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Synthesis and Inhibitory Activity on Platelet Aggregation of 13 В╨Ж-Aza and Other ╨Я ░-Chain Modified BW245C Analogues.

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307
13’-Aza-modified BW245C Analogues
Synthesis and Inhibitory Activity on Platelet Aggregation of 13’-Aza and
Other a-Chain Modified BW245C Analogues
Paul Barraclougha)*,Michael Brockwella),A. Gordon Caldwella),Derek A. Demainea),C. John Harrisa),W. Richard Kinga),
Ray J. Stepneya),Clifford J. Whartona).and Brendan J.R. Whittleb).
Departments of Medicinal Chemistrya) and Pharmacologyb), Wellcome Research Laboratories, Langley Court, South Eden Park Road, Beckenham, Kent
BR3 3BS, U.K.
Received August 5, 1993
Synthese und Plattchenaggregation-hemmende Wirkung von 13’-Aza
und snderer in d e r UKette modifizierter Analoga von BW245C
BW245C analogues which have 15’-keto, -oximino, -sulphinyl, -sulphonyl, methyl, -I-adamantyl, 14’-hydroxy, 16’-hydroxy, 13’-14’-NH=CH, NH-CH2, or -NH-CO groups have been synthesized and evaluated for their
activity in inhibiting platelet aggregation and for their cardiovascular
actions: the 13’-aza analogues 13 and 14 are more potent inhibitors of
human platelet aggregation than BW245C (0.3, 0.6 and O.2xPGl2,respectively) and these inhibitory activities on platelet aggregation increase on
incubation in virro. The prostaglandin mimetic properties of 13 (BW68C)
and 14 (BW361C) were studied in more detail and their platelet inhibitory
and vasodilatory effects found to be of longer duration than those of
BW245C. All other modifications to the @chain of BW245C led to less
potent or inactive compounds.
Analoge von BW245C. die eine I5’-Keto-, -0ximino. -Sulfinyl-, Sulfonyl-, -Methyl-, -( 1-Adamantyl)-, 14’-Oxy-, 16’-0xy, 13’- I4’-N=CH-,
-NH2-CH2- oder eine NH-CHO-Gruppe enthalten, wurden synthetisiert
und auf Plattchenaggregationshemmung und Herzkreislaufwirksamkeit
untersucht. Es wurde gefunden, das die 13’Aza-andogen 13 und 14 st&kere Plattchenhemmener als BW245C (0.3, 0.6 und 0.2 x PGI2) sind. Die
Hemmung steigt bei Inkubation in virro. Die prostaglandinartigen Eigenschaften von 13 (BW68C) und 14 (BW361C) wurden niher untersucht,
und es wurde gezeigt, daE die plattchenhemmende und vasodilatorische
Wirkung Ihger als die von BW245C anhielt. Alle anderen Abiinderungen
an der o-Kette von BW245C ergaben weniger wirksame oder inaktive
Verbindungen.
The hydantoin prostaglandin (PG)analogue, BW245C (l)’.’),is a potent
inhibitor of human platelet aggregation and a powerful vasodilator.
BW245C is also a non-selective DP receptor agonist’) and its effects on
platelets and blood pressure are believed to be mediated, at least in part, by
DP receptors45). These effects are of relatively short duration in vivo. In
rats, for example, the half-life of the hypotensive effect of BW245C is
about 2-10 min.2), depending on the dose employed. Likewise, the inhibition of platelet aggregation observed ex vivo in anaesthetized rabbits, following intravenous infusion of 1, is no longer apparent 30 min after the
termination of the infusion?).
also rapidly excreted and two major metabolites are present
in rat urine. The structures of these metabolites are at
present unknown. Nonetheless we speculated that
BW245C, or a ringla-chain modified metabolite, would
undergo oxidation to a C-15’ ketone by a 15-OH PGDH in
vivo (but more slowly than PGD2) because of the several
potent PG mimetic properties and the bulkier cyclohexyl
group that BW245C possesses. However, neither the substrate specifi~ities~-’~)
of 15-OH PGDH’s, nor the effects of
w-chain modifications’.’ ‘1 on the platelet effects of PGD,
(and BW245C) have been investigated sufficiently for
many reliable predictions concerning metabolic stability or
PG mimetic activity of BW245C analogues to be made.
Analogues 2-15*)(Scheme 1) were therefore selected for
synthesis and pharmacological evaluation to extend our
knowledge of structure-activity relationships of hydantoin
PGD, mimics.
An analogue of BW245C, displaying a longer duration of
action, and also perhaps a more selective DP receptor agonist action, may provide a better probe than BW245C for
investigating the clinical utility of a PG agonist inhibiting
platelet aggregation. Structural modifications to BW245C
were therefore required which would give rise to potent
long-acting platelet aggregation inhibitory properties.
A major metabolic pathway for the deactivation of the pharmacological
effects of PG’s is their oxidation at the 15-hydroxyl group, by 15-hydroxyPG dehydrogenase( 15-OHPGDH)’). A majority of the PGD, metabolites
isolated from monkey and man have indeed undergone oxidation to a C-15
ketone. PGD2 is a poor substrate for most 15-OH PGDH isoenzymes
except for one NADP-linked 15-OH PGDH which is specific for the D
series of PG’s. Most PGD2 is probably initially converted to 9a, 1l!3PGD2
and this reduced PG then undergoes fwiher metabolism by the widely distributed 15-OH PGDH enzymes’).
In contrast only preliminary metabolic studies in rats have
been carried out on BW245C. These studies6)show that 1 is
rapidly converted into at least five metabolites. BW245C is
Arch. Pharm. (Weinheim) 327.307-317 (1994)
Chemistry
Ketone 2 was prepared by oxidation of BW245C (1)’)
with Cr03. Reaction of 2 with NH20H gave oxime 3.
Hydantoins 4-11 were obtained via C,N-disubstituted glycinates 23-29 (Scheme 2). These glycinate intermediates
were synthesized by N-mono alkylation of amine 16’). Of
the alkylating agents (A) employed, bromide 1712),oxirane
*)
All analogues are racemic
0 VCH Verlagsgesellschaft mbH, D-69451 Weinheim, 1994 0365-6233/94D505-0307 $5.00 + .25/0
308
Barraclough and coworkers
x.0
X - NOH
1
8
0
OH
7
H"K""'m
0
ia
12
?%"""-
WZH
HNKN$---(-J
OH
0
Scheme 2
14
Scheme 1
MI3),
and enone 2214)were prepared by known methods.
The iodide 19, bromide 20, and the brosylate 21 were
obtained by the routes shown in Schemes 3, 4, and 5 ,
respectively.
Reduction of ketone 28 with NaBH4 gave the corresponding alcohol, glycinate 29. Glycinates 23-27and 29 were
converted to the hydantoin-esters 30 as shown in scheme 2.
The crude esters 30 were not purified but hydrolysed directly to the corresponding acids.
Acids 4 and 10 were prepared by this method as racemates. Oxidation of sulphide 4 with Nd04 gave sulphoxide
5, while oxidation of 4 with Hz02yielded sulphone 6. Analogues 5, 7, 8, 9 and 11 were obtained as diastereomeric
mixtures. The individual diastereomers of 7, 9 and 11 were
readily separated from their more polar epimers 31 by
HPLC.
C4(CH2)3-
CH-c>
I
The synthesis of the 13'-aza analogue 12, starting from
bromide 3815)and the semicarbazone 3916),is given in
Scheme 6. A key step in the reaction sequence is the condensation of N-aminohydantoin 41 with cyclohexyl glyoxal
42l7.I8)
to yield ketone 43. Similarly, condensation of 41
and aldehyde 4419)gave diester 45 (Scheme 7). The individual diastereomers of 45 (but not 13)could be readily separated by HPLC. 45 was then converted into acids 13 and 14
as shown in Scheme 7. Analogue 15 was prepared from ahydroxy acid 4920321)
by a 3-step route (Scheme 8).
Structure-activity relationships
The ability of the hydantoin PG analogues to inhibit
ADP-induced platelet aggregation was measured in vitro
using human platelet rich plasma and the results are given
in Table 1. Analogues 2-8and 12 are inactive. Compounds
Ns"
l.m
OAC
14
Scheme 3
Arch. Pharm. (Weinheim) 327,307-317(1994)
13'-Aza-modified BW245C Analogues
309
OH
P
Scheme 4
R
SI
36
Scheme 5
43
Scheme 6
44
4s
NaBH CN.
