Synthesis and structure-activity relationships of a new class of selective EP3 receptor agonist, 13,14-didehydro-16-phenoxy analogues of prostaglandin E1

Article abstract of DOI:10.1016/S0968-0896(99)00288-6

A series of 13,14-didehydro-16-phenoxy analogues of prostaglandin E₁ was synthesized and their agonistic activity on EP receptor subtypes was evaluated. 13,14-Didehydro-16-phenoxy-1-decarboxy analogues, 7e and 7f, display highly selective activity on the EP₃ receptor subtype, thus, their utility as a selective anti-ulcer agent can be expected. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0968-0896(99)00288-6

Bioorganic & Medicinal Chemistry 8 (2000) 353±362
Synthesis and Structure±Activity Relationships of a New Class of
Selective EP3 Receptor Agonist, 13,14-Didehydro-16-phenoxy
Analogues of Prostaglandin E1
Youichi Shimazaki, a,* Kazuya Kameo, a Tohru Tanami, a Hideo Tanaka, a Naoya Ono, a
Youichi Kiuchi, a Sentaro Okamoto, b Fumie Sato b and Atsushi Ichikawa c
aMedicinal Research Center, Taisho Pharmaceutical Co., Ltd., 403 Yoshino-cho, Ohmiya, Saitama 330-8530, Japan
bDepartment of Biomolecular Engineering, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama,
Kanagawa 226-8501, Japan
cDepartment of Physiological Chemistry, Faculty of Pharmaceutical Sciences, Kyoto University, Yoshida, Sakyo-ku,
Kyoto 606-8501, Japan
Received 26 August 1999; accepted 23 October 1999
AbstractÐA series of 13,14-didehydro-16-phenoxy analogues of prostaglandin E1 was synthesized and their agonistic activity on
EP receptor subtypes was evaluated. 13,14-Didehydro-16-phenoxy-1-decarboxy analogues, 7e and 7f, display highly selective
activity on the EP3 receptor subtype, thus, their utility as a selective anti-ulcer agent can be expected. # 2000 Elsevier Science Ltd.
All rights reserved.
Introduction
Among the EP3-receptor agonists reported so far, PG
analogues which have a o side chain substituted with a
phenoxy group at the C-16 (PG-numbering) position
such as enprostil8,9 and GR63799910 show enhanced
agonistic activity at the EP3 receptor subtype. Some of
us have recently revealed that the 13,14-didehydro deri-
vatives of natural PGE have a di€erent spectrum of the
biological activities than those of the natural com-
pound.11 These facts prompted us to investigate biolo-
Prostaglandins (PGs) play important roles in control-
ling a multitude of signi®cant physiological processes.
Receptors for the prostanoids have been divided into
®ve classes, EP, FP, IP, DP and TP,1 and recently it has
been reported that the EP receptor can be divided into
four subtypes, EP1, EP2, EP3 and EP4, each of which
mediates di€erent e€ects in various tissues and cells.2 Of
these, the EP3 receptor subtype has been revealed to
mediate contraction of the uterus,3 inhibition of gastric
acid secretion,4 release of neurotransmitters5 and
sodium/water reabsorption in kidney tubules.6
gical activities of 13,14-didehydro PGs having
a
phenoxy-substituted o side chain, and we report here
the synthesis of a series of 13,14-didehydro-16-phenoxy
PGE analogues and their biological evaluation with
regard to EP receptors (Fig. 1).
Numerous EP3 receptor agonists have been developed
in anticipation of their use as a potent antiulcer drug.7
However, many of these also demonstrate agonistic
activity at the other EP receptor(s), which results in
causing side e€ect(s) such as hypotension and diarrhea,
and thus, limiting their clinical use. Therefore, develop-
ment of a highly selective EP3-receptor agonist is an
attractive research subject.
Chemistry
13,14-Didehydro-16-phenoxy PGE1 analogues 5 and
their methyl esters 4 were synthesized by using the
method developed by us, which includes introduction of
the a side chain to exo-enone intermediate 1 by an
intermolecular radical addition reaction as a more
concise reaction than a 1,4-addition reaction using a
cuprate (Scheme 1).12 Thus, exo-enone 1, which was
prepared by the reaction of commercial 2-(N,N-diethyl-
amino)methyl-5-siloxy-2-cyclopentenone13,14 with the
Keywords: prostaglandins; agonists; receptors; substituent e€ects.
*Corresponding author. Tel.: +81-48-6663-1111; fax: +81-48-652-
7254; e-mail: s14801@ccm.taisho.co.jp
0968-0896/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(99)00288-6

354
Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
Figure 1.
Scheme 1.
alkynylaluminium reagent corresponding to the o side
chain having a phenoxy group,15 reacted with the alky-
liodide 2 and n-Bu3SnH in the presence of triethyl-
borane at 0 ꢀC to furnish the adduct 3, which in turn was
protodesilylated using aqueous HF in acetonitrile at
0 ꢀC to a€ord PG methyl esters 4. The ester moiety in 4
was converted into a carboxylic acid group by hydro-
lysis using esterase (PLE) to a€ord PG analogues 5,16
because 4 was unstable under the basic condition.
and the following Knoevenagel condensation reaction
of the resulting aldehyde with malonic acid, was con-
verted into the esters or amides of (E)-6-iodo-2-hexenoic
acid by treatment with NaI and the following con-
densation reaction with alcohol or amine after convert-
ing to the corresponding acid chloride. The iodo side
chain units 2 for the synthesis of other PGs shown in
Scheme 1 and Figure 2 were readily prepared by the
conventional methods.
Similarly, PG analogues 6a±f and 7a±o shown in Figure
2 were synthesized from 1 and the corresponding alky-
liodides by using the intermolecular radical addition
reaction as a key step.
Results and Discussion
The PG analogues prepared here were evaluated for
their ability as an agonist on EP3 receptor by investi-
gating the elastic activities on guinea pig vas deferens17
and the receptor binding assay. Receptor binding a-
nities were determined by the displacement of [3H]-
PGE2 from cloned receptors.18±21 The results, shown in
Tables 1±3, are expressed in terms of nano-molar ED50
The required a side chain units, alkyliodides 2, for syn-
thesizing PG analogues 4b, 5b and 6 were readily pre-
pared according to the procedure shown in Scheme 2.
Thus, (E)-6-chloro-2-hexenoic acid, which was prepared
from commercial 4-chlorobutanol by Swern oxidation
Figure 2.

Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
355
Scheme 2.
or IC50 values, and they show a good relation between
in vitro assay on vas deferens and the binding anity on
EP3.
corresponding acid derivative 5b and thus it loses its
selectivity. Therefore, we next investigated the activities
of the compounds 6a±f which have a bulky ester moiety
or amide group instead of a methyl ester moiety
(Table 2); however, it was found that these modi®ca-
tions reduced the activity on the EP3 receptor although
introduction of a bulky ester group decreased the rate of
hydrolysis: the t-butyl ester derivative 6b was hydro-
lyzed with esterase in plasma at least 3 times more
slowly than the compound 4b.22
As revealed from Table 1, 13,14-didehydro PGE1 has a
similar spectrum of receptor binding to that of PGE2,
except for the activity on EP1.11 Introduction of a
phenoxy group into the o side chain did not enhance the
anity on EP3 but provided a good selectivity for EP3.
Furthermore, the EP3 selectivity was considerably
improved by converting to the corresponding methyl
ester and/or by introducing an unsaturation (double
bond) between the C-2 and C-3 positions. Noteworthy
is the fact that the methyl ester derivatives exhibited
higher activity and selectivity on EP3 than the corre-
sponding carboxylic acid derivatives as observed on
enprostil and its acid derivative.8,9
Based on these results and to design modi®cation suit-
able to maintain the duration of activity in vivo, we
further investigated the ability of 13,14-didehydro-16-
phenoxy PGE1 derivatives having a functional group at
the C-1 position other than a carboxyl group (Table 3),
and it was found that 1-deoxo analogue of 5a or 5b, the
compound 7b or 7e, respectively, exhibited strong and
highly selective activity on the EP3 receptor. The results
shown in Table 3 indicate several features: (1) carbon
number (chain length) of the a side chain should be
Table 1 indicates that the compound 4b is a most potent
and selective analogue. However, the methyl ester
moiety in 4b may be easily hydrolyzed in vivo to the
Table 1. EP receptor e€ects of 13,14-didehydro PGE1 analogues
Compound
R
EP3 (vas deferens) (ED50, nM)
Receptor variants (IC50, nM)a
EP1 EP2
37.7
EP3
1.1
1.2
3.16
EP4
PGE2
13,14-didehydro PGE1
N.T.
