Optimisation of a pyrazole series of progesterone antagonists; Part 1

Article abstract of DOI:10.1016/j.bmcl.2010.04.018

The design and synthesis of a novel series of non-steroidal progesterone receptor antagonists is described. Ligand-lipophilicity efficiency (LLE) was used in the selection of a prototype agent for in vivo pharmacology studies.

Full text of DOI:10.1016/j.bmcl.2010.04.018

Bioorganic & Medicinal Chemistry Letters 20 (2010) 3384–3386
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Optimisation of a pyrazole series of progesterone antagonists; Part 1
*
Kevin N. Dack , Sarah Skerratt, Patrick S. Johnson, Paul A. Bradley, Ian R. Marsh
Department of World-Wide Medicinal Chemistry, Pfizer PharmaTherapeutics Division, Sandwich CT 13 9NJ, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 17 February 2010
Revised 7 April 2010
Accepted 7 April 2010
Available online 13 April 2010
The design and synthesis of a novel series of non-steroidal progesterone receptor antagonists is
described. Ligand-lipophilicity efficiency (LLE) was used in the selection of a prototype agent for
in vivo pharmacology studies.
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Progesterone
Endometriosis
LLE
The progesterone receptor (PR) is a member of the family of li-
gand-activated transcription factors that includes the estrogen
(ER), androgen (AR), glucocorticoid (GR) and mineralocorticoid
(MR) receptors.1 The use of antagonists for the treatment of a vari-
ety of progesterone-related diseases and disorders is of consider-
able interest. Recent studies have shown PR antagonists to have
application in the treatment of endometriosis and uterine fibroids.2
At present, RU-486 (mifepristone, 1) is the only PR antagonist ap-
proved for clinical use.3 RU-486 is an 11b-substituted steroid and
displays potent antagonist activity at other steroidal receptors, in
particular the glucocorticoid receptor (GR).4 This lack of selectivity
limits its chronic use.
As part of a program to identify non-steroidal PR antagonists,
we ran a high-throughput screen (HTS) using a PR binding assay.
Triage of the HTS data was guided by knowledge of target class
ligands. Specifically, compounds containing a cyanoaryl group
were prioritised as this group is a known ketosteroid A-ring iso-
stere in non-steroidal PR ligands such as the PR agonist Tanapro-
get (2)5 (Fig. 1).
4-H, 4-Cl, 4-F or 4-MeO resulted in a >10-fold reduction in PR
antagonist activity. The 3- and 2-CN analogues showed no PR
antagonism at 10 lM. Lersivirine (3) was also devoid of PR activity.
The selectivity profile of compound 5 was assessed and it was
found to be >40-fold selective for PR over AR, ER, GR and MR
(IC50s all >5 lM).
We decided to retain an N-substituent bearing a polar function-
ality in our optimisation work. We hoped this would act not only
as a selectivity handle but as a method of modulating potency,
lipophilicity, and metabolic stability. Previous experience with this
class of compound had shown that lipophilic analogues such as 5
often had an impaired metabolic stability profile.8 Therefore, in or-
der to deliver a candidate compound suitable for in vivo investiga-
tion, we felt it would be crucial to both improve PR potency and
reduce compound lipophilicity. To assess success against these
The HTS identified a series of phenoxypyrazoles, exemplified by
4 and 5 (Fig. 2). These were related to a series of proprietary non-
nucleoside HIV reverse transcriptase inhibitors (NNRTi’s)6–8 that
included the clinical candidate Lersivirine8 (3). In a functional PR
assay, using recombinant human PR expressed in CHO-MMTV-
beta-lactamase, both 4 and 5 were shown to be moderately potent
antagonists (IC50 of 224 nM and 131 nM, respectively).
Follow-up file screening around hits 4 and 5 demonstrated that
4-CN on the phenoxy moiety was superior to other mono-substit-
uents assessed (data not shown). Replacing 4-CN of 5 with either
Figure 1. Superposition of Tanaproget and progesterone-bound PR ligand binding
domain (LBD). The PR-LBD/Tanoproget (2) structure is shown in green (PDB code
1ZUC) and the PR-LBD/progesterone structure is shown in pink (PDB code 1A28).
Hydrogens bonds are shown as black dotted lines.
* Corresponding author. Tel.: +44 1304 648437; fax: +44 1304 651821.
E-mail address: kevin.dack@pfizer.com (K.N. Dack).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.04.018

