The discovery of GSK221149A: A potent and selective oxytocin antagonist

Article abstract of DOI:10.1016/j.bmcl.2007.11.008

Optimisation of a series of oxazole diketopiperazines has led to the discovery of a very potent and selective oxytocin antagonist GSK221149A. GSK221149A has been shown to inhibit oxytocin-induced uterine contractions in the anaesthetised rat.

Full text of DOI:10.1016/j.bmcl.2007.11.008

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 18 (2008) 90–94
The discovery of GSK221149A: A potent and selective
oxytocin antagonist
John Liddle,a,* Michael J. Allen,d Alan D. Borthwick,a David P. Brooks,c
David E. Davies,a Richard M. Edwards,c Anne M. Exall,a Chris Hamlett,a
Wendy R. Irving,a Andrew M. Mason,a Gerald P. McCafferty,c Fabrizio Nerozzi,a
Simon Peace,a Joanne Philp,a Derek Pollard,a Mark A. Pullen,c Shaila S. Shabbir,b
Steve L. Sollis,a Timothy D. Westfall,c Pat M. Woollard,b
Charlene Wuc and Deirdre M. B. Hickeya
aDepartment of Medicinal Chemistry, GlaxoSmithKline Research and Development, Medicines Research Centre,
Gunnels Wood Road, Stevenage, Hertfordshire, UK
bDepartment of Drug Metabolism and Pharmacokinetics, GlaxoSmithKline Research and Development, Medicines Research Centre,
Gunnels Wood Road, Stevenage, Hertfordshire, UK
cDepartment of Urogenital Biology, Cardiovascular and Urogenital Centre of Excellence for Drug Discovery,
GlaxoSmithKline Research and Development, King of Prussia, PA, USA
dDepartment of Molecular Discovery Research, GlaxoSmithKline Research and Development,
New Frontiers Science Park, Harlow, Essex, UK
Received 14 September 2007; revised 1 November 2007; accepted 2 November 2007
Available online 6 November 2007
Abstract—Optimisation of a series of oxazole diketopiperazines has led to the discovery of a very potent and selective oxytocin
antagonist GSK221149A. GSK221149A has been shown to inhibit oxytocin-induced uterine contractions in the anaesthetised rat.
Ó 2007 Elsevier Ltd. All rights reserved.
Preterm labour occurs in 10% of all births worldwide
and is the single largest cause of neonatal morbidity
and death.1 Although it is difficult to estimate the cost
of neonatal intensive care of low birth-weight babies,
it is thought to exceed $5 billion/year in the US alone.2
Oxytocin (OT), a cyclic nonapeptide neurohypophyseal
hormone, binds to OT receptors stimulating contractil-
ity in human myometrium and is widely used for the
induction of labour.3 The density of oxytocin receptors
in myometrium is elevated during pregnancy and it is
believed that a premature increase of such receptors
can initiate preterm labour by sensitising the uterus to
relatively unchanged circulating levels of OT.4,5 The de-
sign of OT antagonists as potential tocolytic agents for
the prevention of premature labour has therefore been
an intense area of research for many years.6 Although
the peptide OT antagonist atosiban has been approved
for clinical use in Europe, it is not orally bioavailable
and has low plasma stability rendering it less likely to
be useful for long-term or preventative treatment.7 Atos-
iban also has a greater affinity for the vasopressin V1a
receptor than the oxytocin receptor.8
We recently reported the diketopiperazine 1 to be a po-
tent and orally bioavailable OT antagonist (Table 1).9
Compounds of this series had been hindered by poor
physicochemical properties, including low solubility
and relatively high logD, which consequently led to rel-
atively high plasma protein binding and cytochrome
P450 interactions, with high clearance in cynomolgus
monkey and human microsomes.
Structure–activity relationships (SAR) had indicated
that both the indanyl and isobutyl fragments contrib-
uted to potency,13 and hence we sought to investigate
the modification of the exocyclic aryl and amide func-
tionality. Herein, we describe our efforts around the
five-membered ring heterocycles to improve the poor
Keywords: Oxytocin; Labour; Diketopiperazine; Tocolytic.
*
Corresponding author. E-mail: john.2.liddle@gsk.com
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.11.008