N.T.
0.07
0.94
>1000
97.7
9.9
9.2
178
44
>1000
5a
4a
5b
4b
0.08
0.12
0.47
6.80
0.26
0.42
>1000
65.8
>1000
>1000
>1000
>1000
>1000
43.3
>1000
5c
4c
1.44
10.3
15.8
N.T.
N.T.
N.T.
27.0
58.9
N.T.
N.T.
N.T.
5d
4d
11.7
13.7
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
>100
enprostil
enprostil (Free acid)
0.07
0.06
1.72
1.87
319
4.01
>1000
>1000
>1000
50.8
aN.T=not tested.

356
Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
Table 2. EP receptor e€ects of 13,14-didehydro PGE1 analogues
Compound
R
EP3 (vas deferens) (ED50, nM)
Receptor variants (IC50, nM)a
EP1 EP2
0.94 37.7
EP3
EP4
9.92
>1000
>1000
PGE2
OMe
0.77
0.47
7.49
1.14
0.42
36.2
4b
6a
>1000
>1000
>1000
>1000
6b
77.6
>100
N.T.
N.T.
N.T.
6c
6d
6e
NH2
NMe2
24.5
55.2
63.9
36.6
N.T.
N.T.
141.8
N.T.
N.T.
>1000
N.T.
N.T.
>1000
N.T.
N.T.
6f
101
>100
N.T.
N.T.
N.T.
aN.T.=not tested.
adjusted to that of natural PGs (7b versus 7a or 7d), (2)
the ole®n geometry of the 2,3-double bond is important
(7e versus 7g), (3) introduction of a triple bond into the
a side chain cause loss of activity (7h and 7l), (4) oxygen
functionality at the C-1 position is necessary or the
activity would disappear (7m±o), and (5) methyl ether
derivatives (7c and 7f) and keto compounds (7j and 7k)
also exhibited the desired activity.
were performed with Perkin±Elmer 2400. TL chroma-
tograms were run on SiG/UV (Wako) using vaniline or
KMnO4 as a visualization agent. Mass spectra were
recorded with a JMS-SX102 (JEOL) mass spectrometer.
All optical rotations were determined at the sodium D
line using a DIP-360 (JASCO).
(3R,4R)-2-methylene-3-[(3R)-3-(t-butyldimethylsiloxy)-4-
phenoxy-1-butynyl]-4-(t-butyldimethylsiloxy)cyclopentan-
1-on (1). To an ice-cooled solution of (3R)-3-(t-butyl-
dimethylsiloxy)-4-phenoxy-1-butyne (3.85 g, 13.9 mmol)
in toluene (28.8 mL) was added n-butyllithium (1.95 M,
in hexane solution, 6.4 mL) under an argon atmosphere.
After stirring for 30 min, to the mixture was added di-
ethylaluminium chloride (0.97 M in hexane, 14.8 mL),
and the resulting mixture was stirred for 30 min at room
temperature. After cooling to 0 ꢀC, to this was added
(4R)-2-(N,N-diethylamino)methyl-4-(t-butyldimethylsil-
oxy)-cyclopent-2-en-1-on (0.25 M in toluene, 38.4 mL).
The resulting mixture was stirred for 15 min at room
temperature and poured into a mixture of saturated
NH4Cl aq (100 mL), hexane (100 mL) and 3 N HCl
(30 mL). The organic phase was separated and the aqu-
eous phase was extracted with hexane. The combined
organic layers were washed with saturated NaHCO3
aq (50 mL), dried over magnesium sulfate, ®ltered
(MgSO4), and concentrated in vacuo. The residue was
puri®ed by column chromatography on silica gel (hex-
With these results in hand, we selected the compound
7e as the most promising candidate to be explored in
vivo experiments, and we carried out investigations
on inhibition of gastric acid secretion in rat23 and anti-
ulcer activity on the stress ulcer.24 The results for 7e are
given in Table 4 together with those for 7f, and it was
found that these analogues are e€ective in both inhibi-
tion of acid secretion and protection against the stress
ulcer.
In conclusion, the PGE analogues having a 13,14-di-
dehydro-16-phenoxy structure synthesized here show
selective EP3-receptor agonistic activity. The pharma-
cological data presented here demonstrate that the
compounds 7e and 7f with their antisecretory and anti-
ulcer activity are potential agents for ulcer therapy.
Experimental
1
ane/diethyl ether (10/1)) to a€ord 3.27 g of 1. oil; H
Melting points were obtained in open capillary tubes
and were uncorrected. The NMR spectra were obtained
on either a Varian XL-200 or a Varian XL-300 spec-
trometer in CDCl3 or DMSO-d6 with Me4Si as an
internal standard. Infrared spectra were recorded on a
Perkin±Elmer 685 spectrometer. Elemental analyses
NMR (CDCl3, 200 MHz) d 0.11 (s, 3H), 0.12 (s, 3H),
0.15 (s, 6H), 0.90 (s, 9H), 0.91 (s, 9H), 2.33 (dd, J=18.0,
7.7 Hz, 1H), 2.72 (dd, J=18.0, 6.5 Hz, 1H), 3.50±3.60
(m, 1H), 3.97±4.09 (m, 2H), 4.23±4.35 (m, 1H), 4.73±
4.83 (m, 1H), 5.56 (dd, J=2.7, 0.6 Hz, 1H), 6.15 (d,
J=3.1 Hz, 1H), 6.85±7.00 (m, 3H), 7.21±7.34 (m, 2H).

Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
Table 3. EP receptor e€ects of 13,14-didehydro PGE1 analogues
357
Compound
R
EP3 (vas deferens) (ED50, nM)
Receptor variants (IC50, nM)a
EP1 EP2
EP3
EP4
PGE2
0.77
>100
1.14
N.T.
0.94
N.T.
37.7
N.T.
9.92
N.T.
7a
7b
7c
7d
1.55
>30
19.1
N.T.
N.T.
>1000
>1000
>1000
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
9.95
7e
7f
0.9
10.8
44.0
5.79
12.6
70
1000
>1000
>1000
>1000
1000
>1000
>1000
>1000
>1000
7g
7h
7i
>100
>100
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
7j
34.0
38.9
7.91
12.46
N.T.
1000
>1000
>1000
7k
7l
>1000
>1000
>1000
>100
N.T.
N.T.
N.T.
7m
7n
>100
>100
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
N.T.
7o
>100
N.T.
N.T.
N.T.
N.T.
aN.T.=not tested.
IR (neat): 2955, 2930, 2886, 2858, 2241, 1737, 1643,
1601, 1589, 1497, 1472, 1389, 1362, 1288, 1251, 1114,
chromatographed on silica gel (hexane/diethyl ether
1
(4/1)) to a€ord 481 mg of 3a. oil; H NMR (CDCl3,
1
1050, 1007, 975, 838, 780, 754 cm
.
200 MHz) d 0.10 (s, 3H), 0.12 (s, 3H), 0.13 (s, 3H), 0.14
(s, 3H), 0.89 (s, 9H), 0.91 (s, 9H), 1.17±1.80 (m, 10H),
2.10±2.32 (m, 1H), 2.17 (dd, J=18.3, 7.4 Hz, 1H), 2.29
(t, J=7.3 Hz, 2H), 2.60±2.74 (m, 2H), 3.66 (s, 3H), 4.00±
4.06 (m, 2H), 4.24±4.35 (m, 1H), 4.72±4.78 (m, 1H),
6.86±6.99 (m, 3H), 7.23±7.32 (m, 2H); IR (neat): 2952,
2931, 2858, 2241, 1747, 1601, 1497, 1464, 1362, 1251,
1172, 1116, 1050, 1007, 976, 838, 780, 755, 692, 670
cm 1; MS (FAB) (+KI) m/z: 669 (MK+).
16-Phenoxy-17,18,19,20-tetranor-13,14-didehydro-PGE1
methyl ester 11,15-bis(t-butyldimethylsilyl ether) (3a).
