 Hz), 1.00 (t, 3H, J = 7.8 Hz), 1.20–1.33
(m, 14H), 1.45 (quintet, 2H, J = 6.8 Hz), 1.52 (sextet,
2H, J = 7.8 Hz), 1.81 (quintet, 2H, J = 7.8 Hz), 1.97
(s, 3H), 2.15 (dt, 2H, J = 2.0 Hz, 6.8 Hz), 4.03 (t, 2H,
J = 6.8 Hz), 6.93 (d, 1H, J = 8.8 Hz), 7.00 (d, 2H,
J = 8.8 Hz), 7.07 (d, 1H, J = 6.8 Hz), 7.21–7.43 (m,
6H), 7.54 (d, 2H, J = 8.8 Hz), 7.71 (d, 1H,
J = 7.8 Hz), 7.88 (d, 1H, J = 2.0 Hz); IR (liquid film)
1742, 1593, 1489, 1458 cmꢀ1; HRMS (ESI, positive)
calcd for C43H50NaO6S2 (M+Na)+ 749.2946; obsd
749.2943.
5.42. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-(phenyloxy-
carbonylamino)tridec-2-yn-1-yl]biphenyl-3-sulfonate (22b)
BF3ÆEt2O (0.11 ml, 0.83 mmol) was added to a solution
of compound 21 (0.60 g, 0.83 mmol) and phenylcarba-
mate (0.34 g, 2.5 mmol) in CH2Cl2 (1.0 ml) at 0 ꢁC.
After stirring for 1 h at room temperature, satd NaH-
CO3 (1.0 ml) was added to the reaction mixture. The
resulting mixture was poured into water and extracted
with ether. The combined organic layer was washed with
brine, dried over MgSO4, and filtered. The filtrate was
evaporated in vacuo to give the crude product, which
was purified by silica gel column chromatography. Elu-
tion with EtOAc/hexane (3:17) afforded the title com-
pound (0.60 g, 88%) as a colorless oil. 1H NMR
(500 MHz, CDCl3) d 0.88 (t, 3H, J = 6.8 Hz), 0.99 (t,
3H, J = 6.8 Hz), 1.18–1.38 (m, 14H), 1.42–1.57 (m,
4H), 1.80 (quintet, 2H, J = 6.8 Hz), 2.15 (t, 2H,
J = 5.9 Hz), 4.01 (t, 2H, J = 5.9 Hz), 5.29 (d, 1H,
J = 6.8 Hz), 5.45 (d, 1H, J = 6.8 Hz), 6.92 (d, 1H,
J = 8.8 Hz), 6.97 (d, 2H, J = 8.8 Hz), 7.00–7.07 (m,
2H), 7.14–7.44 (m, 8H), 7.51 (d, 2H, J = 8.8 Hz), 7.63
(d, 1H, J = 7.8 Hz), 7.92 (s, 1H); IR (neat) 3391, 1741,
1593, 1488, 1458 cmꢀ1; HRMS (ESI, positive) calcd
for C48H53NNaO6S2 (M+Na)+ 826.3212; obsd
826.3222.
5.40. Phenyl 4-[(4-butoxyphenyl)thio]-20-tridec-2-yn-1-ylbi-
phenyl-3-sulfonate (22a)
Triethylsilane (20 ll, 0.12 mmol) and trifluoroacetic
acid (13 ll, 0.16 mmol) were added to a solution of
compound 21 (56 mg, 0.081 mmol) in CH2Cl2
(0.5 ml) at 0 ꢁC. After stirring for 4 h, the reaction

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3559
5.43. 20-(1-Aminotridec-2-yn-1-yl)-4-[(4- butoxyphenyl)
thio]biphenyl-3-sulfonate (25b)
(1:9) afforded the title compound (0.10 g, 72%) as a col-
orless oil. H NMR (400 MHz, CDCl3) d 0.88 (t, 3H,
1
J = 6.8 Hz), 1.00 (t, 3H, J = 6.8 Hz), 1.18–1.33 (m,
18H), 1.34–1.43 (m, 2H), 1.46–1.55 (m, 4H), 1.81 (quin-
tet, 2H, J = 7.8 Hz), 2.02 (t, 2H, J = 7.0 Hz), 4.02 (t, 2H,
J = 6.6 Hz), 6.87 (d, 1H, J = 6.6 Hz), 6.90 (d, 1H,
J = 7.3 Hz), 7.00 (d, 2H, J = 8.1 Hz), 7.22–7.38 (m,
8H), 7.54 (d, 2H, J = 8.1 Hz), 7.78 (d, 1H, J = 8.1 Hz),
7.87 (d, 1H, J = 2.0 Hz).
NaOH (50 mg, 1.2 mmol) was added to a solution of
compound 22b (50 mg, 0.061 mmol) in a mixed solvent
of dioxane (0.30 ml) and H2O (0.10 ml). After stirring
for 8 h at 90 ꢁC, the resulting mixture was purified by sil-
ica gel column chromatography. Elution with MeOH/
CH2Cl2 (1:4) afforded the title compound (12 mg, 40%)
1
as an amorphous solid. H NMR (500 MHz, CD3OD)
d 0.89 (t, 3H, J = 6.8 Hz), 1.00 (t, 3H, J = 6.8 Hz),
1.21–1.38 (m, 14H), 1.42–1.57 (m, 4H), 1.78 (quintet,
2H, J = 6.8 Hz), 2.23 (dt, 2H, J = 2.0 Hz, 6.8 Hz), 4.03
(t, 2H, J = 6.8 Hz), 5.02 (d, 1H, J = 2.0 Hz), 6.86 (d,
1H, J = 8.8 Hz), 7.01 (d, 2H, J = 8.8 Hz), 7.16 (dd, 1H,
J = 2.0 Hz, 8.8 Hz), 7.28 (d, 1H, J = 7.8 Hz), 7.43 (t,
1H, J = 7.8 Hz), 7.48 (t, 1H, J = 7.8 Hz), 7.53 (d, 2H,
J = 8.8 Hz), 7.76 (d, 1H, J = 7.8 Hz), 7.93 (d, 1H,
5.46. Sodium 4-(4-ethoxyphenoxy)-20-(1-hydroxy-1-methyl
tridec-2-yn-1-yl)biphenyl-3-sulfonate (25c)
NaOH (16 mg, 0.39 mmol) was added to a solution of
compound 24 (36 mg, 0.050 mmol) in a mixed solvent
of dioxane (0.30 ml) and H2O (0.10 ml). After stirring
for 6 h at 90 ꢁC, the resulting mixture was purified by sil-
ica gel column chromatography. Elution with MeOH/
CH2Cl2 (1:4) afforded the title compound (16 mg, 47%)
J = 2.0 Hz); IR (KBr) 3691, 1594, 1494, 1458 cmꢀ1
;
HRMS (ESI, negative) calcd for C35H44NO4S2
1
as an amorphous solid. H NMR (400 MHz, CDCl3) d
(MꢀNa)ꢀ 606.2712; obsd 606.2715.