K. N. Dack et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3384–3386
3385
N
O
OH
N
Me
Me
O
N
OH
N
N
1, Mifepristone (RU486)
3, Lersivirine
Me
Me
O
O
R1
N
N
N
N
Me
N
N
H
S
2,Tanaproget
PR Agonist
PR Antagonist HTS hits
4, R1 = H
5, R1 = CH2CH2OH
Figure 2. Reference structures.
R2
O
Cl
O
R2
R2
R2
R2
b
a
R2
d
O
N
O
O
O
O
c
R2
O
R1
O
N
N
N
R2
OH
N
N
N
5
4 R1 = H
e, f,
or g
6-15
Scheme 1. Synthesis of phenoxypyrazoles. Reagents and conditions: (a) NCS, TMSCl, DCM, 0–5 °C; (b) 4-cyanophenol, Cs2CO3, acetone, reflux, 30–80% for steps (a) and (b); (c)
2-hydroxyethylhydrazine, AcOH, 50–80%; (d) H2NNH2ÁH2O, AcOH, 25 °C, 85–95%; (e) R1-halide, KOtBu, 1,2-DME, 0?25 °C to give 6, 12 and 13; (f) ethylbromoacetate, KOtBu,
1,2-DME, 0?25 °C, followed by either hydrolysis using NaOH in MeOH (to give 7) or aminolysis with either saturated NH3/MeOH (to give 8), MeNH2/EtOH (to give 9 and 14)
or Me2NH/MeOH (to give 10); (g), ClCH2SCH3, KOtBu, 1,2-DME, 0?25 °C, followed by Oxone, MeOH, H2O, 25 °C to give 11 and 15 (45–55%).
criteria, we monitored compound ligand-lipophilicity efficiency
(LLE = Àlog (PR IC50) À log D),9 in addition to PR potency.
A range of pyrazole analogues with polar N-substituents were
prepared according to Scheme 1.
Metabolite identification studies with 9 and 11 showed that
oxidation of the ethyl R2 groups were occurring during microsomal
incubation. Replacement of the ethyl groups with cyclopropyl moi-
eties generated mono-methyl amide 14 and methyl sulfone 15.
These compounds retained the improved PR potency and LLE pro-
files of compounds 9 and 11 but had improved metabolic stability.
Because of the potency and LLE advantage, coupled with accept-
able metabolic stability, the amide 14 (PF-02367982) was selected
for further study. It was evaluated in a number of nuclear hormone
receptor binding and functional assays, and was found to be highly
Replacement of the hydroxyl group with a methyl ether (com-
pound 6, Table 1) gave an increase in PR potency, but not LLE. In
addition, the increase in lipophilicity rendered the compound vul-
nerable to increased metabolism. Oxidation of alcohol 5 to acid 7
resulted in PR inactivity. Pleasingly however, simple amides 8–10
restored PR antagonism, with secondary mono-methyl amide 9
having the highest LLE (4.9). Sulfone 11 also demonstrated a good
balance of properties, with an improved potency and LLE profile
over compounds 4 and 5. Aminoethyl 12 and homologated alcohol
13 were detrimental to PR potency.
selective for PR (IC50 >10
lM against GR, AR, MR, ER). It was also
selective (all IC50 values >10
lM) in wide-ligand profiling over a
wide range of >70 targets (CEREP, Bioprint™, http://www.cerep.fr).
The PR potency of 14 was confirmed (IC50 40 nM) with an alkaline