To a solution of 6-iodo-2-hexanoate (965mg, 3.19mmol)
and 1 (800 mg, 1.6 mmol) in toluene (2.0 mL) were
added n-Bu3SnH (0.86 mL, 3.19 mmol) and Et3B (1.0 M
in hexane, 0.16 mL) under an argon atmosphere at
0 ꢀC. After stirring for 4 h, the mixture was directly

358
Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
Table 4. In vivo-pharmacological activities of 13,14-didehydro PGE1
analogues
(ddd, J=6.9, 3.9, 1.9 Hz, 1H), 6.87±7.02 (m, 3H), 7.24±
7.34 (m, 2H); IR (neat): 3402, 2932, 2860, 2238, 1738,
1600, 1588, 1496, 1456, 1385, 1292, 1245, 1173, 1081,
1045, 908, 756, 693, 593, 510 cm 1; MS (FAB) (+KI)
m/z: 427 (MK+); HRMS m/z: 395.2043 (C22H29O6,
395.2046).
The following compounds were synthesized from 1 and
the corresponding alkyliodide 2 according to the same
procedure used for preparation of the compounds 3a, 4a
and 5a described above.
Compound
R
Acidity (T.A.O.a)
(ED30 mg/kg s.c.) (ED50mg/kg p.o)
Anti-ulcer
Famotidine25
7e
0.88b
0.013
0.11
0.036
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 methyl ester (4b). Oil; [a]2d4 24.09 (c
0.842, MeOH); 1H NMR (CDCl3, 300 MHz) d 1.38±
1.86 (m, 6H), 2.13±2.31 (m, 3H), 2.23 (dd, J=18.5,
9.1 Hz, 1H), 2.65 (ddd, J=11.4, 8.3, 1.8 Hz, 1H), 2.76
(ddd, J=18.5, 7.6, 1.3 Hz, 1H), 2.85 (d, J=3.5 Hz, 1H),
3.06 (d, J=5.2 Hz, 1H), 3.71 (s, 3H), 4.08 (dd, J=9.6,
6.9 Hz, 1H), 4.14 (dd, J=9.6, 4.0 Hz, 1H), 4.28±4.40 (m,
1H), 4.75±4.84 (m, 1H), 5.82 (dt, J=15.7, 1.4 Hz, 1H),
6.88±7.04 (m, 4H), 7.24±7.35 (m, 2H); IR (neat): 3412,
2933, 2861, 2241, 1744, 1723, 1656, 1600, 1588, 1496,
1456, 1438, 1385, 1291, 1246, 1174, 1080, 1044, 988,
910, 756, 693 cm 1; MS (FAB) m/z: 401 (MH+); HRMS
m/z: 401.1970 (C23H29O6, 401.1964).
7f
0.046
0.053
aT.A.O.=total acid output.
bED50 mg/kg s.c.
16-Phenoxy-17,18,19,20-tetranor-13,14-didehydro-PGE1
methyl ester (4a). To a solution of 3a (375 mg,
0.557 mmol) in acetonitrile (18.6 mL) was added 46%
HF aq (4.2 mL) at 0 ꢀC. The resulting mixture was stir-
red for 90 min at 0 ꢀC and poured into a stirred mixture
of saturated NaHCO3 aq (97 mL) and AcOEt (150 mL).
The organic layer was separated and the aqueous layer
was extracted with AcOEt (80 mL). The combined
organic layers were washed with brine (50 mL), dried
over MgSO4, concentrated and chromatographed on
silica gel (hexane/diethyl ether (1/9)) to a€ord 211 mg of
4a. white solid; mp 60±61 ꢀC [a]d23 32.75 (c 0.986,
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 (5b). Oil; [a]2d2 21.40 (c 0.995, MeOH);
1H NMR (CDCl3, 300 MHz) d 1.32±1.85 (m, 6H), 2.17±
2.32 (m, 3H), 2.23 (dd, J=18.6, 9.2 Hz, 1H), 2.64 (ddd,
J=11.4, 8.3, 1.8 Hz, 1H), 2.75 (ddd, J=18.6, 7.3,
1.3 Hz, 1H), 2.80±4.40 (br, 3H), 4.08 (dd, J=9.6, 6.8Hz,
1H), 4.14 (dd, J=9.6, 3.9 Hz, 1H), 4.29±4.39 (m, 1H),
4.80 (ddd, J=6.8, 3.9, 1.8 Hz, 1H), 5.82 (dt, J=15.6,
1.4 Hz, 1H), 6.90±7.10 (m, 4H), 7.25±7.34 (m, 2H); IR
(neat): 3392, 2931, 2861, 2242, 1740, 1696, 1652, 1600,
1588, 1496, 1456, 1417, 1291, 1245, 1173, 1080, 1045,
984, 885, 756, 693 cm 1; MS (FAB) (+KI) m/z: 425
(MK+); HRMS m/z: 387.1810 (C22H27O6, 387.1808).
1
MeOH); H NMR (CDCl3, 300 MHz) d 1.22±1.83 (m,
10H), 2.22±2.30 (m, 1H), 2.23 (dd, J=18.5, 9.2 Hz, 1H),
2.29 (t, J=7.5 Hz, 2H), 2.63±2.70 (m, 1H), 2.75 (ddd,
J=18.5, 7.3, 1.2 Hz, 1H), 2.80 (d, J=3.3 Hz, 1H), 2.97
(d, J=5.0 Hz, 1H), 3.65 (s, 3H), 4.08 (dd, J=9.7,
7.1 Hz, 1H), 4.15 (dd, J=9.7, 3.8 Hz, 1H), 4.30±4.39 (m,
1H), 4.77±4.83 (m, 1H), 6.91±7.02 (m, 3H), 7.27±7.34
(m, 2H); IR (neat): 3430, 2933, 2859, 2241, 1741, 1600,
1588, 1496, 1456, 1374, 1246, 1173, 1081, 1046, 885,
757, 693, 511 cm 1; MS (FAB) m/z: 403 (MH+). Anal.
calcd for C23H38O6: C, 68.64; H, 7.51. Found: C, 68.45;
H, 7.55.
16-Phenoxy-4-oxa-17,18,19,20-tetranor-13,14-didehydro-
PGE1 methyl ester (4c). Oil; 1H NMR (CDCl3,
300 MHz) d 1.55±1.91 (m, 4H), 2.08±2.49 (m, 3H), 2.24
(dd, J=18.5, 8.9 Hz, 1H), 2.57 (t, J=6.3 Hz, 2H), 2.70
(ddd, J=11.2, 8.0, 1.9 Hz, 1H), 2.75 (ddd, J=18.5, 7.3,
1.3 Hz, 1H), 3.38±3.52 (m, 2H), 3.67 (t, J=6.3 Hz, 2H),
3.68 (s, 3H), 4.08 (dd, J=9.7, 6.9 Hz, 1H), 4.14 (dd,
J=9.7, 4.0 Hz, 1H), 4.27±4.41 (m, 1H), 4.79 (ddd,
J=6.9, 4.0, 1.9 Hz, 1H), 6.88±7.05 (m, 3H), 7.24±7.36
(m, 2H); IR (neat): 3412, 2929, 2872, 2239, 1741, 1600,
1588, 1496, 1439, 1365, 1329, 1246, 1175, 1080, 1045,
909, 758, 694, 511 cm 1; MS (SIMS) (+KI) m/z: 443
(MK+); HRMS m/z: 405.1920 (C22H29O7, 405.1913).