0.85 (t, 3H, J = 6.8 Hz), 0.95 (t, 3H, J = 6.8 Hz), 1.15–
1.33 (m, 18H), 1.48 (sextet, 2H, J = 6.8 Hz), 1.70–1.78
(m, 2H), 1.96 (t, 2H, J = 7.0 Hz), 3.98 (t, 2H,
J = 6.8 Hz), 6.71 (d, 1H, J = 8.1 Hz), 6.93–6.98 (m,
4H), 7.12 (dt, 1H, J = 8.2 Hz, 2.0 Hz), 7.20 (t, 1H,
J = 8.2 Hz), 7.28 (t, 1H, J = 8.2 Hz), 7.50 (d, 2H,
J = 8.8 Hz), 7.80 (d, 1H, J = 6.6 Hz), 7.85 (d, 1H,
J = 2.0 Hz).
5.44. Phenyl 4-[(4-butoxyphenyl)thio]-20-tridec-2-ynoylbi
phenyl-3-sulfonate (23)
Dess–Martin periodinane (0.20 g, 0.48 mmol) was added
to a solution of compound 19d (0.16 g, 0.24 mmol) in
CH2Cl2 at 0 ꢁC. After stirring for 2 h, satd NaHCO3
(0.2 ml) and satd Na2S2O3 (0.2 ml) were added to the
reaction mixture and stirred for 1 h at room tempera-
ture. The resulting mixture was poured into water and
extracted with ether. The combined organic layer was
washed with brine, dried over MgSO4, and filtered.
The filtrate was evaporated in vacuo to give the crude
product, which was purified by silica gel column chro-
matography. Elution with EtOAc/hexane (1:9) afforded
5.47. 2-(Undec-10-ynyloxy)tetrahydro-2H-pyrane (27)
p-TsOH (26 mg, 0.15 mmol) was added to a solution
of 10-undecyn-1-ol (0.50 g, 3.0 mmol) and 3,4-dihy-
dro-2H-pyran (0.41 ml, 4.5 mmol) in CH2Cl2 at 0 ꢁC.
After stirring for 30 min, satd NaHCO3 (1.0 ml) was
added to the reaction mixture. The resulting mixture
was poured into water and extracted with ether. The
combined organic layer was washed with brine, dried
over MgSO4, and filtered. The filtrate was evaporated
in vacuo to give the crude product, which was purified
by silica gel column chromatography. Elution with
EtOAc/hexane (1:20) afforded the title compound
1
the title compound (0.14 g, 85%) as a colorless oil. H
NMR (400 MHz, CDCl3) d 0.88 (t, 3H, J = 7.3 Hz),
1.00 (t, 3H, J = 7.3 Hz), 1.20–1.38 (m, 14H), 1.43-1.57
(m, 4H), 1.76–1.85 (m, 2H), 2.25 (t, 2H, J = 7.3 Hz),
4.02 (t, 2H, J = 6.6 Hz), 6.89 (d, 1H, J = 8.8 Hz), 6.99
(d, 2H, J = 8.8 Hz), 7.15 (d, 1H, J = 8.1 Hz), 7.21–7.29
(m, 4H), 7.33–7.39 (m, 3H), 7.44 (d, 1H, J = 1.5 Hz,
7.3 Hz), 7.46–7.54 (m, 3H), 7.82 (d, 1H, J = 2.2 Hz),
8.01 (dd, 1H, J = 1.5 Hz, 8.8 Hz); IR (liquid film)
1249, 1457, 1489, 1593, 1647, 2210 cmꢀ1; HRMS (ESI,
positive) calcd for C41H46NaO5S2 (M+Na)+ 705.2684;
obsd 705.2676.
1
(0.69 g, 92%) as a colorless oil. H NMR (400 MHz,
CDCl3) d 1.26–1.42 (brs, 10H), 1.49–1.54 (m, 4H),
1.55–1.61 (m, 4H), 1.69–1.75 (m, 1H), 1.80–1.86 (m,
1H), 1.94 (d, 1H, J = 2.6 Hz), 2.18 (dt, 2H,
J = 2.9 Hz, 10.3 Hz), 3.35–3.41 (m, 1H), 3.48–3.53
(m, 1H), 3.70–3.76 (m, 1H), 3.85–3.90 (m, 1H), 4.58
(t, 1H, J = 2.9 Hz); IR (liquid film) 3312, 2933, 2857,
1033 cmꢀ1; HRMS (EI, positive) calcd for C16H28O2
(MꢀH)+ 251.2011; obsd 251.2017.
5.45. Phenyl 4-(4-ethoxyphenoxy)-20-(1-hydroxy-1-meth-
yltridec-2-yn-1-yl)biphenyl-3-sulfonate (24)
2.0 M solution of methylmagnesiumbromide in ether
(0.21 ml, 0.42 mmol) was added to a solution of com-
pound 23 (0.14 g, 0.21 mmol) in THF (1 ml) at
ꢀ78 ꢁC. After stirring for 2 h, satd NH4Cl (0.2 ml) was
added to the reaction mixture and stirred for 1 h at
room temperature. The resulting mixture was poured
into water and extracted with ether. The combined or-
ganic layer was washed with brine, dried over MgSO4,
and filtered. The filtrate was evaporated in vacuo to give
the crude product, which was purified by silica gel col-
umn chromatography. Elution with EtOAc/hexane
5.48. 2-[[3-(Prop-2-ynyloxy)propyl]oxy]tetrahydro-2H-
pyran (29a)
55 wt% of NaH (84 mg, 4.0 mmol) was added to a
solution of propargylalcohol (61 ml, 1.1 mmol) and
2-[(3-bromopropyl)oxy]tetrahydro-2H-pyran (0.18 g,
0.70 mmol) in DMF at 0 ꢁC. After stirring for 3 h, satd
NH4Cl (1.0 ml) was added to the reaction mixture. The
resulting mixture was poured into water and extracted
with ether. The combined organic layer was washed
with brine, dried over MgSO4, and filtered. The filtrate

3560
T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
was evaporated in vacuo to give the crude product,
which was purified by silica gel column chromatogra-
phy. Elution with EtOAc/hexane (1:20) afforded the ti-
tle compound (0.10 g, 62%) as a colorless oil. 1H
NMR (400 MHz, CDCl3) d 1.46–1.63 (m, 4H), 1.65–
1.75 (m, 1H), 1.77–1.94 (m, 3H), 2.41 (t, 1H,
J = 2.4 Hz), 3.45–3.54 (m, 2H), 3.58–3.67 (m, 2H),
3.89–3.90 (m, 2H), 4.14 (d, 2H, J = 2.4 Hz), 4.59 (dd,
1H, J = 4.4 Hz, 2.8 Hz); IR (thin film) 3291, 3261,
2944, 2870, 2115, 1442 cmꢀ1; MS (EI, positive) m/z:
197 (MꢀH)+.