J. Liddle et al. / Bioorg. Med. Chem. Lett. 18 (2008) 90–94
91
Table 1. Physicochemical properties of 1
hOT pKi10
Table 2. Inhibition of the binding of OT at the human OT receptor10
8.9–9.2
3.4
F
O
O
R
LogDa
HSAb
Sol.
H
N
N
93%
F
H
N
O
O
HN
O
<1 lg/mL
N
CYP450 3A4 IC50 4 lM
HN
O
Compound
R
hOT pKi
O
2a
7.9
1
a Chromatographic lipophilicity measurement.11
b Human serum albumin binding.12
O
3
4
8.2
9.0
physicochemical and pharmacokinetic properties of this
series.
CH3
O
Replacing the difluorophenyl ring with unsubstituted five-
membered ring heterocycles resulted in approximately a
10-fold loss in intrinsic potency (Table 2). Potency was re-
gained on the introduction of a lipophilic group, such as
methyl or bromide, in the 4 or 5 position. In an attempt
to access the putative lipophilic pocket reached by these
substituents, a number of trifluoromethyl heteroaryls
were prepared. The CF3 group was invariably the most
potent substituent but its strongly electron withdrawing
nature led to epimerisation of the exocyclic position in
neutral or mildly basic media rendering the compounds
less attractive for developability reasons.
H3C
CH3
5
8.9
7.4
O
O
O
NHCH3
6
Br
7
9.1
O
Polar groups were poorly tolerated; the methylamide 6
was significantly less potent than the parent furan 3
although large substituents, such as phenyl and pyridyl
(analogue 9), were tolerated. Disubstitution in both
the 4 and 5 positions did not increase potency further.
Methyl substitution ortho to the ring junction was toler-
ated as exemplified by the dimethyl oxazole 14 having
similar potency to the methyl oxazole 15. Although
the SAR trends were consistent across all heterocycles,
the absolute potencies were often significantly different.
The difference in potencies of similarly substituted het-
erocycles may therefore be a reflection of the heterocy-
CF3
8
10.0
8.6
O
Py
9c
10
11
S
Br
cle’s ability to orientate the substituent into
lipophilic pocket of the receptor.
a
8.7
S
Although the introduction of lipophilic substituents
gave favourable binding affinities, unsurprisingly this
was generally accompanied by poor physicochemical
properties (Table 3). Indeed, the furyl and thiophene
derivatives were poorly water-soluble, relatively highly
protein bound and carried undesirable CYP450 interac-
tions (results for the 3A4 isozyme are shown in Table 3).
More polar azoles were able to accommodate lipophilic
substituents whilst maintaining good physicochemical
properties. The oxazole analogues had good aqueous
solubility, low protein binding and minimal CYP450
interaction making the series most attractive for further
investigation.
Br
9.0
S
S
12
8.2
9.4
H3C
S
13b
N
H3C
O
14b
8.5
Established SAR suggested that a range of amides was
tolerated and we therefore envisioned using this func-
tionality as a handle to optimise the pharmacokinetic
profile. Potency and pharmacokinetic data on selected
N
CH3
(continued on next page)

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J. Liddle et al. / Bioorg. Med. Chem. Lett. 18 (2008) 90–94
Table 2 (continued)
selective with respect to the vasopressin V1a receptor
(>10-fold). Oxazoles with polar amide substituents, such
as the alcohol 18, had poor oral exposure in the rat.
Good pharmacokinetic profiles were achieved with
less-polar amides. Tertiary amides 19 and 20 had mod-
erate clearances and volumes of distribution of approx-
imately 1 L/kg resulting in half lives of around 1 h. Both
compounds had acceptable bioavailabilities in rat in ex-
cess of 30%.
Compound
R
hOT pKi
H3C
O
15
8.9
N
16b
9.1
9.8
O
N
The SAR of the isobutyl portion of 1 was sensitive to
relatively minor modifications (Table 4) which had dra-
matic effects on the pharmacokinetic profile. Branching
at the a-carbon was tolerated and resulted in a superior
rat pharmacokinetic profile.
F3C
O
17
N
a tert-Butyl amide rather than isopropylamide.14
b Dimethylamide rather than isopropylamide.14
c Py, 2-pyridyl.
Indeed 22, GSK221149A, had greater oral exposure in
the rat compared to 20 in both DMSO/Peg and
HPMC/Tween formulations, with good bioavailability
and a half life of 1.4 h. Compound 22 also has low to
moderate intrinsic clearance in microsomes from three
pre-clinical species (rat, dog, cynomolgus monkey) and
low intrinsic clearance in human microsomes and is
more potent than 20. Although several amide analogues
of 22 were prepared, with a range of physicochemical
properties, 22 offered the preferred overall profile.
Table 3. Properties of selected compounds
O
R
H
N
N
HN
O
O
With this data in hand, 22, GSK221149A, was selected
for progression to further selectivity and in vivo efficacy
models. GSK221149A is a potent and selective oxytocin
receptor antagonist (Table 5) with no detectable agonist
activity. GSK221149A is greater than 10-fold more po-
tent than atosiban with a far superior selectivity profile
with respect to vasopressin receptors.15
Compound
R
hOT Log HSAb CYP450
Sol.
c
pKi
Da
3A4 IC50 (lg/ml)
CH3
4
9.0 3.3
10.0 3.6
8.7 3.7
96
97
99
7
3
1
34
O
The in vivo activity of GSK221149A was evaluated in
anaesthetised, non-pregnant Sprague–Dawley female
rats.15 Uterine activity was measured as an integral of force
over time. Intravenous administration of GSK221149A
produced a dose-dependent decrease in oxytocin-induced
uterine contractions, with an ID50 = 0.27 0.60 mg/kg
and IC50 = 180 nM.
CF3
8
<1
<1
O
Br
10
S
The synthesis of the diketopiperazines followed previ-
ously described chemistry.16 The isopropyl amides were
conveniently prepared in three steps from the corre-
sponding isonitrile via the Ugi reaction as outlined in
Scheme 1. The intermediate mixture of diastereoisomers
23 was deprotected with 4 M HCl and subsequently cyc-
lised by treatment with base in a one-pot procedure.
Generally, a 2:1 mixture of diastereoisomers was formed
with the desirable (R)-isomer 8 being the minor product
isolable by chromatography.
H3C
O
15
8.9 2.5
63
>100
>240
N
a Chromatographic lipophilicity measurement.11
b Human serum albumin binding (%).12
c Lowest IC50 from CYP450 3A4 using substrates diethoxyfluoroscein
(DEF), 7-{3-(4-phenylpiperazin-1-ylmethyl)benzyl}resorufin (PPR)
or 7-benzyloxyquinoline (7-BQ).
GSK221149A and other tertiary amides were prepared
in four steps via the Ugi reaction as outlined in Scheme
2. A 2:1 mixture of diastereoisomers 24 was formed with
the desirable (R)-diastereoisomer being the minor
product. Hydrogenation of crude 24 furnished the cyc-
lised phenol 25, again enriched with the undesirable
(S)-diastereoisomer. Activation of the mixture 25 with
carbonyl diimidazole followed by the addition of 2 N
HCl promoted epimerisation at the exocyclic position
amides are recorded in Table 4. The oxazole amides 18–
22 had good aqueous solubility (>220 lg/mL), low
plasma protein binding (<80% HSA) and no significant
cytochrome P450 interactions (CYP3A4 IC50 > 100 lM).
As expected, a range of functionality could be incorpo-
rated in the amide without having detrimental effects on
potency. Compounds 15 and 18–22 demonstrated antag-
onist activity in an in vitro functional assay and were