16-Phenoxy-17,18,19,20-tetranor-13,14-didehydro-PGE1
(5a). To a mixture of PLE (sigma, 210 units, suspended
in ammonium sulfate, 150 mL) in phosphate bu€er
(pH=8, 11 mL) was added a solution of 19a (91.2 mg)
in acetone (1 mL) and the mixture was stirred for 15 h at
room temperature. The resulting mixture was acidi®ed
(pH=4) with 0.1 N HCl. After the solution was satu-
rated with ammonium sulfate, the mixture was extrac-
ted with AcOEt (20 mL). The combined extracts were
washed with brine (20 mL), dried over MgSO4, ®ltered,
and concentrated in vacuo. The residue was puri®ed by
column chromatography on silica gel (ethyl acetate/
methanol (10/1)) to a€ord 73.3 mg of 5a. oil;
[a]2d3 16.54 (c 0.734, MeOH); 1H NMR (CDCl3,
300 MHz) d 1.00±1.84 (m, 10H), 2.16±2.36 (m, 1H), 2.23
(dd, J=18.6, 9.0 Hz, 1H), 2.32 (t, J=7.3 Hz, 2H), 2.65
(ddd, J=11.4, 8.4, 1.9 Hz, 1H), 2.74 (ddd, J=18.6, 7.3,
1.2 Hz, 1H), 4.02±4.17 (m, 2H), 4.28±4.38 (m, 1H), 4.80
16-Phenoxy-4-oxa-17,18,19,20-tetranor-13,14-didehydro-
1
PGE1 (5c). Oil; H NMR (CDCl3, 200 MHz) d 1.56±
1.87 (m, 4H), 2.10±2.40 (m, 1H), 2.23 (dd, J=18.5,
9.0 Hz, 1H), 2.57 (t, J=5.9 Hz, 2H), 2.61±2.82 (m, 1H),
2.74 (ddd, J=18.5, 7.3, 1.3 Hz, 1H), 3.38±4.19 (m, 5H),
3.69 (t, J=5.9 Hz, 2H), 4.10 (dd, J=9.7, 6.4 Hz, 1H),
4.15 (dd, J=9.7, 4.4 Hz, 1H), 4.24±4.41 (m, 1H), 4.82
(ddd, J=6.4, 4.4, 1.8 Hz, 1H), 6.82±7.04 (m, 3H),

Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
359
1
7.18±7.36 (m, 2H); IR (neat): 3402, 2930, 2873, 2242,
1739, 1600, 1588, 1496, 1456, 1393, 1293, 1246, 1175,
1083, 1046, 909, 758, 694, 594 cm 1; MS (SIMS) (+KI)
m/z: 429 (MK+); HRMS m/z: 391.1755 (C21H27O7,
391.1757).
MeOH); H NMR (CDCl3, 300 MHz) d 1.26±1.71 (m,
6H), 2.00±2.24 (m, 3H), 2.06 (dd, J=18.2, 7.9 Hz, 1H),
2.52±2.65 (m, 2H), 3.96 (dd, J=9.8, 6.6 Hz, 1H), 3.99
(dd, J=9.8, 5.0 Hz, 1H), 4.09±4.21 (m, 1H), 4.54±4.63
(m, 1H), 5.46 (d, J=5.5 Hz, 1H), 5.69 (d, J=6.0 Hz,
1H), 5.84 (d, J=15.5 Hz, 1H), 6.58 (dt, J=15.5, 6.9 Hz,
1H), 6.87 (brs, 1H), 6.90±6.97 (m, 3H), 7.24±7.35 (m,
3H); IR (neat): 3493, 3458, 3359, 3208, 2938, 2915,
2859, 2234, 1721, 1673, 1640, 1609, 1587, 1492, 1455,
1439, 1402, 1326, 1294, 1276, 1249, 1173, 1150, 1103,
1066, 1046, 901, 877, 765, 697, 665 cm 1; MS (LSIMS)
m/z: 386 (MH+). Anal. calcd for C22H27N1O5: C,
68.55; H, 7.06 N, 3.63. Found: C, 68.49; H, 6.95; N,
3.57.
16-Phenoxy-17,18,19,20-tetranor-4,4,5,5,13,14-hexade-
1
hydro-PGE1 methyl ester (4d). Oil; H NMR (CDCl3,
300 MHz) d 1.65±1.82 (m, 1H), 1.90±2.05 (m, 1H), 2.29
(dd, J=18.5, 9.1 Hz, 1H), 2.23±2.54 (m, 6H), 2.71±2.82
(m, 2H), 3.04±3.11 (m, 1H), 3.15±3.21 (m, 1H), 3.29 (s,
2H), 4.09 (dd, J=9.7, 7.0 Hz, 1H), 4.15 (dd, J=9.7,
3.9 Hz, 1H), 4.29±4.41 (m, 1H), 4.76±4.84 (m, 1H),
6.90±7.03 (m, 3H), 7.25±7.34 (m, 2H); IR (neat): 3402,
2930, 2873, 2242, 1739, 1600, 1588, 1496, 1456, 1393,
1
1293, 1246, 1175, 1083, 1046, 909, 758, 694, 594 cm
;
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 N,N-dimethylamide (6d). Oil; 1H NMR
(CDCl3, 300 MHz) d 1.37±1.96 (m, 6H), 2.12±2.35 (m,
3H), 2.23 (dd, J=18.5, 9.1 Hz, 1H), 2.51 (brs, 2H), 2.65
(ddd, J=11.3, 8.2, 1.7 Hz, 1H), 2.74 (ddd, J=18.5, 7.3,
1.2 Hz, 1H), 3.02 (s, 6H), 4.10 (dd, J=9.8, 6.5 Hz, 1H),
4.13 (dd, J=9.8, 4.8 Hz, 1H), 4.25±4.37 (m, 1H), 4.80
(ddd, J=6.5, 4.8, 1.7 Hz, 1H), 6.23 (d, J=15.0 Hz, 1H),
6.84±7.03 (m, 3H), 6.90 (dt, J=15.0, 7.5 Hz, 1H), 7.24±
7.36 (m, 2H); IR (neat): 3368, 3012, 2933, 2861, 2240,
1744, 1659, 1601, 1496, 1456, 1401, 1292, 1247, 1156,
1081, 1046, 982, 885, 756, 693, 666, 511 cm 1; MS
(LSIMS) (+KI) m/z: 452 (MK+); HRMS m/z:
414.2287 (C24H32N1O5, 414.2281).
MS (SIMS) (+KI) m/z: 437 (MK+); HRMS m/z:
399.1823 (C23H27O6, 399.1808).
16-Phenoxy-17,18,19,20-tetranor-4,4,5,5,13,14-hexade-
1
hydro-PGE1 (5d). Oil; H NMR (CDCl3, 300 MHz) d
1.66±1.80 (m, 1H), 1.84±1.98 (m, 1H), 2.29 (dd, J=18.6,
9.1 Hz, 1H), 2.29±2.58 (m, 7H), 2.70±2.86 (m, 2H), 4.11
(dd, J=9.8, 6.7 Hz, 1H), 4.16 (dd, J=9.8, 4.0 Hz, 1H),
4.31±4.44 (m, 1H), 4.35±5.00 (br, 3H), 4.84 (ddd,
J=6.6, 4.0, 1.7 Hz, 1H), 6.90±7.04 (m, 3H), 7.25±7.35
(m, 2H); IR (neat): 3401, 2930, 2874, 2244, 1731, 1600,
1496, 1456, 1385, 1290, 1246, 1156, 1077, 1044, 907,
756, 693, 667 cm 1; MS (FAB) (+KI) m/z: 423 (MK+);
HRMS m/z: 385.1657 (C22H25O6, 385.1651).
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 N-n-butylamide (6e). Oil; 1H NMR
(CDCl3, 300 MHz) d 0.92 (t, J=7.3 Hz, 3H), 1.27±1.89
(m, 10H), 2.12±2.30 (m, 3H), 2.23 (dd, J=18.6, 9.0 Hz,
1H), 2.65 (ddd, J=11.3, 8.3, 1.8 Hz, 1H), 2.67±2.99 (br,
2H), 2.74 (ddd, J=18.6, 7.3, 1.2 Hz, 1H), 3.24±3.34 (m,
2H), 4.09 (dd, J=9.6, 6.5 Hz, 1H), 4.15 (dd, J=9.6,
4.8 Hz, 1H), 4.26±4.37 (m, 1H), 4.80 (ddd, J=6.5, 4.8,
1.8 Hz, 1H), 5.70 (brt, 1H), 5.77 (d, J=15.3 Hz, 1H),
6.81 (dt, J=15.3, 7.3 Hz, 1H), 6.91±7.02 (m, 3H), 7.26±
7.34 (m, 2H); IR (neat): 3328, 2933, 2863, 2240, 1741,
1669, 1628, 1601, 1549, 1497, 1457, 1374, 1246, 1155,
1081, 1047, 983, 756, 693 cm 1; MS (FAB) (+KI) m/z:
480 (MK+); HRMS m/z: 442.2580 (C26H36N1O5,
442.2594).