5.53. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-12-
(tetrahydro-2H-pyran-2- yloxy)dodec-2-yn-1-yl]biphenyl -
3-sulfonate (30)
1.6
M Solution of n-BuLi in hexane (0.23 ml,
0.36 mmol) was added to a solution of compound 27
(0.10 g, 0.40 mmol) in THF (1.0 ml) at ꢀ78 ꢁC. After
stirring for 10 min, a solution of aldehyde 18d (0.10 g,
0.40 mmol) in THF (1.0 ml) was added to the reaction
mixture via cannula. After stirring for 1 h, satd NH4Cl
(1.0 ml) was added to the reaction mixture. The resulting
mixture was poured into water and extracted with ether.
The combined organic layer was washed with brine,
dried over MgSO4, and filtered. The filtrate was evapo-
rated in vacuo to give the crude product, which was
purified by silica gel column chromatography. Elution
with EtOAc/hexane (1:3) afforded the title compound
5.49. 2-[[5-(Prop-2-ynyloxy)pentyl]oxy]tetrahydro-2H-pyran
(29b)
The title compound was synthesized in the same manner
as the general procedure for 29a. H NMR (400 MHz,
1
1
CDCl3) d 1.40–1.76 (m, 11H), 1.78–1.87 (m, 1H), 2.41
(t, 1H, J = 2.4 Hz), 3.39 (td, 1H, J = 6.6 Hz, 9.6 Hz),
3.47–3.53 (m, 1H), 3.52 (t, 2H, J = 6.6 Hz), 4.13 (d,
2H, J = 2.4 Hz), 4.57 (dd, 1H, J = 2.8 Hz, 4.4 Hz); IR
(thin film) 3261, 2941, 2867, 1354 cmꢀ1; MS (ESI, posi-
tive) m/z: 249 (M+Na)+.
(0.10 g, 66%) as a pale yellow oil. H NMR (400 MHz,
CDCl3) d 1.00 (t, 3H, J = 7.3 Hz), 1.24–1.36 (brs,
10H), 1.45–1.60 (m, 10H), 1.68-1.74 (m, 1H), 1.78–1.85
(m, 3H), 1.99 (d, 1H, J = 5.1 Hz), 2.20 (dt, 2H,
J = 2.2 Hz, 9.5 Hz), 3.35–3.40 (m, 1H), 3.47–3.52 (m,
1H), 3.69–3.75 (m, 1H), 3.84–3.89 (m, 2H), 4.03 (t,
2H, J = 6.6 Hz), 4.56–4.58 (m, 1H), 5.01 (t, 1H,
J = 2.2 Hz), 6.93 (d, 1H, J = 8.8 Hz), 7.01 (d, 2H,
J = 8.8 Hz), 7.07 (d, 1H, J = 6.6 Hz), 7.22 (d, 2H,
J = 8.8 Hz), 7.27–7.43 (m, 6H), 7.55 (d, 2H,
J = 8.8 Hz), 7.80 (d, 1H, J = 7.3 Hz), 7.91 (d, 1H,
5.50. 2-[[7-(Prop-2-ynyloxy)heptyl]oxy]tetrahydro-2H-pyran
(29c)
The title compound was synthesized in the same manner
as the general procedure for 29a. H NMR (400 MHz,
1
J = 1.5 Hz); IR (thin film) 2933, 2857, 1249 cmꢀ1
;
CDCl3) d 1.32–1.42 (m, 6H), 1.48–1.65 (m, 8H), 1.68–
1.75 (m, 1H), 1.78–1.87 (m, 1H), 2.41 (t, 1H,
J = 2.4 Hz), 3.38 (td, 1H, J = 6.6 Hz, 9.6 Hz), 3.47–
3.53 (m, 1H), 3.51 (t, 2H, J = 6.6 Hz), 3.73 (td, 1H,
J = 6.9 Hz, 9.6 Hz), 3.74–3.89 (m, 1H), 4.13 (d, 2H,
J = 2.4 Hz), 4.57 (dd, 1H, J = 3.0 Hz, 4.1 Hz).
HRMS (ESI, positive) calcd for C45H54O7S2 (M+Na)+
793.3208; obsd 793.3193.
5.54. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-[[5-
(tetrahydro-2H-pyran-2-yloxy)propyl]oxy]but-2-ynyl]-
1,10-biphenyl-3-sulfonate (31a)
5.51. 2-[[9-(Prop-2-ynyloxy)nonyl]oxy]tetrahydro-2H-pyran
(29d)
The title compound was synthesized in the same manner
as the general procedure for 30. H NMR (400 MHz,
1
CDCl3) d 1.00 (t, 3H, J = 7.4 Hz), 1.45–1.58 (m, 6H),
1.64–1.73 (m, 1H), 1.75–1.92 (m, 5H), 2.22 (d, 1H,
J = 5.2 Hz), 3.44-3.52 (m, 2H), 3.60 (t, 2H,
J = 6.3 Hz), 3.29–3.87 (m, 2H), 4.03 (t, 2H,
J = 6.5 Hz), 4.17 (s, 2H), 4.56 (brs, 1H), 5.06 (d, 1H,
J = 5.2 Hz), 6.94 (d, 1H, J = 8.3 Hz), 7.02 (dd, 2H,
J = 8.8 Hz, 2.1 Hz), 7.09 (dd, 1H, J = 7.7 Hz, 1.3 Hz),
7.21–7.44 (m, 8H), 7.56 (dd, 2H, J = 8.8 Hz, 2.1 Hz),
7.80 (d, 1H, J = 7.7 Hz), 7.90 (d, 1H, J = 1.9 Hz); IR
(thin film) 3390, 2940, 1872, 1592, 1489, 1457,
1387 cmꢀ1; MS (FAB) m/z: 716 (M)+.