HOAC3
1
Scheme 7
Scheme 8
Arch. Pharm. (Weinheim) 327,307-317 (1994)
OH
OAC
48
W
Barraclough and coworkers
310
Table 1: Inhibition of ADP-induced human platelet aggregation by BW245C and
analogues
Potencies relative to PGD, are. approximate. BW245C is approximately 8 and 0.2
times as potent as PGD2 and PG12, respectively. The relative potencies were calculated from the IC50 values of the compounds. ICso is the concentration required to
reduce the aggregation to 50% of its control amplitude. Compounds 1, 13, 14 and
PGI, have ICS0values (ng . ml-’), of 1.5 rt 0.1 (10). 1.1 f 0.2 (6).0.50 f 0.06 (8).
and 0.30 f 0.04 (10) respectively for (n) experiments. Inactive compounds have
IC5,’s > 200 ng . ml” and relative potencies i0.01 x PGD2. All compounds were
incubated with human PRP for 1 min. PG12 was used as a standard for each batch of
PRP.
[loll is the increase in relative potency on incubation of the compound with
human PRP for 5 or 10 min.
The 15’-epimers (more polar diastereomers) of all the above analogues are less
potent inhibitors of human platelet aggregation. The relative potencies of 47 and 48
for example are 0.01 x and 0.16 x PGI2, respectively. For 52,0.02 x PGD2.
a
9-11 and 15 are only weakly active, whereas the 13’-aza
analogues 13 and 14 are more potent inhibitors of platelet
aggregation than BW245C.
The inactivity of oxime 3 indicates that the C=NOH
group is a poor mimic of the 15’-CH(OH) moiety of
BW245C. Replacement of this essential functionality by
15’-CH222),CO, S(0) or SOz groups also results in loss of
platelet inhibitory properties. Similarly, displacing the 15’OH group of 1 to the 14’ or 16’ position, as in analogues 7
and 8, leads to inactive compounds. When the 15’-H of
BW245C is replaced by a 15’-Me group a 14-fold drop in
potency occurs. A marked decrease in potency (ca. 100fold) has also been reported”) for replacement of the 15’-H
by methyl in PGDz. Substitution of the 15-CH (C6Hll)
moiety of 1 by a (CH& unit also gives rise to a marked
decrease in potency. The weak activity of analogue 11 suggests that bulky 15’-groups, such as 1-adamantyl and tbutyl’) are not compatible with potent inhibition of platelet
aggregation in this series of compounds. Thus, analogues 911, which might be expected’-’’) to be poorer substrates for
15-hydroxy PGDH than BW245C, are also much weaker
agonists at the prostaglandin D2 receptor (DP) which mediates their effects on platelets.
The imino-ketone 12 is inactive and this observation is
not surprising in view of the lack of PG mimetic actions of
ketone 2. This result indicates that replacement of the 13’14’ CH2CH2moiety of 2 by a 13’-14’ N=CH group has no
observable effect on its lack of PG agonism. In contrast the
same structural change in BW245C leads to a small
increase in potency. Thus the potencies of imino-alcohol 13
and 1 are 12 x and 8 x PGD2, respectively. Replacement of
the 13’-CH2 group of BW245C by an NH unit gives rise to
a marked increase in potency. The amino-alcohol 14 is the
most potent (24 x PGD2) analogue of BW245C prepared to
date.
In contrast, the amide 15 is much less potent than 14 or 1.
This finding may reflect the weaker binding of 15 at the
platelet PG receptor, which may be related to less favourable H-bonding interactions, or the adoption of disfavoured
conformations.
The pharmacological properties of the two most potent
13’-aza analogues 13 (BW68C) and 14 (BW361C) were
studied further:
The anti-aggregating activity of these compounds
increased markedly on incubation with human platelet rich
plasma (PRP) or a suspension of platelets washed free of
plasma. Thus when 13 was incubated with human PRP for
5 or 10 min (instead of the normal 1 min period) its antiaggregating potency increased 5 or 11-fold respectively
(Fig. 1). A similar increase (5-fold after 5 min) was
observed with 14. Only smaller increases (3-fold after 10
min) were found with BW245C, PGD2 or PGIz under comparable conditions.
Incubation of 13 or 14 with plasma alone, with subsequent transfer to samples of human PRP, failed to enhance
activity. The above increase in activity with human PRP
therefore cannot be due to the action of plasma enzymes. In
order to investigate the mechanism of these increases in
activity, studies on the stimulation of platelet CAMP levels
were undertaken. PGDz and BW245C inhibit platelet aggre-
Arch. Phorm. (Weinheim) 327,307-317 (1994)
31 1
13'-Aza-modified BW245C Analogues
Irn
4
1
Table 2: Hypotensive activities of BW245C and analogues following bolus intravenous administration io anaesthetized rats.
Relative Potencies'
T
14
0.02
Po
0.3
Values are relative to prostacyclin in the same anaesthetized animal;n = 4.
Half-life (standard error mean f 20%) of hypotensive effects at a
dose that causes a fall in systemic arterial blood pressure of 20 mm
a
.
-.
0
2
4
8
8
10
Hg.
time of incubation (rnin)
Fig. 1: Inhibition of ADP-induced platelet aggregation following incubation of 13 with human PRP. Results, expressed as % inhibition of control
aggregation, are the mean f S.E.M of 4 experiments. and show the
increase in anti-aggregating activity of 13 following incubation in virro.
gation with a concomitant elevation of cAMP levels and
adenylate cyclase activity3).Cyclic AMP levels were determined in aliquots of human platelets following incubation
of several prostanoid agonists using both protein binding
techniques23)and radio-imm~noassay~~).
Incubation with
PG12 led to a rapid rise in cAMP levels, plateau levels of
15-18 times the basal levels being reached within 2 min.
With BW245C and PGD2, a lower maximal plateau of
about 7 times basal value was attained after 2 rnin. These
results provided support for BW245C and PGD2 acting at a
different receptor site (DP) to that of PG12 (EP). The stimulation by 13 and 14 reached levels (12 x basal) between
those achieved by PGI, and PGD2. In addition, these
experiments with 13 and 14 showed that cAMP levels continued to increase over a 10 min incubation period before
reaching maximum stimulation. Thus the time courses of
the cAMP elevating effects of 13 and 14 are the same as
those for their inhibitory properties on platelet aggregation.
Initial attempts to determine the PG receptor selectivities
of 13 and 14 in an isolated rabbit external jugular vein preparation4)were unsuccessf~l~~).
The relaxations induced by
these analogues are prolonged (> 1 h) and a steady state
response could not be attained. However the biphasic dose
response curves obtained suggested actions at more than
one PG receptor.
The cardiovascular effects of compounds 1, 13, and 14
were determined2)in anaesthetized rats and dogs (Tables 2
and 3). These three compounds display similar potencies in
their ability to lower blood pressure following intravenous
administration. The duration of vasodepression caused by
analogues 13 and 14, however, is much longer than that
induced by BW245C. In the dog, both 13 and 14 induced a
long-lasting fall in blood pressure and an increase in heart
rate which were still apparent after 2 h, making determination of the dose response curves difficult.
The ex-vivo anti-aggregating actions of the 13'-aza analogues 13 and 14 were investigated in the anaesthetized rabbit following both intravenous and oral administration.
Despite the lower sensitivity of rabbit platelets, compared
to human platelets, to the anti-aggregating actions of 13 in
vitro, it is a potent inhibitor of ex vivo platelet aggregation
following intravenous infusion (Table 4). The inhibition
was still apparent 2 hours after termination of the infusion.
Similar results were observed with 14 in contrast to those
with BW245C, where inhibition cannot be detected 30 minutes after administration of drug. Cardiovascular changes
were not detected following intravenous infusions of 13 and
14, but the absence of hypotensive effects probably reflects
the insensitivity of this species to such actions of PGDp
agonsts, since BW245C only displayed weak hypotensive
actions. Substantial inhibition of aggregation was also
Table 3: Hypotensive activities of BW245C and analogues following bolus intravenous administration to anaesthetized dogs.
-56*
6-
0.825
-5435
€Of15
1.25
-65*7
A0
0.625
-26s
>30
1.25
-5135
A0
2.5
13
14
Change in arterial blood pressure; n = 2.
Half-life of hypotensive effects.