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 1-ethylpropyl ester (6a). Oil; 1H NMR
(CDCl3, 300 MHz) d 0.88 (t, J=7.4 Hz, 6H), 1.38±1.86
(m, 11H), 2.14±2.30 (m, 2H), 2.23 (dd, J=18.5, 9.2 Hz,
1H), 2.64 (ddd, J=11.5, 8.3, 1.9 Hz, 1H), 2.75 (ddd,
J=18.5, 7.3, 1.2 Hz, 1H), 3.06±3.34 (br, 2H), 4.08 (dd,
J=9.6, 6.9 Hz, 1H), 4.14 (dd, J=9.6, 4.0 Hz, 1H), 4.27±
4.39 (m, 1H), 4.73±4.86 (m, 2H), 5.82 (dt, J=15.6,
1.5 Hz, 1H), 6.87±7.03 (m, 4H), 7.25±7.35 (m, 2H); IR
(neat): 3418, 2970, 2937, 2879, 2242, 1745, 1714, 1653,
1600, 1588, 1497, 1460, 1384, 1246, 1174, 1104, 1081,
1046, 990, 910, 756, 693, 595, 510 cm 1; MS (LSIMS)
(+KI) m/z: 457 (MK+); HRMS m/z: 457.2597 (C27H36
O6K, 457.2590).
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 t-butyl ester (6b). Oil; [a]2d3 18.80 (c 1.64,
MeOH); H NMR (CDCl3, 300 MHz) d 1.40±1.86 (m,
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 N-isopropylamide (6f). Oil; 1H NMR
(CDCl3, 200 MHz) d 1.16 (d, J=6.5 Hz, 6H), 1.31±1.92
(m, 6H), 2.08±2.36 (m, 3H), 2.22 (dd, J=18.5, 9.0 Hz,
1H), 2.65 (ddd, J=11.2, 8.3, 1.9 Hz, 1H), 2.74 (ddd,
J=18.5, 7.2, 1.1 Hz, 1H), 3.96±4.39 (m, 4H), 4.74±4.84
(m, 1H), 5.46 (d, J=7.9 Hz, 1H), 5.72 (d, J=15.2 Hz,
1H), 6.79 (dt, J=15.2, 7.0 Hz, 1H), 6.89±7.03 (m, 3H),
7.23±7.36 (m, 2H); IR (neat): 3306, 2974, 2932, 2861,
2242, 1742, 1667, 1623, 1601, 1548, 1497, 1457, 1367,
1335, 1291, 1246, 1172, 1081, 1046, 983, 884, 756, 693,
667 cm 1; MS (FAB) (+KI) m/z: 466 (MK+); HRMS
m/z: 428.2442 (C25H34N1O6, 428.2437).
1
6H), 1.48 (s, 9H), 2.11±2.31 (m, 3H), 2.23 (dd, J=18.6,
9.2 Hz, 1H), 2.65 (ddd, J=11.3, 8.2, 1.8 Hz, 1H), 2.76
(ddd, J=18.6, 7.3, 1.3 Hz, 1H), 4.09 (dd, J=9.6, 6.8 Hz,
1H), 4.14 (dd, J=9.6, 4.1 Hz, 1H), 4.28±4.39 (m, 1H),
4.76±4.83 (m, 1H), 5.74 (dt, J=15.6, 1.5 Hz, 1H), 6.84
(dt, J=15.6, 7.0 Hz, 1H), 6.90±7.03 (m, 3H), 7.24±7.35
(m, 2H); IR (neat): 3413, 2978, 2933, 2862, 2242, 1746,
1713, 1651, 1600, 1589, 1497, 1456, 1393, 1369, 1318,
1246, 1158, 1081, 1046, 986, 851, 756, 693 cm 1; MS
(FAB) (+KI) m/z: 481 (MK+); HRMS m/z: 443.2430
(C26H35O6, 443.2433).
2-Decarboxy-2-hydroxy-16-phenoxy-17,18,19,20-tetranor-
13,14-didehydro-PGE1 (7a). Oil; 1H NMR (CDCl3,
300 MHz) d 1.20±1.91 (m, 10H), 2.20±2.31 (m, 1H), 2.23
(2E)-16-Phenoxy-17,18,19,20-tetranor-2,3,13,14-tetrade-
hydro-PGE1 amide (6c). Oil; [a]d24
38.49 (c 0.53,

360
Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
(dd, J=18.5, 9.3 Hz, 1H), 2.64 (ddd, J=11.5, 8.4,
1.9 Hz, 1H), 2.74 (ddd, J=18.5, 7.3, 1.3 Hz, 1H), 3.32±
3.55 (br, 2H), 3.61 (t, J=6.5 Hz, 2H), 4.07 (dd, J=9.6,
7.0 Hz, 1H), 4.13 (dd, J=9.6, 3.9 Hz, 1H), 4.25±4.38 (m,
1H), 4.75±4.85 (m, 1H), 6.88±7.05 (m, 3H), 7.25±7.35
(m, 2H); IR (neat): 3369, 2932, 2859, 2242, 1740, 1600,
1588, 1496, 1456, 1403, 1385, 1293, 1246, 1156, 1080,
1047, 909, 757, 693, 510 cm 1; MS (FAB) (+KI)
m/z: 399 (MK+); HRMS m/z: 361.2029 (C21H29O5,
361.2015).
6.90±7.04 (m, 3H), 7.25±7.35 (m, 2H); IR (neat): 3369,
3013, 2931, 2860, 2242, 1742, 1600, 1588, 1496, 1456,
1385, 1292, 1155, 1082, 1046, 973, 909, 755, 692, 667,
594, 510 cm 1; MS (FAB) (+KI) m/z: 429 (MK+);
HRMS m/z: 373.2025 (C22H29O5, 373.2015).
(2E)-2-Decarboxy-2-methoxymethyl-16-phenoxy-17,18,
19,20-tetranor-2,3,13,14-tetradehydro-PGE1 (7f). Oil;
[a]2d2 27.86 (c 0.974, MeOH); 1H NMR (CDCl3,
300 MHz) d 1.30±1.84 (m, 6H), 1.98±2.10 (m, 2H), 2.17±
2.30 (m, 1H), 2.23 (dd, J=18.6, 9.0 Hz, 1H), 2.65 (ddd,
J=11.2, 8.1, 1.8 Hz, 1H), 2.74 (ddd, J=18.6, 7.3,
1.4 Hz, 1H), 2.70±3.06 (br, 2H), 3.31 (s, 3H), 3.85 (dd,
J=6.0, 0.9 Hz, 2H), 4.07 (dd, J=9.6, 7.1 Hz, 1H), 4.13
(dd, J=9.6, 4.0 Hz, 1H), 4.28±4.38 (m, 1H), 4.78 (ddd,
J=7.1, 4.0, 1.9 Hz, 1H), 5.48±5.73 (m, 2H), 6.90±7.03
(m, 3H), 7.26±7.35 (m, 2H); IR (neat): 3401, 2930, 2859,
2241, 1744, 1600, 1588, 1496, 1454, 1385, 1292, 1246,
1154, 1078, 1046, 975, 908, 756, 693, 595, 509 cm 1; MS
(FAB) (+KI) m/z: 425 (MK+); HRMS m/z: 387.2176
(C23H31O5, 387.2172).
2-Decarboxy-2-hydroxymethyl-16-phenoxy-17,18,19,20-
tetranor-13,14-didehydro-PGE1 (7b). Oil; [a]2d3 24.10 (c
1
1.17, MeOH); H NMR (CDCl3, 300 MHz) d 1.00±1.94
(m, 12H), 2.05±2.35 (m, 1H), 2.23 (dd, J=18.6, 9.3 Hz,
1H), 2.65 (ddd, J=11.4, 8.3, 1.7 Hz, 1H), 2.74 (ddd,
J=18.6, 7.3, 1.2 Hz, 1H), 3.20±3.68 (br, 2H), 3.62 (t,
J=6.4 Hz, 2H), 4.07 (dd, J=9.6, 7.1 Hz, 1H), 4.13 (dd,
J=9.6, 3.6 Hz, 1H), 4.27±4.38 (m, 1H), 4.78 (ddd,
J=6.8, 3.8, 1.8 Hz, 1H), 6.87±7.06 (m, 3H), 7.25±7.36
(m, 2H); IR (neat): 3392, 2931, 2858, 2242, 1742, 1600,
1588, 1496, 1456, 1374, 1292, 1246, 1172, 1080, 1047,
910, 756, 693, 594, 510 cm 1; MS (LSIMS) (+KI): 413
(MK+); HRMS m/z: 375.2042 (C22H31O5, 375.2034).