The title compound was synthesized in the same manner
as the general procedure for 29a. H NMR (400 MHz,
1
CDCl3) d 1.30 (brs, 10H), 1.48–1.63 (m, 8H), 1.67–
1.76 (m, 1H), 1.79–1.88 (m, 1H), 2.41 (t, 1H,
J = 2.4 Hz), 3.38 (td, 1H, J = 6.7 Hz, 9.6 Hz), 3.46–
3.53 (m, 1H), 3.51 (t, 2H, J = 6.6 Hz), 3.73 (td, 1H,
J = 6.9 Hz, 9.6 Hz), 3.84–3.90 (m, 1H), 4.13 (d, 2H,
J = 2.4 Hz), 4.57 (dd, 1H, J = 2.8 Hz, 4.4 Hz); IR (liquid
film) 3310, 2934, 2856, 1354 cmꢀ1; MS (EI, positive)
m/z: 281 (MꢀH)+.
5.52. 2-[[11-(Prop-2-ynyloxy)undecyl]oxy]tetrahydro-2H-
pyran (29e)
5.55. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-[[5-
(tetrahydro-2H-pyran-2-yloxy)pentyl]oxy]but-2-ynyl]-
1,10-biphenyl-3-sulfonate (31b)
The title compound was synthesized in the same man-
ner as the general procedure for 29a. 1H NMR
(400 MHz, CDCl3) d 1.27 (br s, 14H), 1.48–1.63 (m,
8H), 1.67–1.75 (m, 1H), 1.78–1.88 (m, 1H), 2.41 (t,
1H, J = 2.4 Hz), 3.38 (td, 1H, J = 6.7 Hz, 9.6 Hz),
3.46–3.53 (m, 1H), 3.51 (t, 2H, J = 6.7 Hz), 3.73 (td,
1H, J = 6.9 Hz, 9.6 Hz), 3.84–3.91 (m, 1H), 4.13 (d,
2H, J = 2.4 Hz), 4.58 (dd, 1H, J = 2.8 Hz, 4.3 Hz);
IR (liquid film) 3311, 2929, 1354 cmꢀ1; MS (ESI, po-
sitive) m/z: 309 (MꢀH)+.
The title compound was synthesized in the same manner
as the general procedure for 30. H NMR (400 MHz,
1
CDCl3) d 1.00 (t, 3H, J = 7.4 Hz), 1.38–1.73 (m, 14H),
1.77–1.86 (m, 2H), 2.14 (dd, 1H, J = 1.5 Hz, 5.2 Hz),
3.39 (td, 1H, J = 6.6 Hz, 9.7 Hz), 3.49 (t, 2H,
J = 6.5 Hz), 3.45–3.52 (m, 1H), 3.73 (td, 1H,
J = 6.8 Hz, 9.7 Hz), 3.82–3.88 (m, 1H), 4.03 (t, 2H,
J = 6.5 Hz), 4.16 (d, 2H, J = 1.6 Hz), 4.55–4.58 (m,

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3561
1H), 5.07 (td, 1H, J = 1.6 Hz, 5.2 Hz), 6.94 (d, 1H,
J = 8.3 Hz), 7.02 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.09
(dd, 1H, J = 1.3 Hz, 7.7 Hz), 7.21–7.44 (m, 8H), 7.56
(dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.79 (dd, 1H, J = 1.3 Hz,
7.7 Hz), 7.90 (d, 1H, J = 2.0 Hz); IR (thin film) 3390,
1593, 1489, 1458, 1387, 1368, 1249 cmꢀ1; HRMS (ESI,
positive) calcd for C42H48O8S2Na (M+Na)+ 767.2688;
obsd 767.2690.
(m, 1H), 3.72 (td, 1H, J = 6.9 Hz, 9.6 Hz), 3.83–3.90
(m, 1H), 4.03 (t, 2H, J = 6.5 Hz), 4.16 (d, 2H,
J = 1.6 Hz), 4.57 (t, 1H, J = 3.5 Hz), 5.06 (td, 1H,
J = 1.6 Hz, 5.1 Hz), 6.94 (d, 1H, J = 8.3 Hz), 7.02 (dd,
2H, J = 2.0 Hz, 8.7 Hz), 7.09 (dd, 1H, J = 1.3 Hz,
7.6 Hz), 7.21–7.44 (m, 8H), 7.56 (dd, 2H, J = 2.0 Hz,
8.7 Hz), 7.79 (dd, 1H, J = 1.2 Hz, 7.7 Hz), 7.91 (d, 1H,
J = 2.0 Hz); IR (thin film) 3389, 1593, 1489, 1388,
1368, 1249 cmꢀ1; HRMS (ESI, positive) calcd for
C48H60O8S2Na (M+Na)+ 851.3627; obsd 851.3649.
5.56. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-[[7-
(tetrahydro-2H-pyran-2-yloxy)heptyl]oxy]but-2-ynyl]-1,10-
biphenyl-3-sulfonate (31c)
5.59. Phenyl 4-[(4-butoxyphenyl)thio]-20-(1,12-dihydroxy-
dodec-2-yn-1-yl)biphenyl-3-sulfonate (32)
The title compound was synthesized in the same man-
ner as the general procedure for 30. 1H NMR
(400 MHz, CDCl3) d 1.00 (t, 3H, J = 7.4 Hz), 1.30–
1.39 (m, 6H), 1.48–1.52 (m, 10H), 1.66–1.74 (m, 1H),
1.77–1.86 (m, 3H), 2.04 (dd, 1H, J = 1.5 Hz, 5.2 Hz),
3.39 (td, 1H, J = 6.6 Hz, 9.5 Hz), 3.46 (t, 2H,
J = 6.6 Hz), 3.46–3.53 (m, 1H), 3.72 (td, 1H,
J = 6.9 Hz, 9.5 Hz), 3.82–3.88 (m, 1H), 4.03 (t, 2H,
J = 6.5 Hz), 4.16 (d, 2H, J = 1.7 Hz), 4.55–4.58 (m,
1H), 5.07 (td, 1H, J = 1.7 Hz, 5.2 Hz), 6.94 (d, 1H,
J = 8.3 Hz), 7.02 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.09
(dd, 1H, J = 1.3 Hz, 7.7 Hz), 7.21–7.44 (m, 8H), 7.56
(dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.79 (dd, 1H,
J = 1.3 Hz, 7.7 Hz), 7.90 (d, 1H, J = 2.0 Hz); IR (thin
p-TsOH (10 mg, 0.01 mmol) was added to a solution
of compound 30 (0.10 g, 0.13 mmol) in MeOH
(1.0 ml) at room temperature. After stirring for 1 h,
Et3N (1.0 ml) was added to the reaction mixture.