Arch. Phorm. (Weinheim) 327,307-317 (1994)
312
Barraclough and coworkers
Table 4: Effect of BW245C and analogues on ADP-induced platelet aggregation, determined ex vivo in the anaesthetized rabbit following 15 min intravenous infusion
Inhb. ol RPbW Platelel Mn.
Conpound
Post-Infurion
hme. (min)
Bw245c
10
3.3
30
m
13
14
MI2
EDmb@g kg-lmin)
10
0.2
30
0.2
10
0.1
30
0.1
10
0.2
30
am
observed within 0.5 hour of oral administration of 13
(500 pg kg-') and remained essentially constant for at least
2.5 h.
In summary the 13'-aza analogues 13 (BW68C) and 14
(BW361C) have been prepared as racemates. Unfortunately
both compounds are gums which have to be stored in dilute
ethanolic solution at -2OOC to prevent spontaneous epimerisation. Nevertheless 13 and 14 are potent inhibitors of
human platelet aggregation (0.3 and 0.6 x PG12, respectively) and vasodepressors with a much longer duration of
action than BW245C. The increased activity of 13 and 14
on incubation with platelets in vitro and their prolonged
cardiovascular and anti-platelet actions in vivo could be the
consequence of slowly-developing associations of the drugs
with their PG receptor sites, followed by their slow dissociations. The pharmacology and SAR's of 13'-aza PGD2 agonists have been investigated in more detail and these studies
will be reported elsewhere.
Experimental Part
Melting points: Koffler hot-stage instrument, uncorrected. - 'H-NMR
spectra: Bruker HFX 90 (90 MHz), Bruker AM-200 (200 MHz) or WM360 (360 MHz) instruments, TMS as internal standard. - E.1. mass spectra:
Kratos MS-25 instrument at 70 eV. FAB mass spectra: as described26).- R
spectra: Perkin Elmer 580 instruments, liquids as thin films. - Thin layer
chromatogramms (t.1.c.): Merck silica gel 60F2s4, developed with 12 or
phosphomolybdic acid. - Flash chromatography: silica gel (230-400 mesh).
- Gas-liquid chromatography (G.1.c.): Pye-Unicam 204 series instrument.
(A) polydimethylsiloxane OV-101 column (B) polycyanopropylmethylphenylmethylsiloxane OV-225 column, flow rate 50 ml . min-' N2, FID
detector. - High performance liquid chromatography (HPLC): Bio-sil (2044 pm) column. CH2CI2-MeOH-HOAce.g. (93:5:2) eluant.
Preparation of Alkylating agents [A]
(f)-Cyclohe.ryloxirune (18)
The following modification of Cohen'sL3'procedure was used: m-chloroperoxybenzoic acid (46.1 g, 85%, 0.23 mol) was added in portions over
1 h to a stirred solution of vinylcyclohexane (25.0 g, 0.23 mol) in dry benzene (250 ml) with external cooling to maintain the temp. at 20°C. The
mixture was stirred for 20 h, and then filtered. The filtrate was poured onto
cold aqueous NaHC03 solution (500 ml of 5% w/v; 0.30 mol), the benzene
layer separated, and the aqueous solution extracted with benzene. The
combined extracts were washed with aqueous Na2S03 solution (80 ml of
5%), then with water and dried over MgS04. Evaporation, and distillation
of the residue, yielded 16.5 g (58%) of 18 as a colourless oil, bp. 62-64OC
(18 mm), lit.27):bp 63-65°C (14 mm).Glc purity 98.9%. R, 4.8 min (A),
85°C - C~H140(126.2).
(rt)-l-Acetoxy-1-cycloheiyl4iodohutane (19)
a ) 4-Chloroburyrylcyclohexane (32)
4-Chlorobutyronitrile (103.5 g, 1.0 mol) was added during I h to the
Grignard reagent prepared from chlorocyclohexane (1 18.5 g, 1.0 mol) and
Mg (24.3 g, I.O g-atom) in dry ether (700 ml). After an additional 1 h of
stirring. the mixture was poured onto crushed ice ( I kg) and conc. HCI
(500 ml). The ether layer was separated and discarded. The aqueous solution was heated I h on a steam bath. On cooling to room temp., an oil separated which was extracted with ether. and dried over MgSO.,. Evaporation, and distillation of the residue, gave 38.5 g (20%) of 32 as a colourless
oil. b.p. 73-75" (0.2 mm).Cmax (film): 1707 cm I . - 'H-NMR (200 MHz.
CDC13): 6 (ppm) = 0.90-1.53 (5H, complex m,aliphatic H), 1.53-1.93 (5H.
complex m, aliphatic H), 2.03 (2H, quint., J = 7 Hz, CICHzCH2), 2.36 (IH,
m, CHCO), 2.66 (2H, t, J = 7 Hz, CHzCO), 3.60 (2H. t. I = 7 Hz, CH2CI).
- Glc purity 98.8%. R, 5.6 min (A), 110°C. - Redistillation gave an analytically pure fraction, b.p. 74575°C (0.2 mm). - C I d I 7 C I O(188.7) Calcd.
C 63.6 H 9.08 Found C 63.8 H 9.14.
h ) I -Acetoxy-l-chloro-l-cyclohexylbutane (34)
Chloroketone 32 (38.0 g. 0.20 mol) was added during 1 h to a suspension of NaBH, (3.9 g, 0.10 mol) in ethanol (250 ml) in which NaOH (1.0
g, 0.03 mol) had been dissolved. The temp. rose to 4OoC and was kept
below 45'C by cooling. After an additional 1 h of stirring, the mixture was
cooled to 10°C and acidified with conc. HCI. Ethanol was removed in
vucuo and water (250 ml) added to the residue. The oily mixture was
extracted with ether (3x). the combined extracts were washed with water
and dried over MgSO,. The ether was evaporated to give 35.0 g (ca. 90%)
of 4-chloro-1-cyclohexylhutanol(33) as a pale yellow oil. A mixture of
crude 33 (35.0 g, 0.18 mol) and acetic anhydride (35.0 g, 0.34 mol) was
heated at 95°C for 1.5 h and then distilled at reduced pressure to yield 29.6
g (ca. 70%) of 34 as a colourless oil, b.p. 86-88OC (0.2 mm). - h a x
(film): 1736 cm-I. - 'H-NMR (200 MHz, CDCI,): 6 (ppm) = 0.88-1.35
Arch. Pharm. (Weinheim) 327,307-317 (1994)
13'-Aza-modified BW245C Analogues
(6H, complex m, aliphatic H), 1.40-1.85 (9H, complex m, aliphatic H),
2.05 (3H. s. CH,). 3.55 (2H. t, J = 7 Hz, CH2Cl), 4.75 (IH, m,HCOAc). Glc purity 99.3% (A), R, 6.0 min (1 10°C). - Redistillation gave an analytically pure fraction, b.p. 87-88°C (0.2 mm). - CI2H2,C1O2(232.7) Calcd. C
61.9 H 9.09 Found 62.0 H 8.95.
c) The chloroester 34 (20.0 g, 0.09 mol) was added to a solution of Nal
(60 g, 0.40 mol) in dry acetone (150 ml)and the mixture stirred and heated
at reflux for 8 h. After cooling NaCl was removed by filtration and the filtrate evaporated. The residue was treated with ether (150 ml) and water
(50 ml) and the org. extract separated and dried over MgSO,. Evaporation
and flash distillation of the residue (oil bath temp. 94-108"C, 0.02 mm)
gave 15.0 g (54%) of 19 as an unstable pale yellow oil. - CI2HZ11O2
(324.2) - Vmax (film): 1734 cm-I. - 'H-NMR (200 MHz, CDCI3): 6 (ppm)
= 0.88-1.35 (6H, complex m, aliphatic H), 1.40-1.90 (9H. complex m, aliphatic H), 2.05 (3H. s, CH3), 3.18 (2H, t, J = 7 Hz, CH21), 4.75 (lH, m,
HCOAc).