(2Z)-2-Decarboxy-2-hydroxymethyl-16-phenoxy-17,18,
19,20-tetranor-2,3,13,14-tetranor-PGE1 (7g). Oil; 1H
NMR (CDCl3, 300 MHz) d 1.25±2.16 (m, 9H), 2.20±
2.29 (m, 1H), 2.23 (dd, J=18.5, 9.1 Hz, 1H), 2.66 (ddd,
J=11.3, 8.2, 1.8 Hz, 1H), 2.74 (ddd, J=18.5, 7.3,
1.3 Hz, 1H), 3.20±3.38 (br, 1H), 3.39±3.62 (br, 1H), 4.07
(dd, J=9.6, 6.9 Hz, 1H), 4.13 (dd, J=9.6, 4.1 Hz, 1H),
4.18 (d, J=6.6 Hz, 2H), 4.26±4.38 (m, 1H), 4.69±4.85
(m, 1H), 5.42±5.67 (m, 2H), 6.89±7.03 (m, 3H), 7.25±
7.35 (m, 2H); IR (neat): 3369, 3014, 2931, 2860, 2242,
1741, 1600, 1588, 1496, 1456, 1385, 1293, 1246, 1155,
1081, 1045, 912, 756, 693, 503 cm 1; MS (FAB) (+KI)
m/z: 411 (MK+); HRMS m/z: 411.1578 (C22H28O5K,
411.1574).
2-Decarboxy-2-methoxymethyl-16-phenoxy-17,18,19,20-
tetranor-13,14-didehydro-PGE1 (7c). Oil; 1H NMR
(CDCl3, 300 MHz) d 1.18±1.88 (m, 12H), 2.20±2.31 (m,
1H), 2.23 (dd, J=18.5, 9.3 Hz, 1H), 2.65 (ddd, J=11.4,
8.3, 1.8 Hz, 1H), 2.74 (ddd, J=18.5, 7.3, 1.3 Hz, 1H),
3.15±3.42 (m, 2H), 3.32 (s, 3H), 3.35 (t, J=6.6 Hz, 2H),
4.07 (dd, J=9.7, 7.1 Hz, 1H), 4.13 (dd, J=9.7, 4.1 Hz,
1H), 4.27±4.39 (m, 1H), 4.74±4.85 (m, 1H), 6.90±7.05
(m, 3H), 7.25±7.36 (m, 2H); IR (neat): 3402, 2931, 2858,
2242, 1746, 1600, 1588, 1497, 1456, 1374, 1292, 1246,
1156, 1083, 1046, 910, 756, 693, 594, 510 cm 1; MS
(FAB) (+KI): 427 (MK+); HRMS m/z: 389.2344
(C23H33O5, 389.2328).
2-Decarboxy-2-hydroxymethyl-16-phenoxy-17,18,19,20-
tetranor-2,2,3, 3, 13,14-hexadehydro-PGE1 (7h). Oil;
1H NMR (CDCl3, 300 MHz) d 1.42±1.90 (m, 6H), 2.17±
2.33 (m, 4H), 2.25 (dd, J=18.6, 9.2 Hz, 1H), 2.70 (ddd,
J=11.4, 8.3, 1.8 Hz, 1H), 2.76 (ddd, J=18.6, 7.3,
1.3 Hz, 1H), 3.17 (d, J=3.6 Hz, 1H), 3.27 (d, J=5.2 Hz,
1H), 4.10 (dd, J=9.7, 6.9 Hz, 1H), 4.15 (dd, J=9.7,
4.1 Hz, 1H), 4.20±4.27 (m, 2H), 4.29±4.40 (m, 1H),
4.76±4.84 (m, 1H), 6.91±7.04 (m, 3H), 7.26±7.35 (m,
2H); IR (neat): 3392, 2936, 2864, 2286, 2233, 1741,
1600, 1588, 1496, 1456, 1375, 1293, 1246, 1153, 1080,
1045, 1016, 911, 757, 693, 595 cm 1; MS (LSIMS)
(+KI): 409 (MK+); HRMS m/z: 409.1429 (C22H26
O5K, 409.1418).
2-Decarboxy-2-hydroxyethyl-16-phenoxy-17,18,19,20-
tetranor-13,14-didehydro-PGE1 (7d). White solid; mp
ꢀ
1
84±85 C; H NMR (CDCl3, 300 MHz) d 1.19±1.85 (m,
14H), 2.20±2.34 (m, 1H), 2.23 (dd, J=18.5, 9.4 Hz, 1H),
2.65 (ddd, J=11.5, 8.3, 1.8 Hz, 1H), 2.74 (ddd, J=18.5,
7.3, 1.3 Hz, 1H), 3.20±3.34 (br, 1H), 3.35±3.48 (br, 1H),
3.62 (t, J=6.6 Hz, 2H), 4.07 (dd, J=9.7, 7.1 Hz, 1H),
4.13 (dd, J=9.7, 4.0 Hz, 1H), 4.26±4.38 (m, 1H), 4.74±
4.83 (m, 1H), 6.85±7.03 (m, 3H), 7.25±7.36 (m, 2H); IR
(neat): 3403, 2920, 2855, 2238, 1740, 1601, 1589, 1499,
1467, 1455, 1412, 1366, 1331, 1301, 1280, 1241, 1196,
1173, 1153, 1116, 1093, 1074, 1051, 1004, 908, 882, 754,
691, 602, 577, 534, 506 cm 1; MS (FAB) (+KI) m/z:
427 (MK). Anal. calcd for C23H30O5: C, 71.11; H, 8.30.
Found: C, 70.96; H, 8.40.
2-Decarboxy-2-(10-hydroxyethyl)-16-phenoxy-17,18,19,20-
tetranor-13,14-didehydro-PGE1 (7i). Oil; 1H NMR
(CDCl3, 300 MHz) d 1.20±1.87 (m, 12H), 1.17 (d,
J=6.2 Hz, 3H), 2.20±2.35 (m, 1H), 2.23 (dd, J=18.5,
9.3 Hz, 1H), 2.65 (ddd, J=11.4, 8.4, 2.0 Hz, 1H), 2.74
(ddd, J=18.5, 7.3, 1.3 Hz, 1H), 3.25±3.44 (br, 2H),
3.71±3.84 (m, 1H), 4.07 (dd, J=9.7, 7.1 Hz, 1H), 4.13
(dd, J=9.7, 4.0 Hz, 1H), 4.26±4.39 (m, 1H), 4.75±4.82
(m, 1H), 6.87±7.03 (m, 3H), 7.25±7.35 (m, 2H); IR
(neat): 3392, 2931, 2858, 2242, 1741, 1600, 1589, 1496,
1456, 1375, 1292, 1246, 1156, 1082, 1047, 909, 756, 692,
(2E)-2-Decarboxy-2-hydroxymethyl-16-phenoxy-17,18,19,
20-tetranor-2,3,13,14-tetra-dehydro-PGE1 (7e). [a]d23
21.96 (c=1.02, MeOH); H NMR (CDCl3, 300 MHz)
1
d 1.30±1.85 (m, 6H), 1.94±2.12 (m, 2H), 2.16±2.34 (m,
1H), 2.23 (dd, J=18.6, 9.1 Hz, 1H), 2.34±2.60 (br, 3H),
2.66 (ddd, J=11.5, 8.2, 1.8 Hz, 1H), 2.74 (ddd, J=18.6,
7.3, 1.4 Hz, 1H), 4.03±4.17 (m, 4H), 4.28±4.38 (m, 1H),
4.78 (ddd, J=6.9, 4.0, 1.8 Hz, 1H), 5.56±5.72 (m, 2H),

Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
361
667, 510 cm 1; MS (FAB) (+KI) m/z: 427 (MK+);
HRMS m/z: 389.2015 (C23H33O5, 389.2022).
(+KI) m/z: 443 (MK+). Anal. calcd for C23H32O4S:C,
68.28; H, 7.97. Found: C, 68.00; H, 8.02.