The resulting mixture was poured into water and ex-
tracted with ether. The combined organic layer was
washed with brine, dried over MgSO4, and filtered.
The filtrate was evaporated in vacuo to give the crude
product, which was purified by silica gel column chro-
matography. Elution with EtOAc/hexane (1:1) affor-
ded the title compound (80 mg, 87%) as a colorless
oil. 1H NMR (400 MHz, CDCl3) d 1.00 (t, 3H,
J = 7.3 Hz), 1.23–1.38 (brs, 10H), 1.45-1.57 (m, 6H),
1.74–1.84 (m, 2H), 2.20 (dt, 2H, J = 2.0 Hz, 9.5 Hz),
3.61 (t, 2H, J = 6.6 Hz), 4.03 (t, 2H, J = 6.6 Hz),
5.01 (s, 1H), 6.93 (d, 1H, J = 8.1 Hz), 7.01 (d, 2H,
J = 8.8 Hz), 7.06 (d, 1H, J = 6.6 Hz), 7.22 (d, 2H,
J = 8.1 Hz), 7.27–7.42 (m, 8H), 7.55 (d, 2H,
J = 8.8 Hz), 7.80 (d, 1 H, J = 8.8 Hz), 7.91 (d, 1H,
film) 3390, 1593, 1489, 1458, 1388, 1368, 1249 cmꢀ1
;
HRMS (ESI, positive) calcd for C44H52O8S2Na
(M+Na)+ 795.3002; obsd 795.3019.
5.57. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-[[9-
(tetrahydro-2H-pyran-2-yloxy)nonyl]oxy]but-2-ynyl]-1,10-
biphenyl-3-sulfonate (31d)
J = 2.2 Hz); IR (thin film) 3388, 2931, 1242 cmꢀ1
;
HRMS (ESI, positive) calcd for C40H46O6S2
The title compound was synthesized in the same man-
ner as the general procedure for 30. 1H NMR
(400 MHz, CDCl3) d 1.00 (t, 3H, J = 7.4 Hz), 1.24–
1.37 (m, 10H), 1.47–1.52 (m, 10H), 1.67–1.75 (m,
1H), 1.77–1.86 (m, 3H), 1.99 (dd, 1H, J = 1.8 Hz,
5.2 Hz), 3.37 (td, 1H, J = 6.7 Hz, 9.6 Hz), 3.46 (t,
2H, J = 6.6 Hz), 3.47–3.53 (m, 1H), 3.72 (td, 1H,
J = 6.9 Hz, 9.6 Hz), 3.83–3.90 (m, 1H), 4.03 (t, 2H,
J = 6.5 Hz), 4.16 (d, 2H, J = 1.7 Hz), 4.57 (t, 1H,
J = 3.6 Hz), 5.07 (dt, 1H, J = 1.7 Hz, 5.2 Hz), 6.94 (d,
1H, J = 8.3 Hz), 7.02 (dd, 2H, J = 2.1 Hz, 8.8 Hz),
7.09 (dd, 1H, J = 1.3 Hz, 7.6 Hz), 7.21–7.44 (m, 8H),
7.56 (dd, 2H, J = 2.1 Hz, 8.8 Hz), 7.79 (dd, 1H,
J = 1.2 Hz, 7.8 Hz), 7.90 (d, 1H, J = 2.0 Hz); IR (thin
(M+Na)+ 709.2624; obsd 709.2612.
5.60. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-[(3-
hydroxypropyl)oxy]but-2-ynyl]-1,10-biphenyl-3-sulfonate
(33a)
The title compound was synthesized in the same manner
as the general procedure for 32. H NMR (400 MHz,
1
CDCl3) d 1.01 (t, 3H, J = 7.4 Hz), 1.49–1.58 (m, 2H),
1.72–1.83 (m, 4H), 3.55 (t, 2H, J = 6.3 Hz), 3.60 (t, 2H,
J = 6.4 Hz), 4.06 (t, 2H, J = 6.4 Hz), 4.14 (d, 2H,
J = 1.4 Hz), 4.98 (s, 1H), 6.97 (d, 1H, J = 8.3 Hz), 7.06
(d, 1H, J = 7.6 Hz), 7.09 (d, 2H, J = 8.7 Hz), 7.20 (d, 2H,
J = 7.9 Hz), 7.32 (t, 2H, J = 7.6 Hz), 7.37–7.45 (m, 4H),
7.56 (d, 2H, J = 8.7 Hz), 7.76 (d, 1H, J = 7.6 Hz), 7.81
(d, 1H, J = 1.8 Hz); IR (thin film) 3385, 2958, 2934,
2873, 1593, 1489 cmꢀ1;MS(FAB, positive)m/z:632(M)+.
film) 3397, 1593, 1489, 1458, 1388, 1368, 1249 cmꢀ1
;
HRMS (ESI, positive) calcd for C46H56O8S2Na
(M+Na)+ 823.3315; obsd 823.3311.