(f)-4-Bromo-2-cyclohexylbufan-2-ol
(20)
3-cyclohexylprop- I-ene-3-one (35)')(6.90 g. 0.05 mol) stirred in dry
ether (10 ml) at 0°C was treated dropwise with a freshly prepared solution
of HBr (4.0 g, 0.05 mol) in dry ether (20 ml). The resulting solution was
stirred at room temp. for 0.5 h, then added slowly to a solution of MeMgI
[fromMe1 (8.5 g, 0.06 mol), Mg turnings (1.5 g, 0.06 g-atom), and dry
ether (35 ml)] so as to maintain a steady reflux. The mixture was stirred at
room temp. for 2.5 h. treated with saturated aqueous NH4C1, filtered
(Celite), and the filtrate dried (Na2S04) and concentrated in vacuo. The
residue was purified by distillation giving 5.6 g (48%) of 20 as a pale yellow oil, b.p. 86-88OC (0.04 mm), m.p. 31-33°C. - Cld319BrO (235.2). - 'HNMR (90 MHz, CDC13): 6 (ppm) = 0.80- I .80 ( I 1H, complex m, C6HI'),
1.20 (3H, s, Me), 1.48 (lH, s, OH, exchang.), 2.04 (2H, m, CH2), 3.50
(2H, m, CH2Br).
I-[2-(4-Bromophenylsu[phonyloxy)erhyl]cyclohepranol(21)
a ) tert-Bury1 I -Hydroxycycloheprylacetare (36)
tert-Butyl acetate (95.0 g, 0.82 mol) was added over 20 min to a stirred
solution of LiNPr'2, prepared from n-BuLi (457 ml of 1.64 M hexane solution, 0.75 mol) and diisopropylamine (76.7 g, 0.76 mol) in dry tetrahydrofuran (600ml) at -78°C under dry N,. The mixture was stirred at -78°C for
1 h. a solution of cycloheptanone (84.1 g, 0.75 mol) in dry tetrahydrofuran
(100 ml) added over 30 min, and then stirring was continued for 3 h while
the mixture was allowed to warm to 0°C. 4 N HCI (ca. 250 ml) was then
added to neutralize the reaction mixture which was diluted with water (400
ml) and ether (400 ml). The org. layer was separated, the aqueous layer
extracted with ether, and the combined extracts dried over MgS04. Evaporation of the solvent and distillation of the residue yielded 138 g (80%) of
36 as a colourless oil, b.p. 93-95°C (0.8 mm), lit.28);b.p. 91-92°C (0.3
mm). - C13H2403 (228.3). - Glc purity 99.2%. R, (A) 7.0 min, 150°C.
- 'H-NMR (200 MHz, CDC13): 6 (ppm) = 1.34-1.91 (12H, complex m,
~ x C H ~1.48
) , (9H, S , ~xCH,),2.38 (2H, S, CHzCO), 3.65 (IH, bn, OH,
exchang).
b) 2-(l-Hydroxycycloheptyl)ethanol (37)
A solution of the ester 36 (130 g, 0.57 mol) in dry ether (200 ml) was
added over 1.5 h to a stirred suspension of L i A I b (16.4 g, 0.43 mol) in
dry ether (800 ml) at 0-5OC. The mixture was stirred at room temp. 96 h,
cooled to 0°C. and ethanol (10 ml) was added over 0.5 h. A saturated
aqueous solution (40 ml) of Na2S04 was then added over 0.5 h and the
suspension stirred for a further 2 h. The mixture was filtered (Celite), the
inorg. salts washed with ether, and the filtrate and washings evaporated.
Distillation of the residue yielded 68 g (75%) of 37 as a colourless syrup,
b.p. 106-110°C (0.1 mm). - C9HI8O2(158.2). - 'H-NMR (200 MHz,
Arch. Pharm. (Weinheim)327,307-317(1994)
313
CDCI,): 6 (ppm) = 1.22-1.80 (15H. complex m, 7xCH2+0H, 1H
exchang.), 3.45 (lH, br. peak, OH, exchang.) 3.86 (2H. t, J = 7 Hz, OCH,).
- Glc purity z 98% R, 6.0 min (B), 170°C.
c) p-Bromophenylsulphonyl chloride (37.0 g, 0.14 mol) was added in
portions over 0.5 h to a stirred solution of diol 37 (22.0 g, 0.14 mol) in dry
pyridine (60 ml) at 0-5°C. The mixture was stirred 3 h at room temp.,
poured onto ice (200 9). and the oil was decanted off. This oil was dissolved in CH2C12(250 ml), washed with water and dried over MgS04. The
CH2C12 was evaporated and the residual oil chromatographed on silica.
Elution with CH2C12 gave 26.2 g (50%) of 21 as a colourless syrup.
- C15H2~Br04S
(377.3). - 'H-NMR (200 MHz, CDCI,): 6 (ppm) = 1.201.77 (12H, complex m, 6xCH2), 1.88 (2H, t, J = 7 Hz, 0-C-CHI), 4.27
(2H, t, J = 7 Hz, OCH2), 7.74 (4H, AB-system, J = 9 Hz, arom. H). This
unstable material darkened and decomposed on standing at room temp. for
several days.
Preparation of Heterocyclic Prosraglandin Analogues.
(f)-3-(3-Cyclohexyl-3-oxopropyl)-2~-dioxo~-imida~olidineheptanoic
acid (2)
A solution obtained by dissolving Cr03 (0.60 g, 6.0 mmol) in 2N H2S04
(10 ml, 20 mmol) was added over 10 min to a stirred solution of 1')(1.0 g,
2.7 mmol) in acetone (20 ml). The mixture was heated at reflux for 3 h,
poured onto ice (30 g), and extracted with ethyl acetate (3x). The extracts
were dried over MgS04, evaporated, and the residual oil was stirred with
10% w/v aqueous NaHCO, (25 ml) solution for 1 h. The resulting aqueous
suspension was washed with ether (2x). the org. extracts discarded, and the
aqueous layer acidified with 2N HCl. The oily mixture was extracted with
ethyl acetate (3x) and the combined extracts dried over MgS04. Evaporation, and crystallisation of the residue from ether at OOC. gave 0.80 g
(80%) of 2 as colourless crystals, mp. 77.5-78.5"C. - C19H33N205 (366.5)
Calcd. C 62.3 H 8.25 N 7.64 Found C 62.5 H 8.34 N 7.49. - 'H-NMR (200
MHz, CDCl,): 6 (ppm) = 0.90-1.90 (20H. complex m,aliphatic H),2.202.60 (3H, m, CHCO + CH2C02), 2.79 (2H, m, CH2CO), 3.25 (IH, m,
NCHH), 3.74 (lH, m, NCHm),4.07 (lH, br. t, J = 5 Hz, COCHN), 8.5
(2H, v.br. peak. NH, OH, exchang.).
(f)-3-(3-cyclohexyl-3-hydroxyiminopropyl)-2,5-dioxo-4-imidazolidinehepranoic acid (3)
Ketone 2 (0.40 g, 1.1 mmol) was dissolved in aqueous Na2C03solution
(0.66 g, 6.2 m mol in 10 ml H20) and NH20H.HCI (0.20 g, 2.9 mmol)
added in a single portion. After stirring for 0.5 h, the mixture was kept at
room temp. in a sealed flask for 4 days, and then heated on a steam bath
for 2 h. The solution was cooled, washed with ether, and the aqueous layer
acidified with 2N HCI. The oily mixture was extracted with CHCI3 (3x).
extracts were dried over MgS04. Evaporation, and recrystallisation of the
residue from EtOAc, gave 0.22 g (53%) of oxime 3 as colourless crystals,
mp 159-160°C. - CI9H3,N3O5(381.5) Calcd. C 59.8 H 8.19 N 11.0 Found
C 59.7 H 8.23 N 10.7. - 'H-NMR (200 MHz. CDC13): 6 (ppm) = 0.90-1.90
(20H. complex m,aliphatic H), 2.25 (2H, t, J = 7 Hz,CH2C02),2.55 (3H,
m, CH2(C=N)CH), 3.24 (lH, m, NCHH), 3.78 (IH, m, N C m ) , 4.07 (IH,
t, J = 5 Hz, COCHN), 8.0 (lH, v.br. peak, OH, exchang.), 9.84 (IH, br.s.,
NH,exchang.).