2-Decarboxy-2-acetyl-16-phenoxy-17,18,19,20-tetranor-
13,14-didehdro-PGE1 (7j). White solid; mp 66.5±67.5 ꢀC;
1H NMR (CDCl3, 300 MHz) d 1.18±1.85 (m, 10H), 2.11
(s, 3H), 2.21±2.30 (m, 1H), 2.24 (dd, J=18.5, 9.2 Hz,
1H), 2.39 (t, J=7.3 Hz, 2H), 2.65 (ddd, J=11.4, 8.4,
1.8 Hz, 1H), 2.74 (ddd, J=18.5, 7.3, 1.3 Hz, 1H), 3.31
(d, J=4.2 Hz, 2H), 4.08 (dd, J=9.7, 7.0 Hz, 1H), 4.13
(dd, J=9.7, 4.0 Hz, 1H), 4.26±4.40 (m, 1H), 4.75±4.85
(m, 1H), 4.75±4.85 (m, 1H), 6.88±7.04 (m, 3H), 7.25±
7.35 (m, 2H); IR (neat): 3367, 3270, 2930, 2858, 2233,
1751, 1708, 1601, 1588, 1499, 1456, 1413, 1371, 1339,
1302, 1294, 1242, 1215, 1201, 1174, 1149, 1104, 1085,
1045, 902, 878, 816, 749, 689, 596, 507 cm 1; MS (FAB)
(+KI) m/z: 425 (MK+). Anal. calcd for C23H30O5: C,
71.48; H, 7.82. Found: C, 71.42; H, 7.92.
2-Decarboxy-2-ethyl-16-phenoxy-17,18,19,20-tetranor-
13,14-didehydro-PGE1 (7n). White solid; mp 51.5±
53.0 ꢀC; 1H NMR (CDCl3, 300 MHz) d 0.86 (t, J=
6.6 Hz, 3H), 1.16±1.85 (m, 14H), 2.22±2.34 (m, 1H),
2.23 (dd, J=18.5, 9.5 Hz, 1H), 2.66 (ddd, J=11.4, 8.4,
1.8 Hz, 1H), 2.74 (ddd, J=18.5, 7.3, 1.4 Hz, 1H), 2.96
(d, J=4.7 Hz, 1H), 3.16 (d, J=3.0 Hz, 1H), 4.07 (dd,
J=9.6, 7.2 Hz, 1H), 4.14 (dd, J=9.6, 3.7 Hz, 1H), 4.27±
4.41 (m, 1H), 4.75±4.84 (m, 1H), 6.86±7.04 (m, 3H),
7.25±7.36 (m, 2H); IR (neat): 3369, 2921, 2850, 2239,
1752, 1729, 1602, 1589, 1500, 1468, 1456, 1371, 1341,
1292, 1244, 1175, 1147, 1127, 1085, 1048, 904, 880, 814,
750, 726, 691, 508 cm 1; MS (FAB) (+KI) m/z: 411
(MK+). Anal. calcd for C23H32O4:C, 74.16; H, 8.66.
Found: C, 73.88; H, 8.60.
(2E)-2-Decarboxy-2-acetyl-16-phenoxy-17,18,19,20-tetra-
nor-2,3,13,14-tetradehydro-PGE1 (7k). Oil; [a]d22
24.99 (c=1.33, MeOH); H NMR (CDCl3, 300 MHz)
2-Decarboxy-2-acetamidomthyl-16-phenoxy-17,18,19,20-
tetranor-13,14-didehydro-PGE1 (7o). Oil; 1H NMR
(CDCl3, 300 MHz) d 1.20±1.85 (m, 12H), 1.98 (s, 3H),
2.17±2.36 (m, 1H), 2.24 (dd, J=18.5, 9.1 Hz, 1H), 2.68
(ddd, J=11.3, 8.2, 1.8 Hz, 1H), 2.74 (ddd, J=18.5, 7.3,
1.3 Hz, 1H), 2.85±3.35 (m, 4H), 4.08 (dd, J=9.7, 6.9 Hz,
1H), 4.13 (dd, J=9.7, 4.4 Hz, 1H), 4.28±4.39 (m, 1H),
4.79 (ddd, J=6.9, 4.4, 1.8 Hz, 1H), 5.67±5.81 (br, 1H),
6.88±7.02 (m, 3H), 7.24±7.34 (m, 2H); IR (neat): 3307,
2930, 2857, 2240, 1742, 1651, 1600, 1588, 1559, 1496,
1456, 1371, 1293, 1246, 1155, 1081, 1045, 909, 756, 693,
598, 509 cm 1; MS (FAB) (+KI) m/z: 454 (MK+);
HRMS m/z: 416.2455 (C24H34 N1O5, 416.2437).
1
d 1.25±1.84 (m, 6H), 2.16±2.30 (m, 3H), 2.23 (dd,
J=18.6, 9.1 Hz, 1H), 2.24 (s, 3H), 2.65±3.00 (br, 2H),
2.64 (ddd, J=11.4, 8.2, 1.8 Hz, 1H), 2.75 (ddd, J=18.6,
7.3, 1.3 Hz, 1H), 4.08 (dd, J=9.7, 6.8 Hz, 1H), 4.13 (dd,
J=9.7, 4.0 Hz, 1H), 4.29±4.39 (m, 1H), 4.79 (ddd,
J=6.8, 4.0, 1.8 Hz, 1H), 6.07 (dt, J=16.0, 1.4 Hz, 1H),
6.77 (dt, J=16.0, 6.9 Hz, 1H), 6.90±7.04 (m, 3H), 7.25±
7.35 (m, 2H); IR (neat): 3401, 2932, 2861, 2242, 1744,
1670, 1625, 1600, 1588, 1496, 1456, 1427, 1364, 1292,
1
1247, 1155, 1081, 1045, 982, 910, 757, 694, 593 cm
;
MS (FAB) (+KI) m/z: 423 (MK+); HRMS m/z:
385.2026 (C23H29O5, 385.2015).
Determination of EP3 activity (vas deferens)
2-Decarboxy-2-propionyl-16-phenoxy-17,18,19,20-tetra-
1
Male guinea-pigs were killed by a blow to the head fol-
lowed by exsanguination. Preparations of vas deferens
were mounted in organ baths for recording of tension
with isometric transducers. The bathing solution was
aerated with 95% O2 and 5% CO2 and maintained at
37 ꢀC. Lengths (20 mm) of vas deferens were suspended
in Krebs solution (no additions) between electrodes for
supra-maximal electrical ®eld stimulation (supramax-
imal voltage 1 ms, 10 Hz for 1 s every 32 s). 13,14-Dide-
hydro PGE1 derivative doses were added cumulatively.
Log concentration-inhibition curves were constructed.
nor-2,2,3,3,13,14-hexadehydro-PGE1 (7l). Oil; H NMR
(CDCl3, 300 MHz) d 1.12 (t, J=7.4 Hz, 3H), 1.45±1.87
(m, 6H), 2.210±2.40 (m, 3H), 2.24 (dd, J=18.6, 9.1 Hz,
1H), 2.55 (q, J=7.4 Hz, 2H), 2.66 (ddd, J=11.2, 8.2,
1.8 Hz, 1H), 2.75 (ddd, J=18.6, 7.2, 1.2 Hz, 1H), 3.17±
3.30 (m, 2H), 4.08 (dd, J=9.5, 6.9 Hz, 1H), 4.14 (dd,
J=9.5, 4.2 Hz, 1H), 4.28±4.43 (m, 1H), 4.74±4.86
(m, 1H), 6.90±7.04 (m, 3H), 7.25±7.36 (m, 2H); IR
(neat): 3419, 2938, 2866, 2211, 1744, 1672, 1600,
1588, 1496, 1458, 1409, 1376, 1293, 1246, 1176, 1079,
1046, 913, 799, 757, 693, 593, 511 cm 1; MS (FAB)
(+KI): 435 (MK+); HRMS m/z: 397.2006 (C24H29O5,
397.2015).
EP1ꢁEP4 receptor binding assay
[5,6,8,11,12,14,15-3H]PGE2 (200 Ci/mmol) was obtained
from Daiichikagaku, [3H] iloprost (14.1 Ci/mmol), and
iloprost were obtained from Amersham Corp. PGE2
was purchased from Funakoshi Phamaceuticals (Tokyo,
Japan). All other chemicals were of reagent grade. The
establishment of Chinese hamster ovary (CHO) cell line
stable expressing each prostanoid receptor has been
described previously.