5.58. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-[[11-
(tetrahydro-2H-pyran-2-yloxy)undecyl]oxy]but-2-ynyl]-1,10-
biphenyl-3-sulfonate (31e)
5.61. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-[(5-
hydroxypentyl)oxy]but-2-ynyl]-1,10-biphenyl-3-sulfonate
(33b)
The title compound was synthesized in the same manner
as the general procedure for 30. H NMR (400 MHz,
The title compound was synthesized in the same manner
as the general procedure for 32. H NMR (400 MHz,
1
1
CDCl3) d 1.00 (t, 3H, J = 7.4 Hz), 1.23–1.38 (m, 14H),
1.48–1.63 (m, 10H), 1.67–1.75 (m, 1H), 1.78–1.86 (m,
1H), 2.00 (dd, 1H, J = 1.2 Hz, 5.1 Hz), 3.38 (td, 1H,
J = 6.7 Hz, 9.6 Hz), 3.46 (t, 2H, J = 6.7 Hz), 3.47–3.53
CD3OD) d 1.01 (t, 3H, J = 7.4 Hz), 1.34–1.41 (m, 2H),
1.47–1.58 (m, 6H), 1.77–1.83 (m, 2H), 3.46 (t, 2H,
J = 6.5 Hz), 3.51 (t, 2H, J = 6.6 Hz), 4.06 (t, 2H,
J = 6.4 Hz), 4.13 (d, 2H, J = 1.6 Hz), 4.99 (brs, 1H),

3562
T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
6.97 (d, 1H, J = 8.3 Hz), 7.06 (d, 1H, J = 7.7 Hz), 7.09
(d, 2H, J = 8.8 Hz), 7.20 (d, 2H, J = 8.1 Hz), 7.30–7.34
(m, 2H), 7.37–7.42 (m, 3H), 7.43 (dd, 1H, J = 1.9 Hz,
8.3 Hz), 7.56 (d, 2H, J = 8.8 Hz), 7.75 (d, 1H,
J = 7.7 Hz), 7.82 (d, 1H, J = 1.9 Hz); IR (thin film)
3383, 1593, 1489, 1458, 1387, 1366, 1249 cmꢀ1; HRMS
(ESI, positive) calcd for C37H40O7S2Na (M+Na)+
683.2113; obsd 683.2130.
3376, 1593, 1489, 1458, 1387, 1367, 1249 cmꢀ1; HRMS
(FAB, positive) calcd for C43H52O7S2Na (M+Na)+
767.3052; obsd 767.3029.
5.65. Sodium 4-[(4-butoxyphenyl)thio]-20-(1,12-dihydr-
oxydodec-2-yn-1-yl)biphenyl-3-sulfonate (34)
NaOH (0.10 g, 2.4 mmol) was added to a solution of
compound 32 (80 mg, 0.12 mmol) in a mixed solvent
of dioxane (1 ml) and water (0.3 ml). After stirring
for 8 h at 90 ꢁC, the resulting mixture was purified by
silica gel column chromatography. Elution with
MeOH/CH2Cl2 (1:5) afforded the title compound
(30 mg, 39%) as an amorphous solid. 1H NMR
(400 MHz, CD3OD) d 1.00 (t, 3H, J = 7.3 Hz), 1.27–
1.42 (brs, 10H), 1.45 (t, 2H, J = 7.3 Hz), 1.50-1.56
(m, 4H), 1.75–1.91 (m, 2H), 2.17–2.21 (m, 2H), 3.52
(t, 2H, J = 6.6 Hz), 4.03 (t, 2H, J = 6.6 Hz), 5.28 (t,
1H, J = 2.2 Hz), 6.82 (d, 1H, J = 8.1 Hz), 7.01 (d, 2H,
J = 8.8 Hz), 7.18-7.24 (m, 2H), 7.32 (dd, 1H,
J = 1.5 Hz, 8.8 Hz), 7.38 (dt, 2H, J = 1.5 Hz, 7.3 Hz),
7.55 (dd, 2H, J = 2.2 Hz, 6.6 Hz), 7.78 (dd, 1H,
J = 1.5 Hz, 8.1 Hz), 7.98 (d, 1H, J = 2.2 Hz); IR
(KBr) 3398, 2929, 1242 cmꢀ1; HRMS (ESI, negative)
calcd for C34H41NaO6S2 (MꢀNa)ꢀ 609.2345; obsd
609.2351.
5.62. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
[(7-hydroxyheptyl)oxy]but-2-ynyl]-1,10-biphenyl-3-sulfo-
nate (33c)
The title compound was synthesized in the same
manner as the general procedure for 32. 1H NMR
(400 MHz, CD3OD)
d 1.01 (t, 3H, J = 7.4 Hz),
1.28–1.37 (m, 6H), 1.47–1.58 (m, 6H), 1.77–1.83 (m,
2H), 3.44 (t, 2H, J = 6.5 Hz), 3.52 (t, 2H,
J = 6.7 Hz), 4.06 (t, 2H, J = 6.4 Hz), 4.12 (d, 2H,
J = 1.6 Hz), 4.99 (brs, 1H), 6.97 (d, 1H, J = 8.3 Hz),
7.06 (d, 1H, J = 7.7 Hz), 7.09 (d, 2H, J = 8.7 Hz),
7.20 (d, 2H, J = 8.0 Hz), 7.30–7.35 (m, 2H), 7.37–
7.42 (m, 3H), 7.43 (dd, 1H, J = 1.9 Hz, 8.3 Hz),
7.55 (d, 2H, J = 8.7 Hz), 7.75 (d, 1H, J = 7.7 Hz),
7.81 (d, 1H, J = 1.9 Hz); IR (thin film) 3386, 1593,
1489, 1458, 1387, 1366, 1249 cmꢀ1; HRMS (ESI, po-
sitive) calcd for C39H44O7S2Na (M+Na)+ 711.2426;
obsd 711.2427.
5.66. Sodium 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
(3-hydroxypropoxy)but-2-yn-1-yl]biphenyl-3-sulfonate
(35a)
5.63. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
[(9-hydroxynonyl)oxy]but-2-ynyl]-1,10-biphenyl-3-sulfo-
nate (33d)
The title compound (22 mg, 40% from 33a) was synthe-
sized in the same manner as the general procedure for
1
The title compound was synthesized in the same manner
as the general procedure for 32. H NMR (400 MHz,
compound 34. H NMR (400 MHz, CD3OD) d 1.04 (t,
1
3H, J = 7.4 Hz), 1.53–1.61 (m, 2H), 1.75–1.86 (m, 4H),
3.60 (t, 2H, J = 6.3 Hz), 3.63 (t, 2H, J = 6.4 Hz), 4.07
(t, 2H, J = 6.4 Hz), 4.20 (d, 2H, J = 1.5 Hz), 5.41 (brs,
1H), 6.87 (d, 1H, J = 8.2 Hz), 7.05 (d, 2H, J = 8.7 Hz),
7.23–7.27 (m, 2H), 7.38 (t, 1H, J = 7.5), 7.44 (t, 1H,
J = 7.5 Hz), 7.59 (d, 2H, J = 8.7 Hz), 7.83 (d, 1H,
J = 7.5 Hz), 8.03 (d, 1H, J = 1.9 Hz); IR (KBr) 11593,
2873, 2931, 2957, 3399 cmꢀ1; HRMS (ESI, positive)
calcd for C29H32NaO7S2 (M+H)+ 579.1487; obsd
579.1484.