(*)-(S*,R*) 3-(3-CycIohexyl-3-hydro~buryl)3,5-dioxo-4-imidazolidineheptanoic acid (9)
Bromide 20 (4.93 g. 20.9 mmol) and diethyl 2-aminononanedioate (16)
(5.2 g, 20.1 mmol) in ethanol (15 ml) were heated at reflux for 20 h. The
solvent was evaporated, water was added, and the mixture basified with
Na2C03. The oily mixture was extracted with ether, the ether extract
washed with water and dried over MgS04. Evaporation gave a yellow oil
(7.4 g) which was chromatographed on silica. Elution with H20-saturated
314
EtOAc gave 4.6 g (55%) of dierhyl 2-(3-cyclohe.ryI-3-hydroxyburylaminojnonanedioate (26) as a pale yellow oil, Rf 0.5 (Si02, Et2O). A solution
of KCNO (1.77 g, 21.9 mmol) in water (6 ml) was added gradually with
cooling and stimng to a solution of amino-diester 26 (4.5 g, 10.9 mmol) in
ethanol (20 ml) and 2N HC1 (1 1 ml. 22 mmol) at 5°C. The mixture was
stirred for 2 h and allowed to stand at room temp. overnight. Most of the
ethanol was evaporated, water was added, and the oil was extracted into
ether. The ether extract was washed with water and dried over MgS04.
Evaporation of the solvent gave an oil which was heated on the steam bath
for 5 h to give the crude ester 30 [R=(CH2)2C(Me)OHC6HII],as a yellow
oily mixture of diastereomers, Rr 0.55 and Rf 0.65 (SO2: CHCI,-MeOH,
9:l). containing some impurities. These esters (4.2 g) were stirred with
0.5N NaOH (50 ml,25 mmol) for 3 h. The insoluble non-acidic material
was removed by washing with ether, the clear alkaline solution acidified
with 2N HCI, and the liberated oil extracted with ether (3x). Evaporation
of the dried (MgS04)extracts gave a pale yellow viscous oil which on triturating in ether gave 3.1 g of a mixture of the epimers 9 and 31 as a
colourless solid, Rr 0.62 (SO2, CHC1,-MeOH-HOAc, 90:5:5). The individual diastereomers were obtained by HPLC (Si02, CH2CI2-MeOHHOAc, 98.25:1.25:0.5) yielding 0.96 g (23%) of the less polar epimer 9,
mp 173-174°C (EtOAc). - C20HMN205(382.5) Calcd. C 62.8 H 8.96 N
7.32 Found C 63.0 H 9.01 N 7.14. - 'H-NMR [200 MHz, CDC13(CD3)2S0(9:l)]:6 = 0.90-1.35 (13H, complex m. aliphatic H), 1.06 (3H. s.
Me), 1.40-1.90 (10H. complex m, aliphatic H), 2.20 (2H, t, J = 7 Hz,
CH2CO), 2.3-2.8 (IH, v.br. peak, OH, exchang), 3.06 (IH, m, NCHH),
3.66(1H,m,NCHH), 3.95 (IH, t, J = 5 Hz,COCHN),9.70(1H, br.s., NH,
exchang.).
1.2 g (29%) of the more polar epimer 31 [n=2, RI=Me, R2=C6HII]was
obtained as a colourless solid, mp. 109-1 11°C (EtOAc-hexane). C2"H3,N205 (382.5) Calcd. C 62.8 H 8.96 N 7.32 Found C 62.9 H 9.12 N
7.22.
In a similar manner reaction of amine 16 (5.2 g) with I-bromo-2cyclohexylthioethane (17)j2) (4.60 g) gave 6.0 g (75%) of diethy12-(2-cyclohexylfhioethy1)aminononunedioare (23) as a yellow oil, RI 0.8 (Silica, H2O
saturated EtOAc) which was converted as above to 3.5 g (64%)of (*)-3[(2-cyclohexy1thio)ethylj-2j-dioxo-4-imida;olidinehepranoic
acid (4). mp.
80-83OC (ether). - CIRH3&04S (370.5) Calcd. C 58.4 H 8.16 N 7.56
Found C 58.1 H 8.12 N 7.78. - 'H-NMR (200 MHz, CDCI3):6 = 1.10-2.05
(20H. complex m,aliphatic H), 2.32 (2H, t, J = 7 Hz,CH2C02). 2.73 (3H,
m, CH2SCH). 3.12 (1H. m, NCHH), 3.88 (IH. m, NCHH), 4.18 (IH, m,
COCHN),9.0(1H, br.s., NH, exchang.).
Barraclough and coworkers
C 62.8 H 8.96 N 7.32 Found C 62.9 H 9.21 N 7.27. - 'H-NMR (360 MHz,
CDCI?): 6 (ppm) = 0.85-2.0 (25H, complex m, aliphatic H), 2.33 (2H. t, J
= 7 Hz,CH2CO), 3.02 (IH, m,NCHH), 3.39 (IH, m, CHOH), 3.69 (IH,
m, NCHY), 4.05 (IH, m, COCHN), 8.35 (IH, br.s., NH,exchang.). - Rf
0.65 (Silica, CHCI3-MeOH-HOAc,90:5:5).
By a similar procedure to that used for the preparation of 26, reaction of
amine 16 (13.1 g) with the brosylate 21 (19.1 g) and LiCl(4.5 g) in ethanol
(40 ml) gave 8.1 g (40%) of dierhyl 2-[2-(1 -hydroxycyclohepryl)erhyl]aminononanedioare (27) as a yellow oil, Rf 0.3 (silica, ether-hexane 3:l).
This was converted as above (see preparation of 9) to 3.1 g (42%) of (k)-3[2-(l-hydroxycyc1ohepryl)efhyl~-25-dioxo~-imida~olidinehep1anoic
acid
(10). mp. 131-132°C (EtOAc). - C19H32N20S
(368.5) Calcd. C 61.9 H 8.75
N 7.60 Found C 61.9 H 8.75 N 7.54. - 'H-NMR (200 MHz, CDC13(CD&SO, 9:l): 6 (ppm) = 1.OO-1.88 (24H. complex m,aliphatic H), 2.12
(2H, t, J = 7 Hz,CH2CO), 3.00 (IH, m, NCHH), 3.53 (IH, m, NCHH),
3.90 (IH, t, J = 4 Hz, COCHN), 9.74 (IH, br.s. NH, exchang.).
(*)-3-[3-(1
-Adamantyl)-3-hydro.rypropy1]-2~-dioxo-4-imidazolidineheprunoic acid ( 11)
Amine 16 (2.59 g, 0.01 mol) and 3-(1-adamantyl)prop-l-en-3-one
(22)14)(1.93 g, 0.01 mol) were mixed at 0°C and set aside at room temp.
overnight giving dierhyl 2-1341-a&manryl)-3-oxopropyl)aminononanedioafe (28) as an oil. A stirred solution of this ketone (4.52 g, 0.01 mol) in
ethanol (35 ml) was treated dropwise at O°C with NaBH4 (0.20 g, 5.3
mmol) in ethanol (25 ml). kept at room temp. for 3.5 h, and then concentrated in vacuo. Water was added, the mixture was brought to pH 6 by N
HCI. and the product was extracted into ether. The extract was washed
with water, dried over MgSO,, and evaporation gave 4.2 g of dierhy12-[3(1-a~man~l)-3-hydroxypropyl]aminononanedioa1e
(29) as an oil consisting of two diastereomers, Rr 0.50 and 0.55 (SO2; CHC13-MeOH 50:l) and
minor impurities which could be removed by chromatography (SO2;
CHC13-MeOH. 50: I). This crude amino-alcohol was treated with KCNO
and 2N HCI, heated on a steam bath for 7 h. and then the resulting ester
hydrolysed with N NaOH, as for the preparation of 9, to give 1.1 g (26%
overall yield from 16) of 11, mp. 155-157°C (CHCI,-EtOAc). C~~H~,&OS
(420.6) Calcd. C 65.7 H 8.63 N 6.66 Found C 65.8 H 8.75 N
6.50. - 'H-NMR (200 MHz, (CD-O2SO):6 (ppm) = 0.90-1.78 (24H, complex m, 12xCH2). 1.87 (3H, br.s, adamantyl H), 2.15 (2H, t, J = 7 Hz,
CH2CO), 2.78 (IH, br.d, J = 9 Hz, CflOH), 3.10 (IH, m, NCHH), 3.25
(IH, br. peak, OH, exchang.). 3.39 (IH, m,NCHH), 4.04 (IH, t, J = 4 Hz,
COCHN), 4.2 (IH, br. peak, OH, exchang.), 10.6 (1H. br.s, NH, exchang.).
Reaction of amine 16 (2.6 g) with oxirane 18 (1.36 g) at 165°C in a
- HPLC also gave 1.1 g (26% overall) of 31 [n=2, Rl=H, RZ=l-adamansealed glass tube for 6 h gave 3.16 g (82%) of diethyl2-(2-cyclohexyI-2- tyl], mp. 178-179OC (CHCI3-Et2O-EtOAc).- C23H3&205(420.6) Calcd. C
hydroxyethy1)aminononanedioate (24) as a yellow oil, Rf 0.40 and 0.45
65.7H8.63N6.66FoundC65.9H8.81 N6.61.