3386
K. N. Dack et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3384–3386
Table 1
Pyrazole analogues 4–15
R2
O
R2
R1
N
N
N
R1
R2
PR IC50 (nM)
Log Db
LLEc
HLM Clint
RLM Clint
a
d
d
Entry
4
5
6
7
8
H
Et
Et
Et
Et
Et
Et
Et
Et
Et
Et
cPr
cPr
224
131
13
>10,000
173
35
886
56
1130
204
47
3.4
3.0
3.3
3.9
<4.0
<2
4.6
4.9
3.2
4.6
4.5
3.7
4.8
4.5
26
25
>150
—
<10
22
35
60
17
27
223
—
>500
—
—
51
144
—
37
65
22
CH2CH2OH
CH2CH2OCH3
CH2CO2H
>3.9
(3.0)
(2.2)
2.6
(2.9)
2.7
1.1
(3.0)
2.5
CH2CONH2
CH2CONHCH3
CH2CON(CH3)2
CH2SO2CH3
CH2CH2NH2
CH2CH2CH2OH
CH2CONHCH3
CH2SO2CH3
9
10
11
12
13
14
15
<10
<7
71
2.6
<10
a
Concentration to inhibit by 50% the fluorescence from the beta-lactamase produced by progesterone (10 nM) stimulation of recombinant human PR expressed in a CHO-
MMTV-beta-lactamase cell line. Geometric mean of at least duplicate determinations.
b
Log D measured in octanol:pH 7.4 buffer. If log D was not obtained, then (clog P) is shown.
c
Ligand-lipophilicity efficiency (LLE) = Àlog (PR IC50) À log D. cLog P was used if measured log D was not available.
d
Clint is the intrinsic metabolic clearance in microsomes, in ll/min/mg of microsomal protein (HLM is human, RLM is rat).
one-induced arborisation of rabbit and cynomolgus macaque
Table 2
Pharmacokinetics of 14 dosed at 0.2 mg/kg iv and po
endometrium.10 Sulfone 15 was subject to further modifications
to optimise PR potency and physicochemical properties, and will
be reported in due course.
Species
Rat
Dog
Cla (mL/min/kg)
Vdb (L/kg)
T (h)c
27
0.9
0.9
78
3
0.4
2.1
90
Acknowledgements
Fd (%)
a
b
c
In vivo clearance after iv dosing.
Volume of distribution at steady state after iv dosing.
Half-life after iv dosing.
This Letter includes the work of a number of people in addition
to the authors. Compound synthesis: Toby Underwood, Simon
Wheeler, Carol Bains, Geoff Gymer, Dan Millns, Tom Findley, Felic-
ity Shaw. Discussions: Alan Stobie. Biology: Nick Pullen, Alex de
Giorgio-Miller and Michelle Tutt. ADME: Peter Bungay.
d
Bioavailability after oral dosing.
phosphatase assay using a human breast cancer cell line (T 47D)
that endogenously expresses PR.
References and notes
The pharmacokinetics of amide 14 was determined in rat and
dog (Table 2). Amide 14 was progressed as a prototype non-steroi-
dal PR antagonist into in vivo pharmacology studies, and was
shown to block progesterone-induced arborisation of rabbit and
cynomolgus macaque endometrium at 3 mg/kg po qd and at
2.5 mg/kg po bid.10 Taken together, these data confirmed the
in vivo pharmacological credentials of amide 14 as a specific PR
antagonist and support the utility of this class of agents in the
treatment of gynecological conditions such as endometriosis and
uterine fibroids.10
In summary, the optimisation of a novel series of non-steroidal
progesterone receptor antagonists using functional activity and LLE
to guide compound selection is described. Starting with the HTS
Hit 4 (R1 = H), we introduced polar side chains at R1 to improve po-
tency and selectivity, lower lipophilicity and increase LLE.
Changing the metabolically vulnerable ethyl groups at R2 to cyclo-
propyls improved the overall metabolic stability. Compound 14
was progressed as a prototype non-steroidal PR antagonist into
in vivo pharmacology studies, and was shown to block progester-
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