CD3OD) d 1.01 (t, 3H, J = 7.4 Hz), 1.23–1.37 (m,
10H), 1.48–1.58 (m, 6H), 1.77–1.83 (m, 2H), 3.44 (t,
2H, J = 6.5 Hz), 3.53 (t, 2H, J = 6.6 Hz), 4.06 (t, 2H,
J = 6.4 Hz), 4.12 (d, 2H, J = 1.6 Hz), 4.99 (brs, 1H),
6.97 (d, 1H, J = 8.3 Hz), 7.06 (d, 1H, J = 7.7 Hz), 7.09
(d, 2H, J = 8.7 Hz), 7.20 (d, 2H, J = 8.2 Hz), 7.32 (t,
2H, J = 7.1 Hz), 7.37–7.41 (m, 3H), 7.43 (dd, 1H,
J = 1.9 Hz, 8.3 Hz), 7.56 (d, 2H, J = 8.7 Hz), 7.75 (d,
1H, J = 7.7 Hz), 7.81 (d, 1H, J = 1.9 Hz); IR (thin film)
3383, 1593, 1489, 1458, 1387, 1366, 1249 cmꢀ1; HRMS
(ESI, positive) calcd for C41H48O7S2Na (M+Na)+
739.2740; obsd 739.2739.
5.67. Sodium 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
[(5-hydroxypentyl)oxy]but-2-yn-1-yl]biphenyl-3-sulfonate
(35b)
5.64. Phenyl 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
[(11-hydroxyundecyl)oxy]but-2-ynyl]-1,10-biphenyl-3-sul-
fonate (33e)
The title compound (24 mg, 33% from 33b) was synthe-
sized in the same manner as the general procedure for
1
compound 34. H NMR (400 MHz, CD3OD) d 1.00 (t,
The title compound was synthesized in the same manner
as the general procedure for 32. H NMR (400 MHz,
3H, J = 7.4 Hz), 1.33–1.40 (m, 2H), 1.48–1.58 (m, 6H),
1.75–1.83 (m, 2 H), 3.47 (t, 2H, J = 6.5 Hz), 3.52 (t,
2H, J = 6.6 Hz), 4.03 (t, 2H, J = 6.4 Hz), 4.15 (d, 2H,
J = 1.6 Hz), 5.37 (brs, 1H), 6.84 (d, 1H, J = 8.2 Hz),
7.01 (d, 2H, J = 8.7 Hz), 7.19–7.23 (m, 2H), 7.34 (dt,
1H, J = 1.2 Hz, 7.5 Hz), 7.40 (dt, 1H, J = 1.2 Hz,
7.5 Hz), 7.55 (d, 2H, J = 8.7 Hz), 7.78 (d, 1H,
J = 7.5 Hz), 7.99 (d, 1H, J = 1.9 Hz); IR (KBr) 3399,
1593, 1494, 1458, 1243 cmꢀ1; HRMS (ESI, negative)
calcd for C31H35O7S2 (MꢀNa)ꢀ 583.1824; obsd
583.1814.
1
CD3OD) d 1.01 (t, 3H, J = 7.4 Hz), 1.21–1.38 (m,
14H), 1.48–1.58 (m, 6H), 1.77–1.84 (m, 2H), 3.44 (t,
2H, J = 6.5 Hz), 3.53 (t, 2H, J = 6.7 Hz), 4.06 (t, 2H,
J = 6.4 Hz), 4.12 (d, 2H, J = 1.5 Hz), 4.99 (brs, 1H),
6.97 (d, 1H, J = 8.3 Hz), 7.06 (d, 1H, J = 7.4 Hz), 7.09
(d, 2H, J = 8.7 Hz), 7.20 (d, 2H, J = 7.8 Hz), 7.32 (t,
2H, J = 7.4 Hz), 7.37–7.42 (m, 3H), 7.43 (dd, 1H,
J = 1.9 Hz, 8.3 Hz), 7.56 (d, 2H, J = 8.7 Hz), 7.75 (d,
1H, J = 7.4 Hz), 7.81 (d, 1H, J = 1.9 Hz); IR (thin film)

T. Nakamura et al. / Bioorg. Med. Chem. 15 (2007) 3548–3564
3563
5.68. Sodium 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
[(5-hydroxyheptyl)oxy]but-2-yn-1-yl]biphenyl-3-sulfonate
(35c)
penicillin G sodium, and 100 lg/mL streptomycin sul-
fate (Invitrogen)) with 125 nM methotrexate. CHO–
S1P1 cells were seeded into a 96-well plate (4 · 105
cells/well) with the selection medium and cultured
overnight. The next day, FBS was removed from the
medium, and the cells were preincubated with 1 mM
3-isobutyl-1-methylxanthine (IBMX) for 5 min and
then stimulated by 30 lM forskolin with or without
100 nM S1P and various amounts of test compounds
for 15 min in the presence of 1 mM IBMX. After
the reaction, the cells were dissolved using 1% Triton
X-100 and the cyclic adenosine 30,50-monophosphate
(cAMP) concentration was measured in a competitive
assay using a cAMP femtomolar kit (CIS Bio Interna-
tional), following the instructions of the manufacturer
(n = 3).
The title compound (28 mg, 40% from 33c) was synthe-
sized in the same manner as the general procedure for
1
compound 34. H NMR (400 MHz, CD3OD) d 1.00 (t,
3H, J = 7.4 Hz), 1.31 (brs, 6H), 1.47–1.57 (m, 6H),
1.75–1.82 (m, 2H), 3.46 (t, 2H, J = 6.5 Hz), 3.52 (t,
2H, J = 6.7 Hz), 4.03 (t, 2H, J = 6.4 Hz), 4.15 (d, 2H,
J = 1.6 Hz), 5.37 (brs, 1H), 6.83 (d, 1H, J = 8.2 Hz),
7.01 (d, 2H, J = 8.7 Hz), 7.19-7.23 (m, 2H), 7.34 (dt,
1H, J = 1.2 Hz, 7.5 Hz), 7.39 (t, 1H, J = 7.5 Hz), 7.55
(d, 2H, J = 8.7 Hz), 7.79 (d, 1H, J = 7.5 Hz), 7.99 (d,
1H, J = 2.0 Hz); IR (thin film) 3399, 1593, 1493, 1458,
1243 cmꢀ1
;
HRMS (ESI, negative) calcd for
C33H39O7S2 (MꢀNa)ꢀ 611.2137; obsd 611.2126.