J. Liddle et al. / Bioorg. Med. Chem. Lett. 18 (2008) 90–94
93
Table 4. Profile of selected oxazoles
Me
O
N
O
2
R
N
HN
O
1
R
H
O
Compound
R1
R2
hOT pKi
Rat PK (iv 2 mg/kg, po 5 mg/kg)
Cla
t1/2
AUCc
Fd (%)
b
15
18
19
20
NHi-Pr
8.9
8.9
8.8
225e
Me
N
218e
OH
35
1.4
1070e
45
N
3633e
1190f
N
O
O
8.7
8.2
23
14
1.1
1.3
33
69
21
22
4200e
N
N
5470e
1800f
ꢀ100
O
9.0
19
1.4
42
IV formulation 5:95 20% polyvinylpyrrolidone in DMSO: 35% sulfobutylether-7-betacyclodextrin in water.
a Clearance (mL/min/kg).
b Half life (h).
c AUC (hr ng/ml).
d Bioavailability.
e PO formulation 5:95 DMSO/PEG400 using amorphous.
f PO formulation: hydroxypropylmethylcellulose (HPMC)/Tween using crystalline material.
Table 5. Binding affinity at recombinant/native receptors
benzotriazol-1-yloxytripyrrolidinophosphonium hexa-
fluorophosphate followed by the addition of morpholine
and subsequent column chromatography yielded homo-
chiral GSK221149A.
Receptor
GSK221149A Ki (nM)
Atosiban Ki (nM)
hOT
rOT
hV1a
hV1b
hV2
0.65
4.1
11
32
>12,000
>10,000
950
0.15
44
In conclusion, GSK221149 has nanomolar affinity for
the oxytocin receptor with >1400-fold selectivity over
the closely related vasopressin receptors. GSK221149A
has a good rat pharmacokinetic profile, low human
microsomal clearance and has been shown to inhibit
oxytocin-induced contraction in vivo in the anaesthe-
tised rat.
330
and yielded the acids 26 with the required (R)-diastereo-
isomer as the major product. Acid activation with
CF3
CF3
CF3
O
O
O
O
OH
O
O
O
a
H
H
NHBoc
O
b, c
N
N
N
N
BocHN
HN
O
O
NH2.HCl
MeO2C
NC
CH3O
O
23
8
Scheme 1. Reagents: (a) triethylamine, MeOH; (b) 4 M HCl in dioxan, MeOH; (c) triethylamine, DCM.

94
J. Liddle et al. / Bioorg. Med. Chem. Lett. 18 (2008) 90–94
O
O
O
N
O
N
O
N
N
H
OH
O
OBn
O
O
OH
H
a
b
H
NHZ
N
N
N
ZHN
MeO2C
O
OBn
NH2.HCl
HN
O
NC
CH3O
O
25
24
c
O
N
O
O
O
N
N
H
OH
O
O
d
N
H
N
O
HN
O
HN
O
26
22
Scheme 2. Reagents and conditions: (a) triethylamine, MeOH; (b) H2, Pd/C, ethanol/acetic acid; (c) carbonyl diimidazole, CH2Cl2 3 h then acetone/
2 N HCl; (d) benzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate, dichloromethane 1 h then morpholine.
10. The method for the binding assay is described in the
following publication Wyatt, P. G.; Allen, M. J.; Chilcott,
J.; Hickin, G.; Miller, N. D.; Woollard, P. M. Bioorg.
Med. Chem. Lett. 2001, 11, 1301.
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