(Silica, CHCI,-MeOH, 50: 1, two diastereomers) which was converted, as
above, to 0.65 g (22%) of (~))-3-(2-Cyclohexyl-2-hydroxyerhyl)-25-dioxo- (~)-3-~(2-CyclohexyIsu1phinyl)e1hyl]-2j-dioxo~-imidaiolidinehepfanoic
acid (5)
4-imidazolidineheptanoicacid (7). mp. 100-102°C. - C I 8 H a 2 O 5(354.5)
Calcd. C 61.0 H 8.53 N 7.90 Found C 60.9 H 8.79 N 7.72. - 'H-NMR (360
The sulphide 4 (0.74 g, 2.0 mmol), water (5 ml), and NaHC03 (0.175 g,
MHz, CDCI,): 6 (ppm) = 0.80-2.10 (21H. complex m. aliphatic H), 2.33
2.1 mmol) were stirred until all the solids had dissolved (ca. 0.5 h). The
(2H, t, J = 7 Hz, CH*CO), 2.98 (IH, m, NCBH), 3.67 (2H, m. CHOH +
resulting solution was added over 0.5 h to a stirred solution of NaI04 (0.44
NCHH), 4.33 (IH, m. COCHN). 7.91 (IH, br.s., NH, exchang.). - Rr0.73
g. 2.1 mmol) in water (8 ml) at 0-SOC; the mixture was stirred at 0-5°C for
(SO2, CHCI3-MeOH-HOAc, 9055). - HPLC also gave 0.41 g (14%) of
2 h and then 16 h at room temp. Inorg. salts (NalO,) were removed by filthe more polar epimer 31 [n=l, RI=H, R2=C6Hll]. mp. 131-134°C. tration, the filtrate was acidified with 2N HCI and the mixture extracted
C18H&05 (354.5) Calcd C 61.0 H 8.53 N 7.90 Found C 60.9 H 8.31 N
with CHCI3 (4x). The extracts were dried over MgS04, evaporated, and
7.70. - Rf 0.66 (SO2,CHCI,-MeOH-HOAc, 9053).
the residual gum crystallised from ether at -10°C to give 0.59 g (76%) of a
Reaction of amine 16 (5.18 g) and the iodoester 19 (6.48 g) by a similar
mixture of the two diastereomers of 5, mp. 75-80°C. - C18H30N205S
procedure to that used for obtaining 26 gave 6.4 g (70%) of dierhyl2-(4- (386.5) Calcd. C 55.9 H 7.82 N 7.25 Found C 55.7 H 7.98 N 7.03. - Rf
ocetoxy-4-cyclohe.ryIburylamino)nonanedioare(25) as a tan oil, Rf0.8
0.35 and 0.40 (Si02; CHCI,-MeOH-HOAC 9 0 5 5 ) . Vmax (KBr): 1 110,
(H20-saturated EtOAc), which was converted as above to 3.2 g (56%) of
985 cm-I. - 'H-NMR (200 MHz, CDCI3): 6 (ppm) = 1.10-2.22 (20H. com(f)-3-(4-Cyclohexyl-4-hydroxybutyI)-2,S-dioxod-imi&zolidineheptanoic plex m,aliphatic H), 2.31 (2H, t, J = 7 Hz, CH2CO), 2.71 (IH, m. CHSO),
acid 8 and 31, [n=3, Rl=H, R2=C6HII].mixture of diastereomers purified
2.82-3.23 (2H, complex m, CH2SO), 3.58 (IH, m, NC-m, 4.05 (IH, m,
but not separated by HPLC, mp. 102-106°C. - C2&,.,N2O5 (382.5) Calcd.
N W . 4.23 (IH, m,COCHN). 8.88 (IH, br.s, NH. exchang.).
Arch. Pharm. (Weinheim)327,307-317 (1994)
315
13'-Aza-modified BW245C Analogues
(f)-3-[(2-C~clohexylsrrlphonyl)ethyl]-2,5-dioxoll-imidazolidinehepta~ic
etheral solution was washed with N HCI. water, and dried over MgS04.
Evaporation gave an oil which was crystallised from ether. The resulting
acid (6)
solid was recrystallised from MeOH-H20 to give 0.86 g (44%) of 43 as
The sulphide 4 (1.0 g, 2.7 mmol)was added to a stirred solution of H,02
colourless plates, mp. 90-92°C. - C20H31N305(393.5) Calcd. C 61.1 H
(3 ml of 30% w/v. 26.5 mmol) in HOAc (12 ml) and then heated at reflux
7.94 N 10.7 Found C 60.8 H 7.65 N 10.5.
for 8 h. After cooling the solution was evaporated in vacuo, and the resid) Ester 43 (0.80 g. 2.0 mmol).2N NaOH (2.2 ml, 4.4 mmol), and water
due azeotroped (2x) with C C 4 to remove traces of HOAc. The residual
(7 ml) were stirred at room temp. for 1 h. The mixture was washed with
gum was triturated in dry ether and the resulting colourless solid crystalether, the aqueous solution acidified with 2N HCI, and the gum extracted
lised from ether to give 0.35 g (32%) of 6, mp. 99-102OC. - C 1 8 H d 2 0 6 S
with ether. The ether extract was washed with water and dried over
(402.5) Calcd. C 53.7 H 7.51 N 6.96 Found C 53.5 H 7.71 N 6.77. Vmax
MgSO,. Evaporation and recrystallisation of the residue from EtOAc and
(KBr) 1350, 1130 cm-I. - 'H-NMR (200 MHz. CDC13): 6 (ppm) = 1.05then MeOH-H,O gave 0.59 g (81%) of 12 as colourless plates, mp. 1502.25 (20H. complex m,aliphatic H), 2.33 (2H, t, J = 7 Hz, CH2CO). 2.90
152°C. - ClRH2,N3O5(365.4) Calcd 59.2 H 7.45 N 11.5 Found C 59.3 H
(IH, m, SCH), 3.16 (IH, m, SCHH). 3.34 (IH, m, SCHH), 3.58 (IH. m,
7.66 N 11.4. - 'H-NMR (200 MHz, CDC13-(CD3)2S0,9:l): 6 (ppm) = 1.2NCfIH), 4.07 (IH. m,N C W , 4.32 (lH, m, COCHN), 9.0 (1H. br.s, NH,
2.1 (20 H, complex m, aliphatic H), 2.26 (2H, t, J = 7 Hz, CH2C02). 2.3exchang.) - Rf 0.5 (SO2 CHC1,-MeOH-HOAc, 9 0 5 5 ) .
3.4 (2H, v.br. peak, OH, NH, exchang.), 3.32 (lH, m, CHCOC=N), 4.38
(IH, t, J = 5 Hz, COCHN), 7.88 (IH. S, CH=N).
(f)-3-[(2-Cyclohexyl-2-oxoethylidene)amino]-2,5-dioxo-4-imidazolidinehepranoic acid (12)
(~)-(S',R')-3-[(2-Cyclohexyl-2-hydroxyethylidene)amino]-2
S-dioxo-4a) (*)-Ethyl 3-Benzylideneamino-2
5-dioxo-4-imidazolidineheptanoare
imidazolidinehepranicacid (13).