5.72. Selectivity profiling among S1P receptors
5.69. Sodium 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
[(5-hydroxynonyl)oxy]but-2-yn-1-yl]biphenyl-3-sulfonate
(35d)
For the measurement of human S1P1 and S1P4 recep-
tor signaling, Gqi5 chimeric G proteins were co-ex-
pressed with each receptor in CHO cells. Both cell
lines and CHO cells stably expressing human S1P2
receptors were established in our laboratory. Fluo3-
AM and pluronic acid (Pluronic F-127 low absor-
bance) were purchased from Molecular Probes. Each
receptor-expressing cell was suspended in HAM med-
ium (Invitrogen) containing 10% dialyzed FBS,
100 U/mL penicillin G sodium, and 100 lg/mL strepto-
mycin sulfate (5 · 104 cells/mL for S1P1, S1P2, and
S1P3, 1 · 105 cells/mL for S1P4), and seeded into a
384-well plate (Corning 3712) with 50 lL/well and cul-
tured for 2 days. The plate was washed using an
ELx405 washer (BIO-TEK INSTRUMENTS) with
reaction buffer (Hanks’ balanced salt solution without
phenol red (Invitrogen), 20 mM Hepes, and 2.5 mM
probenecid, pH 7.4) and 25 lL of the buffer being left
in each well. Twenty-five microliters of another reac-
tion buffer, containing 4 lM Fluo3-AM and 0.04%
(w/v) pluronic acid, was poured into each well and
the plate was cultured for 45–60 min, and then washed
again in the same way, leaving 15 lL of the buffer. Se-
rial-diluted test compounds were dissolved into the
reaction buffer, a 3-fold concentration of the final,
and 15 lL of each one was added into the plate. After
10 min of culturing, the plate was set on a FLIPR384
(Molecular Devices) and 15 lL of reaction buffer
containing S1P was injected into each well. The inten-
sity of the S1P receptor signaling and inhibitory effects
of each of the test compounds were simultaneously
measured using the FLIPR384. The final concentra-
tions of S1P were modified for each receptor, at
100 nM, 100 nM, 10 nM, and 10 lM for S1P1–S1P4,
respectively.
The title compound (24 mg, 33% from 33d) was synthe-
sized in the same manner as the general procedure for
compound 34. H NMR (400 MHz, CD3OD) d 1.04 (t,
1
3H, J = 7.4 Hz), 1.32 (brs, 10H), 1.51–1.61 (m, 6H),
1.79–1.86 (m, 2H), 3.49 (t, 2H, J = 6.5 Hz), 3.57 (t,
2H, J = 6.7 Hz), 4.07 (t, 2H, J = 6.4 Hz), 4.19 (d, 2H,
J = 1.4 Hz), 4.92 (brs, 1H), 6.87 (d, 1H, J = 8.2 Hz),
7.05 (d, 2H, J = 8.7 Hz), 7.23–7.28 (m, 2H), 7.38 (d,
1H, J = 7.5 Hz), 7.43 (t, 1H, J = 7.5 Hz), 7.59 (d, 2H,
J = 8.7 Hz), 7.83 (d, 1H, J = 7.5 Hz), 8.03 (d, 1H,
J = 1.8 Hz); IR (thin film) 3376, 1593, 1489, 1458,
1387,1367, 1249 cmꢀ1; HRMS (ESI, negative) calcd for
C35H43O7S2 (MꢀNa)ꢀ 639.2452; obsd 639.2416.
5.70. Sodium 4-[(4-butoxyphenyl)thio]-20-[1-hydroxy-4-
[(5-hydroxyundecyl)oxy]but-2-yn-1-yl]biphenyl-3-sulfo-
nate (35e)
The title compound (30 mg, 47% from 33e) was synthe-
sized in the same manner as the general procedure for
1
compound 34. H NMR (400 MHz, CD3OD) d 1.04 (t,
3H, J = 7.4 Hz), 1.28-1.42 (m, 14H), 1.52–1.62 (m,
6H), 1.79–1.86 (m, 2H), 3.49 (t, 2H, J = 6.5 Hz), 3.57
(t, 2H, J = 6.7 Hz), 4.07 (t, 2H, J = 6.4 Hz), 4.19 (d,
2H, J = 1.5 Hz), 5.41 (brs, 1H), 6.87 (d, 1H,
J = 8.2 Hz), 7.05 (d, 2H, J = 8.7 Hz), 7.23–7.28 (m,
2H), 7.38 (t, 1H, J = 7.5 Hz), 7.43 (t, 1H, J = 7.5 Hz),
7.59 (d, 2H, J = 8.7 Hz), 7.83 (d, 1H, J = 7.5 Hz), 8.03
(d, 1H, J = 1.9 Hz); IR (thin film) 3376, 1593, 1489,
1458, 1387, 1367, 1249 cmꢀ1; HRMS (ESI, negative)
calcd for C37H47O7S2 (MꢀNa)ꢀ 667.2763; obsd
667.2756.
5.71. cAMP assay
References and notes
CHO cells stably expressing human S1P1 receptors
(CHO–S1P1 cells) were established in our laboratory
and maintained in selection medium (a-minimum
essential medium without ribonucleosides and deoxyri-
bonucleosides (Invitrogen) containing 10% dialyzed
fetal bovine serum (FBS, JRH Biosciences), 100 U/mL
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assay of a primitive derivative showed almost the same IC50
values (data not shown), cAMP assays were mainly used for
the evaluation of subsequent derivatives.
9. Although we tried a [32P] S1P-receptor binding assay for the
measurement of the acceptor specificity of our compounds,
it was unusable because some of our cell strains had an
inadequate acceptor expression amount. Therefore, we
evaluated the acceptor specificity of the compounds by
measuring the inhibition of a compound over the signal
transduction of each acceptor subtype.
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