2-Decarboxy-2-methylthiomethyl-16-phenoxy-17,18,19,20-
tetranor-13,14-didehydro-PGE1 (7m). White solid; mp
75.5±76.5 ꢀC; H NMR (CDCl3, 300 MHz) d 1.21±1.84
1
(m, 12H), 2.08 (s, 3H), 2.20±2.31 (m, 1H), 2.23 (dd,
J=18.5, 9.3 Hz, 1H), 2.47 (t, J=7.2 Hz, 2H), 2.65 (ddd,
J=11.5, 8.4, 1.9 Hz, 1H), 2.75 (ddd, J=18.5, 7.3,
1.3 Hz, 1H), 3.19 (d, J=4.9 Hz, 1H), 3.41 (d, J=3.3 Hz,
1H), 4.07 (dd, J=9.7, 7.2 Hz, 1H), 4.13 (dd, J=9.7,
3.8 Hz, 1H), 4.28±4.40 (m, 1H), 4.76±4.85 (m, 1H),
6.89±7.03 (m, 3H), 7.25±7.35 (m, 2H); IR (neat): 3370,
2926, 2850, 2240, 1748, 1601, 1589, 1499, 1468, 1456,
1370, 1340, 1302, 1292, 1238, 1197, 1175, 1147, 1081,
1048, 909, 881, 812, 748, 689, 545, 507 cm 1; MS (FAB)
The harvested CHO cells expressing each receptor sub-
type were homogenized using a Potter-Elvehjem homo-
genizer in 10 mM Tris±HCl (pH 7.4), containing 10 mM
MgCl2, 1 mM EDTA, 20 mM indomethacin, and
0.1 mM phenylmethylsulfonyl ¯uoride. The membrane

362
Y. Shimazaki et al. / Bioorg. Med. Chem. 8 (2000) 353±362
fraction (250,000Âg pellet) was washed and suspended
in the same bu€er. The standard mixture comprised
4 nM [3H] PGE2 and 80 mg of the membrane prepared
from each receptor-expressing CHO cell type in 100 ml
of 10 mM Mes-NaOH, pH 6.0, containing 10 mM
MgCl2 and 1 mM EDTA (bu€er A). After incubation at
30 ꢀC for 1 h (10 min for EP1 receptor), the reaction was
terminated by the addition of ice-cold bu€er A, after
which the mixture was rapidly ®ltered through a What-
man GF/C ®lter. The ®lter was then washed with ice-
cold bu€er A and the radioactivity associated with the
®lter was measured by a liquid scintillation counter.
Nonspeci®c binding was determined using a 1000-fold
excess of the respective unlabeled PGs in the incubation
mixture. The speci®c binding was calculated by sub-
tracting the non-speci®c binding from the total binding.
References and Notes
1. Kennedy, I.; Coleman, R. A.; Humphrey, P. P. A.; Levy, G.
P.; Lumley, P. Prostaglandins 1982, 24, 667.
2. Coleman, R. A.; Smith, W. L.; Narumiya, S. Pharmacolo-
gical Reviews 1994, 46, 205.
3. Goureau, O.; Tan®n, Z.; Marc, S.; Harbon, S. Am. J. Phy-
siol. 1992, 263, C257.
4. Chen, M. C. Y.; Amirian, D. A.; Toomey, M.; Sanders, M.
J.; Soll, A. H. Gastroenterology 1988, 94, 1121.
5. Hedqvist, P.; Von Euler, U. S. Neuropharmacology 1972, 11,
177.
6. Sonnenburg, W. K.; Smith, W. L. J. Biol. Chem. 1988, 263,
6155.
7. Collins, P. W. J. Med. Chem. 1986, 29, 437.
8. Eglen, R. M.; Whiting, R. L. Br. J. Pharmacol. 1989, 98,
1335.
9. Carpio, H.; Cooper, G. F.; Edwards, J. A.; Fried, J. H.;
Garay, G. L.; Guzman, A.; Mendez, J. A.; Muchowski, J. M.;
Roszkowski, A. R.; Van Horn, A. R.; Wren, D. Pros-
taglandins 1987, 33, 169.
10. Armstrong, R. A.; Marr, C.; Jones, R. L. Prostaglandins
1995, 49, 205.
11. Tanami, T.; Kameo, K.; Ono, N.; Nakagawa, T.; Annou,
S.; Tsuboi, M.; Tani, K.; Okamoto, S.; Sato, F. Bioorg. Med.
Chem. Lett. 1998, 8, 1507.
12. Ono, N.; Yoshida, Y.; Tani, K.; Okamoto, S.; Sato, F.
Tetrahedron Lett 1993, 34, 6427.
13. Okamoto, S.; Kobayashi, Y.; Kato, H.; Hori, K.; Takahashi,
T.; Tsuji, J.; Sato, F. J. Org. Chem. 1988, 53, 5590.
14. Yoshino, T.; Okamoto, S.; Sato, F. J. Org. Chem. 1991, 56,
3205 and references cited therein. This compound is now
commercially available from Nissan Chemical Industries, Ltd
(Japan).
Gastric acid secretion studies in pylorus-ligated rat
Male Wistar strain rats (SLC, Shizuoka, Japan) were
used. They were kept in stainless steel cages and fasted
but were allowed free access to water for 18 h before the
experiments. The fasted animals were anesthetized with
ether and pylorus ligated following abdominal midline
incision. The animals were treated with 13,14-didehydro
PGE1 derivatives for 30 min before the operation. After
4 h the animals were sacri®ced for collection of gastric
juice. The volume and acidity of the gastric juice were
determined. The acidity of gastric juice was measured
by titration with 0.1 N NaOH up to pH 7.0 with phe-
nolphthalein as the indicator. Total acid output (TAO)
was calculated from the product of the acidity and the
volume of the gastric juice.
15. Ono, N.; Kawanaka, Y.; Sato, F. J. Chem. Soc., Chem.
Commun. 1994, 1251.
16. Hazato, A.; Tanaka, T.; Toru, T.; Okamura, N.; Bannai,
K.; Sugiura, S.; Manabe, K.; Kurozumi, S. Nippon Kagaku
Kaishi, 1983, 1390.
Gastric ulcer studies
The animals were fasted for 18 h. RWIS ulcer was
induced according to the general method. Rats were
placed in a stainless steel stress cage and immersed ver-
tically to the level of the xiphoid process in a water bath
maintained at 23 ꢀC. After the immersion for 7 h the
animals were sacri®ced under anesthesia with ether.
17. Lawrence, R. A.; Jones, R. L.; Wilson, N. H. Br. J. Phar-
macol 1992, 105, 271.
18. Watabe, A.; Sugimoto, Y.; Honda, A.; Irie, A.; Namba,
T.; Negishi, M.; Ito, S.; Narumiya, S.; Ichikawa, A. J. Biol.
Chem. 1993, 268, 20175.
19. Honda, A.; Sugimoto, Y.; Namba, T.; Watabe, A.; Irie,
A.; Negishi, M.; Narumiya, S.; Ichikawa, A. J. Biol. Chem.
1993, 268, 7759.
The stomachs were removed and in¯ated with 10 mL of
1% formaline solution and were placed in the same
solution for 15 min. They were then opened along the
greater curvatures. The area of each ulcer in the corpus
and antrum portion was measured under stereoscopic
microscopy, and 13,14-didehydro PGE1 derivatives
were administered 10 min before the immersion.
20. Sugimoto, Y.; Namba, T.; Honda, A.; Hayashi, Y.;
Negishi, M.; Ichikawa, A.; Narumiya, S. J. Biol. Chem. 1992,
267, 6463.
21. Nishigaki, N.; Negishi, M.; Honda, A.; Sugimoto, Y.;
Namba, T.; Narumiya, S.; Ichikawa, A. FEBS Letters 1995,
364, 339.
22. The 13,14-didehydro PGE1 derivative was incubated with
the plasma of Rat (10 mg/ml) at 37 ꢀC for 30 min. After an
extractive work-up, the compounds (hydrolyzed acid and
remaining ester) were detected by HPLC analysis.
23. Shay, H.; Komarov, S. A.; Fels, S. S.; Meranze, D.;
Gruenstein, M.; Siplet, H. Gastroenterology 1945, 5, 43.
24. Takagi, K.; Okabe, S. Jpn. J. Pharmac. 1968, 18, 9.
25. Takagi, T.; Takeda, M.; Maeno, H. Arch. Int. Pharmaco-
dyn. Ther. 1982, 256, 49.
Acknowledgements
We thank Mr. Seki and Miss Saito for technical assis-
tance in EP3 activity and EP receptor selectivity studies,
and Mr. Higuchi and Mr. Nakamura for spectral data.