(40)
a)Ethyl 3-[(2-ace1oxy-2-cyclohexylerhylidene)amino]-2
5-dioxo4-imidaA solution of NaOEt was freshly prepared by dissolving sodium (4.6 g,
0.20 g-atom) in dry ethanol (100 ml). Benzaldehyde semicarbazone (39)16)
(16.3 g, 0.10 rnol), the NaOEt solution (50 ml), and dry EtOH (20 ml)
were heated at reflux for 15 min. Diethyl 2-brotnononanedioate (38)15)
(16.0 g, 0.05 mol) was added, and the mixture was heated at reflux for 0.5
h. NaOEt solution (25 ml) was added, the mixture was heated at reflux for
5 min, bromodiester 38 (8.0 g. 0.025 mol) was then added and the mixture
was heated at reflux a further 0.5 h. The remaining NaOEt solution (25 ml)
and bromodiester 38 (8.0 g, 0.025 mol) were added and the mixture was
heated at reflux for 1 h. Most of the solvent was evaporated in vacuo and
the residue was shaken with 2N HCI and ether. Unreacted 39 (5.4 g) was
filtered off, the ether solution was washed with water and dried over
MgS0+ Evaporation, trituration of the oily residue with ether (20 ml), and
recrystallisation of the resulting solid from cyclohexane gave 13.0 g (36%)
of 40 as colourless needles. mp 92-94°C. C19H2JN304(359.4) Calcd. C
63.5 H 7.01 N 11.7. Found C 63.5 H 7.03 N 11.7. - 'H-NMR (200 MHz,
CDC13): 6 (ppm) 1.22 (3H. t. J = 7 Hz, CH3), 1.27-1.69 (8H, complex m,
4xCH2), 2.03 (2H, m,CH,CHCON), 2.26 (2H. t, J = 7 Hz,CH2CO), 4.10
(2H. q, J = 7 Hz, OCH2), 4.48 (IH, t, J = 5 Hz. COCHN), 7.40 (3H, m.
arom. H), 7.71 (2H, m,arom. H), 8.87 (IH, s, CH=N), 9.15 (IH, br.s, NH,
exchang.).
b) (&)-Ethyl3-Amino-25-dioxo4-imidazolidinehepranoate(41)
Benzylideneamine 40 (5.8 g, 0.016 mol), ethanol (100 ml), and 10%
PdC catalyst (0.6 g) were stirred at 50°C under H2 (10 aun) for 24 h. The
catalyst was removed, the filtrate evaporated and the residue crystallised
from EtOAc-hexane to yield 3.4 g (78%) of 41 as colourless needles, mp.
80-82OC. - CI2H2'N3O4(271.3) Calcd. C 53.1 H 7.80 N 15.5 Found C 53.3
H 8.00 N 15.4. - 'H-NMR (200 MHz. (CD3),SO): 6 (ppm) = 1.18 (3H. t. J
= 7 Hz, CH3), 1.27 (6H, m, 3xCH2), 1.50 (2H, m, CH,). 1.70 (2H, m.
CHzCHCON), 2.26 (2H, t, J = 7 Hz, CH,CO), 3.32 (2H, b r s , NH2,
exchang.), 3.90 (IH. t. J = 5 Hz, CHCON), 4.05 (2H, q, J = 7 Hz, OCHz),
10.5 (IH, br. peak, NH, exchang.).
c) (*)-Ethyl3-[(2-cyclohexyl-2-oxoethylidene)amino]-2
5-dioxo-4-imidazolidine heptanoate (43)
A solution of amine 41 (1.35 g, 5.0mmol) and cyclohexylglyoxal hemihydrate (42)".'*) (0.82 g, 5.5 mmol) in MeOH (8 ml) was heated at reflux
for 1 h. MeOH was evaporated and the residue was dissolved in ether. The
Arch. Pharm.(Weinheim)327,307-317(1994)
zolidineheptanoare (45)
A solution of amine 41 (1.1 g, 4.05 mmol) and 2-acetoxy-2-cyclohexylacetaldehyde (44)'" (0.82 g, 4.47 mmol) in MeOH (8 ml) was heated at
reflux for 1 h. MeOH was evaporated, the residue dissolved in ether, and
the ether solution was washed with N HCI, then water. and dried over
MgS0+ Evaporation gave an oil which was chromatographed on silica.
Elution with CHCI,-MeOH (40: 1) gave a colourless oil (1.8 g) consisting
of 45 and 46,Rr 0.50 and 0.46. respectively, silica, CHCI,-MeOH-HOAc
(95:4:1). HPLC (Lichrosorb Si 60, 10 pM, Dupont No. 48, CH,CI,MeOH-HOAc, 98.5: 1:OS) of this mixture gave the individual diasteree
mers: 0.63 g (35%) of 45 as a colourless viscous oil. - C22H35N306(437.5)
Calcd. C 60.4 H 8.06N 9.60 Found C 60.1 H 8.20 N 9.30; 0.85 g (47%) of
46 as a colourless viscous oil. - C22H3+$& (437.5) Calcd. C 60.4 H 8.06
N 9.60 Found C 60.1 H 8.32 N 9.90.
b) Ester 45 (0.50 g) and 0.5N NaOH (10 ml) were stirred at room temp.
for 1 h. The mixture was washed with ether, the aqueous solution acidified
with N HCI, and the liberated oil extracted with ether (2x). The extracts
were washed with water and dried over MgS04. Evaporation gave 0.37 g
(88%) of 13 as a colourless gum. - Cl8HZ9N3OS
(367.5). - 'H-NMR (200
MHz, CDC13): 6 (ppm) = 0.90-2.10 (21H, complex m, aliphatic H), 2.32
(~H,~,J=~HZ,CH~CO),~.~~(~H,~,J=~HZ,C~OH
Hz,COCHN), 8.23 (1H. d, J = 6 Hz, CH=N). - MS: m/z = 368 (MH+,
FAB). Hazard-avoidskin conraci - compound may cause skin irritation
and facialflushing.On standing (even at OOC) this compound slowly converted to a mixture of 13 and its C-8' epimer 47.Hydrolysis of ester 46 by
a procedure similar to that described above, gave 85% of 47.mp. 84436°C
(MeOH-H20). - C18H2fi305.H20 (385.5) Calcd. C 56.1 H 8.11 N 10.9
Found C 55.9 H 8.18 N 10.8. - 'H-NMR (200 MHz, (CD3)2SO): 6 (ppm) =
0.84-2.0 (21H. complex m, aliphatic H), 2.18 (2H, t. J = 7 Hz, CH2CO).
2.9-3.6 (2H, br. peak, 2xOH. exchang.), 3.78 (lH, t, J = 6 Hz, CHOH),
4.66 (lH, m,COCHN), 7.38 (IH, d, J = 6 Hz, CH=N).
(f)-(S'.R')-3-[(2-Cyclohexyl-2-hydroxyethyl)amino]-2J-dioxo-4-imidazolidineheptanoic acid (14)
A solution of acid 13 (0.29 g, 0.78 mmol) and NaBH3 CN (50 mg, 0.80
mmol) in MeOH.with acetic acid (0.3 ml) was stirred at 20°C for 48 h. The
solvents were removed in vacuo,water (10 ml) was added to the residue,
and the oil extracted with ether (2x). The extracts were washed with water,
dried over MgS04, and evaporated to yield 0.24 g (83%) of 14 as a colourless gum. - C18H31N305(369.5). - 'H-NMR (360 MHz, (CD&SO): 6
316
Barraclough and coworkers
(ppm) = 0.90-1.78 (21H, complex m, aliphatic H), 2.18 (2H, t, J = 7 Hz,
CHzCO), 2.82 (2H, m, CHzNH). 3.20-3.34 (3H. m, CHOH, COzH, 2H
exchang.), 4.04 (1H. t, J = 4 Hz, COCHN), 5.00 (IH, br.t, J = 7 Hz,
NBCHz). - MS: m/z = 370 (MH+, FAB). - Rf 0.6, silica, CHC13-MeOH
(5:l).
In a similar manner reduction of 47 gave 57% of 48, mp. 103-105°C
(EtOAc-hexane). - C18H31N305(369.5) Calcd. C 58.5 H 8.46 N 11.4
Found C 58.7 H 8.67 N 11.4. - 'H-NMR (360 MHz. (CD3)zSO): 6 (ppm) =
0.89-1.81 (21H, complex m, aliphatic H), 2.18 (2H, t, J = 7 Hz, CHzCO),
2.75 (lH, m, NCHH). 2.93 (lH, m, NCHfD, 3.22 (3H, m, CHOH, C02H,
2H exchang.), 4.07 (IH, t, J = 5 Hz, COCHN), 4.99 ( l H , m, NHCH2,
exchang.). - Rf0.6, silica, CHC13-MeOH (5:l).
Pharmacologicalmerhods
Inhibition of Platelet Aggregarion in Virro
Human blood was freshly collected into siliconized (Siloclad: Clay
Adams) plastic (Sterilin Ltd.) vessels containing trisodium citrate (3.15%;
0.1 volume with 0.9 volume blood) and centrifuged (200 g for 15 min) at
room temp. The platelet-rich plasma (PRP) was withdrawn into plastic
containers and kept at room temp. Inhibition of platelet aggregation was
determined by a Born-type aggregometer as describedM)by incubating aliquots (0.5 ml) of the PRP for 1 min at 37OC with or without the prostaglandin analogue prior to addition of sufficient adenosine diphosphate (ADP)
to just cause a non-reversing control aggregation. Dose-inhibition curves
were constructed for each compound and the ICso was calculated as that
required to reduce the aggregation to 50% of its control amplitude. Prostacyclin was used as a standard for each batch of PRP. Comparison of the
(f)-3-[(2-Cyclohexyl-2-hydroxy)acetamido~-2,5-dioxo-4-imidazolidineICso values of analogues with those of prostacyclin then allowed a potency
hepranoic acid (15)
ratio to be calculated; e.g. BW245C/PGI2 is 0.2. The potencies of analogues relative to PGD2 were estimated from potency ratios of PG12/PGDz
a ) (f)-2-Aceroxy-2-cyclohe*ylacericacid (50)
observed p r e v i o u ~ l y in
~ ~human
~ ~ ) PRP. Results in Table 1 show relative
(f)-2-Cyclohexyl-2-hydroxyaceticacid (49)20-2')(3.16 g, 0.02 mol) and
potencies following the 1 min incubation in PRP and also increase in
acetic anhydride (3.0 ml, 0.03 mol) were stirred at 100°C for 4 h. The
potency after 5-10 min incubation in PRP.
cooled mixture was treated with water (30 ml), stirred for 0.5 h, and then
extracted with ether (2x). The extracts were washed with water, dried over
Cardiovascular Acrions in Rats
MgSO,, and evaporated to yield a viscous oil which solidified on scratching. Recrystallisation from petroleum fraction bp. 40-60°C gave 2.28 g
Anaesthesia was induced in male Wistar rats (250-300 g body weight)
(57%) of 50, mp. 85-87OC. lit.29' (L-enantiomer): bp. 135-140°C (0.3 mm).
with sodium pentobarbitone (30 mg . kg.', i.v.) and supplemented (3 mg .
- C1&II6O4(200.2) Calcd. C 60.0 H 8.05 Found C 59.9 H 8.14. - 'H-NMR
kg-') when required. Arterial pressure was recorded from a cannulated
(200 MHz. CDC13): 6 (ppm) = 1.06-1.40 (5H, complex m, aliphatic H),
femoral artery, the resting values being in the range of 100-140 mm Hg.
1.60-2.0 (6H, complex m, aliphatic H), 2.15 (3H, s, CH4.4.88 (IH, d, J =
Rectal temp. was maintained at 37°C by thermistorcontrolled radiant heat.
5 Hz, CHO), 8.7 (IH, vb.r. peak, OH, exchang.).
Each compound was injected into a femoral vein in a volume of 0.25 ml
b) A solution of ethyl chloroformate (434 mg, 4.0 mmol) in dry tetrahyand flushed with 0.2 ml of saline (0.9% w/v). Dose-response relationships
drofuran (2 ml) was added to a mixture of acid 50 (800 mg, 4.0 mmol).
for the fall in mean systemic arterial blood pressure with the prostaglandin
NEtl (404 mg, 4.0 mmol), and tetrahydrofuran (12 ml) at - S T over 40
analogues were constructed and compared to those obtained with prostacymin. The reaction mixture was stirred at -5'C for 20 min and then a soluclin ( E l 2 ) in the same animal, and a potency ratio calculated (Table 2).
tion of amine 41 (1.08 g, 4.0 mmol) in tetrahydrofuran (4 ml) added over
10 min. Stirring was continued for 2 h at -5°C. the mixture allowed to
Cardiovascular studies in rhe dog
stand at room temp. overnight, and then the solvent was removed in vacuo.
Dogs (10-15 kg body weight) were anaesthetized with chloralose
The residue was dissolved in CHzClz (30 ml), washed with 6% NaHC03
(80 mg . kg-' i.v.) and maintained with subcutaneous injections of pentosolution, N HCI, and then water, and dried over MgS0,. Evaporation gave
barbitone (3 mg . kg-I). Drugs were administered via a femoral vein by
1.7 g of crude ester 51 as a colourless gum. This material and 0.5N NaOH
bolus injection and blood pressure was recorded from the right femoral
(20 ml) were stirred at room temp. for 1 h, water (10 ml) was added, and
artery. Results are given in Table 3.
the mixture washed with ether (2x) to remove non-acidic impurities. The
aqueous solution was acidified with 2N HCI, extracted (3x) with ether, and
the extracts dried over MgSO,. Evaporation of the solvent gave an oil (1.4
Ex vivo acriviry
g) which was crystallised from ether to give 350 mg (23% overall from 50)
Determination of rabbit plareler aggregation and concurrent change in
of a mixture of 15 and 52 as a colourless solid. Separation of the diastereosystemic arterial blood pressure
mers by HPLC proved difficult and gave 24 mg (2%) of 15 as colourless
Male rabbits (2-2.5 kg body weight) were anaesthetized with sodium
needles, mp. 159-161°C (EtOAc-hexane). - CI8H2.$J3O6(383.4) Calcd C
pentobarbitone (30 mg . kg-', i.v. followed by maintenance doses of 3 mg .
56.4 H 7.62 N 11.0 Found C 56.5 H 7.81 N 10.8. - 'H-NMR (200 MHz.
kg-') and systemic arterial blood pressure was recorded from a cannula
(CD3)SO): 6 (ppm) = 0.95-1.80 (21H, complex m, aliphatic H), 2.18 (2H.
filled with heparinized saline ( 5 units/ml) in a carotid artery; no heparin
t, J = 7 Hz, CHzCO), 3.75 (IH. d. J = 4 HZ, CHO), 4.05 (lH, t, J = 5 Hz,
was administered to the animal. Hydantoins were administered via a canCHN), 3.6-3.1 (3H, v.br. peak, 2xOH, NH), 10.1 (IH, brs, NH).- Rf0.34,
nula in a femoral vein. Blood samples (1.5 ml) were collected over 1 min
silica, CHCI3-MeOH-HOAc (90:5:5). The above experiment also gave 56
into a plastic syringe containing tri-sodium citrate (3.18%. 0.15 ml) from a
mg (4%) of 52 as colourless needles, mp. 154-156OC (EtOAc-hexane).
cannula inserted into a femoral artery, shaken gently and transferred to an
CI8Hz9N3O6(383.4) Calcd. C 56.4 H 7.62 N 11.0 Found C 56.3 H 7.50 N
Eppendorf plastic tube and spun in a modified Eppendorf centrifuge for 2
10.8. - 'H-NMR (200 MHz, (CD&SO): 6 (ppm) = 0.95-1.85 (21H. comsec (Model 5412. maximum centrifugal force 10 OOO x g). The PRP was
plex m, aliphatic H), 2.20 (2H. t, J = 7 Hz, CHzCO), 3.0-3.6 (3H, v.br.
collected and a 0.35 ml aliquot was transferred to the aggregometer and
peak, 2xOH, NH), 3.80 (lH, d, J = 4 Hz, CHO), 4.05 (lH, t, J = 5 Hz,
incubated at 37°C for 1 min prior to addition of sufficient ADP (20 FM) to
CHN), 11.1 ( l H , br.s, NH). - Rf 0.32, silica, CHC13-MeOH-HOAc
produce near-maximal aggregation. The time-interval between removal of
(9055). - Note: On TLC 15 is less polar than 52 (15: R, = 0.34; 5 2 R, =
blood samples and the transference of the PRP to the aggregometer was
0.32, silica, CHC13-MeOH-HOAc; 9 0 5 5 ) . However, on HPLC 52 is elutless than 1.5 min. Blood samples were removed at 10 min intervals once
ed off the column before 15 (52: R, 35-40min; 15: R, 40-46 min, Biosil,
the blood pressure of the rabbit had reached steady resting levels, 10-20
CHCI2-MeOH-AcOH, 97.5:2:0.5).
min following the preparative surgical procedures. After three control
-
Arch. Pharm. (Weinheim)327,307-317(1994)
13’-Aza-modified BW245C Analogues
blood samples had been prepared and the control aggregation of the platelets measured, the hydantoin was infused intravenously for 15 min from a
Braun slow-infusion apparatus. Blood samples were collected 10 min after
the start of the infusion. Two further 10 min samples post-infusion were
collected prior to commencing further infusions. Results are given in Table
4.
References
1
2
3
4
5
6
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8
9
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synthesis, platelet, chains, inhibitors, modified, activity, bw245c, aggregation, othet, aza, analogues
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